EFFECT OF TWO INVASIVE PLANT SPECIES ON SOME SOIL CHEMICAL PROPERTIES

Department of Botany, Obafemi Awolowo University, Ile-Ife, Nigeria

Received 9th September, 2014; accepted 2nd April, 2015

* Author for correspondence

Abstract

Invasive plants are capable of modifying ecosystem function. However, it is difficult to generalize because impacts often appear to be species and site-specific. In this study, we examined the impacts of two major invasive plant species (Tithonia diversifolia (Hemsl) A. Gray and Chromolaena odorata (L.) R.M. King and H. Robinson) on pH and nutrient pools in the top soil. Sample plots, 5 m x 5 m each, were established on invaded and non-invaded areas at 10 sites in areas invaded by each of these species. In each plot, soil samples were randomly collected at depth 0-15 cm and analysed for chemical properties (pH, organic carbon, exchangeable cations (Ca,Mg, K, Na), nitrogen and phosphorus). One-way ANOVA was used to determine significant difference in soil properties on invaded and non-invaded plots. The results showed that the invaded plots had significantly higher pH, organic carbon, N, P, Ca, Mg, K and Na than the non-invaded plots (p<0.05). There was significant difference on the effects of the two species on the soil properties. It is concluded that these invasive species significantly improved the soil fertility of the invaded sites.

Key words: Invasive plants, soil, soil fertility, Tithonia diversifolia, Chromolaena odorata

Introduction

Alien species are non-native or exotic organisms that occur outside their adaptive and dispersal ranges (Raghubanshi et al., 2005; Bruton and Merron, 1985). According to SCBD (2001), invasive alien species are species introduced deliberately or unintentionally outside their natural habitat, where they have the ability to establish themselves, invade, out-compete natives and take over the new environment. Some of the alien species become invasive when they are introduced deliberately or unintentionally outside their natural habitats into new areas where they express the capability to establish and invade, out-competing native species. These invasives are widely distributed in all kinds of ecosystems throughout the world, and include all categories of living organisms. Nevertheless, plants, mammals and insects comprise the most common types of invasive alien species in terrestrial environments (Raghubanshi et al., 2005). Invasive species have now affected every ecosystem types on the planet and are considered as the second greatest global threat to biodiversity, after habitat destruction (Raghubanshi et al., 2005; Essa et al., 2006).

The invasion of plant species also alters the physicochemical properties of soils in the invaded areas (Dogra et al., 2008). The soils in the invaded areas become nutrient-rich, which generally helps in the growth of invasive species.

El-Ghareeb (1991) studied the allelopathic effect of the invasive plant Tribulus terrestris on surrounding vegetation in an abandoned field of Kuwait. The study demonstrated that besides the growth- inhibitory effect of the plant on other plants, the soil moisture and concentrations of N, P and K were significantly higher in T. terrestris site. Other evidences support the idea that higher resource availability increases the susceptibility to invasion of plant communities (Burke and Grim, 1996; Maron and Connor, 1996). 

Chromolaena odorata and Tithonia diversifolia are two plant species introduced into sub-Saharan Africa which became established and invasive. Tithonia diversifolia, commonly called Mexican sunflower

and native to central America, was probably introduced into Africa as an ornamental (Akobundu and Agyakwa, 1987). It is reported to be present in different countries of Sub-Saharan Africa, in Kenya (Niang et al., 1996), Malawi (Ganunga et al., 1998), Nigeria (Ayeni et al., 1997), Rwanda (Drechsel and Reck, 1998), Uganda and Cameroon (Shokalu, 1997), Zimbabwe (Jiri and Waddington, 1998) and Zambia (Shokalu, 1997., Muoghalu and Chuba, 2005). It is also reported to be widely distributed throughout the humid and sub-humid tropics in Central and South America, Asia and Africa (Sonke, 1997).

Chromolaena odorata (L.) R. M. King and H. Robinson (Eupatorium odoratum L.) (Asteraceae) also known as Chromolaena or Siam weed, is a native of North, Central and South Americas and the West Indies (Henderson, 2001). It has become highly aggressive invasive throughout many tropical countries (McFadyen and Skarratt, 1996). It was introduced to Botanical Gardens of Dacca (India), Java and Peradeniya (Sri Lanka) in the 19th century and for ornamental reasons in Southern Africa in the early 20th century. In West Africa, the plant was accidentally introduced with forestry seeds in Nigeria in 1947 and was deliberately introduced to Ivory Coast in 1952 to control Imperata spp. in coffee and oil palm plantations (De Rouw, 1991) following a recommendation by the famous botanist, Auguste Chevalier. Since introduction, it has become established and invasive. It has been reported to be one of the worst alien invasive plant species in the humid tropics and sub-tropics of the old world (Holm et al., 1977), where it is a menace to agriculture, human health and biodiversity. Its prolific regeneration from seeds frustrates attempts to eliminate the plant manually (De Rouw, 1991).

Tithonia diversifolia and Chromolaena odorata are aggressive colonisers of new sites with high light and adequate moisture conditions. Tithonia diversifolia is now a prominent and fast-growing invasive plant species in Nigeria, inhabiting the rainbelt of the southern part of Nigeria especially the southwest and the coastal areas. It also inhabits the wet part of the Guinea savanna especially along the fringes of the rainbelt (latitude 6-9° N) (Agboola et al., 2005). Chromolaena odorata is one of the most troublesome invasive species in Nigeria and in many tropical countries. The two species, C. odorata and T. diversifolia, form dense stands of 2.3 m high and can eliminate almost other vegetations. In Nigeria, T. diversifolia is fast displacing C. odorata from its habitat.

Though several studies on the detrimental effects of the allelochemicals on plant germination and growth of these species have been reported (Baruah et al., 1994, Tongma et al., 1997, Onwugbuta-Enuyi, 2001, Ismail and Chong, 2002; Bogateh et al., 2006), there has been paucity of studies on the effect of these species on the soil properties of the ecosystem they have invaded. In this study, the effects of C. odorata and T. diversifolia on soil properties of weed-infested sites is reported.

 

MATERIALS AND METHODS

 Study Area

The study was carried out in Ile-Ife in South-western Nigeria. Ile-Ife lies within latitudes 07º30′ N to 07º35′  N and longitudes 04º30′ E to 04º35′  E. The original vegetation of Ile-Ife has been described as lowland forest zone (Keay, 1959), semi-deciduous moist forests (Charter, 1969) and Guineo-Congolian forest drier type (White, 1983). Hall (1969) also described the vegetation as the dry forest sub-group.

The geology of the area is underlain by the Pre-cambrian basement complex of the Southwestern Nigeria. The rock consists of banded gneisis and migmatite quartzites, quartz, mica, schists and related rocks (Smyth and Montgomery, 1962).

            The soils of the area are moderately to strongly leached and have low to medium humus contents, weakly acidic to neutral surface layers and moderately to strongly acidic sub-soils (Smyth and Montgomery, 1962). It is derived from materials of old basement complex that is made up of granitic, metamorphosed sedimentary rock (Hall, 1969).

In each location, a pair of 5 m x 5 m adjacent sample plots was established. One plot of the pair was placed in invaded vegetation (invaded plot) where the invading plant species was dominant and has a high cover and the second plot in a neighbouring vegetation, where the invader had no cover (uninvaded plot). The uninvaded plot was chosen so as to have as similar site conditions as possible to the invaded plot. Five soil samples each were randomly collected to a depth of 0-15 cm from invaded and uninvaded plots using a soil auger. The five samples were bulked for each plot, air-dried and sieved through <2 mm mesh size for chemical analysis.

 

Chemical Analysis

The soil samples were analysed for soil pH, exchangeable cations (Ca, Mg, K, Na), total nitrogen, available phosphorus and organic matter. Soil pH was determined in 0.01M CaCl2 (1:2 soil solution ratio) using a glass electrode pH meter (Pye model 292). Total nitrogen, available phosphorus and exchangeable cations (Ca, Mg, K, Na) analyses were done following procedures outlined in Tel and Rao (1982). The soil Organic matter was determined using Walkey – Black method (Black, 1965). The significant difference test in soil properties between the soil properties of invaded and uninvaded plots was carried out by one-way analysis of variance. The significant difference test between the effects of the two species was also carried out by one-way analysis of variance using the SPSS.

 

RESULTS

The soil pH of invaded sites was significantly higher than that of uninvaded sites (F= 39.421; P<0.05) (Table 2). It was slightly alkaline in the invaded sites (Table 2). The pH of the Tithonia diversifolia invaded sites was not significantly different from that of Chromolaena odorata invaded sites (Table 2). The soil nitrogen concentrations of the Chromolaena odorata and Tithonia diversifolia invaded sites were significantly higher than those of the uninvaded sites (Table 2). The nitrogen concentration of soils of Tithonia diversifolia invaded sites were not significantly different from those of Chromolaena odoratainvaded sites (Table 2).

Discussion

            This study clearly points out the increase in the soil nutrients of the invaded sites as a result of invasion of T. diversifolia and C. odorata when compared with the uninvaded sites, thus rendering the soil of the invaded sites nutrient-rich and more fertile when compared with the uninvaded sites. These findings are in support of the findings of Norgrove et al. (2008) where farmers in Cameroon have reportedly associated the presence of C. odorata with higher soil fertility (Yonghachea, 2005). Likewise, in Ghana, farmers consider both C. odorata and earthworm casts as indicators of good soils and link these mechanistically by stating that C. odorata provides litter input, shade and a moist environment which promotes earthworm activity (Adjei-Nsiah et al., 2004). In southern Cameroon, Norgrove and Hauser (1999) found that weed biomass, dominated by C. odorata, accounted for 76% of variation in earthworm cast production in a cropped field and subsequently, in an adjacent site, Norgrove et al. (2003) found that mulching with C. odorata resulted in an increase in cast production and that these casts were richer in nitrogen and potassium than those derived from non-mulched plots, probably due to feeding on the N- and K-rich C. odorata residue. There is increase in soil organic matter (C) content under stands of invasive plants which has been reported in many previous studies (Scott et al., 2001; Caldwell, 2006; Fickbohm and Zhu, 2006; Heneghan et al., 2006; Vila et al., 2006). Plant invaders can increase levels of soil organic matter through the production of leaf litter and leaching of root exudates, fueling microbial activity associated with the nitrogen cycle and nitrogen fixation (Jones et al., 2003; Liao et al., 2008). Increased soil nitrogen concentrations observed in this study may give invasive plants a competitive advantage over native plants that are better competitors in low-nitrogen soils (Ehrenfeld and Scott, 2001; Evans et al., 2001; Corbin and D’Antonio, 2004).

The invasion of Tithonia diversifolia and Chromolaena odorata also alters the physico-chemical properties of soils in the invaded areas. The soils in the invaded areas become nutrient-rich, which generally helps in the growth of invasive species. It was clear from the results that the values of all soil nutrients were found to be higher in the invaded areas as compared to the control. Minimum change was observed in the case of pH compared to other parameters. El-Ghareeb (1991) studied the allelopathic effect of the invasive plant Tribulus terrestris on surrounding vegetation in an abandoned field of Kuwait. His study demonstrated that besides the growth-inhibitory effect of the plant on other plants, the soil moisture and concentrations of N, P and K were significantly higher in T. terrestris site. In the present study, the amounts of available

nutrients were significantly more in invaded soils as compared to the uninvaded soils. Abundant evidences support the idea that higher resource availability increases the susceptibility to invasion of plant communities (Burke and Grim, 1996; Maron and Connor, 1996). An increase in soil organic matter (C) content under stands of invasive plants has been reported in many previous studies (Scott et al., 2001; Caldwell, 2006; Fickbohm and Zhu, 2006; Heneghan et al., 2006; Vila et al., 2006). This has been attributed to increased biomass production and litterfall (Yelenik et al., 2004) or to reduced litter decomposition rates (Ogle et al., 2003).

Similarly, T. diversifolia and C. odorata produce a lot of biomass and litter fall especially during the dry season which is decomposed and add organic matter and nutrients to the soils of invaded sites, thereby increasing these soil properties.

Nutrient dynamics may also become altered as a result of changes in the physical properties of the soil caused by the introduction of new species (Boettcher and Kalisz, 1989; Finzi et al., 1998; Kelly et al., 1998; Ehrenfeld, 2001). The results of the study justify the use of these species to improve soil fertility and improve crop yields (Ayeni et al., 1997; Drechsel and Reck, 1998; Jama et al., 2000; Norgrove et al., 2003; Adjei-Nsiah et al., 2004).

 

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HYPOGLYCAEMIC AND HYPOLIPIDAEMIC EFFECTS OF TOTAL SAPONINS FRACTION EXTRACTED FROM IRVINGIA GABONENSIS (AUBRY LECOMTE EX O’RORKE) BAILL. STEM BARK IN RATS

Department of Medical Biochemistry, University of Benin, Benin City, Nigeria.

Received 16th May, 2015; accepted 28th June,  2015.

Abstract

The crude aqueous stem bark extract of Irvingia gabonensis has been shown to possess significant long-term hypoglycaemic/antidiabetic, hypolipidaemic and antioxidant effects on normal rabbits and streptozotocin diabetic rats. Phytochemical analysis of this plant revealed the presence of saponins. In this study, sequential solvent extraction and chromatographic methods were used to recover total saponins fraction from I. gabonensis stem bark. The I. gabonensis total saponins fraction (ITSF) was then administered orally for 28 days to normal female rats in three dose regimes; 10, 20 and 40 mg/kg body weight, in groups 2, 3, and 4, respectively. The effects of ITSF on the fasting blood sugar (FBS) and serum lipid profile of the treated rats were compared to untreated normal control (Group 1).  The FBS of 10, 20 and 40 mg/kg saponins-treated groups were not significantly different from the control for the duration of this study. However, the three dose administrations of ITSF caused significant reductions in serum total cholesterol and LDL-cholesterol. Though the saponins-treated rats at the three dose groups exhibited higher triacylglycerol concentrations, the increase was not statistically significant. Similarly, saponins treatment did not significantly alter serum HDL-cholesterol concentration. The reductions observed in serum total cholesterol and LDL-cholesterol in the treated groups indicates the anti-hypercholesterolaemic effects of these saponins.

Key words: Irvingia gabonensis, Saponins, Solvent extraction, Hypolipidaemic, Hypoglycaemic 

Author for correspondence

Department of Biochemistry, Faculty of Life Sciences, University of Benin, Benin City, Nigeria.

Department of Biochemistry, Adekunle Ajasin University, Akungba-Akoko, Ondo State, Nigeria

 

INTRODUCTION

Irvingia gabonensis(Aubry Lecomte ex O’Rorke) Baill. is a species of African trees which belong to the family Irvingiaceae (Ladipo et al., 1996). It is commonly called wild mango, African mango, bush mango, “dika” or “Ogbono” (Adamson et al., 1990). They bear edible mango-like fruits, and are especially valued for their fat and protein-rich nuts. I. gabonensis is indigenous to the humid forest zone from the northern tip of Angola, including Congo, DR Congo, Nigeria, Côte d’Ivoire and south-western Uganda (Etong et al., 2014). I. gabonensis tree grows to a height of 15-40 m, the bole is slightly buttressed. The bark is greyish, smooth or very slightly scaly. The leaves are 5-15 x 2.5-6 cm, elliptic to slightly obovate, one margin often a little more rounded than the other. The flowers are yellowish to greenish-white and are arranged in slender, clustered racemes or small panicles above the leaves. Fruits are yellowish when ripe, broadly ellipsoid and variable in size between varieties, 5-7.5 cm with a yellow, fibrous pulp surrounding a large seed (Orwa et al., 1999). The stem bark of the plant contains phytochemicals like phenols, alkaloids, flavonoids, cardiac glycosides, anthraquinones, saponins, steroids and tannins (Omonkhua and Onoagbe, 2010; Ojo et al., 2014).

Saponins are a diverse group of compounds widely distributed in the plant kingdom. They are characterized by structures containing either a triterpenoid or steroidal aglycone and one or more sugar chains (Francis et al., 2002). Their structural diversity is reflected in their physiochemical and biological properties which are exploited in a number of traditional (as soaps, fish poison and molluscides), industrial and medicinal applications (cholesterol lowering and anticancer, etc.) (Francis et al., 2002). Saponins exert a wide range of pharmacological activities including expectorant, anti-inflammatory, vaso-protective, hypocholesterolaemic, immune-modulatory, hypoglycaemic, molluscidal, antifungal and antiparasitic effects (Sparg et al., 2004). Many studies have established saponins as the active ingredient in many herbal medicines and highlight their contribution to health benefits of foods such as soyabeans and garlic (Matsuura, 2001).

Saponins isolated from various sources are known to have hypoglycaemic effects (Petit et al. 1993; Kim et al., 1998; Yoshikawa et al., 2001). Some of the mechanisms proposed for the hypoglycaemic effect include stimulation of pancreatic â-cells (Petit et al., 1993), delay of glucose absorption in the small intestine and the inhibition of glucose transport across the brush border of the small intestine (Matsuda et al., 1999). Saponins have also been demonstrated to exhibit hypocholesterolemic effects (Zhao et al., 2008). They also reduce low density lipoproteins cholesterol levels (Francis et al., 2002). Saponins may lower serum lipid by forming micelles with dietary fats, enhancing HDL (high density lipoprotein) cholesterol scavenging ability and delaying the absorption of dietary fat by inhibiting pancreatic lipase activity (Hu et al., 2002).

Our previous studies revealed that aqueous extract of I. gabonensis stem bark exhibit significant hypoglycaemic/antidiabetic and hypolipideamic effects. Since saponins have been documented to possess these effects, this study was designed to extract total saponins from I. gabonensis stem bark and determine the effect of this extract on fasting blood sugar and serum lipid profile of rats.

MATERIALS AND METHODS

Chemicals/Reagents

Randox kits for glucose, total cholesterol, total triacylglycerols and HDL-cholesterol (Randox Laboratory Ltd., Ardmore, United Kingdom), silica gel (Sigma, London), methanol, n-hexane, ethyl acetate, butanol and acetic acid (BDH Chemical Limited, Poole, England).

 Plant materials

The stem bark of I. gabonensis was obtained from the local forest at Akungba-Akoko, Ondo State, Nigeria. The collection was done from mid-June to July during daytime. Herbarium specimen, with voucher number UIH 22286 was deposited at the University of Ibadan Herbarium. The bark was dried under a shade for two weeks and ground into powder.

Extraction of saponins

A modification of the method of Hostettmann et al. (1991) was used to extract saponins from I. gabonensis stem bark. One kilogramme (1 kg) ground sample was extracted with 1.5 litres of methanol for 24 h. The methanolic extract was concentrated using a rotary evaporator and partitioned between n-hexane and water (1:2, v/v) in a separating funnel. The mixture was vigorously shaken and allowed to stand for 24 h. The water layer was carefully recovered and concentrated. This concentrated solution was partitioned between ethyl acetate and n-butanol (1:2, v/v). The butanol fraction was concentrated to obtain crude saponins fraction. Concentrated crude saponins extract was applied to a silica gel column. The column was eluted with n-butanol: acetic acid: water (1:1:1 v/v/v). The fractions were collected, pooled together and used for the experiment. The scheme below was employed for the extraction of I. gabonensis total saponins fraction.

 Animals and Experimental Protocol

A total of twenty (20) female adult rats of the Wistar strain, with average weight of 115.7 g were obtained from the Department of Animal Science, University of Ibadan, Nigeria. They were kept in a well aerated room, with 12 h light and 12 h dark cycles. They were allowed food (standard pelleted feed) and water ad libitum and allowed to acclimatize for three weeks before the commencement of the study. Treatment of the animals conformed to the guidelines for the Care and Use of Laboratory Animals (The National Academy of Sciences, 2011). The rats were divided into four (4) groups of five (5) rats each as shown below:

     

The total saponins fractions were orally administered (by gavage) daily for 28 days.

Blood collection

Blood for monitoring fasting blood sugar was drawn from the tail vein of each rat at specific intervals. At the end of 28 days, the rats were sacrificed; the thoracic/abdominal regions were opened to expose the heart and other organs. Blood was obtained through heart puncture. Blood for glucose assay was collected in fluoride bottles while that for lipid profile analysis was collected in plain bottles. The blood samples were allowed to clot on ice and centrifuged at 1,000 X g for 5 minutes; the serum was then separated for analysis. 

Biochemical Analyses

Fasting blood glucose was measured by the glucose oxidase method of Barham and Trinder (1972). Serum triacylglycerol concentration was measured by the method of Tietz (1990). Serum total cholesterol levels and serum HDL-cholesterol concentrations were analyzed according to Richmond (1973) and Lopes-Virella et al. (1977) methods, respectively. Serum LDL-cholesterol level was calculated as described by Friedewald et al. (1972).

Statistical analysis

The differences among groups were analyzed by the one-way analysis of variance (ANOVA). Inter-group comparisons were done using Duncan’s Multiple Range Test (DMRT) with 95% confidence intervals.  The SPSS 15.0 (SPSS Inc., Chicago, Illinois, USA) was used for this analysis. Values are means of 5 determinations ± SEM

RESULTS

Sequential solvent extraction of I. gabonensis stem bark gave a total saponins yield of 0.64%. The effect of I. gabonensis total saponins fraction (ITSF) on the fasting blood sugar (FBS) and serum lipid profile of normal female rats are presented in figures 2 and 3, respectively.

 

Figure 2: Effect of I. gabonensis total saponins fractions (ITSF) on FBS concentration (mg/ml) of normal female rats.

Treatment with saponin fractions in the different dose groups did not significantly alter the fasting blood sugar (FBS) of rats. It is pertinent to note that from day 5 to day 28, the 10 mg/kg saponins treated group exhibited slightly higher FBS concentration compared to normal control, the 20 and 40 mg/kg saponins-treated groups. Nevertheless, all FBS values recorded for the period of 28 days were within normal range.

Figure 3: Effect of I. gabonensis total saponins fraction on serum lipid profile (total cholesterol, triacylglycerols, LDL-cholesterol and HDL-cholesterol) (mmol/l) of normal female rats. Values carrying different letters are statistically different at P<0.05.

Serum total cholesterol and LDL-cholesterol concentrations were significantly (p<0.05) reduced in the three dose groups (10, 20 and 40 mg/kg saponins) of the treated rats compared to untreated normal control. Surprisingly, the reduction in serum total cholesterol was inversely proportional to the dose i.e. the lowest dose (10 mg/kg saponins) produced the greatest reduction in total cholesterol. The serum triacylglycerols concentration of the treated rats increased slightly in all dose groups (especially the 10 and 40 mg/kg saponins groups) compared to untreated control while HDL-cholesterol concentration of the treated rats was not significantly different from control.

 

DISCUSSION AND CONCLUSION

Saponins have been shown to have profound biological effects including anti-hyperlipidaemic (particularly anti-hypercholesterolaemic), anti-obesity, anti-diabetic and anti-oxidant effects. Preliminary phytochemical analysis of I. gabonensis stem bark revealed the presence of saponins (Omonkhua and Onoagbe, 2010). The extraction of saponins from other medicinal plants have shown that the concentration of saponins in different plants and even different parts of the same plant varies. For example, Hassan et al. (2011) reported that the whole plant of Schwenkia americana Linn, rhizome of Asparagus africanus Lam, leaves of Dichrostachys

cinerea Linn, stem bark of Ficus iteophylia Miq and the leaves of Indigofera pulchra Willd gave crude saponins percentage yields of 2.74, 3.59, 1.62, 0.81, and 1.57, respectively. Also, Sri et al. (2011) reported that the leaves, stems and tubers of Anredera cordifolia gave crude saponins yield of 2.81, 0.37 and 4.32, respectively.

            Iivingia gabonensis total saponins fractions (ITSF) did not significantly alter the FBS concentration of female rats. The slightly higher FBS observed at 10 mg/kg treatment may reflect the inability of this low concentration of saponins to exert significant inhibition on glucose absorption (Murakami et al., 1999; Yoshikawa et al., 2001).  Though several saponins have been shown to possess hypoglycaemic effects, under the conditions of this study, the fasting blood sugar of the treated rats was not significantly altered.

The anti-hyperlipidaemic effect of ITSF was obvious in the lower total cholesterol and LDL-cholesterol concentration of the treated female rats. The HDL-cholesterol concentration of the treated rats was slightly lower than the control. It can, however, be observed that the HDL-cholesterol concentration of the 40 mg/kg treatment group was similar to the control. Surprisingly, under the conditions of this study, ITSF increased serum triacylglycerols concentrations. Saponins react with bile acids to form large mixed micelles, thus facilitating the excretion of bile acids. Consequently, serum cholesterol is recruited to form bile acids resulting in a decrease in serum cholesterol concentration (Oakenfull, 1986; Oakenfull and Sidhu, 1990). In addition, some saponins reduce dietary cholesterol absorption, which ultimately decreases serum cholesterol concentration (Stark and Madar, 1993). Saponins have also been observed to delay intestinal absorption of triacylglycerols by inhibiting pancreatic lipase activity (Han et al., 2000; Francis et al., 2002).  While reduction in cholesterol concentration may reflect both increased cholesterol excretion and inhibition of its absorption, reduction in triacylglycerols would normally reflect the inhibition of lipase activities. This inhibition may require a longer period of time, hence the lack of triacylglycerols reduction after 28 days of treatment. Longer and/or higher doses of saponins treatment may be required for triacylglycerols reduction.

This study revealed that sequential solvent extraction of saponins from I. gabonensis stem bark gave a yield of 0.64%. ITSF did not significantly alter FBS concentration in rats but had significant anti-hyperlipidaemic effects. This group of saponins could be important sources of cholesterol lowering agents which could be beneficial in the treatment of hypercholesterolaemia.

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PHYSICOCHEMICAL ASSESSMENT AND POLLEN ANALYSIS OF HONEY FROM NSUKKA, NIGERIA

  Received 22nd January, 2015; accepted 2nd June, 2015

ABSTRACT
The study was aimed at determining the physico-chemical properties and pollen spectra of honey samples from commercial dealers in Nsukka, Nigeria. The physicochemical parameters were analyzed using the official methods of analysis of the Association of Analytical Chemists and the acetolysis procedure was used in analyzing the pollen content. The results of the physico-chemical properties showed that there were significant differences (P< 0.05) in the quantity of the assessed honey parameters from various locations. The free acidity occurred in the range of 2.35 ± 0.16 – 36.30 ± 1.63 meq./kg; pH 3.9 ± 0.01 – 4.7 ± 0.01; ash 0.37 ± 0.00 – 0.454 ± 0.00 %; protein 0.11 ± 0.00 – 2.01 ± 0.00 %; glucose 24.96 ± 0.04- 65.0 ± 0.82 mg/100 g; fructose 28.84 ± 0.03 – 79.0 ± 0.82 mg/100 g; sucrose 1.05 ± 0.01- 35.70 ± 0.14 mg/100 g and HMF 0.29 ± 0.01 – 51.80 ± 0.01 mg/100 g. The assessment of the honey properties indicated that most of these parameters meet international standard for good quality honey, except for  the presence of very high sucrose content in most of the samples. This might suggest possible adulteration with disaccharide sugar.The botanical characterization of the honeys showed that three of eight honey samples were monofloral, two bifloral and six multifloral. Trees and shrubs were major pollen and nectar sources. Eighty-eight pollen types were identified and were derived mostly from trees and shrubs. The non-nectariferous plant sources contributed pollen in the range of 1.62-5.34 % in the honey samples. Important honey plants identified include Elaeis guineensis Jacq., Syzygium guineense (Willd.) DC., Combretaceae-Melatomataceae, Parkia biglobosa (Jacq.) R. Br. ex G. Don f., Nauclea latifolia Sm., Crossopteryx febrifuga (Afzel. ex G.Don) Benth., Prosopis africana (Guill., Perr. & A. Rich.) Taubert, Lannea sp., Brachystegia eurycoma Harms.  and Parinari sp., while the major sources of bee pollen load include Elaeis guineensis Jacq., Alchornea cordifolia (Schumach.) Mull. Arg., Combretaceae-Melatomataceae, Nauclea latifolia Sm., Lannea sp. and Prosopis africana (Guill., Perr. and A. Rich.) Taubert. Further studies are recommended to ascertain other biological components and the possible source of high sucrose.
*Author for correspondence. *Njokuocha, C.  R.  Department of Plant Science and Biotechnology, University of Nigeria, Nsukka, Enugu State, Nigeria.

Key words: Honey plants, Honeybees, nectariferous plants, Southeastern Nigeria,  monofloral, bifloral and multifloral honeys
INTRODUCTION
Honey is a major product resulting from the foraging activities of Honeybees (Apis melifera L.). It may be defined as a sweet natural substance produced by Honeybees from nectarine and/ or extra-floral nectarines or excretions of plant sucking insects on the living parts of plants (Agwu and Njokuocha, 2004). Following a series of processes (Codex Alimentarius Commission, 2001; Nombre et al., 2010), honey is then formed. Honey is consumed directly as food and can serve as nutritional ingredient and plays an important role in the manufacture of numerous food products, confectioneries and drugs. The variability and uniqueness of honey flavour, aroma, nutritional and medicinal properties as well as colour are determined by the sources of nectar and pollen (Dafni et al., 1988; Ramalho et al., 1991; Abu-Tarbousch et al., 1993). It is rich in nutrients such as sugars, organic acids, mineral salts, proteins, vitamins, antioxidants and many biologically active compounds (Jasim  et al., 2007; Chefrour et al., 2009). These substances contribute in making honey one of  the richest natural food substances.
The quality of honey is influenced by a number of factors such as the nectar sources, pollen, climatic conditions, harvesting methods and storage conditions (Perez-Arquillue et al., 1994; Conti, 2007). These factors determine considerably the percentage composition of the physico-chemical properties such as Hydroxymethyl furfural (HMF), simple sugars, moisture, free acidity and ash present in honey. These properties are used in accessing the quality of honey. Besides, they influence the demand and commercial values of such honey (Bryant, 2001; Ohe et al., 2004; Rouff and Bogdanov, 2004; Nombre et al., 2010). The acceptability of any honey on the international market is dependent on the extent to which it meets international standard for good quality honey (Codex Alimentarius Commission, 2001; European Union Council, 2002).
In Nigeria, honey production has a very high potential because of the diverse floral species and complex structure of the Nigerian vegetation. Unfortunately, honey production and apicultural farms in the different regions of the country are few. Most of the honey is harvested from the wild. Because the quantity of honey produced is not adequate for both domestic and industrial uses, the available honey sold to the open market is predisposed to adulteration. To ascertain whether the honey sold in the open market meets international standard, it becomes pertinent to analyze some samples for authenticity.
The pollen content of wild honey is often loaded with a variety of pollen types which may vary in class, from primary pollen to secondary pollen, important minor and minor pollen (Silici and Gokceoglu, 2007). The pollen content of honey provides information on the floral and geographical origin of the honey (Agwu and Akanbi, 1985; Bryant, 2001). Pollen grains also contribute to the nutritional and good qualities of honey because they are important sources of ash, fibre, carbohydrate, vitamins, protein and major essential amino acids such as leucine and lysine (Hassan, 2011).
This study was aimed at determining (1) the physico-chemical properties of the honey samples from Nsukka, Nigeria (2) the common pollen grains in the honey samples and (3) compare the values with international standard.
MATERIALS AND METHODS
Honey samples were obtained from commercial dealers from various locations in Nsukka, Enugu State, Nigeria. The physicochemical analysis was carried out in the Department of Biochemistry, University of Nigeria, Nsukka, while the pollen analysis was done in the Department of Plant Science and Biotechnology of the same University.

