Beyond Ketamine and Phencyclidine: Analytically Confirmed Use of Multiple Novel Arylcyclohexylamines.

BACKGROUND: Methoxetamine and 3-methoxy-phencyclidine are novel arylcyclohexylamines whose use and clinical toxicity are poorly reported in the medical literature. We report a case of analytically confirmed use of both methoxetamine and 3-methoxy-phencyclidine. CASE REPORT: A 27-year-old male presented 10 hours after insufflating an Internet-obtained powder. He was hypertensive, tachycardic, and demonstrated dissociated affect, a delayed verbal response to questions, ataxia, and vertical nystagmus. A urine drug screen was positive for phencyclidine and 11-nor-delta9-THC-9-carboxylic acid. He was admitted and his mental status and blood pressure normalized eight hours later. Blood samples (0, 2, and 3 hours from arrival) and the powders were analyzed by liquid chromatography-quadrupole time-of-flight mass spectrometry. Methoxetamine and 3-methoxy-phencyclidine were detected in all samples (279 ng/ml, 205 ng/ml, and 180 ng/ml for methoxetamine; 167 ng/mL, 131 ng/mL, and 90 ng/ml for 3-methoxy-phencyclidine at 0, 2, and 3 hours, respectively). No phencyclidine or tetrahydrocannabinol was detected. Two powders contained methoxetamine while one contained 3-methoxy-phencyclidine. CONCLUSION: The literature regarding methoxetamine and 3-methoxy-phencyclidine toxicity is limited. Methoxetamine use is associated with altered mental status, ataxia, and hypertension. Toxicity from 3-methoxy-phencyclidine is poorly described. There is no prior case describing serial qualitative analysis. Health care providers should be aware of the novel arylcyclohexylamines and their toxicity.

An Impaired Driver Found to be Under the Influence of Methoxetamine.

The popularity of designer drugs has increased over the past few years as users seek new, cheap and sometimes "legal" ways to get high. This case report focuses on a case that happened in the City of Henderson, Nevada, which involved the designer drug methoxetamine. Methoxetamine is a psychoactive compound that is structurally related to ketamine and reported to have similar effects. These effects include analgesia, cardiovascular and respiratory stimulation, and enhanced skeletal muscle tone. Presented here is a case of a 33-year-old female who was pulled over after almost colliding with a marked police motorcycle, causing the police officer to avoid the collision by running onto a pedestrian sidewalk. Upon stopping the vehicle and questioning the passengers, the officer learned that the driver of the vehicle had ingested methoxetamine earlier in the day. After the driver was taken into custody, a blood sample was drawn and sent to the laboratory for analysis. Initial screening of the blood sample showed presumptive positive results for the amphetamine enzyme-linked immunosorbent assay. The next day, a full scan screen of the blood sample was performed using the gas chromatography-mass spectrometry (GC/MS) and methoxetamine and dextromethorphan were detected. Since the laboratory did not have the ability to confirm methoxetamine, the sample was sent to NMS Labs for analysis. The results from NMS Labs showed a methoxetamine concentration of 160 ng/mL. To date, this is the first DUI case in the state of Nevada where methoxetamine was detected and confirmed. A short time after the NMS results were received, a full SWGTOX validation was performed on a new GC/MS method to confirm methoxetamine along with five synthetic cathinone analytes. After the GC/MS analysis validation was complete, the sample was subsequently reanalyzed for methoxetamine in the toxicology laboratory at the Henderson Police Department Forensic Science Laboratory and the result that attained was 151 ng/mL, which was in line with the result from NMS Labs.

Fatal intoxication with methoxetamine.

