A SEMIAUTOMATED REFLECTANCE COLORIMETRIC METHOD FOR THE DETERMINATIONOF LIPASE ACTIVITY IN MILK

Measurement of heat-stable lipase activity in dairy products relies on methods that are slow or that cannot be used in turbid solutions, whi ch limits their industrial value. A need exists for a rapid, simple, i nformative assay to detect lipase activity in dairy products. In this study, we observed that hydrolysis of p-nitrophenyl esters, monitored by reflectance colorimetry, was linearly correlated to spectrophotomet ry (R(2) = 0.93) and release of titratable FFA (R(2) = 0.92 to 0.97), indicating that chromogenic substrates were useful in determining lipa se activity. However, at the concentrations reported in milk, FFA inhi bited p-nitrophenyl caprylate hydrolysis, which led to an underestimat ion of lipase activity in milk that had previously undergone lipolysis . Milk fat also significantly reduced hydrolysis of the chromogenic su bstrates tested but could be accounted for by a correction equation. T o demonstrate the use of the assay, lipase activity in UHT skim milk i noculated with Pseudomonas fluorescens AFT36 was followed using reflec tance colorimetry and tributyrin agar. Lipase activity increased as ce ll numbers increased during 106 h of incubation. Extracellular lipase activity was detected after 10 h of incubation using reflectance color imetry, but 28 h were required using tributyrin agar. Reflectance colo rimetry and chromogenic substrates allowed a rapid, sensitive, and mea ningful detection of esterase and lipase activity in milk.