Pilot-Scale Protease Production by Bacillus sp. C4 for Silk Degumming Processes

ABSTRACT

The serine protease of Bacillus sp. C-4 has been reported as candidate proteases in the degumming process of the silk industry. The optimization of protease production by Bacillus sp. C4 was carried out using two-level fractional factorial and central composite designs. The investigation was followed by laboratory and pilot-scale protease production using molasses in batch and fed-batch bioreactors to reduce the production cost. The highest protease activity observed (1546.5 U/mL, equivalent to 34.4 U/mL/h) in a 5-L bioreactor was obtained under fed-batch fermentation compared with batch process. The production of protease was subsequently scaled up to a pilot-scale 300-L fed-batch bioreactor, and a maximum protease activity of 1260 U/mL (28 U/mL/h) was observed. The degumming efficiencies of the protease produced by Bacillus sp. C4 in the CCD and bioreactors were 88.3% and 81.6%, respectively, relative to the alkaline degumming method. Furthermore, the stability test of the protease showed that the addition of CaCl₂ prolonged the stability of the protease activity from 4 to 8 weeks at room temperature. Furthermore, the protease can be conveniently preserved at room temperature for 1 month and at 4°C for more than 2 months without the addition of any chemicals/cryoprotectants.

摘要

芽孢杆菌C-4丝氨酸蛋白酶已被报道为丝绸工业脱胶过程中的候选蛋白酶. 采用两级分式析因和中心组合设计，对芽孢杆菌C4产蛋白酶条件进行了优化. 为了降低生产成本，采用分批糖蜜和分批补料生物反应器进行了实验室和中试规模的蛋白酶生产. 与分批法相比，补料分批发酵法在5l生物反应器中的蛋白酶活性最高（1546.5u./mL-1，相当于34.4u./mL-1./h-1）. 随后，将蛋白酶的生产规模扩大到中试规模300-L进料间歇式生物反应器，并观察到最大蛋白酶活性为1260 U./mL-1（28 U./mL-1./h-1）. 与碱性脱胶法相比，C4芽孢杆菌产蛋白酶在CCD和生物反应器中的脱胶效率分别为88.3%和81.6%. 此外，蛋白酶的稳定性试验表明，添加CaCl2可使蛋白酶在室温下的稳定性从4周延长到8周. 此外，在不添加任何化学物质/低温保护剂的情况下，蛋白酶可以在室温下保存一个月，在4℃下保存两个月以上.

KEYWORDS:

• Bacillus sp.
• molasses
• optimization
• protease
• silk degumming
• stability

关键词:

• 芽孢杆菌
• 糖蜜
• 优化
• 蛋白酶
• 真丝脱胶
• 稳定性

Acknowledgments

The authors would like to thank Dr. Patoomporn Chim-anek (Department of Microbiology, Kasetsart University) for providing the Bacillus strain used in this study. This research work was financially supported by the Agricultural Research and Development Agency (ARDA), the Ministry of Agriculture and Cooperatives (MOAC), Thailand.

Supplementary material

Supplemental data for this article can be accessed on the publisher’s website

Disclosure statement

There is no conflict of interest in this study.