Study of the distribution of antidepressant drugs in vitreous humor using a validated GC/MS method.

Vitreous humor has become in recent years an important alternative biological fluid in forensic toxicological analysis especially for the investigation of cases where alcohol and drugs of abuse are involved but there is limited scientific information regarding the distribution of antidepressant drugs in this material. This work aimed to study the distribution of antidepressant drugs in vitreous humor and to estimate the blood/vitreous humor concentration ratios of these drugs. For this purpose, a GC/MS method for the simultaneous determination of 9 antidepressant drugs, namely amitriptyline, nortriptyline, citalopram, clomipramine, fluoxetine, maprotiline, mirtazapine, sertraline and venlafaxine, and 4 of their metabolites, namely desmethylmaprotiline, desmethylmirtazapine, desmethylsertraline, O-desmethylvenlafaxine, was developed and validated. The developed method includes solid-phase extraction followed by derivatization with Heptafluorobutyric Anhydride. For all analytes, LOD and LOQ were 1.50 and 5.00ng/mL, respectively, and the calibration curves were linear within the dynamic range of 5.00-500.0ng/mL (R2≥0.990). The absolute recovery was found to be ≥86.3 % for all analytes. The accuracy (%Er) was found to range between -6.58 and 6.18 %, whereas the precision (%RSD) was less than 10.9 % for all analytes. The developed method was successfully applied to vitreous humor samples from 43 blood positive cases for antidepressant drugs. Whenever antidepressant drugs were detected in blood, they were also detected in the respective vitreous humor samples. The vitreous humor/blood concentration ratios were also calculated and were found to range from 0.04-7.07. Citalopram, mirtazapine, and its metabolite desmethylmirtazapine as well as venlafaxine and its metabolite O-desmethylvenlafaxine were the most identified substances in these samples (n≥4) and their results were better statistically evaluated. Our results suggest that vitreous humor could be an appropriate matrix for the determination of antidepressants in postmortem toxicology.

A GC-MS method for the determination of furanylfentanyl and ocfentanil in whole blood with full validation.

PURPOSE: Fentanyl analogues are popular in recent years among drug addicts and have been related to many overdoses and deaths worldwide. Furanylfentanyl, ocfentanil, acetylfentanyl and butyrfentanyl are among the most common of these drugs. Methods for the determination of furanylfentanyl and ocfentanil by gas chromatography-mass spectrometry (GC-MS) in biological samples do not exist, and therefore, their development would be extremely useful for routine toxicological analysis. METHODS: A GC-MS method was developed and fully validated for the determination of furanylfentanyl and ocfentanil in whole blood. This method was also suitable for the determination of acetylfentanyl and butyrfentanyl. The method included solid-phase extraction after protein precipitation using acetonitrile, and it was applied during the toxicological investigation of forensic cases. Methadone-d 3 was used as internal standard for the quantification of the analytes. RESULTS: The limit of detection and limit of quantification values were 0.30 and 1.0 ng/mL for furanylfentanyl and ocfentanil and 0.15 and 0.50 ng/mL for acetylfentanyl and butyrfentanyl, respectively. The calibration curves were linear (R 2 ≥ 0.993) from 1.00 to 100 ng/mL for furanylfentanyl and ocfentanil and from 0.50 to 50.0 ng/mL for acetylfentanyl and butyrfentanyl. The recoveries were not lower than 85%, while accuracies and precisions were not greater than 6.0% (% error) and 8.0% (% relative standard deviation), respectively, for all four fentanyl analogues. CONCLUSIONS: The developed method is the first one in the literature for the detection of furanylfentanyl and ocfentanil in biological fluids by GC-MS, and it provides very high sensitivity comparable to that by liquid chromatography-tandem mass spectrometry.

A fully validated method for the simultaneous determination of 11 antihistamines in breast milk by gas chromatography-mass spectrometry.

