51B-9

P. RAVIYAN, Department of Food Science & Human Nutrition, Washington State University, Box 646376, Pullman, WA 99164-6376, J. Tang, Department of Biological Systems Engineering, Washington State University, and B. A. Rasco, Department of Food Science and Human Nutrition, Washington State University.

JUSTIFICATION: Lack of rapid methods to validate and/or verify the adequacy of pasteurization processes is a pressing problem. The marker(s) here described have specific and well defined kinetic properties. Residual enzyme activity in the marker rapidly assayed providing a measure of heat exposure. No interference with endogeneous enzymes or other food components on the marker assay system is observed.

OBJECTIVES: Develop and evaluate immobilized a-amylase markers for validating or verifying food pasteurization processes. Study inactivation kinetics of biochemical markers under different processing conditions and compare with the model organism Listeria innocua. Determine marker diffusional and stability properties.

METHODS: a-amylase (1,4-a-D-glucano-hydrolase, EC 3.2.1.1) from Asperigillus oryzae was physically entrapped in 20% polylacrylamide, and cut into 0.5x0.5x0.5 cm pieces, each containing ca. 2 FAU. Enzyme activity was measured spectrophotometrically by reaction with 3,5 dinitrosalicylic acid.

RESULTS: Over 85% activity is retained following immobilization. Enzyme diffusion is slow, with >75% activity retained after 2 hours in 0.02 M phosphate buffer. The gels are easy to handle, firm but not brittle, and can be incorpated into semi-solid foods. The gels will not melt after 10 min in boiling water. The soluble enzyme has a D value of 28.41 min. at 55°C, 8.37 min. at 60°C, 4.45 min. at 65°C, and 2.49 min. at 70°C; and a Z value of 14.62°C. The enzyme entrapped in 20% acrylamide gel has a D value of 79.00 min at 55°C, 20.00 min at 60°C, and 5.62 min. at 65°C and a Z value of 10°C. Thermal inactivation of the enzyme follows first order kinetics. Enzyme recovery after a 7D thermal process for Listeria innocua is within the linear range of the assay, making this a promising tool for monitoring pathogen inactivation.

SIGNIFICANCE: Implantable biochemical marker(s)in foods can be recovered and assayed after pasteurization processes to determine adequacy of thermal processing. These are active, stable, are easy to handle and have predictable thermal inactivation rates. Marker(s) can replace lengthy, costly, microbial testing.