Determination of ecgonine and seven other cocaine metabolites in human urine and whole blood by ultra-high-pressure liquid chromatography–quadrupole time-of-flight mass spectrometry

Ecgonine is suggested to be a promising marker of cocaine (COC) ingestion. A combined mass spectrometry (MS) and tandem MS (MS/MS) method was developed to simultaneously determine ecgonine and seven other metabolites of cocaine in human urine and whole blood with ultra-high-pressure liquid chromatography coupled with quadrupole time-of-flight mass spectrometry. The compounds were extracted from as little as 100 μL of sample by solid-phase extraction with a 96-well μElution solid-phase extraction plate. The protonated molecules or fragment ions at accurate mass acquired in MS mode were used to quantify specific analytes, following by dedicated MS/MS identification. The assay was linear in the range from 5 to 50-100 ng/mL for urine samples, except for ecgonine methyl ester (10-200 ng/mL) and ecgonine (40-400 ng/mL), and was linear from 1-2 to 50 ng/mL for whole blood samples, except for ecgonine methyl ester (20-1,000 ng/mL) and ecgonine (40-2,000 ng/mL). The correlation coefficients were all greater than 0.99. The limits of detection ranged from 0.2 to 16 ng/mL, and the lower limits of quantification ranged from 1 to 40 ng/mL. The repeatability and intermediate precision were 18.1 % or less. The accuracy was in the range from 80.0 to 122.9 %, process efficiencies were in the range from 8.6 to 177.4 %, matrix effects were in the range from 28.7 to 171.0 %, and extraction recoveries were in the range from 41.0 to 114.3 %, except for ecgonine (12.8 % and 9.3 % at low and high concentrations, respectively). This method was highly sensitive in comparison with previously published methods. The validated method was successfully applied to the analysis of real samples derived from forensic cases, and the results verified that, on the basis of data from four positive samples, ecgonine is a promising marker of cocaine ingestion.

Metabolism of levamisole and kinetics of levamisole and aminorex in urine by means of LC-QTOF-HRMS and LC-QqQ-MS

The antihelminthic drug Levamisole can enhance cocaine effects by conversion into the amphetamine-like drug aminorex. We describe an LC-MS method for the determination of levamisole and its metabolite aminorex in human urine. Selectivity is given, calibration curves were linear within the calibration range 2.5–250 ng/mL; limits of the method were LoD 0.51 ng/mL, LoQ 1.02 ng/mL for levamisole and LoD 0.65 ng/mL, LoQ 0.76 ng/mL for aminorex. Precision data was in accordance with the guidelines (intraday precision for aminorex ranged between 5.75 and 11.0 % for levamisole between 8.36 and 10.9 %; interday precision for levamisole 10.9–16.9 % and for aminorex 7.64–12.7 %; accuracy data for levamisole ?1.96 to –14.3 % and for aminorex?11.9 to–18.5 %). The validated method was successfully applied to study the urinary excretion of levamisole after the administration of 100 mg of levamisole orally. Levamisole and aminorex could be detected in post-administration urine samples. Levamisole could be detected up to 39 h after ingestion, while aminorex was detectable up to 54 h. Maximum aminorex concentrations were 45 ng/mL urine. Further metabolites of levamisole after oral ingestion by means of liquid chromatography hybrid quadrupole time-of-flight high-resolution mass spectrometry (LC-QTOF-HRMS) were identified. Only 0.5 % of the ingested drug was quantified as unchanged levamisole in urine. Besides aminorex, five isomers of aminorex and 4 hydroxy-metabolites of aminorex or its isomers were found. Furthermore, levamisole is also hydroxylated and eliminated free or conjugated with sulfate or glucuronide into urine.     

Molecularly imprinted polymer for selective determination of Δ9-tetrahydrocannabinol and 11-nor-Δ9-tetrahydrocannabinol carboxylic acid using LC-MS/MS in urine and oral fluid

