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  • Fast targeted analysis of 132 acidic and neutral drugs and poisons in whole blood using LC-MS/MS.

Fast targeted analysis of 132 acidic and neutral drugs and poisons in whole blood using LC-MS/MS.

Forensic science international (2014-05-06)
Matthew Di Rago, Eva Saar, Luke N Rodda, Sophie Turfus, Alex Kotsos, Dimitri Gerostamoulos, Olaf H Drummer
ABSTRACT

The aim of this study was to develop an LC-MS/MS based screening technique that covers a broad range of acidic and neutral drugs and poisons by combining a small sample volume and efficient extraction technique with simple automated data processing. After protein precipitation of 100μL of whole blood, 132 common acidic and neutral drugs and poisons including non-steroidal anti-inflammatory drugs, barbiturates, anticonvulsants, antidiabetics, muscle relaxants, diuretics and superwarfarin rodenticides (47 quantitated, 85 reported as detected) were separated using a Shimadzu Prominence HPLC system with a C18 separation column (Kinetex XB-C18, 4.6mm×150mm, 5μm), using gradient elution with a mobile phase of 25mM ammonium acetate buffer (pH 7.5)/acetonitrile. The drugs were detected using an ABSciex(®) API 2000 LC-MS/MS system (ESI+ and -, MRM mode, two transitions per analyte). The method was fully validated in accordance with international guidelines. Quantification data obtained using one-point calibration compared favorably to that using multiple calibrants. The presented LC-MS/MS assay has proven to be applicable for determination of the analytes in blood. The fast and reliable extraction method combined with automated processing gives the opportunity for high throughput and fast turnaround times for forensic and clinical toxicology.

MATERIALS
Product Number
Brand
Product Description

Hydrochlorothiazide for peak identification, European Pharmacopoeia (EP) Reference Standard
Indapamide, European Pharmacopoeia (EP) Reference Standard
Bumetanide, European Pharmacopoeia (EP) Reference Standard
Prednisolone for peak identification, European Pharmacopoeia (EP) Reference Standard
Sigma-Aldrich
Ultrapure Acetonitrile
Dimethyl sulfoxide, European Pharmacopoeia (EP) Reference Standard
Phenoxymethylpenicillin, European Pharmacopoeia (EP) Reference Standard
Sigma-Aldrich
Dimethyl sulfoxide, anhydrous, ≥99.9%
Sigma-Aldrich
Amoxicillin, 95.0-102.0% anhydrous basis
Sigma-Aldrich
8-Octanoyloxypyrene-1,3,6-trisulfonic acid trisodium salt, suitable for fluorescence, ≥90% (HPCE)
Supelco
Dimethyl sulfoxide, analytical standard
Sigma-Aldrich
Acetonitrile, ≥99.9% (GC)
Sigma-Aldrich
Hydrochlorothiazide, meets USP testing specifications
Sigma-Aldrich
Dimethyl sulfoxide, puriss. p.a., dried, ≤0.02% water
USP
Residual Solvent Class 2 - Acetonitrile, United States Pharmacopeia (USP) Reference Standard
Sigma-Aldrich
Acetonitrile, ≥99.5% (GC)
USP
Hydrochlorothiazide, United States Pharmacopeia (USP) Reference Standard
USP
Methyl alcohol, United States Pharmacopeia (USP) Reference Standard
Sigma-Aldrich
Methanol solution, suitable for NMR (reference standard), 4% in methanol-d4 (99.8 atom % D), NMR tube size 3 mm × 8 in.
Sigma-Aldrich
Methanol-12C, 99.95 atom % 12C
Supelco
Methanol solution, contains 0.10 % (v/v) formic acid, UHPLC, suitable for mass spectrometry (MS), ≥99.5%
Supelco
Hydrochlorothiazide, Pharmaceutical Secondary Standard; Certified Reference Material
Sigma-Aldrich
Acetonitrile, suitable for DNA synthesis, ≥99.9% (GC)
Sigma-Aldrich
Acetonitrile, suitable for HPLC-GC, ≥99.8% (GC)
Sigma-Aldrich
Sodium hydroxide-16O solution, 20 wt. % in H216O, 99.9 atom % 16O
Hydrochlorothiazide, European Pharmacopoeia (EP) Reference Standard
USP
Indapamide, United States Pharmacopeia (USP) Reference Standard
Supelco
Acetaminophen solution, 1.0 mg/mL in methanol, ampule of 1 mL, certified reference material, Cerilliant®
Sigma-Aldrich
Dimethyl sulfoxide, puriss. p.a., ACS reagent, ≥99.9% (GC)
Sigma-Aldrich
Dimethyl sulfoxide, Hybri-Max, sterile-filtered, BioReagent, suitable for hybridoma, ≥99.7%