Investigations of cigarette smoke dosages in inhalation experiments with Syrian hamsters. I. Concentration of cigarette smoke in the inhalation chamber and of carbon monoxide in the blood.

We tested 20 different smoke inhalation machines and determined that 60% of the gas phase and 40% of the total paritulate matter reached the inhalation chamber; the remainder was lost in the exhaust gases. After determining these concentration percentages, we were able to calculate precisely the dosage of each puff of smoke administered to Syrian hamsters in our smoking machine. We also determined that the CO-Hb content in the blood of the hamsters rose to 60%; this CO-Hb value was a limiting factor for time of exposure to smoke, since too-high concentrations of CO-Hb would have killed the animals. On the basis of these results, we concluded that the animals should be exposed as long as possible to attain a high dose of smoke, but because of toxic effects the exposure time should not exceed 20 minutes, and the CO-Hb content should not exceed 60%.

Investigations of cigarette smoke dosages in inhalation experiments with Syrian hamsters. II. Uptake of cigarette smoke by the animals; comparisons of the influence of the gas phase and the whole smoke of different cigarettes.

An increased CO2 content (CO2 enrichment) in the smoke-air mixture was measured in an inhalation chamber containing Syrian hamsters. Since these additional CO2 concentrations originated from the animals' expiratory air, the level of CO2 (CO2 enrichment) was considered a function of the respiratory capacity. Results obtained with this method of characterizing the respiratory behavior of hamsters in inhalation experiments led us to conclude that the vapor phase of cigarettes reduced respiration up to 45% and the whole smoke, up to 60%. Qualitative and quantitative differences in the composition of the vapor phase and particulate phase as offered in animal inhalation experiments for comparing different cigarettes did not induce measurable differences in the respiratory behavior. Therefore, such inhalation experiments were found to be suitable for comparing the relative effects of different cigarette types. Estimations on the inhaled amounts of TPM led to results that were similar to those obtained in experiments with radioactively labeled smoke.

Investigations on the subchronic toxicity of 2-methoxypropanol-1(acetate) in rats.

Wistar rats were exposed to 2-methoxypropylacetate-1 (2-MPAc-1) vapours in concentrations of 0, 110, 560 and 2800 ppm for (equiv. to 0; 0.6; 3.0 and 14.9 mg/L) for 4 weeks in chambers (6 hours/day; 5 days/week; five male and five female animals per group). The top concentration was equivalent to a 95% vapour saturation at 20 degrees C and the animals reacted to this with a moderate respiratory irritation during the 6 hours exposure times; at 560 ppm these effects were only slight. The top dose was also associated with a significantly reduced body weight development and some hematologic and biochemical alterations of little specificity. The most prominent effect was thymic atrophy. No effects were noted on the testes or on the cellularity in blood or bone marrow. 560 ppm were without systemic effects. Furthermore, 2-methoxypropanol-1 (2-MP-1), 2-MPAc-1 and 2-ethoxyethanol (EE) were administered in parallel by gavage to groups of five male Wistar rats daily for 10 days at near equimolar dose levels (1800, 2600 and 1800 mg/kg per day, respectively). At the end of the administration period the testes were investigated. There was a pronounced testicular atrophy in animals exposed to EE, whereas no adverse effects were observed with 2-MP-1 and 2-MPAc-1. The results of these studies indicate that 2-MP-1 and 2-MPAc-1 which previously had been shown to exert pronounced prenatal toxicity in rabbits and weak prenatal effects in rats are devoid of other forms of systemic toxicity in rats that are typically observed with ethoxyethanol and methoxyethanol.

Dermal oncogenicity study of 2-ethylhexyl acrylate by epicutaneous application in male C3H/HeJ mice.

