UHPLC-ESI-MS/MS method for direct analysis of drugs of abuse in oral fluid for DUID assessment.

An ultra-high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry method for the direct analysis in oral fluid (OF) of several abused drugs and metabolites in a single chromatographic run was set up and validated. Amphetamine, methamphetamine, morphine, O-6-monoacetylmorphine, cocaine, codeine, methylenedioxymethamphetamine (MDMA), methylenedioxyethylamphetamine, methylenedioxyamphetamine, methadone, benzoylecgonine (BEG), Δ9-tetrahydrocannabinol (THC), ketamine, and cocaethylene were determined in a single chromatographic run with no sample pretreatment, after addition of the respective deuterated internal standards. The method was designed to perform a confirmation analysis on the residual OF samples after the preliminary on-site screening test, and it was applied on preservative buffers from different devices (Mavand Rapidstat, Concateno DDS, and Greiner Bio-One) or on neat OF samples. The method was suitable to be applied to the small amounts of sample available for the confirmatory analysis after the preliminary on-site screening or on undiluted OF samples. Limits of detection varied from 5 (morphine) to 0.2 ng/mL (methamphetamine, MDMA, BEG, and cocaethylene). The method was linear for all the substances involved, giving quadratic correlation coefficients of >0.99 in all the different preservative buffers checked. In addition, repeatability and accuracy were satisfactory for the majority of the substances, except for a few cases. The developed method was subsequently applied to 466 residual samples from on-site screening performed by police officers. Of these samples, 74 showed the presence of cocaine and metabolites; THC was detected in 49 samples. Two samples showed codeine and morphine while MDMA was detected in 11 samples and ketamine in four samples.

Liquid chromatography/mass spectrometry analysis of exhaled leukotriene B4 in asthmatic children.

BACKGROUND: The role of leukotriene (LT) B4, a potent inflammatory mediator, in atopic asthmatic and atopic nonasthmatic children is largely unknown. The lack of a gold standard technique for measuring LTB4 in exhaled breath condensate (EBC) has hampered its quantitative assessment in this biological fluid. We sought to measure LTB4 in EBC in atopic asthmatic children and atopic nonasthmatic children. Exhaled nitric oxide (NO) was measured as an independent marker of airway inflammation. METHODS: Fifteen healthy children, 20 atopic nonasthmatic children, 25 steroid-naïve atopic asthmatic children, and 22 atopic asthmatic children receiving inhaled corticosteroids were studied. The study design was of cross-sectional type. Exhaled LTB4 concentrations were measured using liquid chromatography/mass spectrometry-mass spectrometry (LC/MS/MS) with a triple quadrupole mass spectrometer. Exhaled NO was measured by chemiluminescence with a single breath on-line method. LTB4 values were expressed as the total amount (in pg) of eicosanoid expired in the 15-minute breath test. Kruskal-Wallis test was used to compare groups. RESULTS: Compared with healthy children [87.5 (82.5-102.5) pg, median and interquartile range], exhaled LTB4 was increased in steroid-naïve atopic asthmatic [255.1 (175.0-314.7) pg, p < 0.001], but not in atopic nonasthmatic children [96.5 (87.3-102.5) pg, p = 0.59)]. Asthmatic children who were receiving inhaled corticosteroids had lower concentrations of exhaled LTB4 than steroid-naïve asthmatics [125.0 (25.0-245.0) pg vs 255.1 (175.0-314.7) pg, p < 0.01, respectively]. Exhaled NO was higher in atopic nonasthmatic children [16.2 (13.5-22.4) ppb, p < 0.05] and, to a greater extent, in atopic steroid-naïve asthmatic children [37.0 (31.7-57.6) ppb, p < 0.001] than in healthy children [8.3 (6.1-9.9) ppb]. Compared with steroid-naïve asthmatic children, exhaled NO levels were reduced in asthmatic children who were receiving inhaled corticosteroids [15.9 (11.5-31.7) ppb, p < 0.01]. CONCLUSION: In contrast to exhaled NO concentrations, exhaled LTB4 values are selectively elevated in steroid-naïve atopic asthmatic children, but not in atopic nonasthmatic children. Although placebo control studies are warranted, inhaled corticosteroids seem to reduce exhaled LTB4 in asthmatic children. LC/MS/MS analysis of exhaled LTB4 might provide a non-invasive, sensitive, and quantitative method for airway inflammation assessment in asthmatic children.

