TEBUCONAZOLE First draft prepared by E. Bosshard, Federal Office of Public Health, Division of Food Science, Schwerzenbach, Switzerland Explanation Evaluation for acceptable daily intake Biochemical aspects Absorption, distribution and excretion Biotransformation Toxicological studies Acute toxicity Short-term toxicity Long-term toxicity and carcinogenicity Reproductive toxicity Embryotoxicity and teratogenicity Genotoxicity Special studies Skin and eye irritation and skin sensitization Cataract induction Studies on metabolites Acute toxicity Short-term toxicity Embryotoxicity and teratogenicity Genotoxicity Observations in humans Comments Toxicological evaluation References Explanation Tebuconazole is a triazole fungicide which inhibits ergosterol biosynthesis. It was evaluated for the first time by the present Meeting. Evaluation for acceptable daily intake 1. Biochemical aspects (a) Absorption, distribution and excretion Rats were treated with single doses of 2 or 20 mg/kg bw per day of tebuconazole labelled with 14C, either uniformly in the phenyl ring or in the 3,5-triazole ring, with or without pretreatment with unlabelled compound. Tebuconazole was excreted predominantly in the bile and faeces: Male rats with biliary fistulae excreted about 90% of the dose with the bile, and faecal excretion within 72 h after administration was about 80% of the applied dose in males and about 65% in females; urinary excretion amounted to about 16% of the applied dose in males and about 35% in females. The amount excreted was not related to the administered dose. These results indicate that enterohepatic recirculation occurs in intact animals. The sex difference in excretion was observed only with the phenyl-labelled compound. After administration of triazole-labelled tebuconazole, about 70-80% of the administered dose was excreted in faeces and up to 25% in urine. Maximal plasma concentrations were reached 0.5-2 h after administration. The plasma concentration of radioactivity rose more slowly and terminal elimination from plasma proceeded faster after administration of the high dose. Less than 1% of the administered dose was recovered in the tissues two to three days after administration; liver contained the most tissue residues. Male animals in all groups had higher residue levels than females (Weber, 1987, 1988; Chopade, 1992). Uniformly labelled [14C-phenyl]-tebuconazole was administered to a lactating goat at a dose of 15 mg/kg bw per day for three consecutive days. Organ and milk samples were analysed 2 h after the last dose. Residue levels were highest in kidney (4 ppm) and liver (5 ppm); less than 0.1 ppm of residues were found in fat, muscle and milk (Lee & Wood, 1987). The excretion of tebuconazole was studied in laying hens. Of a dose of 10 mg/kg bw per day administered orally on three consecutive days, about 80% was excreted within the first 3.5 h. One-half hour after the last dose, 8 ppm were found in liver, 6 ppm in kidney and 0.15 ppm in eggs (Lee et al., 1988; Ecker & Weber, 1991). The permeability of human and rat skin in vitro to a series of 25% w/w EC formulations of tebuconazole was determined using 14C-tebuconazole and two reference compounds, [4-14C]- hydrocortisone and [4-14C]-testosterone. Within 24 h, up to 37% of a dose of 1.25 g/l tebuconazole in water had permeated human skin, as compared with about 22% of testosterone and up to 5% of hydrocortisone. In the rat, permeation by tebuconazole was lower than that by testosterone but higher than that by hydrocortisone (Brain, 1991). Groups of rats were treated dermally with doses of 0.01, 0.1, 1 or 10 mg/rat of 14C-tebuconazole in ethanol, corresponding to actual doses of 0.604, 5.86, 52.4 or 547 µg/cm2. Radioactivity was measured in the urine, faeces, blood and carcass and at the application site 0.5, 1, 2, 4, 8 and 24 h after treatment to provide an estimate of skin absorption. At doses up to 52.4 µg/cm2, about 60% of the applied dose was absorbed within 24 h; at the highest dose, about 12% was absorbed through the skin. Most of the dose was absorbed within the first few hours of application (Eigenberg, 1991). (b) Biotransformation In the study described above in which rats were treated with tebuconazole labelled with 14C either in the phenyl ring or in the 3,5-triazole ring, with or without pretreatment with unlabelled compound (Weber, 1987, 1988; Chopade, 1992), the main metabolites were the oxidation products of one of the methyl groups of the tertiary butyl moiety, i.e. the alcohol and the carboxylic acid. Metabolism in female animals resulted preferentially in simple oxidation products (e.g. hydroxy and carboxy metabolites) and then conjugation to the glucuronide and sulfate, with only minor cleavage of the triazole moiety. In male animals, the primary oxidation products were further oxidized to triol and keto acid derivatives; in addition, cleavage of triazole occurred, as indicated in trials with triazole-labelled compound. The free triazole accounted for about 5% in the urine of the males and 1.5% in that of females. Parent compound was found in only minor amounts (Ecker et al., 1987; Weber, 1987, 1988; Chopade, 1992). The proposed biotransformation pathway in rats is shown in Figure 1. In the study described above in lactating goats, the metabolic pathway was similar to that found in rats. The major metabolite identified was the tert-butyl alcohol derivative and its conjugate; the parent compound was also found (Lee & Wood, 1987). In the study in laying hens described above, hydroxylation of the tert-butyl group followed by conjugation to the sulfate was the major metabolic pathway (Lee et al., 1988; Ecker & Weber, 1991). 2. Toxicological studies (a) Acute toxicity The acute toxicity of tebuconazole is summarized in Table 1. Table 1. Acute toxicity of tebuconazole Species Sex Route LD50 (mg/kg bw) Reference or LC50 (mg/m3)a Rat (fasted) M Oral > 5000 Heimann & Pauluhn, 1983 F 3930 Rat M Oral 4260 Heimann & Pauluhn, 1983 F 3350 Rat (fasted) M Oral 4000 Ohta, 1991a F 1700 Rat M Intraperitoneal 750 Heimann & Pauluhn, 1983 F 395 Rat M&F Dermal (24 h) > 5000 Heimann & Pauluhn, 1983 Rat M&F Dermal (24 h) > 2000 Ohta, 1991b Rat M&F Inhalation (4 h)b > 818 Heimann & Pauluhn, 1983 Inhalation (5x6 h)b > 240 Rat M&F Inhalation (4 h); aerosol > 371 Pauluhn, 1988 Inhalation (4 h); dust > 5093 Mouse (fasted) M Oral 1615 Heimann & Pauluhn, 1983 F 3023 Mouse (fasted) M Oral 2800 Ohta, 1991c F 5200 Rabbit (fasted) M&F Oral > 1000 Heimann & Pauluhn, 1983 Dog (fasted) M&F Oral 625-1250 Hoffmann, 1983a Sheep (fasted) M&F Oral 625-1250 Hoffmann, 1983b a Concentrations determined analytically b In ethanol and polyethylene glycol The symptoms of acute poisoning described in rats consisted of sedation, locomotor incoordination, spastic gait and emaciation. A study was conducted in male rats to study possible combination effects of tebuconazole with either triadimenol or dichlofluanid. A slightly superadditive effect was observed in combination with triadimenol but not with dichlofluanid (Flucke, 1987).