Date

2014

Author

Ören, Gözde

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The growing interest in enzyme production by the usage of microorganisms continues for easier process control and sustainably higher amount of enzyme production. Food and agricultural industry, which produces large amounts of waste materials, is an excellent resource for enzyme manufacture. Among all, xylanase is considered to have 25 % shares in conjunction with pectinolytic enzymes within food enzyme production and extensively used in extraction/clarification of juices and wines, improvement of bread-quality, and improvement of feed/food digestibility etc. Thus, in this study, Bacillus subtilis and Kluyveromyces marxianus were searched for their potential in xylanase production using mixed culture submerged fermentation method. Effect of inoculum amounts (2-4 ml/100 ml), time (12- 96 h), substrate concentration (10-40 %) and type (arabinose, xylose, sucrose and glucose, synthetic pure xylan and hazelnut shell) as carbon sources, and pH on xylanase production was evaluated at constant temperature of 35oC and agitation of 130 rpm. Xylanase activities of 11.23±0.02 IU/ml and 12.11±0.05 IU/ml were obtained using 1.0 ml of K. marxianus and 1.0 ml of B. subtilis cultures, respectively. Enhanced xylanase activity of 17.93±0.04 IU/ml was produced after 24 h of fermentation with mixed cultures of Bacillus subtilis (0.5 ml) and Kluyveromyces marxianus (0.5 ml). The higher xylanase production of 19.65±0.4 IU/mL was achieved with total inoculum amount of 2.0 ml of Bacillus subtilis (1.0 ml) and Kluyveromyces marxianus (1.0 ml) after 24 h fermentation. Screening experiments revealed that the highest enzyme activities were obtained using arabinose (26.77± 0.5 IU) and xylose (14.92 ± 0.6 IU/ml) as carbon sources after 24 h fermentation at 35oC, pH 6.5 and 130 rpm. Hydrolyzed hazelnut shell (11.38±0.01 IU/ml) was found the second best raw material after the xylose (14.58±0.06 IU/ml). Optimum conditions suggested by Box-Behnken RSM were found as 4 (ml/100 ml) B. subtilis, 4 ml (ml/100ml) K. marxianus, and 40 (% w/v) solid load and pH 7 with maximum activity of 49.5 IU/ml. The determination coefficient (R2) of the model (0.95) and the insignificant lack of fit (p=0.099>0.05) verified that the model fitted well to the experimental data. As a result, xylanase production was increased by 152 % compared to initial unoptimized culture conditions. This study also showed that xylanase production using mixed culture of bacteria and yeast is advantageous over single culturing.

Subject Keywords

Xylanases., Bacillus subtilis., Kluyveromyces., Fermented foods.

URI

http://etd.lib.metu.edu.tr/upload/12616784/index.pdf
https://hdl.handle.net/11511/23272

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Graduate School of Natural and Applied Sciences, Thesis