Measurement of opioid peptides with combinations of reversed phase high performance liquid chromatography, radioimmunoassay, radioreceptorassay, and mass spectrometry |
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| Authors: | G H Fridland D M Desiderio |
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| Institution: | 1. Charles B. Stout Neuroscience Mass Spectrometry Laboratory, University of Tennessee, Memphis 800 Madison Avenue, Memphis, Tennessee 38163, USA;2. Department of Neurology, University of Tennessee, Memphis 800 Madison Avenue, Memphis, Tennessee 38163, USA;3. Department of Biochemistry, University of Tennessee, Memphis, 800 Madison Avenue, Memphis, Tennessee 38163, USA;1. Chemistry Department, University of the Balearic Islands, 07122 Palma de Mallorca, Spain;2. Department of Applied Mathematics and Analysis, University of Barcelona, Spain;3. Department of Chemistry, College of Science, King Khalid University, Abha 61321, Saudi Arabia;1. School of Business and Management, American University of Sharjah, P.O. Box 26666, Sharjah, United Arab Emirates;2. University of Wollongong in Dubai, Blocks 5, 14 & 15, Dubai Knowledge Village, P.O. Box 20183, Dubai, United Arab Emirates;1. Department of Head and Neck Surgery, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA 90024, USA;2. Neurology Department, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA 90024, USA |
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| Abstract: | Novel state-of-the-art mass spectrometric methods have been developed and are now used to identify and to quantify enkephalins and other neuropeptides in biological tissue extracts. As the first step, RP-HPLC gradient elution is performed of a Sep-Pak treated peptide-rich fraction from a tissue extract, and the eluent is monitored by a variety of post-HPLC detectors. In an effort to maximize the structural information that can be obtained from the analysis, UV (200 nm) provides the analog absorption trace; receptorassay analysis (RRA) data of all (90) fractions that are collected are used to construct the profile of opioid-receptoractive peptides; radioimmunoassay (RIA) of selected HPLC fractions at retention times corresponding to the retention time of standards, or in some special cases of all 90-fractions, provides immunoreactivity information; and fast atom bombardment mass spectrometry (FAB-MS) in two modes - corroboration of the (M+H)+ of the expected peptide, or MS/MS to monitor an amino acid sequence-determining fragment ion unique to that peptide in the selected ion monitoring (SIM) mode - provides structural information. As a demonstration of the level of quantification sensitivity that can be attained by these novel MS methods, FAB-MS-MS-SIM of solutions of synthetic leucine enkephalin was sensitive to the 70 femtomole level. This paper discusses RIA versus RRA data, and recent MS measurements of peptides in human tissues. |
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