Enzymatic production of lactulose and epilactose in milk.

The enzymatic production of lactulose was described recently through conversion of lactose by a thermophilic cellobiose 2-epimerase from Caldicellulosiruptor saccharolyticus (CsCE). In the current study, we examined the application of CsCE for lactulose and epilactose production in milk (1.5% fat). The bioconversions were carried out in stirred reaction vessels at 2 different temperatures (50 and 8°C) at a scale of 25 mL volume. At 50°C, 2 highly different CsCE amounts were investigated for the time course of formation of lactulose and epilactose. The conversion of milk lactose (initial lactose content of 48.5 ± 2.1 g/L) resulted in a final yield of 57.7% (28.0 g/L) lactulose and 15.5% (7.49 g/L) epilactose in the case of the approximately 9.5-fold higher CsCE amount (39.5 µkat epilactose, 50°C) after 24 h. Another enzymatic lactose conversion was carried out at low 8°C, an industrially relevant temperature for milk processing. Although the CsCE originated from a thermophilic microorganism, it was still applicable at 8°C. This enzymatic lactose conversion resulted in 56.7% (27.5 g/L) lactulose and 13.6% (6.57 g/L) epilactose from initial milk lactose after 72 h. The time courses of lactose conversion by CsCE suggested that first epilactose formed and afterward lactulose via epilactose. To the best of our knowledge, this is the first time that an enzyme has produced lactulose directly in milk in situ at industrially relevant temperatures.

Biodiversity of refrigerated raw milk microbiota and their enzymatic spoilage potential.

The refrigerated storage of raw milk selects for psychrotolerant microorganisms, many of which produce peptidases and lipases. Some of these enzymes are heat resistant and are not sufficiently inactivated by pasteurisation or even ultra-high temperature (UHT) treatment. In the current study, 20 different raw cow's milk samples from single farms and dairy bulk tanks were analysed close to delivery to the dairies or close to processing in the dairy for their cultivable microbiota as well as the lipolytic and proteolytic potential of the isolated microorganisms. Altogether, 2906 isolates have been identified and assigned to 169 species and 61 genera. Pseudomonas, Lactococcus and Acinetobacter were the most abundant genera making up 62% of all isolates, whereas 46 genera had an abundance of <1% and represent only 6.6%. Of all isolates, 18% belong to hitherto unknown species, indicating that a large fraction of the milk microbiota is still unexplored. The potential of the isolates to produce lipases or peptidases followed in many cases a genus or group specific pattern. All isolates identified as members of the genus Pseudomonas exhibited mainly lipolytic and proteolytic activity or solely proteolytic activity. On the other hand, nearly all isolates of the genus Acinetobacter were lipolytic but not proteolytic. Only 37% of all tested lactic acid bacteria (LAB) showed enzymatic activity at 6 °C and the type of activity was proteolytic in 97% of these cases.