Background - Alcohol is the most abused substance in Western society, resulting in major economic losses and negative health consequences. Therefore, there is a need for a selective and robust detection method for alcohol consumption in various clinical and forensic settings. This study aimed to validate a mass spectrometry method for quantifying phosphatidylethanol (PEth) and perform retrospective data analysis from the patient population of a national reference laboratory.

In postmortem forensic investigation cases where the bladder is voided or dehydrated prior to autopsy, it is possible to wash the bladder with saline and collect the 'bladder wash' and any residual urine for toxicological analysis. While not conventional, this study aims to determine the use of bladder washes as alternative specimens in postmortem forensic toxicology. Comprehensive drug and alcohol analysis was performed on blood, urine, vitreous humor and bladder wash samples. Control studies consisted of matched bladder wash and urine samples for comparison. Authentic applicability studies were performed on bladder wash samples in cases where only blood or no urine samples were available. Bladder wash testing via the routine urine methodology were shown to have the appropriate sensitivity and specificity to serve as an alternative specimen. Specificity of the applicability studies was further improved when comparisons were corrected by evaluating individual analytes jointly with their related parent drug or metabolites. Individual and corrected sensitivity and specificity rates of above 99% were typically observed in both comparisons against urine and blood paired samples. Following drug analysis of 31 cases in which only a bladder wash was available, 57 detections from 23 different analytes were detected that otherwise would have not been obtained. This study demonstrates that standardized collection of the easily accessible bladder wash for postmortem toxicological analysis serves forensic toxicologists and pathologists with invaluable information where urine or other biological specimens are not available.

The long-term stability of drug concentrations in human plasma samples, when stored under normal laboratory conditions over several years, is important for research purposes and clinical re-evaluation, and forensic toxicology. Fifty human plasma samples from a former clinical trial were re-analyzed after storage at -20°C for 11 years. Plasma samples were extracted using solid-phase extraction. Isotope labeled sufentanil-d5 was used as internal standard. Sufentanil plasma concentrations were determined by ultra-performance liquid chromatography with gradient elution, followed by tandem mass spectrometry with electrospray ionization. The linear dynamic range was 25-2500 pg/mL, the limit of detection was 10 pg/mL, and the lower limit of quantification was 25 pg/mL. Intra- and inter-assay error did not exceed 6%. The deviation of the measured sufentanil plasma concentrations between the re-analysis and the first analysis was -63 ± 14% (mean ± SD). Therefore, sufentanil concentrations in human plasma were not stable in samples frozen at -20°C over 11 years.

There is a growing interest for quantification of drugs in capillary blood. Phosphatidylethanol (PEth) is a biomarker for alcohol intake measured in whole blood, thus making it a candidate for capillary sampling. Our laboratory has been running a method for PEth quantification in venous blood since 2016 and we aimed to expand this method to also include capillary dried blood spot (DBS) samples. Two 10 µL volumetric absorptive microsampling (VAMS) devices, Capitainer®B Vanadate and Mitra® were included in the method development and validated. Calibrators and quality controls were spiked during the automatic sample extraction without the VAMS devices present, making it possible to extract and analyze both types of VAMS samples in the same set-up. With the Mitra device all pre-established validation criteria were fulfilled in the measuring range 0.03-4.0 µM (21-2812 ng/mL), including method comparison with our venous blood method. Capitainer fulfilled all validation criteria, except for the accuracy of samples with PEth levels ≥ 0.5 µM (≥ 352 ng/mL) (deviation -17.1 to -20.5%). The correlation analysis between Capitainer and the venous blood results showed no constant bias, but an acceptable small proportional mean difference of -7.6%. Overall, the method validation results for both Capitainer and Mitra were considered acceptable. Both devices were found suitable for the analyses of PEth.

Natural toxins present an ongoing risk for human exposure that requires a rapid and accurate diagnosis for proper response. In this study, a qualitative liquid chromatography-high-resolution mass spectrometry (LC-HRMS) method was developed and validated for the detection of a large, diverse selection of natural toxins. Data-dependent acquisition was performed to identify compounds with an in-house mass spectral library of 129 hazardous toxins that originate from plants, animals, and fungi. All 129 compounds were spiked into human urine, extracted, and evaluated for spectral library matching. Of these, 92 toxins met the quality criteria and underwent validation in urine matrix based on American National Standards Institute guidelines. A generalized workflow for method expansion was developed and enables the rapid addition of relevant compounds to the established method. This LC-HRMS method achieves efficient detection of natural toxins in urine, and the created workflow can rapidly increase compound coverage via method expansion.

