Improved shelf life estimation of UHT milk by prediction of proteolysis

During growth in raw milk, many psychrotrophic bacteria produce proteases that can retain activity following ultra-high temperature (UHT) treatment. In this study, casein and skim milk powder assays for detecting very low levels of protease in UHT milk were optimized, and the suitability of azocasein and fluorescein isothiocyanate-casein (FITC-casein) as substrates was investigated. The strongest correlations of protease activity with proteolysis in stored UHT milk were observed when FITC-casein was used as substrate in the assays. Assays using casein and FITC-casein as substrates yielded the highest activities. To determine sensitivity, crude protease was added at low concentrations to UHT milk, and the milk was assayed for progress of proteolysis over 12 months and for protease activity using the casein and FITC-casein assays. With long assay incubation times, the FITC-casein assay was more sensitive than the casein assay and may be suitable for detecting very low levels of protease activity and predicting progress of proteolysis in stored UHT whole milk.