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The Health Council of The Netherlands advises the Dutch government on scientific issues concerning health and environmental protection. In 1978 a committee of the Council established mechanistic criteria for quantitative risk assessment of carcinogenic substances. For this purpose it adopted the multistage model of carcinogenesis. Based on this model two categories of carcinogens were distinguished. The first category includes complete carcinogens and tumor initiators: these agents induce irreversible modification of DNA and act by a stochastic mechanism. The second category includes promotors and other substances acting as cocarcinogens; these agents act by nonstochastic mechanisms. Stochastic action implies that there is no threshold for the contribution to the carcinogenic effect, whereas nonstochastic action implies a threshold dose. Recently, reevaluation of the method for carcinogenic risk assessment was considered due because of remarkable scientific progress since 1978, such as the identification of cellular oncogenes and tumor suppressor genes, and the partial clarification of mechanisms at the cellular and/or molecular level by which tumor promotors stimulate tumor development. The present paper discusses the Dutch method for carcinogenic risk assessment in the light of the recent scientific developments. It concludes that this method for risk assessment is still a valid and appropriate one.

In 1986 a survey was published by CEFIC on the occurrence of chlorinated solvents in ambient air, in surface water, and in ground water. The present article concentrates on 1,1,1-trichloroethane (1,1,1-T), and puts into perspective the environmental occurrence and the toxicity. Critical toxicological data are briefly discussed. As no evidence of a carcinogenic effect of 1,1,1-T is apparent, the no-adverse-effect levels in chronic inhalation exposure in rats (875 ppm) and mice (1500 ppm) form the basis for the estimation of potential risk to human health. Environmental exposure to 1,1,1-T is mainly via the atmosphere (120 micrograms/day); the contributions of drinking water (2 micrograms/day) and food (3 micrograms/kg) are negligible. Safety margins are calculated by comparing the no-adverse-effect levels in rat and mouse studies with the total body burden. Safety margins are also calculated after converting no-adverse-effect levels into estimated internal dose levels by physiologically based pharmacokinetic modeling. Safety margins vary with the starting point, but are of the order of 10(5) for the general population and more than 10(4) for the population close to industrial activities. It may be concluded that the risk of a potential health effect resulting from environmental exposure to 1,1,1-trichloroethane is negligible.

The herbicieds 2-methyl-4-chlorophenoxy acetic acid (MCPA) and 2-(2-methyl-4-chlorophenoxy) propionic acid (MCPP or mecoprop) were tested for 90 days in rats. The compounds were added to the diet at levels of 0, 50, 400, and 3200 ppm. Growth, food intake, mortality, haematology, blood and liver chemistry, organ weights and histopathology were used as criteria. The main effects of both compounds were growth retardation and elevated relative kidney weights at levels of 400 ppm and more. The 50 ppm dose level can be considered as a no-toxic-effect level in the 90-day study. In subacute dermal studies in rabbits during 3 weeks the dosages were 0, 0.5, 1.0 and 2 g MCPA or MCPP per kg body weight. Therafter followed a recovery period of 2 weeks. Growth, mortality, skin reaction, haematology, organ weights (MCPP) and histopathology were recorded and determined. Both compounds caused slight to moderate erythema at all dose levels, whereas elasticity of the skin was decreased. In both experiments the skin returned to normal during the recovery period. Weight loss was observed at all dose levels. In the MCPA experiment high mortality and histopathological changes in the liver, kidneys, spleen and thymus were recorded at the two highest dose levels. The cause of this could have been either the treatment with MCPA or a dysbacteria infection which developed during the experiment. Oral and intraperitoneal acute toxicity of MCPP for the rat were found to be 1210 and 402 mg/kg, respectively. After a single oral or dermal application of MCPA to the rabbit, the compound was excreted unchanged in the urine.

Four groups, each of 25 male and 25 female weanling rats, were given dietary levels of 0, 0.1, 0.5 and 2.5 ppm MeHgCl for 2 years. Observations were made on behaviour, growth, food intake, haematology, serum enzymes, urinalysis, microsomal liver enzymes, organ weights and histology with special reference to the nervous system, hitochemistry of the kidneys and cerebellum and on tissue Hg concentrations. Significant findings included a slight growth reduction in females at 2.5 ppm, increased relative kidney weight at 2.5 ppm and histochemical changes in kidney enzymes at 2.5 ppm. No effect was seen on the nature or incidence of pathological lesions or tumours at any level. From the results obtained in the short-term, reproduction and long-term studies, the no-toxic effect level for rats appears to be between 0.1 and 0.5 ppm MeHgCl in the diet. Exposure of the Dutch population does not appear to present a health hazard at the moment because the mean intake of total Hg is still far below the intake deemed to be safe.