A blind quality control (BQC) program in blood alcohol analysis was implemented at the Houston Forensic Science Center (HFSC) in September 2015. By mimicking authentic toxicology blood evidence, the laboratory can perform a concurrent evaluation of their technical and administrative casework procedures and test the accuracy and reliability of their volatile analysis method in a format that is blinded to the analyst. From September 2015 to November 2023, HFSC's Quality Division submitted 1228 antemortem whole-blood samples: 292 ethanol-negative samples and 936 ethanol-positive samples at 16 target concentrations (0.051, 0.080, 0.100, 0.110, 0.120, 0.130, 0.150, 0.160, 0.170, 0.180, 0.190, 0.200, 0.230, 0.240, 0.250, and 0.260 g/dL). A second, unopened blood tube in 168 of the 1228 BQCs was also analyzed after 721-1140 days: 24 ethanol-negative samples and 144 ethanol-positive samples at 5 target concentrations (0.080, 0.100, 0.130, 0.180, and 0.240 g/dL). All 316 ethanol-negative samples remained negative. After 42-758 days, the average (median, range) change in ethanol concentration of the 936 positive samples was -1.4% (-1.3%, -12.0% to +8.4%) with a statistically significant difference (P < .001) observed for the gradual decline in blood alcohol concentration (BAC) over time. The average BAC percentage differences per target concentration, ranged from -6.4% (-0.008 g/dL) to +5.7% (+0.011 g/dL), were within HFSC's current measurement uncertainty (9.4% at k = 3), showing no apparent correlation between the change in ethanol and the theoretical target concentration. As the analysis time between the two blood specimens from the same evidence kit extended, the loss in ethanol significantly increased (P < .001).

Opioids are widely prescribed pain medications that have the potential for misuse and abuse. As part of a routine procedure, our laboratory frequently encounters questions from clients/Medical Review Officers (MROs) regarding opioid hair concentrations in relation to the amount of opioid taken as part of a prescription. In this manuscript, we have analyzed a large number of real-world examples of opioid hair concentrations following self-reported consumption of an opioid prescription regimen. This dataset provides a reference point of opioid hair concentrations after an extensive aqueous wash that likely correspond to consumption of an opioid prescription regimen. Practitioners in the field could use this reference to make decisions on the opioid concentration of a hair sample in relation to a client-provided prescription.

Identification of N,N-dimethylpentylone (DMP) in counterfeit "Ecstasy" and "Molly" tablets poses risk to public health due to its adverse effects. Little information is available regarding the pharmacological activity or relevant blood or tissue concentrations of DMP, and even less is known about other structurally related beta-keto methylenedioxyamphetamine analogues on recreational drug markets, such as N-propyl butylone. Here, a novel toxicological assay utilizing liquid chromatography-tandem quadrupole mass spectrometry (LC-QQQ-MS) was developed and validated for the quantitation of DMP and five related synthetic cathinones (eutylone, pentylone, N-ethyl pentylone (NEP), N-propyl butylone, and N-cyclohexyl butylone), with chromatographic resolution from isomeric variants and quantitation performed by standard addition. A forensic series of 125 cases is presented for DMP and related analogs, along with pharmacological activity assessments using monoamine transporter and mouse behavioral assays. The blood concentration range for DMP in postmortem forensic cases was 3.3-4,600 ng/mL (mean: 320±570 ng/mL, median: 150 ng/mL), whereas pentylone, the primary N-desmethyl metabolite of DMP, was identified in 98% of cases with a concentration range 1.3-710 ng/mL (mean±SD: 105±120 ng/mL, median: 71 ng/mL). N-Propyl butylone, a newly identified synthetic cathinone, was quantitated in seven cases (mean±SD: 82±75 ng/mL, median: 50 ng/mL, range: 1.7-200 ng/mL). DMP displayed potent uptake inhibition at the dopamine transporter (IC50=49 nM), with 100-fold weaker potency at the serotonin transporter (IC50=4990 nM). DMP was a locomotor stimulant in mice (ED50=3.5 mg/kg) exhibiting potency relatively similar to eutylone, N-ethyl pentylone, and pentylone. Our results show that DMP is a psychomotor stimulant associated with adverse clinical outcomes leading to death. Forensic laboratories must continue to update testing methods to capture emerging drugs, with specific emphasis on resolution and identification of isomeric species. Following the scheduling of DMP in early-2024, there could be an anticipated market shift towards a new unregulated synthetic stimulant to replace DMP.