Antihistamines are excreted into breast milk in small amounts; however, there are no adequate published studies or data concerning their effects on newborns and safety during breastfeeding. Thus, the development of sensitive and specific methodologies for the determination of antihistamines in breast milk is critical. A simple and sensitive GC-MS method for the simultaneous determination of 11 antihistamines (diphenhydramine, orphenadrine, chlorpheniramine, dimethindene, meclozine, hydroxyzine, loratadine, desloratadine, cetirizine, rupatadine and ebastine) in breast milk was developed and validated. The antihistamines were solid-phase extracted and derivatized with acetic anhydride and n-propanol. Diazepam-d5 , hydroxyzine-d4 and cetirizine-d8 were used as internal standards. Absolute recovery values for all analytes ranged from 70.5 to 120.0%, while the limits of detection and quantification for all analytes were 1.50 and 5.00 ng/mL, respectively. All calibration curves were linear (R2 ≥ 0.990) within the range 5.00-1000.0 ng/mL. Accuracy (Er ) ranged between -7.6 and 7.0%, while precision (RSD) was <12% for all antihistamines. The developed method is suitable for the investigation of antihistamine-related clinical cases, as well as for pharmacokinetic and breastfeeding safety studies.

Fentanyls continue to replace heroin in the drug arena: the cases of ocfentanil and carfentanil.

PURPOSE: Ocfentanil and carfentanil are two potent synthetic opioids that are analogues of fentanyl and are actively involved in the recent fentanyl crisis. The aim of this review is to provide all the available information on these two fentanyl analogues. METHODS: All reviewed information was gathered through a detailed search of PubMed and the World Wide Web using relevant keywords. RESULTS: Like most of the members of the family of fentanyls, they are either sold as heroin to unsuspecting users or used extensively to lace heroin street samples. Despite the fact that ocfentanil was studied clinically in the early 1990s, it did not manage to find its place in clinical practice. On the other hand, carfentanil is mainly used today as an anesthetic agent in large animals. Ocfentanil and carfentanil are used and abused extensively, mainly in Europe and in the United States. As a result, they are the cause of some verified intoxication cases and deaths worldwide. This review provides information concerning chemistry, synthesis, prevalence, pharmacology, and toxicology, as well as the current legal status of these two fentanyl analogues. Analytical methods developed for the determination of ocfentanil and carfentanil in biological specimens and seized materials, as well as related intoxication and lethal cases are also presented. CONCLUSIONS: Ocfentanil and carfentanil are undeniably very dangerous opioid drugs and a very serious matter of concern for public safety. The authorities should take the appropriate actions to avoid the expansion of this threat by taking proper and prompt measures.

Development and validation of a GC-MS method for the determination of hydroxyzine and its active metabolite, cetirizine, in whole blood.

A simple, rapid, sensitive and accurate gas chromatography-mass spectrometric method was developed and validated for the simultaneous determination of hydroxyzine and cetirizine in whole blood. Solid-phase extraction procedure using Bond Elut LRC Certify II columns was used for the isolation of hydroxyzine and cetirizine from 1mL whole blood followed by derivatization with a mixture of acetic anhydride:n-propanol (1:1, v/v). Limits of detection and quantification were 1.50 and 5.00ng/mL, respectively. The assay was linear within the concentration range of 5.00-1000.0ng/mL and the correlation coefficient was R2≥0.993 for both analytes. Absolute recovery was determined at three quality control concentration levels and was found to be at least 87.2% for both substances. Intra-day and inter-day accuracy values for both hydroxyzine and cetirizine were ranged from -1.2 to 3.8% and -2.7 to 2.0%, respectively, at the three concentration levels studied, whereas their respective intra-day and inter-day precision values were less than 9.9 and 6.5%, respectively, in terms of relative standard deviation (%RSD). The developed method was successfully applied for the quantification of hydroxyzine and cetirizine concentrations in whole blood, during the investigation of clinical cases where these two antihistamines were detected.

Metabolites replace the parent drug in the drug arena. The cases of fonazepam and nifoxipam.

