Screening and identification of a Bacillus amyloliquefaciens strain for aqueous enzymatic extraction of medium-chain triglycerides

A strain with a high yield of neutral proteinase and low yield of lipase, resistance to medium chain fatty acid triglycerides is the key to increasing yield and quality of Cinnamomum camphora seed kernel oil (CCSKO) with aqueous enzymatic extraction technology. A bacterial strain, NCU116 isolated from the waste residue produced in CCSKO production through primary screening with plate and secondary screening with shake-flask fermentation. It was found to be suitable for the extraction of CCSKO or other medium-chain triglycerides by using the extraction technology. Its activity of neutral proteinase was 4536.5 U/mL, and only 0.088 U/mL for lipase production. The strain was identified as Bacillus amyloliquefaciens by morphological, physiological, biochemical and 16S rDNA molecular identification. The extracellular enzymes produced by NCU116 included neutral proteinase, pectinase, glucoamylase, cellulase, amylase and lipase. The neutral proteinase had the maximum activity at 50 °C, but was unstable. Its optimum temperature and pH value were approximately 40 °C and 7.0 respectively. Mn²⁺ was an activator of neutral proteinase. The glucoamylase had the maximum activity at 45 °C, and was activated by Ca²⁺, Zn²⁺, Fe³⁺ and Mn²⁺. Its optimum temperatures and pH value were 45–50 °C and 6.0 respectively. The pectinase had the maximum activity at 40 °C, and was activated by Cu²⁺, Fe²⁺, Mn²⁺ and Ca²⁺. Its optimum temperatures and pH value were 35–40 °C and 7.0 respectively. The cellulase had the maximum activity at 35 °C, and was activated by Ca²⁺ and Mn²⁺. Its optimum temperatures and pH value were 30–40 °C and 7.0 respectively.