Physicochemical analysis
The physicochemical properties were analysed according to the methods of Association of Analytical Chemists (AOAC, 1997), while pollen analyses were carried out according to Moore and Webb (1978) and Njokuocha and Nnamani (2009).

Determination of Moisture Content
Ten grams (10 g) of each of the honey samples was obtained (Metter P162N) and heated in an electric heater (Metter, B & T BS 2648) at 70 oC for 2 hours and at 100 oC for the next 4 hours until constant weight was obtained. The percentage moisture content was determined as follows:
W1 – W2 /W0 – W1 x 100
Where, W1 = weight of fresh sample + crucible
W2 = weight of dry sample (solids) + crucible
W0 = weight of crucible
Percentage weight of dry sample (solid) = W1  – Wo / W2 – WO  x 100

Determination of Ash Content
Two grams (2 g) of each of the honey sample was weighed in a pre-weighed crucible and  heated in a muffle furnace (Gallenkamp – Hotspot) and ashed at 600 oC for 3 hrs. The percentage ash content was obtained by dividing the weight of ash by the weight of fresh sample and multiplied by 100 (AOAC, 1997).

Determination of Protein Content
Two grams (2 g) of the honey sample was weighed into the digestion flask and 25 ml of concentrated H2SO4 and a digestion mixture consisting of a speck of selenium, 1 g of anhydrous K2SO4 and 7 g of CuSO4 were added into the flask. The flask was slightly agitated to mix the content and then heated for 2 hrs to digest until the colour changed from green to a clear solution. The digest was allowed to cool and transferred to 100 ml distillation flask and made up to the mark with distilled water. Ten (10) ml of the digest was put in the distillation flask and 10 ml of 40 % NaOH was added to it. The ammonia (NH3) released during the distillation was collected in form of NH4OH  in a  50 ml conical flask containing 20 ml of 4 % boric acid solution and few  drops of methyl indicator. Then, 35 ml of the distillate collected was titrated against standard 0.1 N HCL solution till the colour turned pink. A blank was also run through the above steps. Percentage crude protein content of each of the honey samples was calculated thus:
% crude protein =    (H-B) x N x 0.014 x D x 100 x 6.25
Wt. of honey sample x V
Where:
H = honey sample titration, B = blank titration, N = normality
D = dilution of sample after digestion, V = volume taken for distillation
0.014 = milli equivalent weight of nitrogen

Determination of pH
Ten per cent (10 %)  honey solution was prepared by weighing 10 g of the sample and dissolving this in 100 ml of distilled water and the pH reading was taken with a pH meter (pHep:  Hanna Instruments).

Determination of Hydroxymethylfurfural (HMF)
Ten grams (10 g) of the honey samples was dissolved in 20 ml oxygen free cold water and thereafter transferred to 50 ml volumetric flask and made up to 50 ml. Two (2) ml of the solution was introduced into two test tubes and 5 ml solution of p-toluidine was added to each tube. Thereafter, 1 ml of barbituric acid was added into one test tube and 1 ml of water to the other test tube (blank). The absorbance of test sample was read against the blank at 550 nm using spectrophotometer (Spectro 21D, Pec Medicals, USA).
HMF (mg/100 g) = Absorbance /cell path length x 19.2 (AOAC, 1997).

Determination of free acidity
Five ml of dilute honey sample (10 %) was titrated against 0.02 N sodium hydroxide using 0.3 ml of phenolphthalein indicator. The relative amount of titretable acid was determined thus:
Free Acidity:       Titre value x Normality of NaOH x 4.6
Weight     of sample
Determination of Sugar Content
One gram (1 g) of the honey sample was dissolved in 50 ml of distilled water to form a stock solution for determination of glucose, fructose and sucrose contents. Spectrophotometer used in measuring absorbance was Spectro 21D Pec Medicals USA.

Glucose Content: One mill of the stock solution was added to 1 ml of alkaline copper reagent and boiled for 8 minutes.  One (1) ml of phosphomolybdic acid reagent and 7 ml of water were added to the cooled sample and the absorbance at 420 nm against a blank was taken (AOAC, 1997).

Fructose Content: One  (1) ml each of reagents A  (50 mg Resinol in 50 ml ethanol) and B  (50 ml conc. HCl + 10 ml of water) was added to 1 ml of the stock solution  and heated in a water bath at 80 oC for 8 minutes. On cooling to room temperature, the absorbance was read at 530 nm (AOAC, 1997).

Sucrose Content: Five  (5) mls of 2 % Oscinol was added to 1 ml of stock solution and boiled for 12 minutes. It was made up to 25 ml with distilled water and the absorbance at 620 nm was read (AOAC, 1997).

Pollen Analysis
Ten grams (10 g)  of the agitated honey sample was diluted with 35 mls of acidified water (3 ml Conc. H2SO4 and 997 ml distilled water) to dissolve the colloidal matters and sugars. The sample was centrifuged at 2000 rpm for ten minutes to recover the residue and subsequently acetolysed (Moore and Webb, 1978; Njokuocha and Nnamani, 2009). The final polliniferous residue was suspended in 2 drops of glycerol-alcohol in vial bottles from where samples were taken for routine counting and pollen grain identification under the light microscope. Routine counting was done on the entire area (484 cm2) of the cover slip. Identification of pollen grains was aided by photomicrographs in Bonnefille and Riolett (1980), Y’bert (1979), APLF (1974) and pollen slides in the Environment and Palynology Research Unit, Department of Plant Science and Biotechnology, University of Nigeria, Nsukka.

RESULTS
Physicochemical Characteristics
The mean values of the physicochemical characteristics of the honey samples are summarized in Tables 1 and 2. The mean protein value range varied from 0.11± 0.00i to 2.01 ± 0.00a. The protein analysis showed that the mean quantity of protein in sample 2 was significantly different (P< 0.05) from that of other samples. Similarly, there were significant differences (P< 0.05) in the mean quantity of protein recorded between other samples (Table 1). The mean pH values showed that there was no significant difference (P>0.05) in the quantity of acid recorded in samples 1, 2, 3, 4, 6, 7, 8, 10 and 11, but they varied significantly (P<0.05) from those of 5 and 9. However, sample 3 (P>0.05) did not vary significantly from those of 5 and 9 (Table 1). The mean values varied in range from 3.9 ± 0.10 in sample 9 to 4.7 ± 0.01 in sample 4.  Free acidity also varied considerably, with the highest value of 36.30 ± 1.63 meg/kg recorded in sample 4, followed by samples 2 (31.00 ± 0.82 meg/kg) and 1 (29.00 ± 1.41 meg/kg), while the least value was recorded in sample 11 (2.35 ± 0.16g meg/kg). There was a significant difference (P<0.05) in the mean value of free acidity between sample 4 and other samples (Table 1). The mean value of the ash content showed that samples 3 and 5 were significantly different (P<0.05) from other samples. Samples 1, 2 and 4 did not vary significantly (P> 0.05), but varied significantly (P<0.05) from samples 6, 7, 8, 9, 10 and 11 with the exception of sample 4 which was not significantly different (P>0.05) from that of sample 10 (Table 1). The honey samples varied from 0.37 ± 0.00 % to 0.45 ± 0.00 % with the highest value recorded in sample 3 (0.45 ± 0.00 %) and the least in sample 7 (0.371 ± 0.00h %). The sugar characteristics showed that fructose value was higher than that of glucose in all the samples. The sum of the fructose and glucose in honey samples 4 – 11 was within the acceptable EU commission and Codex Standard (> 60 mg/100g) for blossom honey. However, samples 1- 3 did not meet internationally acceptable standard. The mean values of glucose and fructose showed that sample 8 was significantly different (P<0.05) from those of other samples (Table 2). The percentage sucrose content ranged from 1.05 ± 0.01 % to 35.70 ± 0.14 % in the samples. The mean value of sucrose in sample 11 was significantly (P<0.05) different from those of other samples. The sucrose content in samples 1-3 was within internationally acceptable limits of 5 % for blossom honey, while the values of samples 4-11 exceeded the acceptable limit (Table 2). The moisture content of the samples ranged from 5.85 ± 0.01 % in honey sample 6 to 25 ± 0.82 % in sample 11. The mean value of moisture in sample 11 was significantly different (P<0.05) from those of other samples. There was also a significant difference in the mean values between other samples (Table 2).  The HMF values ranged from 0.29 ± 0.01 mg/kg in samples 1 and 2 to 51.80 ± 0.01 mg/kg in honey sample 11. The mean value of sample 11 was significantly different from those of other samples, while other samples were significantly different from one another (Table 2).
Pollen Analysis
The pollen analysis showed that the honey samples contained between 21 – 36 different pollen types derived from trees, shrubs, herbs and climbers. Apart from the pollen of entomophilous plants which are usually associated with honey samples, pollen grains of zoophilous, amphiphilous and anaemophilous plant species were also recorded in all the honey samples. The pollen analysis showed that the honey samples belong to different floral classes. This was because of the relative contribution of pollen to the honey by different plant species. Honey samples 7, 8 and 10 were monofloral, because of the high dominance of pollen of Elaeis guineensis which contributed over 54 % of the pollen content. Honey sample 5 (dominated by Elaeis   guineensis and Alchornea cordifolia) and honey sample 6 (dominated by Elaeis guineensis and Senna sp.) were classed as bifloral with the dominant pollen types contributing between 20 – 36 % of the total pollen recorded.  Honey samples 1, 2, 3, 4, 9 and 11 were multifloral because the dominant pollen types were more than five.
Some of the important honey plants distinguished in the honey samples included  Elaeis guineensis, Syzygium guineense, Irvingia wombolu, Combretaceae-Melastomataceae, Parkia biglobosa, Nauclea latifolia, Crossopteryx febrifuga, Prosopis africana, Lannea sp., Senna sp., Brachystegia eurycoma and Parinari kirstingii.

DISCUSSION
Physicochemical characteristics
The protein content of honey is regarded as an important nutritional ingredient, although it is generally low compared to the sugar content. The presence of protein in the honey samples indicates that they are nutritionally rich and, therefore, suitable as dietary supplement. The protein values compare favourably with those of organic and non-organic honey samples from Brazil (Sereia et al., 2011). Protein has been reported in honey samples from Kogi State, Nigeria (Oyeleke et al., 2010) as well as in honey samples from Algeria (Chefrour, 2009). The percentage protein in honey varies widely according to the plant sources. These variations are believed to be a function of the floral origin, nectar sources, climate and quantity as well as type of pollen in the honey (Sereia et al., 2011). This may be responsible for the significant differences (P< 0.05) observed in the mean value of the honey samples from various sources.
The pH and free acidity of honey (meg/kg) can affect the texture and palatability as well the stability and shelf life of the honey. The mean pH range in this study is comparable to the acceptable standard pH range (3.42 – 6.1) of honey from the U.S. (White Jr, 1975). Only samples 4, 7 and 10 had pH values higher than the recommended range (3.2 – 4.5) (Nigussie et al., 2012) and they were not significantly different (P> 0.05) from one another. Comparable pH range has been obtained in honey samples from some parts of Nigeria (Adebiyi et al., 2004; Lawal et al., 2009) and Burkina Faso (Nombre et al., 2010). Such an acidic range indicates that the honey samples are capable of inhibiting the growth of many microbial pathogens (Oyeleke et al., 2010).
All the honey samples had free acidity that was less than 50 meq/kg, which is the   maximum limit for good quality honey prescribed by Codex Alimentarius Commission (CODEX, 2001) and the EU Commission Directive (The Council of the  European Union, 2002). This result is comparable to that of Meda et al. (2005) from Burkina Faso and Sereia et al. (2011) who worked on organic and non-organic Apis  mellifera honeys from Brazil. The acidity in honey may originate from the action of the enzyme glucose oxidase which forms gluconic acid. Further acids may be associated with the variation in organic acids derived from different nectar sources, action of bacteria during honey ripening and the amount of mineral constituent present in the honey (White jr., 1975; Ouchemoukh et al., 2007).
The ash content of a honey is determined highly by the characteristics of the soil, floral nectar and pollen spectrum (Bryant, 2001). These factors may have contributed to the significant difference (P< 0.05) observed in the mean ash content of the different honey samples. The percentage ash content is an indication of the mineral composition of honey and a quality criterion for assessing honey’s botanical origin. The ash content of the honey samples varied significantly from one other and none of them exceeded the 0.6 % limit for floral honey. Similar levels of ash content have been reported in multifloral honey from some other parts of Nigeria (Adebiyi et al., 2004; Lawal et al., 2009; Anyansola and Banjo, 2011), Burkina Faso (Meda et al., 2005) and Tigray honey in Ethiopia (Nigussie et al., 2012). However, higher ash contents have been reported in Moroccan and Czeh honeydew honey (Diez et al., 2004).
The low moisture contents of some of the honey samples indicate their degree of   maturity prior to  harvest. Moisture content is an important factor influencing the   stability of honey against fermentation, taste, viscosity and granulation during storage (Nombre et al., 2010). It is affected by the degree of ripeness/maturity, season of collection, climate, extraction technique, storage conditions and moisture content of the original nectar (Nigussie et al., 2012). Apart from samples 8, 10 and 11 which were significantly different (P< 0.05) from others, the average moisture contents of other samples were within the international standard and directives (20%) for floral honey (Codex Alimentarius, 2001; The Council of European Union, 2002). The average moisture content of the honey from this region is attributed to the tropical savanna climate and the fact that most of the honeys in this region are produced during the period of relatively low amount of rainfall when most honey plants are in flower (Keay et al., 1964). The moisture content of the analysed honey samples compare to a considerable extent with the works of Ayansola and Banjo (2011) from southwestern Nigeria and Nombre et al. (2010) from Burkina Faso as well as that of non organic honey fraction from Brazil (Sereia et al., 2011).
The moisture contents of some Nigerian and Burkina Faso and Ethiopian honeys have been found to be considerably higher (Adebiyi et al., 2004; Meda et al., 2005; Lawal et al., 2009; Nigussie et al., 2012). Generally, water is important in assessing the grade of ripeness of honey and its shelf–life. High water content accelerates the rate of fermentation, spoilage, loss of flavour and aroma. It favours rapid growth of in-situ yeasts which results in sour taste, increased water content and higher acetic acid content (Nigussie et al., 2012).
HMF is one of the factors used to ascertain the freshness and degree of deterioration of honey. Apart from honey sample 11 which was highly significantly different (P< 0.001) from others, all the other honey samples have HMF below the limits (40 mg/kg) of internationally acceptable standard (Codex Alimentarius, 2001; The Council of European Union, 2002). This indicates that honey samples 1-10 were still fresh and conform to international standard and directives. It also shows that the sugar content of the honey samples was not affected by storage conditions. This finding compares favourably with similar works conducted in other parts of Nigeria (Lawal et al., 2009, Ayansola and Banjo, 2011) and North-east Algeria (Chefrour et al., 2009). It also agrees to a large extent with that of Martinez-Gomez et. al. (1993), (Spanish Eucalyptus honey) and Sereia et al. (2011) on organic and non-organic Brazilian honeys.
The sugar analysis shows that the monosaccharides (fructose and glucose) were the main sugars in the honey samples. This is in agreement with the findings of other workers (Krell, 1996; Qamer et al., 2008; Ayansola and Banjo, 2011) where fructose and glucose represented 80-95 % of total sugars in original honey. The higher values of fructose than glucose in all the samples are also in agreement with the findings of Crane (1990) and Ayansola and Banjo (2011). The sum of fructose and glucose shows that the values of all the monosacharides in the honey samples fall virtually within the limits of acceptable international honey standard which should not be less than 60 % (Codex Alimentarius Commission, 2001 and Council of European Union, 2002). The minor difference observed in the values of a few samples may be due to processing. Similar findings have been reported in Nigeria and other places (Qamer et al., 2008; Nombre et al., 2010; Ayansola and Banjo, 2011).
observed in the mean ash content of the different honey samples. The percentage ash content is an indication of the mineral composition of honey and a quality criterion for assessing honey’s botanical origin. The ash content of the honey samples varied significantly from one other and none of them exceeded the 0.6 % limit for floral honey. Similar levels of ash content have been reported in multifloral honey from some other parts of Nigeria (Adebiyi et al., 2004; Lawal et al., 2009; Anyansola and Banjo, 2011), Burkina Faso (Meda et al., 2005) and Tigray honey in Ethiopia (Nigussie et al., 2012). However, higher ash contents have been reported in Moroccan and Czeh honeydew honey (Diez et al., 2004).
The low moisture contents of some of the honey samples indicate their degree of   maturity prior to  harvest. Moisture content is an important factor influencing the   stability of honey against fermentation, taste, viscosity and granulation during storage (Nombre et al., 2010). It is affected by the degree of ripeness/maturity, season of collection, climate, extraction technique, storage conditions and moisture content of the original nectar (Nigussie et al., 2012). Apart from samples 8, 10 and 11 which were significantly different (P< 0.05) from others, the average moisture contents of other samples were within the international standard and directives (20%) for floral honey (Codex Alimentarius, 2001; The Council of European Union, 2002). The average moisture content of the honey from this region is attributed to the tropical savanna climate and the fact that most of the honeys in this region are produced during the period of relatively low amount of rainfall when most honey plants are in flower (Keay et al., 1964). The moisture content of the analysed honey samples compare to a considerable extent with the works of Ayansola and Banjo (2011) from southwestern Nigeria and Nombre et al. (2010) from Burkina Faso as well as that of non organic honey fraction from Brazil (Sereia et al., 2011).
The moisture contents of some Nigerian and Burkina Faso and Ethiopian honeys have been found to be considerably higher (Adebiyi et al., 2004; Meda et al., 2005; Lawal et al., 2009; Nigussie et al., 2012). Generally, water is important in assessing the grade of ripeness of honey and its shelf–life. High water content accelerates the rate of fermentation, spoilage, loss of flavour and aroma. It favours rapid growth of in-situ yeasts which results in sour taste, increased water content and higher acetic acid content (Nigussie et al., 2012).
HMF is one of the factors used to ascertain the freshness and degree of deterioration of honey. Apart from honey sample 11 which was highly significantly different (P< 0.001) from others, all the other honey samples have HMF below the limits (40 mg/kg) of internationally acceptable standard (Codex Alimentarius, 2001; The Council of European Union, 2002). This indicates that honey samples 1-10 were still fresh and conform to international standard and directives. It also shows that the sugar content of the honey samples was not affected by storage conditions. This finding compares favourably with similar works conducted in other parts of Nigeria (Lawal et al., 2009, Ayansola and Banjo, 2011) and North-east Algeria (Chefrour et al., 2009). It also agrees to a large extent with that of Martinez-Gomez et. al. (1993), (Spanish Eucalyptus honey) and Sereia et al. (2011) on organic and non-organic Brazilian honeys.
The sugar analysis shows that the monosaccharides (fructose and glucose) were the main sugars in the honey samples. This is in agreement with the findings of other workers (Krell, 1996; Qamer et al., 2008; Ayansola and Banjo, 2011) where fructose and glucose represented 80-95 % of total sugars in original honey. The higher values of fructose than glucose in all the samples are also in agreement with the findings of Crane (1990) and Ayansola and Banjo (2011). The sum of fructose and glucose shows that the values of all the monosacharides in the honey samples fall virtually within the limits of acceptable international honey standard which should not be less than 60 % (Codex Alimentarius Commission, 2001 and Council of European Union, 2002). The minor difference observed in the values of a few samples may be due to processing. Similar findings have been reported in Nigeria and other places (Qamer et al., 2008; Nombre et al., 2010; Ayansola and Banjo, 2011).

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CYTOMORPHOLOGICAL STUDIES OF SOME NIGERIAN ASTERACEAE (COMPOSITAE) -TRIBE INULEAE

Department of Plant Biology and Biotechnology  University of Benin, Benin City, Nigeria.                                                                  

 Received 15th March, 2012; accepted17th May,  2012

Abstract
Cytological and morphological studies were carried out on 26 species distributed in 10 genera of the tribe Inuleae. The pollen fertility test of the species studied was high, ranging from 58% to 100%; this showed that the plants were stable and hybridization was not responsible for their differences.  Chromosome counts of 17 species are reported for the first time from the Nigerian flora  (Anisopappus dalzelii, n=14; Helichrysum albiflorum, n=7; H. mechowianum, n=7;  H. quartinianum, n=7; H. rhodolepis, n=7; Inula klingii, n=9; I. subscaposa, n=9; Laggera braunii, n=10; L. graclis, n=10; L. oloptera, n=10; Mollera angolensis, n=10; Porphyrostemma chevalieri, n=15; Pulicaria undulata, n=10; Sphaeranthus angustifolius, n=10; S. flexuosus, n=10; S. senegalensis, n=10 and Vicoa leptoclada n=9). Chromosome count of n=8 for Gnaphalium luteo-album differed from previous reports. Basic chromosome numbers of 3 genera i.e Gnaphalium x, = 7, 8; Mollera, x = 10 and Porphryostemma x, =15, have been suggested for the first time. The basic chromosome number x=10 is provisionally taken for  the tribe Inuleae and the results from the study agree with the classification of the tribe. The achene morphology of all the taxa studied have been drawn and described for the first time.Distribution maps of all the taxa studied are presented for the first time and they have confirmed that these plants are prevalent in high altitudes. The occurrence of Helichyrsum quartinianum Bak. is reported for the first time in the flora of Nigeria.

Key Words: Chromosome number, achene morphology, base number, Inuleae

INTRODUCTION
The family Asteraceae is one of the largest family amongst the dicotyledons. It comprises of about 1,302 genera and 22,000 species (Turner, 1977). A large number of the Asteraceae are troublesome weeds e.g Carthamus tinctorus. Many are weeds of roadsides, open waste places and pastures.
The earlier intrafamilial classification of the family relied more on traditional data including characters derived from anatomy, chromosome studies, embryology, morphology, palynology, phytochemistry and ultrastructural studies to solve the problems of the family and this intrafamilial systematics of the Asteraceae has been extensively studied by Cronquist (1955, 1977), Heywood et al.(1977), Turner (1977) but the classification into subfamilies and tribal relationships remained unresolved.
Using these traditional techniques, Adams (1963) categorised the family into 13 tribes while Willis and Airyshaw (1973) and Smith (1977) divided the family into 12 tribes. Most of the workers on Asteraceae systematics recognized that the family can be divided into 2 subfamilies – Tubuliflorae (Asteroideae) and Cichoroideae in 12 or 13 tribes. The tribe Inuleae was divided into 2 groups – Inulinae and their distant relation Gnaphaliinae.
In recent times, taxonomists apply new techniques such as DNA hybridization and numerical taxonomy for classification and sorting relationship amongst the family. Using numerical taxonomy, Bremer (1987, 1994) applied an explicit cladistic methodology in developing hypothesis of evolutionary relationships between the family and Anderberg (1989) and Jansen et al. (1991c) scrutinized and analysed the family by means of a computerised parisimony programme (PAUP) and suggested 15 tribes. Bremer, and Jansen and their colleagues (Bremer, 1987; Jansen and Palmer, 1987, 1988; Jansen et al., 1991a, 1991b; Bremer and Jansen, 1992; Michaels et al., 1993; Jansen and Kim, 1996; Lundberg and Bremer, 2003; Funk and Specht, 2007; Funk and Chan, 2009; Funk et al., 2009a, 2009b and Funk, 2010) have used morphological and molecular data to change this- the family is now divided into 42 – 43 tribes and the traditional tribe Inuleae is now divided into two tribes that are not closely related (Gnaphaliieae and Inulinae) (Gupta and Gill, 1986; Funk, 2010). With this new classification, the problem of the family has remained unresolved.
The tribe Inuleae is characterized by having 2 types of florets in each capitulum, anther base sagitate or tailed, leaves alternate or basal; head radiate or discoid; involucral bracts commonly dry and scarious in several rows; foliaceous or petaloid; receptacle naked or scaly. Tubular florets present. According to Gupta and Gill (1986), the tropical taxa of this family have not been well studied and phylogenetic conclusions are not possible since most of the species have not been studied.It is represented in West Africa by 18 genera and 49 species. Of these, 29 species distributed in 12 genera occur in Nigeria (Hutchinson and Dalziel, 1963). In Nigeria, Olorode (1974), Olorode and Olopade (1978), Olorode and Olorunfemi (1978), Olorode and Okoli (1978), Isawumi (1982),  Ayodele (1997),  Adegbite (1995) and Omoigui (1985) have been engaged in the biosystematic studies of this family.
There is no consensus on the circumscription of the tribes into subfamilies neither the number of genera into the tribes nor the relationship amongst the 42-43 recognised tribes on the other hand. As a guide to solving this problem, Kaderit and Jeffrey (2007) and their numerous co-workers have reordered the genera, tribes and subfamilies within the family based on recent morphology and molecular results and this work is now the standard reference for descriptions of the tribes and genera of the family (Funk, 2010).
There is dearth of information on the tribe Inuleae. The aim of this paper is to present the cytomorphological studies of some Nigerian Asteraceae in Nigeria with emphasis on the tribe Inuleae, with 10 genera consisting of 26 species.

MATERIALS AND METHODS
Plant samples for the present study were collected from the different parts of the country and their exact locations have been given in Table I. The plants were identified at the Forest Herbarium (FHI) of the Forestry Research Institute, Ibadan. The identified plants have been deposited at the University of Benin Herbarium. All the herbarium sheets deposited in Forestry Herbarium, Ibadan and the University of Ibadan, Department of Botany herbarium (UIH), Obafemi Awolowo University Herbarium (IFE) belonging to the taxa studied were critically examined and compared with their voucher specimens.The information extracted from the herbarium sheets was supplemented by field observation. A base map designed by Agboola (1979) was used for plotting plant distribution in Nigeria. In order to determine the soil requirements for the species studied, an overlay map for the soil types of Nigeria was prepared after Agboola (1979). Flower buds of different sizes were fixed in 1:3:6 acetic acid, chloroform, ethylalcohol for 12 hours and subsequently squashed in 2 % acetocarmine.Permanent slides were made following the techniques outlined by Gill (1969).
Mature pollen grains were squashed in a drop of glycerol acetocarmine and the slides were left in the oven at 60oC for 24 hours. Grains which were filled with stained protoplasm were considered to be fertile while the small shrivelled, unstained or weakly stained ones were considered as sterile. The grains were examined under different microscopic fields and the % pollen fertility was calculated. The seed outlines were drawn by using Willd Heerburg Type 256575 microscope fitted with a drawing tube and the final details of the seed coat were filled in by examining the seed under a dissecting stereoscopic microscope and a magnifying lamp each equipped with fluorescent light.  The terminology used in describing the seed morphology is that of Martin and Barkley (1961).
RESULTS
The taxa studied along with their location voucher number, base number, ploidy level and percentage pollen fertility and size are summarized in Table I. The arrangement of the genera and species is in alphabetical order.

Anisopappus africanus (Hook F.)Oliv.& Hiern  = Telekia africana Hook. F.
A robust herb up to 60 cm high with striate pubescent pithy stems and yellow flower heads with no ray florets. The present haploid count was 14 (Fig.1. 1). Meiosis and pollen formation were normal with 99% filled pollen grain and an average grain size of 26 µm.

Distribution and Ecology
The plant is restricted in distribution to ferruginous tropical soils and eutrophic brown soils of Nigeria (Map. 1) with annual rainfall of 1,016 mm – 1,524 mm.

Achene Morphology
Achene elliptic; rounded at the top; brown; narrowing toward base; 2 mm long; pappus short and barbed; body hairs present (Fig.2. 1).
Exsiccata
FHI 59503, 62490 Cameroon; FHI 74982 Zaria; FHI 53017, 53111, 53119 Ghana; FHI 30211, 41140 Bamenda; FHI 10266 Buea; FHI 28639, 36262 Victoria; FHI 97831 Burundi; FHI 25078 Mt. Koloishe; FHI 25174 Ikwette; FHI 55502 Obudu; FHI 62495 Mambilla; UIH 516 Cameroon; Morton GC 9529; Ejura; A1529, A1529 Technimann-Nkoranza, Morton A1154, Stone 49 Ogoja; Mann 1313, 1920 Cameroon; Omoigui 097 Ikwette Plateau (Obudu).

