Toxicological Interpretation Research Articles

A study of the pharmacokinetics of clozapine and its metabolites by the dynamics of its distribution in the oral fluid of healthy volunteers.

Clozapine (CLZ) -related accidents or crimes are common in the world. Oral fluid drug detection is a convenient measure of dealing with things like that. There has not been any literature reported detailedly the representation rule of clozapine and its metabolites in oral fluid so far. The study aimed to describe the pharmacokinetics of CLZ and its metabolites N-desmethylclozapine and clozapine-N-oxide in human oral fluid after a single 12.5mg oral dose of CLZ. Twenty-nine volunteers, including 20 males and 9 females, were recruited, and 2mL oral fluid was collected from each participant at post-consumption time-points of prior (zero), 0.5, 1.5, 3, 5, 8, 12, 24, 36, 51, 82, and 130h, respectively. Analytes of interest were extracted with solid-phase extraction and analyzed with liquid chromatography tandem mass spectrometry method. Pharmacokinetic parameters were calculated using the pharmacokinetic software DAS according to the non-compartment model. The maximum concentration, the time of maximum concentration, oral clearance, and the elimination half-life of clozapine were 16.57 ± 9.63ng/mL, 4.53 ± 3.61h, 57.65 ± 23.77 L/h and 53.58 ± 52.28h, respectively. The maximum concentration, the time of maximum concentration, and the elimination half-life of the metabolite N-desmethylclozapine were 3.08 ± 1.19ng/mL, 9.38 ± 9.33h and 62.67 ± 82.57h, respectively; of clozapine-N-oxide were 1.15 ± 0.36ng/mL, 4.53 ± 2.19h and 19.15 ± 23.11h, respectively. It was the first study on the pharmacokinetics of CLZ and its metabolites in the oral fluid of Chinese healthy volunteers, and it provided a basis for the therapeutic drug monitoring and toxicological interpretation in clozapine-related cases.

Difficulties associated with the interpretation of postmortem toxicology.

While postmortem (PM) toxicology results provide valuable information towards ascertaining both the cause and manner of death in coronial cases, there are also significant difficulties associated with the interpretation of PM drug levels. Such difficulties are influenced by several pharmacokinetic and pharmacodynamic factors including PM redistribution, diffusion, site-to-site variability in drug levels, different drug properties and metabolism, bacterial activity, genetic polymorphisms, tolerance, resuscitation efforts, underlying conditions, and the toxicity profile of cases (i.e. single- or mixed-drug toxicity). A large body of research has been dedicated for better understanding and even quantifying the influence of these factors on PM drug levels. For example, several investigative matrices have been developed as potential indicators of PM redistribution, but they have limited practical value. Reference tables of clinically relevant therapeutic, toxic, and potentially fatal drug concentrations have also been compiled, but these unfortunately do not provide reliable reference values for PM toxicology. More recent research has focused on developing databases of peripheral PM drug levels for a variety of case-types to increase transferability to real-life cases and improve interpretations. Changes to drug levels after death are inevitable and unavoidable. As such, guidelines and practices will continue to evolve as we further our understanding of such phenomena.

Atypical postmortem redistribution in chronic methadone consumers.

Available literature demonstrates that methadone is prone to moderate postmortem redistribution, but subject to high interindividual variability in the central to peripheral blood concentration ratios (C/P). In this case series, 10 cases of chronic methadone users displaying C/P < 1 (range 0.26-0.82) are described. Femoral, cardiac and ante-mortem blood concentrations of methadone and its metabolite 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) are reported for all cases, as well as sex, age, case history, results of the pathological investigation, other toxicological findings and cause and manner of death. EDDP blood concentrations, similar in both central and peripheral blood, as well as antemortem blood concentration results in Case 4, demonstrate that this atypical C/P < 1 finding is attributable to postmortem changes and not analytical or pre-analytical artifacts. Case 4 is a particularly instructive example, with femoral blood concentration (966 ng/mL) approximately twice as high as cardiac blood (499 ng/mL) and ante-mortem blood (418 ng/mL, collected 38 min prior to death)-clearly demonstrating that cardiac blood methadone concentration is more representative of the antemortem blood concentration in this case. In Case 4 and four others, toxicological interpretation based on femoral blood concentration alone would have been misleading. Based on these results and evidence from the literature, it is hypothesized that methadone bioaccumulates in the tissues of chronic users and redistributes from thigh tissues into femoral blood, increasing the concentration postmortem. This case series highlights how femoral blood is not always preserved from postmortem changes and that the analysis of multiple blood sources is necessary to avoid a misleading toxicological interpretation-particularly for cases of chronic methadone users.

