RESUMO
Gamma-hydroxybutyrate (GHB) is a central nervous system depressant that has gained popularity as an illicit recreational drug. We describe a case of an elderly woman who was found unconscious in her home. The paramedics initially suspected an intracranial incident. A head computed tomography was negative, as was the initial urinary drug screening. The diagnosis of GHB intoxication was made with the detection of GHB in a urine sample obtained 28-29 hours after the assumed time of intake. Our case underscores the importance of considering drug testing in a broad range of patients and shows that elderly patients may have an extended detection window of GHB.
Assuntos
Depressores do Sistema Nervoso Central , Drogas Ilícitas , Oxibato de Sódio , Humanos , Feminino , Idoso , Oxibato de Sódio/urina , Drogas Ilícitas/urina , Detecção do Abuso de Substâncias/métodosRESUMO
Gamma-hydroxybutyric acid (GHB) represents an important drug in clinical and forensic toxicology, particularly in the context of drug-facilitated crimes. Analytically, GHB remains a major challenge given its endogenous occurrence and short detection window. Previous studies identified a number of potential interesting novel conjugates of GHB with carnitine, amino acids (AA, glutamate, glycine, and taurine), or fatty acids. As a basis for comprehensive studies on the suitability of these novel biomarkers, we developed and validated a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method in human urine. Additionally, already known markers 2,4-dihydroxy butyric acid (2,4-DHB), 3,4-DHB, glycolic acid, succinic acid, succinylcarnitine, and GHB glucuronide were included. The method was fully validated according to (inter)national guidelines. Synthetic urine proved suitable as a surrogate matrix for calibration. Matrix effects were observed for all analytes with suppression effects of about 50% at QC LOW, and approximately 20% to 40% at QC HIGH, but with consistent standard deviation of <25% at QC LOW and <15% at QC HIGH, respectively. All analytes showed acceptable intra- and inter-day imprecision of below 20%, except for inter-day variation of GHB taurine and FA conjugates at the lowest QC. Preliminary applicability studies proved the usefulness of the method and pointed towards GHB glycine, followed by other AA conjugates as the most promising candidates to improve GHB detection. FA conjugates were not detected in urine samples yet. The method can be used now for comprehensive sample analysis on (controlled) GHB administration to prove the usefulness of the novel GHB biomarkers.
Assuntos
Oxibato de Sódio , Humanos , Cromatografia Líquida/métodos , Oxibato de Sódio/urina , Aminoácidos , Espectrometria de Massas em Tandem/métodos , Ácidos Graxos , Hidroxibutiratos/análise , Carnitina , Biomarcadores/metabolismo , Glicina , TaurinaRESUMO
γ-Hydroxybutyric acid (GHB), a neurotransmitter or neuromodulator in the human central nervous system, is often abused in drug-facilitated sexual assaults due to its euphoric and sedative effects. While the analysis of GHB has received continuous attention, its inherent characteristics pose challenges. In the current study, capillary electrophoresis (CE) with capacitively coupled contactless conductivity detection (C4D) was built, and Good's buffers were evaluated as the background electrolytes for CE separation and C4D detection. On this basis, a simple and efficient CE-C4D method was developed for GHB analysis. Through theoretical discussion and experimental optimization, the separation of GHB and related positional isomers α-hydroxybutyric acid (AHB) and ß-hydroxybutyric acid (BHB) was achieved within 4 min using 150 mM 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) as the running buffer. Under the optimized condition, the relative standard deviations of migration time and peak area were less than 1.1% and 4.5%, indicating good precision. The C4D signal of GHB showed a good linear relationship with GHB concentration in the range of 3-300 µM with a determination coefficient of 0.9997, and the detection limit was calculated to be 0.37 µM based on the signal-to-noise ratio of three. Furthermore, liquid-liquid extraction (LLE) and solid-phase extraction (SPE) were comparatively studied for sample matrix purification. Combined with the optimized SPE procedure, the developed CE-C4D method has been successfully applied for the determination of exogenous GHB in spiked beverages and endogenous GHB in human urine.