A.  dalzielii Hutch. = A.chinensis Chev
An erect perennial herb up to 1m high with yellow ray and disc florets in few heads, in rough open grassland. The present haploid count of 14 (Fig.1. 2) is a new count for the species. Meiosis and pollen formation were normal with 100% filled pollen and an average pollen grain size of 26.12 µm.
Distribution and Ecology
This species is common in ferralsols, lithosol and eutophic brown soils of the country (Map 1) with annual rainfall of 1,016 mm – 1,524 mm.
Achene Morphology
Achene cylindric; narrowing towards base; black; round at the top; pappus short nd barbed; body hairs present, 3 mm long (Fig.2. 2).
Exsiccata
FHI 28834 Madaki; FHI 83759 Mando FR.; FHI 43090, 55255, 83666 Zaria; FHI 53162, 94366, 21739 Jos; FHI 25019 Unya (Obudu); FHI 42921 Cameroun; FHI 56667 Sierra Leone; FHI 57684 Oyo; FHI 63612 Kaduna; FHI 49172, 49177 Ghana; FHI 88173 Obudu; FHI 40558 Bamenda; FHI 5976 Ibadan; FHI 46878 Kafanchan; FHI 6833 Agwu (Onitsha) IFE, 1325, 5606, Cameroon; IFE 5618 Cameroon; IFE 945 Chappal- Waddi (Gembu); Omoigui 098 Jos.

Map 1: Distribution map of Anisopappus africanus (•) and A.  dalzielii(î) in Nigeria

Gnaphalium L.
A cosmopolitan genus of 200 species ( Willis and Airyshaw, 1973). In West Africa, there are 3 species. A base number of 7 has been suggested for the genus by Darlington and Wylie (1955).

Gnaphalium indicum L.
A small spreading herb with numerous silvery-wooly flower heads crowded in clusters; pale yellow florets, found on river beds. A haploid count of 7 (Fig.1. 3) was determined. Meiosis was normal.

Distribution and Ecology
The plant is commonly found in lithosol regions of the country (Map 2) with annual rainfall of 1,016 mm – 1,524mm.

Achene Morphology
Achene 1mm long; black; elliptic; arched sideways; pappus soft and delicate; base of pappus encircled by series of semi-transparent pappus scales (Fig.2. 3).

Exsiccata
FHI 63696 Maiduguri; FHI 24342 Tokyo; Omoigui 178 along the bank of River Kaduna; Omoigui 177 along the bank of ABU Dam.

G. luteo-album .L.
An erect or strangling grey-green tomentose herb up to 30 cm tall, found in acidic habitats of sandy stony places. In Nigeria, it is scattered in distribution (Map 2). Its scarcity in the west and southern parts of the country may be due to lack of collection. A haploid count of 8 was determined at metaphase (Fig.1. 4). Meiosis and pollen formation were normal with 93.8% pollen stainability with an average pollen grain size of 19.68 µm.

Achene Morphology
Achenes 1-2 mm long; black; elliptic- oblong; arched sideways; pappus soft and delicate; base of pappus scales by a series of semi-transparent pappus scales; smooth; ending in a blunt base (Fig.2. 4).

Exsiccata
Omoigui 059 Obudu Cattle Ranch; FHI 3694 Maiduguri.

Distribution map of Gnaphalium indicum (•) and G. luteo-album (  )in Nigeria.            Map 2

Helichrysum Mill
A large genus made of 500 species distributed in Southern Europe, tropical and South Africa, Madagascar, South West Asia; Ceylon and Australia with over 200 species found in South Africa only (Willis and Airyshaw, 1973). Many of the species are xerophytes with hairy surface, decurrent leaves. The dried flower heads of some of the species are ornamental. In West Africa, the genus is represented by 14 species, 7 of which occur in Nigeria. Darlington and Wylie (1955) suggested a base number  of 7 for the genus.

Helichrysum albiflorum Mosser
An erect perennial herb up to 30 cm high; stems and leaves pale grey.  A haploid count of 7 was determined for this species (Fig.1. 5) and this is a new report for the genus. Meiosis and pollen formation were normal.

Distribution and Ecology
It is restricted in distribution to lithosols, ferralsols and ferruginous tropical soils of Nigeria (Map3) with annual rainfall of 1,016 mm – 1 524 mm.

Achene Morphology
Achene cylindric; body hairs absent; black 2 mm long; pappus long, thin, minutely pubescent (Fig.2. 5).

Exsiccata
FHI 52872 Victoria; FHI 48261 Cameroon; FHI 28430, 53072 Bamenda; FHI 78340 Bauchi, Shere Mountain; FHI 77604 Ogoja; Omoigui 071 Ikwette Plateau (Obudu).

H. foetidum (L.) Moench
= Gnaphalium foetidum L.
An erect aromatic branched herb with a pithy stem up to 60 cm high; leaves narrow, whitish beneath with bright yellow florets. A haploid count of 7 (Fig.1. 6). Meiosis and pollen formation were normal with 88.4% with an average grain size of 21.56 µm.

Distribution and Ecology
It is of widespread occurrence in alluvial and eutrophic brown soils in the country (Fig. 3) with annual rainfall 1,524 mm – 4 068 mm (Map 3).

Achene Morphology
Achene cylindric; body smooth; brown; pappus long and thin; 3 mm long (Fig.2. 6).

Exsiccata
FHI 25095 Ikwette; FHI 25235 Ogoja; Omoigui 252 Obudu Cattle Ranch; Omoigui 251 Mambilla; IFE 4419 Buea (Equatorial Guinea).
H. globosum Sch. Bip.ex A. Rich.
A perennial herb, whitish, with stiff leaves forming a basal rosette; inflorescence scapose, elongating during flowering with numerous pale brown clustered flower heads. A haploid count of 7 was determined (Fig.1. 7). Meiosis and pollen formation were normal.

Distribution and Ecology
It is of widespread occurrence in alluvial and eutrophic brown soils in the country with annual rainfall 1,524 mm – 4,68 mm.

Achene Morphology
Achene cylindric, body smooth; pappus long and thin; entire achene 3 mm long (Fig.2. 7).

Exsiccata
FHI 93128, 46053, 65548, 63300, 97409 Mambilla; FHI 85097, 10370, 103712 Cameroon; FHI 57352, 3771, 40594 Bamenda; FHI 73744 Ogoja; FHI 66752, 91018 Gembu; IFE 396 Ngel Nyaki; IFE 3083, 2979, 2978, 2970, 1560 Cameroun Mountain; IFE 633 Obudu Cattle Ranch; IFE 7362 Cameroun border; Omoigui 254 Obudu Cattle Ranch;Omoigui 255 Gembu.
Map 3: Distribution map of Helichrysum albiflorum (  ) H. foetidum (•) and H. globosum (î) in Nigeria.

H. mechowianum Klatt.
=  H. hoepfnerianum Vatke
=   H. congolnum Schltr. & O. Hoffm.
A composite perennial herb with flowering scape appearing singly before the leaves almost leafless, usually after fires; high flower heads numerous in a dense terminal cyme yellow tinged red found in grassland. A haploid count of 7 was determined at metaphase stage (Fig.1. 8). This is a new report for the species. Meiosis and pollen formation were normal with 58% filled pollen with an average pollen grain size of 24 µm.

Distribution and Ecology
The plant is restricted indistribution to ferralsols (Map 4) with annual rainfall of 1,016,mm – 1,524 mm.

Achene Morphology
Achene cylindric, body hairs present; base flat; pappus thin and delicate; minutely barbed; 6 mm long; black (Fig.2. 8).

Exsiccata
FHI 48262, 41459, 45144 Mambilla; FHI 4007 Sierra Loene; FHI 10374 Bauchi; FHI 4573 Naraguta; Omoigui 173 Ikwette Plateau (Obudu); Omoigui 218 Gembu.
H. quartinianum Bak.
An annual herb up to 1m high; lemon yellow florets; with silvery hairs all over and the leaves fleshy. This is the first report of its occurrence in Nigeria. A haploid number of 7 was determined at metaphase stage (Fig.1. 9). This is a new report for the species. Meiosis and pollen fertility were normal with 96.3% filled pollen with an average pollen grain size of 32 µm

Distribution and Ecology
It is restricted in distribution to eutrophic brown soils in Nigeria with annual rainfall 1,016 mm – 1,524 mm (Map 4).

Achene Morphology
Achene cylindric; base truncate; body hairs absent; black; pappus long and thin longer than the achene; entire achene 4 mm long (Fig.2. 9).

Exsiccata
FHI 61392, 84550, 84581, 88036 Gembu; FHI 66755, 46074, 65543 Mambilla; FHI 97864 Burundi; Omoigui 178 Mambilla (Gembu).

H. rhodolepis Bak.
An erect perennial herb with simple whitish tomentose stems up to 1 m high; flower heads numerous in a dense terminal cluster, with the inner involucral bracts silvery and the outer reddish; cream coloured florets. The present haploid count of 7 at metaphase (Fig.1. 10) is a new report for the species.

Distribution and Ecology
The plant is restricted in distribution to ferralsols, ferruginous tropical soils to eutrophic brown soils in the country (Map 4) with annual rainfall of 1,016 mm – 1,524 mm.

Achene Morphology
Achene cylindric; body hairs absent; pappus long, thin, minutely barbed; brown; 5 mm long (Fig.2. 10).

Exsiccata
FHI 65266, 81808, 85098 Cameroon; Omoigui 222 Ikwette Plateau; Omoigui 167 Gembu; IFE 6138 Cameroon;  IFE 1055 Vogel (Gembu).
Map 4: Distribution map of Helichrysum  mechowianum(*), H. quartinianum  (î) and H. rhodolepis (•)     in Nigeria.
Inula L.
A large genus composed of 200 species distributed in Europe, Asia and Africa. (Willis and Airyshaw, 1973). Three species occur in West Africa and Nigeria.

Inula klingii O. Hoffm.
An erect perennial herb with stout stem up to 1m high; with lower leaves large, sessile and touching the ground; inner involucral bracts dull yellow; disc florets yellow. The whole plant is dull yellow with a lot of silky hairs. A haploid count of 9 was determined at metaphase stage (Fig.1. 11). This is a new count for the species. Meiosis and pollen fertility were normal with 100% filled pollen and an average pollen grain size of 33.80 µm.

Distribution and Ecology
It is restricted to lithosol regions of the country (Map 5) with annual rainfall of 1,270,mm – 1,524 mm.

AcheneMorphology
Achene elliptic; brown; arched inwards on one side; body hairs present; pappus long and thin, 8 mm long (Fig.2. 11).

Exsiccata
FHI 78734 Zaire; FHI 99025, 56971, 42920 Jos; FHI 80124 Kaduna; Omoigui 096 Jos. 211 Kaura Falls
I. subscaposa S. Moore
An erect perennial herb with slender stems up to 2 m high; arising from a woody stock, ray and disc florets yellow in flower heads. The present haploid count of 9 (Fig.1. 12) is a new report for the species. Meiosis and pollen formation were normal with 75.4% pollen fertility and average pollen grain size of 26 µm.
Distribution and Ecology
It is restricted in distribution to eutrophic brown soils and reddish brown lateritic soils of the country (Map 5) with annual rainfall of 1,016 mm -1,524 mm.

AcheneMorphology
Achene 4 mm long; brown; cylindric; forming a flat base, body hairs present; pappus delicate and long (Fig.2. 12).

Exsiccata
FHI 62868, 63265 Gembu; FHI 44767 Pantisma (Adamawa); FHI 78166 Mambilla; FHI 42818, 63315 Cameroon; Omoigui 172 Gembu.
Map 5: Distribution map of Inula klingii (•) and Inula subscaposa (î) in Nigeria.

Laggera Sch. Bip. ex. Benth
A genus of 20 species distributed in Africa, Arabia, India and South East Asia (Willis and Airyshaw, 1973). Hutchinson and Dalziel (1963) reported 6 species in West Africa, 4 of which were presently investigated. A base number of 10 has been suggested for the genus by Gupta and Gill (1983).

Laggera alata( G. Don)  Sch. Bip. ex Oliv. var. alata
= Blumea olopteraDC
= Erigeron alatus G. Don
A strongly aromatic erect herb up to 2 m high; with florets usually mauve in numerous heads.  A haploid number of 10 was counted at metaphase stage (Fig.1. 13). Meiosis and pollen fertility were normal with 94% pollen stainability and an average pollen grain size of 28 µm.

Distribution and Ecology
It is found in lithosols and eutrophic brown soils of the country (Map 6) with annual rainfall of 1,016 mm – 1,524 mm.

Achene Morphology
Achene cylindric; coarsely ridged; hairy; light brown; pappus double, inner shorter and stiff; pappus with bristles, 9 mm long (Fig.2. 13).

Exsiccata
FHI 51253, 48327, 10388, 10389 Cameroon; FHI 1205 Jos; FHI 39478, 49218, 53123 Ghana; FHI 92852 Mambilla; Omoigui 506 Ikwette Plateau (Obudu).

Laggera aurita (Linn. f).Benth.ex C. B.
= Blumea aurita (Linn. f.) DC
= Conyza aurita Linn. f.
=C. senegalensis Willd.
A green stemmed annual herb, strongly aromatic, densely pubescent with pale yellow florets. A weed of waste places. A haploid count of 10 was determined (Fig.1. 14). Meiosis and pollen fertility were normal with 86.8% filled pollen with an average grain size of 24 µm.

Distribution and Ecology
A common weed of waste and cultivated lands found all over the Northern parts of the country but is restricted to ferralsols and ferruginous tropical soils in the Southern part of the country (Map 6).

Achene Morphology
Achene cylindric; flat; black; body hairs absent; pappus long, thin and barbed; 2 mm (Fig.2. 14).

Exsiccata
FHI 63125 Nsukka; FHI 61275 Kabba; FHI 61866 Ilorin; FHI 61369 Kano; FHI 38301 Borgu (Ilorin); FHI 62043 Adamawa; FHI 103438, 89581 Minna; FHI 24130 Bauchi; FHI 49210, 48736 Ghana; FHI 6212 Onitsha; FHI 3420 Bauchi; FHI 91380 New Bussa; FHI 89104 Mokwa; FHI 93427 Argungun (Sokoto); IFE 1339 IFE Campus; Ife 10789 UI Campus; Lowe 644 Mairabo; 595 Warrah; UIH 1219 Ibadan; IFE 748 Igbetti- Kishi; IFE 1973 Uzairue; FHI 58301 Kainji; Omoigui 257 Jos, 028 Igbetti.

Map 6: Distribution map of Laggera alata (*)and L. aurita () in Nigeria

L. braunii Vatke
An erect perennial herb 60 cm high with large reddish purple, sessile flower heads which are borne in clusters. A weed found in hilly grasslands. A haploid count of 10 was determined (Fig.1. 15).

Distribution and Ecology
A weed restricted to lithosol regions of the country (Map7) with annual rainfall of 1,016 m.

Achene Morphology
Achene 7 mm; flat; elliptic; light brown; body smooth; pappus long, thin and delicate; body curved towards base; base ends in a knob (Fig.2. 15).

Exsiccata
Omoigui 067 Kurra Falls (Jos).
L. gracilis (O. Hoffm. & Musch1.) C. D. Adams
= Blumea alata var. gracilis O. Hoffm.
A non-aromatic glandular herb up to 60 cm long; bushy, with pink florets borne singly in slender branches and peduncles. A haploid count of 10 was determined (Fig.1. 16). This is a new report for the species. Meiosis and pollen formation were normal with 100% filled pollen and an average pollen grain size of 26.1 µm.

Distribution and Ecology

A weed restricted in distribution to lithosols and ferruginous tropical soils of Northern Nigeria (Map 7) with annual rainfall of 1,016 mm – 1,260 mm.
Achene Morphology
Achene 9 mm; oblong-cylindric; body curved inwards at one side; base knobbed; brown; base of pappus encircled with semi-transparent hairs (Fig.2. 16).

Exsiccata
FHI 92400 Liberia; FHI 99017 Jos; FHI 65620, 67265 Borgu (Ilorin); FHI 73634, 43115  Zaria; FHI78857 Kubbani (Zaria); FHI 55109, 80502 Naraguta; FHI 56680 Sierra Leone; FHI42898 Cameroon; FHI 53016,53120 Ghana; FHI 79916 Abuja; FHI 91852 Kaduna; FHI 92087  Ikole; UIH 7472 Yaounde; Omoigui 062 Kurra Falls; 065 Jos.

L. oloptera (DC.) C. D. Adams.                                    = Blumea oloptera DC.                                        = L. oblonga Oliv. & Hiern.                                            = L.marcorrhiza O. Hoffm.  & Muschl
A perennial herb of stony grassland up to 20 cm high; branching near the base from a woody root stock; flowers purple- whitish pink florets. It is restricted in distribution to ferruginous tropical soil (Map 7). A haploid count of 10 (Fig.1. 17) was recorded. This is a new report for the species and it is in line with the base number of 10 suggested for the genus.

Achene Morphology
Achene 1cm long; brown; cylindric; base truncate; pappus long; fine feathery spreading out stiff hairs on both sides of the body (Fig.2. 17).

Exsiccata
Omoigui 076 Igarra Rd (Igarra)

Laggera pterodonta DC.)Sch. Bip ex Oliv. =  Blumea pterodonta DC.
An aromatic glandular weedy herb up to 100 cm high; purplish-pink florets which is restricted in distribution to ferruginous tropical soil and lithosol regions of the country (Map 7). A haploid count of 10 (Fig.1. 18) was determined for the species. Meiosis and pollen formation were normal.

Achene Morphology
Achene 7 mm long; brown; elliptic; arched inwards; tapers to the base; pappus long with 6-8 fine feathery hairs (Fig.2. 18).

Exsiccata
Omoigui 044 Oke-Ibudun (Ondo).
Map 7: Distribution map of Laggera braunii (€ )  L. gracilis (o), L. pterodonta (  ) and L. oloptera ( ) in       Nigeria
Mollera O. Hoffm.
A tropical African genus consisting of 2 species (Willis and Airyshaw, 1973). Hutchinson and Dalziel (1963) reported the occurrence of only 1 species in both West Africa and Nigeria.

Mollera angolensis O. Hoffm.
=  M. punctulata Hiern
An annual herb up to 60 cm high; with yellow ray and disc florets, a weed of fields and open ground. The haploid count of 10 determined for this species at metaphase stage (Fig.1. 19) is a new report for the species and the genus. Meiosis and pollen formation were normal with 98% filled pollen and an average pollen grain size of 26 µm.

Distribution and Ecology
Commonly found in lithosols and eutrophic brown soils of the country (Map 8) with annual rainfall of 1,016 mm -1,524 mm.

Achene Morphology
Achene oblong at the top; narrowing toward the base, marked with lengthwise ridges; brown; body hairs present, 2 mm long (Fig.2. 19).

Exsiccata
FHI 43102, 43093, 46898, 73616, 83661 Zaria; FHI 37205, 40701 Jos; FHI 42756 Vom; FHI 20142, 55261, 84413, 84414 Kaduna; FHI 45044, 56941 Naraguta; FHI83678 Akwanga; Omoigui 124 Jos, 066 Kurra falls.
Map 8: Distribution map of Mollera angolensis (•) in Nigeria

Porphyrostemma Benth. ex Oliv.
A tropical African genus made up of 3 species (Willis and Airyshaw, 1973). In both West Africa and Nigeria, only 1 species was encountered (Hutchinson and Dalziel, 1963). A base number of 15 has been suggested for the genus.
Porphyrostemma chevalieri (O. Hoffm)  Hutch. & Dalz.
=  P.grantii Benth. ex Oliv. var. chevalieri 0. Hoffm.
An annual erect herb from a few centimetres to 30 cm high with pink florets in solitary flower heads, ending in terminal branches.  A haploid number of 15 was determined for this species at metaphase stage (Fig.1. 20). This is a new count for the species. Meiosis and pollen formation were normal with 71% filled pollen and an average pollen grain size of 28 µm.

Distribution and Ecology
It is restricted in distribution to lithosol region of the country (Map 9) with annual rainfall of 762 mm – 1,270 mm (Map 9).

Achene Morphology
Achene oblong-elliptic; black; grooved with lengthwise ridges; body hairs present; pappus thin, delicate and few, and blunt 8 mm long (Fig.2. 20).

Exsiccata
FHI 42886 Yola; FHI 82450 Upper Volta; FHI 21024 Jos; FHI 27204 Vom; FHI 61594, 64709 Gombe; FHI 83552 Zaria; Omoigui 088 Bauchi; Omoigui 214 Jos.
Map 9: Distribution map of Porphyrostemma chevalieri  in Nigeria

Pulicaria Gaertn.
A temperate and warm Eurasian genus made up of 50 – 60 species (Willis and Airyshaw, 1973). It is also found in the tropics and South Africa. Hutchinson and Dalziel (1963) reported the presence of 21 species in both West Africa and Nigeria. Base numbers of 5 and 9 have been suggested for the genus by Darlington and Wylie (1955).
Pulicaria crispa (Forsk.)Oliv.
=Aster crispus Forsk.
An aromatic weed of fields with decumbent branches spreading and ascending up to 30 cm high, stems and leaves silvery; florets light yellow. A haploid number of 10 was counted at metaphase stage (Fig.2. 21). Meiosis and pollen fertility were normal with 59.1% and an average pollen grain size of 21.38 µm.

Distribution and Ecology
This plant is restricted in distribution to ferruginous tropical soils and lithosol zone of the country (Map 10) with annual rainfall of 508 mm – 1,016 mm.

Achene Morphology
Achene 2 mm long; brown; oblong-elliptic; pappus thin anddelicate, no body hairs (Fig.2.21).

Exsiccata
FHI 101924, 101925, 59744 Zaria; FHI 45775 Numan; FHI 10407, 63454 Bauchi; FHI 49895 Katagum; FHI 5296,48737,49204, 48629 Ghana; FHI 37314 Niger; FHI 25928 Giwa (Zaria); FHI 63682 Maiduguri; FHI 64645 Yola; FHI 21136 Kano; Omoigui 141 Bauchi 141 Bauchi; Omoigui 210 Zaria; Omoigui 250 Igboho.
P.undulata (Linn.)C.A. Mey.
=  Inula undulata Linn.
=  P. undulata var. alveolosa (Blatt. & Trab) Maire.
An aromatic weed of fields with decumbent branches, spreading and ascending but much smaller than P. crispa and more hairy; florets yellow. A haploid count of 10 was determined at metaphase stage (Fig.1. 22). This is a new report for the species. Meiosis and pollen fertility were normal with 100% filled pollen with an average pollen grain size of 20.64 µm.

Achene Morphology
Achene 3 mm long; brown elliptic; pappus thin, body hairs absent (Fig.2. 22).

Exsiccata
FHI 75629 Cameroon; FHI 52117, 48491, 53177, 53175 Ghana; Omoigui 073 Sokoto; Moiser 186, 214, 230, 238 Katagum; Dalz.314 Yola; Dalz.34 Borno.
Map 10: Distribution map of Pulicaria crispa (•) and  P. undulata (o) in Nigeria.

Sphaeranthus L.
A genus made up of 40 species, distributed in Africa, Madagascar, Iraq, India, South East Asia and North East Australia (Willis and Airyshaw, 1973). In West Africa and Nigeria, 14 species are known to occur  (Hutchinson and Dalziel, 1963). A base number of 10 has been suggested for the genus by Shetty (1961).

Sphaeranthus angustifolius DC
=  S.nubicus Sch. Bip. ex Oliv. And Hiern
=   S. lelyi Robyns.
A grandular pubescent aromatic erect weed found in cultivated fields, up to 60 cm high; florets red in solitary heads. A haploid count of 10 was determined at metaphase stage (Fig.1. 23) for this species. This is a new report for the species. Meiosis and pollen formation were normal with 8% filled pollen and an average pollen gram size of 26 mm.
Distribution and Ecology
The plant grows in lithosol, semi-arid brown soils of Nigeria (Map11) with annual rainfall of 508 mm – 1,270 mm.

Achene Morphology
Achene cylindric; arched inwards; narrowing toward base; body hairs absent; no pappus, 2 mm; black (Fig.2. 23).

Exsiccata
FHI 23172 Bauchi; FHI 48492, 53010 Ghana; FHI 48126 Katsina; FHI 17662 Kaduna; FHI 80332 Gombe; Omoigui 080 Bauchi; Omoigui 225 Sokoto; Omoigui 223 Kano.

S. flexuosus O. Hoffm.
=  S. brounae Robyns
A much branched annual herb up to 60 cm high; florets purple in triangular heads; an aromatic weed of cultivated fields. The present haploid count of 10 (Fig.1. 24) is new for this species. Meiosis and pollen formation were normal with 99.4% filled pollen and an average pollen grain size of 21.68 µm.

Distribution and Ecology
The plant is restricted in distribution to lithosol regions of the country (Fig. 11) with annual rainfall of  762 mm – 1,270 mm.

Achene Morphology
Achene ovate; narrowing towards base; body pappus absent; 1 mm long; black (Fig.2. 24).

Exsiccata
FHI 69546, 38995 Cameroun; FHI 65472, 10439 Bauchi 081 Yelwa (Bauchi); Omoigui 224 Kano State.

S. senegalensis DC
=    S. lecomteanus O. Hoffm. & Musch1.
A creeping annual herb, half decumbent, weeds of cultivated fields, hairy leaves, florets reddish purple. The haploid count of 10 agrees with the base number of 10 (Fig.1. 25). This is a new report for the species. Meiosis and pollen fertility were normal with 91% filled pollen and an average pollen grain size of 20 µm.

Distribution and Ecology
The plant is restricted to lithosols, ferralsol and ferruginous tropical soils of Northern Nigeria (Map 11) with annual rainfall of 508 mm – 1,016 mm.

Achene Morphology
Achene ovate; narrowing towards base to form a knob; body hairs present; pappus absent; black 1mm long (Fig.2. 25).

Exsiccata                                                    FHI 49185, 49231 Ghana; FHI 64847 Zaria; FHI 36423 Samaru; FHI 65420 Bauchi; FHI 59745 Kajuri (Zaria); FHI 41991 Gurmi (Sokoto); FHI 33006 Hadejia (Kano); FHI 17921 Gulumba (Borno); Omoigui 171 Zaria; Omoigui 226 Shika (Zaria).
Map 11: Distribution map of Sphaeranthus angustifolius (o), S. flexuosus () and S. senegalensis (•) in     Nigeria.

Vicoa Cass.
A small genus made up of 12 species distributed in Northern Europe, tropical Africa to Central Asia (Willis and Airyshaw, 1973). In West Africa and Nigeria, only 1 species was encountered. A base number of 9 has been suggested for the genus by Federov (1969).

Vicoa leptoclada (Webb) Dandy
=   Inula leptoclada Webb.
An erect annual weed up to 1m high; found in cultivated lands; leaves dark green above, pale  beneath, florets yellow. A haploid count of 9 was determined at metaphase stage (Fig.1. 26). This is a new report for the species. Meiosis and pollen formation were normal with 84% filled pollen and an average pollen grain size of 26 µm.

Distribution and Ecology
A well distributed weed found in all parts of the country (Map 12) ranging from alluvial soil to eutrophic brown soils with annual rainfall of 762 mm -1,524 mm.

Achene Morphology
Achene oblong-cylindric; brown; marked with lengthwise ridges; rounded at the base; pappus long and delicate, body hairy, 6 mm long (Fig.2. 26).

Exsiccata
FHI 63492, 77282 Yola GRA; FHI 65454, 78294 Gombe; FHI 29981 Gurara Falls (Abuja); FHI 72896, 94436 Zaria; FHI 30239, 19823, 81020 Ilorin; Ago- Are; FHI 98384 Gombe- Yola Road; FHI 5542 Biu; FHI 41874 Jebba; FHI 40286 Arakanga F. R. (Abeokuta); FHI 65613 Borgu; FHI 25636, 98797 Oyo; FHI 42910 Gurumi (Adamawa); FHI 49898 Katagum; FHI 6116 Lafia; FHI 43148 Samaru; FHI 10516 Bauchi Plateau; FHI 59675
Kaduna; FHI 46956 Bam-Ngelzarma (Borno); FHI 43628 Yankari Game Reserve; FHI 45357 Kainji Dam; 60485 Jalingo; FHI 63754, 27970 Mokwa; FHI 61298 Kabba; FHI 57078 Kablama F. R. (Zaria); FHI 63005 Minna; FHI 62269 Egbe; IFE 1470 Kadawa (Kano); IFE 047 Zaria; IFE 577 Yelwa (Sokoto); IFE 296 New Bussa; Omoigui 153 Yelwa (Bauchi); Omoigui 155 Igbetti; Omoigui 154 Kaduna.
Map 12: Distribution map of Vicoa leptoclada (•) in Nigeria
Discussion
Chromosome Number
Chromosome numbers  have been widely used in systematic investigations of the Asteraceae by Turner and King (1964), King et al. (1976), Solbrig (1977), Gill et al.(1979), Jansen and Stuessy (1980) and Gupta and Gill (1986). The information on chromosome number when used in conjunction with other taxonomic data tends to provide a better understanding of the tribal and generic relationship within the family.Olorode (1974) reported the range of chromosome numbers in Nigerian Asteraceae to be from n=5 in Emilia coccinea to n=36 in Bidens pilosa while Omoigui (1985) reported the range of chromosome numbers of the Southern Nigerian Asteraceae to be n=5 (Emilia coccinea) to n= 48 (Bidens pilosa).  From the results of this study, the chromosome numbers of the tribe Inuleae range from n=7 in Helichysum mechowanium, H.  albiflorum and Gnaphalium indicum to n=15 in Porphyrostemma chevalieri. This range differs widely from the Indian taxa (another tropical flora) with a range of 2n=6 (Pterotheca falconeri) to 2n=120 (Eupatorium odorantum).From this study, the report of n =14 (Anisopappus africanus) agrees with the earlier report of Auquier and Rennard (1975) (2n=28) for the species. The haploid count of 7 for Gnaphalium indicum agrees with the other workers (Gupta and Gill, 1981, Mehra and Remanandan, 1975). Earlier, Banerjee and Sharma (1974), Chatterjee and Sharma (1968) and Mehra and Remanandan (1975) reported a tetraploid 2n = 28 for the species. For G. luteo-album, the haploid count of 8 reported for the species differed from earlier reports of n = 7 (Federov, 1969; Singh, 1972;Turner,1977) and n=14 (Larsen, 1966). Thus the genus Gnaphalium is dibasic with x = 7, 8.
For the genus Helichrysum, the report of n=7 for H. albiflorum confirms the reports ofTurner and Lewis (1965), Hedberg and Hedberg (1977), Mejias and Luque (1987) and Subramanian (1980). For H. foetidum and H. globosum the report agrees with the report of Hedberg and Hedberg (1977) and Thulin (1970) with n=7.         The count of n = 10 for Laggera alata var. ialata agrees with the report of Gupta and Gill (1983), while n = 10 for L. aurita agrees with Mehra et al. (1965), Matthew and Matthew (1975), Bir and Sidhu (1979), Sidhu (1979),Gupta and Gill (1986), Nirmala and Rao (1981) and  Subramanian (1980). Earlier on, Gupta and Gill (1986) reported the presence of B chromosome in the species.The report of n = 10 for Laggera pterodonta confirms the report of Matthew and Matthew (1975, 1979)andBir and Sidhu (1979). Meanwhile, Mehra et al. (1965) had earlier on reported a cytotype of n=9 for the species.The count of n=9 for the genus Inula agrees with the suggestion of Darlington and Wylie (1955) while x=10 for Spharenthus agrees with the report of Shetty (1961).
According to Gupta and Gill (1986), polyploidy and aneuploidy have led to a phylogenetic increase or decrease at diploid level and this seems to have added to the numerical diversity in chromosome of the Indian taxa of the inuleae. This observation also applies to the Nigerian taxa of the Inuleae.