Gabapentin in drugged driving investigations.

The increasing use and misuse of gabapentin pose a major risk to public health and traffic safety. Gabapentin has been approved by the Food and Drug Administration (FDA) since 1993 for adjunctive therapy in the treatment of epilepsy and neuralgia but is increasingly being prescribed for numerous off-label uses including insomnia, anxiety, depression, and migraine. Reported side effects include blurred vision, drowsiness, and loss of coordination. Driving behaviors such as exiting the lane of travel and crashes have been reported in connection to drugged driving investigations concerning gabapentin. To further assist with the toxicological interpretation of gabapentin in driving under the influence of drugs (DUID) scenarios, a review of approximately 108,000 gabapentin-positive DUID cases was conducted. Of those, 858 cases met inclusion criteria and underwent additional evaluation. Blood specimens were screened via enzyme-linked immunosorbent assay (ELISA) and confirmed by liquid chromatography tandem mass spectrometry (LC-MS/MS) for quantitation of gabapentin. This review found an overall DUID gabapentin positivity of 7.9% between January 2020 and December 2022; 17 states from various geographical regions had at least one positive gabapentin DUID case. Observations in six driving and human performance cases where gabapentin was the only drug reported were consistent with the known adverse effects of the medication. Half of the case histories reviewed involved crashes where the driver was determined to be at fault. Additionally, 94% of the cases in this review involved gabapentin in combination with other drugs. The most prevalent drug combinations were opioids and gabapentin present in 64% of cases.

Pharmacokinetics and Absorption, Distribution, Metabolism and Excretion of RGLS4326 in Mouse and Monkey, an Anti–miR-17 Oligonucleotide for the Treatment of Polycystic Kidney Disease

RGLS4326 is a short oligonucleotide inhibitor of microRNA-17 (miR-17) that preferentially distributes to the kidney and displaces miR-17 from translationally active polysomes. Here, we present pharmacokinetics and absorption, distribution, metabolism, and excretion properties of RGLS4326 from mice and monkeys. RGLS4326 was absorbed rapidly after subcutaneous administration, distributed extensively to the kidney and liver, with preferential distribution to the kidney, and cleared rapidly from plasma by tissue uptake and renal excretion. Plasma exposure increased in a dose-proportional manner with no notable accumulation after repeat doses. Plasma protein binding of RGLS4326 across all species tested was between 79% and 96%. RGLS4326 predominantly distributed to the kidney with a long half-life (t1/2; t1/2 ranged from 8-11 days) and no marked (≤twofold) accumulation in kidney and liver after repeat doses. RGLS4326 was minimally metabolized by nucleases, not cytochrome P450 (P450) isozymes, across species and underwent sequential hydrolysis from both 3' and 5' ends to produce chain-shortened metabolites. There were no human unique metabolites observed. Renal excretion was the major route of elimination of RGLS4326, and a significant fraction (50%-79%) of the dose was recovered intact in the urine of mice and monkeys across all dose levels. RGLS4326 is not a substrate, inhibitor, or inducer of P450 isozymes, and it is not a substrate or inhibitor of uptake and most efflux transporters. Thus, RGLS4326 exhibits low potential of mediating drug-drug interactions involving P450 isozymes and drug transporters. SIGNIFICANCE STATEMENT: Pharmacokinetics (PK) and absorption, distribution, metabolism, and excretion (ADME) properties of RGLS4326 were characterized in vivo and in vitro. RGLS4326 shows similar PK and ADME properties across mice and monkeys in vivo and across human and animal matrices in vitro. Subcutaneous administration results in preferential exposure of RGLS4326 to the intended target organ (kidney) to drive maximum target engagement. These studies support the interpretation of toxicology and efficacy studies and help characterize the disposition of RGLS4326 in humans.

Lipophilicity of fentalogs: Comparison of experimental and computationally derived data.