(b) Short-term toxicity Rats Groups of 20 male and 20 female Wistar rats (Bor: WISW) were given tebuconazole at doses of 0, 30, 100 or 300 mg/kg bw per day by intubation for four weeks and observed during a four-week recovery period. The dose of 300 mg/kg bw per day caused mild lethargy and reduced body-weight gain during the treatment period. Doses of 100 mg/kg bw per day and above decreased the haemoglobin concentration and haematocrit values. In females at 300 mg/kg bw per day, the leukocyte count was increased. Clinical chemistry revealed slight, not statistically significant, increases in the activities of glutamate-oxalate and glutamate-pyruvate transaminases in males at 300 mg/kg bw per day, and marked increases in liver enzyme activities (glutamate-oxalate and glutamate-pyruvate transaminases and alkaline phosphatase) in females at the same dose. Treatment at 100 and 300 mg/kg bw per day induced the microsomal enzyme system, and the activities of N- and O-demethylases and cytochrome P450 and the triglyceride concentration in liver were increased. All these changes were reversible. Urinalyses revealed no abnormal findings. At 100 and 300 mg/kg bw per day, the weights of the liver and spleen were increased in animals of each sex and the weight of the kidney in females. Histopathological findings at 300 mg/kg bw per day consisted of fatty changes in the liver and bile-duct proliferation in females; enlargement of the centrilobular hepatocytes was found in male rats. Histopathological changes were also found in the adrenal cortex, consisting of proliferated endothelial cells and an increased incidence of fat vacuoles. Sclerosis of the red pulp of the spleen, associated with sideropenia, was observed in males at 300 mg/kg bw per day; sideropenia was also found in females at 100 mg/kg bw per day. The NOAEL was 30 mg/kg bw per day on the basis of changes in haematological and clinical chemical parameters and organ weights at higher doses (Heimann & Kaliner, 1984). Groups of 10 male and 10 female Wistar rats (Bor: WISW) were exposed to nominal aerosol concentrations of tebuconazole at 0, 5, 50 or 500 mg/m3 in polyethylene glycol by head and nose exposure for 6 h/day, five days per week for three weeks (15 days). The analytical concentrations were 0, 1.2, 11 and 156 mg/m3. About 90% of the particle mass had an aerodynamic diameter of < 5 µm. The treatment had no effect on mortality rate, body-weight gain, haematological or clinical chemical parameters or organ weights; urinalysis showed no abnormal findings. Rats treated with 156 mg/m3 had piloerection after each exposure. Mixed-function oxidases in the liver were induced. At the end of the study, N-demethylase activity in the liver was increased in animals of each sex at the highest dose. Males in this group also had increased O-demethylase activity. No treatment-related gross or histopathological alterations were observed. The NOAEL was 10.6 mg/m3, equivalent to about 2 mg/kg bw per day, on the basis of liver enzyme induction at the highest concentration (Pauluhn, 1985, 1987). Groups of 10 male and 10 female rats (Wistar, BOR:WISW) were given dietary concentrations of tebuconazole (purity, 93.4%; 4.8% symmetrical isomer) at 0, 100, 400 or 1600 ppm, equal to 9, 35 or 172 mg/kg bw per day, for 13 weeks. The treatment had no effect on appearance or behaviour or the findings of ophthalmic or haematological examinations or urinalyses. Food intake was increased in animals of each sex at 1600 ppm. Retardation of body-weight gain was observed at 400 ppm in females during the first six weeks and at 1600 ppm in animals of each sex. Two animals at 1600 ppm died; no deaths occurred in the other dose groups. Few of the changes in clinical chemistry showed a dose-related or time-consistent pattern and were regarded as toxicologically insignificant. An increase in urea concentration at 1600 ppm and a decrease in triglyceride concentrations in animals at 400 ppm and above were seen only after the first four weeks and not at the end of the study. Pronounced increases in N-demethylase activity and cytochrome P450 content were found in male animals at 1600 ppm, and increased liver weights were seen in females at this dose. Histopathological examination revealed an increased incidence of intra-plasmatic vacuoles in the cells of the zona fasciculata of the adrenals (probably lipid accumulation) in some females at 400 ppm and in all females at 1600 ppm. This effect was less pronounced in males because of a higher background incidence of adrenal vacuole formation in control animals (Bomhard & Schilde, 1986). The NOAEL was 100 ppm, equal to 9 mg/kg bw per day, on the basis of retardation of body-weight gain and histopathological changes in the adrenals at higher doses Rabbits Groups of 18 male and 18 female HC New Zealand white rabbits received tebuconazole at doses of 0, 50 or 250 mg/kg bw per day on the shorn skin for 6 h daily for three weeks. The treatment was tolerated with no effects (Heimann & Schilde, 1984). In an additional study, a dose of 1000 mg/kg bw per day was also tolerated with no systemic or local effects (Heimann & Schilde, 1988). Dogs Groups of four male and four female beagle dogs were given tebuconazole (purity, 93.4%; 4.8% symmetric isomer) at dietary concentrations of 0, 200, 1000 or 5000 ppm (equal to 9, 45 or 220 mg/kg bw per day for males and females combined) over 13 weeks. One dog at 5000 ppm was found dead after the first dose, with no previous clinical signs. Treatment did not affect body temperature, pulse rate or neurological signs. Food consumption was reduced at 1000 and 5000 ppm, and body-weight gain was retarded in these groups; a few dogs at 1000 ppm and most of those at 5000 ppm had a deteriorated nutritional status. Ophthalmic examination revealed lens opacities in all animals at 5000 ppm, which first appeared after seven weeks of treatment. An increase in thrombocyte count was found at 5000 ppm, and at the end of the study six of eight animals at this dose had marked anisocytosis. Alkaline phosphatase activity in plasma showed a retarded age-induced fall at 1000 ppm and an increase at 5000 ppm. N-Demethylase activity in the liver had increased slightly by the end of the study in animals at 1000 ppm and had increased markedly in those at 5000 ppm. The cytochrome P450 content of the liver was also elevated at 5000 ppm. In the group at the highest dose, a shift in the composition of serum proteins was observed, with a slightly lower albumin content and a simultaneous increase in the ß-globulin content. Urinalyses revealed no treatment-related effects. Changes in organ weights did not follow a consistent pattern, except that an increase in spleen weights was seen in animals of each sex at 5000 ppm. Histopathological examination confirmed lens degeneration, indicating the induction of cataracts in animals at 5000 ppm. Other histopathological alterations observed in this group included slightly increased accumulation of ferriferrous pigments in Kupffer cells of the liver and of siderocytes in spleen. The NOAEL was 200 ppm, equal to 9 mg/kg bw per day, on the basis of reduced body-weight gain and food consumption and liver enzyme induction at 1000 ppm and higher. The NOAEL for cataract induction was 1000 ppm, equal to 45 mg/kg bw per day (von Keutz & Schilde, 1987a). In a one-year study, groups of four male and four female beagle dogs were given tebuconazole (purity, 96.9%) at dietary concentrations of 0, 40, 200 or 1000 ppm (weeks 1-39) and 2000 ppm (weeks 40-52), equal to 2, 10 or 60 mg/kg bw per day in males and females combined. Concentrations of up to 2000 ppm did not affect survival rate, appearance, behaviour or organ weights, and haematological examination and urinalyses showed no changes. Body temperature, pulse rates and reflexes were normal, as were food and water consumption and body-weight gain. Ophthalmic examination revealed lens opacities in two dogs at 200 ppm and one at 1000 ppm; the opacities appeared between weeks 26 and 32 and were of the same intensity at all subsequent examinations. In the single dog at 1000 ppm which showed this ocular change, corneal opacity was also found, which persisted until the end of the study, whereas the lens opacities disappeared after week 32 of treatment. The lack of a dose-response relationship with regard to lens opacities does not preclude an association with treatment but may be due to the small number of animals in the group and differences in individual sensitivity. None of the other animals in this group had lens stars; in single animals, faint lens stars were already present before treatment started but did not become more pronounced with the treatment. Incipient lens stars observed in animals at 40 and 200 ppm also remained stable, and most disappeared before the end of the treatment period. Lens stars are physiological structures found occasionally in juvenile dogs. Clinical chemical analyses revealed slight, dose-related changes in the activity of alkaline phosphatase: Whereas the age-dependent reduction in activity was similar in control animals and in those at 40 and 200 ppm, the mean activity in animals at 1000/2000 ppm indicated slight induction, resulting in a retardation in the physiological fall in alkaline phosphatase activity. The activity of N-demethylase and the triglyceride content of the liver were