Liquid chromatography-mass spectrometry (LC-MS) methods for detection of multiple drugs of abuse (DoA) in oral fluid (OF) samples are being implemented in many clinical routine laboratories. Therefore, there is a need to develop new multianalyte methods with simple sample pretreatment and short analysis times. The purpose of this work was to validate a method detecting 58 DoA to be used with two different OF sampling kits, the saliva collection system (SCS) from Greiner Bio-One and Quantisal from Immunalysis, using the same sample pretreatment and analytical method. A set of 110 samples collected with the SCS kit was further compared to an high-resolution mass spectrometry (LC-HRMS) method in another laboratory. The method was successfully validated, with precision and accuracy of ≤15% and z-scores of <2 for external controls. Using a sensitive LC-MS-MS instrument, the detection limits were <1 µg/l in neat oral fluid. In the comparative study between the LC-MS-MS and LC-HRMS methods using SCS samples, a good agreement was observed. Discrepancies were limited to lower concentration ranges, attributable to differences in cut-off thresholds between the methods. This work contributes to the development of LC-MS multianalyte methods for OF samples, which are suitable for clinical routine laboratories.

The prevalence of mitragynine (kratom) in forensic toxicology casework has steadily increased over time. Readily available and currently legal, mitragynine is widely used for its stimulant and, depending on concentration, sedative effects. Our laboratory analyzed various fluid and tissue specimens from 51 postmortem cases to investigate the distribution of mitragynine and its active metabolite 7-hydroxymitragynine. Central and peripheral blood concentrations were compared, with an average heart blood to femoral blood ratio being 1.37 for mitragynine and 1.08 for 7-hydroxymitragynine. This ratio >1.0 suggests that mitragynine has some propensity toward postmortem redistribution; however, the difference in concentrations of mitragynine and 7-hydroxymitragynine is not statistically significant. Large average mitragynine to 7-hydroxymitragynine ratios of 30.9 in femoral blood and 32.4 in heart blood were observed compared to average ratios of 14.8 in vitreous humor and 16.9 in urine. In addition, the stability of these two compounds was investigated in both matrix and organic solvent. When stored refrigerated (4°C), mitragynine was stable for up to 30 days and 7-hydroxymitragynine was stable for up to 7 days with an analyte loss of <20%. Following 60 days of refrigerated storage, 7-hydroxymitragynine concentrations dropped over 50% from initial concentrations. Methanolic preparations of mitragynine and 7-hydroxymitragynine were stable following 3 months of storage at -20°C.

Ongoing legalization of cannabis for recreational use contributes to increasing numbers not only of incidents of driving under the influence, but within all forensic fields. In addition, newly emerging cannabinoids such as hexahydrocannabinol (HHC) and the increasing use of cannabidiol (CBD) products have to be addressed. The aims of this study were first to extend laboratory analysis capacity for the "established" cannabinoid ∆9-tetrahydrocannabinol (THC) and its metabolites 11-OH-THC and THC-COOH in human plasma/blood, and second to develop analytical procedures concerning HHC and CBD. An LC-MS/MS method based on the available (low-end) instrumentation was used. Samples (250 µL) were prepared by protein precipitation and solid phase extraction. Chromatographic separation was achieved on a reversed-phase C18 column within 15 min. Detection was performed on a 3200 QTRAP instrument (Sciex) in positive multiple reaction monitoring (MRM) mode. Matrix matched six-point calibrations were generated applying deuterated internal standards for all analytes except HHC. The method was fully validated according to GTFCh guidelines. Linear ranges were 0.5-25 µg/L for THC, 11-OH-THC, HHC and CBD, and 2.0-100 µg/L for THC-COOH, respectively. Limits of detection and limits of quantification were 0.5 and 1.0 µg/L (THC, 11-OH-THC, HHC, CBD), and 2.0 and 4.0 µg/L (THC-COOH). Applicability of plasma calibrations to blood samples was demonstrated. Acceptance criteria for intra- and inter-day accuracy, precision, extraction efficiency and matrix effects were met. No interfering signals were detected for more than 60 pharmaceutical compounds. The presented method is sensitive, specific, easy to handle and does not require high-end equipment. Since its implementation and accreditation according to ISO 17025, the method has proven to be fit for purpose not only in DUID cases but also within post-mortem samples. Furthermore, the design of the method allows for an uncomplicated extension to further cannabinoids if required.