Although fentanyl and a small number of derivatives used for medical or veterinary procedures are well characterized, physiochemical properties have not been determined for many of the newer fentanyl analogs. Partition coefficients (Log P) were determined for 19 fentalogs using the shake-flask method and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Experimentally determined partition coefficients were compared with computationally derived data using six independent software sources (ACD/LogP, LogKOWWIN v 1.69, miLogP 2.2, OsirisP, XLOGP 3.0, ALogPS 2.1). Fentalogs with a wide variety of structural modifications were intentionally selected, yielding Log P values ranging from 1.21 to 4.90. Comparison of experimental and computationally derived Log P values were highly correlated (R2 0.854-0.967). Overall, substructure-based modeling using fragmental methods or property-based topological approaches aligned more closely with experimentally determined Log P values. LC-MS/MS was also used to estimate pKa values for fentalogs with no previously reported data. Lipophilicity and pKa are important considerations for analytical detection and toxicological interpretation. In silico methods allow the determination of physicochemical information prior to certified reference materials being readily available for in vitro or in vivo studies. Computationally derived data can provide insight regarding physiochemical characteristics of future fentalogs and other classes of synthetic analogs that have yet to emerge.

Drug concentrations and interactions – therapeutic or toxic?

Undertaking toxicological analysis in clinical and post-mortem investigations can assist to determine the extent of drug contribution to compliance, impairment, toxicity and potentially death. The overlap in blood concentrations following drug consumption can be significant and relying solely on a ‘number’ to determine whether or not a drug is therapeutic or toxic is difficult. Some poisons will be toxic at any measurable concentration, but for the vast majority of compounds which are detected routinely in medico-legal casework, the assessment of drug contribution in a case is dependent on many factors. These factors include but are not limited to dose, frequency, administration, drug interactions, natural disease and in some specific situations drug stability and redistribution (i.e., post-mortem). This presentation will provide an overview of the most important considerations for better interpretation of forensic toxicology results with demonstrated case examples.

Postmortem investigation of more than 100 consecutive autopsy cases in femoral blood, cardiac blood, brain tissue and muscle tissue

Drug concentrations in peripheral blood are most often used in interpretation of toxicological results in postmortem cases. However, peripheral blood is not always available or suitable for analysis, especially in advanced stages of decomposition. Therefore, other matrices than peripheral blood need to be investigated in order to reveal the cause of death. However, there is no obvious second choice. In this study, we analyzed postmortem femoral blood, cardiac blood, brain tissue and muscle tissue in order to investigate if drug concentrations in these alternative matrices were comparable to femoral blood and thereby useful for toxicological interpretation. A total of 104 consecutive autopsies where toxicological investigation was included were performed in a four-month period. Femoral blood ( n = 89), cardiac blood ( n = 85), brain tissue ( n = 91) and muscle tissue ( n = 104) were analyzed in all cases when available. Quantification of 69 common drugs and metabolites of toxicological interest was performed with four different multi-analyte methods all employing automated sample preparation and UHPLC MS/MS in MRM mode. In 104 autopsy cases, we were able to detect a total of 53 drugs and metabolites. Alternative matrix to femoral blood concentration ratios were calculated for 47 analytes for cardiac blood, 49 analytes for brain tissue and 50 analytes for muscle tissue, respectively. However, about half of the analytes were only detected in < 5 cases. Median ratios of analytes detected in ≥ 5 cases were determined to 0.59–1.9 (median = 1.1) for cardiac blood to femoral blood, 0.57–2.5 (median = 1.5) for brain tissue to femoral blood and 0.67–7.0 (median = 1.4) for muscle tissue to femoral blood. The largest deviation in ratios was observed for THC in muscle tissue. Here muscle tissue to femoral blood ratios varied from 2.3 to 20 with a median of 7.0 ( n = 10). It is, however, a well-known fact that quantitative analysis of THC in muscle tissue is challenging, which our data also support. We therefore recommend that THC in muscle tissue should be reported as “detected” instead of a concentration. Analysis of postmortem cardiac blood, brain tissue and muscle tissue revealed that concentrations were not directly comparable to those of femoral blood. However, 85% of the median ratios of alternative matrix to femoral blood of analytes detected in ≥ 5 cases, where in the range 0.5–2.0, and were therefore not markedly different. This demonstrates that interpretation of postmortem concentration in other matrices than femoral blood can be challenging and must be performed with caution.

