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Zinc oxide particles (ZnOPs) of both nanometer and sub-micron sizes are important components of high demand consumer products such as sunscreen, paint, textile, food packaging, and agriculture. Their ultimate discharge in the aquatic ecosystem is nearly unavoidable. For sustainable use of ZnOPs, there is an urgent need to assess its ecotoxicity using ecological indicator organisms. Moina macrocopa, an important component of the aquatic ecosystem is one such less explored indicator organism. In the present investigation, ZnOPs of two different sizes (250 ± 20 and 500 ± 50 nm) were selected for risk assessment as most of the previous reports were based on the use of 10-100 nm ZnOPs. ZnOPs of 500 nm were more lethal than that of 250 nm size, with respective LC 50 of 0.0092 ± 0.0012 and 0.0337 ± 0.0133 mg/L against M. macrocopa after 48 h of exposure. We further used a sublethal concentration of 500 nm (0.00336 mg/L) and 250 nm (0.00092 mg/L) ZnOPs followed by measurement of enzymatic biomarkers of toxicity (acetylcholinesterase, digestive enzymes, antioxidant enzymes). A size-dependent variation in enzymatic response to 250 and 500 nm ZnOPs was seen. Exposure to ZnOPs inhibited acetylcholinesterase and digestive enzymes (trypsin, amylase), and elevated antioxidant enzymes (catalase, glutathione S-transferase) levels. The exposure also decreased the superoxide dismutase activity and increased that of β-galactosidase. Microscopic investigation revealed the accumulation of ZnOPs in the digestive tract of M. macrocopa that possibly disrupts enzyme activities. The present study will contribute to establishing regulatory policy on the maximum permissible limit of ZnOPs in different water bodies., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Chitosan, a functional biopolymer, was irradiated with 100 kGy gamma irradiation and used to access its physical, antioxidant, plant growth promoting and antimicrobial properties. The molecular weight of chitosan reduced to 82.2 kDa from 337.7 kDa after irradiation. UV-Vis spectroscopy and FTIR results revealed slight changes in chitosan skeleton after irradiation but the degree of acetylation of both chitosan was ~18%. DSC profile indicated a prominent decline in enthalpy and energy for phase transition, TGA indicated shift in decomposition temperature, while XRD analysis showed a reduction in chitosan crystallinity after irradiation. DPPH and ABTS radical scavenging activity of chitosan (2-10 mg/mL) enhanced significantly by 1.25-1.45 and 1.80-3.14 folds after irradiation. There was a considerable improvement in morphological parameters (number of leaves, nodes, height, fresh weight and dry weight) and biochemical parameters (chlorophyll, total soluble sugars and soluble proteins content) of in vitro potato plant in chitosan supplemented medium at 75 mg/L concentration than the control. The minimum inhibitory concentration of normal and irradiated chitosan for Alternaria spp. was 2500 and 2000 mg/L and for Fusarium spp. was 1750 and 1500 mg/L, respectively. IC 50 value of normal and irradiated chitosan for Fusarium spp. was 1387.9 ± 9.2 and 954.3 ± 6.1 mg/L, and for Alternaria spp. was 1536.1 ± 24.3 and 1416.8 ± 3.5 mg/L, respectively., (Copyright © 2019 Elsevier B.V. All rights reserved.)

