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Checking up on MOCA
158
THE CHEMICAL ENVIRONMENT
with structural alterations in DNA, and we have previously mentioned one tentative mechanism for such damage, whereby the cytosine residues in the nucleic acid undergo preferential attack by the mutagen (Cited in F.C.T. 1967, 5, 107). A variety of breaks and spurious reunions become visible in the chromosome chain. A paper has now been published reporting details of the morphological effects of HA upon human leucocyte chromosomes in vitro. Chromosomal damage induced in whole blood cultures bore a direct, though not strictly linear, relation to HA dose and length of treatment. Gaps and constrictions were far more widespread than breaks and exchanges in the genetic material. No clear relation could be established between distribution of aberrations and chromosome length, but the damage appeared to be in some way selective. Thus, if administered during the G2 phase, HA tended to produce aberrations at the same locus, suggesting an effect on normal folding into the metaphase conformation. The results of this work and a similar study on HeLa $3 cells suggest that HA does not induce unscheduled DNA synthesis, its main action probably being to interact with proteins involved in chromosomal rearrangements.
2467. Checking up on MOCA Linch, A. L., O'Connor, G. B., Barnes, J.R., Killian, A. S., Jr. &Neeld, W. E., Jr. (1971). Methylene-bis-ortho-chloroaniline (MOCA): Evaluation of hazards and exposure control. Am. ind. Hyg. Ass. J. 32, 802. MOCA (4,4'-methylene-bis-(2-chloroaniline); 3,3'-dichloro-4,4'-diaminodiphenylmethane) has gained in importance over the past 10 yr as a curing agent for isocyanate polymers and epoxy resins. However, it has been reported to produce kidney irritation in man and the dog (Mastromatteo, J. occup. Med. 1965, 7, 502) and, as an aromatic amine, it is inevitably suspect as a potential carcinogen. Concern has therefore been expressed for the safety of workers handling it. A high incidence of liver, lung and mammary tumours has been reported in rats fed a protein-deficient diet containing MOCA (Steinhoff & Grundman, Naturwissenschaften 1969, 56, 215; Stula et aL Toxic appl. Pharmac. 1971, 19, 380), but the latter authors found no evidence of tumour formation after oral administration of MOCA to dogs for 4 yr (unpublished data, 1971). The evidence presented by Linch et aL (cited above) gives further grounds for hope that exposure to MOCA presents no serious hazard to man. The data presented stem from a health survey of workers exposed to MOCA for varying periods during 17 yr of process development and production. A statistically equivalent control group and other factory personnel were also included in the study. The workers within each group were classified as to general health, frequency and duration of absenteeism, type of illness and degree of exposure to the chemical. Proof of exposure to MOCA was obtained by urine analysis, which showed up interesting personal variations in absorption, metabolism and clearance rates. Analysis for MOCA was complicated by interference from other aromatic amines, but gas chromatography proved a reliable procedure capable of detecting 40 ppb MOCA in urine (b = 109) or 10/zg/ma in air. Air concentrations seldom rose above this level, however. For this reason, inhalation was not a significant route of absorption and biological rather than air monitoring was the more useful measure of exposure.
THE CHEMICAL ENVIRONMENT
159
No malignant tumours were diagnosed in any of the 31 workers who had been actively engaged in MOCA production for more than 15 yr, nor in the 31 controls. There were only two cases of malignancy (with tumours of the larynx and large intestine) in the 178 former MOCA workers, who had experienced some degree of exposure for varying periods of time during the first 6 yr of production, and both of these had been diagnosed before the men began working with MOCA. The incidence of cancer deaths in the total plant population over a 15-yr period was 0.115 ~ and was thus below the national average of 0.139 ~o. This difference may be a reflection of the relatively low age of the men involved in this study. The incidence of other disabilities was closely comparable in the exposed and control groups, with respiratory syndromes (42 and 48~, respectively) and gastro-intestinal problems (18 and 19 ~) heading the list and cardiovascular disease (2 ~ in each group) and haematological disturbances (0.5 and 0 ~o) coming a long way down. No dermatological cases were found in either group. Thus in 209 workers currently or formerly exposed to MOCA for up to 16 yr, there was no evidence of unusual absenteeism, unexpected serious or chronic disease, abnormalities of urinary cytology or an excessive incidence of malignant tumours or deaths. Direct absorption of MOCA through the skin occurred to a sufficient degree in the early days of production to raise urinary excretion levels to 25 mg/litre. At this level, MOCA had no observable haematological effects, in contrast to p-chloroaniline, which produces some degree of methaemoglobinaemia at levels of 10-20 mg/litre. This high level of MOCA absorption was later effectively reduced by improved ventilation and cleaning, the use of protective clothing, the institution of mandatory showers after each shift and other similar measures.
