Alkalophilic Bacillus subtilis ASH produced high levels of xylanase using easily available inexpensive agricultural waste residues such as wheat bran, wheat straw, rice husk, sawdust, gram bran, groundnut and maize bran in solid-state fermentation (SSF). Among these, wheat bran was found to be best substrate. Xylanase production was highest after 72 h of incubation at 37°C and at a substrate to moisture ratio of 1:2 (w/v). The inoculum level of 15% resulted in maximum production of xylanase. The enzyme production was stimulated by the addition of nutrients such as yeast extract, peptone and beef extract. In contrast, addition of glucose and xylose repressed the production of xylanase. The extent of repression by glucose (10%, w/v) was 81% and it was concentration-dependent. Supplementation of the medium with 4% xylose caused 59% repression. Under optimized conditions, xylanase production in SSF (8,964 U of xylanase/g dry wheat bran) was about twofold greater than in submerged fermentation. Thus, B. subtilis produced a very high level of xylanase in SSF using inexpensive agro-residues, a level which is much higher than that reported by any other bacterial isolate.Furthermore, the enzyme was produced at room temperature and with tap water without the addition of any mineral salt in SSF, leading to a marked decrease in the cost of xylanase production, which enhances its industrial potential.

The present study describes the one-step purification and characterization of an extracellular cellulase-free xylanase from a newly isolated alkalophilic and moderately thermophilic strain of Bacillus subtilis ASH.Xylanase was purified to homogeneity by 10.5-fold with ~43% recovery using ion-exchange chromatography through CM-Sephadex C-50. The purified enzyme revealed a single band on SDS-PAGE gel with a molecular mass of 23 kDa. It showed an optimum pH at 7.0 and was stable over the pH range 6.0-9.0. The optimum temperature for enzyme activity was 55 °C. The purified xylanase did not lose any activity up to 45 ºC, however, it retained 80% and 51% of its activity after pre-incubation at 55 ºC and 60 ºC, respectively. The enzyme obeyed Michaelis-Menton kinetics towards birch wood xylan with apparent K m 3.33 mg/ml and V max 100 IU/ml. The enzyme was strongly inhibited by Hg 2+ and Cu 2+ while enhanced by Co 2+ and Mn 2+. The purified enzyme could be stored at 4 ºC for six weeks without any loss of catalytic activity. The faster and economical purification of the cellulase-free xylanase from B. subtilis ASH by onestep procedure together with its appreciable stability at high temperature and alkaline pH makes it potentially effective for industrial applications.

Aim: Yak milk is a type of milk that people are less familiar with due to its remote geographical location which may have significant effects on composition, microbiota and hydrolytic outcome. Present work was designed with the aim to evaluate the antioxidative effect of peptides derived from yak milk caseinate on hydrolysis with three different proteases. Materials and Methods:In this investigation Yak milk casein was hydrolyzed by three commercially available proteases (Trypsin, Pepsin and chymotrypsin). These hydrolysates collected at different hydrolysis times (30 min, 60 min, 90 min, 120 min, 150 min, 180 min, 210 min, 240 min, 270 min, 300 min, 330 min and 360 min) were assayed for their antioxidant activity with respect to the effect of incubation period.Results: Among all the enzyme hydrolysates, the tryptic hydrolysates showed highest antioxidant activity followed by chymotryptic hydrolysates. Further, the peptide samples showing highest activity were subjected to RP-HPLC for their partial characterization. Tryptic and peptic hydrolysates produced peaks mainly in the region of hydrophillic solvent indicating the presence of hydrophillic peptides/peptides. Conclusion:The results indicated that yak milk casein could be a resource to generate antioxidative peptides and be used as multifunctional active ingredients for many value-added functional foods as well as a traditional food protein.

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This paper reports high level production of a cellulase-free xylanase using wheat bran, a cost-effective substrate, under submerged fermentation by alkalophilic Bacillus subtilis ASH. Production of xylanase was observed even at alkaline pH up to 11.0 and temperature 60 °C, although the highest enzyme titer was recorded at neutral pH and 37 °C. The enzyme production under optimized fermentation was 1.5-fold greater than under unoptimized conditions. Pre-treatment of unbleached pulp of 10% consistency with crude xylanase (6 IU/g o.d. pulp) at 60 ºC for 2 h increased the final brightness by 4.9%. The enzyme treatment reduced the chlorine consumption by 28.6% with the same brightness as in the control. A reduction in kappa number and increase in viscosity was observed after enzyme pre-treatment. Scanning electron microscopy revealed loosening and swelling of pulp fibers. The strength properties viz. grammage, fiber thickness, beating degree, tensile index, breaking length, tear index and double fold of the treated pulp were improved as compared to the control pulp. This study reveals the potentialof B. subtilis ASH xylanase as a biobleaching agent for the paper and pulp industry.

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A reliable and reproducible reversed-phase high performance liquid chromatography (RP-HPLC) was developed for the quantitative determination of Remipril drug content from marketed bulk tablets. The active ingredient of Remipril separation achieved with C18 column using the methanol water mobile phase in the ratio of 40:60 (v/v). The active ingredient of the drug content quantify with UV detector at 215 nm. The retention time of Remipril is 5.63 min. A good linearity relation (R2=0.999) was obtained between drug concentration and average peak areas. The limit of detection and limit of quantification of the instrument were calculated 0.03 and 0.09 µg/mL, respectively. The accuracy of the method validation was determined 102.72% by recoveries method.