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Effective enhancement of polylactic acid-degrading enzyme production by Amycolatopsis sp. strain SCM_MK2-4 using statistical and one-factor-at-a-time approaches
Authors:Watsana Penkhrue  Apinun Kanpiengjai  Chartchai Khanongnuch  Kazuo Masaki  Wasu Pathom-Aree  Winita Punyodom
Affiliation:1. Department of Biology, Faculty of Science, Chiang Mai University, Muang District, Chiang Mai, Thailand;2. Division of Biotechnology, School of Agro-Industry, Faculty of Agro-Industry, Chiang Mai University, Muang District, Chiang Mai, Thailand;3. Department of Chemistry, Faculty of Science, Chiang Mai University, Muang District, Chiang Mai, Thailand;4. National Research Institute of Brewing, Higashihiroshima, Hiroshima, Japan;5. Department of Chemistry, Faculty of Science, Chiang Mai University, Muang District, Chiang Mai, Thailand
Abstract:This study aims to find the optimal medium and conditions for polylactic acid (PLA)-degrading enzyme production by Amycolatopsis sp. SCM_MK2-4. Screening of the most effective components in the enzyme production medium by Plackett–Burman design revealed that the silk cocoon and PLA film were the most significant variables enhancing the PLA-degrading enzyme production. After an response surface methodology, a maximum amount of PLA-degrading enzyme activity at 0.74?U?mL?1 was predicted and successfully validated at 95% after 0.39% (w/v) silk cocoon and 1.62% (w/v) PLA film were applied to the basal medium. The optimal initial pH value, temperature, and inoculum size were evaluated by a method considering one-factor-at-a-time. The values were recorded at an initial pH in the range of 7.5–9.0, a temperature of 30–32°C, and an inoculum size of 4–10%. The highest activity of approximately 0.95?U?mL?1 was achieved after 4 days of cultivation using the optimized medium and under optimized conditions in a shake flask. Upscaling to the use of a 3-L stirred tank fermenter was found to be successful with a PLA-degrading activity of 5.53?U?mL?1; which represents a 51-fold increase in the activity compared with that obtained from the nonoptimized medium and conditions in the shake flask.
Keywords:Amycolatopsis  biodegradation  poly(  smallcaps smallerCapital"  >L-lactic acid)  protease  response surface methodology  silk cocoon
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