Study of the teratogenic potential of FD & C Yellow No. 5 when given by gavage to rats.

FD & C Yellow No. 5 (tartrazine) was given to Osborne-Mendel rats by gavage at dose levels of 0, 60, 100, 200, 400, 600 or 1000 mg/kg body weight/day on days 0-19 of gestation. No maternal or developmental toxicity was observed when the rats were killed on day 20. The mean daily food consumption for the entire period of gestation was significantly greater in the females given 1000 mg/kg body weight/day than in the controls, but maternal body-weight gain was not affected. No dose-related effects were observed in implantations, foetal viability or external foetal development. Foetal skeletal and visceral development was similar among foetuses from all groups. At the doses given, FD & C Yellow No. 5 was neither toxic nor teratogenic.

A study of the teratogenic potential of caffeine ingested in drinking-water.

Caffeine dissolved in drinking-water was available ad lib. to Osborne-Mendel rats at dose levels of 0, 0.007, 0.018, 0.036, 0.07, 0.10, 0.15 or 0.20% during days 0-20 of gestation. The corresponding daily caffeine intakes were 0, 10.1, 27.4, 50.7, 86.6, 115.8, 160.9 and 204.5 mg/kg body weight. Dosages of 160.9 and 204.5 mg/kg were associated with decreased implantation efficiency, increased resorptions and decreased mean numbers of viable foetuses. Numbers of runts were significantly increased after dosages of 115.8-204.5 mg/kg/day. Foetal body weight and length were decreased and oedematous foetuses were increased at dosages of 86.6-204.5 mg/kg/day. Contrary to results seen after gavage studies, caffeine available ad lib. in drinking-water produced no dose-related gross anomalies. Only two animals with missing or hypoplastic nails were produced, both in the 160.9-mg/kg group. Sternebral ossification deficiencies were increased at all dose levels except 10.1 mg/kg/day. Skeletal ossification deficiencies were increased in a dose-related manner at the four highest dose levels. Caffeine given by water bottle produced ossification deficiencies similar to those seen after intubation, but at higher dosages.

Potential reversibility of skeletal effects in rats exposed in utero to caffeine.

Groups of Osborne-Mendel rats, killed at three time intervals following mating, were studied to determine whether prenatal skeletal ossification delays observed following low-level caffeine administration represent transient or persistent ossification problems. Group A litters were killed on gestation day 20; group B neonates were killed on post-natal day 0; and group C pups were killed on post-natal day 6. Within each group, dose levels of 0, 0.018, 0.036 or 0.07% caffeine in distilled water were available ad lib. to groups of 30-60 dams from gestation day 0 to day 20. Average daily caffeine consumption was 24.7-29.0 mg/kg body weight for the 0.018% group, 42.7-48.8 mg/kg body weight for the 0.036% group and 70.6-75.1 mg/kg body weight for the 0.07% group. In group A litters, the mean number of viable foetuses was significantly less in the mid-dose and high-dose animals than in the controls. In the same group, the average number of foetuses per litter with at least one sternebral ossification delay was increased significantly in all treated groups and the average number of foetuses per litter with at least two sternebral variations was significantly increased in the mid- and high-dose groups. The percentages of litters containing foetuses with at least two and at least three sternebral variations and the average number of foetuses per litter with at least three sternebral variations were significantly increased only in the high-dose group. Foetuses from the mid- and high-dose groups also had significant increases in certain skeletal defects, namely missing centra and reduced ossification of the dorsal arch. Foetuses from the high-dose group also had significant increases in bipartite supraoccipital, and reduced ossification of the hyoid, metacarpals and metatarsals. In group B, day 0 neonates from all treated groups showed a significantly increased incidence of delayed sternebral ossification (average number of foetuses per litter with one or more missing, incomplete or bipartite sternebrae). The percentages of litters containing neonates with delayed sternebral ossification were also increased significantly in all treated groups. Neonates from the 0.07% level in group B also exhibited a significant increase in the incidence of supernumerary rib bud, and in reduced ossification of the metacarpals, metatarsals and calcaneus bones. Significant increases were also seen, in group B, in the low- and mid-dose animals, respectively, in supernumery rib bud and in reduced ossification of the metatarsals.(ABSTRACT TRUNCATED AT 400 WORDS)

Study of the teratogenic potential of guar gum.