In a comparative study 0, 50, 200, 1000 and 3000 ppm (parts per million) of five acaricides, tetrasul (2,4,5,4′-tetrachlorodiphenylsulphide), V110 (2,4,5,4′-tetrachlorodophenylsulphoxide), tetradifon, chlorfenson and kelthane were given to groups of 10 male and 10 female rats in separate 90-day toxicity experiments. Growth, food intake, mortality, haematology, blood and liver biochemistry, organ weight, histopathology, number of liver parenchymal cells per surface unit and the diameter of liver-cell nuclei were used as criteria. The acaricides investigated, with the exception of kelthane, induced both functionally and structurally the same liver alterations. SER (smooth endoplasmic reticulum) whorls were found in the higher dose-levels of all 5 compounds. In all compounds, except kelthane, enlarged thyroids were found accompanied by histological alterations. Tetrasul was more toxic than V110, followed by tetradifon. Chlorfenson was equally as toxic as tetrasul. Kelthane can be regarded as the most toxic of this group of compounds. The 50-ppm dosage appeared to be a no-toxic effect level only in the case of tetradifon.

In a 90-day feeding study, 4 groups of 10 male and 10 female rats received in the diet 0, 200, 1000 and 5000 ppm 1-naphthaleneacetic acid (1-NAA). Growth and food intake was reduced significantly only in males on 5000 ppm. Haematological examination yielded essentially negative results except for a non-significant reduction at the 1000 and 5000 ppm levels in the leucocyte count, mainly due to a reduction in neutrophils. No significant effects were observed in the renal concentration test, urinalysis, renal histochemistry or histology of a wide range of organs at any level of 1-NAA tested. Increased relative weights of thyroid, testes, brain and liver were confined to the 5000 ppm level. The increase in relative liver weight was not accompanied by histological liver damage and was associated with elevated liver microsomal enzyme activity. The loss of glucose 6-phosphate (G6Pase) and increase in glucose 6-phosphate dehydrogenase (G6PDH) seen histochemically in the centrilobular region of the liver in males on 5000 ppm, accompanied by glycogen depletion in the liver, could however be indicative of liver damage. On the basis of conventional criteria, a no-effect level of 1000 ppm would have been indicated by this study but in view of liver glycogen depletion at all levels tested a no-effect level was not established.

Hexachlorobenzene Toxicity in Pigs. den Tonkelaar, E. M., Verschuuren, H. G., Bankovska, J., de Vries, T., Kroes, R., and van Esch, G. J. (1978). Toxicol. Appl. Pharmacol. 43 , 137–145. A 90-day toxicity study was carried out in which pigs received 0.05, 0.5, 5.0, and 50 mg/kg/day of hexachlorobenzene (HCB). Special attention was given to the induction of porphyria and microsomal liver enzymes and the concentration of HCB in blood and tissues. Animals given the highest dose (50 mg/kg) showed clinical signs associated with porphyria and died during the experiment. At lower dosages these signs were not observed. An increased excretion of coproporphyrin was found in the 5.0- and 0.5-mg/kg groups, as well as induction of microsomal liver enzymes accompanied by increased liver weight (at 5.0 mg/kg) and characteristic histopathological changes in the liver. The concentration of HCB in blood and tissues was elevated at all dosages. Concentrations in fat were approximately 500 times greater than those in blood, while the concentration in liver was higher than in the kidneys and brain. Under the conditions of this experiment, a no-effect level was judged to be 0.05 mg/kg/day of HCB.

Rats received 0, 200, 1000 and 5000 ppm metaldehyde in the diet for 2 years. Reproduction studies over three generations using the same dietary levels were carried out. In the third litter of each generation attention was paid to possible embryotoxic or teratogenic effects. The parameters studied included growth, food intake, behaviour and survival, haematology, clinical biochemistry, organ weight, histopathology, reproductive performance and teratogenicity. At 5000 ppm the relative liver weight was increased and this was accompanied by an increase in liver microsomal enzyme activity. The most striking observation was a dose-related development of posterior paralysis in females due to a transverse lesion of the spinal cord. The latency period was more than 550 days. Three rats with posterior paralysis showed a transverse lesion of the spinal cord. No significant histological damage to other organs was seen. The tumour incidence was not increased in any of the metaldehyde dosage groups. The reproduction study confirmed the findings of the long-term test. Posterior paralysis appeared in at least 50% of the females on 5000 ppm metaldehyde in all 3 generations. Some were affected at 1000 ppm but none at 200 ppm. Histologically, a fracture or distortion of thoracic vertebrae and subsequent compression of the spinal cord was found. The onset of paralysis was related to the time of delivery. The reproductive performance was susceptibility in this respect to metaldehyde. Apart from one male rat on 200 ppm with clinical posterior paralysis without transverse lesions in the spinal cord, this level was without toxic effects both in the long-term and 3-generation reproduction study.

