Introduction

India is a treasure house for endemic flora with various climatic conditions. Nearly 5725 species of flowering plants are widely recognized as endemics plants. Among this endemic flora, 3471 species are found in the Himalayas, 2051 in Peninsular India and 239 in Andaman & Nicobar Islands (Nayar, 1996). Rhynchosia beddomei Baker (Fabaceae) is an endemic plant of the Eastern Ghats and its distribution is recorded in Andhra Pradesh, Tamil Nadu and Karnataka states (Aluri & Kunuku, 2018; Rao & Henry, 1996). In Andhra Pradesh, its distribution is restricted to moist deciduous forests of Seshachalam hill ranges. R. beddomei is a source for the tribal inhabitants of the Seshachalam hill ranges for the treatment of sprains, healing wounds and as an antidote for insect bites (Rao et al., 1996; Reddy, Reddy, Pattanaik, & Raju, 2006). In addition to its endemism, rare distribution and high therapeutic importance, its identification in dried form is a big challenge (Verma, Ahmad, & Singh, 2020). For the identification of an authentic botanical source, pharmacognostic approaches are the only choice (Balasubramaniam, Sekar, & Badami, 2020; Pratap, Sudarsanam, & Nagaraju, 2016). The pharmacognostic characteristics of R. beddomei are not reported yet. The present study was conducted with an aim to establish pharmacognostical data of stem and roots with the scope of macroscopical, microscopical, physicochemical, phytochemical and fluorescence studies.

Description and distribution

Rhynchosia beddomei Baker in Hook. f. Fl. Brit. India. 2.222.1876; Gamble 1: 374.1981.

Erect undershrubs, up to 1.5 m tall, branches white – covered with whitish or greyish hairs. Leaves 3-foliolate, leaflets oblong or oblong-lanceolate, velvety, white pubescent on both surfaces, subcoriaceous, margin entire, acute (Figure 1 A-C; Figure 2 A-B). Flowers in subsessile congested racemes Calyx to 1 cm long, lobes oblong, obtuse, longer than the corolla and silky. Corolla bright yellow. Pod suborbicular, thinly canescent; seed 1, strophiolate. R. beddomei distributed in moist deciduous forests of Seshachalam hills in Southern Eastern Ghats.

Phenology of the plant

Leaf fall is noticed from May to June, followed by new foliage formation from August to October. Flowering is started in November and ends in January, followed by fruiting from February to April.

Materials and Methods

Collection of specimens

The whole plant material was collected from the Seshachalam hill ranges and the plant was identified with the help of Flora of the Presidency of Madras (Gamble, 1936) and crosschecked (Voucher Specimen Number: CR119). A specimen of the herbarium was deposited in the Department of Botany, Sri Venkateswara University, Tirupati, Andhra Pradesh. Different parts were collected and fixed in FAA (Formalin+ Acetic acid + 70% Ethyl alcohol) (Ratio of 5:5:90). After 24 hours, the specimens were dehydrated with graded series of tertiary-butyl alcohol (Sass, 1940), followed by infiltration of the specimens by progressive addition of paraffin wax (Melting point 58-60^o C) until TBA solution attained supersaturation. The specimens were cut into paraffin blocks.

Ethnobotany of the plant

The ethnopharmacological importance of R. beddomei was obtained through conversation in the local Telugu language with nearly 130 tribal informants from Seshachalam hill ranges between the age groups of 40-70 years.

The studied areas are inhabited by Yanadis and Chenchus. The majority of the tribal inhabitants are Chenchus. The survey is aimed to document the data on the local names of the plant, useful plant parts, method of drug preparation, mode of drug administration, dosage, the form of usage and whether the plants used either singly or in combination with other plants, minerals and salts.

Microscopical studies

For microscopical observation, plant specimens were subjected to dehydration, infiltration, supersaturation, sectioning, de-waxing and staining. It is followed by the preparation of microphotographs at different magnifications (Johansen, 1940; Pratap, Sudarsanam, & Prasad, 2014).

Physico-chemical studies

Loss on drying, total ash values, acid insoluble ash values, water-insoluble ash, various extractive values were calculated (Kumari, Pratap, & Murthy, 2020; Yadav & Singh, 2018).

Fluorescence studies

Fluorescence studies used to develop the standards for the measurement of the purity of the powdered drug (Jyothi et al., 2020; Pratap et al., 2014).

Preliminary phytochemical studies

The root and stem powders were subjected to various types of analysis for the identification of secondary metabolites like alkaloids (Dragendorff’s and Mayer’s tests), triterpenes (Libermann Burchard’s test), flavonoids (Aluminium chloride test), anthraquinones (Borntrager’s test), polyphenols (Ferric chloride test), sterols (Salkowski’s test), coumarins (Lacton test), saponins (Foam test) and tannins (Gelatin test) (Harbone, 1973; Kokate, Purohit, & Gokhale, 2008).

Results and Discussion

Ethnopharmacological medications of the plant

The ethnopharmacological uses of R. beddomei root, stem and leaf was collected from the Yanadi and Chenchu tribes from Seshachlam hills (Table 1).

