title: Direct Measurement of Lipase Inhibition by Orlistat Using a Dissolution LinkedIn VitroAssay.
abstract: OBJECTIVE
  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 anin vitroassay to monitor orlistat inhibition from drug release or dissolution samples.
authors.affiliation: New Products Research and Development, GlaxoSmithKline, Parsippany, NJ, USA.
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Lewis
authors.affiliation: New Products Research and Development, GlaxoSmithKline, Parsippany, NJ, USA.
authors.forename: Dongzhou J
authors.initials: DJ
authors.lastname: Liu
keywords: Bioassay; Dissolution; Inhibition; Lipase; Orlistat