Gokare A. Ravishankar

Enhancement of growth and coumarin production in hairy root cultures of witloof chicory (Cichorium intybus L.cv.Lucknow local) under the influence of fungal elicitors

Studies were conducted to determine the possible production of coumarins in hairy root cultures of Cichorium intybus L.cv. Lucknow local under the influence of microbial agents. Pythium aphanidermatum and Phytopthora parasitica var. nicotiana were cultured and their mycelial and medium filtrate were used for the elicitation of coumarin production. The media filtrate of P. parasitica at 1.0% v/v added to the MS basal medium led to the maximum growth of hairy roots of C. intybus, which was 1.57-fold higher than the untreated control on the 28th day of culture, along with maximum esculin and esculetin yields which were 4.06- and 3.71-fold higher than the control on the 28th day. The yields of esculin and esculetin in hairy root cultures of chicory strongly correlated with growth. To check the effect of these fungal elicitors on endogenous polyamine metabolism, titers of total endogenous polyamines were analyzed. It was confirmed that the media filtrate of P. parasitica at 1.0% v/v concentration resulted in maximum accumulation of total endogenous polyamines, wherein endogenous spermine titers were found to be maximum as compared to endogenous spermidine and putrescine titers on the 28th day. Total endogenous spermine in the case of 1.0% MF v/v P. parasitica was 1.3-fold higher than that of the control on the 28th day of culture. Maximum growth index with greater length of primary root (17.61+/-0.18 cm) and greater number of secondary and tertiary roots was recorded for the medium filtrate of P. parasitica at 1.0% v/v concentration. This study also provided an insight into the morphological changes in terms of branching patterns, occurring in roots under the influence of these fungal elicitors.

Putrescine Influences Growth and Production of Coumarins in Hairy Root Cultures of Witloof Chicory (Cichorium intybus L. cv. Lucknow Local)

Abstract. The effect of putrescine (Put) on the growth and production of two coumarins, esculin and esculetin, in hairy roots of chicory (Cichorium intybus L. cv. Lucknow local) was examined. To study the role of Put on growth and production of coumarins, polyamine inhibitors, namely α-dl-difluromethylornithine and α-dl-difluromethylarginine were used at 1 mM concentration. Put treatment at 1.5 mM produced a 1.9-fold increase in the growth of hairy roots, as well as the production of esculin and esculetin. The treatments with polyamine (PA) inhibitors resulted in much lower growth and production of coumarins compared with both 1.5-mM Put treatment and the control. Both free and conjugated PAs were studied over the whole culture period, and conjugates of all three PAs, namely Put, spermidine, and spermine, were higher than free PAs throughout the culture period. The treatments with PA inhibitors showed lower levels of endogenous PAs compared with Put-treated samples. The treatment with 1.5 mM Put showed maximum accumulation of endogenous conjugated Put (2,098 ± 157 nmoles gm−1 fresh weight). The production of esculin and esculetin was strictly correlated with growth in all treatments. Put at 1.5 mM resulted in greater length of primary root (18.3 ± 1.4 cm) as compared with the control (11 ± 0.9 cm) and larger numbers of secondary and tertiary roots.

Effect of the Carotenoid-Producing Alga, Dunaliella bardawil, on CCl4-Induced Toxicity in Rats

Dunaliella bardawil is a carotenoid-producing alga that is being considered for use in nutraceuticals. To evaluate potential protective effects of consumption of this alga, rats were treated with two different doses of D. bardawil (2.5 and 5.0 g kg–1 body weight [bw]) as a biomass suspension daily for 14 days. Animals were tested against Carbon tetrachloride (CCl4; 2 ml kg–1)–induced liver toxicity as measured by various biochemical marker enzymes in liver and blood. All measurements were taken 6 h following the single dose of CCl4. The results of this study show that there was a slight, but statistically significant mean serum enzyme values, with D. bardawil treatment, compared to higher mean values in animals receiving CCl4 alone. Lipid peroxidation is measured by thiobarbituric acid–reactive substance (TBARS) activity was likewise slightly less elevated with algae treatment. The results also demonstrated protection against DNA strand breaks in hepatocytes, as measured by single cell gel electrophoresis. Liver histopathology was less severe with D. bardawil treatment, supporting the apparent protective action of 14-day treatment on hepatic oxidative injury.

Putrescine facilitated enhancement of capsaicin production in cell suspension cultures of Capsicum frutescens.

Putrescine treatment (0.1 mmol/L) influenced enhancement of growth and capsaicin production in the cell suspension cultures of C. frutescens. The administration of polyamine inhibitor DFMA (alpha-DL-difluoromethylarginine) resulted in a reduction of the growth, capsaicin content and the endogenous titres of polyamines (PAs). The capsaicin synthase activity was also higher in the putrescine (Put) treated cultures. Ethylene levels were lower in the cultures treated with putrescine. This study suggested that Put facilitates growth and capsaicin production.

Precursor Biotransformation in Immobilized Placental Tissues of Capsicum frutescens Mill: I. Influence of Feeding Intermediate Metabolites of the Capsaicinoid Pathway on Capsaicin and Dihydrocapsaicin Accumulation

Summary In order to enhance the levels of capsaicinoids in vitro and to optimise the suitable precursor, immobilized placental tissues of Capsicum frutescens were fed with intermediates of the capsaicinoid pathway. Feeding t-cinnamic acid, p-coumaric acid, caffeic acid or ferulic acid (2.5 mM final concentration) resulted in a greater accumulation of capsaicin and dihydrocapsaicin than was achieved with the distant precursors phenylalanine and 1-valine. Of all the precursors, coumaric acid was found to be the best for capsaicin and dihydrocapsaicin formation, resulting in an accumulation of 2890.8 μg/culture and 818 μg/culture, respectively, in 5 days of culture. Maximum conversion to capsaicin and dihydrocapsaicin was observed in cinnamic acid treated cultures, which was 82 % and 27.5 %, respectively. In the present study, a production of 1.188 mg capsaicin/g DW of placental tissue/day was achieved without the addition of any precursor and a production of 5.78 mg capsaicin/g DW of placental tissue/day by treating with coumaric acid; this was 11-fold higher than in chilli pepper fruit. These results demonstrate successful culture of placenta, the site of synthesis of capsaicin, to boost levels of capsaicin and dihydrocapsaicin in vitro .

Augmentation of major isoflavones in Glycine max L. through the elicitor-mediated approach

Abstract - Isoflavone content in soybean seeds was enhanced by the elicitor-mediated approach under field conditions through the floral application of abiotic elicitors-salicylic acid, methyl jasmonate and biotic elicitors-Aspergillus niger and Rhizopus oligosporus. Among isoflavones, daidzein and glycitein were found to be highly responsive to elicitors, with an increase of 53.7% and 78.7%, respectively as compared to control. Highest total isoflavone content (1276.4 mg g-1 of seeds) was observed upon the administration of 0.1 mMsalicylic acid, which is 92.7% higher than in control. This study would be valuable for augmentation of the isoflavone content in soybean seeds in field grown plants for better nutraceutical potential.