Determination of nitrite and nitrate in postmortem whole blood samples of 10 sodium nitrite poisoning cases: The importance of nitrate in determining nitrite poisoning.

Sodium nitrite (NaNO2), a widely used food preservative, has become a popular agent in South Korea for use in committing suicide since the mid-2010s because of its easy access. After ingesting sodium nitrite, nitrite ions oxidize hemoglobin to methemoglobin in red blood cells (RBCs), causing methemoglobinemia which can be fatal depending on the severity. As the number of deaths involving sodium nitrite has increased rapidly over the years, we developed a quantitative analysis method for nitrite and its oxidized form, nitrate, using ion chromatography (IC) with a conductivity detector. A simple ultrafiltration method was used for sample preparation because chloride ions which usually interfere with nitrite in most IC methods were completely separable using the developed analytical method. The limit of detection and lower limit of quantitation of nitrite were 0.5 and 1 mg/L, respectively. Nitrite and nitrate showed good linearity in the range of 1-500 mg/L and 5-500 mg/L, respectively. The established method was successfully applied to 10 authentic sodium nitrite poisoning cases, resulting in low nitrite concentrations (32.4 ± 29.5 mg/L in peripheral blood samples and 20.4 ± 18.7 mg/L in heart blood samples) and high nitrate concentrations (298.0 ± 25.6 mg/L in peripheral blood samples and 252.0 ± 41.3 mg/L in heart blood samples). The imbalance between nitrite and nitrate was due to the extensive conversion of nitrite to nitrate in postmortem bloods, which was confirmed by spiking nitrite into blank blood samples. In conclusion, not only the blood concentrations of nitrite but also those of nitrate should be quantified and considered for the determination of sodium nitrite poisoning, especially in postmortem blood samples.

Major component causing neurological toxicity in acute glufosinate ammonium poisoning: determination of glufosinate, 1-methoxy-2-propanol, and ammonia in serum and cerebrospinal fluid.

OBJECTIVE: To determine the primary contributor to neurotoxicity in patients with glufosinate ammonium (GLA) poisoning, by quantifying glufosinate, 1-methoxy-2-propanol, and ammonia in serum and the cerebrospinal fluid (CSF). MATERIALS AND METHODS: We collected and analysed data from confirmed cases of GLA poisoning between May 2018 and August 2020. Based on the occurrence of neurological complications (mental change, seizure, and central apnoea), patients were assigned to one of two groups: those with complications (NCx) and without (non-NCx) complications. Concentrations of glufosinate, 1-methoxy-2-propanol (1M2P), and ammonia were measured in the serum upon admission and during hospital stay. The concentrations of all these substances were again measured in the CSF following a decline in the mental status or seizure (NCx group) or on the day after hospitalisation (non-NCx group). RESULTS: Of the 20 patients included, ammonia levels in the serum and CSF at onset of altered sensorium in the NCx group (n = 16) were significantly higher than those at one day after hospitalisation in the non-NCx group (n = 4) (p = 0.011 in serum, p = 0.047 in CSF), with its concentration in the CSF being higher than that in the serum in 15/16 cases. The concentration of 1M2P was similar in the serum and CSF (8/16), but the concentrations of glufosinate (7/16) was lower in the CSF than in the serum. In the non-NCx group (n = 4), only ammonia was detectable. CONCLUSIONS: Among patients with GLA poisoning, increased CSF ammonia was significantly correlated with neurological complications.

A novel method for cyanide quantification in human whole blood using ion chromatography with amperometric detection and its application to cyanide intoxication cases.

Cyanide is a highly toxic agent that has been frequently used for suicide in South Korea. It is also used in various industrial fields, such as metal plating, in which many accidental cyanide intoxications have occurred. To overcome the disadvantages of conventional cyanide analysis methods, a simple and fast method for the analysis of cyanide in whole blood using ion chromatography (IC) with amperometric detection was developed in this study. Whole blood samples were deproteinized, diluted, and analyzed using an IC-amperometric detection system. The limits of detection and quantitation were 0.1 and 0.2 mg/L, respectively. The method showed good linearity in the range of 0.2 to 50 mg/L with R2  > 0.99. The intra- and inter-assay precision and accuracy values were <10%. The established method was successfully applied to analyze whole blood samples from three cyanide intoxication cases.