slightly increased in animals at 1000/2000 ppm. Most of the gross pathological findings, such as a dose-related increase in the incidence of livers with marked lobulation in animals treated with 200 ppm and above, were not correlated with histopathological alterations. Histopathological findings included intracytoplasmic vacuoles in cells of the zona fasciculata of the adrenals in animals at 200 and 1000/2000 ppm and slight siderosis in the spleen in most animals at 1000/2000 ppm. The NOAEL was 40 ppm, equal to 2 mg/kg bw per day, on the basis of cataract formation and histopathological changes in the adrenals at higher doses (von Keutz & Schilde, 1987b). In a second one-year study conducted at lower doses, groups of 12 male and 12 female beagle dogs were given tebuconazole (purity, 96%) at dietary concentrations of 0, 100 or 150 ppm, equal to 3 or 4.5 mg/kg bw per day for males and females combined. The treatment did not affect mortality, body-weight gain, food consumption, biochemical, haematological or urinary parameters, ophthalmoscopic findings, gross pathological appearance or organ weights. The only histopathological alteration was slight hypertrophy of adrenal zona fasciculata cells in all animals at 150 ppm; only one control animal had similar changes. The enlargement was accompanied by an increased incidence of large fatty vacuoles. The NOAEL was 100 ppm, equal to 3 mg/kg bw per day, on the basis of histopathological alterations in the adrenals at the higher dose. The NOAEL for cataract induction was 150 ppm, equal to 4.5 mg/kg bw per day (Porter et al., 1989). (c) Long-term toxicity and carcinogenicity Mice Groups of 50 male and 50 female NMRI mice were given tebuconazole (purity, 95%) at dietary concentrations of 0, 20, 60 or 180 ppm, equal to 6, 18 or 53 mg/kg bw per day, for 21 months. Groups of 10 treated animals of each sex were sacrificed after one year of treatment. Appearance, behaviour, mortality, food consumption and body weight were not affected by the treatment. In animals treated at 180 ppm, reduced erythrocyte count and haemoglobin and haematocrit values were observed, particularly at the end of the first year of the study. Sporadic alterations in clinical chemical parameters were observed in all groups. The few dose-dependent changes consisted of an increase in total bilirubin concentration in females treated with concentrations of 60 ppm and higher and decreased cholesterol concentrations in plasma at 180 ppm. Gross pathological examination showed no treatment-related effects. Liver weights were increased in animals at 180 ppm at interim sacrifice and at the end of the study. Histopathological examination revealed a slight increase in the prevalence of lipid-containing periportal vacuoles in the livers of animals treated with 60 or 180 ppm at both interim and terminal sacrifice. No increase in tumour incidence was found. The NOAEL was 20 ppm, equal to 6 mg/kg bw per day, on the basis of histopathological changes in the liver (Bomhard & Ramm, 1988a). In a similar study using higher doses, groups of 50 male and 50 female NMRI mice were maintained on a diet containing tebuconazole (purity, 95%) at concentrations of 0, 500 or 1500 ppm, equal to 85 or 280 mg/kg bw per day, for 21 months. Groups of 10 animals per sex and dose were scheduled for interim sacrifice after 12 months. The mortality rate was not affected by treatment. The incidence of animals with a 'distended abdomen' (probably due to liver enlargement) was increased among those at 1500 ppm. A dose-dependent reduction in body-weight gain was observed in males at 500 and 1500 ppm and in females at 1500 ppm, particularly during the first half of the study. Food intake tended to be higher in treated animals than in controls. Alterations in haematological parameters were found in both treated groups: At 500 ppm and above, reduced mean cell volume and haematocrit values and elevated mean cell haemoglobin concentrations were found in males, and thrombocyte counts were elevated in females. Thromboplastin time was reduced in both treated groups. At 1500 ppm, erythrocyte counts, haemoglobin content and haematocrit values were markedly reduced and thrombocyte counts were elevated, particularly in males. Leukocyte counts were also increased. Parameters of clinical chemistry that showed dose-related changes in both treated groups included pronounced increases in the activities of glutamate-oxalate and glutamate-pyruvate transaminases and alkaline phosphatase, particularly at 1500 ppm. Cholesterol, bilirubin and albumin levels were reduced at 500 and 1500 ppm, but there was no clear dose-response relationship. Liver weights were increased in a dose-related fashion at both doses, and adrenal weights were elevated in females at 1500 ppm. The gross pathological evidence for treatment-related effects on the liver consisted of enlargement and pale appearance of the liver in both treated groups and some enhanced lobulation. Capsular thickening and swelling and an increased incidence of nodular masses were also found at 1500 ppm. Histopathological examination revealed treatment-related alterations, mainly in the liver. The non-neoplastic findings consisted of single-cell and focal necroses, focal inflammation, hepatocytic degeneration, hepatocytic hyerplasia and hypertrophy, fatty vacuolation, bile-duct hyperplasia and steatoses, oval-cell proliferation, extramedullary haematopoiesis and pigment accumulation in Kupffer cells. Many of these alterations were found at both 500 and 1500 ppm. No evidence was found for a compound-related effect on the incidence of benign or malignant tumours after one year of treatment. Increased incidences of tumour-bearing animals were seen, however, at 1500 ppm after 21 months of treatment. Increases were seen in the incidences of both benign and malignant tumours in males and of malignant tumours in females. Statistically significant increases in tumour incidence were found only in the liver. An increased incidence of adenomas was observed at 1500 ppm, with 36% (versus 6% in controls) in males and 4% (versus 0 in controls) in females; the incidences of carcinomas at this dose were 21% (versus 0 in controls) in males and 26% (versus 2% in controls) in females. The incidences of hepatocellular tumours in animals at 1500 ppm were reported to be above those in historical control mice of this strain. The only reported incidences were 0-12% for adenoma and 0-10% for carcinoma in males. The NOAEL was < 500 ppm, equal to < 85 mg/kg bw per day, on the basis of reduced body-weight gain and histopathological changes in the liver. The NOAEL for carcinogenicity was 500 ppm, equal to 85 mg/kg bw per day, on the basis of an increased incidence of liver tumours at 1500 ppm (Bomhard, 1991). Rats Groups of 50 male and 50 female Wistar rats (Bor:WISW) were given tebuconazole (purity, 95%) in the diet at concentrations of 0, 100, 300 or 1000 ppm, equal to 5, 16 or 55 mg/kg bw per day, for two years. Groups of 10 animals per sex and dose were similarly treated and killed at interim sacrifice after 12 months of treatment. Treatment did not affect mortality, appearance, general behaviour of the animals or haematological parameters, and urinalyses showed no changes; no treatment-related damage to the eyes was found. Body-weight gains were lower than those of the control animals at doses of 300 ppm and above. Food consumption was increased among females at 1000 ppm, and water consumption was decreased in females at 300 ppm and above. Clinical examinations revealed several statistically significant changes but no consistent dose-related pattern indicative of an association with treatment, with the probable exception of a marginal increase in the activity of alanine aminotransferase at 1000 ppm. Decreased triglyceride concentration was found in female animals at 1000 ppm, and an increase in spleen weight was found in these animals. Gross pathological examination did not provide any indication of treatment-related effects. Histopathological examination revealed an increased incidence of pigmented Kupffer cells in liver and increased haemosiderin accumulation in the spleen of females at 1000 ppm. The incidence of C-cell adenomas