Encapsulation of pectinase within polyacrylamide gel: characterization of its catalytic properties for continuous industrial uses.

Pectinase as a biocatalyst play a significant role in food and textile industries. In this study, the pectinase was immobilized by encapsulation within polyacrylamide gel to enhance its catalytic properties and ensure the reusability for continuous industrial processes. 9.5% acrylamide and 0.5% N, N'- methylenebisacrylamide concentration gave high percentage of pectinase immobilization yield within gel. The catalytic properties of immobilized pectinase was determined with comparison of soluble pectinase. The immobilization of pectinase within polyacrylamide gel didn't effect catalytic properties of pectinase and both the free and immobilized pectinase showed maximum pectinolytic activity at 45 °C and pH 10. The Michaelis-Menten kinetic behavior of pectinase was slightly changed after immobilization and immobilized pectinase showed somewhat higher Km and lower Vmax value as compared to soluble pectinase. Polyacrylamide gel encapsulation enhanced the thermal stability of pectinase and encapsulated pectinase showed higher thermal stability against various temperature ranging from ranging from 30 °C to 50 °C as compared free pectinase. Furthermore, the surface topography of polyacrylamide gel was analyzed using scanning electron microscopy and it was observed that the surface topography of polyacrylamide gel was changed after encapsulation. The encapsulation of pectinase within polyacrylamide gel enhanced the possibility of reutilization of pectinase in various industries and pectinase retained more than 50% of its initial activity even after seven batch of reactions.

Xylan deterioration approach: Purification and catalytic behavior optimization of a novel ß-1,4-d-xylanohydrolase from Geobacillus stearothermophilus KIBGE-IB29.

The ß-1,4-d-xylanohydrolase is an industry valuable catalytic protein and used to synthesize xylooligosaccharides and xylose. In the current study, ß-1,4-d-xylanohydrolase from Geobacillus stearothermophilus KIBGE-IB29 was partially purified up to 9.5-fold with a recovery yield of 52%. It exhibited optimal catalytic activity at pH-7.0 and 50 °C within 5 min. Almost 50% activity retained at pH-4.0 to 9.0 however, 70% activity observed within the range of 40 °C to 70 °C. The ß-1,4-d-xylanohydrolase showed a significant hydrolytic pattern with 48.7 kDa molecular mass. It was found that the enzymatic activity improved up to 160% with 1.0 mM ethanol. Moreover, the activity of enzyme drastically increased up to 2.3 and 1.5 fold when incubated with Tween 80 and Triton X-100 (1.0 mM), respectively. The ß-1,4-d-xylanohydrolase also retained 72% activity at -80 °C after 180 days. Such a remarkable biochemical properties of ß-1,4-d-xylanohydrolase make it possible to forecast its potential use in textile and food industries.

Immobilization of pectin depolymerising polygalacturonase using different polymers.

Polygalacturonase catalyses the hydrolysis of pectin substances and widely has been used in food and textile industries. In current study, different polymers such as calcium alginate beads, polyacrylamide gel and agar-agar matrix were screened for the immobilization of polygalacturonase through entrapment technique. Polyacrylamide gel was found to be most promising one and gave maximum (89%) immobilization yield as compared to agar-agar (80%) and calcium alginate beads (46%). The polymers increased the reaction time of polygalacturonase and polymers entrapped polygalacturonases showed maximum pectinolytic activity after 10 min of reaction as compared to free polygalacturonase which performed maximum activity after 5.0 min of reaction time. The temperature of polygalacturonase for maximum enzymatic activity was increased from 45°C to 50°C and 55°C when it was immobilized within agar-agar and calcium alginate beads, respectively. The optimum pH (pH 10) of polygalacturonase was remained same when it was immobilized within polyacrylamide gel and calcium alginate beads, but changed from pH 10 to pH 9.0 after entrapment within agar-agar. Thermal stability of polygalacturonase was improved after immobilization and immobilized polygalacturonases showed higher tolerance against different temperatures as compared to free enzyme. Polymers entrapped polygalacturonases showed good reusability and retained more than 80% of their initial activity during 2nd cycles.

Purification and characterization of thiol dependent, oxidation-stable serine alkaline protease from thermophilic Bacillus sp.

Alkaline serine protease was purified to homogeneity from culture supernatant of a thermophilic, alkaliphilic Bacillus sp. by 80% ammonium sulphate precipitation followed by CM-cellulose and DEAE-cellulose ion exchange column chromatography. The enzyme was purified up to 16.5-fold with 6900 U/mg activity. The protease exhibited maximum activity towards casein at pH 8.0 and at 80 °C. The enzyme was stable at pH 8.0 and 80 °C temperature up to 2 h. The Ca2+ and Mn2+ enhanced the proteolytic activity up to 44% and 36% as compared to control, respectively. However, Zn2+, K+, Ba2 +, Co2 +, Hg2+ and Cu2+ significantly reduced the enzyme activity. PMSF (phenyl methyl sulphonyl fluoride) completely inhibited the protease activity, whereas the activity of protease was stimulated up to two folds in the presence of 5 mM 2-mercaptoethanol. The enzyme was also stable in surfactant (Tween-80) and other commercial detergents (SDS, Triton X-100).