The use of molecularly imprinted polymers (MIPs) for solid phase extraction (MISPE) allows a rapid and selective extraction compared with traditional methods. Determination of Δ⁹-tetrahydrocannabinol (THC) and 11-nor-Δ⁹-tetrahydrocannabinol carboxylic acid (THC-COOH) in oral fluid (OF) and urine was performed using homemade MISPEs for sample clean-up and liquid chromatography tandem mass spectrometry (LC-MS/MS). Cylindrical MISPE shaped pills were synthesized using catechin as a mimic template. MISPEs were added to 0.5 mL OF or urine sample and sonicated 30 min for adsorption of analytes. For desorption, the MISPE was transfered to a clean tube, and sonicated for 15 min with 2 mL acetone:acetonitrile (3:1, v/v). The elution solvent was evaporated and reconstituted in mobile phase. Chromatographic separation was performed using a SunFire C18 (2.5 μm; 2.1?×?20 mm) column, and formic acid 0.1 % and acetonitrile as mobile phase, with a total run time of 5 min. The method was fully validated including selectivity (no endogenous or exogenous interferences), linearity (1–500 ng/mL in OF, and 2.5–500 ng/mL in urine), limit of detection (0.75 and 1 ng/mL in OF and urine, respectively), imprecision (%CV <12.3 %), accuracy (98.2–107.0 % of target), extraction recovery (15.9–53.5 %), process efficiency (10.1–46.2 %), and matrix effect (<?55 %). Analytes were stable for 72 h in the autosampler. Dilution 1:10 was assured in OF, and Quantisal? matrix effect showed ion suppression (<?80.4 %). The method was applied to the analysis of 20 OF and 11 urine specimens. This is the first method for determination of THC and THC-COOH in OF using MISPE technology.     

Development and validation of two LC-MS/MS methods for the detection and quantification of amphetamines, designer amphetamines, benzoylecgonine, benzodiazepines, opiates, and opioids in urine using turbulent flow chromatography

In the context of driving ability diagnostics in Germany, administrative cutoffs for various drugs and pharmaceuticals in urine have been established. Two liquid chromatography–tandem mass spectrometry methods for simultaneous detection and quantification of amphetamines, designer amphetamines, benzoylecgonine, benzodiazepines, opiates, and opioids in urine were developed and validated. A 500-μL aliquot of urine was diluted and fortified with an internal standard solution. After enzymatic cleavage, online extraction was performed by an ion-exchange/reversed-phase turbulent flow column. Separation was achieved by using a reversed-phase column and gradient elution. For detection, a Thermo Fisher TSQ Quantum Ultra Accurate Mass tandem mass spectrometer with positive electrospray ionization was used, and the analytes were measured in multiple-reaction monitoring mode detecting two transitions per precursor ion. The total run time for both methods was about 15 min. Validation was performed according to the guidelines of the Society of Toxicological and Forensic Chemistry. The results of matrix effect determination were between 78 % and 116 %. The limits of detection and quantification for all drugs, except zopiclone, were less than10?ng/mL and less than 25 ng/mL, respectively. Calibration curves ranged from 25 to 200 ng/mL for amphetamines, designer amphetamines, and benzoylecgonine, from 25 to 250 ng/mL for benzodiazepines, from 12.5 to 100 ng/mL for morphine, codeine, and dihydrocodeine, and from 5 to 50 ng/mL for buprenorphine and norbuprenorphine. Intraday and interday precision values were lower than 15 %, and bias values within?±?15 % were achieved. Turbulent flow chromatography needs no laborious sample preparation, so the workup is less time-consuming compared with gas chromatography–mass spectrometry methods. The methods are suitable for quantification of multiple analytes at the cutoff concentrations required for driving ability diagnostics in Germany.     

Simultaneous Determination of Methadone,Heroin, Cocaine and their Metabolites in Urine by GC‐MS

Abstract

A gas chromatographic‐mass spectrometric (GC‐MS) method for the determination of methadone, heroin, cocaine, and their metabolites in urine using Selected Ion Monitoring (SIM) was developed. Following a liquid‐liquid extraction with Toxitubes A^® and using their deuterated analogs as internal standards, the analytes were derivatized with 99:1 (v/v) N,O‐bis‐trimethylsilyl‐trifluoroacetamide/trimethylchlorosilane and injected by hand, in the splitless mode, at 240°C and a purging time of 0.75 min. The mass selective detector was kept at 300°C and molecules were ionized in the electron impact mode, using an energy of 70 eV. The detector response was linear for all drugs studied over the range 50–1000 ng/mL.     