and carcinomas of the thyroid was increased in all treated males in comparison with that in controls (0/50), with 2/49 (4%) at 100 ppm, 3/50 (6%) at 300 ppm and 3/50 (6%) at 1000 ppm There was no clear dose-response relationship, and the rates were within the range of spontaneously occurring thyroid C-cell tumours in male Wistar rats. The mean historical incidences of interstitial-cell and C-cell adenomas grouped with parafollicular tumours were reported to be 7.4% (0-19.3) in males and 8.4% (2.5-21.2) in females (Bomhard et al., 1986). The incidence of 0 in concurrent controls is unusually low. In female rats, a higher frequency of endometrial adenocarcinoma was found in comparison with controls (0/50), with incidences of 3/50 at 100 ppm, 2/50 at 300 ppm and 1/50 at 1000 ppm These incidences were also small and not dose-related. The reported historical incidence of uterine adenocarcinomas varied considerably, from 0 to 14.4%, with a mean incidence of 6.3% (Bomhard et al., 1986). The fact that the endometrial tumours found in this study differ morphologically from the typical uterine carcinomas that occur spontaneously in this rat strain precludes a direct comparison, either qualitatively or quantitatively. The absence of a dose-response relationship may indicate, however, that these findings were not treatment-related. The NOAEL was 100 ppm, equal to 5 mg/kg bw per day, on the basis of reduced body-weight gain at higher doses (Bomhard & Ramm, 1988b). (d) Reproductive toxicity Rats Groups of 25 male and 25 female Wistar rats (Bor:WISW) were treated with tebuconazole (purity, 95%) at dietary concentrations of 0, 100, 300 or 1000 ppm, equal to 7, 22 or 72 mg/kg bw per day, over two generations. Treatment did not affect appearance, behaviour, general condition or mortality rates, and fertility and gestation indexes were not altered. At 1000 ppm, the body-weight gain of the parent animals was reduced; food consumption was reduced only among males at the highest dose in the F0 generation and in animals of each sex in the F1 generation. Reductions in the mean litter size at birth, in the viability index (survival until day 5 after birth) and in the lactation index were seen at 1000 ppm, and mean birth weights (only in the F1a generation) and body-weight gain (in both generations) were reduced. None of the pups had grossly apparent malformations, and no treatment-related organotoxic effects were noted on gross and microscopic examination. The NOAEL was 300 ppm, equal to 22 mg/kg bw per day, on the basis of reduced body-weight gain in parents and pups and reduced litter size at the highest dose (Eiben, 1987). (e) Embryotoxicity and teratogenicity Mice Groups of 25 female NMRI/ORIG Kisslegg mice were treated by gavage with tebuconazole (purity, 93.6%) at a dose of 0, 10, 30 or 100 mg/kg bw per day on days 6-15 of gestation. The treatment did not affect mortality rates, body-weight gain or pregnancy rates of the maternal animals. At 30 mg/kg bw per day and higher, a dose-related increase in the incidence of runts was observed, with 20/234 (8.6%) at 30 mg/kg bw per day and 26/202 (13%) at 100 mg/kg bw per day; there were 4/234 (2%) at 10 mg/kg bw per day and 5/236 (2%) in the control group. An increased incidence of malformations was found at 100 mg/kg bw per day, 13/202 (6.5%), which consisted most frequently of cleft palates and individual cases of micrognathia, rib fusion and spinal dysplasia; the incidence of malformations in the control group was 1/236 (0.4%). The incidence of cleft palate at this dose, 6/202 (3%), was also markedly higher than the mean incidence in historical controls, 0.7%. The NOAEL for maternal toxicity and clinical signs was 100 mg/kg bw per day and that for embryotoxicity and teratogenicity was 10 mg/kg bw per day (Renhof, 1988a). In a study to investigate maternal toxicity further, groups of 10 female NMRI/ORIG Kisslegg mice were treated by gavage with tebuconazole (purity, 97.4%) at doses of 0, 10, 20, 30 or 100 mg/kg bw per day on days 6-15 of gestation. Haematological and clinical chemical tests showed no effect of treatment on mortality rates or body weights of the maternal animals; slight reductions in the haematocrit and mean corpuscular volume were seen at 30 and 100 mg/kg bw per day, but with no clear dose-response relationship. The activities of alanine and aspartate transaminases showed slight, usually statistically significant increases in all treated groups but with no consistent relation to dose. Triglyceride levels were increased at 100 mg/kg bw per day. Liver weights were increased at all doses, again with no clear dose-response relationship. Histopathological investigation revealed cytoplasmic vacuolation of the liver at 100 mg/kg bw per day, which was correlated with an increase in lipid content. The dose of 100 mg/kg bw per day thus produced pronounced liver toxicity; the changes in liver enzyme activities, unrelated to dose, in all treated groups indicate that maternal toxicity may have been induced at 10 mg/kg bw per day. The embrotoxic and teratogenic effects thus occur at maternally toxic doses. The NOAEL for maternal toxicity was < 10 mg/kg bw per day (Renhof & Karbe, 1988). Groups of 25 female NMRI/HAN mice were given tebuconazole (purity, 96-98%; different batches used for main and supplementary study) at daily dermal doses of 0, 100, 300 or 1000 mg/kg bw per day for 6 h/day on days 6-15 of gestation. Treatment had no effect on mortality rates, body weight, food consumption or reproductive parameters (numbers of corpora lutea, implantations, resorptions and live fetuses). Examination of the litters revealed no treatment-related effect on mean body weight, and no abnormal findings were detected in the viscera. The frequency of external abnormalities (cleft palate, malposition of hind legs, exencephaly and 'tail cranial bended') at 1000 mg/kg bw per day was not different from that in the control group, with 3% in the controls, 3% at 100 mg/kg bw per day, 1.8% at 300 mg/kg bw per day and 5.3% at 1000 mg/kg bw per day. The increase at the highest dose was due to a higher incidence of cleft palate, with 11/285 (3.9%) at 1000 mg/kg bw per day and 6/301 (2%) in the control group. The incidence of cleft palate in control animals in an independent study was 1.6%. Skeletal examination showed an increased number of supernumerary ribs at 1000 mg/kg bw per day. The NOAEL was 1000 mg/kg bw per day for maternal toxicity and clinical signs and 300 mg/kg bw per day for embryotoxicity and teratogenicity (Becker et al., 1990). Since treatment-related effects on the fetuses appeared to have occurred in the absence of maternal toxicity, a study was initiated that included biochemical and histopathological examinations. Tebuconazole was administered in the same way as described above to groups of 10 mated females. As in the first study, no deaths occurred and no clinical signs were observed in animals treated at 1000 mg/kg bw per day; the treatment had no effect on body-weight gain, food consumption or reproductive parameters. Clinical biochemical examinations revealed increased alanine transaminase activity at 1000 mg/kg bw per day; there was also increased cytochrome P450 content and N-demethylase activity in liver tissue at > 300 mg/kg bw per day and a marginal increase in O-demethylase activity. None of these parameters showed a dose-response relationship. There was a dose-related reduction in relative adrenal weights in all dose groups, which was statistically significant at 1000 mg/kg bw per day; absolute adrenal weights were also reduced in all treatment groups, but not in relation to dose. No effect was seen on liver weights, but histological examination revealed fatty changes in periportal areas in most mice at 300 mg/kg bw per day and in all mice at 1000 mg/kg bw per day. The NOAEL was 100 mg/kg bw per day on the basis of fatty changes in the liver at higher doses (Becker et al., 1990). Rats In a range-finding study, groups of 25 female Wistar rats were treated by gavage with tebuconazole (purity, 99.5%) at doses of 0 or 100 mg/kg bw per day on days 6-15 of gestation. The treatment had no effect on mortality rates, but body-weight gain was retarded. The fertilization