Fonazepam (desmethylflunitrazepam) and nifoxipam (3-hydroxy-desmethylflunitrazepam) are benzodiazepine derivatives and active metabolites of flunitrazepam. They recently invaded the drug arena as substances of abuse and alerted the forensic community after being seized in powder and tablet forms in Europe between 2014 and 2016. A review of all the existing knowledge of fonazepam and nifoxipam is reported, concerning their chemistry, synthesis, pharmacology and toxicology, prevalence/use, biotransformation and their analysis in biological samples. To our knowledge, fonazepam and nifoxipam-related intoxications, lethal or not, have not been reported in the scientific literature. All the available information was gathered through a detailed search of PubMed and the World Wide Web.

Bioanalysis of antihistamines for clinical or forensic purposes.

Antihistamines are a class of drugs that inhibit the action of histamine and are used to alleviate symptoms associated with allergic reactions, but some of them can cause side effects, the most unpleasant and dangerous of which are the sedative effects that may hinder important psychological functions and impair skilled performance. These side effects could decrease safety in certain common and critical tasks, such as driving or operating machinery, leading to accidents. Antihistamines can also cause intoxications, sometimes lethal, especially when co-administered with alcohol or other sedative drugs. Thus, the development of analytical methods for their determination in biological fluids is considered to be useful for the investigation of clinical and forensic cases. These methodologies could also be used for pharmacokinetic studies. Several liquid and a few gas chromatographic methods have been developed for the determination of antihistamines in biological matrices after proper pretreatment procedures. This article reviews the published analytical methodologies that were gathered through the search in PubMed database and the recent developments on isolation or determination of antihistamines in biological materials. Current trends and future perspectives on bioanalysis of antihistamines are also discussed. Copyright © 2016 John Wiley & Sons, Ltd.

Fenethylline (Captagon) Abuse - Local Problems from an Old Drug Become Universal.

Fenethylline is a theophylline derivative of amphetamine having stimulant effects similar to those of other amphetamine-type derivatives. Fenethylline was used as medicament for hyperactivity disorders in children, narcolepsy and depression, but it has also been used as a drug of abuse under the common name of 'captagon'. Unlike other drugs of abuse, the clandestine synthesis of fenethylline is simple, using inexpensive laboratory instrumentation and raw materials legal to obtain. A review of all the existing knowledge of fenethylline is reported, concerning its chemistry, synthesis, pharmacology and toxicology, legislation, its prevalence and use as drug of abuse, as well as its analysis in biological or seized samples. Published or reported captagon-related cases and seizures are also presented. All the reviewed information was gathered through a detailed search of PubMed and the Internet. The primary drug market for fenethylline (as captagon) has traditionally been countries located on the Arabian Peninsula but also North Africa since 2013. In Arab countries, millions of captagon tablets are seized every year which represents one-third of global amphetamines seizures within a year. Furthermore, three of four patients treated for drug problems in Saudi Arabia are addicted to amphetamines, almost exclusively in the form of captagon. Significant information on fenethylline is provided for pharmacologists, toxicologists and forensic pathologists. Fenethylline, although old, has recently been introduced to the drug market, especially in Arab countries. Continuous community alertness is needed to tackle this current growing phenomenon.

Development and validation of a LC/MS method for the determination of Δ(9)-tetrahydrocannabinol and 11-carboxy-Δ(9)-tetrahydrocannabinol in the larvae of the blowfly Lucilia sericata: Forensic applications.

In a number of forensic toxicological cases, Δ(9)-tetrahydrocannabinol (THC) and its metabolite 11-carboxy-delta-9-tetrahydrocannabinol (THCA) are frequently considered as contributor factors to the event. To that, a liquid chromatographic mass spectrometric method is described for the identification and quantitation of THC and its metabolite THCA in the forensically important larvae of L. sericata. Larvae of Lucilia sericata were fortified with varying concentrations of THC and THCA covering the calibration range between 10 and 500pg/mg. For the isolation of the analytes from larvae, several extraction techniques were evaluated and finally liquid-liquid extraction under acidic pH was selected using hexane-ethyl acetate (50:50, v/v) as extraction solvent. For the chromatographic separation, a Waters Symmetry® C18 analytical column was used while the mobile phase was acetonitrile-ammonium acetate (2mM) (30:70, v/v). The detection was performed using electrospray ionization source in negative mode (ESI-) and the selected ions monitored were m/z 313 for THC and m/z 343 for THCA. The proposed method which is simple and sufficiently sensitive for the detection of THC and THCA even in a single larva sampling, assisted the investigation of a forensic case.

