Nikhil B. Gaikwad

Highly efficient in vitro proliferation and genetic stability analysis of micropropagated Ceropegiaevansii by RAPD and ISSR markers: A critically endangered plant of Western Ghats

Ceropegiaevansii McCann (family: Asclepiadaceae), a critically endangered plant of Western Ghats has acquired significant importance due to its medicinal implications, edible tubers, and ornamental flowers. This study deals with the optimization of axillary bud proliferation using nodal explants followed by genetic stability analysis of regenerants. Maximum number of shoots (11.6 ± 1.1) was observed on the Murashige and Skoog (MS) medium supplemented with 6-benzylaminopurine (4.0 mg/l) and indole-3-acetic acid (0.3 mg/l) with 85% shoot multiplication frequency. In vitro-grown shoots were rooted best in 1/2 MS medium supplemented with indole-3-butyric acid (1.0 mg/l) with an average of 10.3 ± 0.9 roots per shoot and 92% rooting frequency. Plantlets were acclimatized best (90%) in a mixture of sterile soil, sand, and coco peat (1:2:1). Micropropagated plants were subjected to random amplified polymorphic DNA and inter simple sequence repeat markers analyses. Collectively, 759 bands were generated which were monomorphic and similar to the mother plant. Findings of this study are the first report on micropropagation and assessment of genetic stability of micropropagated plantlets in C. evansii which suggests that axillary shoot proliferation can safely be used as an effective tool for propagation and conservation of C. evansii.

Phytochemical composition, antioxidant activity and HPLC profiles of Swertia species from Western Ghats.

Swertia chirayita is one of the potential medicinal plants of the family Gentianaceae in traditional medicine. Due to its high demand and scarcity, trade of chirayita is affected by adulterants. Swertia species from Western Ghats were compared with S. chirayita for phytochemical characterisation and antioxidant activities by using different extracts. This study revealed that acetone is the best extraction solvent of phenolic and flavonoid compounds with antioxidant properties as compared with other extracts. S. chirayita showed better antioxidant activity than other species with highest content of phenolics and flavonoids. Among the species from Western Ghats, Swertia minor has better antioxidant properties with higher content of phenolics and flavonoids when compared with S. chirayita. Gallic acid was detected in all species under study by using HPLC analysis. The Swertia species under study showed similar phytochemical properties and antioxidant potential and hence their use as substitute to S. chirayita needs to be further investigated.

RP-HPLC analysis of seco-iridoid glycoside swertiamarin from different Swertia species

Abstract Genus Swertia is valued for its great medicinal potential; mainly Swertia chirayita (Roxb. ex Fleming) H. Karst. is used in traditional medicine for a wide range of diseases. Seco-iridoid glycosides like swertiamarin is referred with enormous pharmacological potentials. The aim of the study was to identify a suitable substitute to S. chirayita by quantifying seco-iridoid swertiamarin from five different Swertia species endemic to the Western Ghats. The reverse-phase high-performance liquid chromatography diode array detector analyses were performed and chromatographic separation was achieved on a Lichrospher 100, C18e (5 µm) column (250–4.6 mm). A mobile phase consisting of acetonitrile and water (25:75) was used for separation. Results indicated that the concentration of the marker compound has been found to vary largely between and within the species from different localities. The content of swertiamarin was the highest in S. chirayita compared to the other species studied herein, advocating the use of Swertia minor as an alternate source to S. chirayita. Graphical abstract

Highly efficient in vitro regeneration, establishment of callus and cell suspension cultures and RAPD analysis of regenerants of Swertia lawii Burkill

Graphical abstract

Quantitative determination of three pentacyclic triterpenes from five Swertia L. species endemic to Western Ghats, India, using RP-HPLC analysis

Aim of this study was to identify pentacyclic triterpenoids betulinic acid (BA), oleanolic acid (OA) and ursolic acid (UA) from five Swertia species endemic to Western Ghats, which are used as substitutes as well as adulterants to Swertia chirayita. Our results indicate that the concentration of active compound vary largely among and within the species from different localities. OA was found in a higher amount in all species compared to BA and UA. From the results, it was clear that BA, OA and UA are present in the endemic species collected from Western Ghats, thus advocating the use of these species as alternate sources to S. chirayita. This in due course may release pressure of exploitation from natural resources of S. chirayita and help to bring it out from an endangered category from conservation point of view.

Seed coat sculpture of subgenus Ceratotropis (Piper) verdc., genus Vigna Savi in India and its taxonomic implications

Abstract The seed coat morphology was studied using scanning electron microscopy in 21 species (of which two are both cultivated and wild) of subgenus Ceratotropis (Piper) Verdc. and one species each from subgenus Plectotropis and subgenus Vigna of genus Vigna. The macromorphological and micromorphological seed characters, which include seed shape, testa texture, hilum, aril, testa pattern and ornamentation of periclinal wall, exhibited well-defined variability among the studied taxa. The smallest seeds were recognized in Vigna subramaniana (1.97 × 1.91 mm) and the largest in Vigna unguiculata (7.02 × 4.45 mm). The smallest hilum was observed in Vigna subramaniana (0.50 ± 0.01 mm) and the largest in Vigna umbellata (3.57 ± 0.05 mm). Novel morphological features such as reticulate, reticulate-foveate, alveolate, colliculate-pusticulate, substriate, ruminate-reticulate and ruminate-channelled reticulations of periclinal wall of the testa cell and hilum structure were observed in addition to those reported in earlier studies. In the present investigation, we reported the seed micromorphology of wild endemic Vigna species, namely Vigna sahyadriana, Vigna subramaniana, Vigna hainiana, Vigna indica, Vigna khandalensis and Vigna konkanensis for the first time. Based on seed characters, the taxonomic identity and species complex in the subgenus Ceratotropis has been discussed. A key to the species has been developed based on seed characters.