Maturation of Mitochondrially Targeted Prx V Involves a Second Cleavage by Mitochondrial Intermediate Peptidase That Is Sensitive to Inhibition by H2O2.

Prx V mRNA contains two in-frame AUG codons, producing a long (L-Prx V) and short form of Prx V (S-Prx V), and mouse L-Prx V is expressed as a precursor protein containing a 49-amino acid N-terminal mitochondria targeting sequence. Here, we show that the N-terminal 41-residue sequence of L-Prx V is cleaved by mitochondrial processing peptidase (MPP) in the mitochondrial matrix to produce an intermediate Prx V (I-Prx V) with a destabilizing phenylalanine at its N-terminus, and further, that the next 8-residue sequence is cleaved by mitochondrial intermediate peptidase (MIP) to convert I-Prx V to a stabilized mature form that is identical to S-Prx V. Further, we show that when mitochondrial H2O2 levels are increased in HeLa cells using rotenone, in several mouse tissues by deleting Prx III, and in the adrenal gland by deleting Srx or by exposing mice to immobilized stress, I-Prx V accumulates transiently and mature S-Prx V levels decrease in mitochondria over time. These findings support the view that MIP is inhibited by H2O2, resulting in the accumulation and subsequent degradation of I-Prx V, identifying a role for redox mediated regulation of Prx V proteolytic maturation and expression in mitochondria.

Monitoring urinary testosterone and epitestosterone levels, and their ratio, in Korean chemical castration subjects using liquid chromatography-tandem mass spectrometry.

In Europe, chemical castration has been adopted as a treatment for paraphilia since the 1930s. Among the various chemical castration agents, luteinizing hormone-releasing hormone (LHRH) agonists are now used widely because of their effectiveness and safety. In South Korea, a legislation of chemical castration to control the sexual impulses of sexual offenders was enforced in July 2011. Most of these subjects are treated with leuprorelin acetate, an LHRH agonist, for chemical castration. Despite this, there are few studies that address the long-term influence of LHRH agonists on testosterone (T) and epitestosterone (E) levels in chemical castration subjects. In order to analyze the urinary levels of T in chemical castration subjects, whose T levels are extremely low, we developed and validated an analytical method for the detection of both T and E in human urine using a liquid chromatography-tandem mass spectrometry (LC-MS/MS) system. The urine samples were hydrolyzed, extracted, and analyzed by LC-MS/MS with electrospray ionization in the positive-ion mode. The limits of detection were 0.02 ng/mL and the limits of quantitation were 0.05 ng/mL, which provided great sensitivity. The established method was applied to urine samples from chemical castration subjects and healthy male volunteers. The chemical castration subjects showed significantly lower urinary T levels than the control subjects. In addition, the urinary E levels were also lower in the chemical castration subjects; however, the T/E ratios were constant and did not show a notable decrease because of the simultaneous decrease in both urinary T and E. The urinary T levels and T/E ratio did not exceed the doping control criteria for exogenous T ingestion for any subject. This study shows the trend of urinary T and E levels in long-term treated chemical castration subjects by establishing a highly sensitive LC-MS/MS method, that provides useful information for monitoring chemical castration.

Simultaneous determination of synthetic cannabinoids and their metabolites in human hair using LC-MS/MS and application to human hair.