Methoxetamine (MXE) is a new synthetic drug of abuse structurally related to ketamine and phencyclidine. A case of a 29-year-old male with acute toxicity related to the analytically confirmed use of MXE is reported. The man was found dead at his residence. Biological material was analyzed using liquid chromatography-tandem mass spectrometry. The concentration of MXE in urine of the deceased was 85 µg/mL. Despite the vial containing the blood sample being destroyed during transportation and the blood leaking out into the cardboard packaging, the blood level of MXE was estimated. After determination of the cardboard grammage (approx. 400 g/m(3) ) and the mean mass of the blood obtained after drying (0.1785 ± 0.0173 g per 1 mL), the estimated blood concentration of MXE was found to be 5.8 µg/mL. The high concentration of MXE in blood and urine and the circumstances of the case indicate an unintentional, fatal intoxication with this substance.

Analytical findings of an acute intoxication after inhalation of methoxetamine.

Methoxetamine (MXE) is increasingly used and abused, as it is frequently presented as being safer than ketamine, and legal. Cases of only MXE consumption being associated with the occurrence of seizures are rarely reported. A single MXE intoxication case by inhalation is described concerning a 21-year-old man, not known to be epileptic, who was found collapsed in his bedroom, supposedly after an epileptic seizure. He was transferred to the Emergency Department of the Henri Mondor Hospital, Aurillac, France. He was conscious, but with a sinus bradycardia (48/min) and an ST-segment elevation on the electrocardiogram, and a slightly increased creatine kinase level (270 U/L) and hyponatremia (127 mmol/L). New seizure activity occurred during hospitalization, but the clinical course in the intensive care unit was favorable. Quantitation of MXE in serum and urine using gas chromatography coupled to mass spectrometry (GC-MS) was developed, as well as a liquid chromatography coupled to tandem mass spectrometry (LC-MS-MS) method for the determination of MXE in hair. Limits of detection and quantification were, respectively, 2 and 10 µg/L for the GC-MS method and both 0.5 pg/mg for the LC-MS-MS method. Concentrations of 30 and 408 µg/L were, respectively, measured in serum and urine. Concentrations of 135 and 145 pg/mg were detected in two 2.5 cm hair strands, consistent with one or several consumptions during the 2 ½ months prior to sampling. A sample of the powder consumed was available and also analyzed. This case illustrates the dangers of this drug, which justify its classification as a narcotic in France since August 2013.

A polydrug intoxication involving methoxetamine in a drugs and driving case.

Methoxetamine ((RS)2-(3-methoxyphenyl)-2-(ethylamino)cyclohexanone)) is becoming a drug of interest among practitioners of forensic toxicology. In this case report, we describe the case background, standard field sobriety tests, sampling, and analysis of this drug in a whole blood sample as well as screening methods and analysis from a driver operating under the influence of intoxicating substances. Methoxetamine was isolated from the blood sample using mixed mode solid phase extraction. After elution and evaporation, the residue was dissolved in mobile phase (consisting of acetonitrile and aqueous formic acid) for analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and gas chromatography-mass spectrometry (GC-MS). The case sample was found to contain clonazepam, 7-aminoclonazepam, carboxy-THC, Ddphenhydramine, and MDMA. The case sample was found to contain 10 ng/mL of the drug (methoxetamine) in whole blood. The results of this drug analysis and previous analyses are discussed in terms of this driver operating under the influence of drugs.

[Acute methoxetamine intoxication--a case report with serum and urine concentrations]. / Ostre zatrucie metoksetamina--opis przypadku z wyznaczonymi stezeniami w surowicy i moczu.

Methoxetamine (MXE) is a novel synthetic drug, structurally related to phencyclidine, with ketamine-like properties. Available in Poland since 2010, with no legal control, is adverti. sed as the "ideal dissociation drug". The aim of this study was to present a case of nasal methoxetamine acute poisoning in a 28-year-old man, the course of treatment, and the method of identification of this substance in serum and urine. In the course of this intoxication extreme agitation and aggression with slight response to benzodiazepines were observed. The patient was confused, hallucinated. In addition, the physical examination re. vealed tachycardia 120/min and normal blood pressure (130/80 mm Hg). The period of acute poisoning was covered by amnesia. The MXE concentrations in serum and urine were determined using liquid chromatography-mass spectrometry (LC-MS-MS) method, and were respectively 270 ng/ml and 660 ng/ml. Confirmed MXE poisoning increases our knowledge about this new substance, providing relevant clinical and analytical data.

