APPLICATION OF Lacticaseibacillus casei JCM 1134T FOR THE PRODUCTION AND PARTIAL ANALYSIS OF BIOACTIVE PEPTIDES ENCODED IN CARABAO MILK

Abstract

This study was conducted to generate and characterize bioactive peptides encrypted in carabao milk through lactic acid bacteria (LAB) proteolysis. Thirteen LAB strains from the genus Lactococcus, Lactiplantibacillus, Latilactobacillus, Pediococcus, Leuconostoc, Weissella, Lacticaseibacillus, Lactobacillus, and Limosilactobacillus were screened for proteolytic activity using litmus milk test (LMT) and pH measurement. Lacticaseibacillus casei JCM 1134T was selected to ferment (3% v/v) carabao milk at 37°C for 5 days. Milk protein hydrolysate from Lacticaseibacillus casei JCM 1134T-fermented carabao milk was purified and separated using hydrophobic interaction chromatography (HIC), ion exchange chromatography (IEC), ultrafiltration (UF), reversed-phase high performance liquid chromatography (RP-HPLC), and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Fractions were tested for antimicrobial (microbial inhibition), antioxidant (DPPH free radical scavenging), and antihypertensive (ACE inhibition) activities. Some fractions only had antimicrobial activity against Bacillus subtilis JCM 1465^T, antioxidant activity decreased as the purification process progressed, and high antihypertensive activity was observed in hydrolysates and retentates from whey and casein proteins. However, when subjected to electrospray ionization time-of-flight mass spectrometry (ESI-TOF MS), all of the peaks were impure and inactive. These peaks underwent another RP-HPLC run but cannot be detected anymore. Further optimization of purification methods to generate bioactive peptides is highly recommended.