2468. Nitrofuran metabolism
Akao, M., Kuroda, K. & Miyaki, K. (1971). Metabolic degradation of nitrofurans by rat liver homogenate. Biochem. Pharmac. 20, 3091. Exploitation of the antibiotic properties of nitrofurans has been restricted by the reported toxic and carcinogenic effects of some of these compounds (Cited in F.C.T. 1967, 5, 426; ibid 1970, 8, 715; Food Chemical News 1971, 13 (20), 31). The present in vitro work was carried out to study the cellular site and rate of metabolism of nitrofurans in general. After separate incubation of seven nitrofurans with the 8500 g supernatant, microsomal suspension or 105,000 g supernatant of rat-liver homogenate, using added NADPHz as a hydrogen donor, metabolic changes were monitored by following optical absorbance and loss of nitro groups. For most compounds, the slow changes that occurred in these two properties under aerobic conditions appeared unrelated. Under anaerobic conditions, the loss of nitro groups and the decrease in absorbance both occurred rapidly and to a comparable degree. Much of the enzyme activity responsible for the major changes in absorbance was associated with the microsomal fraction, though some was also found in the 105,000 g supernatant. It was concluded that the metabolic changes were probably reductive, since metabolism was accelerated under anaerobic conditions. Nitrofuran degradation, at least in vitro, may therefore be associated with the same cytoplasmic structures and the same enzymes that reduce nitrophenyl compounds and azobenzenes, particularly since the presence of a nitrofuran tends to inhibit the metabolism of these other compounds. FOOD l 1 / I - - L
THE CHEMICAL ENVIRONMENT
with structural alterations in DNA, and we have previously mentioned one tentative mechanism for such damage, whereby the cytosine residues in the nucleic acid undergo preferential attack by the mutagen (Cited in F.C.T. 1967, 5, 107). A variety of breaks and spurious reunions become visible in the chromosome chain. A paper has now been published reporting details of the morphological effects of HA upon human leucocyte chromosomes in vitro. Chromosomal damage induced in whole blood cultures bore a direct, though not strictly linear, relation to HA dose and length of treatment. Gaps and constrictions were far more widespread than breaks and exchanges in the genetic material. No clear relation could be established between distribution of aberrations and chromosome length, but the damage appeared to be in some way selective. Thus, if administered during the G2 phase, HA tended to produce aberrations at the same locus, suggesting an effect on normal folding into the metaphase conformation. The results of this work and a similar study on HeLa $3 cells suggest that HA does not induce unscheduled DNA synthesis, its main action probably being to interact with proteins involved in chromosomal rearrangements.
2467. Checking up on MOCA Linch, A. L., O'Connor, G. B., Barnes, J.R., Killian, A. S., Jr. &Neeld, W. E., Jr. (1971). Methylene-bis-ortho-chloroaniline (MOCA): Evaluation of hazards and exposure control. Am. ind. Hyg. Ass. J. 32, 802. MOCA (4,4'-methylene-bis-(2-chloroaniline); 3,3'-dichloro-4,4'-diaminodiphenylmethane) has gained in importance over the past 10 yr as a curing agent for isocyanate polymers and epoxy resins. However, it has been reported to produce kidney irritation in man and the dog (Mastromatteo, J. occup. Med. 1965, 7, 502) and, as an aromatic amine, it is inevitably suspect as a potential carcinogen. Concern has therefore been expressed for the safety of workers handling it. A high incidence of liver, lung and mammary tumours has been reported in rats fed a protein-deficient diet containing MOCA (Steinhoff & Grundman, Naturwissenschaften 1969, 56, 215; Stula et aL Toxic appl. Pharmac. 1971, 19, 380), but the latter authors found no evidence of tumour formation after oral administration of MOCA to dogs for 4 yr (unpublished data, 1971). The evidence presented by Linch et aL (cited above) gives further grounds for hope that exposure to MOCA presents no serious hazard to man. The data presented stem from a health survey of workers exposed to MOCA for varying periods during 17 yr of process development and production. A statistically equivalent control group and other factory personnel were also included in the study. The workers within each group were classified as to general health, frequency and duration of absenteeism, type of illness and degree of exposure to the chemical. Proof of exposure to MOCA was obtained by urine analysis, which showed up interesting personal variations in absorption, metabolism and clearance rates. Analysis for MOCA was complicated by interference from other aromatic amines, but gas chromatography proved a reliable procedure capable of detecting 40 ppb MOCA in urine (b = 109) or 10/zg/ma in air. Air concentrations seldom rose above this level, however. For this reason, inhalation was not a significant route of absorption and biological rather than air monitoring was the more useful measure of exposure.
