TY - JOUR AU - NOOR MOHAMAD ZIN, NOOR SYAFFINAZ AU - AZMI, NUR AIN SABRINA AU - ANUAR, NURSARAH SYAMIMI AU - SHAFIE, SYAHIRAH AIN AU - MAZNAN, MUHAMAMMAD AIMAN FARIS AU - GOH, YONG MENG AU - SAMSULRIZAL, NURDIANA PY - 2022/10/31 Y2 - 2024/03/29 TI - A 96-WELL-PLATE–BASED METHOD FOR THE ESTIMATION OF ALPHA-AMYLASE ACTIVITY USING MINIATURISES 3,5-DINITROSALICYLIC ACID (DNSA) COLORIMETRIC METHOD JF - Malaysian Applied Biology JA - MAB VL - 51 IS - 4 SE - Research Articles DO - 10.55230/mabjournal.v51i4.16 UR - https://jms.mabjournal.com/index.php/mab/article/view/2474 SP - 95-102 AB - <p>The DNSA assay has been widely employed for the in vitro detection and quantification of alpha-amylase inhibitory activity. However, the conventional method is associated with inconsistencies between protocols and requires a large volume of samples and other assay reagents that can compromise accurate quantitation. Therefore, the study aimed to develop a reliable, simple, and rapid analytical method for determining α-amylase activity. The developed method was carried out in 96-well microplates with a total volume of 250 µL and a total assay time of 1 hr, including pre-incubation. The method was validated for linearity, the limit of detection (LOD), the limit of quantitation (LOQ), and precision. A higher coefficient of determination (R<sup>2</sup>) value was observed for the developed method as compared to the conventional method (0.9983 ± 0.0003 vs 0.9667 ± 0.0383). The coefficient of variation (CV%) of each data point was less than 15%, indicating excellent data precision. The optimum assay conditions were identified at 2 U/mL of enzyme solution and 5% (w/v) starch solution at 50 °C incubation temperature with an IC<sub>50</sub> value of 0.026 ± 0.005 mg/mL. It is concluded that the developed method is practical, precise, and accurate for estimating α-amylase inhibitory activity and would provide reproducible results.</p> ER -