An accidental fatal intoxication with methoxetamine.

This paper reports an unintentional death involving the administration of methoxetamine [2-(3-methoxyphenyl)-2-(ethylamino)-cyclohexanone] and offers some reference values from living drug abusers. Methoxetamine is a new recreational drug with a similar structure to ketamine. The deceased was a 26-year-old male with a history of drug abuse; he was found lying on the floor in his apartment. Several "red-line" plastic bags were found, one of which was labeled "2-(3-methoxyphenyl)-2-(ethylamino)-cyclohexanone" and another labeled "Haze." In four cases from living subjects with unknown doses, concentrations of methoxetamine were found from 0.13 to 0.49 µg/g. In three of the cases, the blood samples also contained natural or synthetic cannabinoids. In the autopsy case, a considerably higher concentration of methoxetamine, 8.6 µg/g, was found in femoral blood. In addition, tetrahydrocannabinol and the three different synthetic cannabinoids AM-694, AM-2201, and JWH-018, were present in femoral blood. The circumstances and the high femoral blood concentration of methoxetamine point toward an unintentional, acute fatal intoxication with methoxetamine, although the presence of the three synthetic cannabinoids may have contributed to the death.

Acute toxicity associated with the recreational use of the ketamine derivative methoxetamine.

PURPOSE: Long-term regular use of ketamine has been reported to be associated with severe symptomatic urinary tract problems. Methoxetamine, an arylcyclohexylamine derivative of ketamine, is marketed as a "bladder safe" derivative of ketamine, and no cases of acute toxicity following analytically confirmed methoxetamine use have been reported to date. We report here a case series of three individuals with acute toxicity related to the analytically confirmed use of methoxetamine. CASE SERIES: Three patients aged between 28 and 42 years presented to the Emergency Department (ED) on unrelated occasions having used methoxetamine. Clinical features were suggestive of a "dissociative/catatonic" state similar to that seen with ketamine; in addition, they had clinical features of acute sympathomimetic toxicity with significant tachycardia and hypertension. All were managed with low-dose benzodiazepines and discharged home once their symptoms/signs had settled. TOXICOLOGICAL SCREENING: Serum collected at the time of presentation to the ED was analysed qualitatively and quantitatively by gas chromatography-mass spectrometry. Serum concentrations ranged from 0.09 to 0.2 mg/L; in addition, detectable levels of 6-APB/5-APB were found in one of the patients. CONCLUSIONS: These three analytically confirmed cases demonstrate that acute methoxetamine-related toxicity is associated with both "dissociative" and "sympathomimetic" clinical features. The information from these three cases is useful to clinical pharmacologists, not only in managing individuals with acute methoxetamine toxicity but also in advising the appropriate legislative authorities on the risk of acute harm related to methoxetamine use. Further work is needed to determine whether methoxetamine is more "bladder friendly" than ketamine, as has been suggested by those marketing methoxetamine.

Hyperforin inhibits cancer invasion and metastasis.