Carbon monoxide (CO) is a common gaseous toxin that causes severe poisoning symptoms. Accurate detection of the formation of carboxyhemoglobin (COHb) in the blood is very important for the identification of CO poisoning. In this review, the effects of exogenous toxins, including dichloromethane (DCM), nitrite and hydrogen sulfide, on the determination of COHb by spectrophotometry were summarized by comparing epidemiological data, case studies and analytical methods. The mechanism of the effects of these exogenous poisons on COHb detection is described, and the extent of their influence on the clinical diagnosis and forensic identification of CO poisoning is discussed. We suggest that emergency medicine and forensic science practices need to improve the understanding of these toxins, and optimize clinical diagnosis and evaluation strategies to address the effects of toxins on the determination of COHb.

Dexamphetamine, lisdexamphetamine, and methylphenidate are central stimulant drugs widely used to treat Attention-deficit/hyperactivity disorder (ADHD), but poor adherence may lead to treatment failure and the drugs are also subject to misuse and diversion. Drug analysis in oral fluid may thus be useful for monitoring adherence and misuse. We measured drug concentrations in oral fluid and urine after controlled dosing to investigate detection windows and evaluate the chosen cut-offs. Healthy volunteers ingested single oral doses of 10 mg dexamphetamine (n=11), 30 mg lisdexamphetamine (n=11), or 20 mg methylphenidate (n=10), after which they collected parallel oral fluid and urine samples every 8 hours for 4-6 days. Amphetamine (analytical cut-off, oral fluid: 1.5 ng/mL; urine: 50 ng/mL), methylphenidate (oral fluid: 0.06 ng/mL), and ritalinic acid (urine: 500 ng/mL) were analyzed using fully validated chromatographic methods. The median time from ingestion to the last detection in oral fluid was 67 ± 4.9 (lisdexamphetamine) and 69 ± 8.8 (dexamphetamine) hours for amphetamine and 36 ± 2.5 hours for methylphenidate. This was comparable to urine (77 ± 5.1 hours for lisdexamphetamine, 78 ± 4.5 hours for dexamphetamine, 41 ± 2.4 hours for ritalinic acid). The inter-individual variability in detection times was large, probably in part due to pH-dependent disposition. Using a logistic regression approach, we found similar detection rates as a function of time since intake in urine and oral fluid with the chosen cut-offs, with a high degree of probability for detection at least 24 hours after intake of a low therapeutic dose. This demonstrates the usefulness of oral fluid as a test matrix to assess adherence to ADHD medications, provided the analytical method is sensitive, requiring a cut-off as low as 0.1 ng/mL for methylphenidate. Detection windows similar to those in urine may be achieved for amphetamine and methylphenidate in oral fluid.

The problem of finding a suitable biomarker to widen the detection window of γ-hydroxybutyric acid (GHB) intake remains a challenge in forensic toxicology. Based on previously published results, the present study deals with the evaluation of a fatty acid ester of GHB (4-palmitoyloxy butyrate (GHB-Pal)) in whole blood as a potential biomarker to extend the detection window of GHB use e.g. in drug-facilitated sexual assaults (DFSA). A liquid chromatography-mass spectrometry (LC-MS/MS) method for the quantification of GHB-Pal in whole blood was validated. Whole blood samples were collected from subjects involed in police roadside controls (n=113) and from narcolepsy patients (n=10) after the controlled administration of Xyrem® (sodium oxybate). Both sample collectives were previously tested for GHB using two different methods: ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and gas chromatography-mass spectrometry (GC-MS). In samples from routine police casework, GHB-Pal was detected in 67 out of 113 analysed GHB-positive samples with a mean concentration of 0.8 ng/mL ± 0.5 ng/mL (standard deviation). Among samples that were tested positive for both compounds, no linear correlation was observed between GHB and GHB-Pal concentrations (r=0.508). In contrast, GHB-Pal was not detected in any of the blood samples analysed from the patients. The absence of GHB and GHB-Pal in the patient cohort may be attributed to the time interval between dose intake and blood collection (approx. 3 and 6 h), during which GHB was eliminated from the body. Furthermore, GHB-Pal was only detectable at a GHB concentration of at least 16 µg/mL, which indicates that endogenous concentrations or low GHB doses may not be sufficient for GHB-Pal formation. Due to missing correlation between both compounds and the lack of GHB-Pal detection several hours after GHB administration, it can be assumed that GHB-Pal in blood is not a suitable biomarker to widen the detection window of GHB.

