Abstract
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 [13C]-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.
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Dakin, H. D. The hydrolysis of optically inactive esters by means of enzymes: Part I. The action of lipase upon esters of mandelic acid. The resolution of inactive mandelic acid. J. Physiol. 1903, 30, 253–263.
Mu, H.; Porsgaard, T. The metabolism of structured triacylglycerols. Prog. Lipid Res. 2005, 44, 430–448.
Spiegel, S.; Foster, D.; Kolesnick, R. Signal transduction through lipid second messengers. Curr. Opin. Cell. Biol. 1996, 8, 159–167.
Paradis, M.-E.; Badellino, K. O.; Rader, D. J.; Deshaies, Y.; Couture, P.; Archer, W. R.; Bergeron, N.; Lamarche, B. Endothelial lipase is associated with inflammation in humans. J. Lipid Res. 2006, 47, 2808–2813.
Layer, P.; Keller, J.; Lankisch, P. G. Pancreatic enzyme replacement therapy. Curr. Gastroenterol. Rep. 2001, 3, 101–108.
Carroccio, A.; Iacono, G.; Montalto, G. Pancreatic enzyme therapy in childhood celiac disease: A double-blind prospective randomized study. Dig. Dis. Sci. 1995, 40, 2555–2560.
Brady, M. S.; Garson, J. L.; Krug, S. K.; Kaul, A.; Richard, K. A.; Caffrey, H. H.; Fineberg, N.; Balistreri, W. F.; Stevens, J. C. An enteric-coated high-buffered pancrelipase reduces steatorrhea in patients with cystic fibrosis: A prospective, randomized study. J. Am. Diet Assoc. 2006, 106, 1181–1186.
Schmid, R. D.; Verger, R. Lipases: Interfacial enzymes with attractive applications. Angew. Chem. Int. Ed. 1998, 37, 1608–1633.
Beisson, F.; Tiss, A.; Rivière, C.; Verger, R. Methods for lipase detection and assay: A critical review. Eur. J. Lipid Sci. Technol. 2000, 102, 133–153.
Schmidt, M.; Bornscheuer, U. T. High-throughput assays for lipases and esterases. Biomol. Eng. 2005, 22, 51–66.
Frazer, A. C.; Sammons, H. G. The formation of mono- and diglycerides during the hydrolysis of triglyceride by pancreatic lipase. Biochem. J. 1945, 39, 122–128.
Hills, M. J.; Mukherjee, K. D. Assay for triacylglycerol lipase by a rapid thin-layer chromatographic technique. J. Lipid Res. 1988, 29, 1397–1399.
Briquet-Laugier, V.; Ben-Zeev, O.; Doolittle, M. H. Determining lipoprotein lipase and hepatic lipase activity using radiolabeled substrates. Methods Mol. Biol. 1999, 109, 81–94.
Duque, M.; Graupner, M.; Stütz, H.; Wicher, I.; Zechner, R.; Paltauf, F.; Hermetter, A. New fluorogenic triacylglycerol analogs as substrates for the determination and chiral discrimination of lipase activities. J. Lipid Res. 1996, 37, 868–876.
Hendrickson, H. S. Fluorescence-based assays of lipases, phospholipases, and other lipolytic enzymes—a review. Anal. Biochem. 1994, 219, 1–8.
Brockman, H. L. Triglyceride lipase from porcine pancreas. Methods Enzymol. 1981, 71, 619–627.
Isaac, G.; Jeannotte, R.; Esch, S. W.; Welti, R. New mass-spectrometry-based strategies for lipids. Genet. Eng. (NY) 2007, 28, 129–157.
Byss, M.; Triska, J.; Elhottova, D. GC-MS-MS analysis of bacterial fatty acids in heavily creosote-contaminated soil samples. Anal. Bioanal. Chem. 2007 in press.
Lee, S. H.; Pettinella, C.; Blair, I. A. LC/ESI/MS analysis of saturated and unsaturated fatty acids in rat intestinal epithelial cells. Curr. Drug Metab. 2006, 7, 929–937.
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Published online June 13, 2007
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Hao, G., Yang, L., Mazsaroff, I. et al. Quantitative determination of lipase activity by liquid chromatography-mass spectrometry. J Am Soc Mass Spectrom 18, 1579–1581 (2007). https://doi.org/10.1016/j.jasms.2007.05.019
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DOI: https://doi.org/10.1016/j.jasms.2007.05.019