Simultaneous determination of opiates, methadone, amphetamines, cocaine, and metabolites in human placenta and umbilical cord by LC-MS/MS

LC-MS/MS methods for the quantification of morphine, morphine-3-glucuronide, morphine-6-glucuronide, codeine, 6-acetylmorphine, cocaine, benzoylecgonine, ecgonine methyl ester, hydroxybenzoylecgonine, cocaethylene, amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxymethamphetamine (MDMA), methadone, and 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine in human placenta and umbilical cord were developed and validated. Specimens (1?±?0.02 g) were homogenized with the Ultra-Turrax T8 disperser and centrifuged, and the supernatant was submitted to solid-phase extraction with Oasis MCX cartridges. Chromatographic separation was performed using an Atlantis T3 analytical column (100?×?2.1 mm, 3 μm) and a gradient of 0.1 % formic acid and acetonitrile. Selectivity was verified in 10 different blank specimens. The method was linear from 1–5 to 100–500 ng/g, depending on the analyte. Limits of detection and quantification ranged from 0.5 to 2.5 ng/g and 1 to 5 ng/g, respectively. Method imprecision was ≤15.3 %, except for MDMA at low quality control (18.1 %); accuracy, 87.1 to 114 %; extraction efficiency, 16.3 to 154.0 % (%CV?=?1.8-39.4 %); matrix effect, ?75.7 to 449.9 % (%CV?=?3.5–50 %); and process efficiency, 8.7 to 316.0 %. The method was applied to authentic placenta and umbilical cord specimens from drug-user pregnant women.     

Cannabinoids and metabolites in expectorated oral fluid after 8 days of controlled around-the-clock oral THC administration

Oral fluid (OF) is an increasingly accepted matrix for drug testing programs, but questions remain about its usefulness for monitoring cannabinoids. Expectorated OF specimens (n = 360) were obtained from 10 adult daily cannabis smokers before, during, and after 37 20-mg oral Δ⁹-tetrahydrocannabinol (THC) doses over 9 days to characterize cannabinoid disposition in this matrix. Specimens were extracted and analyzed by gas chromatography–mass spectrometry with electron-impact ionization for THC, 11-hydroxy-THC, cannabidiol, and cannabinol, and negative chemical ionization for 11-nor-9-carboxy-THC (THCCOOH). Linear ranges for THC, 11-hydroxy-THC, and cannabidiol were 0.25–50 ng/mL; cannabinol 1–50 ng/mL; and THCCOOH 5–500 pg/mL. THCCOOH was the most prevalent analyte in 344 specimens (96.9%), with concentrations up to 1,390.3 pg/mL. 11-hydroxy-THC, cannabidiol, and cannabinol were detected in 1, 1, and 3 specimens, respectively. THC was detected in only 13.8% of specimens. The highest THC concentrations were obtained at admission (median 1.4 ng/mL, range 0.3–113.6) from previously self-administered smoked cannabis. A total of 2.5 and 3.7% of specimens were THC-positive at the recommended Substance Abuse and Mental Health Services Administration (2 ng/mL) and Driving Under the Influence of Drugs, Alcohol and Medicines (DRUID) (1 ng/mL) confirmation cutoffs, respectively. THC is currently the only analyte for monitoring cannabis exposure in OF; however, these data indicate chronic therapeutic oral THC administration and illicit oral THC use are unlikely to be identified with current guidelines. Measurement of THCCOOH may improve the detection and interpretation of OF cannabinoid tests and minimize the possibility of OF contamination from passive inhalation of cannabis smoke.     

A high selective and sensitive liquid chromatography–tandem mass spectrometry method for quantization of BPA urinary levels in children

A selective and highly sensitive liquid chromatography–tandem mass spectrometry method has been developed and validated for determination of Bisphenol A (BPA) in human urine using labeled d₆-BPA as internal standard. BPA was purified from human urine by affinity chromatography on solid extraction AFFINIMIP® Bisphenol A cartridges, based on molecularly imprinted polymers. After purification, the samples were analyzed on a Phenomenex Kinetex 100?×?4.6 mm, 2.6 μm particle PFP reversed-phase HPLC column, coupled to a triple quadrupole mass spectrometer by an electrospray ion source. Analyses were performed in the multiple reaction monitoring mode and negative ionization; the product ions at 133.2 and 212.1?m/z for BPA and at 138.2 and 215.0?m/z for d₆-BPA were monitored to assess unambiguous identification. The linearity of the detector response was verified in human urine over the concentration range 0.100–200 ng/mL. The detection limit was calculated as 0.03 ng/mL and the limit of quantification of the method is 0.10 ng/mL. This LC/ESI-MS/MS method was in-house validated evaluating specificity, trueness, within-day and between-days precision. The mean recoveries of BPA from spiked urine samples were higher than 94 % and good reproducibility (relative standard deviations?≤?8.1 %) was observed. The developed method was applied to a pilot study involving 105 children, aged from 6 to 14 years (16 normal weight and 89 obese children), from the Regione Campania (Southern Italy). The aim of this study was to determine the concentrations of BPA in urine of children and possible correlations with childhood obesity.     