Hyperforin (Hyp), the major lipophilic constituent of St. John's wort, was assayed as a stable dicyclohexylammonium salt (Hyp-DCHA) for cytotoxicity and inhibition of matrix proteinases, tumor invasion, and metastasis. Hyp-DCHA triggered apoptosis-associated cytotoxic effect in both murine (C-26, B16-LU8, and TRAMP-C1) and human (HT-1080 and SK-N-BE) tumor cells; its effect varied, with B16-LU8, HT-1080, and C-26 the most sensitive (IC50 = 5 to 8 micromol/L). At these concentrations, a marked and progressive decline of growth was observed in HT-1080 cells, whereas untransformed endothelial cells were only marginally affected. Hyp-DCHA inhibited in a dose-dependent and noncompetitive manner various proteinases instrumental to extracellular matrix degradation; the activity of leukocyte elastase was inhibited the most (IC50 = 3 micromol/L), followed by cathepsin G and urokinase-type plasminogen activator, whereas that of the matrix metalloproteinases (MMPs) 2 and 9 showed an IC50 > 100 micromol/L. Nevertheless, inhibition of extracellular signal-regulated kinase 1/2 constitutive activity and reduction of MMP-2 and MMP-9 secretion was triggered by 0.5 micromol/L Hyp-DCHA to various degrees in different cell lines, the most in C-26. Inhibition of C-26 and HT-1080 cell chemoinvasion (80 and 54%, respectively) through reconstituted basement membrane was observed at these doses. Finally, in mice that received i.v. injections of C-26 or B16-LU8 cells, daily i.p. administration of Hyp-DCHA-without reaching tumor-cytotoxic blood levels-remarkably reduced inflammatory infiltration, neovascularization, lung weight (-48%), and size of experimental metastases with C-26 (-38%) and number of lung metastases with B16-LU8 (-22%), with preservation of apparently healthy and active behavior. These observations qualify Hyp-DCHA as an interesting lead compound to prevent and contrast cancer spread and metastatic growth.

The metabolism of cyclamate to cyclohexylamine in humans during long-term administration.

A group of 14 subjects, who had been identified from 261 volunteers in a 1-week screen as being able to metabolize the sweetener cyclamate to cyclohexylamine (>0.2% of a daily dose), and 31 nonconverters (<0.2% metabolism) were given calcium cyclamate tablets (equivalent to 250 mg cyclamic acid, 3 times daily) for a period of 13 weeks. The metabolism of cyclamate to cyclohexylamine was determined using twice-weekly timed (3 h) urine collections during week 1-3 and 7-13. Urine specimens were collected on all other study days to investigate day-to-day fluctuations in cyclohexylamine excretion. Analyses of the twice weekly timed urine collections showed that subjects recruited as nonconverters essentially remained nonconverters. Of the converters, three showed consistently low metabolism, five showed erratic metabolism, five showed low metabolism initially, which increased during the latter part of the study, and one subject showed consistently high metabolism throughout the study. Analysis of the day-to-day urine specimens showed marked intrasubject variability. The plasma concentrations of cyclohexylamine measured on weeks 1-3 and 7-13 reflected the urine profiles. The highest individual long-term average steady-state excretion values based on the 3-h urine collections and daily samples were 21%, 23%, 25%, 29%, 34%, and 38%. The maximum % metabolism detected in the high converters occasionally reached the value of 60% reported in previous short-term studies, but this high activity was not maintained, and was followed by periods of lower metabolism. The results of this metabolism study support an acceptable daily intake (ADI) of 0-11 mg/kg body weight per day.

Biodistribution and renal excretion of isomers of the cationic tracer, (99m)Tc diaminocyclohexane (DACH): biodistribution of cationic renal tracers.

The buildup of organic anions in the plasma in the uremic state can competitively inhibit the tubular extraction of para-aminohippurate or (131)I ortho-iodohippurate (OIH) and lead to spuriously low measurements of effective renal plasma flow (ERPF). This problem can be circumvented by the use of cationic tracers. The cationic renal tracer, (99m)Tc labeled diaminocyclohexane ((99m)Tc DACH), has a clearance of 80% of OIH in mice but its clearance in humans is relatively low, only 30% of OIH. The (99m)Tc DACH isomer(s) used in prior studies, however, was not clearly defined and may have consisted of a single isomer or a combination of isomers. Since the anionic isomers of some (99m)Tc renal tracers have been shown to have widely different clearances, the biodistribution and urine excretion of the (99m)Tc cis-, trans-S,S, trans-R,R and +/-trans-DACH isomers were compared in Sprague-Dawley rats at 10 minutes and 60 minutes postinjection to determine if one of the (99m)Tc DACH isomers may be a significantly better renal tracer than the others. The red cell binding of (99m)Tc +/- trans-DACH was also determined. All of the isomers showed a high degree of specificity for the kidney with minimal secretion into the gastrointestinal tract. Urine excretion of the 4 tracers, however, was only 38-48% that of OIH at 10 minutes and 66-84% that of OIH at 60 minutes. Red cell binding was 6.9%. Cationic renal tracers have the potential to provide a more accurate measurement of ERPF than anionic tracers. Based on the animal data, however, it is unlikely that any of the (99m)Tc DACH isomers will have a substantially higher clearance in humans than the form of (99m)Tc DACH originally tested. Development of alternative cationic renal tracers is warranted.