rate was not reduced by treatment, but most litter parameters (such as number of implantations, litter size and losses) indicated an adverse effect. More runts and fetuses with malformations (micrognathia, hydronephrosis and hydroureter) were found in treated animals. The NOAEL for maternal toxicity, embryotoxicity and teratogenicity was < 100 mg/kg bw per day (Renhof, 1984). In a similar study, groups of 25 female Wistar rats were treated by gavage with tebuconazole (purity, 93%) at doses of 0, 10, 30 or 100 mg/kg bw per day on days 6-15 of gestation. No treatment-related deaths occurred. At 30 and 100 mg/kg bw per day, dose-related reductions in weight gain were observed throughout the treatment period; at 100 mg/kg bw per day, retarded body-weight gain was observed throughout gestation. Effects on litter parameters included an increased number of losses and a decrease in mean fetal weight at 100 mg/kg bw per day. At this dose there were also more runts and fetuses with malformations (mostly microphthalmia): the runt incidence was 36%, compared with 7% in controls, and the incidence of fetuses with malformations was 7%, compared with 2% in controls. The NOAEL for maternal toxicity was 10 mg/kg bw per day and that for embrotoxicity and teratogenicity was 30 mg/kg bw per day (Renhof, 1985a). Groups of 25 femaleWistar rats were treated by gavage with tebuconazole (purity, 98.3%) at doses of 0, 30, 60 or 120 mg/kg bw per day on days 6-15 of gestation. The dose of 30 mg/kg bw per day affected neither maternal nor fetal parameters. At 60 mg/kg bw per day and above, food consumption was reduced and a dose-related loss of body weight was seen at the beginning of the treatment period. Liver weights showed a dose-related increase at 60 and 120 mg/kg bw per day. Reproductive parameters (numbers of corpora lutea and implantations) were not adversely affected. At 120 mg/kg bw per day, resorptions occurred more frequently, and consequently there were fewer live fetuses than in the control group; mean fetal body weights were lower. This dose induced increased incidences of visceral alterations (consisting of excess fluid in the thoracic cavity), supernumerary ribs, non-ossified cervical vertebrae and incompletely ossified sternebrae, indicating retardation of fetal development. The NOAEL was 30 mg/kg bw per day for maternal toxicity and 60 mg/kg bw per day for embryotoxicity (Becker et al., 1988a). Groups of 25 female Wistar rats were treated dermally with tebuconazole (purity, 97.4%) at doses of 0, 100, 300 or 1000 mg/kg bw per day for 6 h/day on days 6-15 of gestation. The treatment was tolerated with no observed effects on dams or fetuses with respect to the mortality rate, body-weight gain, gestation rate or other reproductive parameters, such as numbers of corpora lutea, resorptions and live fetuses. There was no indication of a treatment-related effect on the incidence or spectrum of malformations (Renhof, 1988b). Rabbits Groups of 15 female Himalayan CHBB:HM rabbits were treated by gavage with tebuconazole (purity, 93%) at doses of 0, 3, 10 or 30 mg/kg bw per day on days 6-18 of gestation. Dams given the highest dose had reduced body-weight gain during the treatment period. Reproductive and fetal parameters were not adversely affected, and no treatment-related increases in the incidence of malformations were observed. The NOAEL was 10 mg/kg bw per day for maternal toxicity and 30 mg/kg bw per day for embryotoxicity and teratogenicity (Renhof, 1985b). Groups of 16 female CHIN hybrid rabbits were treated by gavage with tebuconazole at doses of 0, 10, 30 or 100 mg/kg bw per day on days 6-18 of gestation. Treatment-related effects were seen at 100 mg/kg bw per day, consisting of reduced body-weight gain and food consumption, increased mean post-implantation loss and slightly reduced mean fetal body weight. External examination of fetuses showed an increased incidence of malformations, including peromelia (the most frequent malformation), agenesis of claws and cleft palate. Of 90 fetuses born to dams treated at 100 mg/kg bw per day, eight (9%) were malformed, and five of these (6%) had peromelia. Peromelia was seen in none of a total of 346 fetuses in the other three groups combined. At 100 mg/kg bw per day, the percentage of fetuses with absent or incomplete phalangeal ossification was also increased. In combination with the reduction in mean fetal body weight, these findings indicate delayed maturation of fetuses at the highest dose. The NOAEL was 30 mg/kg bw per day for maternal toxicity, embryotoxicity and teratogenicity (Becker et al., 1988b). (f) Genotoxicity Numerous studies have been conducted on the genotoxicity of tebuconazole. The results are summarized in Table 2. No genotoxic activity was found in any study. (g) Special studies (i) Skin and eye irritation and skin sensitization Tebuconazole did not irritate the skin of rabbits, and administration of 50 mg into the conjunctival sac did not induce irritation of the eye (Heimann & Pauluhn, 1983; Sheets, 1988; Märtins, 1990). In another study, instillation of 100 mg into the conjunctival sac of rabbits caused slight irritation of the conjunctiva (Eigenberg, 1988). Tebuconazole did not reveal skin-sensitizing potential in a maximization test in guinea-pigs (Heimann, 1983). The negative result was confirmed in two further tests for sensitization in guinea-pigs, but with the less sensitive Buehler method (Heimann, 1987; Sheets, 1990). Table 2. Results of tests for the genotoxicity of tebuconazole End-point Test system Concentration Purity Results Reference of tebuconazole (%) In vitro Reverse S. typhimurium TA98, 20-12 500 µg/platea 97 Negative Herbold, 1983a, mutation 100, 1535, 1587 1990 Reverse S. typhimurium TA98, 37.5-2400 µg/plateb 96.6 Negative Herbold, 1988a,b mutation 100, 1535, 1537, 1538 Reverse S. typhimurium 15.6-500 µg/plate 98 Negative Ohta, 1991d mutation E. coli (WPuvrA) 31.3-1000 µg/platec 156-5000 µg/plated Forward Chinese hamster 80-100 µg/mle 96.6 Negative Lehn, 1988 mutation ovary cells 12.5-200 µg/mlc (hprt locus) Recombination B. subtilis (spores) 0.3-20 µg/disc 98 Negativec,d Ohta, 1992 repair capacity H17 (rec+), M45 (rec-) Cytogenicity Human lympho cytes 3-30 µg/mlc 96.6 Negative Herbold, 1988c 30-300 µg/mld DNA polymerase E. coli 625-10 000 µg/plate 97.1 Negativec,d Herbold, 1983b repair capacity (K12)p 3478 (pol-) W 3110 (pol+) Unscheduled Rat hepatocytes 0.5-25.2 µg/ml 96.5 Negative Cifone, 1988 DNA synthesis Sister chromatid Chinese hamster 4-30 µg/mlc 96.5 Negative Putman, 1987 exchange ovary cells 15-120 µg/mld Table 2 (contd) End-point Test system Concentration Purity Results Reference of tebuconazole (%) In vivo Micronucleus Mouse bone marrow Single oral dose: 2000, 95.3 Negative Herbold, 1985 formation 500 or 200 mg/kg bwf Dominant lethal Male mouse Single oral dose: 93.5 Negative Herbold, 1986 mutation 2000 mg/kg bw a Toxic at doses > 500 µg/plate b Toxic at doses > 60 µg/plate c In the absence of metabolic activation d In the presence of metabolic activation e Toxic at doses > 75 µg/ml with and without activation f Doses reduced in second and third trials owing to inhibition of erythropoiesis by the test compound (ii) Cataract induction Groups of four male and four female Forest of Dean cats received whole-body exposure to tebuconazole (purity, 95.8%) in polyethylene glycol and ethanol at mean analytical concentrations of 61 or 309 mg/m3 (99% of particles with an aerodynamic diameter of <5 µm) for 6 h/day for four weeks, followed by an observation period of 15 weeks. Control animals were exposed to aerosols of either the vehicle or about 20 mg/m3 Sclex (KNJ 0953) (positive controls). The treatment did not induce clinical symptoms and did not affect mortality rates or body-weight gain. Cataracts due to lens fibre degeneration were found in all animals in the positive control group during the observation period. Exposure to tebuconazole did not result in cataract induction, but three females at 309 mg/m3 and one positive control animal had yellow-tinged spots along the lens fissure. Examination of 42 untreated female cats aged 7-12 months showed that these ocular changes were not common spontaneous alterations; no