Development and Validation of a Sensitive LC-MS/MS Method for the Simultaneous Analysis of Three-Tropane Alkaloids in Blood and Urine

A simple and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the simultaneous analysis of three tropane alkaloids in blood and urine. After 1 mL of a blood or urine sample was extracted using a liquid–liquid extraction method with ethyl acetate at pH 8 and homatropine as the internal standard, the tropane alkaloids were separated. An Allure PFP propyl column (50 mm × 2.1 mm, 5 µm) separated the tropane alkaloids using an acetonitrile-buffer solution (20 mmol/L ammonium acetate and 0.1% formic acid, pH 4) (70:30) as the mobile phase at a flow-rate of 0.2 mL/min in isocratic mode, with the LC-MS/MS in the positive ionization mode. For each compound, detection was related to two daughter ions (scopolamine: m/z 304.4 → 138.1 and 155.9; atropine: m/z 290.3 → 124.0 and 93.1; anisodamine: m/z 306.3 → 140.1 and 91.1; and homatropine: m/z 276.3 → 124.3 and 142.1). The tropane alkaloids exhibited excellent linearity in the range of 0.05–100 ng/mL in blood and 0.2–100 ng/mL in urine, with a limit of detection range from 0.02 to 0.05 ng/mL for biological materials. The extraction recoveries of atropine, scopolamine, and anisodamine were more than 53% in the blood and urine; the interday and intraday RSDs were less than 10%; the within-day and between-day accuracy were between ?9.8% and +8.8%. The present method is simple and rapid, as shown by its application to a clinical case. This method is useful for routine analysis of tropane alkaloids in cases of suspected tropane alkaloid poisoning.     

Liquid Chromatographic Determination of Glyoxal and Methylglyoxal from Serum of Diabetic Patients using Meso‐Stilbenediamine as Derivatizing Reagent

Abstract

Meso‐stilbenediamine has been used as derivatizing reagent for liquid chromatographic (LC) determination of glyoxal (Go), methylglyoxal (MGo), and dimethylglyoxal (DMGo) at pH 3. Liquid chromatographic elution and separation was carried out from the column Kromasil 100 C‐18, 5 µm (15×0.46 mm i.d.) with methanol: water:acetonitrile (59:40:1, v/v/v) with a flow rate of 1 mL/min and ultraviolet detection at 254 nm. The linear calibration curves were obtained for Go, MGo, and DMGo within 0.97–4.86 µg/mL, 1.52–7.6 µg/mL, and 1.41–7.08 µg/mL with detection limits of 48 ng/mL, 76 ng/mL, and 70.8 ng/mL, respectively. The method was applied for the determination of Go and MGo from serum of patients suffering from diabetes and ketosis. The amounts of Go and MGo found were 0.150–0.260 µg/mL and 0.160–0.270 µg/mL with coefficient of variation (C.V.) 2.6–4.7% and 2.5–4.6%, respectively. The results obtained were compared with normal subjects with Go and MGo contents of 0.025–0.065 µg/mL and 0.030–0.070 µg/mL with C.V 1.5–4.9% and 1.6–4.8% in the serum.     

Cannabinoids in oral fluid by on-site immunoassay and by GC-MS using two different oral fluid collection devices