A carcinogenicity bioassay of 2-ethylhexyl acrylate (2-EHA) was conducted by applying 25 microliters 86.5%, 21%, or 2.5% 2-EHA in acetone three times a week to the clipped dorsal skin of male C3H/HeJ mice (80 per group) over their lifetime. Another group was treated with a 43% 2-EHA solution for 24 weeks and thereafter observed for lifetime (stop-test). An untreated group and a group that received only the diluent acetone served as controls. Treatment-related changes in the skin indicative of irritation (scaling, scabbing, hyperkeratosis, hyperplasia) were found in all 2-EHA-treated groups. These lesions were reversible in the 43% group immediately after treatment was stopped, and in the 2.5% group after the 11th week of treatment. Only in the 86.5% and 21% test groups showing chronic irritative skin damage was there a high incidence of nepolastic skin lesions (papillomas, carcinomas, and melanomas) with no dose dependency. In contrast, no skin tumors were found in the control groups, in the group treated with 2.5% 2-EHA for lifetime or in the group treated with 43% 2-EHA for about 6 months and observed for lifetime.

Lung deposition, lung clearance and renal accumulation of inhaled cadmium chloride and cadmium sulphide in rats.

Rats were exposed 6 h/day over 10 days to 0.3 mg/m3 of water soluble cadmium chloride and 0.2, 1.0 and 8.0 mg/m3 of insoluble cadmium sulphide, then killed at intervals over a 3-month period for serial measurements of lung, renal and faecal cadmium. CdCl2 and high-dose CdS animals showed a transient increase in lung weight. Clearance of both compounds was biphasic. Approximately 40% of deposited material was cleared during the 10-day exposure period. For CdCl2, only 9% of the lung burden was cleared rapidly after the last exposure (half-life 1.0 days) and 47% slowly (half-life 87 days), leaving a residual lung burden of 44%. For CdS, 41% of the lung burden was cleared rapidly (half-life 1.4 days) and 40% slowly (half-life 42 days), leaving a final residue 19%. In the CdS high-dose group, the retention of CdS in the lung was greater than that in the CdS low-dose groups, indicating that clearance mechanisms may possibly have been impaired in the high-dose group by too great a lung burden. For both compounds, faecal cadmium was initially high. Renal accumulation of cadmium was substantial for CdCl2 during the exposure period and continued over the following months until it represented approximately 35% of the total cadmium cleared from the lung. For CdS, renal accumulation was only 1% of the amount cleared from the lung. The bioavailability of Cd from CdS is thus poor, the majority being cleared from the lungs and excreted in the faeces. However, the bioavailability of inhaled CdS measured as cadmium in the kidney is greater than the bioavailability of orally ingested CdS.

Uptake of carbon monoxide in blood of miniture pigs and other mammals.

Different affinities of blood for CO were observed in in vivo and in vitro experiments. Hamsters showed the greatest relative affinity (1.0), the corresponding value for rats was 0.8, for pigs 0.74 and for rabbits 0.58. The upward shift of the saturation curves was dependent on the rate of respiratory exchange per unit of body weight, e.g. the curves were less steep for pigs than for rabbits. After increased motor activity the equilibrium saturation of CO in blood is reached more rapidly. From the results of in vitro experiments the following affinity constants for the blood of different species could be calculated: pig K = 130, hamster K = 181, rat I = 141, rabbit K = 109. For equal atmospheric CO concentrations the saturation values were lower (37-40%) in experiments in vitro than in vivo. Similar saturation values were obtained with light excluded, when O2 concentration was decreased to the level of alveolar oxygen concentration. With regard to the affinity of blood for CO, the blood of domestic pigs did not differ from that of minature pigs in in vitro experiments.

Developmental toxicity in rats after inhalation exposure of di-2-ethylhexylphthalate (DEHP).

Di-2-ethylhexylphthalate (DEHP) was investigated in Wistar rats for developmental toxicity after head-nose exposure to aerosol concentrations of 0, 0.01, 0.05 and 0.3 mg/l for 6 h per day from gestation day 6 through 15. A range finding study revealed peroxisome proliferation in the liver of the dams throughout exposure levels of 0.2, 0.5 and 1.0 mg/l with an increasing trend. 0.3 mg/l was therefore regarded as an exposure level leading to peroxisome proliferation as a marker for maternal effects. All concentrations were tolerated without clinical signs of maternal toxicity. The fetuses of 20 animals per exposure group were investigated for structural defects. Five additional animals per group were allowed to litter and the offsprings were raised and observed for postnatal signs of toxicity. No significant developmental toxicity or changes in the postnatal physical development were observed. DEHP is assumed not to exhibit developmental toxicity under the experimental conditions employed.

