Assessment of chemical and genetic variability in Tanacetum gracile accessions collected from cold desert of Western Himalaya.

Genetic diversity is essential for survival and adaptation of high altitude plants such as those of Tanacetum genus, which are constantly exposed to environmental stress. We collected flowering shoots of ten accessions of Tanacetum gracile Hook.f. & Thomson (Asteraceae) (Tg 1-Tg 10), from different regions of cold desert of Western Himalaya. Chemical profile of the constituents, as inferred from GC-MS, exhibited considerable variability. Percentage yield of essential oil ranged from 0.2 to 0.75% (dry-weight basis) amongst different accessions. Tg 1 and Tg 6 were found to produce high yields of camphor (46%) and lavandulol (41%), respectively. Alpha-phellendrene, alpha-bisabool, p-cymene and chamazulene were the main oil components in other accessions. Genetic variability among the accessions was studied using RAPD markers as well as by sequencing and analyzing nuclear 18S rDNA, and plastid rbcL and matK loci. The polymorphic information content (PIC) of RAPD markers ranged from 0.18 to 0.5 and the analysis clustered the accessions into two major clades. The present study emphasized the importance of survey, collection, and conservation of naturally existing chemotypes of medicinal and aromatic plants, considering their potential use in aroma and pharmaceutical industry.

In vitro propagation and withaferin A production in Withania ashwagandha, a rare medicinal plant of India.

Withania ashwagandha, belonging to the family Solanaceae, is an important medicinal herb of India with restricted geographic distribution. It is a rich source of withaferin A (WA) and other bioactive withanolides. In the present study a rapid in vitro mass propagation protocol of W. ashwagandha was developed from nodal explants. Nodal explants were cultured on MS medium supplemented with various concentrations and combinations of plant growth regulators (PGRs). The highest number of regenerated shoots per ex-plant (33 ± 2.7) and highest WA (13.4 ± 1.15 mg/g of DW) production was obtained on MS medium supplemented with 5.0 µM 6-benzyladenine (BA) and 1.0 µM Kinetin (Kn). In vitro raised shoots were further rooted on half-strength MS medium containing 2.0 µM Indole-3-butyric acid (IBA) and analyzed for WA production. The rooted plantlets when transferred to poly bags in the greenhouse showed 90 % survival frequency. Levels of WA were higher in the in vitro and ex vitro derived shoot and root tissues as compared to field grown mother plants. In an attempt to further maximize WA production, shoot cultures were further grown in liquid MS medium supplemented with 5.0 µM 6-benzyladenine (BA) and 1.0 µM Kinetin (Kn). Root cultures were grown on half strength MS liquid medium fortified with 2.0 µM of IBA. WA production in the liquid cultures was significantly higher compared to the static composition of the same media. This protocol, first of its kind in this plant, can be successfully employed for conservation, proliferation and large-scale production of WA. The regenerated plants can also be used in traditional medicine as an alternative to naturally collected plants.

Evaluation of nutritional and antioxidant status of Lepidium latifolium Linn.: a novel phytofood from Ladakh.

Lepidium latifolium Linn. (perennial pepperweed) is one of the preferred phytofoods among cold arid region of Ladakh, India and its leaves contribute significantly to people's diet. This study was conducted to determine its nutritive value and antioxidant activity. Plant samples from three different locations were selected in the present study. Results showed that this plant is an excellent source of glucosinolates, notably sinigrin that is present in very high amount (∼70-90%). Its value ranged from 149 to 199 µg per g fresh weight. Fatty acid composition analysis showed that its leaves were abundant in unsaturated fatty acids, specifically linolenic acid (18∶3) whose percentage is about 50%. Higher glucose and crude protein along with higher nitrogen to sulfur ratio, supplements the nutritive value of this plant. Based on total phenol, flavanoids, free radical scavenging activity and DNA protective activity showed that this ecotype of perennial pepperweed contains high antioxidant properties. The percentage inhibition for O2(-) scavenging activity ranged from 41.3% to 83.9%. Higher content of phenols (26.89 to 50.51 mg gallic acid equivalents per g dry weight) and flavanoids (38.66 to 76.00 mg quercetin equivalents per g dry weight) in leaves could be responsible for the free radical scavenging activity of this plant. Depending upon the location of the plants, variations were observed in different activities. Based on the systematic evaluation in this study, preparations of Lepidium latifolium from Ladakh can be promoted as substitute to dietary requirements.

Protocol for in vitro regeneration and marker glycoside assessment in Swertia chirata Buch-Ham.

We have developed a rapid in vitro propagation system, via axillary shoot formation from nodal explants of Swertia chirata Buch Ham. Culture medium supplemented with 2 mg/L. BAP is best for direct shoot regeneration initially formed adventitious buds from axils of the nodal explants after 30 days. The reduced BAP concentration 0.5 mg/L proliferate shoots effectively. Kept the number of hyperhydrated shoots to minimal and induced on an average 22-38 shoots per flask (4.3 cm average length). The regenerated shoots (5- to 6-m long) formed roots very well in Murashige and Skoog (MS) medium devoid of any growth regulator and followed by acclimatization of plants in pre-sterilized sand containing 1% Trichoderma viride and Azatobactor chrococcum as bioinoculants. The regenerated plants don't show any genomic alterations. This protocol also outlines procedure of assessment of marker iridoid glycosides (amarogentin and amaroswerin) from callus, roots, multiple shoots, regenerated plants, and mother plant. High propagation frequency, reproducibility of procedure, molecular, and phenotypic and chemical stability ensures the efficiency of the developed protocol.