Oral fluid (OF) enables non-invasive sample collection for on-site drug testing, but performance of on-site tests with occasional and frequent smokers’ OF to identify cannabinoid intake requires further evaluation. Furthermore, as far as we are aware, no studies have evaluated differences between cannabinoid disposition among OF collection devices with authentic OF samples after controlled cannabis administration. Fourteen frequent (≥4 times per week) and 10 occasional (less than twice a week) adult cannabis smokers smoked one 6.8 % ?⁹-tetrahydrocannabinol (THC) cigarette ad libitum over 10 min. OF was collected with the StatSure Saliva Sampler, Oral-Eze, and Draeger DrugTest 5000 test cassette before and up to 30 h after cannabis smoking. Test cassettes were analyzed within 15 min and gas chromatography–mass spectrometry cannabinoid results were obtained within 24 h. Cannabinoid concentrations with the StatSure and Oral-Eze devices were compared and times of last cannabinoid detection (t _last) and DrugTest 5000 test performance were assessed for different cannabinoid cutoffs. 11-nor-9-Carboxy-THC (THCCOOH) and cannabinol concentrations were significantly higher in Oral-Eze samples than in Stat-Sure samples. DrugTest 5000 t _last for a positive cannabinoid test were median (range) 12 h (4–24 h) and 21 h (1–?≥?30 h) for occasional and frequent smokers, respectively. Detection windows in screening and confirmatory tests were usually shorter for occasional than for frequent smokers, especially when including THCCOOH ≥20 ng L^?1 in confirmation criteria. No differences in t _last were observed between collection devices, except for THC ≥2 μg L^?1. We thus report significantly different THCCOOH and cannabinol, but not THC, concentrations between OF collection devices, which may affect OF data interpretation. The DrugTest 5000 on-site device had high diagnostic sensitivity, specificity, and efficiency for cannabinoids.     

2-Nitro-6-monoacetylmorphine: potential marker for monitoring the presence of 6-monoacetylmorphine in urine adulterated with potassium nitrite

6-Monoacetylmorphine (6-MAM), being a unique metabolite of heroin, is routinely tested in urine samples to monitor heroin use. However, detection of 6-MAM-related opiates such as morphine is known to be affected by in vitro urine adulteration using oxidizing adulterants such as potassium nitrite. This study aimed to investigate the fate of 6-MAM after exposure to nitrite and to identify any formed oxidation products that may potentially be used for monitoring heroin abuse despite nitrite adulteration. Potassium nitrite (0.05 M and 0.6 M) was reacted with 6-MAM (5–10,000 ng/mL) in both water and blank urine with pH adjusted to range from 3 to 8. Following reaction at room temperature for varying periods, the reaction mixtures were monitored by both the CEDIA® Heroin Metabolite (6-AM) immunoassay and liquid chromatography-mass spectrometry (LC-MS) methods. Structural elucidation of the isolated oxidation products was based on mass spectrometry and nuclear magnetic resonance spectroscopic evidence. Nitrite, under acidic environment (pH?相似文献   

Liquid chromatography/tandem mass spectrometry for the qualitative and quantitative analysis of illicit drugs and medicines in preserved oral fluid

A qualitative and quantitative analytical method was developed for the simultaneous determination of 24 illicit drugs and medicines, in preserved oral fluid samples collected with the StatSure Saliva Sampler? collection device. The samples were prepared by liquid‐liquid extraction followed by liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis. The chromatographic separation was performed with an Atlantis T3 (100 × 2.1 mm i.d., 3 µm) reversed‐phase column using an acetonitrile/2 mM ammonium formate buffer pH 3.4 gradient and the MS/MS detection was achieved with two precursor‐product ion transitions per substance. The method was fully validated, including specificity and capacity of identification, limit of detection (0.2–2.1 µg/L), limit of quantitation (0.8–6.4 µg/L), recovery (34–98%), carryover, linearity (the method was linear in the range 1–200 µg/L), intra‐assay precision (coefficient of variance (CV) <20% for 20 µg/L and CV <10% for 100 µg/L) and inter‐assay accuracy (mean relative error <15%) and precision (CV <20%). The method showed to be specific and sensitive. It has already been successfully used in four proficiency tests and subsequently applied to oral fluid samples collected from road traffic volunteers in the driving population of Portugal (districts of Lisbon, Coimbra and Porto), within the DRUID project. Copyright © 2009 John Wiley & Sons, Ltd.     

Quantitation of perfluoroalkyl acids,parabens and cotinine from single low volume serum sample by validated analytical method