A tri-enzyme mixture of cellulase, pectinase and xylanase present in fermentation broth of Aspergillus niger culture was directly recovered by solvent precipitation and co-immobilized on magnetic nanoparticles (MNPs) by cross linking with glutaraldehyde. A 2:1 ratio of isopropanol:total protein in the fermentation broth gave a precipitate with maximal activity recovery of the three enzymes. The co-immobilization on MNPs was optimized at 30 mM glutaraldehyde for 3 h incubation, and 3:1 MNPs:enzyme ratio, yielding a maximum activity recovery of cellulase (80.25 ± 1.03%), pectinase (84.76 ± 1.71%) and xylanase (75.62 ± 0.76%), respectively. The tri-enzyme co-immobilized MNPs were characterized by particle size analysis, FTIR, XRD, VSM and SEM studies, and showed enhanced thermal stability as evident from lower k d , higher t 1/2 and D-value in the range of 55-75 °C. The tri-enzyme co-immobilized MNPs showed a shift in optimum temperature from 55 to 60 °C with improved pH tolerance along with a slender rise in kinetic constants and enhanced thermodynamic parameters. It was stable for 36 days at 5 °C, and retained >90% activity up to 4 cycles. The tri-enzyme co-immobilized MNPs were successfully utilized for extraction of piperine from black pepper, preparation of sugarcane cell protoplasts, and clarification of papaya juice with prominent yield and reusability vis-à-vis traditional methods., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The present article describes extraction of chitosan from prawn shells waste and its application in thermal stabilization of Fusarium sp. ICT SAC1 cutinase by non-covalent and covalent conjugation. Extracted chitosan represented 78.40% degree of deacetylation (DDA), a molecular weight of 173 kDa and was soluble in 1% acetic acid with 2.8 ± 0.15% insoluble matter. The structural (FTIR, NMR and XRD) and thermal characterization (DSC and TGA) indicated unique properties for chitosan. Plausible chitosan structure was also deduced. The water and fat binding capacities were 923% and 598.05% while 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) and 1,1-diphenyl-2-picrylhydrzyl radicals scavenging activity was 60.62 and 11.83 μM Trolox-Equivalent/ml. The K m and V max values of free cutinase were 0.82 mM and 20.64 mM/min which increased by 14.63 and 17.07%; and 27.18 and 43.94% after non-covalent and covalent conjugation, respectively. A marginal increment in thermal inactivation constants and energy (k d , t 1/2 , D and E d value) were also noticed for cutinase-chitosan conjugates. The enthalpy, free energy and entropy values increased marginally in covalent conjugate vis-à-vis non-covalent conjugated and free cutinase. A reduction in α-helix, random coils and β-sheets content was noted after conjugation., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Cutinase, a member of α/β-fold hydrolase family possess potentially diverse applications in several industrial processes and products. The present work aims towards thermo-stabilization of cutinase from novel source Fusarium sp. ICT SAC1 via non-covalent interaction with polysaccharides. Although all six polysaccharides chosen for study enhanced the thermal stability, pectin was found to be most promising. The interaction protocol for cutinase with pectin was optimized sequentially with respect to the ratio of enzyme to pectin, solution pH, and buffer strength. Cutinase-pectin conjugate under optimized conditions (1:12, pH-6.5, 50mM) showed enhanced thermal stability as evident from lower inactivation rate constant, higher half-life and D-value within the 40-55°C. A slender rise in K m and V max values and enhanced thermodynamic parameters of cutinase-pectin conjugate were observed after non-covalent interaction. Entropy values were 1.5-fold higher for cutinase-pectin conjugate at each temperature suggesting an upsurge in number of protein molecules in a transition activated state. Positive values of entropy for both forms of cutinase suggested a rise in disordered conformation. Noticeable conformational changes in cutinase after conjugation with pectin were confirmed by FTIR as well as fluorescence emission spectra. An increment in helix to turn conversion was observed in complexed cutinase vis-à-vis free cutinase., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Macromolecular cross-linked enzyme aggregates (M-CLEAs) of α-amylase were prepared by precipitation and subsequent cross-linking. The non-toxic, biodegradable, biocompatible, renewable polysaccharide based macromolecular cross-linkers viz. agar, chitosan, dextran, and gum arabic were used as a substitute for traditional glutaraldehyde to augment activity recovery toward macromolecular substrate. Macromolecular cross-linkers were prepared by periodate mediated controlled oxidation of polysaccharides. The effects of precipitating agent, concentration and different cross-linkers on activity recovery of α-amylase CLEAs were investigated. α-Amylase aggregated with ammonium sulphate and cross-linked by dextran showed 91% activity recovery, whereas glutaraldehyde CLEAs (G-CLEAs) exhibited 42% activity recovery. M-CLEAs exhibited higher thermal stability in correlation with α-amylase and G-CLEAs. Moreover, dextran and chitosan M-CLEAs showed same affinity for starch hydrolysis as of free α-amylase. The changes in secondary structures revealed the enhancements in structural and conformational rigidity attributed by cross-linkers. Finally, after five consecutive cycles dextran M-CLEAs retained 1.25 times higher initial activity than G-CLEAs., (Copyright © 2015 Elsevier B.V. All rights reserved.)