One image is worth more than a thousand words: producing an atlas of medical signs for teaching clinical and forensic toxicology

Clinical and forensic toxicology are critically involved in the acquisition of basic skills to correctly suspect intoxication, collect biological and non-biological materials for toxicological analysis, comprehend the complexities inherent to laboratory activity, and understand the fundamentals of toxicokinetics and toxicodynamics that underlie the interpretation of results. This works presents a pedagogical innovation proposal for the teaching of clinical and forensic toxicology based on a compilation of more than 3 000 cases where the image was fulcra for suspicion. The experience in this article follows the model practiced in bachelors, masters, and PhD degrees, as well as in other continuing training courses, where we are teaching toxicology for more than 15 years. All these levels of education are considered fundamental to the sound development of this science. This approach aims also to offer strength to the intervention of the true toxicologist in all the toxicological phases, besides the classic analytical chemistry. Indeed, it is impossible to provide effective clinical and forensic toxicological interpretations without a proper and broad education, and not thinking exclusively in terms of laboratory techniques. In the future, it will be interesting to evaluate knowledge retention and to propose a database of videos of signs related to intoxications. KEY POINTS A pedagogical innovation proposal for the teaching of forensic and clinical toxicology is presented. A universal and never-ending atlas of phtotographs related to signs of intoxications have been compile. Offering to our students an integrated teaching of clinical and forensic toxicology is crucial since both are grounded in analogous toxicological principles and are mutually dependent.

Adsorption of Therapeutic and Recreational Drugs During Prolonged Storage of Plasma Samples in Gel Separator Tubes.

Hospital samples collected in gel separator tubes are often submitted to forensic toxicology laboratories for analysis in impaired driving and death investigations. Drug adsorption to the gel separator material may lead to underestimation of the drug concentration present at the time of sample collection, potentially affecting the interpretation of analytical results. Using liquid chromatography--tandem mass spectrometry (LC--MS-MS), decreases in plasma concentration of 53 drugs and metabolites relevant to forensic toxicology casework were investigated in samples stored in BD Vacutainer® PSTTM tubes for up to 3 months. After storage for only 1 day, approximately 50% of the drugs and metabolites had significantly lower concentrations in plasma separation tubes (PSTs) compared to non-gel tubes (up to 27% lower). After storage for 3 months, approximately 75% of the drugs and metabolites had significantly lower concentrations in PSTs compared to non-gel tubes (up to 69% lower). Fentanyl, carfentanil, ketamine, diphenhydramine and several antidepressants were among the drugs most susceptible to adsorption. Central nervous system stimulants (e.g., methamphetamine and amphetamine) as well as naturally-occurring and semi-synthetic opioids (e.g., morphine, hydromorphone and oxycodone) were among the drugs least susceptible to adsorption and displayed only minimal relative decreases in concentration (if any) over the 3-month sample storage period. The potential for decreases in drug concentration due to adsorption of drugs to the gel material should be considered for toxicological interpretation based on the analysis of a sample collected in a gel separator tube.

Homicidal poisoning series in a nursing home: retrospective toxicological investigations in bone marrow and hair.

Homicidal poisonings remain rare and can be difficult to detect, especially in the elderly or in medical settings. In this atypical poisoning series, a young nursing assistant purposely poisoned thirteen residents of a nursing home and killed ten of them. The medications used were a mix of psychotropic medications (cyamemazine, loxapine, tiapride, risperidone, and mirtazapine), under liquid formulation, which were inducing malaise and coma. The forensic investigation included analysis of blood, urine, hair, and bone marrow and exhumations of seven corpses up to 3years after the inhumation. Hair collected from a hairbrush of a cremated victim have been analyzed. Bone marrow sample preparation was based on a liquid/liquid triple extraction. Hair were incubated after decontamination overnight at 55°C in methanol. Segmentation was possible for seven samples, except for delayed exhumation samples (n = 4) and hairbrush hair sample (n = 1). The extracts were then analyzed using gas chromatography coupled with mass spectrometry (GC-MS) for unknown screening and using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) for a targeted screening and quantification. Screenings revealed the presence of the same mix of psychotropic medications. Cyamemazine, mirtazapine, loxapine, tiapride, and risperidone hair concentrations were 6-17,458pg/mg, 74-1271pg/mg, 9-1346pg/mg, 13-148pg/mg, and 3-5pg/mg, respectively. Cyamemazine bone marrow concentrations were 229 and 681ng/g and 152-717ng/mL in blood. Patients' medications were also identified and quantified. This poisoning series provide analytical data that could support subsequent toxicological result interpretation in similar forensic cases.