ABSTRACT

In this paper, we present the results of an analytical method that has been recently developed, validated and successfully applied in a biomonitoring approach. In the environmental pollutant studies it is desirable that the analytical method can determine multiple classes of compounds from a single, small volume sample. The presented analytical method with a simple sample pre-treatment allows the quantitation of 13 perfluoroalkyl acids (PFAAs), 6 parabens and cotinine (used as nicotine biomarker) from a single, small volume of 100 µL serum sample by liquid chromatography-triple quadrupole mass spectrometer (LC-MS/MS). The limits of quantitation (LOQ) for PFAAs, parabens and cotinine were 0.10–0.50, 0.20–0.80 and 0.10 ng/mL, respectively. Besides sensitivity the method has excellent trueness/accuracy and repeatability. The trueness of the method for the determination of PFAAs ranged from 95% to 106% and the repeatability (as RSD %) from 0.6% to 5.6%. The accuracy and RSD for parabens were 73–120% and 1.3–9.7%, respectively, and 100–106% and 1.3–3.5 % for cotinine. Biomonitoring data reveals the presence of several PFAAs and parabens in serum samples of Finnish population. The total concentrations for PFAAs and parabens were from 2.0 to 33 ng/mL and from <LOQ to 1100 ng/mL, respectively. Nearly all non-smokers had the serum cotinine concentration below 1.0 ng/mL, which can be suggested as the cut point for cotinine concentration to identify smoking.     

SPE and LC-MS/MS determination of 14 illicit drugs in surface waters from the Natural Park of L’Albufera (València, Spain)

A simple and robust method using solid-phase extraction (SPE) and liquid chromatography tandem mass spectrometry (LC-MS/MS) for the simultaneous determination of 14 drugs of abuse and their metabolites (cocainics, amphetamine-like compounds, cannabinoids, and opiates) in surface waters has been developed. Seven SPE adsorbents (Oasis HLB, Oasis MCX, Oasis Wax, Supelselect HLB, Strata-X, Strata-XCW), amount of sorbent bed, water volume, and pH were investigated. The highest recoveries, as well as the simplest protocol, were obtained for Oasis HLB cartridges (6 mL/200 mg) using 250 mL of water. The proposed method was linear in a concentration range from 0.03–6 to 300–60,000 ng/L depending on the compound, with correlation coefficients higher than 0.998. Matrix effects have been studied in surface water samples, and several isotope-labeled internal standards have been evaluated as a way to compensate the signal suppression observed. Limits of detection (LODs) and quantification (LOQs) ranged from 0.01 to 1.54 ng/L and from 0.03 to 5.13 ng/L, respectively. Recoveries were 71–102% at the LOQ level and 77–104 at 50 ng/L. The intra-day and intermediate precisions were from 1% to 8% and from 2% to 11%, respectively. The present work reports for the first time the occurrence of drugs of abuse residues in surface water samples from the Natural Park of L’Albufera (Valencia, Spain). Codeine, cocaine, benzoylecgonine, ecgonine methylester, amphetamine, 3,4-methylendioxy methamphetamine, morphine, and methadone were quantified with median values of 11.10, 0.02, 5.59, 0.08, 0.21, 0.75 and 0.14 ng/L respectively, and 11-nor-9-carboxy-Δ9-tetrahydrocannabinol was detected in one sample at levels <LOQ.     

An Enzyme Immunoassay for Determining Immunoreactive Trypsinogen (IRT) in Dried Blood Spots on Filter Paper Using an Ultra-Microanalytical System

Cystic fibrosis (CF) is a severe autosomal recessive disorder. It is caused by mutations in the CF transmembrane conductance regulator gene. Early diagnosis of CF can be carried out by determining high immunoreactive trypsinogen (IRT) blood values in newborns. A simple sandwich-type ultramicroELISA assay (UMELISA®) has been developed for the measurement of IRT in dried blood spots on filter paper. Strips coated with a high affinity monoclonal antibody directed against IRT are used as solid phase, to ensure the specificity of the assay. The assay is carried out within 20 h. The useful rank of the curve is 0–500 ng/mL, and the lowest detectable concentration is 4.8 ng/mL. Intra- and inter-assay coefficients of variation were lower than 10%. The recovery mean value was 100.3 ± 11.2%. Cross-reactivity with proteins structurally related to IRT (α2-macroglobulin, α1-antitrypsin, and human chymotrypsin) was lower than the detection limit of the assay. Four thousand four hundred six newborn samples from the Cuban Newborn Screening Program were analyzed, and the mean IRT concentration was 12.8 ng/mL. Higher IRT values were obtained when samples were eluted overnight. Regression analysis showed a good correlation with the commercially available AutoDELFIA® Neonatal IRT kit (n?=?3948, r =?0.885, ??=?0.976, p?<?0.01). The analytical performance characteristics of our UMELISA® TIR Neonatal suggest that it can be used for the neonatal screening of CF.     