In July 1984 the Minister of Welfare, Public Health and Culture, representing the Dutch government, sent a request to the Health Council of the Netherlands to advise on the health risks presented by environmental exposure to several carcinogenic substances. One of these substances was vinyl chloride (VC). On the basis of a working document prepared by the National Institute of Public Health and Environmental Hygiene, a committee of the Health Council of The Netherlands prepared a report concerning a health risk assessment of VC which was published in May 1986. A short review is presented of the available data and the considerations that formed the basis for the risk assessment of the carcinogenicity of VC to humans. The advice was based mainly on human data from epidemiological studies of workers occupationally exposed to VC. The committee concludes that continuous exposure to 0.001 mg/m 3 VC corresponds to an additional cancer mortality risk of 10 −6 per lifetime. The Dutch government considers this additional risk to the general population to be acceptable.

Pentachlorophenol (PCP) was fed to rats at dietary levels of 0, 25, 50 and 200 ppm for 12 weeks. Growth was decreased in the group of female rats fed 200 ppm. The treatment had no effect on food intake and behaviour. Liver weight was increased at the 50 and 200 ppm dose levels accompanied by an increased activity of microsomal liver enzymes. In week 6 higher haemoglobin and haematocrit values were found in the groups of males fed 50 and 200 ppm. In week 11 the haemoglobin values and the number of erythrocytes were decreased in the same groups of males. A possible explanation is discussed. A striking dose-related decrease of calcium deposits in the kidney is found. The no-toxic-effect level for all criteria is considered to be 25 ppm.

A reporduction study over 3 generation of rats was carried out in which groups of 20 female and 10 male rats received in the diet 0, 0.1, 0.5 and 2.5 ppm MeHgCl. The parameters studied included growth, food intake, haematology, serum and urinalysis, organ weights and reproductive performace. No effect was exerted on fertility index, lactation index or on the 21-day body weights of pups but the viability index was imparied at 2.5 ppm in the F1 and F2 generations. Weight gain reductions oberserved at 12 weeks for the 2.5 ppm level were not accompanied by reductions in food intake. At 6 months, Fla females on 2.5 ppm showed a reduced leucocyte count whilst P males on 0.5 and 2.5 ppm showed an increase in neutrophils and a decrease in lymphocytes. The relative weights of the kidneys, heart, spleen, brain and thyroid were increased at 2.5 ppm and in some cases the increases of kideny weights were inconsistently seen at the 0.1 and 0.5 ppm levels. No significant histological changes were seen at any level. In a special 7-week study involving the F3a generation, weanling rats obtained from the four different F2a groups, each comprising 20 females and 10 males, were all transferred to diets containing 25 ppm MeHgCl. Toxicity signs were evident at 7 weeks. No evidence was obtained of increased susceptibility to the toxicity of MeHgCl in successive generations.

The acaricide tetrasul (2,4,5,4′-tetrachlorodiphenylsulphide) is moderately toxic in a number of animal species. The oral LD50 for the rat is 3.65 g/kg body weight. Guinea pigs are considerably more sensitive (LD50 = 0.50 g). After intraperitoneal injection of a suspension in oil the substance is less toxic than after oral administration, due to poor absorption. When tetrasul was fed to rats at dietary levels of 0, 10, 25, 50, 200, 1000 and 3000 ppm for at least 2 years, there was a reduction of growth and food intake at the 1000- and 3000-ppm levels. Reduction of haemoglobin occurred at a 6-month stage with dose levels of 200 ppm and above. Stridor occurred at dose levels of 50 ppm and above, and the incidence was dose-related. Survival was shorter for the 1000- and 3000-ppm groups. Liver and thyroid weights were increased at all stages. In the liver characteristic histological alterations were observed with the formation of SER (smooth endoplasmic reticulum) whorls at the 200 ppm and higher levels. In the thyroid hyperplastic changes were observed at the 1000- and 3000-ppm levels. The 10-ppm level was without any adverse effect and in the 25-ppm group adaptive changes occurred in the liver of a small number of animals. In the three-generation reproduction study using the same dosage the reproductive performance in the 1000- and 3000-ppm groups was diminished. There was a tendency to decreased fertility in the males of the 50- and 200-ppm groups. The histological picture of the testes was normal.