Determination of benzimidazole anthelmintics in animal-derived biological matrices.

This paper describes an easy multiresidue procedure for the determination of 8 benzimidazole anthelmintics (Albendazole, Albendazole sulphone, Albendazole sulphoxide, Ox-bendazole, Ossibendazole, p-OH-Fenbendazole, Fenbendazole, Flubendazole) in foodstuffs of animal origin for human consumption. According to the proposed procedure, 10 g of homogenised sample were extracted with acetonitrile in ultrasonic bath, the organic layer was evaporated to dryness and the residue dissolved in HCl 0.005 M. After an initial washing with hexane to remove fats, the aqueous layer was purified on SPE C2 column and the analytes were eluted with methanol, evaporated to dryness and reconstituted with phosphate buffer/acetonitrile (50/50). The solution obtained was analysed by HPLC with diode array detector (DAD); this detector provides useful information about the peaks present in routine samples, because it gains not only retention times of analytes, as well as their UV-spectra. Benzthiazuron was used as internal standard and the method was validated in the range 10-100 ppb, as requested by the imposed limits. The mean recoveries in spiked samples ranged between 70% and 95%, for 10 and 100 ppb respectively. The developed method resulted sensitive, simple and fast. It is particularly suitable for laboratories that execute screening analysis on a great number of samples, observing the EC regulations that establish the benzimidazole Maximum Residue Limits (MRLs) in food.

A snapshot of workplace drug testing in Italy.

The Italian Decree on Health and Safety at Work (81/08) prescribes mandatory drug tests for jobs which pose safety hazards to others. Workplace drug testing is performed in accordance with the Provision of the Government-Regions Conference, 2008. The aim of our survey was to examine the prevalence of drug use and the main drug findings in a sample of Italian workers performing hazardous jobs. From September 2009 to February 2011, 551 urine samples were collected in 42 Italian companies. Sample collection was carried out at the workplace by qualified laboratory personnel sent from the Institute of Occupational Medicine of the Catholic University (UCSC) of Rome. The workers to be tested were informed the day before, as the law requires. The samples were checked for adulteration, coded, and sent immediately to the laboratory of the UCSC Forensic Toxicology Analytical Unit. The screening test was an immunoassay. The positive samples proceeded to the confirmatory analysis with liquid chromatography-tandem mass spectrometry (LC-MS/MS). The urine samples were analyzed for cannabis, opiates, amphetamines, methamphetamines, cocaine, methadone, and MDMA. Out of 16 samples .9% screened positive; only 4 of them (0.7%) were confirmed with the LC-MS/MS. Confirmed results included cocaine (2 samples), cannabis (1 sample), both cocaine and cannabis (1 sample). The prevalence of positive samples was lower than expected. Such finding cannot be explained by a low reliability of the testing procedure but could be due to test scheduling. More positive cases might be found performing short-notice random testing.

Ion trap liquid chromatography/tandem mass spectrometry analysis of leukotriene B4 in exhaled breath condensate.

The objective of this study is the measurement of leukotriene B7 (LTB4), a potent inflammatory mediator, in exhaled breath condensate by using liquid chromatography/mass spectrometry (LC/MS and LC/MS/MS). Condensation of exhaled breath is a non-invasive method to collect airway secretions. Deuterated (d4)-LTB4 was used as internal standard. The MS and MS/MS behavior of LTB4 and LTB4-d4 was studied by electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) in both positive and negative ion polarity mode. Preliminary results show that monitoring negative ions in ESI mode has the best sensitivity for both LTB4 and LTB4-d4. Therefore, negative ESI was chosen, and the [M-H]- ions at m/z 335 and 339 were selected for quantification. The lower limit of quantification for LTB4, expressed as the lowest point of the calibration curve, was 100 pg/mL. Using this technique, we measured LTB4 in exhaled breath condensate in two healthy subjects, four asthmatic patients on anti-inflammatory treatment, and four asthmatic patients who were not on anti-inflammatory drugs. Exhaled LTB4 concentrations were detected only in asthmatic patients who were not on anti-inflammatory therapy. This method is potentially useful for non-invasive assessment of airway inflammation, but the sensitivity of the technique needs to be improved.