Biomedical subjects
Publications and source records attributed to V Cirimele.
Hair testing is an effective method for identifying chronic drug abusers. The procedure involves a decontamination step, acid hydrolysis in presence of deuterated internal standards, liquid-liquid extraction and analysis by gas chromatography coupled to mass spectrometry. Hair analysis has a wide window of detection ranging from months to years and provides informations concerning the severity and pattern of an individual's drug use.
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Hair samples were obtained from 14 subjects admitted 2 or 3 months previously to a detoxification center. All reported an history of intravenous heroin abuse. After decontamination by two dichloromethane washes, about 50 mg hair were pulverized in a ball mill and incubated at 56 degrees C overnight in 1 mL 0.1 HCl. After neutralization, buprenorphine analyzed by RIA was in the range of 0.01 to 0.47 ng/mg. To confirm buprenorphine, liquid chromatography was used. After neutralization, drugs were extracted with toluene at pH 8.5 during a 3-step extraction procedure. A portion of the reconstituted residue was injected into a Lichrosorb CN column, with a mobile phase of phosphate buffer (pH 4.0)-acetonitrile-1-heptane sulfonic acid-butylamine (85:17:2:0.01, v/v). Detection was achieved by coulometry, and the potential of the electrodes was 0.15 and 0.50 V, respectively. Linear calibration curves were obtained from 0.02 to 2.0 ng/mg with a correlation coefficient r > 0.99 for both drugs. The detection limit for the major metabolite was about 0.01 ng/mg and 0.02 ng/mg for buprenorphine, using a 50 mg hair sample. Recovery (at 0.2 ng/mg) was 54 and 62% for norbuprenorphine and buprenorphine, respectively. Drugs concentrations in hair were in the range 0.02-0.59 and not detected--0.15 ng/mg for buprenorphine and norbuprenorphine, respectively. Results suggest that a dose-response relationship exists between the concentration of buprenorphine in hair and the administered dose.
We have tested several solvents for the decontamination of control hair contaminated by cocaine. Methylene chloride was the most efficient solvent, eliminating more than 60% of contaminating cocaine. Two washes were used, additional washes were not effective in removing more cocaine. Sixteen hair and three pubic hair samples obtained from drug abusers were tested by different extraction procedures: acid, alkaline and enzymatic hydrolyses and extraction by methanol. This study revealed that acid and enzymatic hydrolyses gave higher extraction recoveries for drugs than methanolic extraction. After alkaline hydrolysis and methanolic extraction, extracted samples were dirty and the chromatograms obtained showed an important interfering background. We concluded that acid hydrolysis is the extraction method of choice.
A method for cannabis testing in human whole blood is presented. The procedure involves extraction of a 2 mL specimen acidified with 0.2 mL of 10% acetic acid in a silanized vial in presence of deuterated internal standards into hexane:ethyl acetate. After evaporation to dryness the drug and its metabolite were derivatized by methylation with iodomethane in tetrabutylammonium hydroxide-dimethyl sulphoxide (DMSO). The derivatives were extracted into isoocatane for analysis by gas chromatography-mass spectrometry (GC-MS). The limit of quantification was 1.0 and 0.5 ng/mL for delta 9-tetrahydrocannabinol (THC) and 11-nor-9-carboxy-delta 9-tetrahydrocannabinol (THC-COOH), respectively.
delta 9-Tetrahydrocannabinol (THC), cannabidiol (CBD), and cannabinol (CBN) are three constituents of the 16 that can be currently isolated from some Cannabis spp plants. Their identification in decontaminated hair can indicate exposure to cannabis. In this study, we propose a rapid, simple, and direct (without derivatization) screening procedure for the simultaneous identification and quantitation of CBD, CBN, and THC in hair of chronic cannabis abusers. Hair samples were washed with methylene chloride, hydrolyzed with sodium hydroxide, extracted with n-hexane-ethyl acetate (9:1, v/v), evaporated to dryness, and injected directly on a gas chromatographic-mass spectrometric system operating in electron-impact mode. THC-d3 was used as the internal standard. Thirty hair samples were tested. CBD was detected 23 times, CBN was detected 22 times, and THC was detected five times. Concentrations ranged from 0.03 to 3.00 ng/mg (mean, 0.44 ng/mg), from 0.01 to 1.07 ng/mg (mean, 0.13 ng/mg), and from 0.1 to 0.29 ng/mg hair (mean, 0.15 ng/mg) for CBD, CBN, and THC, respectively. These results show that this new screening procedure is suitable for the detection of CBD and CBN in the hair of cannabis abusers.
