Food Microbiology

Biogenic amines (BAs) are a group of low-molecular mass organic bases produced through decarboxylation of free amino acids during microbial fermentation. Due to the toxicological effects of BAs on human health, it is necessary to monitor the levels of BAs in foods. In order to conduct the rapid and sensitive detection of BAs in foods, an enzymatic assay using copper containing monoamine oxidase (AMAO2) and flavin adenine dinucleotide containing putrescine oxidase (APUO) from Arthrobacter aurescens has been developed. To improve the stability and combine the activity of both enzyme, cross-linked enzyme aggregates (CLEAs) technique was applied, AMAO and APUO have been combined as cross linked enzyme aggregates (combi-CLEAs) using glutaraldehyde, as cross linker. Through the optimization of the formation condition, the combi-CLEAs were produced with a yield of 82% at a large scale preparation. Comparing to free enzyme, combi-CLEAs exhibited a broader pH range, pH 6-8 for tyramine, pH 7-10 for putrescine, with higher activity. The stability of combi-CLEAs was also improved by more than 2-fold compared to that of free enzymes. Kinetics analysis indicated that combi-CLEAs had the similar kinetic patterns with free enzyme. Although a reduced catalytic efficiency was detected in combi-CLEAs, the substrate inhibition was mitigated. The combi-CLEAs was further tested to determine the concentration of mixed BAs, the enzymatic assay result showed that the change of kinetics parameters has no effects on the measurement. Thus, enzymatic assay using combi-CLEAs could be utilized for the BA detection in foods.