Developmental toxicity of 2-butin-1,4-diol following oral administration to the rat.

Developmental toxicity of 2-butin-1,4-diol was determined in groups of 18-22 pregnant Wistar rats at dose levels of 10, 40 and 80 mg/kg bw/day administered by gavage from days 6 to 15 pc. At 80 mg/kg bw/day food consumption and maternal body weight were reduced and one dam died during the treatment period. At this dose level the incidence of affected fetuses per litter with accessory 14th ribs was increased. This variation is assessed as an embryotoxic effect resulting from non-specific stress on the dams. No teratogenic effects were caused by 2-butin-1,4-diol. The NOAEL on the maternal and the developing organism was 40 mg/kg bw/day.

Tri-isobutylphosphate: a prenatal toxicity study in rats.

To assess the prenatal toxicity to rats of the anti-foaming agent, tri-isobutylphosphate (CAS 126-71-6), a study was conducted in which daily dosages of 0, 100, 300 and 1000 mg/kg were administered to different treatment groups by gavage from day 6 to 15 of pregnancy. Dams were killed and foetuses examined on day 20 of pregnancy. Maternal effects during the dosing period included a dosage-related increase in the frequency, persistence and severity of post dosing salivation in all test groups and significantly increased water consumption at 1000 mg/kg. Bodyweight gain at 1000 and 300 mg/kg was lower than that of controls but the differences were not statistically significant. The lowest dosage of 100 mg/kg could be considered as the maternal 'lowest observed adverse effect level' (LOAEL) or 'no observed adverse effect level' (NOAEL) according to whether increased salivation is perceived to be a true toxic effect or simply a reaction to the taste of the test material. Neither litter values nor the prevalence of foetuses with abnormalities indicated any embryotoxic effects (including teratogenicity) at any dosage. The most notable feature of the results was the occurrence of a cluster of foetuses with the congenital abnormality referred to as 'hunched posture syndrome' or 'squat foetus syndrome'. However, the incidence of this finding was similar to that noted among background data for the same strain and, in the absence of any other embryotoxic findings, was considered likely to have arisen coincidentally.

Chronic toxicity and oncogenicity of inhaled methyl acrylate and n-butyl acrylate in Sprague-Dawley rats.

No exposure-related clinical signs or lesions of systemic toxicity and no oncogenic responses were observed in male and female Sprague-Dawley rats exposed by inhalation to methyl acrylate (MA) or n-butyl acrylate (BA) vapours, at concentrations of 0, 15, 45 and 135 ppm. The rats were whole-body-exposed 6 hr/day, 5 days/wk, for 24 consecutive months. There was a 6-month post-exposure observation period for subgroups of BA-exposed rats. Atrophy of the neurogenic epithelial cells and hyperplasia of reserve cells were observed in the nasal mucosa of all MA- and BA-exposed groups. These changes were dose related and mainly affected the anterior part of the olfactory epithelium. Opacity and neovascularization of the cornea were seen in all MA-exposed groups and in the group exposed to 135 ppm BA. These toxic effects of the olfactory epithelium and cornea were attributed to the known irritancy of MA and BA. In the BA subgroups kept for a 6-month post-exposure observation, reconstructive effects, such as replacement of altered olfactory epithelium with respiratory epithelium, and partial regression of corneal neovascularization were observed.

Study on the developmental toxicity of orally administered isobutylidenediurea in rats.