Enantiospecific determination of PNU-83894 and its major metabolite, PNU-83892, in plasma, and its application to the characterization of the enantioselective pharmacokinetics of PNU-83894 in the dog.

A chiral method for the simultaneous analysis of the (+)- and (-)-enantiomers of PNU-83894 and its metabolite, PNU-83892, in plasma was developed to characterize the enantioselective pharmacokinetics of PNU-83894, a potential anticonvulsant candidate. The method involves solid-phase extraction (phenyl column) of the enantiomers from plasma followed by direct enantioselective separation on a beta-cyclodextrin HPLC chiral column and UV detection at 230 nm. The linear range for this method was found to be 12.5 ng/ml to 5.00 microg/ml and the intra- and inter-assay precision and accuracy for each enantiomer were <11% in all cases. The validity of this assay was also demonstrated by its application to the pharmacokinetic evaluation of PNU-83894 in the dog.

Long-term toxicity and carcinogenicity study of cyclamate in nonhuman primates.

Twenty-one monkeys (cynomolgus, rhesus, African green) were fed cyclamate (100 mg/kg and 500 mg/kg) in the diet five times per week from a few days after birth and continuing for up to 24 years. Malignant tumors were diagnosed in three 24-year-old cyclamate monkeys; these were metastatic colon carcinoma (rhesus; 500 mg/kg), metastatic hepatocellular carcinoma (cynomolgus; 500 mg/kg), and a small, well differentiated adenocarcinoma of the prostate (cynomolgus; 100 mg/kg). Benign tumors were found at necropsy in three females; these were adenoma of the thyroid gland (rhesus; 100 mg/kg) and two cases of leiomyoma of the uterus (rhesus; 100 mg/kg and 500 mg/kg). No tumors were detected in an age-matched control group of 16 monkeys. Examination of the testes revealed complete testicular atrophy in one of the old cyclamate monkeys, and focal germ cell aplasia (Sertoli-only tubules) in two other cyclamate monkeys. Focal spermatogenic interruption (maturation arrest) at various germ cell levels mixed with normal spermatogenesis was observed in both the cyclamate-treated and the control monkeys, all of which were over 20 years old. Measurements of terminal cyclohexylamine concentrations showed that three of the males dosed with cyclamate at 500 mg/kg were high converters, with plasma concentrations comparable to the levels that produce testicular atrophy in rats. However, only one of the three high converters showed histologic evidence of irregular spermatogenesis. The overall conclusion is that the testicular abnormalities and the sporadic cases of different malignancies found after more than 20 years of dosing do not provide clear evidence of a toxic or carcinogenic effect of sodium cyclamate in monkeys.

Test chamber exposure of humans to 1,6-hexamethylene diisocyanate and isophorone diisocyanate.