Microscopical studies of stem

The stem is angular in transactional outline with short ridges of varying length (Figure 3). The stem has a thin, uninterrupted epidermal layer of small thick walled cells with prominent cuticle. The cortical zone is narrow, comprising of four or less layers of parenchyma cells with dense tannin contents. Inner to the cortex is a thick, undulate continuous cylinder of sclerenchyma cells (fibers) enclosing the vascular cylinder (Figure 4).

The vascular tissue consists of secondary phloem and secondary xylem. Secondary phloem is wide and continuous, the phloem elements being diffuse in distribution. Wide, circular tanniniferous idioblasts are frequently seen in the phloem. Secondary xylem occurs in the thick, hollow cylinder; it has narrow, angular or circular, thick walled vessels and libriform fibres. The vessels are in radial multiples and are 20 µm wide. Along the inner circumference of the xylem cylinder, these are triangular clusters of small, compact cells abutting the primary xylem strands.

When the sections of the stem is viewed under the polarized light microscope, calcium oxalate crystals were evident in a different part of the stem (Figure 6; Figure 5) as well as in the midrib. The crystals are mostly prismatic type of rectangular and cuboidal shape. Less frequently, they are druses. In the midrib, rhomboidal type crystals are located along the outer margin of the sclerenchyma cylinder (Figure 5). In the stem, the crystals occur in the pith and cortex; they are also seen in the phloem parenchyma (Figure 7; Figure 6).

Microscopical studies of root

Thin and thick roots were studied and both the roots have secondary thickening.

The thin root (Figure 9; Figure 8) has a fairly wide fissured periderm and a narrow cortex with a discontinuous layer of sclerenchyma elements inner with periderm. The secondary phloem is fairly wide and continuous. Secondary xylem is a solid, wide circular cylinder, comprising of xylem fibres wide dilated xylem rays, wide, thin walled, circular, solitary vessels which diffusely radiate from the centre towards the periphery. The diameter of the vessels is narrow in the centre and wider towards the periphery (Figure 9). The wide vessels are 70 µm in diameter.

The thick root measuring 2.5 mm is similar to the thin root, but there are some differences due to more amount of secondary tissues (Figure 12; Figure 11; Figure 10). The thick root has a wider, fissured super periderm followed by a cortical zone where parenchyma cells and sclereids are mixed at random.

Secondary phloem consists of regular radial files of phloem elements. The phloem rays are narrow and less conspicuous. The secondary xylem has more vessel frequency, the vessels are diffuse and spread towards the periphery with an increasing diameter (Figure 12). Xylem fibers of thin walled and thick walled cells occur in alternating circles. The wide vessel elements are 50 µm in diameter.

Powder microscopic studies of stem

Stem powder (Figure 15; Figure 14; Figure 13) contains an abundance of fibers and a few vessel elements. The fibers are all narrow, thick walled and lignified. The fibers 600-750 µm long and 10 µm wide. No pits are evident on the fibers.

In addition to fibers, vessel elements are also seen in the powder. They are short and cylindrical. They are 90 µm wide and 130 µm long. They have dense elliptical pits on the lateral walls (Figure 15). The perforation is simple and horizontal.

Powder microscopic studies of root

The root powder has fibers similar to those of the stem. Vessel elements are more frequent in the root than in the stem. The vessel elements vary from narrow to wide; some of the narrow elements have a short tail. The perforation plate is simple and oblique (Figure 16 ). The wider vessel elements have no tails; their perforation plate is horizontal (Figure 17). Lateral wall pits are circular, alternate and densely crowded. The narrow vessel elements are 120-150 µm long. The wider elements are 150 µm long.

Diagnostic Characters

Stem

• Tanniniferrous circular idioblasts are prominent in the phloem region.

• Prismatic, rectangular and cuboidal shaped Calcium oxalate crystals are found in the pith, cortex and phloem parenchyma.

• Vessels are short, cylindrical and with elliptical pits on the lateral wall

Root

• Presence of secondary growth.

• Narrow vessels in the centre and wider (7µm) towards the periphery in the thin root, whereas wider vessels (50 µm) in the thick root.

• Narrow vessel elements have short drawn out ends and wider elements without drawn out ends.

Ash and extractive values of different parts of R. beddomei

Ash and the Extractive values of a drug provide an opinion on the inorganic composition and other impurities present along with the drug. The results are given in Table 3; Table 2.

Preliminary phytochemical Studies

The present study is intended to unveil the preliminary phytoconstituents from aqueous, ethanol and chloroform extracts of root and stem parts from R. beddomei. The study revealed alkaloids, carbohydrates, cardio glycosides, flavonoids, glycosides, lignins, phenols, saponins and tannins (Table 4).

Fluorescence studies

Fluorescence analysis of the powder produced from the root and stem parts was studied and recorded with different extracts, solutions and reagents (Table 7; Table 6; Table 5).

Conclusions

The study signifies the pharmacognosy of an endemic medicinal plant, R. beddomei, with important diagnostic characters for the authentication of species. These findings are the source for the development of quality control parameters to overcome substitutes and adulterants. The present study is also aimed to document the ethnomedicinal uses on R. beddomei. It will be a great asset if this knowledge of ethnomedicinal and pharmacognostic evaluation is subjected to scientific validation by advanced pharmacological and clinical studies. Such investigations will bring to light the newer drugs of plant origin for the treatment of various ailments.

Funding Support

The authors declare that they have no funding support for this study.

Conflict of Interest

The authors declare that they have no conflict of interest for this study.