AH-7921: the list of new psychoactive opioids is expanded.

AH-7921 is a structurally unique synthetic opioid analgesic that has recently entered the drug arena in Europe, the USA, and Japan. Although it was synthesized and patented in the mid-1970s, it was first identified in a seized sample purchased via the Internet in July 2012 and formally brought to the attention of the European Union early warning system in August 2012 by the United Kingdom. Several in vitro experiments and animal model studies established the morphine-like analgesic action of AH-7921 as a µ-opioid receptor agonist that has been found to be several times more potent than codeine and at least as potent as morphine. This novel psychoactive substance has already led to eight non-fatal intoxications and 16 deaths in Sweden, the United Kingdom, Norway, and the USA. Thus, AH-7921 is a current public health risk, and better international collaboration, effective legislation and continuous community alertness are needed to tackle this current growing problem. The aim of this review is to summarize the current knowledge about this drug concerning its chemistry, pharmacology, and toxicology, as well as its international legal status. The limited existing analytical methodologies for the determination of AH-7921 in biological samples are also presented. Published or reported AH-7921-related cases, fatalities, or intoxications, and self reports from drug users are reviewed.

Development and validation of a GC/MS method for the simultaneous determination of levetiracetam and lamotrigine in whole blood.

A sensitive and accurate gas chromatography-mass spectrometric method was developed and validated for the simultaneous determination of levetiracetam and lamotrigine in whole blood. A solid-phase extraction (SPE) procedure using HF Bond Elut C18 columns followed by derivatization using N-methyl-N-tert-butyldimethylsilyl-trifluoroacetamide (MTBSTFA) with 1% tert-butyldimethylsilyl chloride (TBDMSCl) was used. In this assay, levetiracetam-d6 was used as internal standard. Limits of detection and quantification were 0.15 and 0.50 µg/mL, respectively, for both analytes. The method was proved to be linear within the concentration range of 0.50-50.0 µg/mL (R(2) ≥ 0.992) for both analytes. Absolute recovery was found to be at least 90.0 and 97.2% for levetiracetam and lamotrigine, respectively. Intra-day and inter-day accuracy values for both analytes were ranged from -6.5 to 4.2 and -6.6 to 3.0%, respectively, whereas their respective precision values were less than 11.4 and 8.3%. The developed method was successfully used in our laboratory for quantification of levetiracetam and lamotrigine blood concentrations during the investigation of forensic cases where these antiepileptic drugs were involved. This method could also be used for therapeutic drug monitoring purposes.

A fully validated method for the determination of lacosamide in human plasma using gas chromatography with mass spectrometry: application for therapeutic drug monitoring.

A simple gas chromatographic method with mass spectrometry detection was developed and validated for the determination of lacosamide in human plasma. Lacosamide and the internal standard, levetiracetam-d6, were extracted from 200 µL plasma, by a solid-phase extraction through HF Bond Elut C18 columns, and derivatized using N-methyl-N-tert-butyldimethylsilyltrifluoroacetamide with 1% tert-butyldimethylsilylchloride in acetonitrile. The limit of quantification was found to be 0.20 µg/mL and the assay was linear up to 20.0 µg/mL with correlation coefficient ≥0.994. The intra- and interday precision values were <4.1% in terms of relative standard deviation (%) and the values of intra- and interday accuracy were found to be within -7.2 and 5.3% in terms of relative error (%). Absolute recovery of the method for lacosamide was determined at three concentration levels and ranged from 92.5 to 97.6%. The developed method uses small volumes of plasma and proved to be simple, rapid, and sensitive for the determination of lacosamide in plasma. This method can be used in routine every day analysis of plasma samples obtained from patients who follow respective antiepileptic treatment and for the investigation of clinical and forensic cases where lacosamide is involved.