Tetrasul (2,4,5,4′-tetrachlorodiphenylsulphide) was tested for 90 days in mice, rats, guinea pigs, rabbits, mini-pigs and chicken, in 6 separate experiments but using the same experimental design. Tetrasul was added to the diet at 0, 50, 200, 1000 and 3000 ppm. Growth, food intake, mortality, haematology, blood and liver chemistry, organ weights and histopathology were used as criteria. The liver was the target organ in all species examined. Although effects on thyroid weight and histopathology were found, these could not be evaluated properly, due to differences in iodine content of the diet. If only the most sensitive criteria, namely histopathological changes in the liver and the induction of microsomal liver enzyme activity are taken into account, then mini-pigs and rats appeared to be the most sensitive species for tetrasul. Guinea pigs, rabbits and chicken followed very closely, whereas mice were the least sensitive. The 50-ppm dosage level produced some effects on the liver cell structure in all species except in chicken.

In two experiments an immunosuppressive effect of triphenyltin acetate (TPTA) in female guinea pigs was detected with immunochemic, serologic, histologic, and fluorescent antibody techniques. In the first experiment the animals were given 0 or 15 ppm TPTA in the diet for 49 and 77 days. In the second experiment the animals received the same diet for 104 days. At 21 and 7 days before the end of the TPTA feeding, half of the animals received an injection of tetanus toxoid. Some animals were allowed to recover for 14 days after the 104-day experimental period. Immunochemically and serologically a decreased immunologic response was observed in the TPTA-fed group after tetanus toxoid stimulation, and no signs of recovery were seen after 14 days. No data are available for later stages. In the first experiment, after 77 days, a decrease in plasma cells was observed in the spleen and the cervical, mesenteric, axillary, and popliteal lymph nodes. With the aid of the fluorescent antibody technique a decrease in immunologically active cells in the popliteal lymph nodes of the TPTA-fed animals could be detected in the second experiment. No signs of recovery could be demonstrated after 14 days. In the other lymphatic tissues no differences were observed. It is concluded that TPTA should be regarded as an immunosuppressive agent.

A toxicological investigation was carried out in order to repeat and extend the findings of Bukhman (Vop. Pitan. 1962, 21, 68) concerning the toxicity of propyl gallate in fasting rats. Rats were given a diet containing 0·002 or 0·004% propyl gallate or α-tocopherol. After 13 wk some of the animals were sacrificed, while the remaining rats were placed on a partial starvation diet until death. The survival time was recorded. Chemical determinations of water, protein and fat content were made on the sacrificed animals as well as on those that died. No difference in survival time or in protein and fat utilization was found between the control groups and the groups which received propyl gallate or α-tocopherol. The results of Bukhman could not be confirmed.

Short-term toxicity studies have been carried out in rats and guinea-pigs fed diets containing triphenyltin acetate (TPTA), triphenyltin hydroxide (TPTH) or triethyltin hydroxide (TETH) for 90 days at levels ranging from 0 to 50 ppm. The lowest dietary levels found to retard growth in rats and guinea-pigs respectively are as follows: 5 and 10 ppm for TETH; 25 and 5 ppm for TPTA; 50 and 20 ppm for TPTH. An increase in mortality was found in the following groups: rats on 10 ppm or more of TETH; guinea-pigs on 20 and 50 ppm TPTA and on 50 ppm TPTH. In rats and guinea-pigs given TETH, paralysis and pronation of the hindlegs developed. These toxic signs were not found in the TPTA- and TPTH-treated animals. This observation was correlated with the finding that only the TETH-treated animals developed interstitial oedema of the central nervous system. The water content of the brain and spinal cord was increased in rats and guinea-pigs at respective minimum levels of 5 and 10 ppm of TETH and in guinea-pigs on 20 ppm TPTA. In female rats given 50 ppm TPTA or TPTH the water content of the spinal cord was only increased. The changes in organ weights which could be related to the administration of the organotin compounds were confined to an increase of brain weight in rats and guinea-pigs given TETH, and in guinea-pigs given 5 ppm or more of TPTA and a decrease of thymus and/or spleen weights in rats and guinea-pigs receiving TETH. Guinea-pigs given 5–20 ppm TPTA or 2·5–20 ppm TPTH displayed lymphopenia accompanied in a number of animals by histological changes in the lymphopoietic system, viz. atrophy of the white pulp of the spleen was found. This possibly induced a decrease of general resistance, so that a mycotic infection developed in the animals. Hitherto, damage to the central nervous system was considered to be the most sensitive criterion for assessing the toxicity of organotin compounds, but the present work shows that the effect on lymphopoiesis is a more sensitive index of toxic action for triphenyltin compounds.