Dhananjay Kumar Singh

PsAP2 an AP2/ERF family transcription factor from Papaver somniferum enhances abiotic and biotic stress tolerance in transgenic tobacco

The AP2/ERFs are one of the most important family of transcription factors which regulate multiple responses like stress, metabolism and development in plants. We isolated PsAP2 a novel AP2/ERF from Papaver somniferum which was highly upregulated in response to wounding followed by ethylene, methyl jasmonate and ABA treatment. PsAP2 showed specific binding with both DRE and GCC box elements and it was able to transactivate the reporter genes in yeast. PsAP2 overexpressing transgenic tobacco plants exhibited enhanced tolerance towards both abiotic and biotic stresses . Real time transcript expression analysis showed constitutive upregulation of tobacco Alternative oxidase1a and Myo-inositol-1-phosphate synthase in PsAP2 overexpressing tobacco plants. Further, PsAP2 showed interaction with NtAOX1a promoter in vitro, it also specifically activated the NtAOX1a promoter in yeast and tobacco BY2 cells. The silencing of PsAP2 using VIGS lead to significant reduction in the AOX1 level in P. somniferum. Taken together PsAP2 can directly bind and transcriptionally activate NtAOX1a and its overexpression in tobacco imparted increased tolerance towards both abiotic and biotic stress.

Antiproliferative and antimicrobial efficacy of the compounds isolated from the roots of Oenothera biennis L.

Oenothera biennis L., commonly known as evening primrose, harbours the flavonoids, steroids, tannins, fatty acids and terpenoids responsible for a diverse range of biological activity, such as antitumour, anti‐arthritic and anti‐inflammatory effects. In addition to the previous reports from aerial parts of this plant, studies related to antiproliferative or antimicrobial activity from the roots are warranted.

Advances in Boerhaavia diffusa hairy root technology: a valuable pursuit for identifying strain sensitivity and up-scaling factors to refine metabolite yield and bioactivity potentials

The present study reports the Agrobacterium rhizogenes-mediated hairy root induction in of an ethno-medicinally significant herb–Boerhaavia diffusa L., for elucidating the underlying competence regarding its biosynthetic (i.e. boeravinone B and eupalitin) and bioactivity (antibacterial, antioxidant and anti-inflammatory) potentials. Host plant-specific receptiveness towards A. rhizogenes strains and disparity in compatibility threshold of leaf and nodal explants were evident. Only leaf explants responded, attaining hairy root induction with the ATCC 15834 followed by A4 and SA79 strains in reducing order of transformation efficiency. The growth behaviours differed amongst independent rhizoclones, and two clones of A4 (RBH) and ATCC 15834 (RBT8) origin demonstrated higher growth potentials. Polymerase chain reaction amplification of rol genes confirmed their transformed nature. Optimization of the appropriate solvent and reverse phase high-performance liquid chromatography parameters relating to the targeted metabolite production in the selected RBH and RBT8 clones revealed higher accumulation of eupalitin with the RBH clone having the best result of 1.44 times greater yield over the control root. Compared to the selected rhizoclones, the control roots however showed higher boeravinone B content. Devising a modified “stirred-tank” reactor through equipping with marine impellers and ring spargers facilitated high-density RBH root biomass yield with 6.1-fold and 1.15-fold yield increment of the boeravinone B and eupalitin respectively compared to shake-flask cultures. Considering the control roots, the RBH clone revealed analogous antioxidant/antibacterial activities with improved anti-inflammatory potential. The hairy root mediated higher production of boeravinone B and eupalitin could be achieved for the first time in bioreactor.

Development, optimization, and characterization of a novel tea tree oil nanogel using response surface methodology

Abstract Purpose: To develop and optimize nanoemulsion (NE)-based emulgel (EG) formulation as a potential vehicle for topical delivery of tea tree oil (TTO). Methodology: Central composite design was adopted for optimizing the processing conditions for NE preparation by high energy emulsification method viz. surfactant concentration, co-surfactant concentration, and stirring speed. The optimized NE was developed into emulgel (EG) using pH sensitive polymer Carbopol 940 and triethanolamine as alkalizer. The prepared EG was evaluated for its pH, viscosity, and texture parameters, ex vivo permeation at 37 °C and stability. Antimicrobial evaluation of EG in comparison to conventional gel and pure TTO was also carried out against selected microbial strains. Results and discussion: Optimized NE had particle size and zeta potential of 16.23 ± 0.411 nm and 36.11 ± 1.234 mV, respectively. TEM analysis revealed the spherical shape of droplets. The pH of EG (5.57 ± 0.05 ) was found to be in accordance with the range of human skin pH. EG also illustrated efficient permeation (79.58 μL/cm2) and flux value (JSS) of 7.96 μL cm2/h through skin in 10 h. Viscosity and texture parameters, firmness (9.3 ± 0.08 g), spreadability (2.26 ± 0.06 mJ), extrudability (61.6 ± 0.05 mJ), and adhesiveness (8.66 ± 0.08 g) depict its suitability for topical application. Antimicrobial evaluation of EG with same amount of TTO as conventional gel revealed broader zones of growth inhibitions against all the selected microbial strains. Moreover, EG was also found to be nonirritant (PII 0.0833). These parameters were consistent over 90 d. Conclusion: TTO EG turned out to be a promising vehicle for the topical delivery of TTO with enhanced therapeutic efficacy.