Assuntos
Oxibato de Sódio , Bebidas/análise , Condutividade Elétrica , Eletrólitos , Eletroforese Capilar/métodos , Humanos , Hidroxibutiratos , Oxibato de Sódio/urinaRESUMO
New biomarkers indicating the abuse of drugs and alcohol are still of major interest for clinical and forensic sciences. The endogenous neurotransmitter and approved drug, gamma-hydroxybutyric acid (GHB), is often illegally used for drug-facilitated crimes by spiking GHB into alcoholic beverages. Analytical detection windows of only 6 h in blood and 12 h in urine are often too short to provide reliable proof of GHB ingestion. Therefore, new biomarkers are needed to prove exogenous GHB administration. Previously, amino acid GHB conjugates were discovered in an untargeted metabolomics screening and fatty acid esters with GHB were recently discussed as promising biomarkers to enlarge the analytical detection time windows. However, the development of analytical methods is still slowed down since reference compounds for targeted screenings are still missing. In this paper, we describe simple procedures for the rapid synthesis and purification of amino acid GHB conjugates as well as fatty acid esters, which can be adopted in analytical and clinical/forensic laboratories. Structural characterization data, together with IR, 1 H-nuclear magnetic resonance (NMR), 13 C-NMR, high-resolution mass spectra (MS), and MS/MS spectra in positive and negative ionization mode are reported for all obtained GHB conjugates and GHB conjugate precursors.
Assuntos
Oxibato de Sódio , Aminoácidos , Biomarcadores , Hidroxibutiratos/urina , Laboratórios , Oxibato de Sódio/urina , Espectrometria de Massas em Tandem/métodosRESUMO
BACKGROUND: For the interpretation of concentrations of gamma-hydroxybutyrate (GHB) in post-mortem specimens, a possible increase due to post-mortem generation in the body and in vitro has to be considered. The influence of different storage conditions and the specimen type was investigated. METHOD AND MATERIAL: Post-mortem GHB concentrations in femoral venous blood (VB), heart blood (HB), serum (S) from VB, urine (U), cerebrospinal fluid (CSF) and vitreous humour (VH) were determined by gas chromatography-mass spectrometry after derivatisation. Various storage conditions, that is 4 °C or room temperature (RT) and the addition of sodium fluoride (NaF), were compared during storage up to 30 days. Additionally, bacterial colonisation was determined by mass spectrometry fingerprinting. RESULTS: Twenty-six cases without involvement of exogenous GHB were examined. GHB concentrations (by specimen) at day 0 were 3.9-22.1 mg/L (VB), 6.6-33.3 mg/L (HB), < 0.5-18.1 mg/L (U), 1.1-10.4 mg/L (CSF) and 1.7-22.0 mg/L (VH). At 4 °C, concentrations increased at day 30 to 5.6-74.5 mg/L (VB), 4.6-76.5 mg/L (HB) and < 0.5-21.3 mg/L (U). At RT, concentrations rose to < 0.5-38.5 mg/L (VB), 1.2-94.6 mg/L (HB) and < 0.5-37.5 mg/L (U) at day 30. In CSF, at RT, an increase up to < 0.5-21.2 mg/L was measured, and at 4 °C, a decrease occurred (< 0.5-6.5 mg/L). GHB concentrations in VH remained stable at both temperatures (1.2-20.9 mg/L and < 0.5-26.2 mg/L). The increase of GHB in HB samples with NaF was significantly lower than that without preservation. No correlation was found between the bacterial colonisation and extent of GHB concentration changes. CONCLUSION: GHB concentrations can significantly increase in post-mortem HB, VB and U samples, depending on storage time, temperature and inter-individual differences. Results in CSF, VH, S and/or specimens with NaF are less affected.