Basic Chromosome Number
A perusal of the cytological literature revealed that the proposals for the basic numbers of many genera are based primarily on the lowest known gametic number. For genera with high gametic number, it has often been the practice to consider it a multiple of low number and which might no longer exist.
According to the systematic revision of the group Inulineae (sensu amplo) by Merxmuller et al., (1977), the primary basic chromosome number has been suggested to be x =10.
From this study, there is the prevalence of x=7 (7 species), x=9 (3 species) and x=10 (11 species). The results from this study agree with the results of the Indian Inuleae (Gupta and Gill, 1986), where x= 10 is considered the ancestral basic number of the tribe Inuleae and x= 9 which is found in Inula and Vicoa might have been derived through reduction from x= 10 and the higher basic number of x=15 in Porphyrostemma must have arisen from a multiple of low numbers which might no longer exist. According to Gupta and Gill (1981), if adequate chromosomal data are available for a genus, it becomes easier to decide its basic number.
Distribution Maps
According to Gill (1981), the study of geographical distribution of plants which is based on critical examination of herbarium specimens and the relevant recorded data is of great interest to phytogeographers, taxonomists and geobotanists as this helps in tracing the evolutionary history of the flora or a taxon of a particular region. The plasticity and habitat requirements of a taxon are best judged by the extent of its distribution. From the distribution maps of the plants (Maps 1-12) studied, the plants were found to be restricted to higher altitudes of the country (11 species) that is Gnaphalium luteo-album,Helichrysum albiflorum, H. foetidum, H. globosum, H. mechowianum, H. rhodolepis, H. quartinianum, Inula klingii, I. subscaposa, Laggera alata var. alata, L braunii and L. gracilis and the others to Northern Nigeria (savanna regions of the country). They are Gnaphalium indicum, Mollera angolensis, Porphyrostemmma chevalieri, Pulicaria crispa, P. undulata, Sphaeranthus angustifolius, S. flexuosus and S. senegalensis). Vicoa leptoclada and Laggera pterodonta  have a widespread distribution.

Achene Morphology
The fruits or achenes of members of this family are crowned by a pappus composed of numerous bristly hairs. The pappus hairs which are either uniform or variable in length in different species have been observed by many workers (Keay et al.,1964; Faust, 1972; Jones, 1973, 1976; Isawumi, 1982, 1999; Ayodele, 1987, 1992) to be a good morphological character for taxonomic evaluation of the family. The result of this study on the shape and structure of the achene agrees with the report of Ayodele (1992) that the pappus hairs showed projections of various shapes on superficially smooth-looking surfaces.
Heywood et al.(1977) noted a correlation between the pappus structure and growth habit of some members of the family. They observed that the herbaceous members of the family produced large size fruits and the entire length of the pappus hairs had more projections than the shrubby species with their scanty or totally smooth pappus (Fig.2. 1-26). The result of this study does not support this correlation between growth habit and the pappus structure. All the plants studied were herbaceous in nature. The variation of the pappus structure could rather be an efficient means of dispersal of the achenes by wind.

Conclusion
For any meaningful phylogenetic inferences or reclassification to be made about this tribe in Nigeria, the other remaining plants in the tribe will have to be investigated.

Acknowledgements
The author acknowledges the efforts of the Late Prof. Dr. L. S. Gill, who supervised this work (1985-1990) and Prof. Omotoye Olorode for the use of his laboratory at the Obafemi Awolowo University, Ile-Ife (1990-1992).
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CYTOMORPHOLOGICAL STUDIES OF SOME NIGERIAN ASTERACEAE-TRIBE VERNONIEAE

Department of Plant Biology and  Biotechnology, University of  Benin, Benin City,  Nigeria.

Received 2nd April, 2012; accepted  23rd July, 2012

ABSTRACT
Cytomorphological studies of 59 taxa distributed in 34 species, 1 subspecies, 1 variety and 6 genera of the tribe Vernonieae have been studied. Pollen fertility test of the species studied revealed that meiosis was normal with regular bivalent pairing. Chromosome counts of 11 species, 1 subspecies and 1 variety  (Gutenbergia nigritiana n=10, G. rueppellii n=10, Vernonia calvoana subsp calvoana n=10, V. camporumn=10, V. gerberiformis n=10, V. guineensis  var. cameroonica n=10, V. iodocalyx n=9, V. nigritiana n=9,  V. phillipsoniana n=9, V.richardianan=10, V. saussureoides n=9, V. stenocephala n=9, V. subaphyllan=9 ). The chromosome number of V. tenoreana (n=9) differs from previous reports. This has revealed the first cytotype in the tribe and the count of n=11 for V. migeodiisuggest a new basic number of 11 for the genus, which is reported for the first time from the Nigerian flora. The information from chromosome  and pollen grain size did not support the change of genera for the affected species. The achene morphology of all the taxa studied has been drawn and described for the first time. Distribution maps of all the taxa studied are presented for the first time and they revealed that members of this tribe are prevalent in higher altitudes. There was no correlation between chromosome number and the annual habit nor between achene morphology and growth habit of members of the tribe.The occurrence of Centratherum punctatum Cass.is reported for the first time from Nigeria.

Keywords: Chromosome number, Pollen size, Achene morphology, Vernonieae

INTRODUCTION
The family Asteraceae is one of the largest families among the dicotyledonous plants comprising of about 1,302 genera and 22,000 species (Turner, 1977) and its members enjoy a cosmopolitan distribution with striking concentration in the temperate regions. Although Bentham’s classification of the family (Bentham, 1873a), has long been accepted as a comprehensive system and the family is monophyletic, there are some modifications to this. The phyletic relationships of various tribes have been proposed by more recent systematists (Cronquist, 1955, 1977; Wagenitz, 1976; Heywood et al.,1977).
All the early workers in plant classification recognized the Compositae as a group at some level (Tournefort, 1700; Berkley; Vaillant,1723 and Le franc, 1760 ) and in every type of analysis the family was monophyletic (Small, 1919; Bremer, 1987; Jansen and Palmer, 1987; Hansen, 1991a, 1991b; Michaels et al., 1993; Lundberg and Bremer, 2003). This is possibly because morphologically the family is well characterized; flowers (florets) arranged on a receptacle in centripetal heads and surrounded by bracts, anther thecae fused at the margins to form a ring but the filaments are  free pollen pushed out by the style, calyx (when present) developed into a pappus and the fruit is an achene (Cypsela).
Family-wide treatments are few; Bremer’s (1994) cladistic analysis was the first revision of the whole family. According to Funk (2010), the morphology did not generate enough data to resolve many of the issues which bothered on the circumscription of the genera into sub-families on one hand and the number of monophyletic tribes and the relationship amongst the recognized tribes on the other hand as well as the circumscription of the genera into tribes.                                    Cassini (1816) and Bentham (1873b) placed the Heliantheae at the centre, Vernonieae and Eupatorieae at one end and the Mutisieae and Cichorieae at the other end. Tribes such as Cardueae, Cichorieae, Senecioneae, Astereae and Anthemideae were pretty much the same as they were reported by Cassini (1816) and Heywood et al. (1977). They thought Heliantheae was the most primitive tribe of the family and assumed that the ancestors were perennial herbs or shrubs. Kadereit and Jeffery (2007) and their numerous co-workers reordered the genus, tribe and subfamilies within the family based on recent morphology and molecular  results and this work is now regarded as the standard reference for descriptions of the tribes and genera of the family. There are now 42-43 tribes in the family instead of the traditional 12-13 tribes.
Using morphology and molecular results, Funk et al. (2009) have agreed with the placement of the tribe Vernonieae but the problem of the family remains unresolved. This confusion is highly evident in the tribe Vernonieae. Members of the tribe have leaves which are commonly alternate; heads discoid, florets never yellow; involucral bracts imbricated in several series or rarely, few 1-2 series; corolla tubular and regular anthers not tailed at the base; style branches slender; filiform or alternating e.g., Elephantopus, Scolymus, Vernonia. The tribe is predominantly tropical and the members are mostly herbs and shrubs with homogenous heads.
The tribe Vernonieae is represented worldwide by 1,456 species in 70 genera, of which 34 are monotypic and the type genus Vernonia comprising about 1,000 species (Jones, 1977) while Turner (1977), Bremer (1994), Nordenstam (1998) and Isawumi (1996) recognized 98 genera and 1,300 species. However, Robinson (1996) recognized 111 genera worldwide for the tribe. Recently, Robinson (2007) recognized 118 genera distributed in 15 sub-tribes. According to Isawumi (1999), new tribes and genera or resurrected genera are being established for the tribe, especially in the Eastern hemisphere.  Oliver and Hiern (1877) reported 78 species for members of the tribe in Africa while Jones (1977) has identified over 200 species for the tribe. Recently, Isawumi (2008) recognized 35 genera and 299 species for the African Vernonieae with the type genus Vernonia having 59 species. The first comprehensive work on this tribe in West Africa was by Adams (1963) who recognized 10 genera and 74 species. Isawumi (1996, 1996a) recognized 15 genera and 42 species for the West African Vernonieae. In Nigeria, 37 species have been reported by Hutchinson and Dalziel (1963).
Using morphology, Robinson (1990) resurrected the genus Baccharoides and transferred three species of Vernonia to the genus; Isawumi (1993) transferred 12 species of Vernonia to the same genus on the basis of the style base being surrounded by the nectar and inner involucral bracts with expanded foliose appendages which may be white or variously coloured. Isawumi et al. (1996) recognised 25 species and 26 infraspecific taxa in the genus Baccharoides on the basis of morphological characters such as the endothecal tissue being polarized (Dormer, 1962). Ayodele and Olorode (2005) have cautioned on these unnecessary changes based on limited characters. They believed that these characters could be an adaptation strategy of the different species. Isawumi (2008) noted, however, that the separation of Baccaroides from Vernonia s. i. by Robinson (1990) and Isawumi et al. (1996) could not be sustained mainly on adaptation strategies alone because the group was resurrected as a genus based on many characters. The genus Vernonia could have gradually reduced in heterogeneity and size and eventually become a monophyletic genus. Consequently, many species are being separated from Vernonia s. i. in the neotropical and paleotropical regions of the world as seen in the works of Robinson (1996, 1999, 2007) and Isawumi (2008) and this has affected the Nigerian flora with change of names. For example, Vernonia gerberiformis is now known as Linzia gerberiformis, Vernonia nigritiana is now known as L. nigritiana, Vernonia ituriensis var. occidentalis is now known as L. ituriensis var. occidentalis, Vernonia migeodii is now known as Vernonia strummigeodii, Vernonia bamendae is now known as Orbivestus bamendae and Vernonia  cinerea  is known as Cyanthillium cinereum. Drury and Watson (1966) have cautioned on these changes, and that taxonomy should strive to be a source of useful generalizations as it provides a springboard for future research. Good classifications in fact are those which prove to have a high predictive value. They are not likely to be achieved unless taxonomists try to take account of all available comparative observations. This means that it is illogical to apply only a few so-called technical characters that are available until all other details about the species are considered.  Chromosome numbers have been widely used in systematic investigations of the Asteraceae (Turner and King, 1964;King  et al., 1976; Solbrig, 1977; Turner, 1977; Gill et al., 1979; Jansen and Stuessy, 1980; Gupta and Gill, 1983). These information on chromosome numbers when used in conjunction with other taxonomic data tend to provide a better understanding of the tribal and generic relationships within the family.        Despite the usefulness of chromosome data in tackling problems of systematic relationships, phylogeny and evolution of related plant groups, our present knowledge on the cytology of many of the tribes of the tropical Compositae is inadequate and scattered. The bulk of the data is confined to the chromosome number reports on taxa from the temperate regions (Matthew and Matthew, 1983). Cytologically, this is the least known tribe in the family with a little more than 100 species so far examined for their chromosome numbers (Matthew and Matthew, 1983). In Nigeria, this is the only tribe that has been extensively studied by Isawumi (1982), Ayodele (1987,1992, 1995, 1997, 1999,2000) and Omoigui (1985, 1992). The cytology of 34 species, 1 subspecies belonging to 6 genera (Centratherum, Elephantopus, Ethulia, Gutenbergia, Triplotaxis and Vernonia) have been investigated.
For the purpose of this work, I have followed Smith’s (1977) traditional classification of the family in which he divided the family into 12-13 tribes. The aim of this work is to use the information from chromosome data as a reliable character to verify these re-classifications.

Materials and Methods
The plant materials used in this study were collected from different parts of the country and the specific locations where they have been collected have been given.The herbarium sheets deposited at the national herbarium belonging to the taxa studied were critically examined and compared with the voucher specimens. Information from literature, herbarium and field survey were compiled and used to plot the distribution maps.
For plotting the distribution maps, a base map designed by Agboola (1979) was used in order to determine the requirements for the soil types of Nigeria which was prepared after Agboola (1979). Meiotic chromosomes were studied from microsporocytes. Young flower buds were fixed in 1:3:6 acetic alcohol chloroform. The fixed material was transferred to 70% ethyl alcohol after 24 hours and subsequently squashed in 2 % acetocarmine. Preparation of slides was made following the technique of Gill (1969) to determine the pollen fertility. Mature anthers from unopened anther buds were squashed in 2 %  acetocarmine glycerol and left overnight in an oven at 60OC. Pollen which did not take up stain and were irregularly shaped were taken as sterile. About 500 pollens were examined for each taxa and the % fertility was calculated.
Morphology of the achene was drawn by using Willd Heerburg Type 256575 microscope fitted with drawing tube and the final microscope details of the seed coat were filled by examining the seed under a dissecting microscope. The terminology used for describing the achene morphology was that of Martin and Barkley (1961).
Results
The exact localities, their chromosome numbers, base number, ploidy level, pollen fertility and pollen size of some Nigerian Asteraceae (Tribe Vernonieae) are summarized in Table I.
Centratherum  Cass.
A genus of 20 tropical species, represented by one species in West Africa. Base numbers of 8 and 9 have been suggested for the genus.

Centratherum punctatum Cass.
A tropical genus made up of 20 species (Willis and Airy-shaw, 1973). An occasional garden plant throughout and now locally established in wet places. A haploid count of 16 was determined at Metaphase 1 (Fig.1. 1 ). Meiosis and pollen formation were normal with 92% filled pollen and  an average pollen grain size of 32.41 µm.

Achene Morphology
Achene 1-2 mm long; black; elliptic; cylindric; narrowed towards the base; rough marked with lengthwise ridges; pappus absent (Fig.2. 1).

Exsicatta
Omoigui 012 Uniben; Omoigui 015 School of Forestry, Jos.

Elephantopus L.
A tropical genus made up of 32 species (Willis and Airyshaw, 1973). In West Africa, the genus is represented by 3 species and all species are encountered in Nigeria (Hutchinson and Dalziel, 1963). A base number of 11 has been suggested for the genus by Baldwin and Speese (1955).

Elephantopus mollis Kunth
A tap rooted, erect branched coarsely hairy perennial herb with wrinkled leaves usually up to 50 cm high. It has a wide distribution range. It can be found in ferralsols, lithosols, ferruginous tropical soil, alluvial and eutrophic brown soils of Nigeria with annual rainfall of 1,016 mm – 3,048 mm (Map 1).
A haploid number of 11 was counted during metaphase (Fig.1. 2). Meiosis and pollen formation were normal with 98% filled pollen and an average pollen grain size of 40 µm.

Achene Morphology
Achene flat; elliptic – cylindric; narrowing toward base, black, with lengthwise ridges, body hairs present, pappus double hairs; thin few and delicate, with the secondary hairs more permanent (Fig.2. 2).

Exsiccata
FHI 79176 Abuja; FHI 80128 Nimbia (Kaduna); FHI 77067, 77764 Mambilla; FHI 63599 Minna; FHI 49208, 7554, 41630, 30549 Cameroon; FHI 15413, 36408, 65703, 30158 Calabar, FHI 55417, 21716 Jemaa; FHI 49211, 49198, 49303, 53000 Ghana; FHI 52453, 10325, 42840, 82924, 10324, 10323; 35463, 290, 4409 Cameroon; FHI 72447 Nsukka; FHI 53658 Gabon; 40035, 7343, Owo; FHI 6514, 59450 Zaria; FHI 34937 Oshun (Oyo); FHI 98328 Burundi; FHI 96395 Saki; FHI 6514, 70546 Oban; FHI 92857 Serti; FHI 67189 Igala (Kwara); FHI 4820 Umuahia; FHI 82809 Mokwa (Niger); FHI 50882 Liberia; FHI 56023 Abak; FHI 62337 Sarduna; IFE 1374, 1315 Iseyin; IFE 1371 Kumba (Cameroon); IFE 4 O.A.U. Campus Hall 2, IFE 242 Erin – Odo Hills (Oyo); Omoigui 271 Obudu Cattle Ranch; Omoigui 274 Kurra Falls; Omoigui 145 Obudu.
Elephantopus spicatus B. Juss.ex Aubl.
=   Pseudelephantopusspicatus B. Juss. Ex Aubl.
A tap rooted weed of stony roadside; erect, stiff much branched herb up to 45 cm high with the leaves at the top, spiny with small white florets in oblong heads. It is restricted in distribution to alluvial and lithosol regions of the country with annual rainfall of 1,270 mm – 1,524 mm.  (Map1). Outside Nigeria, it extends southwards up to Sierra Leone.
A haploid count of 11 (fig.1. 3) was determined. Meiosis and pollen formation were normal with 90% filled pollen and an average grain size of 28 µm.

Achene Morphology
Achene flat, elliptic, narrowing toward the base; brown; marked with lengthwise ridges; pappus forming 2 – 4 hairs in each awn; 5.5 mm long; body hairs absent (Fig.2. 3).

Exsiccata
FHI 56641, 4701, 4000, 4061 Sierra Leone; FHI 68372 Lagos; 60584, 70313 Ijebu Ode; FHI 77526 Ibadan; IFE 1377, 1316 Lagoon (Ikorodu); Omoigui 148 Kurra Falls.

 

Ethulia Linn. f
A tropical American and African genus made up of 10 species (Willis and Airyshaw, 1973). In west Africa and Nigeria, only 1species is found (Hutchinson and Dalziel, 1963).
E. conyzoides Linn.f.
An erect branched aromatic herb 60 cm high, with numerous small flower heads with reddish florets; a weed found in wet places. A very common weed which grows in all types of soils in the country (Map 2) with annual rainfall of 508 mm – 3,048 mm. Outside Nigeria, it extends up to Senegal.
A haploid count of 20 was determined at metaphase (Fig.1. 4). Meiosis and pollen formation were normal with 96% filled pollen and an average pollen grain size of 36.24 µm.
Achene Morphology  Achene 2 mm long; brown; deeply grooved by lengthwise ridges; ovate cylindric; narrowing towards base and ending with a small notch (Fig.2. 4).
Exsiccata FHI 88669 Akamkpa (C.R.S); FHI 65511 Yola; FHI 65163 Onitsha; FHI 65465, 55350 Gombe; FHI 46425 Ihiala; FHI 65624 Borgu; FHI 77143, 77375 Mambilla; FHI 96922, 99542, 99518 Umuahia; FHI 448871, 94888 Epe; FHI 94850 94846 Egbado; FHI 77375, 89143 FHI 89135 Itu; FHI 70399 Kwara; FHI 41094, 30385, 30696 Cameroon; FHI 19439 Ibadan; FHI 67616 Enugu; FHI 101507 Kaduna; FHI 49593 Katagum; FHI 53036 Akure; FHI 40746 Jos; FHI 47187, 87596 Pankshin; FHI 38313 River Osse; FHI 40746 Jos; FHI 47187, 87596 FHI 48762, 53009 Ghana; FHI 50475 Ivory Coast, FHI 41992 Sokoto; FHI 48258 R. Ogun; FHI 78206 Mubi; FHI 4564 Ogoja; FHI 39815, 79826 Mokwa (Niger); FHI 87653 Kwale; FHI 69380 Bauchi; FHI 39929 Calabar; FHI 26370 Ibadan; FHI 91243 Obudu; FHI 43945 Okitipupa; FHI 5544 Borno; FHI 69389, 34335 Bauchi; FHI 54297, 59331 Kaduna; FHI 18516, 94910 Ede; FHI 83963 Funtua; FHI 93027 Ikom; FHI 95454 Oguta; FHI 58553 Ijebu; FHI 54297, 79527 Kaduna; FHI 63607, 63608 Minna; 6299 Benue; FHI 87970 Gembu; FHI 89234 Nsukka; FHI 94335 Bauchi; FHI 8118 Ilorin; FHI 61299 Kabba; FHI 99144 Kano; Omoigui 149 Jos; Omoigui 290 Ibadan; Omoigui 293 – Bank of River Ovia (Lagos Road). Gutenbergia  Sch. Bip
A tropical African genus made up of 20 species (Willis and Airyshaw, 1973). There are 4 species in West Africa out of which 3 are found in Nigeria.
G. nigritiana.(Benth.)Oliv.& Hiern = G. nigritanum.Benth.
An erect weedy herb up to 60 cm high with puberulous striate branches; florets purple which is commonly found in lithosols, ferralsols and ferruginous tropical soil regions of the country (Map 3) with annual rainfall of 762 mm – 1,524 mm.Outside Nigeria, it extends to Guinea.
A haploid count of 10 was determined for the species (Fig.1. 5). Meiosis and pollen formations were normal with 75% filled pollen and an average grain size of 50 mm. Achene Morphology Achene oblong – linear; brown; upper end truncate; cup-shaped; body smooth; pappus absent; sides usually with fine lengthwise ridges 2 mm – 3 mm long (Fig.2. 5).
Exsiccata FHI 46897, 56885, 25978, 57038 Zaria; FHI 53744, 7871, 65596, 16219, 57306 Olokemeji; FHI 103429 Oturkpo; FHI 89755 Kajola (Oyo); FHI 58866, 86964, 50734, 63165, 66598 Enugu; FHI 84853 Imeko (Ogun); FHI 100643 Jos; FHI 60214, 87065 Ogoja; FHI 25900 Giwa (Zaria); FHI 99444, 89274 Obudu; FHI 66916 Uzebba; FHI 4691, 5330 Sierra Leone; FHI 38460, 32264 Gwari (Borno); FHI 23633 Uzariaue; FHI 52213 Abeokuta; FHI 8924, 89224, 73131 Nsukka; FHI 91909 Ikole (Ondo); FHI 88302 Okenne, FHI 90229 Ado – Ekiti, FHI Ibadan; FHI 88390 Kabba; FHI 935528 Ifon (Ondo), FHI 86504, 102664 Oka (Ondo);  FHI 51755 Ghana; FHI 40064 Owo; FHI 21862 Jemma; FHI 44715 Gurara (Niger); FHI 20094 Riyom; FHI 66516 Keffi; FHI 7031 Kagoro, FHI 85274 Kishi (Oyo); FHI 47358 Jos. FHI 70027 Igala; FHI 99041 Abuja; FHI 48104 Onitsha; FHI 67280 Egba; FHI 68076 Egbado; FHI 19278 Osi (Ilorin), FHI 92095 Gongola; Omoigui 074 Kaurra falls; Omoigui 30 Igbetti; Omoigui; 027, Kaduna State.
Gutenbergia rueppellii (Sch. Bip) =Ethlulia rueppellii (Sch. Bip)
An erect branched annual herb up to 30 cm high with the leaves whitish below, florets purple. It is a weed of cultivated fields of lithosol regions of Nigeria (Map3). Two populations were studied and they both had a haploid number of 10 (Fig.1. 6). Meiosis and pollen formation were normal with 94% filled pollen and an average pollen grain size of 56 mm.
Achene Morphology Achene  flat; oblong; narrowed downwards to a truncate base; top cup-shaped; body smooth; no pappus; light brown; marked with lengthwise ridges, 0.5 mm – 1 mm long (Fig 2.6).

Exsiccata

FHI 84961 Zaria; FHI 91189 Jos; FHI 94809 Wusasa; FHI 55257 Kaduna; FHI 55047 Kujama; Omoigui; 156 Bauchi; Omoigui; 106 Jos.

 

Triplotaxis  Hutch
A tropical African genus of 3 species (Willis and Airyshaw, 1973) represented by one species in West Africa. A base number of 8 has been suggested for the genus by Mangenot and Mangenot (1962).
Triplotaxis stellulifera (Benth.)Hutch.
= Herderia stellulifera Benth
An erect or procumbent annual herb, up to 60 cm high with pinkish-purple florets. It is commonly found as a weed of roadside and clearings of lowlands in south-western part of the country. It is restricted in distribution to ferralsol regions of Nigeria (Map 2). Eight bivalents were counted at Metaphase I (Fig.1. 7).Meiosis and pollen formation were normal with 95.4% filled pollen and an average pollen grain size of 24.93 µm.

Achene Morphology
Achene 2-3 mm long; brown; elliptic; body hairs present (Fig. 2. 7).

Exsiccata
FHI 12309 Sakponba (Benin), FHI 7141,FHI320131 Ubiaja Forest Reserve (Benin); FHI 33257 Okomu Forest Reserve (Benin); FHI 8776, 9151 Brenan (Okomu); Omoigui 048 Benin (Iyanomo);FHI 74834, 66127 Umuahia; FHI 33257 Ubiaja; FHI 72305, 32013, Benin; FHI 39279, 95265 Sapoba; FHI 6139 Okigwe; FHI 95708 Akamkpa; FHI 95652, 95693Bende; FHI Okpekpho; FHI 70244 Iyekuselu; FHI 72582 Owan, Ozalla, F.R., FHI 79618, 88974 Okomu, F.R., FHI 79618, 88974 Okomu F.R., FHI 87743, 67083 Ikom; FHI 91972 Ondo; FHI 72565 Iyekorhnomwon, Uronigbe F.R., FHI 56401 Calabar; FHI 82621 Ororekpe F.R., FHI 47171 Ijebu; FHI 82625, 64564, 69807, 60972 Ilesha; FHI 69890, 69722, 58478 Ugo-Ugboko Rd; FHI 41700, 53070 Gabon; FHI 13687 Ivory Coast; UIH 15916 Benin; UIH 16098 Ikom; UIH 3202 Omo F.R., UIH 19708 Calabar; UIH 17405 Cameroon; UIH 20129 Ikenne; UIH 3198 Buea (Cameroon); UIH 3200 Akure; Schleter 1239 Ishagamu; Schlecter 7141, 32013 Sapoba; Schleter 3257 Ubiaja F.R., Brenan 8776, 9151 Okomu F.R., Adam 12065 Guinea; Elliot 3933 Deighton 396, 3235, Thomas 1144; Jordan 315 Sierra Leone; Linder 127, Baldwin 6716, 6738, 10309 Liberia; Boughey GC 14523, 13516, 13550, 14795 Ivory Coast; Morton A350, GC9422, Irvine 533, 1660, 2218 Ghana.

 

Vernonia  Schreb
A large genus made up of 100 species distributed in America, Asia and Australia (Willis and Airyshaw, 1973). There are 60 species in West Africa out of  which 40 are known to occur in Nigeria (Hutchinson and Dalziel, 1963). A base number of 9 has been suggested for the genus by Darlington and Wylie (1955) and 17 has been suggested for the new world species by Jones (1979).

V. ambigua Kotschyi and Peyr.
An erect coarse annual herb up to 60 cm high with ribbed pubescent stems, florets purple. A weed of waste places widely distributed in lithosols, ferralsols and ferruginous tropical soils of Nigeria (Map 4). A haploid count of 10 was determined at metaphase (Fig.1. 8). Meiosis and pollen fertility were normal with 86% filled pollen and an average pollen grain size of 42 µm.

Achene Morphology
Entire achene 6 – 7 mm long; oblong – cylindric; arched sideways; light brown; pappus longer than the body; the base of pappus encircled by a series of semi-transparent scales; body hairy (Fig.2. 8).

Exsiccata
FHI 49165. 49177, 49190, 53008 Ghana; FHI 34657 Lagos; FHI 61401 Yola; FHI 19843, 30238 Ilorin; FHI 61600; 15775 Gombe; FHI 48359, 77283 Adamawa; FHI 54373 Kaduna; FHI 27962, 79964 Mokwa; FHI 796, 89869, 73624 A Zaria; FHI 60772 Igbetti; FHI 55042 Kaduna – Kontogora Road; FHI 21663 Jemma; FHI 21417 Kano; FHI 46953 Borno; FHI 45539 Yelwa (Borno); FHI 79899, 79632 Agare (Niger); IFE 7098 Mokwa – Bida Road; IFE 668 Kainji; IFE 7098 Mokwa; IFE 248 Katsina; IFE 406 Niger Rep; Omoigui 129 Jos; Omoigui 302 Bauchi; Omoigui 304 Kurra Falls.

Vernonia biafrae Oliv.and Hiern                                                = V. seneciodes A. Chev.
A climbing shrub with ribbed branches of florets. It is found in ferralsols, ferruginous tropical soils and eutrophic brown soils of Nigeria (Map 4). Two populations were encountered and they both had a haploid number of 9 (Fig.1. 9). Meiosis and pollen fertility were normal with 88% filled pollen and an average pollen grain size of 41 µm.