Methadone, Buprenorphine, Oxycodone, Fentanyl and Tramadol in Multiple Postmortem Matrices.

Peripheral blood (PB) concentrations are generally preferred for postmortem toxicological interpretation, but some autopsy cases may lack blood for sampling due to decomposition or large traumas, etc. In such cases, other tissues or bodily fluids must be sampled; however, limited information exists on postmortem concentrations in matrices other than blood. Pericardial fluid (PF), muscle and vitreous humor (VH) have been suggested as alternatives to blood, but only a few studies have investigated the detection of opioids in these matrices. In this study, we aimed to investigate the detection of methadone, buprenorphine, oxycodone, fentanyl and tramadol in postmortem samples of PF, skeletal muscle and VH, in addition to PB and cardiac blood and if drug concentrations in these alternative matrices were comparable to those in PB and thereby useful for interpretation. In most of the 54 included cases, only one opioid was detected. Methadone, oxycodone, fentanyl and tramadol were detected in all of the alternative matrices in almost all cases, while buprenorphine was detected less often. For methadone, the concentrations in the alternative matrices, except in VH, were relatively similar to those in PB. Larger variations in concentrations were found for buprenorphine, oxycodone and tramadol. Quantitative analyses appeared useful for fentanyl, in all of the alternative matrices, but only four cases were included. Toxicological analyses of opioids in these alternative postmortem matrices can be useful for detection, but quantitative results must be interpreted with caution.

Circumstances, Postmortem Findings, Blood Concentrations and Metabolism in a Series of Methoxyacetylfentanyl-Related Deaths.

Methoxyacetylfentanyl is one of many fentanyl analogs available as new psychoactive substances. It have been encountered in both the European Union and the United States, and existing literature suggest that methoxyacetylfentanyl is around 3- to 5-fold less potent than fentanyl. The aim of the present work was to combine case information with blood concentrations and abundance of urinary metabolites to investigate the importance of these parameters for toxicological interpretation. Quantification of methoxyacetylfentanyl in femoral blood was performed by LC--MS-MS and urinary metabolites were analyzed by LC--QTOF-MS with and without hydrolysis with β-glucuronidase/arylsulfatase. For confirmation of identified metabolites, methoxyacetylfentanyl was incubated with hepatocytes for up to 5 hours and analyzed with the same method as the urine samples. In eleven postmortem cases (27 to 41 years old and including one female) methoxyacetylfentanyl was reported in femoral blood. The cause of death was intoxication by methoxyacetylfentanyl alone or in combination with other drugs in all but one case, where death was attributed to acute complications of an underlying heart disease but with possible contribution from methoxyacetylfentanyl. In total, 27 urinary metabolites were found, including eight glucuronides. Major biotransformations were O-demethylation, dealkylation to form the nor-metabolite, mono- and dihydroxylations of the phenethyl moiety, as well as combinations thereof. The most abundant metabolites in hydrolyzed urine included O-desmethyl-, O-desmethyl-phenethyl-hydroxy-, O-desmethyl-phenethyl-hydroxymethoxy- and nor-methoxyacetylfentanyl. Differences in the abundance of methoxyacetylfentanyl and its major metabolites could be interpreted to indicate fatal intoxications in abstinent or chronic users. We postulate that urinary concentrations of methoxyacetylfentanyl and two metabolites, in combination with the methoxyacetylfentanyl concentration in femoral blood, might be good indicators of the time between administration and death as well as prior use.

11-Year Study of Fentanyl in Driving Under the Influence of Drugs Casework†.

