Sushama A. Patil

Mechanochemical growth of a porous ZnFe2O4 nano-flake thin film as an electrode for supercapacitor application

Herein, we are reporting a simple, economic, easy to handle, scalable and reproducible mechanochemical i.e. rotational chemical bath deposition (R-CBD) approach for the synthesis of well adhered nano-flake ZnFe2O4 thin films (NFs-ZnFe2O4) with uniform morphology on a stainless steel (SS) substrate, in comparison with nano-grain ZnFe2O4 thin films (NGs-ZnFe2O4) prepared using a conventional CBD approach. The influence of rotation on the evolution of the nano-flake morphology in NFs-ZnFe2O4 is also investigated. The porous NFs-ZnFe2O4 thin films demonstrated excellent pseudocapacitor properties with higher specific capacitance of 768 F g−1 at high current density of 5 mA cm−2, stability upto 5000 cycles (88% retention), higher energy density (106 W h kg−1) and power density (18 kW kg−1) compared to NGs-ZnFe2O4. The results were also found to be higher than those reported earlier for MFe2O4 based systems.

Dilute acid pretreatment of rice straw, structural characterization and optimization of enzymatic hydrolysis conditions by response surface methodology

Efficient conversion of fermentable sugars from cheap lignocellulosic biomass is a current need in viable ethanol production technology. In the present study, agricultural waste biomass such as rice straw was pretreated by using 0.5% sulfuric acid for 60 min at 121 °C in an autoclave. A statistical experimental design like central composite design (CCD) was used for optimization of the enzymatic hydrolysis conditions to achieve a significant reducing sugar yield using commercial cellulase. The optimal conditions for acid pretreated rice straw were found to be 40 FPU g−1 enzymes loading, 17.50% biomass loading at 50 °C for 72 h. The reducing sugar yield was 0.359 g g−1, achieved at the optimized conditions. Experimental results under optimum conditions fit well with CCD model predictions. The structural and morphological changes in native and dilute acid treated rice straw substrate were evaluated by FTIR, XRD and SEM analysis. The XRD pattern of biomass revealed an increase in the crystallite size and crystallinity index of pretreated biomass. Scanning electron micrography reported surface porosity and a distorted structure due to pretreatment. HPTLC analysis of sugars like glucose and xylose in hydrolysate produced after enzymatic hydrolysis was determined.

Evaluation of crocin and curcumin affinity on mushroom tyrosinase using surface plasmon resonance.

Tyrosinase inhibitors have potential applications in the cosmetics and food industries for preventing browning reactions and also as therapeutic drugs for neurodegenerative diseases such as Parkinsons. In this article, crocin and curcumin were evaluated as mushroom tyrosinase inhibitors. Results showed that, both compounds strongly inhibited the diphenolase activity than monophenolase. The IC50 values for diphenolase activity were estimated to be 0.11 mM and 0.18 mM for crocin and curcumin respectively. The binding kinetics of crocin and curcumin was studied with mushroom tyrosinase using surface plasmon resonance (SPR). Tyrosinase was immobilized on the gold surface of a Biacore sensor chip through amine coupling. Binding of inhibitors was analyzed by SPR without the need to further modify the surface or the use of other reagents. The binding constant KD (M) for mushroom tyrosinase obtained was 1.21×10(-4) M for crocin and 1.64×10(-4) M for curcumin, while showing a higher affinity for L-DOPA 1.95×10(-8) M, a substrate for tyrosinase (positive control). The study reveals the SPR sensors ability to detect binding of the inhibitors.

