Determination of 4,4'-methylenebis(2-chloroaniline) in urine by liquid chromatography with ion-paired solid-phase extraction and electrochemical detection.

This highly specific and sensitive method for measuring urinary 4,4'-methylenebis(2-chloroaniline) (MBOCA) involves liquid chromatography with electrochemical detection. Before chromatography, urine samples are prepared by ion-paired solid-phase extraction on a disposable octadecylsilica column with acidic methanol solution containing 1-heptanesulfonic acid. This enhances the specificity of the method. Mean overall recovery ranged from 97.1% to 99.5% at added MBOCA concentrations of 20 and 100 micrograms/L in urine. Sensitivity for urine was 1 microgram/L. The intra-assay CV was 2.2% at a MBOCA concentration of 100 micrograms/L. We believe that this method is acceptable for routine measurement of MBOCA in urine from individuals exposed to this industrial chemical.

Determination of the N-acetyl metabolites of 4,4'-methylene dianiline and 4,4'-methylene-bis(2-chloroaniline) in urine.

A sensitive and specific gas chromatographic/mass spectrometric assay is described for the determination of N-acetyl 4,4'-methylene dianiline (N-acetyl MDA) and N-acetyl 4,4'-methylene-bis(2-chloroaniline) (N-acetyl MbOCA) in urine. The method is based on the solvent extraction of the compounds together with deuterium-labelled internal standards, the compounds being separated and detected by capillary gas chromatography/mass spectrometry as their pentafluoropropyl derivatives. The method has been applied to the detection of N-acetyl MbOCA and N-acetyl MDA in the urine of workers occupationally exposed to MbOCA and MDA. The results show that whilst N-acetyl MbOCA is a relatively minor urinary metabolite a significant proportion of MDA is excreted as the N-acetylated compound.

Efficacy of urinary monitoring for 4,4'-methylenebis (2-chloroaniline).

The aromatic amine 4,4'-methylenebis (2-chloroaniline) (MBOCA), an animal carcinogen, is used commercially as a curing agent for isocyanate-containing polymers. It is structurally similar to other aromatic amines that cause bladder cancer in occupationally exposed workers. Since the late 1970s, MBOCA users have relied on urinary monitoring as the primary method of assessing MBOCA exposure in the workplace. This paper (1) outlines uncertainties about MBOCa's metabolism in humans that complicate interpretation of urinary MBOCA results; (2) describes alternative laboratory techniques for measuring MBOCA in urine; and (3) discusses observations from site visits concerning the practical application of urinary monitoring. Recommendations to improve the efficacy of monitoring programs for urinary MBOCA are outlined.

Assessment of occupational exposure to 4,4'-methylene-bis-(2-chloroaniline) "MOCA" by a new sensitive method for biological monitoring.

A new specific and sensitive method for the urinary determination of 4,4'-methylene-bis-(2-chloroaniline) ("MOCA"), a known carcinogen in rats, mice and dogs, has been developed. After a brief study on rats to determine some peculiarities in "MOCA" urinary excretion, this method was used to assess occupational exposure in French industrial firms. Both the manufacture of "MOCA" and its use as a curing agent in the production of polyurethane elastomers were surveyed. "MOCA" excretion levels were distributed from non-detectable (less than 0.5 microgram/l) up to 1600 microgram/l. Concentrations of N-acetyl metabolites, when present, were largely lower than "MOCA" levels. The results seemed to reflect workers' overall exposure fairly. Preventive measures following analytical determinations often led to an obvious lowering of excretion levels.

Structure elucidation and in vitro reactivity of the major metabolite of 4,4'-methylenebis(2-chloroaniline) (MBOCA) in canine urine.

The structure of the major urinary metabolite of 4,4'-methylenebis(2-chloroaniline) (MBOCA) in dogs was identified and the reactivity of the metabolite was characterized in vitro. Arylsulfatase but not beta-glucuronidase hydrolyzed the metabolite in a time- and enzyme concentration-dependent manner. Electron impact mass spectrometry following derivatization and transesterification indicated that the major metabolite was ring hydroxylated and fast atom bombardment mass spectrometry confirmed the molecular weight as a sulfate ester. Proton nuclear magnetic resonance studies indicated that the ring substitution was ortho to an amine. These analytical and enzymatic data supported the proposed structure of the major urinary metabolite of MBOCA in dogs as 5-hydroxy-3,3'-dichloro-4,4'diaminodiphenylmethane-5-sulfate. Protein and DNA binding in vitro and mutagenicity were investigated. During hydrolysis with arylsulfatase, time- and enzyme concentration-dependent protein binding and time-dependent DNA binding were observed. Mutagenicity during enzymatic hydrolysis in the presence of Salmonella typhimurium TA1538 with up to 20 micrograms metabolite/plate was negative and at 50 micrograms/plate the metabolite was cytotoxic. These results indicated that the metabolite was the sulfate conjugate of a reactive molecule. This study demonstrated that the major metabolite of MBOCA in canine urine is an orthohydroxysulfate and thus is similar to the major metabolites of benzidine, 2-naphthylamine, and 4-aminobiphenyl.

Biological monitoring of workers exposed to 4,4'-methylenebis (2-chloroaniline) (MBOCA).

A biological monitoring programme has been developed for assessing occupational exposure to 4,4'-methylenebis (2-chloroaniline) (MBOCA) in a factory which manufactures polyurethane elastomers. In a systematic programme of biological monitoring over a five year period urinary MBOCA concentrations have been used to provide evidence of absorption of MBOCA. Improvements in the handling and use of MBOCA have coincided with a steady reduction in the urinary MBOCA concentrations.

The determination of 4,4'-methylenebis (2-chloroaniline) in urine by electron capture gas chromatography.

The development and use of a gas chromatographic method for monitoring workers who are occupationally exposed to 4,4'-methylenebis(2-chloroaniline) (MBOCA) is reported. The increase in ether-extractable MBOCA on mild hydrolysis is determined. The analytical conditions for a specific and sensitive electron capture gas chromatographic method that will measure both "free" MBOCA and "total" MBOCA are established. It is suggested that workers should be monitored by the measurement of "total" urinary MBOCA.