Assuntos
Mudanças Depois da Morte , Oxibato de Sódio/sangue , Oxibato de Sódio/líquido cefalorraquidiano , Oxibato de Sódio/urina , Manejo de Espécimes , Temperatura , Corpo Vítreo/química , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Fluoreto de Sódio , Fatores de TempoRESUMO
Gamma (γ)-hydroxybutyric acid (GHB) has been reported to be an endogenous compound in the mammalian brain. It used to treat symptoms of alcohol, opioid, and drug withdrawal and cataplexy of narcolepsy. However, it is often used for criminal purposes because it is colorless, tasteless, and has short half-life. For this reason, there is a need for a method of distinguishing between endogenous and exogenous GHB administration. Therefore, urine from rat before administration of GHB and GHB urine after the single intraperitoneal injection of GHB as 30 mg/100 g were collected from Sprague-Dawley rats (7 weeks old, 10 males and females). Negative control urine, urine from individuals suspected of taking GHB, and urine from victims who were GHB-involved crime were collected. In urine samples, GHB was extracted with two-step SPE and collected fraction was derivatized and analyzed by GC/MS and GC/C/IRMS. In GC/MS and GC/C/IRMS analysis of rat urine, there was a statistically significant difference between urine from rat before administration of GHB and GHB rat urine (p < 0.05). In GC/MS analysis of human urine samples, there was no significant difference among human urine groups (negative control, suspects' urine, and victims' urine), but in GC/C/IRMS analysis of human urine samples, there was a statistically significant difference among human urine groups (p = 0.0001). Through these results, GC/C/IRMS can be more effective tool to identify endogenous and exogenous GHB in urine than GC/MS. This study can build a drug management system in forensic investigation agency and offer interpretation method to forensic science and court.
Assuntos
Toxicologia Forense/métodos , Cromatografia Gasosa-Espectrometria de Massas/métodos , Oxibato de Sódio/urina , Adulto , Animais , Isótopos de Carbono/análise , Humanos , Ratos Sprague-Dawley , Adulto JovemRESUMO
In case of drug-facilitated sexual assault (DFSA), the evidence is frequently anecdotal, with few investigations based on scientific evidences being carried out and thus most cases are diagnosed as an acute drug or alcohol intoxication. The reason may lay in the lack of specific knowledge by the victim on the possibility to retrospectively study the allegedly events and to the absence of standardized and shared protocols among health, forensic and police subjects. On this basis, in 2015 the Unit of Forensic Toxicology of University of Florence and the Sexual Assaults Centre in Hospital Careggi have fixed a common protocol to be applied in case of DFSA. The purpose of the study was to describe the results of the application of the shared protocol for toxicological findings among women seeking health care after sexual assault, and to assess the relationship with so-called proactive DFSA drugs. We conducted a study on female patients above 18 years of age consulting the Sexual Assault Centre between 2010 and July 2018. Among the 256 patients included, 37.1% was positive at least for a substance. Alcohol was the most detected substance (57 cases), followed by Cannabis (19 cases), cocaine (15 cases) and opiates/methadone (heroine: 5; morphine:1; methadone: 6); benzodiazepines and amphetamine were found in 13 and in 2 cases, respectively. Only case of gamma-hydroxybutyrate (GHB) consumption was observed while new psychoactive substances were not detected. Among the patients suspecting proactive DFSA, sedative drug findings, not explained by voluntary intake, were encountered.
Assuntos
Vítimas de Crime/estatística & dados numéricos , Delitos Sexuais/estatística & dados numéricos , Detecção do Abuso de Substâncias , Transtornos Relacionados ao Uso de Substâncias/epidemiologia , Adolescente , Adulto , Concentração Alcoólica no Sangue , Criança , Cromatografia Líquida , Feminino , Toxicologia Forense , Humanos , Itália/epidemiologia , Pessoa de Meia-Idade , Entorpecentes/sangue , Entorpecentes/urina , Estudos Retrospectivos , Oxibato de Sódio/sangue , Oxibato de Sódio/urina , Espectrometria de Massas em Tandem , Adulto JovemRESUMO
This article reports the concentrations of gamma-hydroxybutyrate (GHB) in femoral blood and bladder urine in a case series of drug intoxication deaths (N = 37). GHB was determined in blood (B-GHB) and urine (U-GHB) by a GC-FID-GBL method and 30 mg/L was used as a cut-off concentration for reporting positive results. The mean (median) and range of GHB concentrations in bladder urine were 2,818 mg/L (1,900 mg/L) and 120-13,000 mg/L, respectively. These concentrations were appreciably higher than those in femoral blood, 637 mg/L (260 mg/L) and 30-9,200 mg/L, respectively. Urine/blood ratios of GHB were highly variable (mean 8.99, median 5.33 and range 0.16-29.3). GHB is rapidly metabolized and cleared from the bloodstream, whereas there is no metabolism occurring in the urinary bladder. In five autopsy cases, U-GHB was lower than B-GHB, which suggests that these individuals died before equilibration of the drug in all body fluids and tissues. In the other 32 deaths, U-GHB was higher than B-GHB, sometimes appreciably higher, which points towards a longer survival time after intake or administration of GHB. The analysis of urine extends the window of detection of GHB by several hours compared with blood samples, depending in part on when the bladder was last voided before death. Furthermore, the urinary concentration of GHB gives a hint about the concentration in blood during the time that the urine was produced in the kidney and stored in the bladder since the previous void.