Achene Morphology
Entire achene 3-4 mm long; cylindric; thin; brown; marked with fine lengthwise ridges; ending in a knob; pappus long, thin (Fig.2. 9).

Exsiccata
FHI 61208 Omuaran (llorin); FHI 14914 Sierra Leone; FHI 931309, 78103 Mambilla; FHI 10459, 10458 Buea; FHI 25021 Unya (Obudu); FHI 51368 Ogoja; FHI 77220 Gembu; Omoigui 102 Obudu Cattle Ranch, Omoigui 325 Kaurra Falls.

Vernonia calvoana Hook f. subsp calvoana

= Stengelia calvoanaHook f.
A shrub up to 5 m high with velvety pithy branches with rather large sessile leaves; florets purple in heads nearly 3 cm across with white appendages in the middle and involucral bracts. The plant is common in ferralsol and ferruginous tropical soils of the country (Map 5) with annual rainfall of 1,270 mm – 1,524 mm.
The present haploid count of 10 was determined (Fig.1. 10).  Meiosis and pollen fertility were normal with 83.9% pollen stainability and an average pollen grain size of 53.6 µm.
Achene Morphology
Achene cylindric; black; marked with lengthwise ridges; rounded at the base; pappus long and delicate; body smooth; 1 cm long (Fig.2.10).
Exsiccata
FHI 50317 Ekona; FHI 99230 Ogbomosho; FHI 90451 Akamkpa; FHI 10515,  59518 Cameroon; FHI 89449 Ibarapa (Oyo); FHI 10465 Buea; Omoigui 107 Obudu Cattle Ranch.
Vernonia camporum A. Chev.
An erect woody herb up to 6 m high, with little branched stems, under surface of leaves and inflorescence tomentose, inflorescence branched with numerous sessile narrowly turbinate flower heads crowded in long spikes; Florets purple. It is restricted in distribution to soils ranging from alluvial to eutrophic brown soil regions of the country with an annual rainfall of 1,016 mm – 1,524 mm (Map 6).
A haploid count of 10 was determined at anaphase stage (Fig.1. 11). Meiosis and pollen formation were normal with 90% filled pollen and an average pollen grain size of 40 µm.Achene Morphology
Achene  cylindric, body densely pubescent, ending in a wide knob; pappus long and barbed, 8 mm long; black (Fig.2. 11).

Exsiccata
FHI 19762, 49180, 49188, 52994, Ghana; FHI 19263, 30242 Ilorin; FHI 94272 Jos; FHI 28958, 92799 Serti; FHI 991625 New Bussa; FHI 55120 Naraguta; FHI 68816 Gombe; FHI 37311 Kwakute Minna; Omoigui 156 Jos.

Vernonia cinerea (L.) Less
=Conyzacinerea L.
An annual,erect pubescent herb up to 1m long with alternate leaves and  much branched terminal inflorescence of small reddish purple-mauve heads, stem ribbed. A common weed of cultivated fields, roadsides and wet places. It is more abundant in the Southwestern part of the country in ferruginous tropical soil and ferrasols (Map 5). Two populations were studied and both had a haploid number of 9 (Fig.1. 12). Meiosis and pollen formation were normal with 80.1% filled pollen and average pollen size of 28.2 µm.

Achene Morphology
Achene 1-3 mm long; light brown; oblong; coarsely ridged; pappus feathery; base of pappus encircled by a series of semi-transparent hair-like scales; no body hairs (Fig.2. 12).

Exsiccata
Kennedy Nupe; Bater 1081 Lokoja; Thomas 5821 Logas; Dalz 1166 Ibadan; Omoigui 033 Oreghene (Benin); Omoigui 008 Ibadan; FHI 98948, 100200, 6160 Ibadan; FHI 9611, 87123, 48979, 49145, 49583, 96133, Obudu; FHI 96303 Ibarapa; FHI 83005, 100035 Ijebu-ode; FHI 99540 Abraka; FHI 95434, 96281 Oguta; FHI 6159 Benue; FHI 27580, 41792, 811123 Ilorin; FHI 49160, 95033, 90751 Owo; FHI 52548, 66235 Umuahia; FHI 10466 Warri; FHI 49206, 96443 Iseyin; FHI 82950, 83005 Badagry; FHI 78540, 87851 Calabar; FHI 50873 Lokoja; FHI 86277, 52486, 53046, 83054 Egbado; FHI 43294 Degema; FHI 70784 Igala; FHI 88973 Benin, FHI 190744 Oja-Odan; FHI 89563 Kajola; FHI 72259 Nsukka; FHI 70459 Enugu; FHI 94859 Epe; FHI 56594, 27580, 96686 Ifesowapo; FHI 92197 Akamkpa; UIH 3219 Sokponba; UIH 1170 Aviele; UIH 3215 Lagos; UIH 19663 Bida; UIH 19350 Brazil; Omoigui 008 Ibadan; Barter 1081Nupe; Parsons 3 Lokoja; Dalziel 1166 Lagos; Newberry 58 Ibadan; Franqueville 56, Berhaut 670 Senegal; Davey 180, 256 Mali, Esp. Santo 1058 Port Guinea; Adam 1790 Guinea; Vogel 108, Thomas 2095, Deighton 854, 2118, 5379, 5923  Sierra Leone; Baldwin 92139, Adam 16054 Liberia; Boughey GC. 18497 Ivory Coast; Brown 306, Andoh 5674, 5054, Bose 251, Adams 4130 Ghana; Warnecke 233 Togo; Chev. Dahomey; Morton GC 7130 Cameroon.

Vernonia cistifolia O. Hoffm.
A shortly pubescent under-shrub up to 60 cm high, with red purple florets in cymes forming a panicle. The plant grows in ferralsols, ferruginous tropical soil and eutrophic brown soils of the country with annual rainfall of 1,270 mm – 1,524 mm (Map 6).
A haploid count of 9 (Fig.1. 13) was determined for the species. Meiosis and pollen fertility were normal with 92% pollen stainability and an average pollen grain size of 42 µm.
Achene Morphology
Achene 5 mm long; light brown; cylindric, body covered by dense hairs, which elongates to form the pappus, base ending in a knob; pappus thin, delicate forming an obconic structure at the tip (Fig.2. 13).
Exsiccata
FHI 21026 Jos, FHI 52287 Vom; Omoigui 137 Obudu Ranch; Omoigui 178 Gembu.
Vernonia colorata (Willd) Drake
=Eupatorium colorotumWilld
=V. sengalensisLess
A shrub up to 13 m high, inflorescence broadly rounded, white scented florets. It is restricted in distribution to ferruginous tropical soils and lithosol regions of the country with annual rainfall of 1,016 mm – 1,526 mm (Map 6). The present haploid count was 9 (Fig.1. 14). Meiosis and pollen fertility were normal with 100% filled pollen and an average grain size of 57.64 mm.

Achene Morphology
Achene 9 mm long; dark brown; elliptic – cylindric ending with a white knob, body hairs absent, deeply grooved with lengthwise ridges; pappus with stiff upward directed hairs (Fig.2. 14).

Exsiccata
FHI 21365, 21208 S. Rhodesia; FHI 48347, 39015 Cameroon; FHI 10451, 64873, 14157 Olokemeji F.R., FHI 30156 Borgu F.R.; FHI 8092, 39493, 49181 Ghana; FHI 58588 Zungeru; FHI 59735 Jemma; FHI 22512, 92065 Abeokuta; FHI 91212 Shere Hills; FHI 13153 Bauchi Plateau; FHI 91950 Ado Ekiti (Ondo); FHI 3200 Abuja; FHI 68382 Ado-Odo (Badagry); FHI 67951 Imeko (Egbado); FHI 35985 Ibadan – Abeokuta; FHI 64812 Upper Ogun F.R., FHI 1989 Ikole; IFE 2077 Jarawa Hills (Bauchi); IFE 1471 Olokemeji F.R.; IFE 576 Omo-Ede; UIH 11918 Ikole; Omoigui 160 Jos.
Vernonia conferta Benth
A tree with a slender stem up to 15 m high; pale rusty tomentose on branchlets and infloresence with large distinctly petiolate sinuate-margined heads forming a very large terminal erect panicle, found in secondary forest.This plant is restricted in distribution to alluvial soils of Southern Nigeria with annual rainfall of 1,270 mm – 1,524 mm (Map 6).
A haploid count of 9 (Fig.1. 15) was observed at metaphase stage from two populations. Meiosis and pollen formation were normal with 98% filled pollen and an average pollen grain size of 46 µm.
Achene Morphology
Achene oblong – cylindric; with a tapering base; hairy; arched sideways; pappus in a ring of white, minutely plumose bristles; brown; 7 mm long (Fig.2. 15).
Exsiccata
FHI 81572, 51435 Ivory Coast; FHI 67872 Kurmi (Mambilla); FHI 38532 Udo (Okomu F.R.); FHI 35559, 82000 Cameroon; FHI 10469, 40479 Bamendae; FHI 40479 Udi Plateau; FHI 16337 Shasha F.R., FHI 69923 Ehor; FHI 78804 Obudu Plateau; FHI 48180 Calabar; FHI 51280 Mamfe; FHI 23081 Central Africa; FHI 10471 Victoria; FHI 10472 Uganda; IFE 3028 Obudu Plateau, IFE 1231 Oyo; IFE 2341 Idanre; Omoigui 190 Obudu Cattle Ranch.

Vernonia galamensis Willd Cass
= V. pauciflora (Willd) Less
= Conyza pauciflora Willd.
An erect annual herb up to 60 cm high with striate pubescent stems, florets blue. A common weed of ferralsols, ferruginous tropical soils and lithosol regions of the country with annual rainfall of 762 mm-1,270 mm (Map 7).
A haploid count of 9 was determined from two populations for this species (Fig.1. 16). Meiosis and pollen fertility were normal with 92% pollen stainability and 40 µm average pollen grain size.
Achene Morphology
Achene oblong; tapering towards base; coarsely ridged; light brown; pappus longer than the body; pappus of delicate white hairs which are minutely plumose; 4 mm long (Fig.2. 16).
Exsiccata
FHI 101059 Jos; FHI 65634 Borgu; FHI 84449; 94457, 46896 Zaria; FHI 84448 Funtua; FHI 64649, 68833, 68804 62347 Gombe; FHI 30237 Ilorin, FHI 81225, 63752 Mokwa; FHI 63491 Yola; FHI 21667 Anchau (Zaria); FHI 78853, 47970 Samaru; FHI 81753 Ivory Coast; FHI  91610, 91655, 27963 New Bussa; FHI 20500 Igbetti; FHI 67965 Ayangba (Kwara); FHI 42330 Upper Ogun F. R.; FHI 41815 Jebba; FHI 49193, 53086, 49196, 52981, 53981 Biu; FHI 33005 Hadejia (Kano); FHI 59678 Kaduna; FHI 81633 Upper Volta; FHI 62263 Egbe (Kabba); FHI 63830 Bukana; FHI 25643 Oyo; FHI 55041, Kaduna; Omoigui 245 Igbetti.
Vernonia gerberiformis Oliv.and Hiern.
A perennial herb with smooth ribbed simple or sparingly branched stems 45 cm high in clump arising from a fleshy rhizome, with large solitary blue heads. The plant grows in ferralsols, lithosols and ferruginous tropical soil regions of the country with annual rainfall of 1,016 mm-1,524 mm (Map 7).
Ten bivalents were counted at metaphase stage (Fig.1. 17). Meiosis and pollen fertility normal with 90% filled pollen and 64 µm average pollen grain size.
Achene Morphology
Entire achene 2.6 cm long; dark brown; cylindric, ending in a knob; deeply grooved with lengthwise ridges; body hairy; pappus thin, barbed and 1.6 cm long (Fig.2. 17).
Exsiccata
FHI 48244 Cameroon; FHI 10493 Bauchi Plateau; FHI 7642 Zaria; FHI 20185 Jos – Zaria; FHI 49973 Katagum; Omoigui 120 ABU (Zaria).

Vernonia glaberrima Welw ex O. Hoffm.
A much-branched shrub of about 2 m high, leaves scabrid florets in numerous heads, found scattered in open sandy grass land. It is distributed in ferruginous tropical soils, lithosols and ferralsols regions of the country with annual rainfall 254 mm-1,524 mm (Map 7).
A haploid count of 9 (Fig.1. 18) was determined which is in line with the base number of 9. Meiosis and pollen formation were normal with 89% filled pollen and an average pollen grain size of 29.95 µm.

Achene Morphology
Entire achene 9 mm long; dark brown; rectangular; coarsely ridged; pappus white; longer than body hairs; hairs 6 – 7 mm long, feathery with very fine secondary hairs, base of pappus encircled by semi transparent scales (Fig.2. 18).

Exsiccata
FHI 40155 Gwari (Zaria); FHI 19969 Katagum; FHI 10476 Bauchi; FHI 10175; 25731 Samaru; FHI 2292 Kaduna Jos Rd; FHI 55794 Naraguta; FHI 62981 Gurara Falls; FHI 62921, 1485, 306180 Yeboa; UIH 3225, Brinin Gwari; UIH 12357 Afaka F.R., UIH 3226 Ropp (Jos); UIH 16964 Lafia; Omoigui 052 Ogbonna: Meikle 1344, 1342 Birnin Gwari; Lely 65 Naraguta; Morton K. 346 Jos. Chev. 13237 Guinea; Elliot 5102, Burbridge 543, 551, Deighton 1352, 5417 Sierra Leone.Chev. 23953 Benin Rep; Maitland 1402, 1519, 1698 Boughey GC 11098, Morton K. 197 Cameroon; Omoigui 116 Jos, Omoigui 052 Ogbonna (Agenebode), 053 Bauchi-Ring Road- Jos.

Vernonia guineensis Benth var. cameroonica C.D. Adams
An erect herb with densely grey or brownish pubescent stems up to 50 cm high, florets purple. The plant grows in alluvial and eutrophic brown soils of the country with annual rainfall 1,016 mm – 1,524 mm (Map 8).
A haploid number of 10 was determined at metaphase stage (Fig.1. 19). Meiosis and pollen fertility were normal with 95.5% filled pollen and an average pollen grain size of 54 µm.

Achene Morphology
Achene oblong – cylindric;. Light brown; body narrows to the base; several sided; hairy pappus long and numerous; 8 mm-1cm long (Fig.2. 19).

Exsiccata
FHI 10479, 40617, 30038 Bamenda; FHI 34467 Nguroje; FHI 44095 Maisamari; FHI 61522, 93134 Gembu; FHI 3340 Cameroon; FHI 4750 Sierra Leone; FHI 103036, 5594 Shaki; FHI 92853, 46183. 66705 Mambilla; FHI 73673 Ogoja; IFE 643 Obudu Cattle Ranch; Omoigui 108 Obudu Cattle Ranch; Omoigui 241 Gembu.

Vernonia guineensis Benth var. guineensis C.D. Adams
= V. firma Oliv. and Hiern
A herb with erect stems arising from a perennial stock having numerous fusiform roots, florets purple. The plant is found mainly in lithosol regions of the country with an annual rainfall of 1,016 mm – 1,524 mm (Map 8).
A haploid number of 10 was determined at metaphase stage (Fig.1. 20). Meiosis and pollen formation were normal with 77% filled pollen and an average pollen grain size of 52 µm.

Achene Morphology
Entire achene 1.1 cm long; light brown; oblong; tapering towards the base; pappus longer than the body; coarsely ridged; body hairs present (Fig.2. 20).

Exisccata
FHI 10494 Bauchi Plateau 52338 Bafut – Ngemba; FHI 51134 Ivory Coast; FHI 2253, 48437 Zaria, IFE 6658 Keffi Rd Kaduna; IFE 1743 Mayo – Daga (Mambilla); Omoigui 240 Jos.
Vernonia iodocalyx O. Hoffm.
An erect shrub with coarsely dentate; petiolate leaves, flower heads purple. The plant grows widely in ferralsol and ferruginous tropical soils of the country with rainfall of 1,524 mm (Map 9).
The haploid count of 9 was determined at metaphase stage (Fig.1. 21). Meiosis and pollen formation were normal with 88% filled pollen and an average pollen grain size of 58 µm.

Achene Morphology
Achene  cylindric; black; marked with lengthwise ridges; body hairs present; base flat; 8 mm long (Fig.2. 22).

Exsiccata
FHI 10494 Bauchi, Plateau; FHI 52338 Bafut – Ngemba; FHI 51134 Ivory Coast; FHI 22953, 48437 Zaria; IFE 6658 Keffi – Kaduna; IFE 1743 Mayo – Ndaga (Mambilla); Omoigui 242 Obudu Cattle Ranch.

Vernonia ituriensis Muschl var. occidental C. Jeffery
= V. glabra var. occidental C.D. Adams
= V. glabra (Steez.) Vatke.
An erect herb up to 1m high with blue florets. It is restricted in distribution to lithosols and eutrophic brown soils of the country with annual rainfall of 1,016 mm – 1,524 mm (Map 9).A haploid count of 10 was observed (Fig.1. 22). Meiosis and pollen formation were normal.
Achene Morphology
Achene 2 cm long; dark brown; elliptic – oblong; deeply grooved with lengthwise ridges; body hairs present, pappus with stiff upward directed barbed hairs; base ending with a white wide knob (Fig.2. 22).

Exsiccata
FHI 992386 Abuja; FHI 13157 Bauchi Plateau; FHI 7781, 77073 Maisamari; FHI 62384, 84528 Gembu; FHI 10478 Buea, FHI 92750 Mambilla; FHI Bamenda; FHI 4822 Cameroon; FHI 57099 Jos; IFE 1572 Bamenda; Omoigui 105 Jos; 135 Kurra Falls.

 

V. migeodii S. Moore.

=V. courtetii O. Hoffm.
=V. plumbaginifolia Chev.
=V. ambigua Kotschyi and Peyr.
A perennial herb arising from a woody root stock with several sparsely pubescent stems up to 1-5 mm high; leaves sessile; rounded at the base; glandular; purple florets when young turning white during maturity; a common roadside  plant of savanna region. In Nigeria, it is restricted in distribution to ferruginous tropical soils and ferrasols regions of the country (Map 4). A haploid count of 11 (Fig.1. 23) was determined at Metaphase 1. Meoisis and pollen formations were normal with 95.8% pollen stainability and average pollen grain size of 41 µm.
Achene Morphology
Achene 7-9 mm long; light brown; oblong; pappus purple tinged; feathery 6-7 mm long with fine secondary hairs; base of pappus encircled by a series of semi- transparent flat scales (Fig.2. 23).

Exsiccata
FHI 22952 Igabi (Zaria); FHI 25768 R. Benue, Dalz 32 Anara F.R. (Zaria) Phillips 2550 Lagos; Thomas 33 Awka; Thomas 367 428 Obu, Rowland Ogbomosho; Morton 25 Onitsha- Enugu; FHI 10484 Ikom; Omoigui 172 Benin-Akure Rd; Omoigui 21 Onitsha-Awka.

Vernonia nestor S.  Moore
= V. chariensis O. Hoffm
A perennial herb with stiff stems 60 cm high branched only at the top, leaves densely crowded and rather small. Stems are densely clothed with long, silky hairs, florets mauve in numerous heads. The plant is restricted in distribution to lithosol regions of the country with annual rainfall of 1,016 mm – 1,270 mm (Map 10).
A haploid number of 9 (Fig.1. 24) was determined for this species. Meiosis and pollen fertility were normal with 70% pollen stainability and an average pollen grain size of 36 µm.

Achene Morphology
Achene obconic; covered with long body hairs; pappus long, light brown, 5 mm long (Fig.2. 24).

Exsiccata
FHI 92726, 77258A Mambilla; FHI 99023 Jos; FHI 97930 Jos- Kaduna Road; FHI 56961 Barakin Ladi; FHI 101007, 101005 Pankshin; FHI 10485 Bamenda; FHI 44044 Anara F.R. (Kaduna); FHI 59584 Zaria; FHI 84565 Jos; Omoigui 135 Kurra Falls (Jos).

Vernonia nigritiana Oliv and Hiern.
An erect branched annual herb with leaves highly scabrid; involucral bracts coloured bright red on inner surface, spreading in heads up to 3 mm long. The plant is widely found in lithosols, ferralsols and ferruginous tropical soils of Nigeria with an annual rainfall of 762 mm – 1,270 mm (Map 10).
A haploid count of 9 was determined at metaphase stage (Fig.1. 25) for the species. Meiosis and pollen fertility were normal with 100% filled pollen with an average grain of 63.2 µm.

Achene Morphology
Entire achene 1cm long; dark brown body; oblong – cylindric; body gradually narrows to the base forming a knob; body hairs absent, pappus longer than the achene with delicate hairs (Fig.2. 25).

Exsiccata
FHI 46665, 52212 Olokemeji F.R., 28930 Karamatin (Mambilla); FHI 64144, 31613 Ilorin; FHI 32005 Gwari; FHI 3162 Gwada (Niger); FHI 5661 Kabba; FHI 79979 Abuja, FHI 29793, 64775 Bornu; FHI 30796, 43561 Owo; FHI 53045, 49202 Ghana; FHI 45978 Igbetti; FHI 71550 Pategi; FHI 83974 Zaria Kaduna Road; FHI 42036 Farin Dube (Kano); FHI 71400 Kwara (Aro); FHI 55045 Kujama (Zaria); FHI 13978 Ivory Coast; FHI 45492 Igabi; FHI 80073 Mokwa; FHI 35218 Afaka F.R.; FHI 43101 Zaria; FHI 57686 Oyo; FHI 39331 Giwa (Zaria); FHI 38197 Shaki (Oyo); F.R. 76997 Mambilla; FHI 84861; FHI Birnin Gwari (Zaria); FHI 61341 Kabba; FHI 59343 Kaduna; FHI 39256 Jemma; FHI 94484 Zaria – Old  Kano Road; FHI 38423 Ole F.R. (Kwara); FHI 21790 Pankshin; FHI 23734 Keffi FHI 91558 New Bussa; FHI 5325 Sierra Leone; FHI 87063 Ogoja; FHI 90004, 84566 Gashaka; FHI 81655 Upper Volta; FHI 88518 Isanlu (Kwara); FHI 48324 Cameroon; FHI 2991 Borgu F.R. FHI 40307 Shaki; FHI 19275 Osi (Kwara); IFE 514 Old Oyo Game Reserve; FHI 21071 Borgu Game Reserve; FHI 6496 Jos Water Works; IFE 6636 Pankshin; IFE Zaria Funtua Road; Omoigui 237 Kurra falls.

 

Vernonia  perrotettii Sch. Bip.
An erect much branched annual herb up to 40 cm high, with magenta florets, leaves scabrid – steulose. It is restricted in distribution to alluvial and lithosol regions of the country with annual rainfall of 1,016 mm – 1,574 mm (Map 10).
A haploid count of 10 was determined (Fig.1. 26). Meiosis and pollen fertility were normal with 99% pollen stainability and 48 µm average pollen grain size.
Achene Morphology
Achene 9 mm long; light brown; cylindric; narrowing  the base; deeply grooved by lengthwise ridges; body hairy with upward directed hairs; pappus long and barbed (Fig.2. 26).
Exsiccata
FHI 62002 Yola; FHI 14004, 18416 Zaria; FHI 49201 Ghana; FHI 98137 Burundi; FHI 99021 Jos, FHI 69533 Cameroon; FHI 27969, 63733 Mokwa; FHI 8338 Ivory Coast; FHI 98955, 98958 Mokwa – Boda Road; FHI 76922 A.B.U. Campus; FHI 83094 Kaduna; FHI 84684 Birnin Gwari; FHI 89613, 91980 Nsukka; FHI 91973 Shika F.R. FHI 65915 Enugu; FHI 21523, 3413, 24680, 27314 Akwa; FHI 41915 Riyom; FHI 94280, 94370 Jos FHI 36714 Udi; FHI 16446 Dambata (Kano); FHI 100, 0982 Keffi FHI 6117 Lafia; FHI 10492 Bauchi Plateau; FHI 47534 Bukuru; FHI 32979 Agulu (Onitsha); FHI 64901 Kaduna – Jos; FHI 62229 Egbe (Kabba); FHI 55092, 1197 Naraguta; F.R., FHI 65617 Borgu; FHI 79525 Nimba; FHI 38983 Maiduguri; FHI 59595 Kaduna; IFE 766 Mokwa – Jebbard; IFE 355 Kaduna – Kontagora Road;  IFE 405 Niger Rep; IFE 116 Keffi; IFE 6756 Benin Republic; Omoigui 099 School of Forestry, Jos.

Vernonia philipsoniana Lawalree.
=V. lappoides O. Hoffm.
=V. hoffmanniana Hutch and Dalz.
A perennial herb, woody at the base, stems ribbed and loosely hairy, branched below the infloresence, florets purple in few flower heads. It is restricted in distribution to lithosol regions of the country with annual rainfall of 1,016 mm – 1,270 mm (Map 9). A haploid number of 9 (Fig.1. 27) was determined at metaphase stage. Meiosis and pollen fertility were normal with 100% pollen stainability and an average pollen grain size of 48 µm.

Achene Morphology
Achene oblong elliptic; coarsely ridged; light brown; pappus longer than the body hairs; densely pubescent; hairs short plumose with very fine secondary hairs; pappus with bristles, body ending in a knob; 1cm long (Fig.2. 27).

Exsiccata
Omoigui 126 Jos.

Vernonia richardiana (O. Ktze) P. Sermolli
= Cacalia richardana O. Ktze
=V. sereti De Wild
= V. myriocephala A. Rich.
A shrub 2 m high, unbranched below, with sessile undulated margined leaves, flower heads subsessile in divaricately branched panicles, florets bluish purple to mauve. The plant is restricted in distribution to lithosol and eutrophic brown soils of the country with annual rainfall of 1,270 mm-1,524 mm (Map 12).
A haploid count of 10 (Fig.1. 28) was determined for the species. Meiosis and pollen fertility were normal with 97% filled pollen and an average grain size of 44 µm.

Achene Morphology
Achene 9 mm long; light brown; body elliptic ending with knob, body hairs short, pappus present (Fig.2. 28).

Exsiccata
FHI 53049 Ghana; FHI 33121, 93226 Mambilla; FHI 13156 Bauchi Plateau; FHI 10503 Ropp; FHI 69570, 48221 Cameroon; Omoigui 228 Toro (Bauchi), Omoigui 229 Gembu.
Vernonia saussureoides Hutch.
A perennial weed of hilly grassland with erect glabrous pithy stems up  to 60 cm high, arising together from a woody stock, floret deep blue. It is restricted in distribution to lithosols and eutrophic brown soils of the country with annual rainfall of 1,016 mm – 1,524 mm (Map 12).
A haploid count of 9 (Fig.1. 29) was determined for the species. Meiosis and pollen fertility were normal with 72% filled pollen with an average pollen grain size of 62 µm.

Achene Morphology
Achene oblong; brown, body hairs present, marked with lengthwise ridges, body ending in a knob, pappus long and barbed, 1.4 cm long (Fig.2. 29).

Exsiccata
FHI 37087, 59307 Pankshin; FHI 10497, 30466 Bamenda; FHI 7270, 5691 Naraguta F. R; FHI 10498, 1317 Bauchi Plateau; FHI 44764 Pantisana (Adamawa); FHI 62716 Yelwa; Omoigui 092 Kurra Falls.

Vernonia smithiana Less
A perennial herb in hilly grassland with erect slender pale-silky stems up to 45 cm high arising from a woody stock, florets pinkish turning white in hemi-spherical heads crowded in dense terminal corymbs. It is restricted in distribution to ferruginous tropical soils and eutrophic brown soils of Nigeria with annual rainfall of 1,016 mm-1,524 mm (Map 11). A haploid count of 10 (Fig.1. 30) was determined at metaphase stage and meiosis was normal for the species.

Achene Morphology
Achene oblong-cylindric; black with lengthwise ridges; rounded at base; pappus long and delicate; body smooth 5 mm-1 cm long (Fig.2. 30).

Exsiccata
FHI 10499, 22167, 3004, 40610, 38862, 41118, Bamenda; FHI 48916 Gembu; FHI 34857, 29386 Bafut-Ngemba; FHI 66733, 63292, 92847 Mambilla; FHI 53624 Gabon; FHI 48341 Cameroon; FHI 44884 Maisamari; FHI 62748 Yelwa; FHI 79251 Obudu Plateau; IFE 1668 Nguroje; IFE 1700 Mayo-Ndaga, Omoigui 146 Ikwette Plateau (Obudu); Omoigui 281 Gembu.

Vernonia stenocephala Oliv.
= V. oocephala Bak
An erect under-shrub up to 60 cm high, almost unbranched from the base, with whitish florets in numerous crowded heads. It is restricted in distribution to lithosol and eutrophic brown soil regions of the country with annual rainfall of 1,016 mm – 1,524 mm (Map 10).The haploid count of 9 was determined at M1 (Fig.1. 31) for this species. Meiosis and pollen fertility were normal with 74% filled pollen with an average pollen grain size of 48 µm.

Achene Morphology
Achene 1 cm long, black, cylindric; ending in a knob; body hairy; marked with lengthwise ridges, long, delicate not barded (Fig.2. 31).

Exsiccata
FHI 56979, 99006, 91190, 79830 Jos; FHI 13155, 10500 Bauchi Plateau; FHI 59306 Kurra, Omoigui 093 Jos.
Vernonia stenostegia (Stapf.)Hutch and Dalz.
= Candida stenostegia Stapf.
A roadside weed up to 1.5 cm high, sometimes leafless below, florets purple, with wrinkled leaves. The plant is restricted in distribution to ferralsols, ferralginous tropical soils and lithosol regions of the country with rainfall of 1,016 mm-1,524 mm (Map 11).
A haploid count of 10 was determined at metaphase stage (Fig.1. 32). Meiosis and pollen fertility were normal with 96% filled pollen and an average pollen grain size of 40 µm.

Achene Morphology
Achene 1 cm long; black; cylindric; ending in a knob; body hairy; marked with lengthwise ridges; pappus long; delicate not barbed (Fig.2. 32).

Exsiccata
FHI 56976, 99006, 91190, 79830 Jos; FHI 13155, 10500 Bauchi Plateau, FHI 59306 Kurra, Omoigui 093 Jos.