Novel psychoactive substances (NPS) have historically been difficult compounds to analyze in forensic toxicology. The identification, detection and quantitation of these analytes and their metabolites has been difficult due to their rapid emergence, short life span and various potencies. Advancements in analytical instrumentation are fundamental to mitigating these NPS challenges by providing reliable identification and sensitivity. This review discusses the pros and cons of various analytical instruments that have played a pivotal role in NPS analysis. As analytical technology advanced, the ability to analyze for NPS became easier with high resolution mass spectrometry; however, traditional immunoassays are still beneficial for some NPS classes such as benzodiazepines. Over 200 articles from 2010-2023 were reviewed, and 180 were utilized for this review. Journal articles were categorized according to the technology used during analysis: immunoassay, gas chromatography mass spectrometry, liquid chromatography mass spectrometry-low resolution, and liquid chromatography mass spectrometry-high resolution to allow for quick references based on a laboratory's technologies. Journal articles were organized in table format to outline the authors, NPS drug classes, and instrumentation used, among other important information.

The synthetic cathinone (SC) 3,4-methylenedioxy-α-pyrrolidinohexanophenone (MDPHP), is structurally correlated to the 3,4-methylenedioxypyrovalerone (MDPV). In recent years, the number of intoxication cases has increased even if little is known about the pharmacokinetics properties. The post-mortem (PM) distribution of MDPHP remains largely unexplored. In these reports, MDPHP levels were quantified in blood, gastric content and urine. This study aimed to describe the MDPHP PM distribution in several specimens, i.e. central and peripheral blood (CB and PB), right and left vitreous humor (rVH and lVH), gastric content (GCo), urine (U) and hair. The samples were collected from a cocaine-addicted 30-year-old man with a PM interval estimated in 3-4 h. Autopsy examination revealed unspecific findings, i.e. cerebral and pulmonary edema. No injection marks were observed. Toxicological analyses were performed using a multi-analytical approach: headspace gas chromatography for blood alcohol content (BAC); gas chromatography-mass spectrometry (GC-MS) for the main drugs of abuse; liquid chromatography-tandem mass spectrometry (LC-MS/MS) for benzodiazepines and new psychoactive substances (NPS). BAC was negative (0.02 g/L). MDPHP concentrations were: 1,639.99 ng/mL, CB; 1,601.90 ng/mL, PB; 12,954.13 ng/mL, U; 3,028.54 ng/mL, GCo; 1,846.45 ng/mL, rVH; 2,568.01 ng/mL, lVH; 152.38 (0.0-1.5 cm) and 451.33 (1.5-3.0 cm) ng/mg, hair. Moreover, hair segments were also positive for 3,4-dimethylmethcathinone (DMMC < limit of quantification: 0.01 ng/mg), α-PHP (0.59 ng/mg, 0.0-1.5 cm; 3.07 ng/mg, 1.5-3.0 cm), cocaine (6.58 ng/mg, 0.0-1.5 cm; 22.82 ng/mg, 1.5-3.0 cm), and benzoylecgonine (1.13 ng/mg, 0.0-1.5 cm; 4.30 ng/mg, 1.5-3.0 cm). MDPHP concentrations were significantly higher than those reported in the literature for fatal cases. For these reasons, the cause of death was probably the consumption of a lethal amount of MDPHP. Because CB and PB were similar, PM redistribution was not relevant.