Prior to 2017, heroin and other prescription opioids were the most prevalent opioids implicated in driving under the influence of drugs (DUID) investigation cases, and fentanyl was rarely included in the scope of toxicological analysis. Fentanyl has become the most frequently identified opioid in DUID cases, with many suspected heroin cases turning out to be only fentanyl. A review of fentanyl-positive DUID cases at NMS Labs was performed to provide prevalence information, change in concentration, patterns of combined drug use, indicators of impairment and driving behavior in order to assist with the toxicological interpretation of DUID scenarios involving fentanyl. Fentanyl-positive DUID cases received between January 2010 and December 2020 were examined. Blood results were confirmed and quantitated for fentanyl, norfentanyl and acetylfentanyl using a liquid chromatography--tandem mass spectrometry analysis with a limit of quantitation of 0.10, 0.20 and 0.10 ng/mL, respectively. Of 153,234 blood cases examined for DUID over 11 years, fentanyl was confirmed positive in 6,779 (4.4%) cases. However, there were significant changes in positivity over time. Fentanyl percentage positivity increased from 0.60% in 2010 to 12% in 2020. Of 5,976 confirmed fentanyl-positive cases in 2018-2020, blood concentrations >4.0 ng/mL were observed in 44% (2018), 55% (2019) and 59% (2020) of cases. Polypharmacy was common with 87% of blood samples confirmed positive for fentanyl and at least one other compound. Stimulant was the most commonly identified drug class in cases where at least one additional drug class was present. This study illustrates the importance of including fentanyl in a routine blood DUID panel.

Case analysis of kinetics investigations in toxicity studies of pesticides to identify the nonlinearity of internal exposure and the influences of nonlinearity on the toxicological interpretation of pesticide residue

The nonlinearity of internal exposure to 8 pesticides was investigated in toxicity studies using kinetics to identify nonlinearity visually and to investigate the influence of nonlinearity on toxicological evaluation. Data were obtained from risk assessment reports published by the Food Safety Commission (FSCJ). Nonlinearity was defined using 2 indicators: the lowest visual inflection point (LVIP) and the second lowest visual inflection point (SVIP) of kinetics by drawing a linear distribution chart. The area under the curve and 24-h urine concentrations were stable parameters used to identify the LVIP/SVIP. The sampling timing affected the blood concentrations, and the LVIP/SVIP was detected for 6 pesticides using the parent compounds or their metabolites as analytes. The subproportional nonlinearity was significant for these pesticides. The LVIP/SVIP values were consistent in the same species up to a 1-year period, but the values showed species-specific differences in several compounds. In all compounds found to be nonlinear, apical outcomes were observed at the SVIP or above. The presence of nonlinearity was recognized by the FSCJ. The recognition influenced their judgment of no-observed-adverse-effect levels (NOAELs) for carcinogenicity or health-based guidance values, indicating the importance of appropriate kinetics to identify the nonlinearity for toxicological evaluation of pesticide residue.

The European Human Biomonitoring Initiative (HBM4EU): Human biomonitoring guidance values for selected phthalates and a substitute plasticizer.

Ubiquitous use of plasticizers has led to a widespread internal exposure of the European population. Until today, metabolites are detected in almost every urine sample analysed. This raised the urgent need for a toxicological interpretation of the internal exposure levels. The European Human Biomonitoring Initiative (HBM4EU) contributes substantially to the knowledge on the actual exposure of European citizens to chemicals prioritised within HBM4EU, on their potential impact on health and on the interpretation of these data to improve policy making. On that account, human biomonitoring guidance values (HBM-GVs) are derived for the general population and the occupationally exposed population agreed at HBM4EU consortium level. These values can be used to assess phthalate exposure levels measured in HBM studies in a health risk assessment context. HBM-GVs were derived for five phthalates (DEHP, DnBP, DiBP, BBzP and DPHP) and for the non-phthalate substitute Hexamoll® DINCH. For the adult general population, the HBM-GVs for the specific metabolite(s) of the respective parent compounds in urine are the following: 0.5mg/L for the sum of 5-oxo-MEHP and 5-OH-MEHP; 0.19mg/L for MnBP, 0.23mg/L for MiBP; 3mg/L for MBzP; 0.5mg/L for the sum of oxo-MPHP and OH-MPHP and 4.5mg/L for the sum of OH-MINCH and cx-MINCH. The present paper further specifies HBM-GVs for children and for workers.