Vernonia subaphylla Bak.
A perennial herb with few round based flower heads, florets reddish purple terminal on the branches of an almost leafless stem about 45 cm high, arising from a woody stock, usually flowering after burning. The plant is restricted in distribution to ferruginous tropical soil and lithosol regions of the country with annual rainfall of 1,270 mm – 1,524 mm (Map 11).A haploid count of 9 was determined at metaphase stage (Fig.1. 33). Meiosis and pollen fertility were normal with 98% filled pollen and an average pollen grain size of 60 µm.

Achene Morphology
Achene 1cm; body oblong – cylindric; body curved inwards at one side; base knobbed; brown hairy; pappus long, hairs, stiff narrow toward base (Fig.2. 33).

Exsiccata
FHI 40609, 10501 Bamenda, FHI 63333 Cameroon, FHI 75254 Malawi, IFE 3055, 696 Obudu Plateau, IFE 1783 Cameroon, Omoigui 143 Obudu Cattle Ranch.

Vernonia tenoreana Oliv.
An erect shrub up to 2 m high with heads up to 3 cm across; florets white; leaves not wrinkled, creamy coloured; sparingly found in open places. The plant is commonly found in lithosols, ferralsols, alluvial and ferriginous tropical soils of the country with annual rainfall of 1,016 mm-1,524 mm (Map 12). A haploid count of 9 (Fig.1. 34) was determined at M1. Meiosis and pollen formations were normal with 91.48% pollen fertility and an average pollen grain size of 46.74 µm.
Achene Morphology
Entire achene 1-1.5 cm long, light brown, oblong, coarsely ridged, pappus purple tinged and longer than body hairs, hairs 9 mm long feathery with a blunt base (Fig.2. 34).
Exsiccata
FHI 46400 Igbetti, FHI 58089 Jos, FHI 57694, 47285 Oyo, FHI 64223, 14831, 71579, 88571 Ilorin, FHI 43893, 6314, 31589 Ibadan, FHI 101003 Pankshin, FHI 8446 Panchanu; FHI 96356 Ibarapa; UIH 11452 Ibadan, UIH 10619 Oyo, UIH 3250 Abeokuta; Omoigui 042, 065 Ibadan; Summmerhayes 34 Makurdi, Tama Bauchi; Millen 129 Lagos; Millen 94 Abeokuta, Chev. 14062, 3833 D’overy 214 Guinea Bissau, Jac-rel 1991 Guinea; Chev.22786 Benin Rep; Omoigui 042 Igbetti; 043 FRIN Ibadan, 007 Ibadan.

Vernonia undulata Oliv and Hiern
A perennial herb with erect leafy stems up to 60 cm high from a woody stock; florets purple in distinctly stalked heads. The plant grows is in ferrasols, ferruginous tropical soil, lithosols and eutrophic brown soils of Nigeria with annual rainfall of 1,016 mm – 1,524 mm (Map 9).  A haploid count of 9 was determined at metaphase stage (Fig.1. 35). Meiosis and pollen fertility were normal with 71% filled pollen and an average pollen grain size of 32 µm.

Achene Morphology
Achene rounded; deeply grooved in to several sides with minute hairs; black; 3–5 mm long (Fig.2. 35).

Exsiccata
FHI 37310 Minna; FHI 148335 Mambilla; FHI 63331 Cameroon; FHI 84584, 65575 Gembu; FHI 46815 Jos; FHI 70100 Igumala (Bauchi Plateau); FHI 23878, 63208, 63210 Gashaka; FHI 10467 Bauchi Plataeu; FHI 4753 Sierra Leone; FHI 25924, 25921 Giwa (Zaria); FHI 76117 Ethiopia; Omoigui 082 Ikwette Plateau (Obudu); Omoigui 180 Gembu.
Discussion
Chromosome numbers have been widely used in the systematic investigations of the Asteraceae. The chromosome numbers of members of this tribe have been studied (Chaung et al., 1963; Coleman, 1968; Peng and Hsu, 1978; Morton, 1962; Mangenot and Mangenot, 1957,1962; Jones, 1980). According to Gupta and Gill (1986), the tribe is inadequately known cytologically, particularly its tropical taxa.
The range of the chromosome number of the Nigerian Vernonieae is from n=8 (Triplotaxis stellulifera) to n=16 (Centratherum punctatum), an introduced plant. Chromosome numbers of 12 species of this tribe have been recorded for the first time from the Nigerian flora while 2 species differ from earlier reports. The chromosome number of n=9 reported for V. tenoreana differs from the earlier report of n=10 by Ayodele (1992). Thus, 2 cytotypes are present in this species.The report of n=11 for V. migeodii is the first report of this number in the old world Vernonieae. Thus, a new base number of x = 11 is suggested for the genus.
Chromosome numbers have been found to be constant in Elephantopus (n=11). The count of n=11 agrees with the earlier works (Chaung et al., 1963; Coleman, 1968; Peng and Hsu, 1978;  Jones, 1979) while the count of n=11 for E. spicatus confirms the earlier reports (Federov, 1969; Mehra et al., 1965; and Thulin, 1970). The count of n=8 for Triplotaxis stellulifer agrees with Mangenot and Mangenot (1962) while the count of n=16 for Centratherum punctatum agrees with earlier workers (Turner and King, 1964; Coleman, 1970). The count of n=20 for Ethulia conyzoides agrees with the report of  Pilz(1980).  Jones (1979) recorded a count of n=10 and the presence of 2-4 B chromosomes in the species.
The report of chromosome number n=9 and n=10 for many of the species studied for this genus agrees with the base number of 9 and 10 suggested for the old world taxa. The report of n=11 for V. migeodii is the first report of the number in the genus Vernonia.
Isawumi (1996) moved the species V. migeodii to another genusVernoniastrum migeodii. Similarly, he moved V.colorata (n=9) = Gymnanthenum coloratum; V.conferta (n=9) =Monosis conferta; V. gerberiformis (n=10) = Linzia gerberiformis, V. nigritiana (n=10) = Linzia nigritiana; V. ituriensis var. occidental (n=10) = L. ituriensis var. occidental; Vernonia cinerea (n=9) = Cyanthallium cinereum and the others such as V. biafrae (n=9) and V.guineensis var. cameroonica (n=10), V. calvoana, subsp calvoana (n=10) that are left in the genus maintain this number. Until more members of the genera are studied, this character which is the most reliable character in taxonomic studies could not justify the transfer.
The report of n=10 for V. ambigua agrees with Ayodele (1992), n=9 for V. cistifolia (Jones, 1982); n=9 for V. colorata(Mangenot et al., 1957 and Ayodele, 1992); V. cinerea n=9 (Federov, 1969; Moore, 1974; Goldblatt, 1981; Gupta and Gill, 1983); n=9 for V. conferta (Mangenot et al., 1957; Ayodele, 1992); n=9 for V. galamensis (Ayodele, 1992);  n=9 for V.guineensis var. cameroonica (Mangenot and Mangenot, 1957,1962); n=10 for V. ituriensis var. occidental (Riley and Hoffman, 1961; Turner and Lewis, 1965; Jones, 1979,1982); n=9 for V. nestor (Ayodele, 1992); n=9 for V. perrotetti (Miege, 1960; Jones, 1982 and Ayodele, 1992); n=10 for V. smithania (Jones, 1982); n=10 for V. stenostegia (Ayodele, 2000) and n=9 for V. undulata (Morton, 1966).
Majority of the plants studied had n=9 (17 species), n=10 (12 species and variety), n=8 (2 species) and n=11 (3 species) in Vernonia and Elephantopus. Elephantopus (X=11,13) which seems to have 11 as the most successful number does not share any basic number with any other genus in the tribe and n=11 for V. migeodii is quite unusual as this is the first record of that number in the genus  Vernonia. The basic chromosome numbers represented in the tribe are X=7,8,9,10,11,13 and 17 of which X=9 and 10 are predominant in the old world species (Jones, 1977). Though a large number of the species of the central and largest genus Vernonia are unknown chromosomally but the new world taxa are based on X=17, which is derived through polyploidy followed by aneuploidy. The basic chromosome number x=10 and x=9 were suggested to be the primary basic numbers of the tropical Vernonieae (Jones, 1977) and this agreed with the Indian (another tropical) taxa of the tribe Vernonieae (Gupta and Gill, 1986) whereas the Nigerian taxa are opposite i.e., x=9 is more prevalent as such x=9 is suggested as the primary basic number of the tribe. Basic chromosome number of 8 which is rare in the old world species was never observed in the Indian taxa (Gupta and Gill, 1986) while x=8 was found in Triplotaxis stellulifera and Centratherum n=16 and this would have arisen as a result of polyploidy. In the tribe, polyploidy, aneuploid and phylogenetic increase or decrease at the diploid level have added to the numerical diversity in chromosomes in some of the genera. It has gone to the extent of polybasic nature, leaving no clearcut indication of the original basic number.
Earlier, Isawumi (2008) based on floral characters made some changes. He moved 12 species from the genus Vernonia  to other genera e.g V. migeodii to Vernoniastrum migeodii and the chromosome number of (n=11) is yet to be recorded in any tropical Vernonia. He also moved Vernonia gerberiformis to Linzia gerberiformis (n=10), V. ituriensis var. occidental to L. ituriensis var. occidental (n=10) and V. nigritiana to L. nigritiana (n=9). From these results, the genus Linzia is already dibasic with basic chromosome number x=9, 10 and the monophyletic status of this genus is already in doubt. He moved Vernonia cinerea to Cyanthillium cinerum (n=9). x=9 is in line with the result of other members of groups that were left in the genus e.g.Vernonia biafrae.The results of this work support the caution of earlier workers (Drury and Watson, 1965; Ayodele and Olorede, 2005) on the use of limited characters in creating changes in the taxonomy of plants. In the  family, a correlation has been suggested between low chromosome number and the annual habit by many workers (Stebbins, 1950; Babcock, 1947; Clausen, 1951; Gupta and Gill, 1986). Babcock(1947) observed  in the genus Crepis that a decrease in chromosome number was associated with the annual habit. Solbrig (1977) supported this view that lower numbers were more prevalent among annuals compared to the perennials as well as woody elements.This correlation does not conform with the Nigerian Vernonieae as almost all the species studied were herbs and they had chromosome number in the same range as the shrubs (Vernonia calvoana var. calvoana n=10, V. colorata n=10, V. conferta n=9, V.richardiana n=10, V. stenostegia n=10 and V. tenorena n=9). The importance of pollen grain size and morphology in the characterization of plant species has been stressed by many authors (Stebbins, 1950; Olorode and Baquar, 1976). Clausen(1962) reported the use of pollen size as a reliable character for distinguishing two species of Betula. In the family Asteraceae, Olorode and Torres (1970) used this character to delimit artificial hybrids obtained from the genus Zinnia while Adegbite (2010) used the same character in the delimitation of hybrids obtained from the genus Aspilia in Nigeria. The results from this study have shown that the pollen grain size could be used in delimiting species at the species level. The pollen grain size of the members of this tribe ranged from 28 µm (Elephantopus spicatus) to 60 µm (Vernonia richardiana, V.stenostegia and V.saussureoides). The result from this character did not support the changes, for example, Vernonia gerberiformis=Linzia gerberiformis had (46 µm), V. ituriensis var. occidetalis=L. itureinsis var. occidentalis (48 µm) and V. nigritiana=L. nigritiana (36 µm). V. biafrae 42µm and V. cinerea (40 µm).
The pollen fertility test revealed that meiosis was normal with regular bivalent pairing and this was evidenced with the high pollen fertility which confirmed that hybridization was not responsible for the differences in their chromosome number.
Heywood et al. (1977) and Ayodele (1995) noted a correlation between the pappus structure and growth habit of some members of the family. They observed that the herbs produced large sized fruits and had more projections of the pappus than the shrubby species with large fruits. The result of this study sugests that the variation in the structure of the pappus could be one of the efficient means of dispersal. This agrees with the view of Gupta and Gill (1986) who attributed the remarkable success of the family to their efficient  mode of dispersal and diversity of habitats.
The results from the distribution maps of all the taxa studied would help researchers in Nigeria to solve one of the difficulties of studying the members of the tribe (collection of plant material) (Ayodele, 1995). From the distribution maps presented, 13 species are restricted to higher altitudes (Vernonia cistifolia, V. camporum, V. guineensis var. cameroonica, V. guineensis var. guineensis, V. iodocalyx, V. smithiana, V. subaphylla, V. undulata, V. stenocephala, V. nestor, V. phillipsoniana,Elephantopus mollis  and E. spicatus). Three species are restricted to Southern Nigeria (Triplotaxis stellulifera, V. calvoana var. calvoana and V. conferta, V. migeodii). Species restricted to Northern Nigeria are V. gerberiformis, V. perrottetii and V.richardiana. Species of wide-spread occurrence include Ethulia conyzoides, Gutenbergia  nigritiana, G. rueppellii,  V. biafrae, V. cinerea,V. galamensis, V. nigritiana, V. stenostegia and V. tenoreana.
Conclusion

The author supports the views of Drury and Watson (1995) and Ayodele and Olorode (2005) that before any reclassification of the family, more results from different areas of biology should be obtained and analysed.

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VALUE OF MEDICINAL PLANTS: A SCALE DEPENDENT ON TIME AND RACE BEING GUEST LECTURE PRESENTED AT THE 2015 ANNUAL CONFERENCE OF BOTANICAL SOCIETY OF NIGERIA (BOSON) HELD AT THE UNIVERSITY OF LAGOS, AKOKA, NIGERIA.

Introduction

That humans from time immemorial have continued to harness the medicinal potentials of plants is in compliance with the will and commandment of the Almighty God. As recorded in the Holy Bible (Ezekiel 47:12), a verse concerning “trees” states that: “Their fruit will be for food and their leaves for medicine”. Although it is not known exactly when the use of plants as medicines actually started, it is thought to have resulted from human instinctive tendencies and by trial and error. This postulate is supported by the evidence provided by animals living in the wild – they eat plants that heal them and avoid plants that do them harm. With increasing development of civilization, the knowledge of herbs became transmitted verbally and experientially.
In the written record, the study and use of medicinal plants dates back to over 5,000 years to the Sumerians. The first Chinese herb book written about 2,700 BC, listed 365 medicinal plants and their uses. Ma Huang (Ephedra spp) from which ephedrine was sourced was included. Hippocrates the ‘Father of Medicine’ (460-370 BC) produced 79 books and 59 treatises on which modern medicine is said to be founded. Hippocrates advocated the use of some herbal drugs along with fresh air, rest and proper diet to help the body’s own ‘life force’ eliminate the health problem. In the first Century A.D., a Greek physician Dioscorides produced a long treatise on the properties and uses of over 500 medicinal plants. This work remained an authoritative reference work used until about the 17th Century.
Following the invention of printing in the 15th Century, hundreds of books on medicinal plants were published e.g. Theophrastus’ Historia Plantarum and Dioscorides’ De MateriaMedica. The 15th, 16th and 17th Centuries were the great age of Botanical medicine books (Herbals) many of them available for the first time in English other than Greek or Latin. The most popular of the English herbalists was John Gerard, a Tudor Surgeon, apothecary to King James I, and superintendent of the Gardens of the Court of Queen Elizabeth, where he cultivated over 1,000 herbs. Gerard published the herbal book, “The Herballe” or “General Historie of Plantes” in 1597 (Lust, 1979).
The pre-eminent position held by Botanical Medicine began to be slowly eroded in the 17th Century. This was due to the introduction of active chemical drugs like arsenic, iron, mercury, copper sulphate and sulphur by the alchemist Paracelsus. Added to this was the rapid development of chemistry and other physical sciences in the 18th and 19th centuries. Thus chemotherapy became the orthodox system of the 20th Century.
The use of herbs in Africa must be as old as the continent itself. The Egyptians of 1,000 BC used garlic, castor oil, mint, etc. The hieroglyphic records from the early Egyptian culture have provided evidence on the evolution of the use of aromatic plants to produce essential oils. Until the advent of European explorers dating back to the 15th Century, most Africans had no alternative to traditional medicine which included phytotherapy.  As an example, the Yoruba race has the history of its traditional medicine well rooted in the spiritual pedagogy called “IFA”, which tells us that the father of herbalism is Osanyin. According to “IFA”,

Department of Botany University of Ibadan, Ibadan, Nigeria.

pedagogy, which is an account of African wisdom in the custody of the Yoruba race, Osanyin was bought as a slave with 2,000 cowries (“egbaa”) to take care of the garden of Orunmila the Master of spiritual Therapy. Orunmila instructed Osanyin to weed the garden but Osanyin did not do so because he knew the magical and medicinal properties of the plants in the garden. When Orunmila inspected the garden and found that Osanyin had not done anything his anger kindled. “Osanyin”, Orunmila called, “the garden I asked you to weed you have refused to weed it”. Osanyin replied that he did not know which of them he should weed.When Orunmila demanded to know why, Osanyin started to show him the medicinal and magical properties of the plants in the garden. “This one is the leaf for good luck, I must not weed it”, he said. “This leaf is for procuring pregnancy, this leaf is to stop the machination of born-to-die children, this leaf is to cure ailments, this leaf is to enable one secure a good wife, which of them do you think I should weed?” (Dopamu, 1977). Impressed by Osanyin’s outstanding magical and medicinal knowledge of plants, Orunmila later proclaimed him free and made him his companion. According to Komolafe (1998), Osanyin existed and practised herbalism much earlier in creation than the famous Egyptian herbalist and physician called Inhotep(2,980 BC).  A native doctor from Ijebu-Ode, Dr. Joseph Odumosu (1863-1911) was the first person to publish two books in Yoruba traditional medicine. “Iwe Egbogi” (Book on Herbs) and “Iwe Iwosan” (Book on Healing) were published about 1895.
In South Africa traditional medical systems are passed on by word of mouth from one generation to the next. These medical systems and their herbal animal and mineral material medica have ancient origins dating back to palaeolithic times (Van Wyk et al., 1997). Traditional healers are known as “Inyanga” and “Isangoma” (Zulu) referring to herbalists and diviners, respectively. In modern times some healers practise both arts.
Africans regard medicinal plants as sources of vital energy and to a great extent a participatory entry rather than a lifeless object used in healing. Africans believe that plants have a spiritual power or life force which can be harnessed for the benefit of the patient. This belief agrees with that of Paracelsus the transcendental alchemist who acquired a lot of knowledge of the healing powers of plants from traditional European herbalists. In spite of the fact that the practice of herbalism is still associated with magic in some cultures or races worldwide, scientifically we know that leaves, barks and roots of medicinal plants have therapeutic properties of their own and will not need any additional agent or forces to make them potent. This is the basis of modern herbalism.

Resurgence of Herbalism
Despite the predominant influence of western allopathic medicine, there has been a resurgence of herbalism all over the world. What has led to this development? Dhawan (1997) provided the following reasons:
i.    high cost of development and inadequacy of synthetic drugs.
ii.    inadequate availability of modern drugs to cover total population in large countries.
iii.    low priority of third world diseases by Drug Houses in developed countries.
iv.    better local acceptability of traditional drugs
v.    iatrogenic diseases and toxicity of synthetic drugs.
vi.    economic benefit from local cultivation of medicinal plants.
According to a United Nations report (Dees, 1993:28), United States government health agencies alone spent more than $8 million from 1986-1993 to collect thousands of medicinal plant species in more than twenty-two countries. This action must have been predicated on the need to source new drugs from plants. Masood (1997) reported that the sale of medicinal plants have grown by nearly 25% in India over the past ten years, the highest growth rate in the world then. Germany is Europe’s largest consumer of such plants and spends £1.4 billion (US $2.2 billion) annually. France is second (£116 million) and the United Kingdom third (£88 million). Wagner (1988) estimated that 30-40% of all medical doctors in France and Germany relied on herbal preparations as their primary medicines.  The demand for medicinal plants by pharmaceutical companies alone was estimated to be 25% of prescription drugs in the U.S.A. – containing plant extracts or active principles prepared from higher plants (Farnsworth et al., 1986). Mander (1998) reported that 27 million people consume medicines sourced from indigenous plants in Kwazulu-Natal, South Africa. Farnsworth (1988) also reported that 74% of the plant-derived compounds currently used in pharmaceuticals retain the same or a similar use by traditional healers. In realisation of the widespread use of herbal medicines across the globe, the World Health Organization in 1985 estimated that perhaps 80% of the world population relies on herbs for primary health-care needs (Farnsworth et al., 1985).

Medicinal Plants of Different Races
Azadirachta indica (Meliaceae): Neem

Neem is native to India. The fruits, seeds, oil, leaves, root and bark have been used in the Indian Ayurvedic and Unani systems of medicines. For centuries, millions of Indians have cleaned their teeth with neem twigs, smeared skin disorders with neem leaf juice, taken neem tea as a tonic and placed neem leaves in their beds, books, grain bins and cupboards to keep away bugs. Neem has relieved so many different pains, fevers, infections and other complaints that it has been called “the village pharmacy” (Vietmyer, 1992).
Azadirachta indica has been introduced to tropical Africa, America, Saudi Arabia and the Caribbean. Scientific studies have shown that neem parts (fruits, seeds, oil, leaves, bark and roots) have uses such as general antiseptics, antimicrobials, treatment of urinary disorders, diarrhoea, malaria, skin diseases, infected burns and inflammatory diseases. Owing to its multifarious uses to humans, the United Nations declared neem as the “Tree of the 21st Century”.
Andrographis paniculata (Acanthaceae)  commonly known as “king of bitters”, A. paniculata has been used for centuries in Asia to treat gastrointestinal and upper respiratory infections, fever, herpes, sore throat and other chronic infectious diseases. It grows abundantly in southeastern Asia, India, Sri Lanka, Pakistan and Indonesia – but cultivated extensively in China, Thailand, East and West Indies and Mauritius (Sandberg, 1994). In Chinese medicine, A. paniculata is used to rid the body of heat as in fevers and to dispel toxins from the body. In Scandinavian countries it is commonly used to prevent and treat common colds. The plant was introduced into Nigeria more than a decade ago and thrives in home gardens.
Studies have confirmed that the plant has a broad range of pharmacological effects: analgesic, anti-inflammatory, antibacterial, antipyretic, antithrombotic (blood clot preventative), antiviral, cardio-protective, depurative (cleans and purifies the system, particularly the blood), digestive, expectorant, hepatoprotective, hypoglycemic, immune enhancement, laxative, sedative and vermicidal (kills intestinal worms).  Cancer cell-killing ability of A. paniculata was demonstrated against human epidermoid carcinoma of the skin lining and against lymphocytic leukemia (Talukdar and Banerjee, 1968).

Aloe vera (Liliaceae/Asphodelaceae)
The healing properties of Aloe vera have been known for thousands of years. The ancient Egyptians believed that aloes had magic healing power and so assigned them to royal status second only to the Pharaoh. The plant was well known not only to the Egyptians but also to Roman, Greek, Indian and Arab cultures. Arab traders introduced the use of aloe to other parts of the world. Jesuit missionaries planted it around settlements in the New World starting probably in Barbados in 1590. Legend has it that the Indians of both Central America and Mexico were so amazed at Aloe vera’s ability to relieve so many ailments – cough, abscess, arthritis, cataracts, diabetes, genital herpes, gangrene, stomach pain, varicose veins, leg ulcers, etc., that they called the plant with its long leaves reaching towards heaven “The Hand of God” (Dublin, 1999).
Aloe vera is widely used in parts of the world for digestive problems, as laxative, wound healing, haemorrhoids, burns and scalds, psoriasis, eczema, scabies, ulcers and asthma. Aloe ferox widely distributed along the eastern parts of South Africa, is the country’s contribution to world medicine. A commercial laxative medicine, Cape aloe is the dried bitter yellow juice which exudes from just below the leaf surface.

Ageratum conyzoides (Asteraceae):  Goat weed
Ageratum conyzoides is widely used in traditional medicine by various cultures worldwide. In India it is used as a bacteriocide, antidysenteric and antilithic and in Asia, South America as bacteriocide. In Central Africa the plant is used to treat pneumonia but the most common use is for treating wounds and bones. In Cameroon and Congo, traditional use is for the treatment of fever rheumatism, headache and colic. Traditionally, the plant is extensively used in Brazil, where aqueous leaf extracts or whole plants are used to treat colic, colds, fevers, diarrhoea and rheumatism (Ming, 1999).
Pharmacological investigations have shown that plant extracts had activities against Staphylococcus aureus, Escherichia coli and  Pseudomonas aeruginosa. Analgesic action in rats was also reported. (Bioka et al., 1993). Aqueous extract of the whole plant showed clinical control of arthritis, as there was decrease in pain and inflammation after treatment for a week (Matos, 1988).

Ptychopetalu molacoides (Olacaceae): Muirapuama
This small tree is a native of the Brazilian Amazon and other parts of the Amazon rainforest. The indigenous people use the stem bark and root from young plants to treat neuromuscular problems, the root-and-bark tea taken to treat sexual debility, rheumatism, cardiac and gastrointestinal weakness. It has been listed in the Brazilian Pharmacopoeia since 1950 as a neuromuscular tonic for weakness and paralysis, chronic rheumatism sexual impotency and central nervous system disorders.
The herb was taken to England by early explorers and has since been listed in the British Herbal Pharmacopoeia for dysentery and impotence. Similarly, the herb is used in Germany, parts of Europe and the United States for impotence, depression, menstrual cramps and central nervous disorders. Researches on biological activities and clinical tests have confirmed its effectiveness in treating sexual impotence and disorders of the nervous system. Extracts of the plant have adaptogenic, analgesic,antistress and beneficial effects on the Central Nervous System.

Harpagophytum procumbens (Pedaliaceae): Devil’s Claw
This is a weedy perennial plant found in sandy places in the north-western South Africa, Namibia and Botswana. The common name ‘Devil’s claw’ is derived from the fruits which have numerous long arms with sharp hooked thorns as well as two straight thorns on the upper surface. The fruit is dispersed by animals. The thick fleshy roots are sliced, dried and used as medicine.
Devil’s claw has a long history of use traditionally to treat rheumatism  and arthritis and as a general health tonic. It is also used for digestive disorders and lack of appetite. Ointment made from the root is applied to sores, ulcers and boils. (Watt and Breyer-Brandwijk, 1962).
The fruits are the most commonly used in magic and medicine. Owing to the shape of the fruit and in accordance with the doctrine of signature, the Ghanaian local names refer to the fruit as “hanging breast”. Many African herbalists believe that since the shape of the fruit is like a breast that is the signature, that it will be useful for gynaecological problems. In Senegal, young girls before puberty are given a decoction of the ripe fruit to promote ample development of their breasts. In Ivory Coast the breasts are massaged with ointment of the fruit pulp to increase milk flow. Powdered fruit is prepared as a poultice for treatment of possibly mastitis or breast cancer.  In South Africa, the dried fruit is powdered and used as a dressing for ulcers, sores, syphilis and is topically applied for rheumatism. Antimicrobial activity has been demonstrated. Fresh fruit should not be eaten as it is strongly purgative and causes blisters in the mouth and on the skin. Unripe fruit is said to be poisonous.

Medicinal Plants as Food and as Medicine
Hippocrates, the father of modern medicine counselled, “Let your food be your medicine, let your medicine be your food”. Examples of plants which double as food and medicine are:

Persea gratissima (Lauraceae):  Avocado
Avocado is a native tree of Central America where it was cultivated as a staple food by the Aztecs and Mayas of Mexico, long before the Spanish explorers first saw it early in the 16th Century. The fruit was taken to southern Spain in 1601 and got to Mauritius in 1780. From there it spread to Asia mostly in the mid-19th Century and now found in most tropical and subtropical countries of the world.
In ancient times avocado pulp was used as a pomade to stimulate hair growth, and to help heal wound. Native Americans treated dysentery and diarrhoea with avocado seeds.  In Mexico and Central American countries, infusion of the leaves are taken for their digestive and anti-flatulent properties.
Although avocado contains more fat than any other fruit except olive its fat is mostly monounsaturated fat (oleic acid) which helps to protect good HDL cholesterol while wiping out the bad LDL cholesterol that clogs the arteries. Avocado oil is good for eczema, psoriasis, dandruff and hairlessness (Cawood, 2002). Eating avocado fruit helps to lower blood pressure because of its high potassium content. Leaf decoction has sedative effect. For centuries Chinese physicians have prescribed the fruit juice for colic and chills in the stomach. The Japanese employ the same remedy to treat ulcer in the intestines (Bakhru,1999). Bark decoction is taken in Congo (Brazzavile) for cough and in Gabon for chest pain (Burkill, 1985).

Allium sativum(Liliaceae): Garlic
Garlic is believed to have originated in Central Asia, and was known to the Chinese as early as 3,000 BC. It was cultivated in ancient China, Egypt, Greece and Rome as food and as medicine. It has since spread to all parts of the world.
Traditionally, garlic has been used for ailments such as asthma, leprosy, bronchitis, cough, piles, intestinal worms and arteriosclerosis. The health building qualities of garlic has been recognized for centuries all over the world. Khnoum Khoufouf, the builder of one of the oldest pyramids (4,500 BC), decreed that all his workers should take garlic every day so as to maintain their health and strength. In Ayurveda, a decoction of garlic boiled in milk is considered a wonderful drug for tuberculosis.
Garlic is one of the best remedies for lowering high blood pressure. In Russia and Britain garlic is recommended for treatment of rheumatism. In Japan, garlic extract tested on patients with lumbago and arthritis produced beneficial effects; probably due to its anti-inflammatory property. It has been used effectively for skin disorders e.g. pimples, wounds and ulcers as well as sexual debility. Studies in China, Canada and United States have linked garlic with anti-cancer properties. France is supposed to have one of the lowest incidences of cancer due to large amount of garlic consumed there. Garlic eaters in Bulgaria are virtually free from any cancer. In fact Hippocrates had prescribed eating of garlic for uterine tumors between the 4th and 5th centuries BC. Also, folk literature from India (circa 450 AD) speaks about garlic as a cure for abdominal tumors (Dublin, 1999). Antiglycaemic, antibiotic, antidiuretic, anti-inflammatory, anti-cancer, analgesic, antiatheromatous, diaphoretic and anthelmintic properties have been confirmed.