An isocyanate generation apparatus was developed and stable isocyanate atmospheres were obtained. At a concentration of 5 micrograms 1,6-hexamethylene diisocyanate (HDI) per m3 the precision was found to be 7% (n = 5). Three volunteers were each exposed to three different concentrations of HDI (11.9, 20.5, and 22.1 micrograms/m3) and three concentrations of isophorone diisocyanate (IPDI) (12.1, 17.7, and 50.7 micrograms/m3), in an exposure chamber. The duration of the exposure was 2 h. Urine and blood samples were collected, and hydrolysed under alkaline conditions to the HDI and IPDI corresponding amines, 1,6-hexamethylene diamine (HDA) and isophorone diamine (IPDA), determined as their pentafluoropropionic anhydride amides (HDA-PFPA and IPDA-PFPA). The HDA- and IPDA-PFPA derivatives were analysed using liquid chromatography mass spectrometry with thermospray monitoring negative ions. When working up samples from the exposed persons without hydrolysis, no HDA or IPDA was seen. The average urinary excretion of the corresponding amine was 39% for HDI and 27% for IPDI. An association between the estimated inhaled dose and the total excreted amount was seen. The average urinary elimination half-time for HDA was 2.5 h and for IPDA, 2.8 h. The hydrolysis condition giving the highest yield of HDA and IPDA in urine was found to be hydrolysis with 3 M sodium hydroxide during 4 h. No HDA or IPDA could be found in hydrolysed plasma (< ca 0.1 micrograms/l).

A pharmacokinetic study of phenylcyclohexyldiethylamine. An analog of phencyclidine.

The pharmacokinetics of three phencyclidine analogs--phenylcyclohexyl-diethylamine (PCDE), phenylcyclohexylethylamine (PCE), and phenylcyclohexylamine (PCA)--were determined in rats after intravenous administration of each drug. Because PCE and PCA are major metabolites of PCDE, their plasma levels were also measured after administration of PCDE. Similarly, PCA concentrations was determined after administration of PCE. The data were combined and analyzed by nonlinear regression procedures using compartmental and noncompartmental models to determine the kinetic parameters of PCDE metabolism. The object was to estimate the kinetic constants for the metabolic sequence, PCDE to PCE to PCA. A 6-compartment model (two pools for each analyte) that included saturable components for the conversion of PCDE to PCE and PCE to PCA gave the best fit to the combined data. Despite large uncertainties for some microparameters, useful estimates were obtained for clearances, distribution volumes, and fraction of PCDE or PCE converted to PCE and PCA in vivo under nonsaturating conditions. The estimated fraction of PCDE converted to PCA and the apparent Km value for the conversion of PCDE to PCE were comparable to values obtained in vitro with microsomal preparations, suggesting that metabolic studies in vitro provide reasonable predictors of the biotransformation process in vivo for this class of compounds.

Blood pressure changes and sympathetic function in rats given cyclohexylamine by intravenous infusion.

Intravenous infusion of cyclohexylamine (30-120 mg/kg) caused a dose-dependent increase in blood pressure in urethane-anesthetized rats. The increase in blood pressure was inversely related to the duration of the infusion. The blood pressure returned to baseline values (+/- 5 mm Hg) within 60 min of the end of the infusion after doses of 30 and 60 mg/kg were administered over 20 and 40 min. The plasma concentrations of cyclohexylamine were related linearly to the administered dose and decreased only 27% between the end of the infusion and 60 min later. The concentration-effect relationship showed clockwise hysteresis, indicative of tachyphylaxis, as has been reported in humans (Eichelbaum et al., 1974). Administration of bolus doses of tyramine at the end of the infusion or 60 min later demonstrated the presence of an indirect sympathomimetic response, although this was attenuated to a greater extent by high doses and more rapid infusions of cyclohexylamine. An almost complete loss of response to tyramine was found only in animals given 120 mg/kg over 40 min. The presence of a tyramine response 60 min after the infusion of 60 mg/kg occurred when there was an essentially complete reversal of the hypertensive effect of cyclohexylamine. These data indicate that the hypertensive effect of the indirectly acting sympathomimetic amine cyclohexylamine occurs primarily during rapid increases in plasma concentrations. Tachyphylaxis develops rapidly after the cessation of the infusion which is probably due largely to reuptake of released noradrenaline at low doses and depletion of releasable noradrenaline at high doses.

The metabolism of cyclamate to cyclohexylamine and its cardiovascular consequences in human volunteers.

