Utilization of Renewable Agricultural Resources for Ectoine, Xylanase and Cellulase Production from Newly Isolated Halophilic Bacteria

Abstract

Industrial production of high value biomolecules is frequently limited by substrate costs. The present investigation deals with an economically alternative strategy of utilizing renewable agricultural resources as substrates for halophilic bacteria. Several biopolymer degrading halophilic bacteria were isolated from saline regions of coastal Karnataka, India. Halomonas sp. PS6 (GenBank ID: KC295600) produced ectoine (mg/gdw) - on glucose (71.3), xylan (45.2), CMC (22.6), inulin (11.8), chitin (8.1) and carob (22.7). This shows the ability of the halophilic bacterium to utilize bio-polymers in the synthetic medium for growth and ectoine production. Strain PS6 produced substantial amounts of ectoine on agro-residues (mg/gdw) - rice bran (60.2), wheat bran (36.2), sugarcane bagasse (34.7), corn cobs (25.5), groundnut shells (18.3) and coir pith (14.1). 85.4 mg/gdw ectoine was produced at 12% w/v NaCl and 70.5 mg/gdw ectoine at 37 ºC. Using response surface methodology, twofold increase in ectoine was seen (112.3 mg/gdw) at rice bran 50 g/l, NaCl 110 g/l and temperature 37 ºC with validity of 98%. Brachybacterium sp. PS3 (GenBank ID: JQ425852) produced xylanase (1.37 U/ml) in MM63 medium. Higher activity (4.2 U/ml) was seen with xylan, yeast extract and peptone. Strain PS3 produced xylanase (U/ml) - on wheat bran (0.9), corn cobs (0.75), rice bran (0.7 U/ml), sugarcane bagasse (0.44), groundnut shells (0.48) and coir pith (0.4). Using Plackett-Burman design and response surface methodology, a maximum xylanase activity of 8.23 U/ml at pH 9.0, wheat bran 40 g/l, NaCl 90 g/l and corn cobs 30 g/l was obtained with validity of 95.2% and over four-fold increase. PS3 xylanase exhibited highest activity at pH 9.0 and 55 °C and stability up to 4 M NaCl. Halomonas sp. PS47 (GenBank ID: JQ425853) produced cellulase (0.0076 U/ml) on basal MM63 medium. Higher activity (0.14 U/ml) was seen with CMC and combination of yeast extract and peptone. Strain PS47 produced higher cellulase activities (U/ml) - on wheat bran (0.079), corn cobs (0.06), rice bran (0.059 U/ml), sugarcane bagasse (0.04), and groundnut shells (0.049). By statistical optimisation, cellulase activity of 0.35 U/ml at wheat bran 50 g/l, yeast extract 3 g/l and MgSO4.7H2O 0.4 g/l was observed with validity of 95.8% and 3 three-fold increase. PS47 cellulase exhibited highest activity at pH 7.5 and 50 °C and stability up to 4 M NaCl. The present investigation assumes significance in the ability of halophilic bacteria to survive in a wide range of salinity and yield optimum levels of high value biomolecules like compatible solutes and saline hydrolases using cheap agricultural (lignocellulosic) resources.