Assuntos
Toxicologia Forense/métodos , Mudanças Depois da Morte , Oxibato de Sódio , Detecção do Abuso de Substâncias/métodos , Transtornos Relacionados ao Uso de Substâncias/mortalidade , Adulto , Autopsia , Bases de Dados Factuais , Feminino , Humanos , Masculino , Intoxicação/mortalidade , Oxibato de Sódio/sangue , Oxibato de Sódio/intoxicação , Oxibato de Sódio/urina , Manejo de EspécimesRESUMO
A two-dimensional ion chromatography system was developed for the determination of γ-hydroxybutyrate (GHB) in human urine samples. Ion exclusion chromatography was used in the first dimensional separation for elimination of urine matrices and detection of GHB above 10mgL-1, ion exchange chromatography was used in the second dimensional separation via column-switching technique for detection of GHB above 0.08mgL-1. Under the optimized chromatographic conditions, the ion exclusion and ion exchange chromatography separation system exhibited satisfactory repeatability (RSD<3.1%, n=6) and good linearity in the range of 50-1000mgL-1 and 0.5-100mgL-1, respectively. By this method, concentrations of GHB in the selected human urine samples were detected in the range of 0-1.57mgL-1. The urine sample containing 0.89mgL-1 GHB was selected to evaluate the accuracy; the spiked recoveries of GHB were 95.9-102.8%. The results showed that the two-dimensional ion chromatography system was convenient and practical for the determination of GHB in human urine samples.
Assuntos
Cromatografia por Troca Iônica , Troca Iônica , Oxibato de Sódio/urina , Urinálise/métodos , Humanos , Reprodutibilidade dos TestesRESUMO
Recently, phase-II-metabolites of γ-hydroxybutyric acid (GHB), namely GHB-ß-O-glucuronide and GHB-4-sulfate, were implemented in the scope of drug testing methods The clearance of GHB from the circulation is extremely fast due to its incorporation into the metabolic pathway of the citrate cycle. The elimination half-life of GHB from blood was reported to be dose dependent between 30 and 50min resulting in narrow detection windows of less than 12h after illicit administration or cases of drug facilitated sexual assault regardless of the biological matrix used. As sulfated metabolites tend to show prolonged half-lives and slower elimination kinetics compared to unmodified or glucuronidated drugs, the potential of GHB-4-sulfate in prolonging the detection of GHB administration was assessed. Its urinary concentrations were determined in n=100 samples from athletes and n=50 samples from sport students, and the resulting data were used to calculate a preliminary reference population-based threshold for urinary GHB-sulfate concentration. The threshold was then compared to concentrations found in post-administration urine samples collected from 3 volunteers who administered GHB within the setting of a clinical trial. Due to the large inter-individual variability of concentrations found in the reference population, GHB-4-sulfate itself was not suitable to prolong the detection times for GHB applications, even when specific gravity-corrected values were used. Therefore, a metabolomics-based approach was applied to the reference population samples and evaluated regarding other urinary metabolites that potentially correlate with the urinary excretion of GHB-4-sulfate and GHB-ß-O-glucuronide in order to find a suitable marker to normalize urinary concentrations. The most promising candidate was found at a molecular mass of 321.0696 and was preliminarily identified as ß-citryl-glutamic acid.