Developmental toxicity of isobutylidenediurea (IBDU) was determined by oral administration to Wistar rats. The substance was administered as an aqueous suspension to 22-24 pregnant rats per group by gavage in daily doses of 100, 400 and 1000 mg/kg body weight from day 6 post-coitum (p.c.) to day 15 p.c. The control group received the vehicle only (0.5% aqueous carboxymethyl cellulose solution). There were no substance-related effects in the dams concerning food consumption, body weight, body weight gain, uterine weights and clinical or autopsy observations even at the highest dose of 1000 mg/kg body weight/day. The reproduction data revealed no biologically relevant differences between the control and treated groups. The incidence and type of the foetal external, soft tissue and skeletal findings, which were classified as malformations, variations and/or retardations observed in the treated foetuses were similar to the concurrent and/or historical control data. Thus, under the conditions of this study, no signs of maternal toxicity or embryo/foetotoxicity were induced by IBDU and the no-observable-adverse-effect level on the maternal and developing organism was 1000 mg/kg body weight/day.

Subchronic inhalation toxicity study of 2-ethylhexanol vapour in rats.

A 90-day subchronic inhalation toxicity study was performed on Wistar rats in accordance to OECD testing guidelines to evaluate the toxicological profile of 2-ethylhexanol, potential target organs, and a no-observable-adverse-effect-level (NOAEL). 10 males and 10 females per group were exposed to 2-ethylhexanol vapours at concentrations of 15, 40 and 120 ppm (the latter corresponding to the vapour saturation at 20 degrees C) 6 hours/day for 90 days. The respective controls inhaled clean air under the same conditions. No substance-related adverse effects were observed for body weight, body weight gain, mortality, organ weights, clinical biochemistry and haematological parameters including clotting time. Cyanide-insensitive palmitoyl-CoA oxidation, a marker for peroxisome proliferation, was found elevated in a subchronic study in Fischer 344 rats after gavage application of 500 mg/kg but not under the conditions of this 90-day subchronic inhalation study. There were no findings related to the treatment with 2-ethylhexanol either at necropsy or at histological examination. The highest concentration tested under these conditions (120 ppm) was found to be the NOAEL for male and female rats.

Studies on the prenatal toxicity of N,N-dimethylformamide in mice, rats and rabbits.

Prenatal toxicity studies with N,N-dimethylformamide (DMF) in rabbits, rats and mice were carried out using the oral (gavage), dermal, inhalation and ip injection routes of administration. Administration of DMF by gavage led to an increase in malformations in rats and mice in the absence of overt maternal toxicity. The lowest-observable-effect level was 182 mg/kg body weight/day in mice and 166 mg/kg body weight/day in rats. After dermal administration a dose-dependent incidence of teratogenicity was observed in rats at 94-944 mg/kg/body weight/day in the absence of overt maternal toxicity. In rabbits dermal administration led to a steeper increase in the dose-response relationship and at 400 mg/kg body weight/day to a clear teratogenic effect in the presence of slight maternal toxicity. The 200 mg/kg body weight/day dose appeared to be the no-adverse-effect level. Inhalation in rats caused foetotoxicity and embryolethality at 287 ppm. A clear teratogenic effect was shown in rabbits at 450 ppm and a marginal effect at 150 ppm. The no-effect level for does and foetuses was 50 ppm. Ip injection in mice caused clear teratogenicity at 944 mg/kg body weight/day and slight embryotoxicity at 378 mg/kg body weight/day. The rabbit appears to be more sensitive than the rat to DMF-related prenatal toxicity and should, therefore, be used as the basis for the evaluation of teratogenic risk in humans.

Di-(2-ethylhexyl) phthalate: a short-term repeated inhalation toxicity study including fertility assessment.

