Kinetic colorimetric assay of lipase in serum.
We describe a kinetic colorimetric method for assaying lipase (EC 3.1.1.3) activity in serum by using a natural long-chain fatty acid 1,2-diglyceride. In the presence of colipase, deoxycholate, and calcium ions, pancreatic lipase hydrolyzes the clear substrate solution to produce a 2-monoglyceride, which in turn releases glycerol by the action of a 2-monoglyceride lipase. Glycerol is then assayed by a sequence of enzymatic actions (glycerol kinase, glycerol phosphate oxidase, and peroxidase) that produce a violet quinone monoimine dye with peak absorption at 550 nm. The method features zero-order reaction kinetics, provides a simple and rapid assay with an extended dynamic range, is specific and precise, gives results that correlate well (r greater than or equal to 0.99) with those of methods in which emulsified triolein is the substrate, and lends itself readily to automation. For all these reasons, the method seems highly suitable for routine use in clinical laboratories.
Fossati
P
P
Research and Development Laboratory, Bayer Diagnostici SpA, Milan, Italy.
Ponti
M
M
Paris
P
P
Berti
G
G
Tarenghi
G
G
D002118:Calcium / Q000494:pharmacology; D002863:Chromogenic Compounds; D003089:Colipases; D003124:Colorimetry / Q000379:methods* / Q000706:statistics & numerical data; D003840:Deoxycholic Acid / Q000494:pharmacology; D004075:Diglycerides / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D007700:Kinetics; D008049:Lipase / Q000097:blood*; D008823:Micelles; D011786:Quality Control; D013053:Spectrophotometry
D002863:Chromogenic Compounds; D003089:Colipases; D004075:Diglycerides; D008823:Micelles; D003840:Deoxycholic Acid; D008049:Lipase; D002118:Calcium