Determination of methyl isopropyl hydantoin from rat erythrocytes by gas-chromatography mass-spectrometry to determine methyl isocyanate dose following inhalation exposure.

Methyl isocyanate (MIC) is an important precursor for industrial synthesis, but it is highly toxic. MIC causes irritation and damage to the eyes, respiratory tract, and skin. While current treatment is limited to supportive care and counteracting symptoms, promising countermeasures are being evaluated. Our work focuses on understanding the inhalation toxicity of MIC to develop effective therapeutic interventions. However, in-vivo inhalation exposure studies are limited by challenges in estimating the actual respiratory dose, due to animal-to-animal variability in breathing rate, depth, etc. Therefore, a method was developed to estimate the inhaled MIC dose based on analysis of an N-terminal valine hemoglobin adduct. The method features a simple sample preparation scheme, including rapid isolation of hemoglobin, hydrolysis of the hemoglobin adduct with immediate conversion to methyl isopropyl hydantoin (MIH), rapid liquid-liquid extraction, and gas-chromatography mass-spectrometry analysis. The method produced a limit of detection of 0.05 mg MIH/kg RBC precipitate with a dynamic range from 0.05-25 mg MIH/kg. The precision, as measured by percent relative standard deviation, was <8.5%, and the accuracy was within 8% of the nominal concentration. The method was used to evaluate a potential correlation between MIH and MIC internal dose and proved promising. If successful, this method may be used to quantify the true internal dose of MIC from inhalation studies to help determine the effectiveness of MIC therapeutics.

Cross-sectional study on N,N-dimethylformamide (DMF); effects on liver and alcohol intolerance.

PURPOSE: There are still concerns regarding occupational exposure to hepatotoxic DMF. This study was designed to evaluate possible liver damaging effects of DMF under current workplace conditions in synthetic fibres industries. METHODS: Among other laboratory parameters, liver function parameters (alkaline phosphatase (ALP), aspartate aminotransferase, alanine aminotransferase and gamma-glutamyltransferase), the mean corpuscular erythrocyte volume (MCV) and carbohydrate-deficient transferrin (CDT) of the workforce of two companies present at the days of study were investigated. Internal exposure to DMF was assessed via three different biomarkers [sum of N-methylformamide and N-hydroxymethyl-N-methylformamide, N-acetyl-S-(N-carbamoyl)cysteine (AMCC) and 3-methyl-5-isopropylhydantoin (MIH)]. Alcohol consumption was assessed by means of direct ethanol metabolites (ethylglucuronide and ethylsulfate). RESULTS: None of the tested liver enzyme activities showed a positive association with any of the three exposure markers, nor did CDT and MCV. CDT was negatively associated with AMCC and the ALP activity negatively with all three exposure markers. Changes in liver function are seen mainly in conjunction with ethanol consumption but also with increasing body weight and age. MCV was associated with smoking. Almost half of the workers stated to experience alcohol flush reaction. CONCLUSION: The present study indicates that long-term exposure to DMF, which was specified by median urinary AMCC levels of 4.84 mg/g creatinine and DMF haemoglobin adduct levels of 60.5 nmol/MIH/g globin, respectively, does not result in any adverse liver effects. In contrast, these DMF exposure levels still elicit certain alcohol intolerance reactions.

GC-NPD and GC-MS analysis of preserved tissue of Bhopal gas disaster: evidence of methyl carbamylation in post-mortem blood.

Twenty-five preserved autopsy blood samples of Bhopal toxic gas exposed victims were analysed by gas chromatography (GC) coupled with either Nitrogen-Phosphorous detector (NPD) or mass spectrometer (MS) for the presence of methyl carbamyl valine in terms of valine methyl hydantoin (VMH). 84% of these samples showed a positive test for VMH on GC-NPD and the identity of the peaks were further confirmed on GC-MS. The concentration of VMH in the gas-affected positive blood samples ranged from 2.56 to 51.28 nanomoles. These results indicate entry of methyl isocyanate (MIC), one of the constituents of the toxic cloud caused by the disaster, into the blood stream of victims who had inhaled gas.

