Effect of psychrotrophic microorganisms on the plasmin system in milk

The psychrotrophic bacteria that are present in raw milk produce heat-stable proteases that affect the plasmin system and, in turn, the quality of the processed milk and other dairy products. Three bacterial strains, Pseudomonas spp. SRM21A and SRM28A and Pseudomonas fluorescens M3/6, were inoculated at a level of approximately 10(3) cfu/ml into reconstituted nonfat dry milk and incubated at 4 degrees C for up to 9 d. From each sample, casein and whey fractions were obtained and electrophoresed to visualize protein hydrolysis and plasmin activity. Colorimetric assays were used to quantify plasmin-related activities. The casein SDS-PAGE gels showed that, by 5 d of incubation, caseinolytic activity was visible in whey fractions of all three strains at the same molecular mass range as commercial bovine plasmin, which was used as a control. Colorimetric assays indicated that plasmin activity in the casein samples decreased as incubation time increased; plasmin activity in the whey samples increased, peaking at 5 to 7 d. Plasminogen activity decreased over time in the casein fraction and increased in the whey fraction. Protein immunodetection indicated crossreactivity between anti-bovine plasminogen and the plasmin and plasminogen from whey samples collected at 5 and 7 d from the incubated milk. The growth of the Pseudomonas strains tested and their concomitant production of proteases caused the release of plasmin and plasminogen from the casein micelle into the whey fraction.