Zingiber officinale (Zingiberaceae): Ginger
Ancient Chinese sailors chewed ginger to prevent seasickness. Ancient Indians adopted the same practice and introduced it to Arab traders who took the rhizome to ancient Greece. The Greeks ate ginger wrapped in sweetened bread after big meals to settle their stomachs. Ginger was introduced into West Indies (Jamaica), Africa, Brazil and other warmer parts of the world. People from all over the world have relied on ginger to aid digestion, improve circulation, calm nausea and soothe headaches and other pains. Apart from the prevention of motion sickness which researchers have confirmed, clinical studies have shown ginger to be effective in controlling post-operative nausea. For thousands of years, herbalists have prescribed ginger for heartburn, bloating and remedy for poisoning. Researchers in Denmark suggest that ginger could relieve everyday muscle pain and migraines. In South Africa dried rhizomes or extracts of ginger are mainly used as stomachics and tonics to treat indigestion, flatulence and nausea. In Northern Nigeria ginger “chita” is consumed a lot for different health problems especially severe cold and catarrh.

Doctrine of Signatures
In ancient times people must have experimented with plant cures. But how would they know what plant to use? Probably in their search for an answer to this question they developed the concept of the plant “signature”. As a spiritual philosophy based on their belief that plants were created on earth for the good of humankind, the key to human use of plants was hidden in the form i.e. the signature of the plant itself. Therefore, by closely looking for the label from God or intuitively one would recognize the uses of the plants for the benefit of life. This line of reasoning is in obedience to the injunction of Allah that the plants shall be signs for the people who can reason and who can understand (Glorious Quran 13:4).
Signature plants were probably first recognized in ancient China where they correlated plant features to human organs e.g. red and bitter = heart, green and sour = liver, yellow and sweet = spleen.
In Europe signature plants emerged during the Middle Ages. The most famous advocate of signature of plants was the surgeon and philosopher, PhilippusAureolus Theophrastus Bombastus von Hohenheim, a Swiss citizen who later adopted the Latin name Paracelsus. He published the theory of the Doctrine of Signatures in the 16th century and travelled throughout Europe to Asia and Egypt curing people with herbs. The Doctrine was well developed during the European Renaissance as the concept paralleled the widespread belief in an over-all unity of Nature. Examples are: 1. Ginseng root shaped like a man’s body hence used as a general human panacea.

Adansonia digitata (Ose) has massive structure. To enhance physical development of a child, bathe the body with the bark decoction. If the head is bathed with the water extract, it becomes enlarged.

Herbal Medicine, Scientific Evidences and Nigeria
Undoubtedly, traditional medicine and the practitioners among the Nigerian ethnic groups have from time immemorial impacted positively on the health care delivery systems in Nigeria. Being most familiar with Yoruba herbalism, examples of medicinal plants used by the people and which have scientific support are:
1.    Ocimum gratissimum (Efinrin) is used for diarrhoea by bruising the leaves in water, straining and drinking the extract. Scientific study has shown that the plant is rich in a volatile oil which contains up to 75% thymol – an anti microbial agent. The anti-diarrhoea effect is attributed to thymol. (El-said et al., 1969).
2.    Garcinia kola (Orogbo):  Yoruba people associate regular consumption of bitter kola to longevity. Workers have found that extracts of the seeds possess remarkable anti-hepatotoxic and hence hepatoprotective properties. Iwu et al. (1990) produced Kolaviron, a defatted alcoholic extract of the seeds which they found to prevent liver damage in experimental animals challenged with different poisons. Kolaviron, an antioxidant of Garcinia kola, protects the body against oxygen-free radicals. The multiple physiological functions of liver underscore the fact that a healthy liver in all probability will enhance wellness and longevity.
3.    Senna alata (Asuwon pupa): The leaves of this plant are boiled or macerated and taken with cold pap as a mild purgative. The leaves are also used for the treatment of some skin diseases like eczema. The purgative principles in the plant are anthraquinones and the leaves have antimicrobial activity (Iwu, 1993). These findings justify the ethnomedicinal uses.
4.    Rauwolfia vomitoria (Asofeyeje) is used in the treatment of mentally – disturbed patients. The patient is given a root decoction or soaked in locally distilled gin or administered in powdered form and taken with maize pap. The alkaloid reserpine which possesses hypotensive, antihypertensive and sedative properties have been reported in the plant, thus the use for mentally disturbed patients can be justified scientifically (Iwu and Court, 1982).
5.    Ageratum conyzoides (Imi-esu): This is a popular herb for treating bruises, wounds and sores among the Yoruba. Research has shown that the plant has some wound – healing promoting substances like conyzagenin, and 5 –methoxynobiletin (Adesogan, 1987).
6.    Chewing sticks: Although the use of chewing sticks is popular in Africa, the favourite ones used by the Yoruba are Zanthoxylum zanthoxyloides (Orin ata), Massularia acuminata (PakoIjebu), Terminalia glaucescens (Orin idi), Anogeissus leiocarpus(Orin Ayin), Garcinia kola (Orogbo) and Vernonia amygdalina(Ewuro). These plants have been shown to have antimicrobial activities against oral microbial flora albeit in varying degrees: Zanthoxylum > Anogeissus > Terminalia > Vernonia > Massularia > Garcinia. The chewing sticks destroy microbes present in the mouth and have also been reported to contain not only fluoride but also silicon tannic acid, sodium bicarbonate and other natural plaque-inhibiting substances. (Akpata  and Akinrimisi, 1977; Ogunbodede, 1991).
7.    Momordica charantia (Ejirin): This climbing plant is used for managing diabetes and for constipation, skin problems and diarrhoea. Laboratory studies have established the hypoglycaemic effects (Day et al., 1990). Aqueous extracts of the root and leaves have been shown to have appreciable antibacterial activity against the bacteria Escherichia coli, Staphylococcus aureus, Salmonella spp. and Bacillus subtilis.
8.    Capsicum frutescens (Ata ijosi): In Nigeria the Yoruba are known to relish foods with lots of pepper owing to their belief that pepper is essential for healthy living, meaning Pepper promotes longevity.  This statement has been supported by a scientific investigation abroad which indicated that pepper is cardiotonic. It helps in preventing heart disease, atherosclerosis (hardening of the arteries). Apart from enhancing blood circulation, it contains antioxidant compounds.
9.    Terminalia cattapa (Indian almond): Decoction of reddish brown fallen leavesis prescribed to sickle cell disease patients. Study showed that ethanol extract of the fallen leaves exhibited the highest anti-sickling activity, while extracts of other leaves still attached to the tree caused lysis of the red blood cells. This result justified the choice of fallen leaves in traditional medicine (Moody et al., 2002).

Conclusion
Over the centuries, every culture population or race had its own understanding and ways of healing, predominantly using medicinal plants. The nature of plants found in the immediate environments of the people, their customs and beliefs determined the form of healing which varied across countries and races. However, as human development characterized by population range expansion across the globe progressed, it afforded interactions between cultures. Concomitantly, medicinal plants also spread or were introduced to areas other than their places of origin. Many of the herbal remedies of old have since been adopted and adapted by conventional Western allopathic medicine, simply due to the fact that they are effective. Thus the inestimable value of medicinal plants to health care systems in the world has increasingly become appreciated. As an example, the United States of America has been using herbal drugs up to 25% on average every year for decades in prescriptions dispensed in its public pharmacies.
Regrettably, the therapeutic potentials of medicinal plants are yet to be fully harnessed in many populations especially in Africa as the knowledge of herbalism is still held in secrecy by the indigenous people. It has been estimated that a country like Nigeria uses less than 5% of plant-derived drugs in its health care. This is an underutilization of medicinal plant resources. Ironically, the medicinal plants are disappearing or endangered due to indiscriminate and unsustainable collection from the wild and loss to prevailing climatic changes. In light of this, it is time for African countries, especially Nigeria, the largest black nation in the world, to act fast in remedying the situation. Time is ticking fast! Time is not on our side!

References
Adesogan, E.K. (1987). Illumination, wisdom and development through Chemistry.University of Ibadan, Inaugural Lecture. 37pp.

Akpata, E. S. and Akinrimisi, E. (1977). Antibacterial activity of extracts of some African chewing sticks.Oral Surgery, Oral Medicine, Oral Pathology, 44 (5): 717-728.

Bakhru, H.K. (1999). Foods that heal: The natural way to good health. Orient Paperbacks, Delhi 215pp.

Bioka, D., Banyikwa, F.F. and Choudhuri, M.M. (1993). Analgesic effects of a crude extract of Ageratum conyzoides in the rat. ActaHort. 332: 171-176. Council (USA).

Burkill, H.M. (1985). The useful plants of West Tropical Africa.Royal Botanic Gardens, Kew.2nd edition, Vol. 3 Families J-L, 221pp.

Cawood, F.W. (2002). Eat and Heal. FC8A Peachtree City.
Daes, Erica-Irene (1993). Study on the protection of the cultural and intellectual property of indigenous peoples. UNDOCE/CN, 4/Sub 2/1993/28.

Dopamu, P.A. (1977). The practice of magic and medicine in Yoruba traditional religion.Unpublished Ph.D Thesis, University of Ibadan.

Dublin, R. (1999). Miracle food cures from the Bible. Reward Books, Paramus, NJ, USA. 410pp.

Farnsworth, N.R. and Soejarto, D.D. (1985).Potential consequence of plant extinction in the United States on the current and future availability of prescription drugs.EconomicBotany, 39: (3) 231-240.

Farnsworth, N.R. (1988). Screening plants for new medicines, Pages 83-977. In: E.O. Wilson, ed. Biodiversity. National Academy Press, Washington, DC.

Farnsworth, N.R., Soejarto, D. D. and Bingel, A. S.  (1986). Medicinal plants in therapy. Bulletin of the World Health Organization. 63 (6):  965-981.

Foley, S. (2006).Pharmacy in ancient China.College of Pharmacy, Washington State University U.S.A. htt://toxipedia.org/display/toxipedia/shentnung.

Iwu, M.M. (1993). Handbook of African Medicinal Plants. CRC Press, London. 435pp.

Iwu, M.M. and Court, W.E. (1982). Stem bark alkaloids of Rauwolfia vomitoria, Journal of Medicinal Plant Research (Planta Med.) 45, 105-111.

Iwu, M.M., Igbok, O.A., Okunji, C.O. and Tempesta, M.S. (1990). Antidiabetic and aldose reductase activities of bioflavanories of Garcinia kola.J. Pharm. Pharmacol. 42, 290-2.

Komolafe, K. (1998). Impact of traditional medicine on the healthcare delivery systems in Nigeria. Sunday Punch, Page 6, March 8, 1998.

Lust, J. (1979).The Herb Book. Benedict Lust Publications, U.S.A. 660pp.

Mander, M. (1998).Marketing of Indigenous Medicinal Plants in South Africa: A case study in Kwazulu – Natal FAO, Rome, 151pp.

Masood, E. (1997). Medicinal plants threatened by over-use. Nature Vol. 385, 3, February, 1997.

Matos, F.J.A. (1988). “Plantasmedicinais: Boldocolonia e mentrasto O povo Univ. Aberta, Fortaleza, 27, Jan. 1988 p. 2-3.

Ming, L.C. (1999). Ageratum conyzoides.A tropical source of medicinal and agricultural products p. 469-473. In: J. Janick (ed.). Perspectives on new crops and new uses.ASHS Press, Alexandria, VA.

Moody, J.O., Segun, F., Aderounmu, A. and Omotade, O. (2003).Antisickling activity of Terminalia cattapa leaves harvested at different stages of growth. Nig. Journal of Natural Products and Medicine 7, 30-32.

Murray, M.T. (2002). The pill book guide to natural medicines.Bantam Books, New York. 1074pp.

Ogunbodede, E. (1991). Dental care: The role of traditional healers. World Health Forum 12(4), 443-444.

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Shellard, E.J. (1979). The significance of research into medicinal plants. In: Proceedings of African Medicinal Plants (ed. Sofowora, A.). University of Ife Press pages 98-111.

Talukolar, P.B. and Banerjee, S. (1968). Studies on the stability of andrographolide.Indian J. Chem. 6: 252-254.

Ting, L. H. (1987). Developing new drugs related to Chinese medicinal plants. Newsletter of Medicinal and Aromatic Plants FAO, pp 46-58.

Trease, G.E. and Evans, W.C. (1986). Pharmacognosy. Macmillan Publishers Ltd., London.

Van Wyk, B.E., Van Oudtshoorn, B. and Gericke, N. (1997).Medicinal Plants of South Africa. Briza Publications, S. Africa 304pp.

Vietmeyer, N. (1992). Neem: A tree for solving global problems. National Academy Press, Washington D.C. 141 pages.

Wagner, H. (1988). An interview of Prof. H. Wagner, Herbal Gram, 17, 16-17.

Watt, J.M. and Breyer-Brandwijk, M.G. (1962).The Medicinal and Poisonous Plants of Southern and Eastern Africa.2nd Edition. Livingstone. London.

FULFULDE NAMES OF PLANTS OF MAMBILLA PLATEAU, NIGERIA

Department of Botany, University of Lagos, Lagos, Nigeria.

Received 28th August, 2013; accepted 2nd June, 2014

ABSTRACT

A rapid survey of plants of Mambilla Plateau, Nigeria, was carried out and a compilation of their Fulfude names was made. A total of 183 plant species comprising 2 pteridophytes and 181 angiosperms was documented. The compilation revealed elements of folk taxonomy in naming of plants in Fulfude language. It is expected that this report will assist field workers in the area for on-the-spot identification of plants in the field and facilitate further research works.

 

INTRODUCTION

According to Hepper (1966), the first known botanical collection from Mambilla Plateau was by Mr. and Mrs. Gates in 1947, followed by LAtilo and Daramola in January 1955. In 1958, a more comprehensive collection was made by Hepper and Daramola in conjunction with J.W.F. Chapman (Chapman and Chapman, 2001). After these, only a few collections have been made from the forest. On Mambilla Plateau, the most diverse forest is Ngel Nyaki comprising over 146 vascular plant species many of which are trees, and (near-) endemic to the Afromontane Region of White (1983) (Dowsett, 1989). In 1989, a summary of forest structure and species composition of Ngel Nyaki was published by Dowsett with particular reference to its size and floral diversity. However, aside from work done by Chapman (2008), relatively little is known about the taxonomy and ecology of Mambilla Plateau.

Generally, in vegetation studies, on-the-spot field identification of plant species is crucial whether it is taxonomic or ecological work. This is due to the fact that it facilitates data collection and organization and it also saves time. However, the paucity of handy lists of local flora has been a challenge to biodiversity studies in Nigeria especially in pioneering work on sites which had not been previously explored. Also, local flora exists only as a part of the general texts scattered over several pages of the flora of west tropical Africa. In such situations, it could be a challenging experience gathering data on plants and animals which cannot be identified by names. Experience from working in this area has shown that this challenge can be overcome by consulting with medicinal plant practitioners, farmers, pastoralists and village elders for indigenous names of collected plants in the field while the corresponding scientific names are obtained from a reputable herbarium.

The book published by Gbile (1984) has been found to be useful in the field when working in Hausa- and Yoruba-speaking areas. This study was conducted to compile the Fulfulde names of plants found in Mambilla Plateau with their corresponding scientific names so that identification of these plants in the field can be made easier for workers in the area.

MATERIALS AND METHODS

Study Site

Mambilla Plateau (6°47’43″N, 11°11’58″E) is located in Sardauna Local Government Area of Taraba State, Nigeria (Figure 1 & 2). The plateau is an extension of Bamenda highlands (Cameroon) into the Nigerian territory. It is the highest plateau in Nigeria with an average elevation of about 1,500 m above sea level in the south-eastern corner of Taraba State and may reach 1,800 m above sea level in Chabbal Hendu peaking at Gangriwal (2,149 m above sea level) which is the highest mountain in Nigeria. It covers an area of 3,100 km2 (Chapman, 2008) overlying basement complex rocks with some tertiary basalts derived from trachytic lavas occurring in some places e.g. Nguroje (Wikipedia, 2013). It has a rough and rugged terrain with rolling hills interspersed with streams and river valleys and deep gorges. Hill tops are covered by grasses and rocky outcrops. The high grassland is grazed by cattle while the stream valleys are covered with sub-montane forests which provide habitats for a variety of wildlife. The climate is temperate –like with a temperature that never reaches 250 C. There are two distinct seasons. The rainy season runs from March to November (Ihuma et al., 2011) with a mean annual rainfall exceeding 1,850 mm per annum while dry season runs from December to February.

 Sample Collection

Plant specimens were randomly collected from Ngel Nyaki Forest Reserve (NNFR), and forests around Inkiri-Anterre, Yerimaru, Yelwa and Zongo Ajiya. The plants were tagged, pressed and labelled. Initial identification of the plant samples was done using Flora of West Tropical Africa (Hutchinson and Dalziel, 1954, 1958) and Nigerian Trees (Keay et al., 1960, 1964). The scientific names of the identified plants were recorded. All the plants collected were taken to users who are familiar with the local flora (a team of villagers comprising the chief, his deputies, traditional herbalists, farmers, hunters and an interpreter) owing to the fact that they utilize the plants for various cultural purposes. Focused discussion sessions were held to record the Fulfulde names of the plants collected. Fulfulde names of the plants were recorded and a list of Fulfulde names of both identified and unidentified specimens was compiled. The specimens were then taken to the herbarium at Forestry Research Institute of Nigeria, Ibadan for further authentication and voucher specimens were deposited. A comprehensive list of both scientific and Fulfulde names of the plant specimens was then prepared.

RESULTS AND DISCUSSION

A total of 183 plant species distributed in 59 families were encountered during the sampling period (Table 1). Two of these, Cyathea dregei Kunze and Pteridium aquilinum (L.) Kuhn, are pteridophytes while others are spermatophytes. Of all the angiosperm species, the family Fabaceae was the most abundant constituting 21.86% of the samples while the family Moraceae and Combretaceae comprised 5.46% and 4.92% of the total species, respectively. Table 2 shows the distribution of species by family.

Naming of plants in Fulfulde language reflects an aspect of folk taxonomy (Olorode, 1984) in which characteristics other than floral and morphological characteristics are used in naming plants. For instance, Cola Schott & Endl. species are generally referred to as ‘goro’ but Garcinia kola Heckel is specifically called ‘namijin goro’ (i.e. male cola). Another example is seen in the genus Dioscorea L. in which all the different species represented in the study sites are known by the same name (‘mbulunji’) in Fulfulde language.

ACKNOWLEDGEMENTS

We thank Alhaji Sani Mohammed (Galadima Nguroje) for facilitating the study by introducing us to the village heads of Inkiri-Anterre, Zongo Ajiya, Yelwa and Yerimaru and for his hospitality all the time we visited him. We appreciate Buba Yerimaru who served as transportation officer with his motorcycle and as an interpreter. We are also grateful to all the communities visited. This study was supported by University of Lagos Central Research Grant No: M2010/07.

REFERENCES

Chapman, H. (2008). The Nigerian Montane Forest Project. Tropinet, 19(1):7-9.

Chapman, J. D.  and Chapman, H. M. (2001|). The Forest Flora of Taraba and Adamawa States, Nigeria: An ecological account and plant species checklist. Department of Plant and Microbial Sciences, University of Canterbury, Christchurch, New Zealand. 237pp.

Dowsett, R. J. (1989). A preliminary natural history survey of Mambilla Plateau and some lowland forests of Eastern Nigeria. Tauraco Research Report No. 1: 1-56.

Gbile, Z. (1984). Vernacular names of Nigerian plants: Yoruba. Forestry Research Institute of Nigeria, Ibadan. 101pp.

Hepper, F. N. (1966). Outline of the vegetation and flora of Mambilla Plateau. Bull. IFAN 28: 91-127.

Hutchinson, J. and Dalziel, J. M. 1954. Flora of West Tropical Africa. Vol. 1. The Whitefriars Press Ltd. London.

Hutchinson, J. and Dalziel, J. M. (1958). Flora of West Tropical Africa. Volume 1, Part 2. Crown Agents for Overseas Government and Administrations, Millbank, London. 828pp.

Ihuma, J. O., Chima, U. D. and Chapman, H. M. (2011). Tree species diversity in a Nigerian montane forest  ecosystem and adjacent fragmented forests. ARPN Journal of Agricultural and Biological  Science, 6(2):17-22.

Keay, R. W. J., Onochie, C. F. A. and Stanfield, D. P. (1960). Nigerian Trees. Vol. 1. Department of Forest    Research, Ibadan, Nigeria.

Keay, R. W. J., Onochie, C. F. A. and Stanfield, D. P. (1964). Nigerian Trees. Volume II. Department of Forest   Research, Ibadan. 495pp.

Olorode, O. (1984). Taxonomy of West African Flowering Plants. Longman Publishers. 158pp.

White, F. (1983). The Vegetation of Africa. UNESCO, Paris. 356pp.

 

LEGENDS TO FIGURES
Figure 1: Map of Nigeria showing the relative position of Taraba state.
Figure 2: Map of Mambilla Plateau showing the study sites.

DISTRIBUTION AND POLLEN MORPHOLOGY OF SOME SPECIES OF GREWIA LINN. IN NIGERIA

Department of Botany, University of Ibadan, Ibadan, Nigeria

Received, 9th June, 2014; accepted, 15th December, 2014

*Author for correspondence

ABSTRACT
The distribution and pollen morphology of some Grewia species in Nigeria were studied.  These species occupy a wide range of habitats; six species (G. malacocarpa Mast., G. barombiensis K.Schum., G. brunnea K.Schum., G. coriacea Mast., G. hookerana Exell & Mendonca. and  G. oligoneura Sprague) are found in the lowland rain forests of the Southern parts of the country, eight species (G. barteri Burret, G. bicolor Juss., G. cissoides Hutch&Dalz., G. flavescens Juss., G. lasiodiscus K.Schum, G. venusta Fresen., G. tenax (Forsk) Fior and G. villosa Willd) were present in the Guinnea savannah dry land of the Northern part of the country while two species (G. carpinifolia Juss. and G. mollis Juss.) occur in both ecological zones in the country.  Grewia bicolor Juss. is the only species restricted to the high mountains of the Northern part of the country among the Savannah species.  The pollen results reveal two pollen types: the Microcos and Grewia types. The Microcos pollen type consists of the following characters: tricolporate with short colpi, small-sized pollen of 17.5µm-21.0µm x 15.0µm-20.0µm (PA x ED); sub-prolate to prolate shape; exine is thin (0.5 µm -1.50µm) and fine reticulations; lumina shape range from round, angular and elongated. Parent plants with this pollen type are natural to the lowland rainforest zone. The Grewia type consists of pollen, parent plants of which are natural to southern Guinea savannah, as well as G. mollis and G. carpinifolia which occur in both ecological zones.  The pollen grains of the Grewia type are mainly tricolporate but G. venusta is three to four colporate; they possess long colpi; are medium (30.0µm) to large (55-67.5µm x 57.5-82.5µm) in size. Pollen shapes include sub-prolate, prolate and oblate-spheroidal; exine is generally thick (0.87µm-3.0µm), exine pattern is reticulate, coarse reticulate and sometimes striato-reticulate; lumina shapes are distinctly polygonal or irregular, and contain bacules. These characters are discussed in relation to the taxonomy of the genus in Nigeria.

Keywords:  Grewia, Distribution, Palynology, Nigeria.
Introduction
Grewia Lin. (Tiliaceae) consists of about 280-300 species well represented in Tropical Africa, Asia and Australia (Cronquist, 1981; Chung et al., 2003).  In West Africa, 17 species occur; sixteen of these are recorded for Nigeria (Hutchinson and Dalziel, 1954).  Nigeria is a major centre of diversity for Grewia in West Tropical Africa (Czarnecka et al., 2006). Six species are known to occur in lowland rainforest, eight
in dry woodland savannah while two species occupy both ecological zones (Hutchinson and Dalziel, 1954).  The lowland rainforest species are distinguished by presence of panicle inflorescence while the savannah
species are distinguished by the presence of cymose inflorescence (Burret, 1926; Hutchinson and Dalziel, 1954; Bayer and Kutbitzki, 2003; Cowie et al., 2011).
Taxonomic studies of the genus have been carried out by many authors using morphological, anatomical, palynological and molecular data (Burret, 1926; Chattaway, 1934; Cronquist, 1981; Chang and Miau, 1989; Watson and Dallwitz, 1992; Alverson et al., 1997; Thorne,1998; Bayer et al., 1999; Judd et al.,
2000; Bayer and Kutbitzki, 2003; Chung et al., 2003).   In particular, morphological data have provided taxonomic information at family, subfamily, generic, subgeneric and species level for different groups of plants (Stuessy, 1990).  Perveen et al. (2004), highlighted the morphological features of the pollen grains of Grewia in the study of the family Tiliaceae as follow: 3 colporate aperture; rugulate/finely reticulate to coarse reticulate tectum and prolate or subprolate pollen shapes.  El-Husseini  (2006) reported the importance of the colpi ends, pollen size, muri ridge shape, equatorial view and size of lumina and stated that spheroidal pollen grains are peculiar to the Grewia type.  Perveen and Qaiser  (2007) recognized two pollen types: Corchorus depressus type and Corchorus tridens type in their study of the subfamily Grewioideae.  Pollen data have been used effectively at generic and subgeneric or sectional levels on numerous occasions within tribes (Vezey et al., 1994).  Although considerable works have been done on the species of the genus, data on Nigerian species are sparse.   The aim of this study was to investigate the ecological distribution and pollen morphology of some species of Grewia in Nigeria with a view to providing additional information for the delimitation of the species.  The results would provide a better understanding of the relationship between and among the species in Nigeria.

Materials  and Methods
Specimens of Grewia species were collected during field trips to different locations within Nigeria.  Flower buds and floral specimens were preserved in 50% ethanol.  Voucher specimens were made for all collections using standard herbarium procedures and deposited in the Herbarium of the Department of Botany, University of Ibadan, Ibadan, Nigeria (UIH).  The various locations visited are shown in Figure 1.  The herbarium specimens were studied for their distributional data in the Forest Herbarium Ibadan (FHI), Ibadan, and the University of Ibadan Herbarium (UIH).

Distribution and Ecology
Data obtained during the study of specimens in the herbarium and notes taken during field trips provided useful information for the distributional ecology of the genus in Nigeria.

Pollen Morphology: Flower and floral buds preserved in 50% ethanol from the field collections and in the case of species that were not collected from the field, flower and floral buds from well identified herbarium specimens were assessed for pollen morphology using acetolysis method according to the procedures described by Erdtman (1952, 1960) and Ayodele (2005).  The floral buds were crushed with a glass rod in centrifuge tube. Three millilitre of freshly prepared acetolysis mixture (9 parts acetic anhydride to 1 part concentrated Tetraoxosulphate VI acid) was added to the content in the tubes.  The content was heated in a water bath from 700C to boiling point, stirring occasionally.  The centrifuge tubes and content were left in boiling water for 3 minutes and then centrifuged at 4,000 r.p.m. for 5 minutes while still hot.  The supernatant was decanted into Acetolysis waste bottle.  Some water was then added to the sediments in the tubes and shaken vigorously using a whirl mixer.  Few drops of methylated spirit were added to remove the foam formed and centrifuged again.  The supernatant was decanted.  The washing with water and centrifuging were repeated four times.  Fifty per cent glycerine was added and left to stand for two hours.  The tubes were shaken vigorously using a whirl mixer and centrifuged at 4,000 r.p.m. for 10 minutes.  The supernatant was decanted off finally and the tube was inverted over filter paper and left overnight.  One hundred per cent glycerol was added to the tubes and shaken.  This was then poured into labeled storage vials.  The pollen grains were mounted in unstained glycerin jelly and observations were made with a Fisher scientific illumination microscope at (E 40; 0.65) and oil immersion (E 100; 1.25) using 10x eye piece. The measurement was based on 20 readings from each specimen. Photomicrographs were taken using Leica CME with Digital Microscope Eyepiece attachment and Photo Explorer 8.0 SE Basic software. Terminologies used were in accordance with Erdtman (1952), Moore et al.  (1991) and Perveen et al. (2004).  All slides are deposited in the Herbarium of the Department of Botany, University of Ibadan, Ibadan, Nigeria.

Results
Distribution and Ecology
Nigerian lowland rainforests and Guinea Savannah provided suitable habitats for the sixteen species in the genus Grewia. Six species, G. malacocarpa, G. barombiensis, G. brunnea, G. coriacea,    G. hookerana and G. oligoneura are confined to the high rainforest of the Southern part of Nigeria while eight species  (G. barteri, G. bicolor, G. cissoides, G. flavescens, G. lasiodiscus, G. venusta,   G. tenax and G. villosa) are found in the Guinea Savannah in the Northern part of Nigeria. Two species (G. carpinifolia and   G. mollis) occupy both ecological zones (Fig.1).  Some of the Nigerian species have narrow distributional range, e.g., G. villosa and G. tenax occur in the drier regions of Northern Guinea Savannah while  G. hookerana and G. oligoneura are found only in the South-West and south-East high forests, respectively (Table.1).   The 16 species studied are tentatively divided into three groups based on their ecological preferences viz: Group A: G. malacocarpa, G. barombiensis, G. brunnea, G. coriacea, G. hookerana and G. oligoneura  all distributed in the rainforest ecological zone; Group B:  G. barteri,  G. bicolor, G. cissoides, G. flavescens, G. bicolor, G. lasiodiscus, G. tenax,  G. venusta, G. villosa  distributed in the savannah zone and Group C consisting of  G. carpinifolia and G. mollis  found in both ecological zones.

Pollen analysis:
Two pollen types and 5 subtypes are recognized for the Nigerian species of Grewia based on light microscopy (Fig. 2.).

