According to Stace (1965), the leaf is perhaps the most anatomically varied organ in angiosperms and it provides a variety of anatomical features that can be employed as useful taxonomic characters. Many research works have been done on the physiology, chemotaxonomy and medicinal use of the species of these genera and other genera in the family but information on the anatomy and taxonomy is still fragmentary.  Therefore the aim of this work was to determine the taxonomic significance of leaf epidermis and other anatomical features with a view to further establishing a stable taxonomic character among the selected genera.

Plant specimens used were collected in different ecological areas in Abeokuta (Ogun State), Ibadan, Sepeteri and Iseyin (Oyo State) South-western Nigeria. The collected specimens are: Bauhinia rufescens, B. tomentosa, Caesalpinia bonduc, C. pulcherima, Danielia ogea, D. oliveri, Senna acutifolia, S. alata, S. fistula, S. hirsuta, S. obtusifolia, S. occidentalis, S. podocarpa, S. siamea and S. siberiana.  Identification was done at Forestry Herbarium Ibadan (FHI) and University of Ibadan Herbarium (UIH). Voucher specimens were deposited in the two herbaria. Quantitative characters assessed include leaf length and width (taken at the widest point) while qualitative characters such as leaf shape, margin, base and apex were also assessed in situ.  Micro-characters such as cell wall thickness, size of epidermal cell, stomata size and index were also measured.

 

Preparation of leaf epidermal surfaces

Leaf  epidermal  morphology  was studied  using  fresh  specimens.  About 5 mm² to 1 cm² leaf portions were obtained from the standard median portion of the leaves. Three to five specimens of each species were used depending on geographical spread of the species except for those known from only one or two localities. Epidermal peeling was carried out according to the procedure of Johansen (1940) as modified by Jayeola and Thorpe (2000). The peeled specimens were later stained with Safranin and counter stained with Fast green and then cleared in clove oil and mounted with DPX^®. The slides were appropriately labeled and examined under a light microscope Olympus BX51 while photomicrographs of the micro-morphological features were taken at a magnification of X400 using a photomicrograph Olympus BX 51with installed digital camera optics.

For statistical analysis, 10 epidermal cells and 10 stomata were chosen randomly for each species and measured using a micrometer eyepiece. For each quantitative character, the mean and standard error were determined for all the taxa. The stomata index (SI) was calculated based on the formula derived by Metcalfe and Chalk (1979).

 

                                                      

 

Where, SI = Stomata index, S = Number of stomata per unit area, E = Number of epidermal cells in the same unit area.

Tables 1 and 2 and Figures 1 to 4 show the results obtained from this study while Plates 1-30 show photomicrographs of the abaxial and adaxial surfaces of the plant specimens.  Generally, the leaves of the genera (Bauhinia, Ceasalpinia, Daniellia, Senna) were alternate, rarely opposite and mostly entire (Table 1).  Leaves were mostly glabrous in all species except in S. hirsuta and S. alata which were pubescent. The apices were emarginate, acute, acuminate, retuse, or apiculate. The leaflet sizes showed considerable variations within and among the genera with the largest recorded in S. fistula (14.8 cm) and the smallest in B. rufescens (0.8 cm) (Figure 1). The lowest leaf length/width ratio 1:1 was recorded in B. rufescens, B. tomentosa and S. obtusifolia (Table 1), while the highest was recorded in S. acutifolia.  Trichome bases were, however, noticed in some species occurring on their adaxial or abaxial surface. The leaf epidermal cells were more often polygonal (Table 2) but sometimes irregular. Polygonal cells occurred most often on the abaxial surfaces but were also noticed on the adaxial surfaces of C. bonduc, D. ogea, S. alata, S. fistula, S. obtusifolia, S. hirsuta and S. siamea (Plates 1-30). Sometimes, polygonal cells were seen interspersed with irregular cells as seen in C. pulcherima, D. ogea, D. oliveri, S. alata and S. podocarpa.  Stomata were amphistomatic in all the species and have commonly anomocytic, anisocytic and paracytic types apart from the rare to occasional occurrence of other stomata types and abnormalities found on the adaxial surface of D. oliveri. Trichomes were absent in most of the species except for the presence of whip-like and non-glandular trichomes found on both surfaces of  S. hirsuta  and  S. alata.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

  

There was a wide variation in the number and distribution of stomata found in all the species.  Classification of different types of stomata complexes was based on the number and position of the subsidiary cells and the ontogeny of the cell types. According to Richard et al. (2007), stomata comprise two elongated guard cells bracketing a stomatal pore, and often but not always, surrounded by one to many subsidiary cells. The presence of desmocytic stomata in D. oliveri distinguished it from other species. This was corroborated by the findings of Metcalfe and Chalk (1979). Also, absence of stomata on the adaxial surfaces of S. alata, S. fistula, S. siamea, and S. siberiana made them distinct from other species in the genus. The cell shape and cell wall patterns vary considerably among these genera and based on these two characters, the species of the genera was divided into groups of five; those with curved anticlinal wall, straight anticlinal wall, curved and straight anticlinal wall, irregular and polygonal cell shapes.

The occurrence of curved walls in most of the species agreed with the suggestion of Stace (1965) and Richard  et al. (2007) that curved wall was a mesomorphic character and that environmental conditions such as humidity play a significant role in determining the pattern of anticlinal wall.  The relative abundance and variation of the trichomes was of taxonomic importance in the genera.  Almost all the species in these genera were glabrous which distinguished them from other species which were pubescent.  However, the presence of simple, long, interwoven trichomes which covers the epidermal cells and stomata in S. alata and S. hirsuta distinguished them from other species in the same genus. The presence of many short non-glandular trichomes in B. tomentosa and S. siberiana makes it easy to be separated from other species. Metcalfe and Chalk (1979) opined that trichomes frequency and size are environmentally controlled while Stace (1965) reported that hairs are constant in species, and when present showed a constant range of form and distribution useful in diagnosis. Some overlap features noted in some species and genera within the sub-family perhaps still explain the affinity in their relationship despite the recent taxonomic divergence.

Preponderance of stomata on the abaxial surfaces than adaxial surfaces is a mechanism to reduce water loss through transpiration (Adegbite, 2008). The foliar epidermal features of some members of the genera of the sub-family Caesalpinoideae is of taxonomic importance since they can be separated and classified according to their stomata and cell wall shape.  Based on the striking similarities and differences noted in the morphological and anatomical features, it is hereby suggested that further research such as phytochemical analysis and molecular studies be carried out to further delimit the species, as this work serves as baseline upon which other research could stand.

The author(s) have not declared any conflict of interests.

The effort of Mrs. Olanloye Oluwafunmilayo Adebimpe of the Department of Biological Sciences, Federal University of Agriculture, Abeokuta, Ogun State on the photomicrographs is greatly acknowledged.