Kinetic behaviour of pancreatic lipase inhibition by wine

Abstract

This study determined the in vitro inhibitory effects of Mukuzani (dry red) wine and its ethyl acetate/aqueous fractions against pancreatic lipase (PL). Also, it characterises the kinetic behaviour of such inhibition. To current knowledge, no studies have been conducted to describe the kinetic behaviour of PL inhibition by wine or the combined effect on the PL activity of some individual phenolic compounds present in wine. Additionally, the influence of different winemaking methods on PL activity is unknown. Lipase activity was determined by the titrimetric assay method. The kinetic analysis was determined by using the graphical method via Lineweaver-Burk plots. The mode of inhibition (MOI) was determined by looking at the interception pattern and crossing linear lines for the reciprocal data of PL activity with and without inhibitors versus olive oil concentration. The mode of inhibition identified for the wine sample was mixed-mode inhibition. In this type of inhibition, the value of Km (244.329±10.214 μmol·mL-1) was higher, and Vmax (80875.4±3489.754 μmol·mL-1·hour-1) was lower compared to the value of Km (170.901±7.544 μmol·mL-1) and Vmax (88735±4036.741 μmol·mL-1·hour-1) for the non-inhibited PL. This inhibitor decreased the PL activity rate and reduced the affinity between the substrate (olive oil) and enzyme (PL). Selected wine showed predominantly competitive inhibition. The value of inhibition constant Ki1 was equal to 40.556±1.932, and Ki2 was equal to 179.361±8.678 μmol·mL-1, meaning that binding affinity to the pancreatic lipase was higher than to the enzyme-substrate complex. The liquid/liquid extraction method was used to fractionate phenolic compounds of wine into aqueous and ethyl acetate fractions. Fractionated phenolic compounds of wine un-competitively inhibited PL activity. They were considered mutually nonexclusive inhibitors, while a synergistic relationship was found among them. The kinetic parameters of apparent Vmax and Km decreased. Due to the uncompetitive inhibition, the same factor decreased these kinetic parameters. The results obtained herein allow one to conclude that red wine made with the Kakhetian winemaking method can inhibit pancreatic lipase in vitro and play a role in body weight management.