Importancia de la dosificación plasmática de fármacos antiepilépticos en el tratamiento de la epilepsia: estudio de 65 pacientes / Significance of plasmic dosification of antiepileptic drugs in the management of epilepsia: study of 65 cases

Se presentan los resultados del primer trabajo cubano sobre dosificación de antiepilépticos en plasma y tratamiento de la epilepsia, de carácter interdisciplinario y en colaboración por 4 instituciones. Se estudiaron 65 adultos que padecían crisis epilépticas parciales, tonicoclónicas generalizadas o ambas, con más de 1 año, por lo menos, de observación en la Consulta Especial de Epilepsia. Se utilizó la cromatografía líquida de alta resolución para la dosificación plasmática de 3 drogas: defenilhidantoína (DFH) carbamazepina (CBZ) y fenobarbital (FB). El 58,4 % de los pacientes tenían un control total de sus crisis al momento de la determinación; el 32,3 un control parcial y el 9,2 no estaba controlado. La relación entre el control clínico y las cifras de antiepilépticos en plasma fue adecuada. Los pacientes no contolados tenían una concentración promedio más baja (entre 11,72 y 13,87 *g/mL) que los controlados (24,5 *g/mL para la DFH. Las cifras de DFH por debajo de 15 *g/mL se comportaron como subterapéuticas en nuestra serie. La concentración media de FB fue de 13,87 *g/mL y la de CBZ de 5,50. La DFH sola, la CBZ sola y la DFH asociada al FB fueron igualmente efectivas para el control de la crisis. Las dosis terapéuticas de DFH se encontraron en 4,8 mg por kg de peso corporal, y las de CBZ en 10,9. SE presentan casos demostrativos y se comentan los avances que han representado estas técnicas para el tratamiento de la epilepsia, por lo que se sugiere la introducción de las mismas en nuestro país al nivel provincial por la organización de salud cubana

Spiromustine analogues. Relationships between structure, plasma stability, and antitumor activity.

Spiromustine is a hydantoin-containing nitrogen mustard currently in Phase I clinical trial. Since the in vitro plasma half-life of this compound (6.4 min, 37 degrees C, pH 7.4) appeared to be influenced by the hydantoin ring, analogues containing 2-5 methylene spacer groups between this ring and the nitrogen mustard moiety were prepared and evaluated for hydrolytic stability and antitumor activity. Stability correlated with structure and pKa values. The proximity of the hydantoin ring to the mustard function was a stabilizing factor. Activity against murine P-388 leukemia was demonstrated and a gradual decrease in this activity was observed as the hydrolytic instability increased. A relationship between analogue structure and a mass spectral rearrangement ion was identified.

Direct enantiomeric resolution of mephenytoin and its N-demethylated metabolite in plasma and blood using chiral capillary gas chromatography.

A gas chromatographic method was developed for the determination of the R- and S- enantiomers of the anticonvulsant, mephenytoin, and its N-demethylated metabolite, 5-phenyl-5- ethylhydantoin ( PEH ), in plasma and blood. Direct enantiomeric separation of mephenytoin and its internal standard was obtained using a chiral capillary column ( Chirasil -Val) followed by nitrogen specific detection. However, resolution of the enantiomers of PEH and its internal standard required propylation at the 3-position of the hydantoin ring prior to analysis. Similar linear and reproducible standard curves were obtained from both plasma and blood over the concentration range 50 ng/ml to 5 micrograms/ml, and above 100 ng/ml the reproducibility was less than 8% (coefficient of variation). Pronounced stereoselective differences in the plasma concentration--time curves for both mephenytoin and PEH were observed in a normal subject who received a single oral dose of 300 mg racemic mephenytoin. The peak plasma level of S-mephenytoin was only one-fifth that of the R-enantiomer and its elimination half-life was less than 3 h compared to over 70 h for R-mephenytoin. Similarly, S- PEH levels were barely detectable whereas concentrations of R-metabolite steadily increased over 4-6 days before slowly declining.

Gas-chromatographic determination of mephenytoin and desmethylmephenytoin, after off-column alkylation.

We describe a gas-liquid chromatographic method for determining mephenytoin and its active metabolite, desmethylmephenytoin, in human serum. 5-Methyl-5-phenylhydantoin is used as the internal standard. The method involves extraction of the drugs by adsorption onto charcoal and off-column derivatization to their pentyl derivatives. Peak height and concentration are linearly related and the day-to-day CV for therapeutic concentration is about 2 to 6%. No interferences by endogenous compounds or drugs commonly used for seizure control have been encountered.

Initial studies on the penetration of spirohydantoin mustard into the cerebrospinal fluid of dogs.

A gas-chromatographic-mass-spectrometric method using selected ion monitoring has been developed to detect and quantitate levels of spirohydantoin mustard (SHM) as low as 1 ng per injected sample. The ions chosen for analysis of the drug were at m/e 154, 156, and 299. Using this procedure, the distribution of SHM in the blood and cerebrospinal fluid (CSF) of dogs following intravenous administration has been studied. The initial blood decay of SHM is rapid with a half-life of 1.78 and 1.85 min for 2 different dogs. SHM enters the CSF and, after reaching a peak value at approximately 10 min, exhibits a decay curve similar to that found in blood.