N glutaryl l phenylalanine p nitroanilide

Total protease activity was determined by using azocasein as a substrate [19] . The gut extract was mixed with the substrate and incubated for 50 min. Then, 200 μl of 5 % TCA was added and centrifuged. To the supernatant, equal volumes of 1 N NaOH were added and absorbance was read at 450 nm. Units for total protease activity (UA) were calculated by using the equation unit activity (UA)=ABS 450 nm/[time (min)×volume of enzyme (ml)]. Trypsin activity was measured by incubating the gut extract with Nα-benzoyl-DL-arginine pnitroanilide (Sigma-Aldrich), and chymotrypsin activity was measured by incubating the gut extract with N-glutaryl-L-phenylalanine p-nitroanilide (Sigma-Aldrich) [20] . Elastase activity of H. armigera gut proteases was measured by incubating the gut extract with N-succinylalanine-alanine-alanine p-nitroanilide (Sigma-Aldrich) [21] . The gut extract was mixed with respective substrates and incubated for 20 min at 37 °C. Then, 300 μl of 30 % acetic acid was added and stand for 10 min. The samples were centrifuged, and absorbance was read at 410 nm using UV-visible spectrophotometer (Hitachi U-2900, Japan). One unit of enzyme activity was defined as the amount of enzyme catalyzing the hydrolysis of 1 μmol substrate per minute at 30 °C.