Reverse Phase-ultra Flow Liquid Chromatography-diode Array Detector Quantification of Anticancerous and Antidiabetic Drug Mangiferin from 11 Species of Swertia from India

Background: Genus Swertia is valued for its great medicinal potential, mainly Swertia chirayita (Roxb. ex Fleming) H. Karst. is used in traditional medicine for a wide range of diseases. Mangiferin one of xanthoids is referred with enormous pharmacological potentials. Objective: The aim of the study was to quantify and compare the anticancerous and antidiabetic drug mangiferin from 11 Swertia species from India. The study also evaluates hierarchical relationships between the species based on mangiferin content using multivariate analysis. Materials and Methods: The reverse phase-ultra flow liquid chromatography-diode array detector analyses was performed and chromatographic separation was achieved on a Lichrospher 100, C18e (5 μm) column (250–4.6 mm). Mobile phase consisting of 0.2% triethylamine (pH-4 with O-phosphoric acid) and acetonitrile (85:15) was used for separation with injection volume 20 μL and detection wave length at 257 nm. Results: Results indicated that concentration of mangiferin has been found to vary largely between Swertia species collected from different regions. Content of mangiferin was found to be highest in Swertia minor compared to other Swertia species studied herein from the Western Ghats and Himalayan region also. The same was also evident in the multivariate analysis, wherein S. chirayita, S. minor and Swertia paniculata made a separate clade. Conclusion: Conclusively, the work herein provides insights of mangiferin content from 11 Swertia species of India and also presents their hierarchical relationships. To best of the knowledge this is the first report of higher content of mangiferin from any Swertia species.

Molecular authentication of the traditional medicinal plant “Lakshman Booti” (Smithia conferta Sm.) and its adulterants through DNA barcoding

Background: Smithia conferta Sm. is an annual herb widely used in Indian traditional medical practice and commonly known as “Lakshman booti” in Sanskrit. Morphological resemblance among the species of genus Smithia Aiton. leads to inaccurate identification and adulteration. This causes inconsistent therapeutic effects and also affects the quality of herbal medicine. Aim: This study aimed to generate potential barcode for authentication of S. conferta and its adulterants through DNA barcoding technique. Materials and Methods: Genomic DNA extracted from S. conferta and its adulterants was used as templates for polymerase chain reaction amplification of the barcoding regions. The amplicons were directed for sequencing, and species identification was conducted using BLASTn and unweighted pair-group method with arithmetic mean trees. In addition, the secondary structures of internal transcribed spacer (ITS) 2 region were predicted. Results: The nucleotide sequence of ITS provides species-specific single nucleotide polymorphisms and sequence divergence (22%) than psbA-trnH (10.9%) and rbcL (3.1%) sequences. The ITS barcode indicates that S. conferta and Smithia sensitiva are closely related compared to other species. Conclusion: ITS is the most applicable barcode for molecular authentication of S. conferta, and further chloroplast barcodes should be tested for phylogenetic analysis of genus Smithia. Abbreviations used: BLASTn: Basic Local Alignment Search Tool for Nucleotide; MEGA: Molecular Evolutionary Genetic Analysis; EMBL: European Molecular Biology Laboratory; psbA-trnH: Photosystem II protein D1- stuctural RNA: His tRNA gene; rbcL: Ribulose 1,5 bi-phosphate carboxylase/ oxygenase large subunit gene.

Vigna khandalensis (Santapau) Raghavan et Wadhwa: a promising underutilized, wild, endemic legume of the Northern Western Ghats, India

Vigna khandalensis (Santapau) Raghavan et Wadhwa, is an endemic legume belonging to subgenus Ceratotropis (Asian Vigna), genus Vigna of the family Fabaceae. It is commonly known as ‘Ranmung’, ‘Jungli mung’ or ‘Badamung’. V. khandalensis is sporadically distributed at high altitude areas in the Northern Western Ghats, India. It is an annual, erect, tall herb growing along roadsides, hilly slopes, pond sides and near human settlements. The legume seeds are utilized by human beings at the time of famine. Surprisingly, the plant remains neglected and underutilized although its high nutritional value. The scant information on V. khandalensis leads us to investigate this underexploited and underutilized legume as a food source for human. The present paper deals with geographical distribution, taxonomy, ecology, agro-morphological characterization, nutritional content and phytochemical analysis. The results obtained in the present investigation provide information on agro-morphological traits and nutritional potential of V. khandalensis. Furthermore, it is promoted as a legume ‘Crop for the Future’ for food security.