Angelines Cruz

Target screening and confirmation of 35 licit and illicit drugs and metabolites in hair by LC–MSMS

A liquid chromatography-tandem mass spectrometry (LC-MSMS) target screening in 50mg hair was developed and fully validated for 35 analytes (Δ9-tetrahidrocannabinol (THC), morphine, 6-acetylmorphine, codeine, methadone, fentanyl, amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxymethamphetamine, benzoylecgonine, cocaine, lysergic acid diethylamide, ketamine, scopolamine, alprazolam, bromazepam, clonazepam, diazepam, flunitrazepam, 7-aminoflunitrazepam, lorazepam, lormetazepam, nordiazepam, oxazepam, tetrazepam, triazolam, zolpidem, zopiclone, amitriptyline, citalopram, clomipramine, fluoxetine, paroxetine and venlafaxine). Hair decontamination was performed with dichloromethane, and incubation in 2 mL of acetonitrile at 50°C overnight. Extraction procedure was performed in 2 steps, first liquid-liquid extraction, hexane:ethyl acetate (55:45, v:v) at pH 9, followed by solid-phase extraction (Strata-X cartridges). Chromatographic separation was performed in AtlantisT3 (2.1 mm × 100 mm, 3 μm) column, acetonitrile and ammonium formate pH 3 as mobile phase, and 32 min total run time. One transition per analyte was monitored in MRM mode. To confirm a positive result, a second injection monitoring 2 transitions was performed. The method was specific (no endogenous interferences, n=9); LOD was 0.2-50 pg/mg and LOQ 0.5-100 pg/mg; linearity ranged from 0.5-100 to 2000-20,000 pg/mg; imprecision <15%; analytical recovery 85-115%; extraction efficiency 4.1-85.6%; and process efficiency 2.5-207.7%; 27 analytes showed ion suppression (up to -86.2%), 4 ion enhancement (up to 647.1%), and 4 no matrix effect; compounds showed good stability 24-48 h in autosampler. The method was applied to 17 forensic cases. In conclusion, a sensitive and specific target screening of 35 analytes in 50mg hair, including drugs of abuse (THC, cocaine, opiates, amphetamines) and medicines (benzodiazepines, antidepressants) was developed and validated, achieving lower cut-offs than Society of Hair Testing recommendations.

LC-MS/MS method for the determination of nine antidepressants and some of their main metabolites in oral fluid and plasma. Study of correlation between venlafaxine concentrations in both matrices.

In this paper, a fast, sensitive and selective LC-MS/MS method is described for the simultaneous determination of amitriptyline, imipramine, clomipramine, fluoxetine, paroxetine, sertraline, fluvoxamine, citalopram and venlafaxine, as well as some of their main metabolites (nortriptyline, desipramine, norclomipramine and norfluoxetine), in oral fluid and plasma. The sample (0.2 mL) was extracted with an automated solid-phase extraction system (ASPEC XL), using mixed mode OASIS MCX cartridges. Chromatographic separation was performed in a Sunfire C18 IS column (20 mmx2.1 mm, 3.5 microm), using a gradient of acetonitrile and ammonium formate (pH 3; 2 mM) as mobile phase, which allowed the elution of all the compounds in less than 5 min. The method has been fully validated in both specimens. This method was initially applied to the analysis of oral fluid and plasma samples from patients on antidepressant treatment in order to assess for which compounds it was likely to find a good correlation between both matrices. The best results were obtained for venlafaxine, so the study was extended for this compound, comparing the ratio between oral fluid and plasma concentrations (ROF/PL) in five patients on venlafaxine treatment when both samples were collected simultaneously on four different occasions. An important inter and intraindividual variability was found in oral fluid concentrations for 150 mg dose (mean=287.5 ng/m, range 58.8-531.2 ng/mL) and for 75 mg dose (mean=186.3 ng/mL, range=82.1-289.2 ng/mL). R(OF/PL) was calculated for each patient on the four different occasions, showing also a high variability (CV=24.2-69.6%).

