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Journal of Biomolecular Screening
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A Method for Screening Enzyme Inhibitors Using Size Exclusion Chromatography and ESI-LC-MS/MS

Sonal Mathur

FR 8.5, Pharmaceutical and Medicinal Chemistry, Saarland University, Saarbruecken, Germany

Jung Dae Park

Department of Chemistry, Division of Molecular and Life Science, Pohang University of Science and Technology, Pohang, Korea

Dong H. Kim

Department of Chemistry, Division of Molecular and Life Science, Pohang University of Science and Technology, Pohang, Korea

Rolf W. Hartmann

FR 8.5, Pharmaceutical and Medicinal Chemistry, Saarland University, Saarbruecken, Germany

A pilot study was performed for the development of a method to screen compound libraries using an electrospray mass spectrometer interfaced with liquid chromatography (LC). The mixture of compounds was obtained by combining low-molecular weight inhibitors of carboxypeptidase A (CPA), a representative zinc-containing proteolytic enzyme. After the incubation of the mixture with CPA, the enzyme-bound compounds were separated by size exclusion chromatography (SEC) from unbound compounds. The separation of compounds was affected by LC. Three compounds were identified, which represent the tight binding inhibitors of the library. These compounds were quantitated using an automatic switching valve to avoid the interference of buffer salts with the detection of analytes. The quantitated amounts of the compounds were found to be in good accordance with the Ki values. (Journal of Biomolecular Screening 2005:30-35)

Key Words: library screening • size exclusion chromatography • quantitation • ESI-LC-MS/MS • carboxypeptidase A

Journal of Biomolecular Screening, Vol. 10, No. 1, 30-35 (2005)
DOI: 10.1177/1087057104270270
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