Lipase Activity Inhibition by Miang Extract

Based on the findings of a previous study [25] (link), lipase activity was determined by measuring the free fatty acid released after the enzyme – olive oil reaction. Briefly, 3 mL of olive oil was mixed with 2.5 mL of deionized water or the Miang extract, 1 mL of 100 mM sodium phosphate buffer pH 6.5, and 0.5 mL of Tween 80. The mixture was then vigorously mixed using a magnetic stirrer for 15 min to obtain an emulsion. The porcine pancreatic lipase (PPL) (100 U) was added to the emulsified mixture and incubated on a 150-rpm rotary shaker at 37 °C for 30 min. At the end of the incubation period, 3 mL of 95% ethanol was added prior to the mixture, which was then titrated with 50 mM NaOH using an automatic potentiometric titrator. The end point for the titration was set at pH 9.0. One unit of lipase activity was defined as the amount of enzyme that catalyzed the hydrolysis of triglycerides to release 1 microequivalent of fatty acids in 1 min under standard assay conditions. To determine the percentage of the inhibitory concentration of Miang against PPL, the PPL activity in the reaction was initially set up as 200 U. The PPL activity was assayed in the presence of various concentrations of the Miang extract as the inhibitor and compared to that without the presence of an inhibitor. The IC₅₀ value was determined from the regression curve and expressed as g/100 mL of Miang extract.