Grewia type
This is typical of the savanna species and the two species occupying both ecological zones.
The pollen grains are large to medium in size, isopolar and radially symmetrical; Polar view (30.0 – 67.5 µm); Equatorial diameter (18.8 – 82.5µm). Polar/Equatorial diameter ratio: 89 – 175% (Table 3);  oblate spheroidal, subprolate and prolate; Ambient circular, trilobed or triangular; 3-colporate except G. venusta 3-4 colporate (Table 2); Colpi long (20.0 – 45.0) µm, colpi ends acute, open or acute/open; Ora(endoaperture) lalongate, 6.0  -15.0µm broad,  rectangular; Sexine thicker than nexine; Nexine 0.50 – 2.5µm; Exine 1.06 – 2.75µm thick.  The pollen grains are elliptic, circular, oblong and oval in equatorial view; striate, reticulate or coarse reticulate sculpture with bacules; lumina small or large (0.90-13.1)µm in diameter, distinct; lumina shape polygonal to irregular, muri ridge entire or wavy, with or without bacules (Table 2& 3).

Three subtypes are recognized on the basis of pollen size, pollen class and exine sculpture.

Subtype I
G. mollis        pollen grains are large; oblate-spheroidal in shape; Ambient is circular while colpi are provided with acute ends; ora (endoaperture) are la-longate, 9.9 µm broad; exine pattern is coarsely reticulate; lumina are large (13.4 µm in width) and distinct; pollen is provided with baculate muri ridge (Table 2 & 3).

Subtype II
G.barteri, G. cissoides, G. carpinifolia, G. flavescens, G. lasiodiscus,  G. venusta.
Pollen grains medium; subprolate or prolate; Ambient circular (G. carpinifolia, G. flavescens and G. venusta),  triangular (G. barteri, G. bicolor, G. cissoides and G. lasiodiscus)); 3 or 3-4 colporate (G. venusta); colpi length 31.9- 34.2 µm (G. venusta) or  20.0 – 26.8 µm (G. flavescens, G. lasiodiscus, G. tenax), colpi ends acute/open (G. carpinifolia and G. lasiodiscus) and acute/ open or acute (G. cissoides, G. flavescens, G. venusta), Ora (endoapperture) rectangular; Exine pattern coarse reticulate and striate – coarse reticulate (G. flavescens), lumina and muri ridge baculate with wavy margin (Table 2 & 3).

Subtype III
Grewia bicolor, G. tenax and G. villosa
Pollen grains medium, prolate; Tricolporate; Colpi short (G. tenax) to long with ends acute/open or open (G. villosa); exine pattern reticulate rarely coarse reticulate (G. bicolor), not baculate, muri ridge wavy or entire (G. villosa) (Table 2 & 3).
Measurements (µm):    Range
Mean+ Standard Error
The group B taxa are restricted to the northern part of Nigeria.  Grewia barteri and G. venusta are the most widespread species in the savannah area  and are found in the North central, North west, North east and derived Savannah areas of Oyo State, South west Nigeria while  G. tenax and G. villosa were recorded only in Adamawa and Bornu States in the far North east of Nigeria.  Two morphologically similar species, G. bicolor and G. venusta, could be distinguished by the pollen morphology which differs from one another based on the presence of bacules on tectum, ambient, equatorial view and colpi ends. G. bicolor is mainly found in rocky areas on high mountains in the savannah vegetation in Niger, Kastina, Kaduna, Bauchi and Plateau States.  Ware (1990) noted that the occurrence of taxa in rocky areas represent not only edaphic seclusion from the rest of the genus but also a case of parallel evolution into similar but distinguished bedrock areas.  The present study corroborates‘ the xerophytic nature of the group B taxa as reported by Czarnecka et al. (2006).  In group C taxa G. carpinifolia and G. mollis occur widely across the South to the Northern part of Nigeria.  This conforms to the report of Brian and Standfield (1966) and Hutchinson and Dalziel (1954) on the occurrence of both species in the Savannah and the rainforest vegetation in Nigeria.
Grewia  hookerana and G. oligoneura are represented by one collection each from the Southern part of Nigeria in the Forest Herbarium Ibadan. The collection of both species was made in 1947 and 1946, respectively.  The field trip to Akila village, Ijebu east, Ogun State (South-West) and Ikom, Edo State (South-South) where both species were earlier collected did not produce any positive result. This might be an indication that the species are threatened in the country as the earlier sites of collections are presently Cocoa and Cola plantations.  Distribution data can show patterns of spatial isolating mechanism which could indicate the ecological basis for further isolation (Sundberg and Stuessy, 1990).   The distributional diversity observed in this study may have been due largely to variation in the climatic conditions of the different geographical zones in Nigeria.
The palynological evidence obtained from this study shows the naturalness of the species in group A as depicted by ecology with their subprolate or prolate, 3 colporate grains and exine sculpture (Plates 1A-H ).  The pollen morphology combined with geographical distribution and ecology of the group A taxa justify the separation of the group from other taxa in the genus. However, based on pollen morphology, G. carpinifolia is better placed in group B taxa than in group C due to similarities observed in its exine sculpture with that of G. flavescens.  Oblate-spheroidal and large pollen grain of Grewia mollis, distinguishes it from G. carpinifolia and the rest of group B taxa but similarity in exine sculpture justifies its placement in Grewia subtype I.  Although the pollen morphological characters of G. mollis are similar (closely related) to the pollen type recorded by Perveen and Qaiser (2007) for subfamily Tiliodeae, the lumen is baculate as against perforated reported by Perveen and Qaiser (2007). Striato-reticulate and baculate exine sculpture is diagnostic for G. flavescens contrary to reticulate and perforated tectum reported in the Egyptian species by El-Hussieni (2006).  The results obtained from the pollen morphology of the Microcos pollen type conform to the report of the light microscopic pollen morphological characters reported by Perveen  et al. (2004) in the World pollen and spore flora study of Malvaceae (s.l) but differs in colpi length which was 15.7-17.1µm as against 14.0-36.0µm reported by Perveen et al. (2004).  In the present study, it was noted that geographical distribution and ecology correlated with differences in the pollen morphology of the species studied and can be used as basis for the determination of the relationship of the taxa at various levels.  Blackmoore  (1982b), Suarez-Cervera et al.  (1995) and Pozhidaev (2000) stated that the importance of the different surface sculpturing of the exine has been next to the number and position of apertures in providing an amazing wealth of detailed taxonomic value.  The present study recognizes two pollen types and five subtypes in the Nigerian taxa of Grewia based on pollen size and exine ornamentation.  This study supports the separation of Microcos from Grewia as a distinct genus (Burret, 1926; Chattaway, 1934; Bayer et al., 1999; Chung et al., 2003).  Geographical, ecological and pollen morphological data of some species of Grewia in Nigeria have provided valuable characters for solving the species delimitation problems and are useful for the identification of the morphologically closely related species.  The characters presented in this study are consistent within species and distinguished between species in a group.

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INVESTIGATION INTO THE MICROORGANISMS ASSOCIATED WITH THE DETERIORATION OF AMARANTHUS HYBRIDUS (AMARANTH) (Linn) UNDER STORAGE.

Department of Biology, Ibrahim Badamasi  Babangida University, Lapai, Niger State.

Received, 9th August, 2014 ; accepted, 30th October, 2014

ABSTRACT
The incidence of microorganisms and deterioration in vegetables may be expected to reflect the sanitary quality of the processing steps and the microbiological condition of the raw product at the time of processing. This study investigated microorganisms associated with the deterioration of Amaranthus hybridus under two different storage conditions: refrigeration (4oC) and at room temperature (28oC). Fresh samples of Amaranthus hybridus were obtained from three different sources (farm, market and vegetable vendors) in Lapai town, Niger State, Nigeria. Microorganisms associated with vegetables sampled were isolated on Potato Dextrose Agar (PDA) and identified. The results showed that samples stored at room temperature deteriorated completely at the 5th day of storage while those stored at the refrigerating temperature remained fresh. The deterioration was marked by loss of green colour to mushiness of the vegetables and defoliation. The bacteria isolated were Escherichia coli, Bacillus subtilis, Pseudomonas aeroginosa, Proteus mirabilis and Staphylococcus aureus. The fungal isolates were Aspergillus niger, Aspergillus flavus, Mucor pusillus and Aspergillus fumigatus. The pH of the samples was found to increase with increase in number of days of storage at the two conditions of storage. The moisture content of samples stored at room condition increased and was found to have contributed to deterioration.

Key words: Amaranthus hybridus, deterioration, fungal and bacterial species

INTRODUCTION
Amaranthus hybridus L, popularly called “Amaranth or pigweed”, is an annual herbaceous plant,1- 6 feet high. The leaves are alternate, petioled, 3 – 6 inches long, dull green, and rough, hairy, ovate or rhombic with wavy margins. The flowers are small, with greenish or red terminal panicles. Taproot is long, fleshy red or pink. The seeds are small and lenti-cellular in shape; with each seed averaging 1 – 1.5 mm in diameter and 1,000 seeds weighing 0.6 – 1.2 g. It is rather a common species in waste places, cultivated fields and barnyards. In Nigeria, A. hybridus leaves combined with condiments are used to prepare soup (Oke, 1983; Mepha et al., 2007). In Congo, the leaves are eaten as spinach or green vegetables (Dhellot et al., 2006). It is called “Aleho” in Hausa, “Inine” in Igbo, “Tete abalaye” in Yoruba  (Fayemi, 1999).
Deterioration refers to any change in the condition of food in which the food becomes less palatable or even toxic. These changes may be accompanied by alteration in taste, smell or appearance (Effiuvwevwere,  2000).
The incidence of microorganisms in vegetables may be expected to reflect the sanitary quality of the processing steps and the microbiological condition of the raw product at the time of processing. For 100 years, vegetables contaminated in the field have been recognized as a source of human infection. Many of the viruses (Engle and Altoveras, 2000), bacteria and protozoan on vegetables which have caused food poisoning are derived from human faeces. However, pathogenic microorganisms of human origin may also be present in minimally processed vegetables as the minimal technological processing may be unable to remove the original contamination resulting from air, soil, water, insects, animals, workers, harvesting and transportation equipment. Certain fungal species such as Aspergillus, Fusarium, and Penicillium are commonly occurring filamentous fungi that grow in vegetables and their growth may result in production of toxins known as mycotoxins, which can cause ill-health in humans from allergic responses to immune suppression and cancer (Baiyewu et al., 2007). The ability of public health agencies to identify through enhanced epidemiological and surveillance techniques, raw vegetables as probable sources of infectious microorganisms has undoubtedly resulted in increased numbers of outbreaks.
Storage fungi can cause decrease of germination capability, loss in weight, discoloration of kernels, heating and mustiness, chemical and nutritional changes, and mycotoxin contamination. They can change fat quality of peanuts by hydrolytic enzymes producing free fatty acids and glycerol. Altogether, these changes  could lead to  lower quality of leafy vegetables (Kendar and Rolle, 2004).
Amaranths are nutritious vegetables largely consumed in Nigeria; many tend to lose their value due to lack of adequate knowledge in deterioration and poor storage methods. This study aimed to determine the microorganisms and other physicochemical factors causing deterioration in Amaranthus hybridus under storage.

MATERIALS AND METHODS
Collection and Storage of samples.
Fresh samples  of Amaranthus hybridus were obtained from three different places (farm, market and vegetable vendors)  in Lapai town, Niger State, Nigeria, and were kept in sterile bags. Plant identity was confirmed by botanists in the Herbarium of the Department of Biological Sciences, Ibrahim Babadamasi Babangida University, Lapai. Only samples of good quality, free from disease injury, were used in this study. They were properly cleaned   and stored in sterile open plastic containers in the laboratory under two different environmental conditions: at 4oCin the refrigerator and 28+ 2oC laboratory room temperature (Kitinoja and Kader, 2003).

Determination of  pH of the vegetable samples
The change in pH of the stored vegetables was determined before and during storage period. Twenty grams (20 g) of the sample was weighed at days 1, 3, 5, 7and 9 into sterile beakers. The sample was crushed; 20 ml of sterile distilled water was added, and allowed to rest for 30 min after which the pH was determined. Three replicates of each were made.

Determination of degree of deterioration of samples during storage
The severity of deterioration during storage at days 1, 3. 5, 7 and 9 was determined. Change in colour, moldiness and defoliation of the leaves were observed and recorded using the rating scale 0 to 5(Bhuiyan and Croft, 2011).

Moisture content determination
The Association of Official Analytical Chemists (A.O.A.C) (1990) method was used to determine the moisture content of the samples before storage(day1) and at the end of storage period(day9). Evaporating dishes of known weights were used. One gramme of each sample was weighed into each dish. All weighed samples were kept in the oven at 70°C for 24 hours to dry up. The samples were brought out and weighed again. Loss in weight of sample was represented by loss in water content and was calculated thus:
Weight of dish = Xg
Initial weight of dish + sample = Yg
Initial weight of the sample = Y – X = Pg
Final weight of dish + sample = Zg
Final weight of the sample = Z – X g = Qg
% moisture loss = (P – Q)g / P X 1 X 100

Isolation of pathogenic fungi and bacteria species from the deteriorating samples
The edge of infected samples of Amaranthus hybridus collected from the farm, market and vendor labeled and L1, L2 and L3, respectively, were excised and cut into 1mm pieces and surface-sterilized in 0.1% Mercury Chloride for  one minute and rinsed in four successive changes of sterile distilled water and the sections were then blotted dry on clean, sterile paper towels. They were plated on Potato Dextrose Agar (PDA) and Nutrient Agar (NA) in 3 replicates and incubated for 36 hours at 32 ± 2oC under 12-h photoperiod as in the method of Rangaswami and Bagyaraj (2007).

Isolation and identification of the fungal and bacterial isolates
Isolation, characterization and identification of the microorganisms were carried out using colonial, morphological and biochemical characteristics. The fungal isolates were identified based on examination of the colonial heads, phlalides, conidiophores and presence or absence of foot cells or rhizoids. Growth colonies of stock isolates were repeatedly sub-cultured three times on PDA plates using aseptic techniques to obtain pure isolates of organisms. Fungi isolated were identified using Fungi Families of the World Mycological monographs by Samsonand Reenen-Hoekstra (1988). Stock cultures were prepared using slant Potatoes Dextrose Agar in sterile McCartney bottles and preserved at 4OC in a refrigerator (Amadi and Adebola 2008). Incidence of foliar diseases was monitored throughout the period of the experiment. The bacterial species were identified using the following biochemical tests:  coagulase test, catalase test, mortility test, indole test and sugar fermentation test, according to the method of Rangaswami and Bagyaraj (2007).

RESULTS
Deterioration of stored vegetables
Table 1 shows the deterioration of vegetable samples stored at room temperature28+ 2OC and at 4OC refrigeration. At day 1, there was no noticeable infection; leaves were green and fresh under both conditions of storage. At day 3, the same observations were made for samples stored at 4OC but at room temperature, the colour of leaves  collected from farm and market changed to greenish yellow with less than 15% defoliation. However, the leaves of samples collected from vendors were completely yellow, with slight moldiness and 15 to 35% defoliation. At day 5, the samples stored in refrigerator were still green and looked fresh while samples collected from vendors and stored at room temperature turned blackish, heavily moldy and with over 67% defoliation. Followed closely were the samples collected from the market and stored at room temperature. At days 7 and 9, all samples in the refrigerator showed slight moldiness and the leaves turned yellowish green whereas samples under room temperature were moldy with almost complete defoliation.

pH Change during Storage
Generally, gradual change in pH of samples (Table 2) was observed under the two conditions in which the samples were stored. Samples collected from the farm and stored in refrigerator had initial pH of 5.87 before storage,  which increased to 6.35 at the end of the storage period. The same trend was observed in other samples collected from the market.
Moisture Content (MC)
Increase in the moisture content of Amaranthus hybridus stored at room temperature (Table 3) was observed. Samples collected from farm had initial value of 80.12% MC but increased to 84.04% at the end of day 9. The MC of the samples collected from vendors was the least with initial value of 80.56% and final value of 80.82%.

Fungi and Bacteria species Isolated
A total of six fungal species from four different genera were isolated and identified (Table 4). These fungi were Aspergillus niger, A. flavus, A. fumigatus, Penicillium notatum, Mucor pusillus  and Rhizopus stolonifa. These species were isolated from all the samples except M. pusillus and P. notatum that were not present in samples collected from farm and market.  A total of five bacteria species (Table 5)from five genera were isolated and identified from samples collected (Bacillus subtilis, Pseudomonas aeroginosa, Escherichia coli, Proteus mirabilis and Staphylococcus aureus). Out of these bacterial species, only E. coli and Bacillus subtilis were isolated from all the samples collected.

Table   1: Deterioration of vegetable Samples stored under two different conditions

 

T1= Refrigeration temperature 4oC    T2 = Room temperature 28 + 2oC   L1, L2 and L3= Leaves

Numbers in the table are from rating scale adapted from Bhuiyan and Croft, 2011.

Table 2: pH of A. hybridus during storage at room temperature(28oC) and in refrigerator(4oC)

Table 3:  Moisture content of A. hybridus stored at room temperature (28 + 2oC)

Table 4: Fungi species isolated from Amaranthus hybridus   samples

Keys: L1, L2 and L3= Leaves    + = presence ,  _ = absence

Table 5:  Bacterial species isolated  from Amaranthus hybridus

Keys: L1, L2 and L3= Leaves    + = presence , _ = absence

DISCUSSION
The results showed that deterioration began in Amaranthus samples collected from vendors and market and stored at room temperature before those collected from the farm. The vegetable samples stored at refrigerating temperature (4oC) took 7days before any evidence of deterioration was sighted. A lowering of the storage temperature has resulted in a considerable lengthening of the storage life. This result might probably be due to the arrest of the activities of the microorganisms at this temperature and the biological processes responsible for the break-down of the produce are greatly retarded by a lowering of the storage temperature. This was also reported by Kitinnoja and Kader (2003) that the microorganisms tend to be slow in their activities due to low temperature so that increase  in the shelf life of the vegetables increased.
The high amount of microbial deterioration observed with samples taken from Lapai market, vendors and stored at room temperature might be because the environment in which the vegetable was exposed had been polluted with high fungal density. This is in agreement with the study of Adebanjo and Shopeju (1993) and Ofor et al.(2009), who separately reported that bacterial and fungal contamination of vegetables might be from polluted air from the surrounding, soil, irrigation water and handling processes by man before storage.
The deterioration observed in this work was evident by loss of green colour to mushiness of the leaves and very high microbial count. The colour changes varied according to the storage condition. The samples stored at room temperature (28 + 2oC, 60% relative humidity) changed colour from day 3 of storage while the same samples stored at refrigerating temperature (4oC) began from day 7. Different factors, such as respiration, transpiration, translocation and metabolic activity, might contribute to the break-down. Kendar  and Rolle (2004) reported loss of green pigments as a post-harvest deterioration of leafy vegetables and microorganisms as agents of deterioration.
The fungal species isolated from this study have earlier been reported by Eaton and Groopman(1994) and Baiyewu et al.(2007). A. niger was the most commonly occurred among the fungi isolated. Earlier workers had reported this fungus as one that is commonly found on grape fruits ( Chulze et al.,2006), apples (Oelofse et al., 2006) and tomatoes (Yildz and Baysal,2006). Bali et al.(2008) reported that A. niger  caused post harvest deterioration in orange and lime fruits in the field.  Eboh and Okoh (1980) reported Fusarium sp, Aspergillus flavus, Aspergillus niger and Mucor species as the organisms found associated with decayed leafy vegetables. So, the deterioration of the Amaranthus hybridus   samples was not unconnected with these isolates. Anon (1995) reported that the high moisture contents of  Amaranthus hybridus coupled with its richness in minerals and vitamins must have encouraged the growth of this microbe on the vegetable leading to its deterioration. Other factors which may be responsible for the observed change in the deterioration rate might be the activity of various enzymes and the growth rate of micro-organisms at room temperatures.

CONCLUSION
Microorganisms naturally present on all foodstuffs can result from contamination from outside elements such as wind, soil, water, insects and handling during harvest. They can also become contaminated during growing, harvesting and transportation to the market (Akintobi et al., 2011). The occurrence of fungal deterioration of vegetables was also recognised as a source of potential health hazard to man and animals. This is due to their production of mycotoxic compounds which are capable of causing mycotoxicoses in man following ingestion (Effiuvwevwere, 2000). It is, therefore, necessary and important that both the farmer who harvests the vegetables into bags for transportation, the marketers and consumers take necessary and appropriate precautions in preventing the contamination of vegetables offered for sales.

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MORPHOLOGICAL AND PHYTOCHEMICAL EVALUATION OF ACACIA SIEBERIANA VAR WOODII (FABACEAE) STEM BARK.

Department of Pharmacognosy and Drug Development, Ahmadu Bello University, Zaria, Nigeria.

Received 27th May, 2014; accepted 30th December, 2014

*Author for correspondence

ABSTRACT
Acacia sieberiana var. woodii, popularly known as white thorn tree in English, is a savannah tree indigenous to Africa but adapted to Portugal and India due to its immense medicinal values. Stem bark of the plant is utilized extensively as a remedy for stomach ache and ulcers. Morphologically, the stem bark is hard, rough and woody, yellowish-brown to brown with grayish-black scales externally. It possesses a choking smell and an astringent taste when chewed. The chemo-microscopical studies revealed the presence of cellulose, lignified and suberized cell walls impregnated with prismatic calcium oxalate crystals, tannins and starch. Preliminary phytochemical studies revealed the presence of steroids, triterpenes, alkaloids and tannins. These features will serve as useful diagnostic tools for identifying and differentiating A. sieberiana var. woodii from its closely related varieties, A. sieberiana var. sieberiana, and A. sieberiana var. villosa.

Keywords: Morphological, Chemo-microscopical, Phytochemical, Evaluation, Acacia sieberiana var woodii, Stem Bark
INTRODUCTION
Acacia is the largest genus within the family Fabaceae (formerly Leguminosae) with 1,352 species presently identified, and has great importance for industry, rural development and conservation (James et al., 2006). The plant genus has had a long history of its medicinal uses in the treatment of various ailments including diarrhoea, skin infections and stomach ache.
Chemical and biological studies of Acacia species have revealed that they are important producers of secondary metabolites, possessing antitumor, antibacterial and hypotensive activities (Jesus et al., 2007; Gaara et al., 2008; Napar et al., 2012; Saurabhi et al., 2012). Acacia sieberiana (Kyal and Boatwr) has three varieties namely, A. sieberiana var. sieberiana (DC.), A. sieberiana var. woodii (Burtt Davy) and A. sieberiana var. villosa (A.Chev).
Acacia sieberiana var. woodii is a savannah tree 3-25 m in height, bole straight to 6 m long by 1m diameter with a rather rounded crown and trunk 6 m high. The bark is rough to touch and yellowish with grey-brown scales. The branches bear short to massive 10 cm long straight spines (Orwa et al., 2009). Aqueous extracts of the roots, stem bark and fruit pulp have been used traditionally in the management of inflammatory conditions, stomach disorders, bilharzias, fever, diarrhoea and leprosy (Orwa et al., 2009; Christiana et al., 2012). Despite claims for the use of the stem bark in the traditional remedies of various ailments, few researches have been carried out to set standards for the proper identification and differentiation of the plant from other closely related members of the genus and varieties.

MATERIALS  AND METHODS
Collection and Identification of A. sieberiana var. woodii
The plant specimen was collected from Basawa area of Sabon Gari Local Government Area of Kaduna State, Nigeria in May, 2013 and identified and authenticated at the Herbarium Unit of the Department of Biological Sciences, Ahmadu Bello University, Zaria. A voucher number, 900248, was assigned.

Preparation of A. sieberiana var. woodii Stem Bark
Stem bark of A. sieberiana var. woodii was air-dried under shade and pulverized. The powdered plant material was then transferred into an air-tight container for proper storage before use.

Morphological Studies of A. sieberiana var. woodii Stem Bark
Macroscopical/ Organoleptic Studies
This was carried out using the impression of the sense organs in accordance with standard procedures (WHO, 2011).
(a)    Colour: Fresh sample of the stem bark was placed on a white board and observed under daylight.         Both the outer and inner surfaces were observed and photograph of the sample was taken. The         colour of the powdered sample was also observed and recorded.
(b)    Odour: The powdered stem bark was placed in a clean and dry beaker and the odour was detected         by inhaling over the plant material.
(c)    Taste: A small quantity of the powdered plant material was chewed to detect its test.
(d)    Surface texture: The outer and inner surfaces were felt between fingers to observe for its roughness         or smoothness.

Chemo-Microscopical Studies of A. sieberiana Stem Bark
The powdered sample of the stem bark was placed in a clean beaker. A small quantity of sodium hypochlorite was added to the sample in the beaker and placed in a hot water bath for 30 minutes. The sample was thoroughly washed with fresh water and used for the chemo-microscopical examination. The mountant used was glycerol. It was observed under a light microscope following standard procedures (WHO, 2011).
(a)    Cellulose cell walls: Iodinated zinc chloride (2 drops) was added to the cleared sample on a slide,         and this was allowed to stand for a few minutes. 25% Sulphuric acid (1 drop) was added and a cover-        slip was applied before observation.
(b)    Suberized cell walls: Sudan red (2 drops) was added to the cleared sample on a slide, a coverslip was         applied and this was gently heated over flame for a few seconds.
(c)    Lignified cell walls: A few drops of phloroglucinol was added to the cleared sample and allowed to         stand until almost dry. 25% sulphuric acid (1 drop) was added and covered with a coverslip.
(d)    Calcium carbonate and calcium oxalate: To the cleared sample, a few drops of acetic acid were added         and covered with a coverslip. This was observed under the microscope. A few drops of hydrochloric         acid were added.
(e)    Tannin: A single drop of ferric chloride solution was added to the cleared sample and covered with a         coverslip. This was observed under the microscope.
(f)    Starch: A few drops of iodine solution were added to the cleared sample and covered with a coverslip.
(g)    Inulin: A drop each of 1-naphthol and sulphuric acid was added to the cleared sample and covered         with a coverslip. This was then observed under the microscope.
(h)    Hydroxyanthraquinones: A drop of potassium hydroxide was added to the cleared sample and         covered with a coverslip.
Preliminary Phytochemical Studies of A. Sieberiana var. woodii Stem Bark
Extraction of the stem bark
The powdered stem bark (20 g) was weighed and transferred into an extraction thimble and extracted sequentially using hexane and methanol (70%) in Soxhlet apparatus at 60oC for 12 hours.
The methanolic extract collected was transferred into a crucible and placed over hot water bath for evaporation (Das et al., 2010).

Phytochemical Studies
Test for Phenolic Compound
(a)    Ferric chloride test: The extract (0.5 ml) was dissolved in 10 ml of water each and filtered. A few         drops of ferric chloride were added to the filtrate and the colour reaction was observed (Sofowora,         2008).
(b)    Lead sub-acetate test: Lead sub-acetate solution (3 drops) was added to the extract solution and the        reaction was observed (Evans, 2009).
(c)    Gold-Beater skin test: The powdered stem bark (2 g) was placed in 10 ml of 50% alcohol and filtered.
The gold-beater skin was soaked in 2% HCl, rinsed with water and transferred to the extract for 5 minutes. The skin was then removed, washed with water and placed in a solution of ferrous sulphate and observed (Evans, 2009).

Test for Saponins
(a)    Frothing test: The extract (2 ml) was dissolved in 10 ml of water, shaken vigorously for 30 seconds         and then allowed to stand for 30 minutes before observation (Sofowora, 2008).

Test for Steroids/ Triterpenes
(a)    Salkowski test: Chloroform (2 ml) and a few drops of sulphuric acid were added to about 2 ml of the         extract and the reaction was observed (Sofowora, 2008).
(b)    Lieberman-Burchard test: Acetic anhydride (1ml) was added to 1 ml of the extract. A few drops of         sulphuric acid were then added to the solution supernatant and the reaction was observed (Sofowora,         2008).

Test for Flavonoids
(a)    Shinoda test: The extract (0.5 g) was dissolved in 2 ml of 50% methanol. A few drops of magnesium         fillings and 3 drops of hydrochloric acid were added and the reaction observed (Evans, 2009).
(b)    Sodium hydroxide test: A few drops of sodium hydroxide were added to 5 ml of the extract and the         reaction was observed and recorded (Evans, 2009).

Test for Cardenolides
(a)    Kella-killiani test: The extract (2 ml) was dissolved in glacial acetic acid containing ferric chloride         and 1 ml of sulphuric acid was added to the solution. The reaction was observed (Sofowora, 2008).
(b)    Kedde test: 2% 3,5-dinitrobenzoic acid (2 ml) in 95% alcohol was added to  the extract. A few drops of         5% sodium hydroxide were added to the solution and the reaction was observed (Evans, 2009).

Test for Alkaloids
(a)    Mayer test:  Potassium mercuric iodide was added to the extract solution and the reaction was         observed (Evans, 2009).
TABLE I: Preliminary phytochemical studies of A. sieberiana var. woodii stem bark

DISCUSSION
Stem bark of A. sieberiana var. woodii was macroscopically observed to be in line with the report of Orwa et al. (2009) who described the plant as rough, hard and woody with yellowish-brown colour and possessing greyish-black scales externally.
Chemomicroscopically, the stem bark of A. sieberiana var. woodii was observed to contain cellulose cell wall along with lignified and suberized cell walls. Prismatic calcium oxalate crystals along with tannins and starch were also observed. This observation is in line with some studies of related species that have been reported to possess prismatic calcium oxalate, tannins, starch and phloem fibres as in Bauhinia purpurea, A. leucophloea and A. podalyrrifolia (Duarte and Wolf, 2005; Gupta et al., 2010; Gupta et al., 2012).
The preliminary phytochemical studies of the methanolic extract of A. sieberiana var. woodii stem bark showed the presence of flavonoids, as reported by Hegenauer and Renpe (1993), who observed that legumes are particularly rich in flavonoids.  The presence of alkaloids, tannins, steroids and triterpenes in the stem bark is also similar to previous reports on the genus Acacia (Jesus et al., 2007; Shittu et al., 2010). Anthraquinones, cyanogenic and cardiac glycosides were absent in the present study but were reported to be present in the stem barks of closely related species (Kubmarawa et al., 2007; Chaudhary et al., 2009; Anjaneyulu et al., 2010; Suman et al., 2011; Okpanachi et al., 2012; Deshpande, 2013).

Conclusion
The present study on the stem bark of A. sieberiana var. woodii will be useful in distinguishing it form other related species.

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