Assuntos
Glucuronídeos/urina , Hidroxibutiratos/urina , Oxibato de Sódio/urina , Detecção do Abuso de Substâncias/métodos , Sulfatos/urina , Biomarcadores/urina , Estudos de Casos e Controles , Meia-Vida , Humanos , MetabolômicaRESUMO
SSADH deficiency (SSADHD) is a rare autosomal recessively inherited metabolic disorder. It is associated with mutations of ALDH5A1 gene, coding for the homotetrameric enzyme SSADH. This enzyme is involved in γ-aminobutyric acid (GABA) catabolism, since it oxidizes succinic semialdehyde (SSA) to succinate. Mutations in ALDH5A1 gene result in the abnormal accumulation of γ-hydroxybutyrate (GHB), which is pathognomonic of SSADHD. In the present report, diagnosis of SSADHD in a three-month-old female was achieved by detection of high levels of GHB in urine. Sequence analysis of ALDH5A1 gene showed that the patient was a compound heterozygote for c.1226G > A (p.G409D) and the novel missense mutation, c.1498G > C (p.V500 L). By ALDH5A1 gene expression in transiently transfected HEK293 cells and enzyme activity assays, we demonstrate that the p.V500 L mutation, despite being conservative, produces complete loss of enzyme activity. In silico protein modelling analysis and evaluation of tetramer destabilizing energies suggest that structural impairment and partial occlusion of the access channel to the active site affect enzyme activity. These findings add further knowledge on the missense mutations associated with SSADHD and the molecular mechanisms underlying the loss of the enzyme activity.
Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/genética , Deficiências do Desenvolvimento/genética , Succinato-Semialdeído Desidrogenase/deficiência , Ácido gama-Aminobutírico/análogos & derivados , Sítios de Ligação , Simulação por Computador , DNA/genética , Feminino , Células HEK293 , Heterozigoto , Humanos , Lactente , Modelos Moleculares , Mutação/genética , Mutação de Sentido Incorreto , Linhagem , Oxibato de Sódio/urina , Succinato-Semialdeído Desidrogenase/genética , Ácido gama-Aminobutírico/metabolismoRESUMO
BACKGROUND: γ-hydroxybutyrate (GHB) has a high potential for illicit use; overdose of this compound results in sedation, respiratory depression and death. Tolerance to the hypnotic/sedative and electroencephalogram effects of GHB occurs with chronic GHB administration; however, tolerance to respiratory depression has not been evaluated. GHB toxicodynamic effects are mediated predominantly by GABAB receptors. Chronic treatment may affect monocarboxylate transporters (MCTs) and alter the absorption, renal clearance and brain uptake of GHB. OBJECTIVES: To determine effects of chronic GHB dosing on GHB toxicokinetics, GHB-induced respiratory depression, and MCT expression. METHODS: Rats were administered GHB 600 mg/kg intravenously daily for 5 days. Plasma, urine and tissue samples and respiratory measurements were obtained on days 1 and 5. Plasma and urine were analyzed for GHB by LC/MS/MS and tissue samples for expression of MCT1, 2 and 4 and their accessory proteins by QRT-PCR. RESULTS: No differences in GHB pharmacokinetics or respiratory depression were observed between days 1 and 5. Opposing changes in MCT1 and MCT4 mRNA expression were observed in kidney samples on day 5 compared to GHB-naïve animals, and MCT4 expression was increased in the intestine. CONCLUSIONS: The lack of tolerance observed with GHB-induced respiratory depression, in contrast to the tolerance reported for the sedative/hypnotic and electroencephalogram effects, suggests that different GABAB receptor subtypes may be involved in different GABAB-mediated toxicodynamic effects of GHB. Chronic or binge users of GHB may be at no less risk for fatality from respiratory arrest with a GHB overdose than with a single dose of GHB.
