Effect of carbon and nitrogen sources on in vitro flower and fruit formation and withanolides production in Withania somnifera (L.) Dunal.

We studied the influence of sucrose and nitrogen concentration on in vitro flowering and fruit setting in elongated shoots of Withania somnifera. BA (1.5 mg/l) and IAA (0.3 mg/l) on MS medium supplemented with 4% sucrose showed 67% of in vitro flower induction frequency, 9 flowers/shoot, 4 fruits/shoot and 11 seeds/fruit in elongated-shoots. Different concentrations of nitrogen sources (L-glutamine, adenine sulphate, ammonium nitrate, potassium nitrate and sodium nitrate 5-25 mg/l) were tested in combination with 4% sucrose and BA at 1.5 mg/l and IAA at 0.3 mg/l. Highest number of flowers (20 flowers/shoot; 2.2-fold) and fruits (16 fruits/shoot; 3.39-fold), fruit setting (12 seeds/fruit; 1.08-fold) at a higher frequency (88%) were achieved on MS medium augmented with 15 mg/l adenine sulphate with same PGRs and sucrose concentration. The maximum production of withanolide A (0.68 mg/g DW) and withanolide B (0.77 mg/g DW) was recorded in in vitro fruits. Highest accumulation of withaferin A (2 mg/g DW) was quantified from in vitro flowers, whereas, it was low in in vitro fruits (0.49 mg/g DW withaferin A). However, withanone (0.23 mg/g DW) was found accumulated uniformly in both in vitro flowers and fruits compared to control.

An investigation on the cytotoxicity and caspase-mediated apoptotic effect of biologically synthesized silver nanoparticles using Podophyllum hexandrum on human cervical carcinoma cells.

Now-a-days synthesis and characterization of silver nanoparticles (AgNPs) through biological entity is quite interesting to employ AgNPs for various biomedical applications in general and treatment of cancer in particular. This paper presents the green synthesis of AgNPs using leaf extract of Podophyllum hexandrum Royle and optimized with various parameters such as pH, temperature, reaction time, volume of extract and metal ion concentration for synthesis of AgNPs. TEM, XRD and FTIR were adopted for characterization. The synthesized nanoparticles were found to be spherical shaped with average size of 14 nm. Effects of AgNPs were analyzed against human cervical carcinoma cells by MTT Assay, quantification of ROS, RT-PCR and western blotting techniques. The overall result indicates that AgNPs can selectively inhibit the cellular mechanism of HeLa by DNA damage and caspase mediated cell death. This biological procedure for synthesis of AgNPs and selective inhibition of cancerous cells gives an alternative avenue to treat human cancer effectively.

Optimization of carbon source for hairy root growth and withaferin A and withanone production in Withania somnifera.

This study optimized carbon sources in half MS liquid medium for maximum biomass accumulation and withanolides production in hairy root culture of Withania somnifera. The highest production of withaferin A and withanone was achieved when sucrose and sucrose+glucose were used individually as carbon sources. The hairy root suspension culture supplemented with a lower level of sucrose (2%) favored hairy root biomass accumulation (1.41 g DW) followed by sucrose+glucose (2+1) when compared with other carbon sources in half MS liquid medium after 40 days of culture. The hairy roots grown on sucrose (4%) enriched half MS liquid medium stimulated higher production of withaferin A (2.21 mg/g DW) and withanone (2.41 mg/g DW) on the 40th day of culture, followed by sucrose+glucose (4+1%) compared with glucose, fructose, maltose and other combinations tested.

Optimization of elicitation conditions with methyl jasmonate and salicylic acid to improve the productivity of withanolides in the adventitious root culture of Withania somnifera (L.) Dunal.

Adventitious root cultures derived from leaf derived callus of Withania somnifera (L.) Dunal were treated with methyl jasmonate and salicylic acid independently. Biomass accumulation, culture age, elicitation period, and culture duration were optimized for higher withanolides production in the two best-responding varieties collected from Kolli hills (Eastern Ghats) and Cumbum (Western Ghats) of Tamil Nadu, India. Between the two elicitors, salicylic acid (SA) improved the production of major withanolides (withanolide A, withanolide B, withaferin A, and withanone) as well as minor constituents (12-deoxy withastramonolide, withanoside V, and withanoside IV) in the Kolli hills variety. Treatment of root biomass (11.70 g FW) on 30-day-old adventitious root cultures with 150 µM SA for 4 h elicitor exposure period resulted in the production of 64.65 mg g(-l) dry weight (DW) withanolide A (48-fold), 33.74 mg g(-l) DW withanolide B (29-fold), 17.47 mg g(-l) DW withaferin A (20-fold), 42.88 mg g(-l) DW withanone (37-fold), 5.34 mg g(-l) DW 12-deoxy withastramonolide (nine fold), 7.23 mg g(-l) DW withanoside V (seven fold), and 9.45 mg g(-l) DW withanoside IV (nine fold) after 10 days of elicitation (40th day of culture) when compared to untreated cultures. This is the first report on the use of elicitation strategy on the significant improvement in withanolides production in the adventitious root cultures of W. somnifera.