Direct Measurement of Lipase Inhibition by Orlistat Using a Dissolution Linked In Vitro Assay
|Daniel R Lewis1 and Dongzhou J Liu2*|
|1New Products Research and Development, GlaxoSmithKline, Parsippany, NJ, USA|
|2Department of Chemical & Biochemical Engineering, Rutgers University, Piscataway, NJ, USA|
|Corresponding Author :||Dongzhou J Liu
Department of Chemical & Biochemical Engineering
Rutgers University, 1500 Littleton Rd
Parsippany, NJ, USA
E-mail: [email protected]
|Received July 24, 2012; Accepted September 04, 2012; Published September 11, 2012|
|Citation: Lewis DR, Liu DJ (2012) Direct Measurement of Lipase Inhibition by Orlistat Using a Dissolution Linked In Vitro Assay. Clinic Pharmacol Biopharm. 1:103. doi:10.4172/2167-065X.1000103|
|Copyright: © 2012 Lewis DR, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.|
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.