Optimization of l‐DOPA production by Brevundimonas sp. SGJ using response surface methodology

l‐DOPA (3,4‐dihydroxyphenyl‐l‐alanine) is an extensively used drug for the treatment of Parkinsons disease. In the present study, optimization of nutritional parameters influencing l‐DOPA production was attempted using the response surface methodology (RSM) from Brevundimonas sp. SGJ. A Plackett–Burman design was used for screening of critical components, while further optimization was carried out using the Box–Behnken design. The optimized levels of factors predicted by the model were pH 5.02, 1.549 g l−1 tryptone, 4.207 g l−1 l‐tyrosine and 0.0369 g l−1 CuSO4, which resulted in highest l‐DOPA yield of 3.359 g l−1. The optimization of medium using RSM resulted in a 8.355‐fold increase in the yield of l‐DOPA. The anova showed a significant R2 value (0.9667), model F‐value (29.068) and probability (0.001), with insignificant lack of fit. The highest tyrosinase activity observed was 2471 U mg−1 at the 18th hour of the incubation period with dry cell weight of 0.711 g l−1. l‐DOPA production was confirmed by HPTLC, HPLC and GC‐MS analysis. Thus, Brevundimonas sp. SGJ has the potential to be a new source for the production of l‐DOPA.

Production of camptothecine using whey by an endophytic fungus: standardization using response surface methodology

Fusarium oxysporum kolhapuriensis, a novel endophytic fungi isolated from Nothapodytes nimmoniana Mabb. Grahm, was found to produce camptothecine (CPT) using whey as a complex medium. The highest production of CPT was attained using statistical methods Response Surface Methodology (RSM). Central Composite Design (CCD) was used to optimize the complex medium and culture conditions for maximum production of CPT by the fungus. The optimized medium that yielded 283 ± 0.27 mg l−1 of CPT contained 70% (v/v) of acid whey and 2% (w/v) malt extract. The other two culture parameters optimized through RSM were temperature (30 °C) and period of incubation (6 days). The production of CPT was confirmed by analytical techniques such as HPTLC, HPLC and LC-HRMS. This cost effective optimized medium using RSM might be useful for large scale CPT production which will ultimately reduce the further downstream processing cost.

Interaction of small molecules with fungal laccase: A Surface Plasmon Resonance based study.

Laccases have a great potential for use in industrial and biotechnological applications. It has affinity towards phenolics and finds major applications in the field of bioremediation. Here, Surface Plasmon Resonance (SPR) as a biosensor with immobilized laccase on chip surface has been studied. Laccase was immobilized by thiol coupling method and compounds containing increasing number of hydroxyl groups were analyzed for their binding affinity at various concentrations in millimolar range. The small molecules like phloroglucinol (1.532×10(-8) M), crocin (3.204×10(-3) M), ascorbic acid (8.331×10(-8) M), kojic acid (6.411×10(-7) M) and saffron (3.466×10(-7) M) were studied and respective KD values are obtained. The results were also confirmed by inhibition assay and IC50 values were calculated. All these molecules showed different affinity towards laccase in terms of KD values. This method may be useful for preliminary screening and characterization of small molecules as laccase substrates, inhibitors or modulators of activity. This method will be useful for rapid screening of phenolics in waste water because of high sensitivity.

Screening of inhibitors for mushroom tyrosinase using surface plasmon resonance.

Tyrosinase inhibitors have been used as whitening or antihyperpigment agents because of their ability to suppress dermal-melanin production. In the present study, screening and kinetic evaluation of various small molecules were performed on mushroom tyrosinase (MT) using surface plasmon resonance. The binding constant KD (M) values obtained for tannic acid, phloroglucinol, saffron, catechol, and pyrogallol are 1.213 × 10(-4), 7.136 × 10(-5), 3.111 × 10(-5), 1.557 × 10(-5), and 7.981 × 10(-6) M, respectively. Pyrogallol has been found to display high affinity for MT, whereas catechol, saffron, and phloroglucinol have been found to bind with low affinity. MT shows considerable changes in the secondary structure in the presence of inhibitors. The study reveals the Biacore/SPR sensors ability in the rapid identification and characterization of inhibitors for MT. The methodology described here can be used to rapidly screen and optimize various lead compounds for other enzymes and elucidate structure function inter-relationships between various enzymes.