THE CHEMICAL ENVIRONMENT
159
No malignant tumours were diagnosed in any of the 31 workers who had been actively engaged in MOCA production for more than 15 yr, nor in the 31 controls. There were only two cases of malignancy (with tumours of the larynx and large intestine) in the 178 former MOCA workers, who had experienced some degree of exposure for varying periods of time during the first 6 yr of production, and both of these had been diagnosed before the men began working with MOCA. The incidence of cancer deaths in the total plant population over a 15-yr period was 0.115 ~ and was thus below the national average of 0.139 ~o. This difference may be a reflection of the relatively low age of the men involved in this study. The incidence of other disabilities was closely comparable in the exposed and control groups, with respiratory syndromes (42 and 48~, respectively) and gastro-intestinal problems (18 and 19 ~) heading the list and cardiovascular disease (2 ~ in each group) and haematological disturbances (0.5 and 0 ~o) coming a long way down. No dermatological cases were found in either group. Thus in 209 workers currently or formerly exposed to MOCA for up to 16 yr, there was no evidence of unusual absenteeism, unexpected serious or chronic disease, abnormalities of urinary cytology or an excessive incidence of malignant tumours or deaths. Direct absorption of MOCA through the skin occurred to a sufficient degree in the early days of production to raise urinary excretion levels to 25 mg/litre. At this level, MOCA had no observable haematological effects, in contrast to p-chloroaniline, which produces some degree of methaemoglobinaemia at levels of 10-20 mg/litre. This high level of MOCA absorption was later effectively reduced by improved ventilation and cleaning, the use of protective clothing, the institution of mandatory showers after each shift and other similar measures.
2468. Nitrofuran metabolism
Akao, M., Kuroda, K. & Miyaki, K. (1971). Metabolic degradation of nitrofurans by rat liver homogenate. Biochem. Pharmac. 20, 3091. Exploitation of the antibiotic properties of nitrofurans has been restricted by the reported toxic and carcinogenic effects of some of these compounds (Cited in F.C.T. 1967, 5, 426; ibid 1970, 8, 715; Food Chemical News 1971, 13 (20), 31). The present in vitro work was carried out to study the cellular site and rate of metabolism of nitrofurans in general. After separate incubation of seven nitrofurans with the 8500 g supernatant, microsomal suspension or 105,000 g supernatant of rat-liver homogenate, using added NADPHz as a hydrogen donor, metabolic changes were monitored by following optical absorbance and loss of nitro groups. For most compounds, the slow changes that occurred in these two properties under aerobic conditions appeared unrelated. Under anaerobic conditions, the loss of nitro groups and the decrease in absorbance both occurred rapidly and to a comparable degree. Much of the enzyme activity responsible for the major changes in absorbance was associated with the microsomal fraction, though some was also found in the 105,000 g supernatant. It was concluded that the metabolic changes were probably reductive, since metabolism was accelerated under anaerobic conditions. Nitrofuran degradation, at least in vitro, may therefore be associated with the same cytoplasmic structures and the same enzymes that reduce nitrophenyl compounds and azobenzenes, particularly since the presence of a nitrofuran tends to inhibit the metabolism of these other compounds. FOOD l 1 / I - - L