In a study of the 28-day inhalation toxicity of di-(2-ethylhexyl) phthalate (DEHP) aerosols, 9-wk-old Wistar rats, 27 males (mean weight 226 g) and 17 females (mean weight 155 g) per group, were exposed in head-nose inhalation systems to DEHP aerosols of respirable particle size (mass median aerodynamic diameter < or = 1.2 microns) or air (controls). Exposure for 6 hr per day, 5 days per wk for 4 wk to target concentrations of 0, 0.01, 0.05 and 1.0 mg/litre gave estimated doses of 230, 11 and 2.3 mg/kg/day for the males, and 360, 18 and 3.6 mg/kg/day for females, on the assumption of 100% deposition and absorption. Clinical investigation and blood chemistry parameters did not reveal any treatment-related effects. At the end of exposure a statistically significant (16%) increase in relative lung weights, accompanied by increased foam-cell proliferation and thickening of the alveolar septi, was found in the males of the highest dose group. Absolute liver weights were significantly (8.75%) increased in females and relative liver weights were increased in both sexes in the highest dose group, but there were no corresponding histological effects. All these effects were reversed during the 8-wk post-exposure period. No testicular toxicity was observed histologically and no impact on mating performance and male fertility was detected after two matings of treated males with untreated females, 2 and 6 wk after the end of exposure. Electron microscopic examination of liver samples from two male and two female rats per group at the end of exposure and after the 8-wk post-exposure period did not reveal clear substructural changes that could be attributed to exposure or to peroxisome proliferation. The no-observed-effect level for all exposure-related findings was 0.05 mg/litre under the conditions used.

Long-term inhalation toxicity of N-vinylpyrrolidone-2 vapours. Studies in rats.

In previous subchronic studies inhaled N-vinylpyrrolidone-2 (NVP) was haemotoxic, hepatotoxic and irritant to the nose. In the first of two long-term studies, study A, Sprague-Dawley rats were exposed by inhalation to 0, 5, 10 or 20 ppm NVP (6 hr/day, 5 days/wk) for 24 months. Satellite groups were killed after 3, 12 or 24 months. In study B, female Sprague-Dawley rats were exposed to 0 or 45 ppm NVP for 3 months and killed at 3 or 12 and 24 months post-exposure. In study A, survival was unaffected, but reduced body weight gain, haemotoxicity, effects on clinical chemistry parameters indicative of hepatotoxicity, increased liver weight, hepatocellular carcinomas, necrosis, reparative hyperplasia, adenomas and adenocarcinomas of the nasal cavity, and squamous cell carcinomas of the larynx were seen. Increased tumour incidence was seen only in the liver and upper respiratory tract. In study B, the effect of NVP on body weight evident at 3 months disappeared before 1 yr, but effects on liver pathology persisted throughout the subsequent 21-month exposure-free period, and a few liver tumours were seen at 2 yr. As NVP gave negative results in a battery of in vitro and in vivo genotoxicity tests, it appears that the tumours that arose were manifestations of a non-genotoxic mechanism.

Subchronic inhalation and oral toxicity of N-vinylpyrrolidone-2. Studies in rodents.

N-Vinylpyrrolidone-2 (NVP) is a monomeric compound used as an industrial intermediate. Nine of 11 studies previously reported involved exposure of rats (two different strains), mice or hamsters to NVP by the inhalation route at concentrations of up to 120 ppm (6 hr/day, 5 days/wk) over a period of 1 wk to 12 months. The remaining two studies involved exposure of rats to NVP through the drinking water or by gavage at dose levels of up to 100 mg/kg body weight/day. Reduced body weight gain was seen in rats exposed by inhalation to 5 ppm or more for 3 months and in mice and hamsters exposed to 45 ppm for only 1 day. Effects were seen on haematological (reduced haemoglobin, erythrocyte count, haematocrit) and clinical chemistry parameters (specially raised gamma-glutamyltransferase activity and decreases in plasma protein), liver weight increase and liver lesions (centrilobular single-cell necrosis and foci of hepatocellular alteration) in rats and mice but not hamsters. Rats exposed to 40 mg/kg body weight/day NVP or more for 3 months by gavage developed similar liver changes. Atrophy of olfactory epithelium and hyperplasia of nasal respiratory epithelium was seen in rats exposed by inhalation to 5 ppm NVP for 7 wk but not in response to 1 ppm for 13 wk (no observed-adverse-effect level, NOAEL). These studies indicated that the upper respiratory tract and the liver are the main targets for NVP toxicity.

Toxicity of aliphatic amines: structure-activity relationship.