A group of 194 diabetic patients were given calcium cyclamate (1 g/day as cyclamic acid equivalents) for a period of 7 days. Blood and urine samples were collected to determine the formation of cyclohexylamine, which is an indirectly acting sympathomimetic amine. Blood pressure and heart rate were recorded before and after treatment. Urine samples were collected each day and analyzed for cyclamate (to check compliance) and cyclohexylamine (to monitor the development of metabolizing activity). After 7 days intake most individuals (78%) did not excrete significant amounts of cyclohexylamine (less than 0.1% of the daily dose of cyclamate) but a small number (8; 4% of the group) excreted more than 20% of the daily dose as cyclohexylamine in the urine. Similar interindividual variations were found in the plasma concentrations of cyclohexylamine after 7 days intake of cyclamate, with 8 individuals having concentrations of 300-1942 ng/ml. The changes in cardiovascular parameters in these 8 subjects between pre- and postdosing were similar to those found in 150 subjects with plasma cyclohexylamine concentrations less than 10 ng/ml. Twenty of the subjects were restudied after receiving calcium cyclamate for 2 weeks at a daily dose equivalent to 2 g of cyclamic acid (0.66 g tds). Plasma concentrations of cyclohexylamine, heart rate, and blood pressure were measured every 30 min for a period of 8 hr (one dose interval) after the final dose. Twelve patients had plasma concentrations of cyclohexylamine greater than 10 ng/ml (89-2043 ng/ml) at the start of the dose-interval investigations. There were no transient increases or decreases in plasma concentrations of cyclohexylamine which might have resulted in a transient change in blood pressure or heart rate. These data indicate that the metabolism of cyclamate (2 g/day) to cyclohexylamine would not affect blood pressure or heart rate even in individuals with high metabolizing ability.

Liquid membrane work-up of blood plasma samples applied to gas chromatographic determination of aliphatic amines.

A technique for sample work-up and enrichment using a supported liquid membrane in an automated flow system, connected to a gas chromatograph, was used for the determination of aliphatic amines in human blood plasma. The amines studied were N,N-dimethylethylamine, triethylamine, N-methylmorpholine, cyclohexylamine and N,N-dimethylcyclohexylamine. An efficient clean-up of the complex plasma matrix was achieved, resulting in identical blank chromatograms for plasma samples and aqueous solutions. Different parameters influencing the efficiency and selectivity of the extraction procedure were experimentally studied and theoretically explained. The detection limit depends on the extraction flow-rate and the available sample volume. With 1 ml of sample and a flow-rate giving an extraction time of 16 min, the detection limit was ca. 5 ppb (5 micrograms/l); with 4 ml of sample and a lower flow-rate, sub-ppb detection limits could be reached in ca. 3 h. Linear calibration curves up to 500 ppb were obtained. Blood plasma samples from volunteers exposed to N,N-dimethylethylamine in air were analysed, and the results compared favourably with independent measurements by another method.

Quantitative liquid chromatographic determination of bromadoline and its N-demethylated metabolites in blood, plasma, serum, and urine samples.

Bromadoline and its two N-demethylated metabolites were extracted into ether:butyl chloride after the addition of internal standard and basification of the various biological fluids (blood, plasma, serum, and urine). These compounds were then extracted into dilute phosphoric acid from the organic phase and separated on a reversed-phase chromatographic system using a mobile phase containing acetonitrile and a buffer of 1,4-dimethylpiperazine and perchloric acid. The overall absolute extraction recoveries of these compounds were approximately 50-80%. The background interferences from the biological fluids were negligible and allowed quantitative determination of bromadoline and the metabolites at levels as low as 2-5 ng/mL. At mobile phase flow rate of 1 mL/min, the sample components and the internal standard were eluted at the retention times within approximately 7-12 min. The drug- and metabolite-to-internal standard peak height ratios showed excellent linear relationships with their corresponding concentrations. The analytical method showed satisfactory within- and between-run assay precision and accuracy, and has been utilized in the simultaneous determination of bromadoline and its two N-demethylated metabolites in biological fluids collected from humans and from dogs after administration of bromadoline maleate.