such finding was seen in vehicle controls. The etiology of this finding and its toxicological relevance remain unclear. The NOAEL was 61 mg/m3, equivalent to about 5 mg/kg bw per day, on the basis of ocular effects other than cataracts of unknown etiology at the highest concentration (Märtins et al., 1990). Groups of four female beagle dogs were treated by head-nose exposure to tebuconazole (purity, 97.1%) in polyethylene glycol and ethanol at target aerosol concentrations of 0, 150 or 800 mg/m3 for 4 h/day for six weeks and observed for eight weeks. The analytical concentrations were 163 and 914 mg/m3; about 90% of the particles had an aerodynamic diameter of < 3 µm. No vehicle control group was included in the study. The treatment did not affect mortality rates. Most animals at 914 mg/m3 began salivating immediately after exposure, and single animals also made tussive noises; these effects were reversed within 2 h. Retardation of body-weight gain was observed in both treated groups during the second half of the study, but with no dose-effect relationship. A lung function test revealed a marginal decrease in the mean minute volume in both treated groups and a slight reduction in the mean partial pressure of oxygen. The body temperature of treated animals was reduced during exposure, probably due to central nervous system depression caused by the ethanol vehicle. Increased spleen weight observed at 914 mg/m3 was the only change in organ weights. No gross pathological or histopathological changes were observed, and ophthalmic examinations gave no indication of cataract formation. The NOAEL was 163 mg/m3, equivalent to 23 mg/kg bw per day, on the basis of clinical effects. The NOAEL for cataract induction was 914 mg/m3, equivalent to 125 mg/kg bw per day (Märtins, 1991). 3. Studies on metabolites (a) Acute toxicity The acute toxic effects of several metabolites of tebuconazole are summarized in Table 3. Table 3. Acute toxicity of metabolites of tebuconazole Metabolite Species Sex Route LD50 (mg/kg bw)a Reference Carboxy metabolite Rat F Oral > 4000 Astroff & Hagen, 1992 Hydroxy metabolite Rat F Oral > 5000 Sheets & Phillips, 1992 1H-1,2,4-triazole Rat M Oral 1375 Flucke, 1978 Rat M&F Oral 1650 Thyssen & Kimmerle, 1976 a Concentrations determined analytically (b) Short-term toxicity In a three-month study, groups of 15 male and 15 female Wistar rats were fed diets containing 1 H-1,2,4-triazole (purity, 99.6%) at concentrations of 0, 100, 500 or 2500 ppm, equal to 8, 38 or 212 mg/kg bw per day. Appearance, mortality rates, behaviour, clinical chemistry parameters and the results of urinalysis, thyroid function tests (protein-bound iodine) and gross pathological examination were not affected by the treatment. At 2500 ppm, temporary convulsions occurred; body-weight gain and food consumption were reduced, particularly in males; and haematological parameters (haemoglobin concentration, haematocrit, mean corpuscular volume and mean corpuscular haemoglobin) were reduced in males. Histopathological changes, confined to the liver, consisted of fatty accumulation in three males at 2500 ppm (Bomhard et al., 1979). (c) Embryotoxicity and teratogenicity Groups of 25 pregnant Bor:WISW Wistar rats were treated orally with 1,2,4-triazole (purity, 95.3%) on days 6-15 of gestation at doses of 0, 10, 30 or 100 mg/kg bw per day. Treatment at 100 mg/kg bw per day caused a reduction in body-weight gain of the dams, reduced fetal weight and an increased number of runts. At the same dose, the incidence of malformations, consisting mainly of eye deformities such as anophthalmia and micro-phthalmia, was increased over that in concurrent or historical controls (Renhof, 1988c). In a similar study, groups of 25 pregnant Bor:WISW Wistar rats were treated orally with 1,2,4-triazole at doses of 0, 100 or 200 mg/kg bw per day. The treatment impaired body-weight gain in both treated groups but was statistically significant only at 200 mg/kg bw per day. At this dose, an increased number of resorptions and a reduction in fetal weight were observed. In both treated groups, a dose-related increase in the incidence of runts was observed; the incidence of malformations (mainly hydronephrosis and cleft palate) was increased only at 200 mg/kg bw per day (Renhof, 1988d). (d) Genotoxicity The 1 H-1,2,4-triazole metabolite of tebuconazole (purity, 99.7%) was tested for its ability to induce reverse mutation in Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537. Doses of 10-5000 µg/plate were ineffective, in either the presence or the absence of metabolic activation (Poth, 1989). 4. Observations in humans Employees working in the production of tebuconazole, who were under continous medical supervision, did not show exposure-related effects (Kollert, 1987). Comments After oral administration of tebuconazole to rats, 65-80% of the dose was eliminated by the biliary and faecal route, whereas elimination in urine amounted to about 16-35%. Males had a greater biliary and faecal elimination than females. Biotransformation proceeded by oxidation reactions, resulting in hydroxy, carboxy, triol and ketoacid metabolites and conjugates as well as triazole. Administration of acute oral doses to rats induced sedation, abnormal gait and emaciation. Tebuconazole has low acute toxicity and has been classified by WHO (1992) as unlikely to present an acute hazard in normal use. In a 13-week study, rats were fed tebuconazole at concentrations of 0, 100, 400 or 1600 ppm. The NOAEL was 100 ppm, equal to 9 mg/kg bw per day, on the basis of retardation of body-weight gain and histopathological changes in the adrenal glands at higher doses. In two one-year studies in dogs fed diets containing 0, 40, 200 or 1000 ppm or 0, 100 or 150 ppm, an NOAEL of 100 ppm for the two studies combined, equal to 3 mg/kg bw per day, was determined on the basis of histopathological alterations in the adrenal glands at 150 ppm and above and cataract production at 200 ppm and above. Two 21-month studies of toxicity and carcinogenicity were conducted in mice, at dietary concentrations of 0, 20, 60 or 180 ppm or 0, 500 or 1500 ppm. The NOAEL was 20 ppm, equal to 6 mg/kg bw per day, on the basis of fatty changes in the liver. At 1500 ppm, pronounced liver toxicity and an increased incidence of liver tumours were observed. This tumorigenic potential is not considered relevant to humans. In a two-year study of toxicity and carcinogenicity in rats treated at dietary concentrations of 0, 100, 300 or 1000 ppm, the NOAEL was 100 ppm, equal to 5 mg/kg bw per day, on the basis of reduced body-weight gain at higher doses. There was no evidence of carcinogenicity. In a two-generation study in rats fed dietary concentrations of 0, 100, 300 or 1000 ppm, the NOAEL was 300 ppm, equal to 22 mg/kg bw per day, on the basis of reduced body-weight gain in the parental generation and adverse effects on litters at higher dose. Teratogenicity was investigated in mice, rats and rabbits. In mice, doses of 0, 10, 20, 30 or 100 mg/kg bw per day were administered. Increased enzyme activities in liver were observed at all doses but were not dose-related, so that there was no clear NOAEL for maternal toxicity. In mice fed doses of 0, 10, 30 or 100 mg/kg bw per day, a higher incidence of runts was seen at 30 mg/kg bw per day and a higher incidence of malformations (mainly cleft palate) at 100 mg/kg bw per day. The NOAEL for embryotoxicity and teratogenicity was thus 10 mg/kg bw per day. In rats treated with doses of 0, 10, 30 or 100 mg/kg bw per day or 0, 30, 60 or 120 mg/kg bw per day, the NOAEL was 10 mg/kg bw per day for maternal toxicity on the basis of reduced body-weight gain at higher doses. Embryotoxicity and an increased incidence of malformations (mainly microphthalmia) and visceral and skeletal variations were found at 100 mg/kg bw per day and above, giving an NOAEL for embryotoxicity and teratogenicity of 60 mg/kg bw per day. In rabbits treated at doses of 0, 3, 10 or 30 mg/kg bw per day or 0, 10, 30 or 100 mg/kg bw per day, the NOAELs were 10 mg/kg bw per day for maternal toxicity and 30 mg/kg bw per day for embryotoxicity and teratogenicity. At 100 mg/kg bw per day, embryotoxicity and an