Scale-up studies on immobilization of lactoperoxidase using milk whey for producing antimicrobial agent

Abstract

Hypothiocyanite (OSCN-), produced by lactoperoxidase (LPO) in the presence of SCN- and H2O2, inhibits the growth of bacteria. This inhibition is called by LPO system (LPOS). Our laboratory scale study in previous experiment showed that whey immobilized on SP-Sepharose Fast Flow (SP-FF) could produce OSCN- continuously. Then, the purpose of this study is to scale up continuous production of OSCN- using immobilized whey. Immobilized whey was generated by circulating various amounts of whey through SP-FF. To generate OSCN-, 10 ml of the substrate solution containing 0.5 mM SCN- and 0.5 mM H2O2, was circulated through immobilized whey and followed by washing with pure water. The next cycle was done by circulating a fresh 10 ml of substrate solution at the same concentration. The result indicated that a stable immobilization efficiency of more than 90% was achieved in the SP-FF circulated with 300 ml or less of whey per gram of SP-FF. When stored at 4°C, immobilized whey retained 80% LPO activity until 3 weeks storage. The reaction solution discharged from immobilized whey was observed to contain approximately 0.4 mM OSCN-. The experiment using 1.0 g of immobilized whey produced a stable 0.4 mM OSCN- production and antimicrobial activity for at least 6 cycles. The increase in resin volume accompanied by the increase in whey volume resulted the extension of a stable OSCN- production. The experiment using recycled SP-FF did not affect to the stability of OSCN- production and antimicrobial activity. These results may open the way for the large-scale production of OSCN-.