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.

Albumin adducts and urinary metabolites resulting from occupational exposure to 1,5-naphthalene diisocyanate.

OBJECTIVES: 1,5-Naphthalene diisocyanate (NDI) is used in the plastic industry as a curing agent. 1,5-Naphthalene diisocyanate is classified as a sensitizing agent. The objective of this study has been to develop biomonitoring methods for the evaluation of exposure to NDI. MATERIAL AND METHODS: We obtained blood and urine samples from a group of 20 male workers exposed to NDI. The workers answered a questionnaire about their exposure history, job description, the number of years with the company and the time spent working with NDI over the 10 days of the study. Total plasma, albumin, and urine were analyzed for the presence of 1,5-naphthalenediamine (NDA) after acid hydrolysis using gas chromatography-mass spectrometry (GC-MS). RESULTS: 1,5-Naphthalenediamine was found in about 60% of the samples obtained from the workers. 1,5-Naphthalenediamine was obtained after acid hydrolysis of plasma, albumin, and urine at levels up to 1.5 pmol NDA/mg of plasma proteins, 1.15 pmol NDA/mg of albumin, and 55.3 pmol NDA/ml of urine, respectively. CONCLUSIONS: 1,5-Naphthalenediamine found in urine correlates best with the plasma levels (r = 0.91, p < 0.01). The albumin-adduct levels did not correlate with the NDI-specific immunoglobulin E (IgE) or total IgE present in the workers. The adduct and metabolite levels correlate with the air levels of NDI. Int J Occup Med Environ Health 2017;30(4):579-591.

The lessons of Bhopal [toxic] MIC gas disaster scope for expanding global biomonitoring and environmental specimen banking.

Bhopal Toxic gas tragedy represents one of the worst chemical accidents of the world. Autopsy and toxicological studies, apart from presenting evidence of acute and even chronic cyanide toxicity, provided a unique example of the incriminated chemical being traced to the bodies of the victims. The entry of methyl isocyanate (MIC) into the blood stream was established by the presence of carbamoylated end-terminal amino acids of haemoglobin and other tissue proteins. The presence of MIC trimer and a few other identified as well as unidentified tank residue constituents in the blood and viscera further established a close nexus of the products of pyrolysis of MIC in the aerosol inhaled by the victims. The Bhopal studies exemplify the scope for biological monitoring (BM) and environmental specimen banking (ESB) in chemical accidents as part of the global efforts.

Albumin adducts, hemoglobin adducts and urinary metabolites in workers exposed to 4,4'-methylenediphenyl diisocyanate.

4,4'-Methylenediphenyl diisocyanate (MDI) is the most widely used isocyanate in the manufacture of polyurethanes. MDI has been implicated as one of the major causes of occupational asthma. Hydrolysis of MDI can yield 4,4'-methylenedianiline (MDA), which is a suspected human carcinogen. Thus the need to monitor occupational exposure to MDI is of great significance. The use of air monitors alone has been found to be insufficient and there is a need for sensitive markers of recent and long-term exposure. We obtained biological samples from a group of 20 workers exposed to MDI vapor during the manufacture of polyurethane products. The air levels of MDI in the factory were measured using personal, work room and work station monitors. In most cases the levels were below detection limits. The blood and urine samples were analyzed for the presence of adducts and metabolites using GC-MS methods. Urinary base-extractable metabolites were found above control levels in 15 of the 20 workers and ranged from 0.035 to 0.83 pmol MDA/ml. The level of the acetylated metabolite N'-acetyl-4,4'-methylenedianiline (AcMDA) ranged from 0.13 to 7.61 pmol/ml. The amount of MDA released after acid hydrolysis was on average 6.5 times higher than the amount of free MDA and AcMDA present in urine. MDA was detected as a hemoglobin (Hb) adduct in all of the 20 subjects. The level ranged from 70 to 710 fmol/g Hb. In one individual the Hb adduct of AcMDA was detected. This is the first time a Hb adduct of AcMDA has been detected after occupational exposure to MDI. This is a further piece of evidence for the biological availability of the suspected human carcinogen MDA from in vivo hydrolysis of MDI. Plasma albumin conjugates of MDI can cause the onset of respiratory disorders in both man and animal models. Thus we investigated the presence of plasma protein adducts. The plasma MDA levels ranged from 0.25 to 5.4 pmol/ml. Up to 120 fmol/mg were found to be covalently bound to albumin.

Biomonitoring of workers exposed to 4,4'-methylenedianiline or 4,4'-methylenediphenyl diisocyanate.

4,4'-Methylenedianiline (MDA) and 4,4'-methylenediphenyl diisocyanate (MDI) are important intermediates in the production of polyurethanes. In order to biomonitor people exposed to low levels of MDA or MDI we have developed sensitive methods to measure hemoglobin (Hb) adducts and urine metabolites. Adducts and metabolites from 33 workers exposed to MDA and 27 workers exposed to MDI were analyzed by gas chromatography-mass spectrometry after hydrolysis, extraction and derivatization with heptafluorobutyric anhydride. Hb adducts of MDA were detected in 31 out of the 33 MDA workers and both MDA and N-acetyl-MDA (AcMDA) were found in 20 of these individuals. The detection limit for MDA was 20 fmol and for AcMDA 100 fmol/sample, which correspond to an absolute detection limit of approximately 1 fmol MDA and 5 fmol AcMDA, respectively. In the urine of workers exposed to MDA both MDA and AcMDA were found in all samples, with the exception of five where only MDA was detected. Acid hydrolysis of the urine samples yielded an approximately 3-fold higher concentration of MDA than the sum of MDA and AcMDA found after base hydrolysis. MDA but not AcMDA found in urine and in Hb correlate well, except for three outliers. In one workers the Hb adduct level of MDA was very low compared to the urine levels. Two workers had very high levels of MDA as Hb adducts but very low levels as urine metabolites. The former case indicates that the workers were recently exposed to higher levels of MDA. The latter case suggests a relatively low recent exposure. The air levels of MDA, monitored using personal air monitors, were below the detection limit. It was possible, however, to determine exposure to MDA for all workers with the methods presented in this publication. Workers exposed exclusively to MDI were studied. Exposure levels, as monitored using personal air samplers, were below the detection limit of 3 micrograms/m3, with the exception of three individuals. In 10 of the MDI workers, hydrolyzable Hb adducts of MDA (57-219 fmol/g Hb) were found. Except for four subjects, the presence of MDA (0.007-0.14 nmol/l) and AcMDA (0.08-3 nmol/l) was detected in all urine samples after base treatment. Following acid hydrolysis of the urine, higher levels of MDA (0.7-10 nmol/l) were found than the sum of free MDA and AcMDA. According to the present data, it was possible to detect exposure to MDI in a greater number of individuals by analyzing urinary metabolites than by measuring Hb adducts or air monitoring.

Determination of 4,4'-methylenediphenyldianiline (MDA) and identification of isomers in technical-grade MDA in hydrolysed plasma and urine from workers exposed to methylene diphenyldiisocyanate by gas chromatography-mass spectrometry.

Gas chromatography-mass spectrometry using chemical ionization with ammonia as reagent gas monitoring both positive and negative ions was applied. Negative-ion monitoring using ammonia and the pentafluoropropionic anhydride (PFPA) derivatives were chosen owing to low detection limits and good separation for the isomers studied. Technical-grade methylenediphenyldiioscyanate (MDI) was analysed and three isomers, 4,4'-, 2.4'- and 2,2'-methylenediphenyldianiline (MDA), were determined in addition to methylated MDA. Plasma and urine from an exposed worker were hydrolysed and analysed and the MDA isomers were identified in the biological samples.