Hair is one of the key samples for judging drug abuse in the field of forensic science. However, few studies have examined synthetic cannabinoids and their metabolites in human hair. Synthetic cannabinoids are a class of chemicals that bind to cannabinoid receptors, but they differ structurally from the cannabinoids found in cannabis. They have been sold sprayed on dried, shredded plant material under brand names such as "Spice" since the 2000s. In South Korea, synthetic cannabinoids have been widely distributed since 2009 and many types detected up to now. Unlike traditional drugs such as methamphetamine and cannabis, the abuse trends of synthetic cannabinoids were variable by regions and changed according to the times. If new types of synthetic cannabinoids become popular which has been altered in some structures, it becomes difficult to identify using exist analytical method. Therefore, it is important to develop a new analytical method for synthetic cannabinoids currently being abused in society. In this study, we developed simultaneous analytical methods for the detection of 18 synthetic cannabinoids and 41 of their metabolites in authentic human hair samples using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Selectivity, linearity, limits of detection (LODs), limits of quantification (LOQs), precision, accuracy, matrix effect, recovery, and process efficiency were evaluated, and all results were acceptable. Additionally, the distribution of synthetic cannabinoids in the head hair of Korean drug abusers from 2016 to 2018 was investigated. Hair samples from 43 individuals suspected of synthetic cannabinoid use were provided by law enforcement agencies. The drugs detected most prevalently in the head hair of Korean drug abusers were AB-CHMINACA and JWH-210.

Determination of boldenone in postmortem specimens including blood and urine samples using LC-MS/MS.

Boldenone (BOLD), one of androgenic anabolic steroids (AAS), although banned in humans, is still available illegally. AAS abuse has previously been associated with various cardiovascular adverse events including acute myocardial infarction, arrhythmia, and sudden death. In this study, the concentration of BOLD was determined in postmortem specimens from the corpse of a human male who intentionally injected BOLD undecylenate into his shoulder muscle. In addition, the endogenous levels of BOLD in the blood and urine samples of young human males have been reported. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method with solid-phase extraction (SPE) was developed and validated for the analysis of BOLD in blood, muscular tissue and urine samples. The validation parameters including linearity, accuracy, precision, matrix effect, and recovery were satisfactory. The concentrations of BOLD in the blood of 20 young human males who didn't take BOLD were under the limit of quantitation (LOQ, 0.5 ng/mL). Additionally, the mean level of BOLD in the urine samples was 3.19 ± 1.65 ng/mL (range: 0.37˜6.02 ng/mL). The concentrations of BOLD in the victim's blood from the femoral vein and heart were 140.44 and 25.74 ng/mL, respectively. On the other hand, those in the muscular tissue from the injection site and the urine sample were 142.3 ng/g and 3474 ng/mL, respectively.

Detection of 11-nor-9-carboxy-tetrahydrocannabinol in the hair of drug abusers by LC-MS/MS analysis.

Cannabis is the second most commonly abused illicit drug after methamphetamine in South Korea. To prove cannabis consumption, 11-nor-9-carboxy-tetrahydrocannabinol (THC-COOH), the metabolite of tetrahydrocannabinol (THC), was screened for in a hair analysis. In this study, we developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis method, which was used to analyze authentic hair samples in 2017. Possible contaminants on the surface of hair samples were eliminated by washing twice each with 2mL of methanol and distilled water. After adding an internal standard (THC-COOH-d3), the hair samples (about 20mg each) were digested with 1M NaOH, extracted twice with mixed organic solvents (n-hexane:ethyl acetate), and analyzed by an LC-MS/MS system. Identification and quantification of THC-COOH and THC-COOH-d3 were performed using a multiple reaction monitoring (MRM) mode at m/z 245 and 191 and m/z 248, respectively (quantifier ions are underlined). The following validation parameters were evaluated: selectivity, linearity, limit of detection (LOD), limit of quantification (LOQ), precision, accuracy, matrix effect, and recovery. The LOD and LOQ of the method was 0.1pg/mg. Good linearity was achieved for THC-COOH in the range from 0.1 to 20pg/mg. The method showed an acceptable precision and accuracy, both of which were less than 15% at the three concentrations of THC-COOH (0.2, 1, and 10pg/mg). THC-COOH showed ion suppression at these three concentrations. The concentrations of THC-COOH in the authentic hair samples ranged from 0.10 to 27.30pg/mg (total 586 cases), and its concentrations were classified as low, medium, or high ranges, i.e., 0.10-0.39pg/mg, 0.39-1.99pg/mg, or 1.99-27.30pg/mg, respectively, according to statistical evaluation. This method showed the possibility of replacing the existing gas chromatography-tandem mass spectrometry (GC-MS/MS) analysis. However, further development of our LC-MS/MS method is necessary in order to meet the recommended 0.05pg/mg cut-off.