Assuntos
Transportadores de Ácidos Monocarboxílicos/biossíntese , Insuficiência Respiratória/induzido quimicamente , Oxibato de Sódio/efeitos adversos , Oxibato de Sódio/farmacocinética , Animais , Células Cultivadas , Hipnóticos e Sedativos/efeitos adversos , Hipnóticos e Sedativos/sangue , Hipnóticos e Sedativos/farmacocinética , Hipnóticos e Sedativos/urina , Masculino , Ratos , Oxibato de Sódio/sangue , Oxibato de Sódio/urina , Fatores de Tempo , ToxicocinéticaAssuntos
Overdose de Drogas/sangue , Drogas Ilícitas/sangue , Citrato de Sildenafila/sangue , Oxibato de Sódio/sangue , Adulto , Bebidas Alcoólicas/análise , Cromatografia Líquida de Alta Pressão , Contaminação de Medicamentos , Overdose de Drogas/urina , Homossexualidade Masculina , Humanos , Drogas Ilícitas/análise , Drogas Ilícitas/urina , Masculino , Parceiros Sexuais , Citrato de Sildenafila/análise , Citrato de Sildenafila/urina , Oxibato de Sódio/análise , Oxibato de Sódio/urina , Espectrometria de Massas em TandemRESUMO
Date-rape drugs have the potential to be used in drug-facilitated sexual assault, organ theft and property theft. Since they are colorless, tasteless and odorless, victims can drink without noticing, when added to the beverages. These drugs must be detected in time, before they are cleared up from the biofluids. A simultaneous extraction and determination method in urine for GHB, ketamine, norketamine, phenobarbital, thiopental, zolpidem, zopiclone and phenytoin (an anticonvulsant and antiepileptic drug) with LC-MS/MS was developed for the first time with analytically acceptable recoveries and validated. A 4 steps liquid-liquid extraction was applied, using only 1.000mL urine. A new age commercial C18 poroshell column with high column efficiency was used for LC-MS/MS analysis with a fast isocratic elution as 5.5min. A new MS transition were introduced for barbital. 222.7>179.8 with the effect of acetonitrile. Recoveries (%) were between 80.98-99.27 for all analytes, except for GHB which was 71.46. LOD and LOQ values were found in the ranges of 0.59-49.50 and 9.20-80.80ngmL(-1) for all the analytes (except for GHB:3.44 and 6.00µgmL(-1)). HorRat values calculated (between 0.25-1.21), revealed that the inter-day and interanalist precisions (RSD%≤14.54%) acceptable. The simultaneous extraction and determination of these 8 analytes in urine is challenging because of the difficulty arising from the different chemical properties of some. Since the procedure can extract drugs from a wide range of polarity and pKa, it increases the window of detection. Group representatives from barbiturates, z-drugs, ketamine, phenytoin and polar acidic drugs (GHB) have been successfully analyzed in this study with low detection limits. The method is important from the point of determining the combined or single use of these drugs in crimes and finding out the reasons of deaths related to these drugs.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Hipnóticos e Sedativos/urina , Extração Líquido-Líquido/métodos , Compostos Azabicíclicos/química , Compostos Azabicíclicos/isolamento & purificação , Compostos Azabicíclicos/urina , Barbitúricos/química , Barbitúricos/isolamento & purificação , Barbitúricos/urina , Ciências Forenses , Humanos , Hipnóticos e Sedativos/química , Hipnóticos e Sedativos/isolamento & purificação , Ketamina/química , Ketamina/isolamento & purificação , Ketamina/urina , Limite de Detecção , Modelos Lineares , Fenitoína/química , Fenitoína/isolamento & purificação , Fenitoína/urina , Piperazinas/química , Piperazinas/isolamento & purificação , Piperazinas/urina , Piridinas/química , Piridinas/isolamento & purificação , Piridinas/urina , Estupro , Reprodutibilidade dos Testes , Oxibato de Sódio/química , Oxibato de Sódio/isolamento & purificação , Oxibato de Sódio/urina , ZolpidemRESUMO
Detection of gamma-hydroxybutyric acid (GHB) became crucial in many clinical and forensic settings due to its increasing use for recreational purposes and drug-facilitated sexual assault. Its narrow window of detection of about 3-12 h in urine represents a major problem. Analogous to ethyl glucuronide, the recently identified GHB-glucuronide exhibits a longer window of detection than the parent drug. It appeared reasonable that a sulfonated metabolite of GHB (GHB-SUL) will also be formed. Due to the lack of an appropriate standard, GHB was incubated with a human liver cytosolic fraction to produce GHB-SUL. Following development of a liquid chromatography/tandem mass spectrometry (LC-MS/MS) assay to measure GHB and GHB-SUL, authentic urine samples (n = 5) were tested for GHB-SUL. These investigations revealed detectable signals of both GHB and GHB-SUL, strongly indicating that GHB is not only glucuronidated but also sulfonated. Given that sulfonated metabolites generally have longer half-life times than the corresponding free drugs, GHB-SUL may serve as a biomarker of GHB misuse along with its glucuronide.