5-(2-aminopropyl)indole: a new player in the drama of 'legal highs' alerts the community.

ISSUES: 5-(2-aminopropyl)indole (5-IT) is a new psychoactive substance, a 'legal high', that recently invaded the drug arena in Europe and has already led to numerous intoxications and fatalities. Knowledge upon its pharmacology and toxicity is non-existent or restricted; the only available information involves very few published scientific articles, official reports from the European Monitoring Centre for Drugs and Drug Addiction and drug abusers' experiences expressed in online drug forums. APPROACH: A review of the existing knowledge on 5-IT is reported, concerning its chemistry and synthesis, its pharmacological and toxicological aspects, as well as information concerning the fatal and toxic consequences of its use. The existing methodologies for the determination of 5-IT in biological and seized samples as well as its legal status are also presented. All the relative data were gathered through a detailed search of PubMed and the Internet. KEY FINDINGS: No original studies have investigated and/or confirmed its pharmacological properties, acute and chronic toxicity, physiological and behavioural effects or the dependence potential of the drug. Thus, it is difficult to specify the physical effects of 5-IT in humans. This drug is a phenomenon with global significance for public health as its use can lead to intoxication and fatalities. IMPLICATIONS: Significant information on 5-IT is provided for pharmacologists, toxicologists, forensic pathologists and regulatory authorities. CONCLUSION: 5-IT is a current public health challenge. Better international collaboration, effective legislation and continuous community alertness are needed to tackle this current growing phenomenon.

2C-I-NBOMe, an "N-bomb" that kills with "Smiles". Toxicological and legislative aspects.

Substituted phenethylamines are a class of designer drugs that have recently emerged in the drug abuse market. Such substances remain legal to use, possess, and supply until these compounds become classified as scheduled. 2C-I-NBOMe or 25I-NBOMe is the N-benzyl-derivative of the iodo-substituted dimethoxy-phenethylamine (2C-I) that appeared recently in the drug market under the street name "N-Bomb". Due to its high potency, intoxications and fatal cases related to 2C-I-NBOMe use are increased worldwide. The use and trafficking of this substituted phenethylamine is banned only in some countries. A comprehensive review was performed using PubMed and Medline databases, together with additional non-peer reviewed information sources, including books and publications of state authorities in different countries, regarding chemistry, availability, pharmacology, and toxicology of 2C-I-NBOMe. Intoxications or lethal cases, published or reported, as well as the current legislation on this newly introduced drug are also reviewed.

A "krokodil" emerges from the murky waters of addiction. Abuse trends of an old drug.

"Krokodil" is the street name for the semi-synthetic opioid derivative desomorphine. Although an old drug, it re-staged on "drug arena" during the last decade causing detrimental effects to its users. Despite the fact that Russia and other former Soviet Republics were the initial plagued countries, "krokodil" arrived in Europe and United States lately, as a substitute of the relative expensive, and in many cases unavailable, heroin. It can be easily manufactured in home-environment from codeine and causes significant health problems, even deaths worldwide. The aim of this review is to summarize the current knowledge about this drug, concerning its chemistry, synthesis, pharmacology and toxicology. Published or reported "krokodil" related cases, fatalities or intoxications, as well as self reports from drug users are reviewed. The existing analytical methodologies for the determination of desomorphine in biological samples as well as its legal status are also presented.

The clavicle bone as an alternative matrix in forensic toxicological analysis.