Recently, tetrahydrocannabinol (THC) isomers and other semi-synthetic cannabinoids have been introduced into the consumer market as alternatives to botanical cannabis. To assess the prevalence of these potential new analytical targets, a liquid chromatography-tandem mass spectrometry confirmation method was developed for the quantitation of seven cannabinoid metabolites and the qualitative identification of four others in urine. The validated method was applied to authentic urine specimens that screened positive by immunoassay (50 ng/mL cutoff; n=1300). The most commonly observed analytes were 11-nor-9-carboxy-Δ8- and Δ9-THC (Δ8- and Δ9-THCCOOH), with the combination of the two seen as the most prominent analyte combination found. In addition to these metabolites, Δ1°-THCCOOH was observed in 77 specimens. This is the first study to report Δ1°-THCCOOH in authentic urine specimens, with this analyte always appearing in combination with Δ9-THCCOOH. Cross-reactivity studies were performed for (6aR,9R)-Δ1°-THCCOOH using the Beckman Coulter Emit® II Plus Cannabinoid immunoassay and demonstrated cross reactivity equivalent to the Δ9-THCCOOH cutoff, providing added confidence in the reported prevalence and detection patterns. Additionally, 11-nor-9(R)-carboxy-hexahydrocannabinol (9(R)-HHCCOOH) was the most abundant stereoisomer (n=12) in specimens containing HHC metabolites alone (n=14). This is in contrast to 9(S)-HHCCOOH, which was the predominant stereoisomer in specimens containing Δ8- and/or Δ9-THCCOOH. Although HHC and Δ1°-THC metabolites are emerging toxicology findings, based on these specimens collected between April 2022 and May 2024, an analytical panel containing Δ8- and Δ9-THCCOOH appears to be sufficient for revealing cannabinoid exposure within workplace monitoring and deterrence programs.

Since the opioid epidemic was declared in 2017, postmortem fentanyl cases and the need for interpretation of their results have increased. Postmortem redistribution is one of the factors to consider when interpreting cases. There have been several previous studies regarding fentanyl postmortem redistribution; however, these studies either have small sample sizes or were conducted prior to the declaration of the opioid epidemic which may cause conflicting results and not be reflective of current trends. This study includes fentanyl central/peripheral blood ratios from 748 cases from both Harris County, TX and Orange County, TX spanning from January 2009 to June 2022. Because the data set was determined to be non-normally distributed, a Kruskal-Wallis test was used for statistical comparisons. There were statistically significant differences between epidemic cases from the Harris County Institute of Forensic Sciences and the Orange County Crime Laboratory, central/peripheral ratios from pre-epidemic and epidemic years, and in cases where medically-related administration of fentanyl was documented when compared to cases where there was no documentation of licit fentanyl use. Various factors that could impact postmortem redistribution were evaluated (age, gender, polydrug use, etc.) and no clear trend or observation was made from the data. Based on the results of this study, there is still no clear indication as to what caused the increase in central/peripheral ratios, but it may be related to an increase in illicit fentanyl use.

Novel psychoactive substances (NPS) continue to emerge in the marketplace and are often found as substances in traditional illicit drug materials, and users are often unaware of the presence of other drugs. The proper identification and confirmation of the exposure to a drug is made possible when a biological specimen is collected and tested. Sweat is an alternative biological matrix of great interest for clinical, and forensic analysis. One of the reasons is attributed to its expanded drug detection window, enabling a greater monitoring capacity, and provision of information on prospective drug use. However, the concentrations of drugs in sweat samples are often low, which requires highly sensitive and selective methods. Disposable pipette tips extraction (DPX) is a new miniaturized solid phase extraction technique capable of efficiently extracting analytes from biological specimens, providing high recoveries, and requiring minimized solvent use. This study describes the development and optimization of two methods for the extraction of basic and neutral psychoactive substances from sweat samples using GC-MS and Design of Experiments (DoE). The following extraction parameters were optimized by DoE techniques: sample volume, elution solvent volume, washing solvent volume, sample aspiration time, elution solvent aspiration time, and number of cycles performed, including the elution step. It was possible to design a simple extraction protocol that provided optimized recoveries for both basic and neutral compounds. The sum of analyte areas increased at a rate of 54.7% for compounds of basic character and 39.2% for compounds of neutral character. Therefore, our results were satisfactory, demonstrating that DPX can be successfully used for extracting the target drugs from sweat samples.

Diphenhydramine has been available for decades in non-prescription formulations for the treatment of allergic reactions, insomnia and symptomology associated with colds. In addition, dimenhydrinate, a precursor to diphenhydramine, is available in preparations for the treatment of nausea and vomiting. Diphenhydramine and other first-generation antihistamines are being replaced by second- and third-generation antihistamines which are associated with fewer side effects, notably the lack of drowsiness; however, there are still a variety of therapeutic uses that have persisted in both adults and children. In this study, postmortem blood concentrations of diphenhydramine were determined, by liquid chromatography tandem mass spectrometry, in seven children with concentrations ranging from 0.051 to 2.6 mg/L. The cause of death in two cases was attributed, at least in part, to diphenhydramine toxicity while diphenhydramine detection in five cases was considered incidental to the cause of death.