Interaction of small molecules with human tyrosinase: A surface plasmon resonance and molecular docking study

Studies on tyrosinase have recently gained the attention of researchers due to the enzymes biological functions and potential applications in food and cosmetic applications. In this present study, screening and kinetic evaluation of small molecules (>300Da) on human tyrosinase was carried out using surface plasmon resonance and molecular docking studies. Four molecules showed significant binding were further characterized. The binding constant KD (M) values obtained for crocin, curcumin, tannic acid, pyrogallol and hydroquinone are 5.60×10-5, 1.52×10-3, 6.45×10-5, 1.34×10-5 and 2.433×10-7M respectively. In silico docking studies using autodock indicated significant binding and revealed the binding pocket residues for all molecules. The study shows the Biacore/SPR sensors ability for screening and detection of inhibitors for human tyrosinase. This study can be used to rapidly screen and optimize various lead compounds as binders/inhibitors/modulators of human tyrosinase enzyme activity.

Statistical optimization of process parameters for inulinase production from Tithonia weed by Arthrobacter mysorens strain no.1

Tithonia rotundifolia is an easily available and abundant inulin rich weed reported to be competitive and allelopathic. This weed inulin is hydrolyzed by inulinase into fructose. Response surface methodology was employed to optimize culture conditions for the inulinase production from Arthrobacter mysorens strain no.1 isolated from rhizospheric area of Tithonia weed. Initially, Plackett- Burman design was used for screening 11 nutritional parameters for inulinase production including inulin containing weeds as cost effective substrate. The experiment shows that amongst the 11 parameters studied, K2HPO4, Inulin, Agave sisalana extract and Tithonia rotundifolia were the most significant variables for inulinase production. Quantitative effects of these 4 factors were further investigated using Box Behnken design. The medium having 0.27% K2HPO4, 2.54% Inulin, 6.57% Agave sisalana extract and 7.27% Tithonia rotundifolia extract were found to be optimum for maximum inulinase production. The optimization strategies used showed 2.12 fold increase in inulinase yield (1669.45 EU/ml) compared to non-optimized medium (787 EU/ml). Fructose produced by the action of inulinase was further confirmed by spectrophotometer, osazone, HPTLC and FTIR methods. Thus Tithonia rotundifolia can be used as an eco-friendly, economically feasible and promising alternative substrate for commercial inulinase production yielding fructose from Arthrobacter mysorens strain no.1.

Exploration of surface plasmon resonance for yam tyrosinase characterization

Tyrosinase is a ubiquitous enzyme in nature. It catalyzes the reaction in melanin formation. In this investigation, tyrosinase from a Yam tuber (Amorphophallus paeoniifolius) was extracted and purified by ultra-centrifugal filtration followed by ion exchange chromatography. The purified enzyme was obtained with purification fold of 12.65 and specific activity of 60.25 U/mg. The molecular weight of purified enzyme was confirmed to be 45 kDa by SDS-PAGE. The enzyme activity was optimal at pH 6 and 30 °C with Km 10 mM. Further the kinetic study of purified yam tyrosinase was carried out using Surface Plasmon Resonance method. In brief, enzyme was immobilized on chip surface by amine coupling method and different small molecules were analyzed for their binding affinities at different concentrations (mM). The affinity of each compound for yam tyrosinase was different with KD values as tannic acid (5.13 × 10-5), gallic acid (2.05 × 10-8), ascorbic acid (0.004544), caffeic acid (3.09 × 10-9), pyrogallol (2.13 × 10-4) and catechol (1.09 × 10-4). The kinetics data results were confirmed by inhibition assays and IC50 values were calculated. This data will help to study the role of tyrosinase in hyperpigmentation which will create an avenue for tyrosinase inhibitors. Tyrosinase inhibitors have wide range of applications in cosmetics, medical and food industries.