Detection of coca alkaloids in oral fluid from coca leaf (tea) consumers: using solid phase extraction to improve validation parameters and widen the detection window.

Hygrine and cuscohygrine, two coca leaf alkaloids, have been previously proposed as markers to differentiate legal and illegal cocaine consumption. This is a very common problem in some countries of South America, where the consumption of coca leaves has a long tradition. Analytical methods focusing on the assessment of coca leaf alkaloids, such as cuscohygrine, hygrine, tropacocaine and t-cinnamoylcocaine, in oral fluid are virtually non-existent in forensic toxicology laboratories worldwide due to their lack of application. However, the problem of differentiating legal and illegal cocaine use in criminal justice, DUID (drug-impaired driving) and WDT (workplace drug testing) programs is growing. Therefore, researchers are obliged to develop methods to measure coca leaf alkaloids (cuscohygrine, hygrine and t-cinnamoylcocaine) in biological matrices for further validation for routine analyses in forensic toxicology laboratories. This work aims to optimize a previously published separation method by protein precipitation in oral fluid by using solid-phase extraction (SPE) coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) operating in multiple reaction monitoring (MRM) mode. The use of SPE allowed the matrix effect and the background to be reduced in the chromatograms due to the obtained cleaner extracts. Consequently, improved detection and quantification limits were reached. Findings showed that the detection windows for coca leaf alkaloids were longer than three hours in real oral fluid samples from volunteers who drank a cup of coca tea. These detection windows are quite higher than those previously obtained when using the method based on separation by protein precipitation.

Disappearance of codeine, morphine and 6-MAM in hair after cessation of abuse.

Research on the determination of drugs of abuse in hair has established that drugs can be detected in hair even long after cessation of use. The purpose of this study was to analyze hair samples from chronic opioid users who were beginning a controlled drug cessation program. The study population (n = 15) is involved in a drug rehabilitation program in Santiago de Compostela, Spain. Over a 6-month period, subjects provided hair samples at 2-month intervals, with the first sample collected on the day they began the program. Codeine, morphine, and 6-MAM were analyzed by GC/MS (LOQ = 0.2 ng/mg). Hair tresses were divided into 1 cm segments and analyzed for all analytes 0-1 cm corresponding to the proximal portion to the scalp Following cessation of opioid use, traces of codeine, morphine, and 6-MAM still remained in the newly growing hair segments for a specified period. After 2 months, still 27 % of the users tested positive, and at 4 months, 20 % were positive but only for 6-MAM. However, after 6 months of abstinence, the results were negative for all analytes.

Sexual Assault: Approach to Reality in the Area of Santiago de Compostela (Galicia, Spain) through a 12-Year Retrospective Study.

Sexual assault constitutes a severely traumatic experience that impacts the lives of far too many victims each year. The underlying behaviors of the offenders are often associated with psychological, physical and social distress, and the use of psychotropic substances was found in a good number of cases. A descriptive and retrospective review of sexual assault cases has been undertaken to identify trends in the toxicology findings in Drug-Facilitated Sexual Assault in Santiago de Compostela over the past 12 years. During this period, a total of 69 cases were referred to the Forensic Toxicology Service as sexual assault cases. The sex and age distribution of the cases showed that females between the ages of 14 and 65 years constituted the group most frequently submitted to sexual assault, with a peak of 55.1% in the 18- to 30-year age group. Alcohol consumption was positive in 77.1% of positive cases determined, followed by drugs (34.4%) and illicit drugs (26.2%). Our results showed a high percentage of alcohol consumption in sexual assault cases.

Determination of Seven Antidepressants in Pericardial Fluid by Means of Dispersive Liquid-Liquid Microextraction (DLLME) and Gas Chromatography-Mass Spectrometry (GC/MS).

Although blood is often used to detect and quantify the presence of drugs, there are some instances where samples obtained from other biological matrices, like pericardial fluid (PF), are necessary since adequate blood samples may not be available. PF is an epicardial transudate, which contains plasma components that include toxicological substances making this sample useful when blood samples are not available. This fluid is a well preserved postmortem sample and can easily be collected in larger amounts without significant contamination, compared with other body fluids. Although studies involving PF began around the 1980's, the adequacy of such fluid as a biological matrix has been poorly investigated. Antidepressants are frequently detected in postmortem samples from forensic cases. Nowadays, they constitute some of the most commonly prescribed drugs worldwide. A total of seven antidepressants (venlafaxine, mirtazapine, olanzapine, paroxetine, sertraline, fluoxetine and citalopram) were evaluated in this study. A new extraction method involving dispersive liquid-liquid microextraction (DLLME) is presented in which chloroform and acetonitrile are determined to be the best extraction and dispersing solvents. The experimental design was achieved using StatGraphics 18. The Response Surface Methodology enabled us to know the optimal volume for the two solvents used in the DLLME. The detection technique used was gas chromatography-mass spectrometry (GC-MS) with electron impact ionization as ionization source. A temperature gradient has been used and the total chromatographic separation time was 19.43 min. Validation results met the international validation guidances (FDA). Under the optimal condition, the method offered good validation parameters showing a new efficient, simple, rapid, and sensitive method. The analytical method was applied to thirty-one pericardial fluid samples. Twenty-one samples were positive with concentrations between 0.19 and 8.48 µg/mL. Venlafaxine and olanzapine were the antidepressants most frequently found.

Determination of benzodiazepines in pericardial fluid by gas chromatography-mass spectrometry.

In Forensic Toxicology it is sometimes impossible to obtain a valid blood sample to perform toxicological analysis due to several factors like advanced state of decomposition, severe burns, bleed to death…. Pericardial Fluid has already been studied during the last years and has been proposed as a valid specimen for toxicological tests. Over the years, the consumption of benzodiazepines spread among the drug dependent population and became noticeable in drug facilitated assault cases and road accidents. Improvement of the analytical methodology required for detecting the presence of these drugs in biological samples is of great importance for forensic toxicology, in order to correctly diagnose an exposure or a poisoning. In this study, 9 benzodiazepines (diazepam, nordiazepam, midazolam, bromazepam, oxazepam, temazepam, lorazepam, clonazepam and alprazolam) have been determined in pericardial fluid. For this purpose a solid phase extraction (SPE) was carried out using Bond Elut Certify cartridges. After the derivatization of six of the nine benzodiazepines, gas chromatography coupled to a selective mass detector was used as the technique for the separation of the analytes. The method developed was fully validated for the 9 analytes and was applied to real samples of pericardial fluid received at the Forensic Toxicology Service of the University of Santiago de Compostela. Finally, they were compared with blood results looking for the existence of a possible correlation between both biological samples.