Quantitative determination of lipase activity by liquid chromatography-mass spectrometry.
We developed a novel in vitro lipase assay based on the quantitation of fatty acids by liquid chromatography-mass spectrometry. Oleic acids enzymatically released from triolein substrates were isolated from the reaction mixture by reverse-phase chromatography, ionized in negative mode electrospray mass spectrometry and quantitated with the aid of [(13)C]-oleic acid internal standard. The enzymatic activity was measured by monitoring oleic acid productions at multiple time points. This method overcomes the substrate and pH limitations of conventional techniques and thus serves as a generic lipase activity assay.
Hao
Gang
G
Altus Pharmaceuticals, Cambridge, Massachusetts 02139, USA. ghao@altus.com
Yang
Lan
L
Mazsaroff
Istvan
I
Lin
Melanie
M
D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004789:Enzyme Activation; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D008049:Lipase / Q000737:chemistry*; D019301:Oleic Acid / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013379:Substrate Specificity
D019301:Oleic Acid; D008049:Lipase