Cogent note on raised safety concern on concurrent uses of herbal formulations

Herbal therapy is well known holistic therapeutic approach from ancient times to improve emotional, mental and spiritual integrality because not having considerable side effects or adverse effects upon consumption and administration. Although medicinal plants or herbal formulations are widely proposed and considered to be safe, however, they can be potentially be hepatotoxic if not processed properly for final formulation stage to be used for consumption. Hence, simultaneous use of herbs with therapeutic drugs is considered or accepted these days which lead to increases the clinical potential of herb-drug interactions to treat many fatal diseases like hepatitis, cancer and various neurological disorders etc. However, these random or self-guided use of herbal medicine and supplements can be risky because most of times, they are not subject to review by the FDA. Hence, we need healthy attempts to find out possible reasons for studying streamlined clinical toxicological interpretations and clinical pharmacological approaches of herbal products before considering them for their further use as complementary and alternative medicine combination therapies to treat any diseases. Keywords: Herbal products, Herbal medicines, Herbal remedies, Medicinal plants.

Putrified tissue analysis: A challenge for Forensic toxicologists

The postmortem toxicology helps in determining the cause of death and information on events immediately before death. Forensic toxicologists receive various types of biological samples with putrefaction process which pose a greater challenge in chemical analysis. Interpretation of analytical toxicology results must incorporate pharmacokinetics and toxicology of the agents in question, the circumstances under which death occurred including the mechanism of exposure. The aim of present study is to describe the type of biological samples submitted for toxicological analysis in forensic toxicology section and factors associated in their analysis.

Impact of Sequencing Depth and Library Preparation on Toxicological Interpretation of RNA-Seq Data in a "Three-Sample" Scenario.

While RNA-sequencing (RNA-seq) has emerged as a standard approach in toxicogenomics, its full potential in gaining underlying toxicological mechanisms is still not clear when only three biological replicates are used. This "three-sample" study design is common in toxicological research, particularly in animal studies during preclinical drug development. Sequencing depth (the total number of reads in an experiment) and library preparation are critical to the resolution and integrity of RNA-seq data and biological interpretation. We used aflatoxin b1 (AFB1), a model toxicant, to investigate the effect of sequencing depth and library preparation in RNA-seq on toxicological interpretation in the "three-sample" scenario. We also compared different gene profiling platforms (RNA-seq, TempO-seq, microarray, and qPCR) using identical liver samples. Well-established mechanisms of AFB1 toxicity served as ground truth for our comparative analyses. We found that a minimum of 20 million reads was sufficient to elicit key toxicity functions and pathways underlying AFB1-induced liver toxicity using three replicates and that identification of differentially expressed genes was positively associated with sequencing depth to a certain extent. Further, our results showed that RNA-seq revealed toxicological insights from pathway enrichment with overall higher statistical power and overlap ratio, compared with TempO-seq and microarray. Moreover, library preparation using the same methods was important to reproducing the toxicological interpretation.

Aortic Strain -Threshold Analysis of Non-Cancer Outcome from Arsenic Exposure

Background: While arsenic has been shown in many studies to behave as a threshold carcinogen, analyses of non-cancer outcomes (e.g., cardiovascular) have generally been presented only using linear models. Alternative models have received little consideration. We choose here to examine aortic strain data, a known risk factor for hypertension and cardiovascular disease, on arsenic-exposed workers with alternative analytic modeling.Method: A previously published study (Karakulak et al., 2016) of aortic strain measurements in arsenic-exposed workers had demonstrated that aortic strain was markedly reduced among workers with high levels of arsenic exposure (urinary arsenic in ug/L). The original analysis was limited to a linear model without model validation. We have analyzed the data using alternative models.Results: Examination of the linear analysis and its residuals demonstrated significant heteroscedascity with unequal variances, particularly at urinary levels of &gt; 160 ug arsenic per liter (ug/L). The test of equal variances from samples with &gt;160 and samples &lt;= 160 demonstrated inequality (p &lt; 0.0001). The better alternative model was a step-function with a mean aortic strain of 11.3% for those with urinary arsenic &lt;= 160 ug/L and with a mean aortic strain of 5.3% for those with urinary arsenic &gt; 160 ug/L. A further stratified analysis with bins of 40 ug/L showed no effect on aortic strain for the urinary arsenic range of &lt;= 160 ug/L and, separately, no effect for the urinary arsenic range of &gt; 160 ug/L.Conclusion: Aortic elasticity parameters show an adverse effect from arsenic exposure only at urinary exposure levels of &gt; 160 ug/L and not below, indicating a step-function rather than a linear function. Restriction of analyses to a linear no-threshold model may have blinded the finding of a more useful demonstration of a dose-response pattern that may offer different toxicological interpretations.