Screening And Optimization Of Lactobacillus Fermenting Goat Milk For Production Of Antioxidant Peptides

The increase of free radicals can lead to the imbalances of oxidation and antioxidation in the body, which causes a series of diseases and threatens people’s health. Several studies have shown that some proteins can be hydrolyzed by lactobacillus and many peptides including antioxidant peptides are produced in the process of fermentation. These antioxidant peptides not only have good oxidation resistance, but also show the advantage of non-toxic. Based on the researches home and abroad, the preparation method of antioxidant peptides mainly focuses on enzyme hydrolysis method, while there are rarely reports about fermented method using lactobacillus. Furthermore, as the king of milk around the world, goat milk has not been fully utilized. This study used goat milk, the superior resources of our province, as the resources to develop new types of functional goat milk, which provided technical support for the diversity of goat milk products.In this study, we investigated the DPPH radical scavenging activity of 25 lactobacillus strains and 4 strains were selected with higher scavenging rate.16S rDNA sequence method was used to identify the 4 strains. The objective strain-Lactobacillus plantarum L60 was confirmed by bile salt and gastric intestinal juice tolerance test, simulated digestion test as well as sensory evaluation, which showed stronger tolerance and stability as well as higher producing ability. The fermentation conditions and media components were optimized using single-factor experiment and response surface methodology. The main results are as follows.1. The all 25 lactobacillus strains showed scavenging activity of DPPH radical range from 21.21% to 69.14%. Compared with others, L60, L49, L61 and LA5 contained higher antioxidant ability. The results of 16S rDNA showed that 4 strains were all belong to lactobacillus. L60 was Lactobacillus plantarum, L49 was Lactobacillus salivarius, and L61 and LA5 were Lactobacillus casei.2. The survival rate of Lactobacillus plantarum L60 was much higher than other 3 strains in bile salt and gastric intestinal juice tolerance test, which indicated that L60 showed more powerful resistance of poor environment. In simulated digestion test, the peptides produced from the fermented goat milk using L60 were more stable. Moreover, the sensory evaluation of L60 fermented milk was preferable. From the above, the Lactobacillus plantarum L60 was selected.3. The effects of inoculum size, goat milk density, fermented time and temperature on DPPH scavenging rate and superoxide scavenging rate were studied using single factor experiment. Keeping goat milk density unchanged, Box-Bnhnken experiment were designed. The optimal fermented condition was 5% inoculum size,14% goat milk, the fermented temperature was 41℃ and fermented time was 12.3h. There were no significant differences between verification and predicted value, DPPH scavenging rate reached 76.61±0.43%, superoxide scavenging rate reached 31.48±0.52%, which were improved 7.76% and 3.17% compared with control group, respectively.4. The effects of several carbon sources, prebiotics, nitrogen sources and mineral salt on antioxidant activity of goat milk fermented by L60 were studied using single factor experiment. According to Plackett-Burman and factorial experiment, peptone, NaCl and glucose showed significant effects on antioxidant activity. The optimized media components were determined based on Box-Behnken test:1.02% NaCl,0.82% peptone and 0.72% glucose. The verification result was consistent with the predicted one. DPPH scavenging rate reached 80.23±0.43%, the viable count of L60 was (1.09±0.0072)×109CFU/mL, which was significantly higher than before.5. The concentration of peptides in samples after optimization from fermented goat milk using L60 was 0.67mg/mL, which was more than before. While the IC50 was 0.9436mg/mL, which decreased 0.2618 mg/mL.In this study, the optimized fermented goat milk using Lactobacillus plantarum L60 had strong antioxidant ability with high DPPH radical scavenging activity, which had certain theoretical signicance and practical value for the development of new functional goat milk products, as well as the expansion of high-value utilization of goat milk sources.