Cross-Linked Enzyme Aggregates (CLEA) is known as one of the best enzyme immobilization technique nowadays. In this study, amylase extract from Zophobas morio (super mealworm) larvae was immobilized using acetone as the precipitant, glutaraldehyde as the cross-linker and bovine serum albumin as the additive. The characteristics of the produced CLEA were compared to the free soluble amylase, in terms of pH and temperature optimum and stabilities. The results displayed that CLEAand free amylase achieved an optimum temperature at 55°C and 45°C, respectively. CLEA-amylase also had showed greater stability against high temperature as compared to a free enzyme which had lost most of its activity when the temperature was set beyond 45°C. In comparison, at 65°C, CLEA-amylase still retained 73.2% of its activity. Results also revealed that CLEA-amylase has a pH optimum at 11, while it is pH 7 for free enzyme. Similarly, CLEA-amylase was more stable than the free form, over a wider range of pH, particularly at higher pH of 9, 10 and 11. Recyclability study showed that CLEA-amylase could retain 14.9% of its residual activity after 6 times of repeated uses. Since it is reusable, future works might include the evaluations of using CLEA-amylase at the industrial level, remarkably in detergent applications.
Many studies have been done on various species of insects to investigate their potential use in industries. This is because insects have high protein content which could be further manipulated. Due to its eating habit, Zophobas morio larvae, also known as super mealworm has been shown to have high amylase activity. In this study, amylase from super mealworm has been immobilized via Cross-Linked Enzyme Aggregates (CLEA) technique and its kinetic performance, evaluated. CLEA is one of the best immobilization method with respect to enzyme stability and reusability. Kinetic performance of both free and CLEA-amylase were evaluated based on the Michaelis-Menten model. Results obtained based on Hanes-Woolf, LineweaverBurk, Eadie-Hofstee and Hyperbolic Regression plots showed that the kinetic parameters, Vmax and KM, changed upon immobilization. For CLEA-amylase, Hanes-Woolf plot showed the bestfitted model based on R2 with Vmax= 1.068 mM/min and KM= 0.182 mM, however, LineweaverBurk plot was used to obtain the kinetic parameters for free amylase, with Vmax and KM of 17.230 mM/min and 2.470 mM, respectively. Thus it is observed that upon immobilization, Vmax for amylase dropped appreciably, however, much lower substrate concentration is needed to saturate the enzymatic sites to reach its maximum catalytic efficiency. The result from this study might open the new path in discovering the potential use of insects in industrial applications, for example, making use of the recovered enzymes in the detergent industry.