Rutin protects t-butyl hydroperoxide-induced oxidative impairment via modulating the Nrf-2 and iNOS activity

BACKGROUND Rutin (quercetin-3-O-rutinoside), a flavonoid, is predominantly found in the buckwheat, cranberries, mulberry and citrus fruits. It is used as a restorative in the preparation of herbal medicine, multivitamin and known to reduce the fate of heart attack. HYPOTHESIS We aimed to elucidate whether rutin attenuates oxidative stress and its possible mechanism of action in ameliorating the deleterious effect of t-BHP. We also provide evidence that rutin protects the antioxidant status of erythrocytes and liver via Nrf2 and iNOS pathway from oxidative stress. STUDY DESIGN/METHOD Human erythrocytes and mice liver were used for the evaluation of rutins effect against t-BHP induced oxidative stress. The non-enzymatic (GSH, MDA, -CO, -SH) and enzymatic stress markers (SOD, CAT, GPx, GR and GST) were estimated by the colorimetric method. The level of Nrf2, iNOS, liver marker enzymes, triglycerides, cholesterol, HDL-cholesterol, albumin, BUN was measured using ELISA kits. Reactive oxygen species (ROS) was quantified using flow cytometry and fluorometry. RT-PCR was used for the quantification of Nrf2 and iNOS expression levels in the liver tissue of mice. In silico studies were done through receptor-ligand binding interaction. RESULTS Pre-treatment with the rutin ameliorated the toxic effect of t-BHP by modulating the basal level of GSH, -SH, MDA and -CO significantly (p < 0.01) with respect to untreated control. Rutin also protected the erythrocytes against the t-BHP-induced oxidative stress as evidenced by augmented activity of antioxidant enzymes (CAT, SOD, GPX, GR and GST). Furthermore, at the highest tested concentration (16.3 µM), it protected the morphology of the erythrocytes by decreasing the ROS level (p < 0.01). In addition, the lower MEF values of rutin (0.520 ± 0.005) alone or along with t-BHP (0.630 ± 0.021) indicated its non-toxic and protective behavior. The qPCR analyses revealed that t-BHP potently up-regulates the iNOS and down regulate the Nrf2 expression which was ameliorated with rutin treatment in a dose-dependent manner like silymarin. CONCLUSION Our findings demonstrate that rutin potentiates its beneficial aspect by displaying a profound role in iNOS-Nrf2 signaling pathway. Accordingly, it may be concluded that the dietary factors wherein rutin is an ingredient could be helpful in the maintenance of the intracellular redox-homeostasis and thus may be effective against oxidative stress related secondary complications.

Antioxidant and Choline Esterase Inhibitory Activity of Phenolic Rich Extracts from Bombax ceiba L. Flowers

Cite this article: Sinha S, Kumar B, Singh DK, Luqman S, Singh M, Singh A. Antioxidant and Choline Esterase Inhibitory Activity of Phenolic Rich Extracts from Bombax ceiba L. Flowers. Free Radicals and Antioxidants. 2018;8(2):135-40. ABSTRACT Background: Cognition impairment is the most recurrent form of dementia in aged people indicated by permanent neuronal loss and atypical behaviour. Disparities in cholinergic pathway have been reported as major cause of cognition impairment, where deficiency of acetylcholine occurs due to hydrolysis of acetylcholine by cholinesterases. Bombax ceiba (Bombacaceae) commonly known as silk cotton tree is an imperative plant of tropical and subtropical region which have been mentioned in the traditional systems of medicine. Objectives: We have investigated the cholinesterase and antioxidant activity of B. ceiba extracts. Besides these activity evaluations, preliminary phytochemical testing and quantification of total phenolic and flavonoids have also been performed. Method: Extracts from B. ceiba flowers in hexane and ethanol were prepared by cold maceration method. In vitro cholinesterase activity was estimated with the help of Ellman’s reagent, antioxidant potential of extracts was determined by well known method such as DPPH, FRAP, reducing power and total antioxidant power assay. Total phenolics and flavonoids were quantified by colorimetric method. Results: Finding of present study indicated that B. ceiba extracts are rich in polyphenolic contents (19.10 ± 0.74 QcE and 28.01 ± 1.28 GaE). Results of antioxidative potential evaluation suggested that these extracts have high free radical scavenging potential (65.49 ± 2.49%) and are also able to reduce iron like radicals (93.78 FSE). Beside antioxidant potential B. ceiba extracts also inhibited the cholinesterase effectively (IC50 31.22 ± 1.42 μg/ml). Conclusion: Current investigation on B. ceiba flowers indicated that phenolic rich extracts could be used in development of effective plant-based cholinesterase inhibitors.