Siamese neem (Azadirachta indica A. Juss
var.
siamensis Valeton
) (
A. indica
) leaf extract, a traditional ayurvedic medicine, has been reported to exhibit antipyretic, antibacterial, antidyslipidemic, and antihyperglycemia effects. This study investigated the mechanism of hypocholesterolemic effect of methanolic extract of Siamese neem flowers in
in vitro
studies and in Caco-2 cells. Pancreatic cholesterol esterase and 3-hydroxy 3-methylglutaryl-CoA (HMG-CoA) reductase activities were assessed. Cholesterol micelle formation was prepared for
in vitro
cholesterol physicochemical property analyses, micelle size and solubility, and transport of cholesterol into the Caco-2 cells. The expression of niemann-pick C1 like 1 (NPC1L1), and its major regulator, peroxisome proliferator-activated receptor δ (PPARδ), were determined by western blot and real time polymerase chain reaction, respectively. A. indica flower extract inhibited pancreatic cholesterol esterase activity and increased cholesterol micelles size. Uptake of cholesterol into Caco-2 cells was inhibited by
A. indica
flower extract in a dose-dependent manner. In addition,
A. indica
extract inhibited HMG-CoA reductase activity, resulting in low level of intracellular cholesterol accumulation, together with increased cytosolic NPC1L1 protein expression and decreased PPARδ gene expression. In conclusion,
A. indica
flower extract has cholesterol-lowering effects by inhibiting intestinal cholesterol absorption, interfering micellar cholesterol formation, and attenuating cholesterol synthesis. As such,
A. indica
flower extract has potential for developing into nutraceutical product for prevention of hypocholesterolemia.