Anupam Tiwari

Photo-induced biosynthesis of silver nanoparticles from aqueous extract of Dunaliella salina and their anticancer potential

The synthesis of silver nanoparticles (AgNPs) via green route, using biological entities is an area of interest, because one of the potential applications in the nanomedicine. In the present study, we have developed photo-induced, ecofriendly, low cost method for biosynthesis of the stable silver nanoparticles using aqueous extract of Dunaliella salina (AED) which act as both reducing as well as stabilizing agent. Biosynthesis of the AgNPs was optimized as: sunlight exposure (30min), AED (5% (v/v)) and AgNO3 (4mM). Biosynthesis of AgNPs was monitored by using UV-Vis spectroscopy which exhibited sharp SPR band at 430nm after 30min of bright sunlight exposure. SEM and TEM analyses confirmed the presence of spherical AgNPs with average size of 15.26nm. Crystalline nature of AgNPs was confirmed by SAED and XRD analyses where Braggs reflection pattern at (111), (200), (220) and (311) corresponded to face centered cubic crystal lattice of metallic silver. FTIR analysis revealed the involvement of various functional groups present in AED. AFM analysis confirmed the average surface roughness of synthesized AgNPs as 8.48nm. AgNPs were also screened for anticancer potential using assay of calcein AM/PI, Annexin/PI and cancer biomarkers against cancer cell line (MCF-7), while normal cell line (MCF-10A) were kept as control. Interestingly, anticancer potential was comparable to the known anticancer drug (Cisplatin), and was not detrimental to the normal cell line. Therefore, such green synthesized AgNPs may be explored as anticancer agent.

Role of calcium in the mitigation of heat stress in the cyanobacterium Anabaena PCC 7120.

The effects of exogenously added CaCl2 (0.25mM) on photopigments, photosynthetic O2-evolution, antioxidative enzyme activity, membrane damage, expression of two heat shock genes (groEL and groES) and apoptotic features in Anabaena 7120 under heat stress (45°C) for up to 24h were investigated. Heat stress lowered the level of photopigments; however, Ca2+--supplemented cultures showed a low level reduction in Chl a but induced accumulation of carotenoids and phycocyanin under heat stress. Photosynthetic O2-evolving capacity was maintained at a higher level in cells from Ca2+-supplemented medium. Among the antioxidative enzymes, superoxide dismutase activity was unaffected by the presence or absence of Ca2+ in contrast to increases in catalase, ascorbate peroxidase and glutathione reductase activities in cells grown in Ca2+-supplemented medium. Lower levels of lipid peroxidation were recorded in Anabaena cells grown in Ca2+-supplemented medium in comparison to cells from Ca2+--deprived medium. Target cells grown in Ca2+-deprived medium developed apoptotic features in the early stages of heat shock, while Ca2+ application seemed to interfere with apoptosis because only a few cells showed such features after 24 h of heat exposure, indicating a role for Ca2+ in maintaining cell viability under heat stress. There was also continuous up regulation of two important heat shock genes (groEL and groES) in Ca2+-supplemented cultures, exposed to heat shock, again indicating a role for Ca2+ in stress management.

Proline biosynthesizing enzymes (glutamate 5-kinase and pyrroline-5-carboxylate reductase) from a model cyanobacterium for desiccation tolerance

Drought is the most important abiotic stress, challenging sustainable agriculture globally. For desiccation being the multigenic trait, a combination of identified genes from the appropriate organism may render crop tolerant to the water stress. Among the compatible solutes, proline plays multifaceted role in counteracting such stress. The genes encoding proline biosynthesizing enzymes, glutamate 5-kinase (G5K), and pyrroline-5-carboxylate reductase (P5CR) from the low-desiccation-tolerant cyanobacterium Anabaena sp. PCC 7120, were cloned and overexpressed in Escherichia coli BL21(DE3) individually. The recombinant E. coli cells harboring G5K, failed to exhibit enhanced desiccation tolerance relative to those with P5CR that showed increased growth/survival over the wild type. This may be ascribed to the overexpression of the reductase gene. Multiple sequence alignment showed P5CR to be conserved in all the organisms. We hypothesize that P5CR gene from high-desiccation-tolerant cyanobacteria may be adopted as the candidate for making transgenic N2-fixing cyanobacterium for paddy fields and/or crop development in future.

Mass cultivation of Dunaliella salina in a flat plate photobioreactor and its effective harvesting

Mass cultivation of Dunaliella salina was standardized in a flat plate photobioreactor followed by a vertical flat plate photobioreactor. Maximum biomass productivity (14.95 ± 0.43 mgL-1d-1 dry cell weight) was achieved in the latter at inoculum concentration of OD680nm = 0.1, 100 µmolm-2s-1 light illumination and 1.0 L min-1 aeration. Semicontinuous operation with varying KNO3 and NaHCO3 concentrations resulted highest biomass productivity (17.85 ± 0.55 mgL-1d-1) at 0.50 mM NaHCO3 and 15 mM KNO3. However, maximum lipid (16.36 ± 1.18% dry cell weight) was achieved at 0.75 mM NaHCO3 and 10 mM KNO3. Flocculation studies employing potash alum, FeCl3·6H2O or pH showed harvesting efficiencies exceeding 90% in 0.75 mM potash alum or FeCl3·6H2O or pH 11, but they yielded low concentration factor (<5) and were detrimental (Fv/Fm < 0.50). A combination of 0.50 mM FeCl3·6H2O and pH 9 was found as most suitable flocculating strategy with maximum concentration factor (>14) and least damaging (Fv/Fm > 0.54).

Effect of Picking Stages on Fruit and Seed Development in Okra [Abelmoschus esculentus (L.) Moench] Cultivars Kashi Pragati and Kashi Kranti

A field experiment was conducted to study the effect of picking on fruit and seed development in okra (Abelmoschus esculentus L. Moench) cv. Kashi Pragati and Kashi Kranti. Data on fruit and seed characters were recorded from 4 to 40 days after flowering (DAF) at an interval of 2 days. Quick fruit development occurs between 4 to 8 DAF and fruits picked at this stage were of optimum fresh weight, length, diameter and free from thick pericarp thus making it consumable. Fresh weight of filled seeds was found to be maximum at 18 days after flowering in Kashi Pragati (7.61 g) whereas it was 22 days after flowering in Kashi Kranti (6.90 g), while dry weight of seeds per pod and 100 seed weight were maximum at 24 days after flowering. The physiological maturity of seeds was attained after 26 days after flowering when the germination percentage in both the cultivars was above 75%. However, to obtain optimum seed yield with optimum germination percentage and vigour, the fruits should be picked on at 38 days after flowering stage.