A new method for determining residual coagulant activity in cheese was deve
loped. Cheese samples were dispersed in 0.1 M trisodium citrate, under whic
h conditions chymosin was stable for at least 5 h. On incubation of such di
spersions with a synthetic heptapeptide substrate (Pro-Thr-Glu-Phe-[NO2-Phe
]-Arg-Leu) at 37 degrees C and pH 3.2, a single peptide ([NO2-Phe]-Arg-Leu)
was produced from the substrate by the action of coagulant in the cheese a
nd quantified by reverse-phase HPLC with detection at 300 nm. This peptide
was produced at a constant rate. The concentration of this peptide was prop
ortional to the amount of active enzyme present and the assay was capable o
f measuring chymosin activities in buffer as low as 3.66 x 10(-4) rennet un
its (RU) ml(-1). The assay was highly repeatable with a coefficient of vari
ation of 4.3%, and was applied to a number of commercial cheeses for which
residual coagulant activities in the range 11.1-20.1 RU kg(-1) were determi
ned. The assay developed in this study is a simple, rapid, sensitive and re
producible method for determining residual coagulant activity in cheese and
other dairy products. (C) 1999 Published by Elsevier Science Ltd. All righ
ts reserved.