increased incidence of external malformations (mainly peromelia) were observed. Tebuconazole has been studied in a range of tests for genotoxicity in vivo and in vitro. The Meeting concluded that there was no evidence of genotoxicity. An ADI was established on the basis of an NOAEL of 100 ppm in a one-year dietary study in dogs and a 100-fold safety factor. Toxicological evaluation Levels that cause no toxic effect Mouse: 20 ppm, equal to 6 mg/kg bw per day (21-month study of toxicity and carcinogenicity) < 10 mg/kg bw per day (maternal toxicity in a study of teratogenicity) 10 mg/kg bw per day (embryotoxicity in a study of teratogenicity) Rat: 100 ppm, equal to 5 mg/kg bw per day (two-year study of toxicity and carcinogenicity) 10 mg/kg bw per day (maternal toxicity in a study of teratogenicity) 60 mg/kg bw per day (embryotoxicity in a study of teratogenicity) Rabbit: 10 mg/kg bw per day (maternal toxicity in a study of teratogenicity) 30 mg/kg bw per day (embryotoxicity in a study of teratogenicity) Dog: 100 ppm, equal to 3 mg/kg bw per day (one-year study) Estimate of acceptable daily intake for humans 0-0.03 mg/kg bw Studies that would provide information useful for continued evaluation of the compound Further observations in humans References Astroff, A.B. & Hagen, L.L. (1992) Acute toxicity study with HWG 2443 (a metabolite of Tebuconazole Folicur(TM)) in female rats. Unpublished report Ref. No. 92-012-PB, prepared by Miles Inc., Agricultural Division, Toxicology, Stillwell, KS, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Becker, H., Vogel, W. & Terrier, C. (1988a) Embryotoxicity study (including teratogenicity) with HWG 1608 technical in the rat. Unpublished report Ref. No. R 4451, prepared by Research & Consulting Co. AG (RCC), Itingen, Switzerland. Submitted to WHO by Bayer AG, Leverkusen, Germany. Becker, H., Vogel, W. & Terrier, C. (1988b) Embryotoxicity study (including teratogenicity) with HWG 1608 technical in the rabbit. Unpublished report Ref. No. R 4323, prepared by Research & Consulting Co. AG (RCC), Itingen, Switzerland. Submitted to WHO by Bayer AG, Leverkusen, Germany. Becker, H., Vogel, O., Terrier, C., Biedermann, K. & Luetkemeier, H. (1990) Embryotoxicity study (including teratogenicity) with HWG 1608 technical in the mouse (dermal application). Unpublished report Ref. No. R 5116, prepared by Research & Consulting Co. AG (RCC), Itingen, Switzerland. Submitted to WHO by Bayer AG, Leverkusen, Germany. Bomhard, E. (1991) HWG 1608--Toxic dose range carcinogenicity study in NMRI mice (supplement to study T 601 89 53 with administration in diet over a 21-month period). Unpublished report Ref. No. 16376 A, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Bomhard, E. & Ramm, W. (1988a) HWG 1608--Study for cancerogenicity in NMRI mice (administration in diet for up to twenty-one months). Unpublished report Ref. No. 16376, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Bomhard, E. & Ramm, W. (1988b) HWG 1608--Study for chronic toxicity and cancerogenicity in Wistar rats (administration in diet for two years). Unpublished report Ref. No. 16375, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Bomhard, E. & Schilde, B. (1986) HWG 1608--Subchronic toxicological study with rats (feeding for thirteen weeks). Unpublished report Ref. No. 15211, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Bomhard, E., Löser, E. & Schilde, B. (1979) 1,2,4-Triazole: Subchronic toxicological study with rats (feeding study over three months). Unpublished report Ref. No. 8667, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Bomhard, E., Karbe, E. & Loeser, E. (1986) Spontaneous tumours of 2000 Wistar TNO/W. 70 rats in two-year carcinogenicity studies. J.E.P.T.O. 7: 1/2, pp. 35-52. Brain, K.R. (1991) In vitro skin permeability of tebuconazole. Unpublished report Ref. No. M 7591, prepared by An-eX Analytical Services Ltd., Cardiff, United Kingdom. Submitted to WHO by Bayer AG, Leverkusen, Germany. Chopade, H.M. (1992) Addendum I. [Phenyl-U-14C] HWG 1608: Study of biokinetic behavior in the rat. Response to EPA requests and inquiries. Unpublished report Ref. No. MR 97439-1 Metab I/2, prepared by Miles Inc., Stilwell, KS, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Cifone, M.A. (1988) Mutagenicity test on HWG 1608 techn. in the rat primary hepatocyte unscheduled DNA synthesis assay. Unpublished report Ref. No. R 4111a, prepared by Hazelton Laboratories America Inc., Kensington, MD, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Ecker, W. & Weber, H. (1991) [Chlorophenyl-U-14C] tebuconazole absorption, distribution, excretion and metabolism in laying hens. Unpublished report Ref. No. PF 3587, prepared by Bayer AG, Leverkusen, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Ecker, W., Brauner, A., Klein, O. & Weber, H. (1987) Folicur: Metabolism part of the general metabolism study in the rat. Unpublished report Ref. No. PF 2907 and MR 97438. Metab I/3, prepared by Bayer AG, Leverkusen, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Eiben, R. (1987) HWG 1608--Two-generation study in rats. Unpublished report Ref. No. 16223, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Eigenberg, D.A. (1988) Primary eye irritation of Folicur (HWG 1608) technical in albino rabbits. Unpublished report Ref. No. 1003, prepared by Mobay Corp., Stilwell, KS, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Eigenberg, D.A. (199) Dermal absorption of 14C HWG 1608 technical in rats. Unpublished report Ref. No. MR 97470. Metab I/5, prepared by Mobay Corp., Stilwell, KS, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Flucke, W. (1978) Determination of LD 50--Compound: 1,2,4-Triazole. Unpublished letter report by Bayer AG, Institute of Toxicology. Submitted to WHO by Bayer AG, Leverkusen, Germany. Flucke, W. (1987) HWG 1608 and KWG 0519 (c.n. Triadimenol)/HWG 1608 and KUE 13032c (c.n. Dichlofluanid) combination toxicity study. Unpublished report Ref. No. 15747, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Heimann, K.G. (1983) HWG 1608--Study for skin-sensitising effect on guinea pigs. Unpublished report Ref. No. 12024, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Heimann, K.G. (1987) HWG 1608 technical--Study of skin sensitization effect on guinea pigs (Buehler patch test). Unpublished report Ref. No. 16238, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Heimann, K.G. & Kaliner, G. (1984) HWG 1608--Study of the subacute oral toxicity to rats. Unpublished report Ref. No. 13028, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Heimann, K.G. & Pauluhn, J. (1983) HWG 1608 study for acute toxicity. Unpublished report Ref. No. 12168, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Heimann, K.G. & Schilde, B. (1984) HWG 1608--Subacute study of dermal toxicity to rabbits. Unpublished report Ref. No. 12669, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Heimann, K.G. & Schilde, B. (1988) HWG 1608 technical--Subacute dermal study of toxicity to rabbits. Unpublished report Ref. No. 12669a (Addendum to report No. 12669), prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Herbold, B.A. (1983a) HWG 1608--Salmonella/microsome test to evaluate for point mutagenic effect. Unpublished report Ref. No. 12086, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Herbold, B.A. (1983b) HWG 1608--Pol test on E. coli to evaluate for harmful effects on DNA. Unpublished report Ref. No. 11902, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Herbold, B.A. (1985) HWG 1608--Micronucleus test on the mouse to evaluate for mutagenic effect. Unpublished report Ref. No. 13159, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Herbold, B.A. (1986) HWG 1608--Dominant lethal test on the male mouse to evaluate for mutagenic effect. Unpublished report Ref. No. 14985, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Herbold, B.A. (1988a) HWG 160--Salmonella/microsome test to evaluate for point mutagenic effects. Unpublished report Ref. No. 16383, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Herbold, B.A. (1988b) HWG 1608--Salmonella/microsome test using TA 1538 to evaluate for point mutagenic effects (addendum to final report Ref. No. 16383). Unpublished report Ref. No. 16383 A, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Herbold, B.A. (1988c) In vitro cytogenetic study with human lymphocytes for the detection of induced clastogenic effects. Unpublished report Ref. No. 16395, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Herbold, B.A. (1990) HWG 1608--Salmonella/microsome test for point-mutagenic effects (appendix to Report 12086). Unpublished report Ref. No. 12086A, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Hoffmann, K. (1983a) HWG 1608 acute toxicity to the dog after oral administration. Unpublished report Ref. No. 11974, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Hoffmann, K. (1983b) HWG 1608 acute toxicity to the sheep after oral administration. Unpublished report Ref. No. 11970, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. von Keutz, E. & Schilde, B. (1987a) HWG 1608--Subchronic study of toxicity to dogs with oral administration (thirteen-weeks feeding study). Unpublished report Ref. No. 15763, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. von Keutz, E. & Schilde, B. (1987b) HWG 1608--Study of chronic toxicity to dogs after oral administration (twelve-month feeding study). Unpublished report Ref. No. 16211, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Kollert, W. (1987) Report dated 10 November 1987, HWE 1608--Internal experiments, Bayer Medical Department, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Lee, S.G.K. & Wood, S.E. (1987) The metabolism of Folicur in dairy goats. Unpublished report Ref. No. MR 94882, prepared by Mobay Corporation, Kansas City, MO, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Lee, S.G.K., Hanna, L.A., Johnston, K., Wood, S.E. & Leimkuehler, W.M. (1988) The metabolism of 14C-Folicur in chickens. Unpublished report Ref. No. 87 156, prepared by Mobay Corp., Kansas City, MO, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Lehn, H. (1988) HWG 1608--Mutagenicity study for the detection of induced forward mutations in the CHO-HGPRT assay in vitro. Unpublished report Ref. No. 16749, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Märtins, T. (1990) HWG 1608--Study for skin and eye irritation/corrosion in rabbits. Unpublished report Ref. No. 12168 B (addendum report to report No. 12168), prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Märtins, T. (1991) HWG 1608 (tebuconazole)--Subacute inhalation toxicity to dogs--Study for cataracts. Unpublished report Ref. No. 20884, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Märtins, T., Pauluhn, J. & Krötlinger, F. (1990) HWG 1608 (tebuconazole)--Subacute inhalation toxicity to cats--Study for cataracts. Unpublished report Ref. No. 19644, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Ohta, K. (1991a) HWG 1608 technical--Acute oral toxicity study on rats. Study No. 91A016. Unpublished report prepared by Nihon Bayer Agrochem K.K., Tokyo, Japan. Submitted to WHO by Bayer AG, Leverkusen, Germany. Ohta, K. (1991b) HWG 1608 technical--Acute dermal toxicity study on rats. Study No. 91A012. Unpublished report prepared by Nihon Bayer Agrochem K.K. Tokyo, Japan. Submitted to WHO by Bayer AG, Leverkusen, Germany. Ohta, K. (1991c) HWG 1608 technical--Acute oral toxicity study on mice. Study No. 91A017. Unpublished report prepared by Nihon Bayer Agrochem K.K., Tokyo, Japan. Submitted to WHO by Bayer AG, Leverkusen, Germany. Ohta, K. (1991d) HWG 1608--Reverse mutation assay (Salmonella typhimurium and Escherichia coli). Unpublished report Ref. No. RA 91036, prepared by Nikon Bayer Agrochem K.K., Tokyo, Japan. Submitted to WHO by Bayer AG, Leverkusen, Germany. Ohta, K. (1992) HWG 1608--Rec- assay with spores in the bacterial system. Unpublished report Ref. No. RA 92007, prepared by Nihon Bayer Agrochem K.K., Tokyo, Japan. Submitted to WHO by Bayer AG, Leverkusen, Germany. Pauluhn, J. (1985) HWG 1608--Study for subacute inhalation toxicity to rat for three weeks (exposure 15 x 6 hours). Unpublished report Ref. No. 13305, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Pauluhn, J. (1987) HWG 1608--Study for subacute inhalation toxicity to rat for three weeks (exposure 15 x 6 hours). Histopathological examinations. Unpublished report Ref. No. 13305A (addendum to Ref. No. 13305), prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Pauluhn, J. (1988) HWG 1608--Study for acute inhalation toxicity to the rat to OECD-guideline No. 403. Unpublished report Ref. No. 16345, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Porter, M.C., Jasty, V., Troup, C.M. & Hartnagel, R.E. (1989) Safety evaluation of HWG 1608: Chronic (1 year) feeding study in dogs. Unpublished report Ref. No. R 4781, prepared by Toxicology Department, Miles Inc., Elkhart, IN, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Poth, A. (1989) Salmonella typhimurium reverse mutation assay with 1H-1,2,4-triazole. Unpublished report Ref. No. R 4859, prepared by Cytotest Cell Research GMBH & Co. KG, Rossdorf, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Putman, D.L. (1987) HWG 1608--Sister chromatid exchange assay in Chinese hamster ovary (CHO) cells. Unpublished report Ref. No. 953, prepared by Microbiological Associates Inc., Bethesda, MD, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Renhof, M. (1984) HWG 1608--Study for embryotoxic effects on rats after oral administration. Unpublished report Ref. No. 12457, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Renhof, M. (1985a) HWG 1608--Study for embryotoxic effects on rats after oral administration. Unpublished report Ref. No. 13273, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Renhof, M. (1985b) HWG 1608--Study for embryotoxic effects on rabbits after oral administration. Unpublished report Ref. No. 13287, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Renhof, M. (1988a) HWG 1608--Study for embryotoxic effects on mice following oral administration. Unpublished report Ref. No. 16527, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Renhof, M. (1988b) HWG 1608--Study for embryotoxic effects on rats after dermal administration. Unpublished report Ref. No. 17089, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Renhof, M. (1988c) Triazole--Investigations into embryotoxic effects on rats after oral administration. Unpublished report No. 17401, Ref. No. 50 19 339, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Renhof, M. (1988d) Triazole--Investigations into embryotoxic effects on rats after oral administration. Unpublished report No. 17402, Supplement to Ref. No. T 50 19 339, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Renhof, M. & Karbe, E. (1988) HWG 1608--Supplementary study for maternal toxicity on mice following oral administration. Unpublished report Ref. No. 16511, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Sheets, L.P. (1988) Primary dermal irritation of technical grade Folicur in rabbits. Unpublished report Ref. No. 1066, prepared by Mobay Corp., Stilwell, KS, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Sheets, L.P. (1990) Dermal sensitization study with technical grade tebuconazole (Folicur) in guinea pigs. Unpublished report Ref. No. 5052, prepared by Mobay Corp., Stilwell, KS, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Sheets, L.P. & Phillips, S.D. (1992) Acute oral toxicity study with HWG 2061 (a metabolite of tebuconazole Folicur(TM)) in female rats. Unpublished report Ref. No. 6685, prepared by Miles Inc., Agricultural Division, Toxicology, Stilwell, KS, USA. Submitted to WHO by Bayer AG, Leverkusen, Germany. Thyssen, J. & Kimmerle, G. (1976) 1,2,4-Triazole--Occupational toxicology study. Unpublished report Ref. No. 5926, prepared by Bayer AG, Wuppertal, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Weber, H. (1987) [Phenyl-U-14C] HWG 1608: Study of biokinetic behaviour in the rat. Unpublished report Ref. No. PF 2859. Metab II/15, prepared by Bayer AG, Leverkusen, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany. Weber, H. (1988) [Phenyl-U-14C] HWG 1608: Ganzkörperautoradiographische Verteilung der Radioaktivität in der Ratte. Unpublished report Ref. No. PF 2962. Metab II/16, prepared by Bayer AG, Leverkusen, Germany. Submitted to WHO by Bayer AG, Leverkusen, Germany.
See Also: Toxicological Abbreviations