临床药理学与生物药剂学

抽象的

Direct Measurement of Lipase Inhibition by Orlistat Using a Dissolution Linked In Vitro Assay

Daniel R Lewis and Dongzhou J Liu

Purpose: To develop a bio-assay that would be able to directly test gastrointestinal and/or dissolution samples to determine lipase activity and inhibition by Orlistat.
Methods: Enzyme assays were performed with porcine pancreatic lipase and para-Nitrophenyl Palmitate (pNPP) in pH 8.0 reaction buffer at 37°C. Substrate hydrolysis was monitored by absorbance changes at 410 nm. The dissolution of two Orlistat formulations was tested with a USP II apparatus. Samples were HPLC analyzed to determine release profile in addition to being diluted and directly assayed for inhibitory effect.
Results: The lipase-pNPP system demonstrates linearity and Michalis-Menten kinetics with a Km=2.7 ± 0.2 μM and Kcat = 0.019 s-1. Orlistat showed highly potent and time dependent inhibition with 5 ng/ml effecting 50% activity after 5 minutes in the Lipase-pNPP system. Dissolution studies showed a correlation of the drug release profile to the inhibitory effect of dissolution samples in the assay.
Conclusions: The lipase-pNPP method can be used as an in vitro assay to monitor orlistat inhibition from drug release or dissolution samples.