Assuntos
Adjuvantes Anestésicos/química , Hidroxibutiratos/química , Oxibato de Sódio/química , Sulfatos/química , Adjuvantes Anestésicos/urina , Cromatografia Líquida , Humanos , Hidroxibutiratos/urina , Espectrometria de Massas , Oxibato de Sódio/urina , Detecção do Abuso de Substâncias , Sulfatos/urinaRESUMO
BACKGROUND: Many patients present to emergency departments (EDs) with an altered state of consciousness. Fast exclusion of gamma hydroxybutyrate (GHB)-associated intoxication in these patients may optimize diagnostic and therapeutic algorithms and decisions in the ED. METHODS: Between January and March 2014, a novel enzymatic test system was used to quantify GHB in blood and urine samples of suspected intoxicated patients in the ED of the University Hospital. The underlying causes for suspected intoxication and the diagnostic and therapeutic measures were documented and analysed retrospectively. RESULTS: GHB measurements were performed in 13 patients with suspected ingestion during a 3-month study period. GHB was positive in six patients showing serum levels between 61.8 mg/l and 254.8 mg/l, and GHB was tested negative in seven patients with a range of 0.3-6.2 mg/l (upper reference limit 6.1 mg/l). Additional intoxication was found in five of six GHB positive (83%, alcohol n = 2 and other drugs n = 5) and in six of seven negative-tested patients (86%, alcohol n = 5 and other drugs n = 1). CONCLUSION: GHB quantification in the ED provides specific additional information for intoxication, which can lead to more precise diagnostic and therapeutic decisions and may also be important for legal aspects. We believe that GHB analysis in unconscious patients with suspected intoxication may improve the efficient treatment of intoxicated patients.
Assuntos
Overdose de Drogas , Drogas Ilícitas/sangue , Drogas Ilícitas/urina , Oxibato de Sódio/sangue , Oxibato de Sódio/urina , Detecção do Abuso de Substâncias/métodos , Adulto , Tomada de Decisões , Overdose de Drogas/sangue , Overdose de Drogas/urina , Serviço Hospitalar de Emergência/normas , Feminino , Alemanha , Meia-Vida , Humanos , Drogas Ilícitas/toxicidade , Limite de Detecção , Masculino , Estudos Retrospectivos , Oxibato de Sódio/toxicidadeAssuntos
Adjuvantes Anestésicos/intoxicação , Oxibato de Sódio/intoxicação , Acidentes Domésticos , Adjuvantes Anestésicos/sangue , Adjuvantes Anestésicos/urina , Pré-Escolar , Escala de Coma de Glasgow , Humanos , Masculino , Respiração Artificial , Oxibato de Sódio/sangue , Oxibato de Sódio/urina , Inconsciência/induzido quimicamente , Vômito/induzido quimicamenteRESUMO
γ-Hydroxybutyric acid (GHB) is a powerful central nervous system depressant, currently used in medicine for the treatment of narcolepsy and alcohol dependence. In recent years, it has gained popularity among illegal club drugs, mainly because of its euphoric effects as well as doping agent and date rape drug. The purpose of the present work was the development of a rapid analytical method for the analysis of GHB in innovative biological matrices, namely dried blood spots (DBSs) and dried urine spots (DUSs). The analytical method is based on capillary zone electrophoresis with indirect UV absorption detection at 210 nm and capillary wall dynamic coating. The background electrolyte is composed of a phosphate buffer containing nicotinic acid (probe for detection) and cetyltrimethylammonium bromide (CTAB, reversal of electroosmosis in wall dynamic coating). The influence of probe and CTAB concentration, together with buffer pH, on migration time and signal response was investigated. Under the optimized conditions, analytical linearity and precision were satisfactory; absolute recovery values were also high (>90 %); the use of dried matrices (DBSs and DUSs) was advantageous as an alternative matrix to classical ones. No interferences were found either from the most common exogenous or from endogenous compounds. This analytical approach can offer a rapid, precise and accurate method for GHB determination in innovative biological samples, which could be important for screening purposes in clinical and forensic toxicology. Graphical Abstract CE method, by combined indirect UV detection and dynamic coating, for GHB determination in DBSs and DUSs.