SARs may enable the evaluation of the toxic potential of chemicals by drawing conclusions from available data on structurally-related chemicals, thus reducing the need for further testing. The Advisory Committee on Existing Chemicals of Environmental Relevance (BUA) [1,2] of the German Chemical Society (Gesellschaft Deutscher Chemiker [GDCh]) has compiled data on the toxicity and ecological impact for several groups of chemicals [3, 4]. In the present review, some common toxicological properties for aliphatic amines were revealed after evaluation and comparison of the toxicity data.

Assessment of structurally related chemicals: toxicity and ecotoxicity of acrylic acid and acrylic acid alkyl esters (acrylates), methacrylic acid and methacrylic acid alkyl esters (methacrylates).

BUA compiled the available data on toxicity and ecotoxicity for several acrylic and methacrylic acid esters and their corresponding acids. A comparison of these data revealed a qualitative similarity in the toxicological and ecotoxicological properties of the compounds considered. The data indicate that methacrylates are less reactive than the corresponding acrylates.

Developmental toxicity of dimethylacetamide in rabbits following inhalation exposure.

(1) Dimethylacetamide was tested for developmental toxicity after inhalation exposure of pregnant Himalayan rabbits. Fifteen female rabbits per main group were exposed to dimethylacetamide vapours at concentrations of 0, 0.2, 0.7 or 2.0 mg/l (equivalent to 0, 57, 199.5 or 570 ppm) and five female rabbits per satellite group to 0 or 2.0 mg/l 6 h/day from day 7 post-insemination (p.i.) to day 19 p.i. All animals were observed until day 29 p.i. (2) No signs of maternal toxicity were seen in the does of the main groups (body weight and gross pathology) or in the does of the satellite groups (body weight, blood chemistry, histopathological findings of the liver). (3) Fetotoxic effects were caused at a concentration of 0.7 mg/l (e.g., increased skeletal variations) and 2.0 mg/l (e.g., significantly decreased fetal and placental weights, increase in soft tissue and skeletal variations). At 2.0 mg/l, there were also signs of a weak teratogenic effect expressed as a marginal, statistically not significant increase in soft tissue malformations (regarding the heart and great vessels). No compound-related effects were observed in the fetuses after exposure to 0.2 mg/l. (4) The highest concentration tested under these conditions (2.0 mg/l) was found to be a no-observable-adverse-effect-level (NOAEL) for the maternal Himalayan rabbit, whereas 0.2 mg/l was defined as the NOAEL for the developing organism.

Subchronic toxicity studies of 3-methyl-1-butanol and 2-methyl-1-propanol in rats.

1. 90-day subchronic toxicity studies with 3-methyl-1-butanol (MEB) and 2-methyl-1-propanol (MEP) were performed on rats to evaluate the toxicological profile of the compounds under conditions of drinking water studies, to identify the potential target organs, and to determine no-observable-adverse-effect-levels (NOAELs) respective of the substances. The test substances were administered to groups of 10 male and 10 female Wistar rats in drinking water at concentrations of 0, 1000 p.p.m. (about 80 mg/kg/d), 4000 p.p.m. (about 340 mg/kg/d) and 16,000 p.p.m. (about 1250 and 1450 mg/kg/d of MEB and MEP respectively). 2. 16,000 p.p.m. was found to be the maximal concentration for both alcohols applicable to rats in drinking water. Higher concentrations had an influence on palatability and could thus not be tested in drinking water studies. 3. At 16,000 p.p.m. MEB a marginal increase in the red blood cell count as well as a slight decrease in the mean corpuscular volume and the mean corpuscular hemoglobin content was observed in males only. These changes are considered to be treatment-related, although the toxicological significance of these findings is unclear. No other substance-related effects were found on body weight (b.w.), mortality, various parameters of clinical chemistry, organ weights, gross pathology and histopathology. 4000 p.p.m. MEB did not cause any substance-induced changes. Therefore, the NOAEL of MEB was defined as 4000 p.p.m. for male and 16,000 p.p.m. for female rats under conditions of oral application via drinking water. 4. MEP concentrations up to and including 16,000 p.p.m. did not induce any signs of toxicity and were therefore defined as the NOAEL respective of this substance for rats under conditions of drinking water application.