Page 112 - Advances in Forensic Applications of Mass Spectrometry - Jehuda Yinon

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2.4.2.2 Insulin
Identification, detection, and quantitation of insulin in blood are very impor-
tant in forensic toxicology. The presence of bovine or porcine insulin in the
blood of a nondiabetic person may serve as evidence of poisoning. Since
insulin and C-peptide are released from proinsulin, the determination of
both products may be of relevance. Very high level of human insulin without
elevation of the C-peptide indicates exogenous administration. When both
insulin and C-peptide are present in high concentrations, the presence of
pancreatic tumor or administration of insulin-releasing drugs, like sulfonyl-
ureas, is possible. The first LC/MS method for insulin was published by
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Stocklin et al. Insulin and its deuterated analog were isolated by immu-
noaffinity chromatography and solid-phase extraction. The LOD of 3 pmol/l
(about 17 ng/l) was reported. The method was used for hemolyzed post-
mortem forensic blood samples, for which immunoassays gave inconsistent
results. A straightforward LC/ESI/MS method for determination of human,
bovine, and porcine insulin, as well as C-peptide, was published by Darby
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et al. The drugs were isolated from acidified plasma with C18 SPE car-
tridges, separated on a 150 ¥ 2.1 mm C18 column and detected with ESI/MS
+3
(ion trap). Multiply charged molecular ions (M + 3H) and (M + 4H) +4
were monitored. Both ions were selected for quantitation since the intensity
ratio was variable between runs. The LOQ was 0.4 mg/l for insulin and 0.1
mg/l for C-peptide; the comparison with radioimmunoassay showed full
agreement of values. The stability of insulin from blood stored in various
conditions was studied. Zhu et al. developed a LC/ESI/MS/MS assay for
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the quantitation of human insulin, its analog, and the catabolites in plasma.
The compounds were isolated through precipitation, followed by SPE. The
quantitation range was 1 to 500 mg/l.

2.4.3 Muscle Relaxants

Muscle relaxants usually belong to classes of quaternary ammonium com-
pounds of high polarity and low thermal stability. Therefore, liquid chroma-
tography is a method of choice for their separation.
Farenc et al. applied LC-ESI-MS for the determination of rocuronium
(a neuromuscular blocking agent used widely during general anesthesia) in
the plasma of patients. The drug and internal standard (verapamil) were
extracted from plasma with dichloromethane and separated on an ODS
column in ACN–0.1% TFA (gradient elution). Protonated quasi-molecular
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ions for both compounds were monitored. The LOQ was 25 mg/l. Ballard
et al. developed a LC/qTOF/MS/MS method for determination of pancu-
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ronium, vecuronium, tubocurarine, rocuronium, and succinylcholine in
postmortem tissues. These drugs were isolated using a combination of solvent

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