Determination of AB-CHMINACA and its metabolites in human hair and their deposition in hair of abusers.

Despite global efforts to control the abuse of synthetic cannabinoids, the high-level of turnover from the market impedes regulation, endangering public health. N-[(1S)-1-(aminocarbonyl)-2-methylpropyl]-1-(cyclohexylmethyl)-1H-indazole-3-carboxamide (AB-CHMINACA) is the most popular synthetic cannabinoid in South Korea since its introduction in 2014. Nonetheless, few studies have been carried out on AB-CHMINACA and its metabolites, and its deposition in human hair. The purpose of this study was to develop and validate an analytical method for detection of AB-CHMINACA and its six metabolites in hair using a liquid chromatography tandem mass spectrometry (LC-MS/MS) system, for forensic applications. The methanol extracts of hair samples were evaporated, filtered, and analyzed by LC-MS/MS with electrospray ionization in positive ion mode. The limits of detection and quantification ranged from 0.5 to 10pg/mg and 2 to 50pg/mg, respectively. Good linearity was achieved within the range of 5-1000pg/mg or 10-1000pg/mg depending on the analyte. Intra- and inter-assay precision and accuracy values were below 15%. No significant variation was observed using different sources of hair matrices. These validation results proved the selectivity, accuracy and reproducibility of the method. The established method was applied to 37 authentic samples from suspected synthetic cannabinoid users. AB-CHMINACA and its two metabolites, AB-CHMINACA M2 and AB-CHMINACA M4, were detected. The concentration of the parent drug was much higher than those of its metabolites, and the amount of AB-CHMINACA M2 was greater than that of AB-CHMINACA M4 in all samples. No other metabolites were detected in the samples.

An LC-MS/MS method for the simultaneous determination of 15 antipsychotics and two metabolites in hair and its application to rat hair.

In recent years, the inappropriate use of antipsychotics by young Korean men has become a social problem. As military service exemptions are given for mental illness, some men pose as mental health patients to avoid military service. In order to verify the authenticity of mental illnesses, we developed simultaneous analytical methods for the detection of 15 antipsychotics and 2 of their metabolites in hair using liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. The target drugs were modafinil, atomoxetine, aripiprazole, benztropine, buspirone, duloxetine, gabapentin, oxcarbazepine, topiramate, escitalopram, paliperidone, ziprasidone, lamotrigine, clonazepam, levetiracetam, and metabolites of oxcarbazepine and clonazepam. To remove possible contaminants on the hair surface, hair samples were washed twice with methanol and distilled water, and then were extracted with methanol overnight at 38°C. Desipramine-d3 was used as an internal standard. LC-MS/MS analysis was performed on an Agilent 1290 Infinity UHPLC coupled to an AB Sciex Qtrap® 5500 MS/MS. The total chromatographic run time was 14min. The following validation parameters were evaluated: selectivity, linearity, limit of detection (LOD), limit of quantification (LOQ), precision, accuracy, matrix effect, and recovery. The LOD and LOQ values for all analytes, except modafinil, ranged from 0.2 to 10pg/mg hair and from 0.2 to 20pg/mg hair, respectively. Good linearity was achieved for most of the analytes in the range of 20-200pg/mg hair. The method showed acceptable precision and accuracy, which were less than 15%, as well as satisfactory matrix effects and recoveries. Furthermore, this method was also applied to the analysis of rat hair samples. The study in rats showed that the concentrations of atomoxetine and aripiprazole in pigmented hair were significantly higher than those in non-pigmented hair. However, no significant difference was observed in the concentration of topiramate between pigmented and non-pigmented hair. This method will be useful in monitoring the inappropriate use of antipsychotics in suspects posing as mental health patients. However, further research is necessary before applying this method to authentic hair samples from mental health patients.