Assuntos
Depressores do Sistema Nervoso Central/sangue , Depressores do Sistema Nervoso Central/urina , Teste em Amostras de Sangue Seco/métodos , Eletroforese Capilar/métodos , Oxibato de Sódio/sangue , Oxibato de Sódio/urina , Urinálise/métodos , Adulto , Teste em Amostras de Sangue Seco/economia , Eletroforese Capilar/economia , Feminino , Humanos , Limite de Detecção , Masculino , Modelos Moleculares , Urinálise/economia , Adulto JovemRESUMO
γ-Hydroxybutyrate (GHB) is a drug of abuse with a strong anesthetic effect; however, proving its ingestion through the quantification of GHB in biological specimens is not straightforward due to the endogenous presence of GHB in human blood, urine, saliva, etc. In the present study, a surrogate analyte approach was applied to accurate quantitative determination of GHB in human urine using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in order to overcome this issue. For this, (2)H6-GHB and (13)C2-dl-3-hydroxybutyrate were used as a surrogate standard and as an internal standard, respectively, and parallelism between the surrogate analyte approach and standard addition was investigated at the initial step. The validation results proved the method to be selective, accurate, and precise, with acceptable linearity within calibration ranges (0.1-1µg/ml). The limit of detection and the limit of quantification of (2)H6-GHB were 0.05 and 0.1µg/ml, respectively. No significant variations were observed among urine matrices from different sources. The stability of (2)H6-GHB was satisfactory under sample storage and in-process conditions. However, in vitro production of endogenous GHB was observed when the urine sample was kept under the in-process condition for 4h and under the storage conditions of 4 and -20°C. In order to facilitate the practical interpretation of urinary GHB, endogenous GHB was accurately measured in urine samples from 79 healthy volunteers using the surrogate analyte-based LC-MS/MS method developed in the present study. The unadjusted and creatinine-adjusted GHB concentrations in 74 urine samples with quantitative results ranged from 0.09 to 1.8µg/ml and from 4.5 to 530µg/mmol creatinine, respectively. No significant correlation was observed between the unadjusted and creatinine-adjusted GHB concentrations. The urinary endogenous GHB concentrations were affected by gender and age while they were not significantly influenced by habitual smoking, alcohol drinking, or caffeine-containing beverage drinking.
Assuntos
Oxibato de Sódio/química , Oxibato de Sódio/urina , Adulto , Cromatografia Líquida/métodos , Feminino , Humanos , Limite de Detecção , Masculino , Pessoa de Meia-Idade , Detecção do Abuso de Substâncias/métodos , Espectrometria de Massas em Tandem/métodos , Adulto JovemRESUMO
In recent years, drug-facilitated crime (DFC) has become an increasing problem. A minimum list of 80 analytes to be monitored in such cases has been proposed by the Society of Forensic Toxicologists (SOFT) including the recommended minimum performance limits (RMPL). In the present study, two liquid chromatography-tandem mass spectrometry-based screening procedures, one in positive (method I) and one in negative (method II) electrospray ionization mode were developed and validated. Gradient elution was performed on a ZORBAX Eclipse XDB-C18 column after protein precipitation of the urine samples. Detection was carried out in the scheduled multiple reaction monitoring (MRM) mode monitoring two transitions per compound. A total of 100 analytes (91 basic in method I and nine acidic in method II) could be identified using the described procedure. No interferences were observed in 30 tested blank urine samples. The RMPLs were achieved for all analytes and ranged from 1 ng/mL for fentanyl to 10 µg/mL for γ-hydroxybutyrate (GHB). Matrix effects (ME) were evaluated using the same 30 urine samples and ranged from -90 % for tetrazepam to >6,000 % for the 11-nor-9-carboxy-tetrahydrocannabinol (THC-COOH). The relative standard deviations of ME were below 25 % for the vast majority of analytes. Results for urine specimens from nine authentic DFC cases were always negative with exception of drugs prescribed to the victims. Reanalysis with the developed procedure of 24 urine samples, with a positive screening result during routine clinical toxicology analysis, confirmed the routine findings. In an excretion study after a single oral doxylamine dose (30 mg), the parent drug and its nor metabolite could be detected in urine specimens from a young female volunteer for 10 days. The developed procedure allows a selective and sensitive screening of urine samples for almost all recommended analytes relevant in DFC cases.