Although human blood is the reference medium in the field of forensic toxicology, alternative matrices may be required when traditional specimens are not available, especially in the investigation of cases involving decomposing remains. Clavicle bone may provide an appropriate sample of choice since it can easily be obtained at autopsy after the removal of the breastplate for the inspection of the thoracic viscera. To the author's knowledge, this is the first time that clavicle bone is used as an alternative matrix for the detection of drugs. The present study aimed to investigate the suitability of clavicle bone as an alternative matrix for the detection of opiates. Opiates were assayed using a gas chromatography-mass spectrometry in the selected ion monitoring mode. Morphine-d6, codeine-d6 and 6-MAM-d3 were used as internal standards for the determination of morphine, codeine and 6-MAM, respectively. A GC/MS method was developed and validated for the determination of opiates in clavicle samples. Morphine, codeine and 6-MAM were successfully separated in spiked samples allowing for their detection at low levels without interferences from the matrix. Chromatographic run time was 11 min and the tested linearity ranged from 5 to 500 ng/g (r2 > 0.99) for all analytes. The method was further applied in clavicle samples of drug-related cases. Its validation parameters and the application of the developed method in clavicle samples from drug addicts, prove its suitability for the detection of opiates and potentially other drugs.

A validated GC/MS method for the determination of amisulpride in whole blood.

A sensitive GC/MS method for the determination of amisulpride in whole blood was developed, optimized and validated. Sample preparation included solid-phase extraction using HF Bond Elut C18 cartridges and further derivatization with heptafluorobutyric anhydride (HFBA). The limits of detection and quantification were 3.00 and 10.0 µg/L, respectively. The calibration curves were linear up to 1000 µg/L (R(2)≥0.991). Absolute recovery ranged from 94.2 to 101%. Accuracy was found to be between -8.7 and 1.9% and imprecision was less than 10.0%. The developed method covers the generally accepted therapeutic range but it can also cover levels above them. This makes our method suitable for the determination of amisulpride not only for clinical purposes on psychiatric patients, but also during the investigation of forensic cases where amisulpride is involved.

Toxicological analysis of formalin-fixed or embalmed tissues: a review.

During the autopsy of forensic cases, when there is no suspicion of drug use or chemical exposure, biological fluids may not be obtained for toxicological analysis, while specimens of tissues may be collected and preserved in a formalin solution for histological examination. When specific questions arise after the burial, the only possible options are the exhumation of an embalmed body or the toxicological analysis of the formalin-fixed specimens. The drug concentrations in these specimens can be altered due to the extraction efficiency and/or the chemical activity of the formalin solutions used during chemical fixation or embalming process. The aim of this paper is to review the published studies about the determination of specific groups of drugs in formalin-fixed or embalmed specimens and their stability after chemical fixation or embalming process. The analytical aspects of this determination are also discussed. The stability of drugs in formalin environment and the possible reaction of the drugs with formaldehyde, which is a highly reactive chemical substance, should always be considered during post-mortem/post-embalming forensic analysis. The additional analysis of the formalin solution in which the tissue was preserved is considered necessary. The identification and the evaluation of the possible degradation products or chemical derivatives are extremely useful during the interpretation of the results.

Diagnostic value of cardiac troponin I in postmortem diagnosis of myocardial infarction.

Troponin I (cTnI) is a very sensitive biochemical marker for the diagnosis of myocardial infarction (MI). Cardiac troponin (cTnI or cTnT) has nearly absolute myocardial tissue specificity, thereby reflecting even microscopic zones of myocardial necrosis. The aim of this study is to evaluate the pericardial fluid levels of cTnI in medicolegal autopsy cases where patients died of MI and compare them with cases where patients died of other causes. This study included 89 cases selected during a 1-year period from medicolegal autopsies. These were classified into 4 groups: (A) myocardial infarction (n = 28), (B) salt water drowning (n = 20), (C) death resulting from injury in the respiratory system (n = 16), and (D) other causes of death (n = 25), excluding MI. The mean concentrations of cTnI were 1067.03 mg/dL for group A, 546.98 mg/dL for group B, 398.75 mg/dL for group C, and 577.47 mg/dL for group D. In cases with MI (group A), there was a significant difference in the levels of cTnI compared with the other cases. More research needs to be done in order for a cutoff level to be determined.