Determination of naphthalen-1-yl-(1-pentylindol-3-yl)methanone (JWH-018) in mouse blood and tissue after inhalation exposure to 'buzz' smoke by HPLC/MS/MS

The disposition of the cannabimimetic naphthalen‐1‐yl‐(1‐pentylindol‐3‐yl)methanone (JWH‐018) in mice following inhalation of the smoke of the herbal incense product (HIP) ‘Buzz’ is presented. A high‐pressure liquid chromatography with electrospray ionization triple quadrupole mass spectrometer (HPLC/MS/MS) method was validated for the analysis of JWH‐018 in the specimens using deuterated Δ⁹‐tetrahydrocannabinol (d₃‐THC) as the internal standard. JWH‐018 was isolated by cold acetonitrile liquid–liquid extraction. Chromatographic separation was performed on a Zorbaz eclipse XDB‐C₁₈ column. The assay was linear from 1 to 1000 ng/mL. Six C57BL6 mice were sacrificed 20 min after exposure to the smoke of 200 mg ‘Buzz’ containing 5.4% JWH‐018. Specimen concentrations of JWH‐018 were: blood, 54–166 ng/mL (mean 82 ± 42 ng/mL); brain, 316–708 ng/g (mean 510 ± 166 ng/g); and liver, 1370–3220 ng/mL (mean 1990 ± 752 ng/mL). The mean blood to brain ratio for JWH‐018 was 6.8 and ranged from 4.2 to 10.9. After exposure, the responses of the mice were consistent with cannabinoid receptor type 1 activity: body temperatures dropped 7.3 ± 1.1 °C, and catalepsy, hyperreflexia, straub tail and ptosis were observed. The brain concentrations and physiological responses are consistent with the hypothesis that the behavioral effects of ‘Buzz’ are attributable to JWH‐018. Copyright © 2012 John Wiley & Sons, Ltd.     

Determination of Gingerols in Ginger by Ultra-High Performance Liquid Chromatography-Tandem Mass Spectrometry

A rapid and sensitive ultra-high performance liquid chromatography–tandem mass spectrometry method was developed and validated for the determination of 6-, 8-, and 10-gingerol in the rhizomes of Zingiber officinale Rosc. The chromatographic separation was achieved isocratically on a C₁₈ (2.1 mm × 100 mm; 1.7 µm) column using acetonitrile:water (90:10 v/v) at a flow rate of 0.25 mL/min. Mass spectrometric transitions for 6-gingerol, 8-gingerol, and 10-gingerol occurred at m/z 277.2467/177.1470, 305.3196/177.1617, and 333.3614/177.1666, respectively. The analysis time was 3.0 min and the elution of 6-, 8-, and 10- gingerol occurred at 1.27, 1.51, and 1.80 min. The linear dynamic range was established over the concentration range of 2 and 1000 ng/mL for the analytes. The limits of detection and quantitation for 6-gingerol, 8-gingerol, and 10-gingerol were 0.921, 0.856, and 1.069 ng/mL, and 2.951, 2.727, and 3.013 ng/mL, respectively. The relative standard deviation and accuracy were found to be satisfactory. The results showed significant variation (0.176%–0.290% w/w) in gingerols among twelve accessions and that the Patna and Lucknow varieties contained the highest concentrations and were concluded to be superior. These results may be used to devise strategies for cultivating this plant for large scale production.     

Simultaneous and sensitive LC–MS/MS determination of tetrahydrocannabinol and metabolites in human plasma

Cannabis is not only a widely used illicit drug but also a substance which can be used in pharmacological therapy because of its analgesic, antiemetic, and antispasmodic properties. A very rapid and sensitive method for determination of ?⁹-tetrahydrocannabinol (THC), the principal active component of cannabis, and two of its phase I metabolites in plasma has been developed and validated. After solid-phase extraction of plasma (0.2 mL), the clean extracts were analyzed by tandem mass spectrometry after a 5-min liquid chromatographic separation. The linear calibration ranges were from 0.05 to 30 ng?mL^?1 for THC and 11-nor-?⁹-carboxy-tetrahydrocannabinol (THC-COOH) and from 0.2 to 30 ng?mL^?1 for ?⁹-(11-OH)-tetrahydrocannabinol (11-OH-THC). Imprecision and inaccuracy were always below 7 and 12 % (expressed as relative standard deviation and relative error), respectively. The method has been successfully applied to determination of the three analytes in plasma obtained from healthy volunteers after oral administration of 20 mg dronabinol.