title: Screening of inhibitors of angiotensin-converting enzyme (ACE) employing high performance liquid chromatography-electrospray ionization triple quadrupole mass spectrometry (HPLC-ESI-QqQ-MS).
abstract: Angiotensin-converting enzyme (ACE) plays a key role in regulating blood pressure in the body by converting the angiotensin I (AI) into angiotensin II (AII). Angiotensin II is a potent vaso-active peptide that causes arterioles to constrict, resulting in increased blood pressure. A rapid and sensitive method for the identification of inhibitors of ACE was developed, and optimized employing HPLC-ESI-QqQ-MS. In this assay, angiotensin I substrate was converted into the product angiotensin II with the catalytic action of ACE. A calibration curve for depleting concentration of angiotensin I was developed and linearity of R2=0.999 with a remarkably low concentration of substrate range 20-200nM. The limit of detection and quantification of angiotensin I was found to be 1.93 and 5.84nM, respectively. The enzymatic reaction was optimized for incubation time, concentration, and volume of enzyme and substrate. All reactions were performed at 37°C at pH7.5 with standard incubation time of 20min. Two standard inhibitors, Captopril and Lisinopril, were checked through the newly developed method for their inhibitory potential, and their IC50values were found to be 3.969 and 0.852μM, respectively. Reproducibility and precision analysis of different experiments showed <9.9% RSD. The developed method can be used for the identification of new ACE inhibitors.
authors.affiliation: H.E.J. Research Institute of Chemistry, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Pakistan; Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Pakistan. Electronic address: musharraf1977@yahoo.com.
authors.forename: Syed Ghulam
authors.initials: SG
authors.lastname: Musharraf
authors.affiliation: H.E.J. Research Institute of Chemistry, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Pakistan.
authors.forename: Muhammad Salman
authors.initials: MS
authors.lastname: Bhatti
authors.affiliation: H.E.J. Research Institute of Chemistry, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Pakistan; Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Pakistan; Department of Chemistry, College of Science, King Saud University, Riyadh 1145, Saudi Arabia.
authors.forename: Muhammad Iqbal
authors.initials: MI
authors.lastname: Choudhary
authors.affiliation: H.E.J. Research Institute of Chemistry, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Pakistan; Dr. Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Pakistan.
authors.forename: Atta-Ur
authors.initials: AU
authors.lastname: Rahman
mesh_terms: D000803:Angiotensin I / Q000378:metabolism; D000804:Angiotensin II / Q000378:metabolism; D000806:Angiotensin-Converting Enzyme Inhibitors / Q000737:chemistry* / Q000494:pharmacology; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D017706:Lisinopril / Q000737:chemistry / Q000494:pharmacology; D010455:Peptides / Q000378:metabolism; D007703:Peptidyl-Dipeptidase A / Q000378:metabolism*; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods
chemical_list: D000806:Angiotensin-Converting Enzyme Inhibitors; D010455:Peptides; D000804:Angiotensin II; D000803:Angiotensin I; D017706:Lisinopril; D007703:Peptidyl-Dipeptidase A
keywords: Angiotensin I; Angiotensin-converting enzyme; Captopril; Inhibition; LC-QqQ-MS; Lisinopril; Multiple reaction monitoring (MRM)