This paper compares the methods used in Sevilla, Spain with those used in Strasbourg, France for analyzing cannabinoids (delta 9-tetrahydrocannabinol [THC] and 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid [THCCOOH]) in human hair. The Sevilla procedure involved the following steps: After washing the hair samples with methylene chloride at 37 degrees C for 15 min, hydrolysis was performed with 11.8N KOH (in the presence of THC-d3 and THCCOOH-d3) for 10 min at laboratory temperature, followed by the addition of maleic acid, extraction with n-hexane-ethyl acetate, and derivatization with heptafluorobutyric anhydride and hexafluoroisopropanol. Concentrations within the Spanish population ranged from 0.06 to 7.63 ng/mg and from 0.05 to 3.87 ng/mg for THC and THCCOOH, respectively. The Strasbourg procedure involved the following steps: Samples were decontaminated twice with methylene chloride, then pulverized and dissolved in 1N NaOH in the presence of THC-d3 and THCCOOH-d3. After cooling, acetic acid was added, and the samples were extracted with n-hexane-ethyl acetate, followed by a cleanup step with NaOH and HCl. Derivatization with pentafluoropropionic anhydride-pentafluoro-propanol was followed by gas chromatographic-mass spectrometric analysis using negative chemical ionization. Concentrations within the French population ranged from 0.10 to 2.17 ng/mg and from 0.05 to 0.39 ng/mg for THC and THCCOOH, respectively. In order to validate both analytical methods, 10 real hair samples from known drug abusers were analyzed by both procedures. The differences in the results ranged from 2 to 43% (mean, 25.7%) for THC and from 0 to 73.7% (mean, 36.8%) for THCCOOH. After comparing the cannabis results in hair in Spain (70 cases) and France (98 cases), we found quantitative as well as qualitative differences in cannabis consumption in the two populations.
A 38-year-old male was found comatose at home with an empty package of Rohypnol (2-mg tablets, 10 per package) near the body. Many other drugs, which had recently been stolen from a pharmacy, were also found. The judge in charge of the investigation of the stolen pharmaceuticals asked our laboratory to determine if the subject was a chronic user of flunitrazepam, leading our laboratory to develop a procedure for the detection of this benzodiazepine in human hair. The method involved decontamination of hair samples with dichloromethane, incubation in Sorenson buffer (pH 7.6) in the presence of diazepam-d5 used as internal standard, direct liquid-liquid extraction with diethylether-chloroform (80:20, v/v), derivatization with heptafluorobutyric anhydride and analysis by gas chromatography coupled to mass spectrometry in the negative chemical ionization mode of detection. Flunitrazepam and its major metabolite, 7-amino-flunitrazepam, were both detected. The concentrations determined in the proximal hair segment were 89.5 and 24.0 pg/mg for flunitrazepam and 7-amino-flunitrazepam, respectively.
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A fatality that was due to massive ingestion of the thioxanthene neuroleptic (Z)-cis-clopenthixol (zuclopenthixol, Z-CPT) is described. The total toxicological screening and the quantitation of both the ingested drugs and its inactive isomer (E)-trans-clopenthixol (E-CPT, produced by in vivo isomerization) in postmortem fluids and viscerae were produced by high-performance liquid chromatography (HPLC)-diode array detection. Drug confirmation was carried out by HPLC-mass spectrometry with an ionspray interface. Although death occurred 40 h after the drug intake, postmortem blood concentrations were 391 and 275 mg/mL for Z-CPT and E-CPT, respectively (50 to 100 times the usual therapeutic values). The cause of death was suicide, and the manner was acute neuroleptic overdosage.
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