Optimization of concurrent production of xylanolytic and pectinolytic enzymes by Bacillus safensis M35 and Bacillus altitudinis J208 using agro-industrial biomass through Response Surface Methodology.

Application of crude xylanolytic and pectinolytic enzymes in diverse industrial processes make these enzymes commercially valuable and demand their production process to be cost-effective. Out of four different agrowaste biomass, wheat bran (WB) and citrus peel (CP), when amended as fermentation substrates, respectively induced the highest xylanolytic enzymes and pectinolytic enzymes from both, B. safensis M35 and B. altitudinis J208. Further, the simultaneous amendment of WB and CP yielded concurrent production of these cellulase free xylanolytic and pectinolytic enzymes. Hence, the quadratic model was developed using the Central Composite Design of Response Surface Method (CCD-RSM). The model gave the concentration values for WB and CP substrates to be amended in one single production medium for obtaining two optimized predicted response values of xylanase activity and pectinase activity units, which were further practically validated for the xylanase and pectinase production responses from the optimized production medium (OPM). These practically obtained response values from OPM were found to be in accordance with a range of 95% predicted intervals (PI) values. These observations verified the validity of the predicted quadratic model from RSM and suggested that both xylanase and pectinase enzymes can be induced concurrently from both of the bacterial strains.

An Arthrobacter citreus strain suitable for degrading ε-caprolactam in polyamide waste and accumulation of glutamic acid.

ε-Caprolactam-a toxic xenobiotic compound present in industrial polyamide waste was found to be degraded by caprolactam-degrading bacteria. Arthrobacter citreus was able to utilize up to 20 g ε-caprolactam/l as the sole source of carbon more efficiently as compared to the other Gram positive caprolactam-degrading bacteria Rhodococcus rhodochrous and Bacillus sphaericus. The cells of A. citreus remained viable in medium up to 40 g caprolactam/l. The degradation of 10 g caprolactam/l by A. citreus, when supplied as the sole source of carbon and nitrogen lead to the formation of 6-aminocaproic acid which was detected in broth and there was also an increase in the ammonium content. One of the other metabolites found to consistently accumulate in extracellular medium during the utilization of caprolactam by A. citreus was glutamic acid, though not reported in case of other caprolactam-degrading bacteria. A. citreus could metabolise caprolactam to form non toxic products such as 6-aminocaproic acid and glutamic acid which are amino acids of physiological and commercial importance. In the presence of 6-aminocaproic acid, the rate of caprolactam utilization by A. citreus was decreased but not inhibited and the viable count of cells was found to increase using both the substrates simultaneously. A. citreus was also suitable for degradation of caprolactam in presence of low phosphate as prevalent in soil, and in sterile soil without the supplementation of any other carbon or nitrogen, as well as in native non sterile soil where other microorganisms are present.

Use of Nonspecific, Glutamic Acid-Free, Media and High Glycerol or High Amylase as Inducing Parameters for Screening Bacillus Isolates Having High Yield of Polyglutamic Acid.

Out of fifty-five Bacillus isolates obtained from ten different regional locations and sources, seven showed the ability to consistently produce specific extracellular polymeric substance (EPS) on rich as well as synthetic but nonspecific media which did not contain glutamic acid. The isolates were identified as either Bacillus licheniformis or Bacillus subtilis. The EPS from all isolates was resistant to alpha protease, proteinase K, and was thus of high molecular weight. Further it was detected after SDS-PAGE by methylene blue but not by coomassie blue R staining as in case of proteins with high proportion of acidic amino acids. Cell-free EPS, after acid hydrolysis, showed absence of carbohydrates and presence of only glutamic acid. Thus the native the EPS from all seven isolates was confirmed to be gamma polyglutamic acid (PGA) and not exopolysaccharide. The Bacillus isolate T which produced maximum polymer on all media tested had higher amylase: protease activity as compared to other strains. If inoculum was developed in rich medium as compared to synthetic medium, the PGA produced increased by twofold in the subsequent synthetic production medium. Similarly, use of inoculum consisting of young and vegetative cells also increased the PGA production by twofold though amount of inoculum did not affect yield of PGA. Though PGA was produced in even in the absence of glutamic acid supplementation in the production medium by all isolates, the yield of PGA increased by fourfold in the presence glutamic acid and the maximum yield was 30 g/l for isolate K. The supplementation of glutamine instead of glutamic acid into the medium caused an increase in the viscosity of the non-Newtonian solution of PGA.