Liquid chromatography tandem mass spectrometry determination of selected synthetic cathinones and two piperazines in oral fluid. Cross reactivity study with an on-site immunoassay device

Since the past few years, several synthetic cathinones and piperazines have been introduced into the drug market to substitute illegal stimulant drugs such as amphetamine and derivatives or cocaine due to their unregulated situation. These emerging drugs are not usually included in routine toxicological analysis. We developed and validated a LC-MS/MS method for the determination of methedrone, methylone, mephedrone, 3,4-methylenedioxypyrovalerone (MDPV), fluoromethcathinone, fluoromethamphetamine, 1-(3-chlorophenyl)piperazine (mCPP) and 3-trifluoromethylphenylpiperazine (TFMPP) in oral fluid. Sample extraction was performed using Strata X cartridges. Chromatographic separation was achieved in 10min using an Atlantis(®) T3 column (100mm×2.1mm, 3μm), and formic acid 0.1% and acetonitrile as mobile phase. The method was satisfactorily validated, including selectivity, linearity (0.2-0.5 to 200ng/mL), limits of detection (0.025-0.1ng/mL) and quantification (0.2-0.5ng/mL), imprecision and accuracy in neat oral fluid (%CV=0.0-12.7% and 84.8-103.6% of target concentration, respectively) and in oral fluid mixed with Quantisal™ buffer (%CV=7.2-10.3% and 80.2-106.5% of target concentration, respectively), matrix effect in neat oral fluid (-11.6 to 399.7%) and in oral fluid with Quantisal™ buffer (-69.9 to 131.2%), extraction recovery (87.9-134.3%) and recovery from the Quantisal™ (79.6-107.7%), dilution integrity (75-99% of target concentration) and stability at different conditions (-14.8 to 30.8% loss). In addition, cross reactivity produced by the studied synthetic cathinones in oral fluid using the Dräger DrugTest 5000 was assessed. All the analytes produced a methamphetamine positive result at high concentrations (100 or 10μg/mL), and fluoromethamphetamine also at low concentration (0.075μg/mL).

Quantification of selected synthetic cannabinoids and Δ9-tetrahydrocannabinol in oral fluid by liquid chromatography-tandem mass spectrometry.

An LC-MS/MS method for the quantification of the synthetic cannabinoids JWH-200, JWH-250, JWH-073, JWH-018, HU-211, CP 47,497 and CP 47,497-C8, and THC in oral fluid was developed and validated. Samples (0.5 mL) were extracted using Strata X cartridges (Phenomenex). Chromatographic separation was achieved with a Sunfire™ IS column (20×2.1 mm, 3.5 μm) (Waters Corp.), with formic acid 0.1% and acetonitrile as mobile phase. A different chromatographic gradient was applied for the separation of the analytes depending on the ionization mode employed, with a total chromatographic run of 14 min. Detection was performed in a Quattro Micro™ API ESCI (Waters Corp.), using electrospray in the positive mode (ESI+) for JWH-200, JWH-250, JWH-073, JWH-018 and THC, and ESI- for HU-211, CP 47,497, and CP 47,497-C8. Validation of the method included the assessment of selectivity, linearity (0.1-2.5 to 200 ng/mL), limits of detection (0.025-1 ng/mL) and quantification (0.1-2.5 ng/mL), imprecision (%CV≤14.4%), accuracy (91.8-109.7% of target concentration), extraction recovery (65.4-105.6%) and Quantisal recovery (56.1-66.7%), and matrix effect (neat oral fluid: -56.0% to 38.5%; oral fluid in Quantisal buffer: -15.1% to -71.7%). The application of this method to oral fluid samples from roadside testing will provide unique information on the use of these new synthetic drugs by Spanish drivers.

An LC-MS/MS methodological approach to the analysis of hair for amphetamine-type-stimulant (ATS) drugs, including selected synthetic cathinones and piperazines

