Universität zu Köln

Thomas, Andreas ORCID: 0000-0003-1199-0743, Schaenzer, Wilhelm and Thevis, Mario (2014). Determination of human insulin and its analogues in human blood using liquid chromatography coupled to ion mobility mass spectrometry (LC-IM-MS). Drug Test. Anal., 6 (11-12). S. 1125 - 1133. HOBOKEN: WILEY. ISSN 1942-7611

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Abstract

The qualitative and quantitative determination of insulin from human blood samples is an emerging topic in doping controls as well as in other related disciplines (e.g. forensics). Beside the therapeutic use, insulin represents a prohibited, performance enhancing substance in sports drug testing. In both cases accurate, sensitive, specific, and unambiguous determination of the target peptide is of the utmost importance. The challenges concerning identifying insulins in blood by liquid chromatography coupled to ion mobility mass spectrometry (LC-IM-MS) are detecting the basal concentrations of approximately 0.2ng/mL and covering the hyperinsulinaemic clamps at>3ng/mL simultaneously using up to 200L of plasma or serum. This is achieved by immunoaffinity purification of the insulins with magnetic beads and subsequent separation by micro-scale liquid chromatography coupled to ion mobility / high resolution mass spectrometry. The method includes human insulin as well as the synthetic or animal analogues insulin aspart, glulisine, glargine, detemir, lispro, bovine, and porcine insulin. The method validation shows reliable results considering specificity, limit of detection (0.2ng/mL except for detemir: 0.8ng/mL), limit of quantification (0.5ng/mL for human insulin), precision (CV<20%), linearity (r>0.99), recovery, accuracy (>90%), robustness (plasma/serum), and ion suppression. For quantification of human insulin a labelled internal standard ([[H-2(10)]-Leu(B6,B11,B15,B17)] human Insulin) is introduced. By means of the additional ion mobility separation of the different analogues, the chromatographic run time is shortened to 8min without losing specificity. As proof-of-concept, the procedure was successfully applied to different blood specimens from diabetic patients receiving recombinant synthetic analogues Copyright (c) 2014 John Wiley & Sons, Ltd.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Thomas, Andreas UNSPECIFIED orcid.org/0000-0003-1199-0743 UNSPECIFIED Schaenzer, Wilhelm UNSPECIFIED UNSPECIFIED UNSPECIFIED Thevis, Mario UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-425087
DOI:	10.1002/dta.1710
Journal or Publication Title:	Drug Test. Anal.
Volume:	6
Number:	11-12
Page Range:	S. 1125 - 1133
Date:	2014
Publisher:	WILEY
Place of Publication:	HOBOKEN
ISSN:	1942-7611
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language DOPING CONTROL ANALYSIS; ISOTOPE-DILUTION ASSAY; IMMUNOAFFINITY PURIFICATION; DEGRADATION-PRODUCTS; GROWTH-HORMONE; VITREOUS-HUMOR; HUMAN URINE; PEPTIDE; QUANTIFICATION; IDENTIFICATION Multiple languages Biochemical Research Methods; Chemistry, Analytical; Pharmacology & Pharmacy Multiple languages
URI:	http://kups.ub.uni-koeln.de/id/eprint/42508