Amphetamine-type-stimulants (ATS) are the second most commonly used group of illicit drugs worldwide. However, in the last few years, new psychoactive substances (NPS) with stimulant effects have appeared on the illegal market, which are not detected with traditional analytical methods. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination in hair of classic ATS (amphetamine, methamphetamine, 3,4-methylenedioxymethamphetamine and 3,4-methylenedioxyamphetamine), synthetic cathinones (methylone, methedrone, mephedrone, 3,4-methylenedioxypyrovalerone, (±)-4-fluoromethamphetamine and 4-fluoromethcathinone), synthetic piperazines (1-(3-chlorophenyl)piperazine (mCPP) and 3-trifluoromethylphenylpiperazine), and medicines (trazodone and phenazone) that produce mCPP as a metabolite, was developed and fully validated. Hair samples (30 mg) were incubated in acid methanol (0.1% HCl) and extracted by a mixed-mode solid-phase extraction. Chromatographic separation was performed using an Atlantis T3 (3 µm; 2.1x50 mm) analytical column, and ammonium formate 2 mM pH 3 and acetonitrile as mobile phase. The method was validated, including selectivity (no endogenous or exogenous interferences); linearity (2-20 to 2000-4000 pg/mg); limits of detection (0.2 to 5 pg/mg) and quantification (2 to 20 pg/mg); accuracy (93.4-109.4% of target concentration); imprecision (%CV<11.6%); extraction recovery (40.5-92.1%); matrix effect (24.1-227.2%); process efficiency (9.8-165.7%) and stability in the autosampler (-14.5% of loss). The method was applied to the analysis of 16 hair samples. Amphetamine (n=7; 69.1-777.1 pg/mg), methamphetamine (n=3; 120.4-1,538.9 pg/mg), MDA (n=2; 27.8-135.4 pg/mg) and MDMA (n=8; 73.4-3,654.5 pg/mg) were found. Moreover, 10 positive results for mCPP were detected (341.7->4000 pg/mg); however, in all cases trazodone identification (2085.3->4000 pg/mg) probed a licit origin of mCPP. Copyright

Determination of colchicine in biological fluids by reverse-phase HPLC. Variation of colchicine levels in rats☆

A reverse-phase HPLC method for the determination of colchicine in biological fluids is proposed. Blood, liver and kidney colchicine concentrations were determined in rats at different times following intraperitoneal (i.p.) injection of 10 mg/kg of the drug. Colchicine was extracted from the samples studied using dichloromethane at pH 8. The dry extract was redissolved in the mobile phase and analyzed with simultaneous detection at 254 and 350 nm.

A sequential spectrophotometric method for the determination of paraquat and diquat in plasma

Abstract A sequential spectrophotometric method for the simultaneous determination of paraquat and diquat in plasma using second-derivative spectroscopy after ion-pair liquid-liquid extraction was developed. The paraquat and diquat in the last extract are reduced with alkaline sodium dithionite to coloured complexes whose absorption peaks appear at a different wavelength. The two herbicides can thus be determined sequentially over the wavelength ranges 454.3—446.4 nm (diquat) and 460—396 nm (paraquat), respectively.

Analysis of Salvinorin A in urine using microextraction in packed syringe and GC–MS/MS

BACKGROUND The aim of this work was to develop and validate a method for the determination of Salvinorin A in human urine using microextraction by packed sorbent (MEPS) and GC-MS/MS. RESULTS The technique uses a sample volume as low as 0.2 ml, and the analyte was extracted using a C18 sorbent. The method showed to be linear between 20 and 1000 ng/ml and presented a LOD of 5 ng/ml. Intra- and inter-day precision and accuracy were acceptable. Absolute recoveries ranged from 71 to 80%. CONCLUSION GC-MS/MS with MEPS demonstrated to be a fast and simple procedure for the quantification of Salvinorin A in urine. This is the first time that GC-MS/MS with MEPS was used for the determination of this compound in biological fluids. Furthermore, the device could be reused for up to 80 extractions, which accounted for a lower cost of analysis.

Simultaneous Determination of Diazepam and its metabolites in Plasma by High-Performance Liquid Chromatography

Abstract A reversed phase high-performance liquid chromatographic method (HPLC) for the simultaneous determination of diazepam and its three active metabolites, nordazepam, oxazepam and temazepam, in plasma was proposed. The compounds were isolated by solid-phase extraction. The chromatographic mobile phase was metanol-water (55:45, v/v) at a flow rate of 1 mL/min. UV detection was performed concurrently at 240 and 254 nm.

Sequential Second Derivative Spectroscopy of Cocaine and Adulterants in Street Drug Samples. Part I: Cocaine, Procaine, and Lidocaine

Abstract A sequential spectrophotometric method for the resolution of mixtures of cocaine, procaine and lidocaine in powder samples is proposed. The analytes are determined sequentially in the following order: 1st-Procaine, 2nd-Cocaine and 3rd-Lidocaine. The three determinations are carried out in phosphate buffer at pH 7.8. This solvent was selected as alternative to 0.1M HCl and NaOH, because the spectral characteristics, solutions stability and reproductibility were suitable.