Guar gum in the diet at 0, 1, 2, 4, 7.5 or 15% was available ad lib. to male and female Osborne-Mendel rats for 13 wk before mating, during mating and throughout gestation. During gestation, the females consumed 0, 0.7, 1.4, 2.7, 5.2 or 11.8 g guar gum/kg of body weight/day, respectively. The animals were killed on gestation day 20. No behavioural effects were seen in any of the treated dams, and no females died during the experiment. Pregnant females in the treated groups consumed less food than the controls during gestation days 0-20 but the decrease was significant only in the 4 and 7.5% groups and was not dose related. Ingestion of guar gum before mating did not affect fertility. In the dams fed 1-7.5% guar gum, there was no effect on the number of corpora lutea or implantations. The dams fed 15% guar gum had slightly fewer corpora lutea and implantations than the controls but no effect was seen on implantation efficiency in this group. The number of viable foetuses/litter was also reduced slightly but not significantly in the 15% group, but since the number of resorptions was not affected, this decrease appears to be an effect of the decreased number of corpora lutea. There was no compound-related effect on foetal development or sex distribution. No terata were seen.

Study of the teratogenic potential of gum arabic.

Gum arabic in the diet at 0, 1, 2, 4, 7.5 or 15% was available ad lib. to male and female Osborne-Mendel rats during premating and mating and throughout gestation. During gestation, the treated females consumed from 683 mg gum/kg body weight/day in the 1% group to 10,647 mg gum/kg/day in the 15% group. The animals were killed on gestation day 20. There were no dose-related changes in maternal findings, number of foetuses, foetal viability or external, visceral or skeletal variations. No terata were seen.

Study of the teratogenic potential of FD & C Red No. 40 when given by gavage to rats.

Osborne-Mendel rats were intubated with FD & C Red No. 40 at dose levels of 0, 30, 75, 150, 300, 600, or 1000 mg/kg body weight/day on days 0-19 of gestation. No developmental toxicity was observed when the animals were killed on day 20 of gestation. No dose-related changes were seen in maternal daily observations, food consumption, body-weight gain or implantations, or in foetal viability, body weight, body length, sex distribution or external variations. Skeletal and soft-tissue development appeared similar in foetuses of all groups. The isolated increases that occurred in the number of male foetuses, number of females with two or more resorptions, number of litters with three or more sternebral variations and incidence of 14th rib bud are considered random occurrences and were not related to dosage.

Study of the teratogenic potential of FD & C yellow No. 5 when given in drinking-water.

FD & C Yellow No. 5 was available to pregnant Osborne-Mendel rats throughout gestation at dose levels of 0.05, 0.1, 0.2, 0.4 or 0.7% in solution in distilled drinking-water. Based on fluid consumption, the rats received 67.4, 131.8, 292.4, 567.9 and 1064.3 mg FD & C Yellow No. 5/kg body weight/day. Distilled water served as the control. No dose-related changes were seen in mean daily food consumption or maternal body-weight gain. Starting during the second trimester of gestation, fluid consumption was significantly greater in the rats given 0.7% FD & C Yellow No. 5 than in the controls. The females were killed on gestation day 20. No dose-related changes were seen in maternal clinical findings, implantations, foetal viability or foetal size (weight and length). No dose-related foetal terata were seen. Neither visceral development nor skeletal development (sternebral and other skeletal bones) was affected by the dye. The small numbers of statistically significant increases in skeletal variations in the 0.05 and 0.4% levels are considered random because they are not dose related.

Teratogenic potential of FD & C red no. 3 when given in drinking water.

FD & C Red No. 3 (erythrosine), a commonly used food additive, was administered to pregnant Osborne-Mendel rats to study its teratogenic potential. Dosing solutions of 0.05, 0.1, 0.2 or 0.4% in distilled water were available at all times and corresponded to daily doses of 64, 121, 248 and 472 mg FD & C Red No. 3/kg body weight. Distilled water served as the control. On gestation day 20, the animals were killed and caesarean sections were performed. The treated animals consumed less fluid than did the control animals, but only random decreases were statistically significant and no dose relationship was seen. Only the 0.2% group consumed significantly more feed than the controls during gestation. Maternal weight gain during days 0-20 was not significantly affected in any group. No dose-related changes were seen in maternal clinical findings, implantations, foetal viability, foetal size (weight and length) or visceral development. No dose-related teratogenesis was seen. Skeletal development was not affected; the few statistically significant increases in skeletal variations were not dose related and were considered to be random. FD & C Red No. 3 was neither foetotoxic nor teratogenic at the dose levels tested in drinking water.

Effect of intratesticular injection of sodium fluoride on spermatogenesis.

The potential of sodium fluoride to affect spermatogenesis in the rat was assessed by intratesticular injection. Experimental rats' left testis was injected with sodium fluoride (50, 175 and 250 ppm) in vehicle (0.9% physiological saline); control testes were injected with vehicle. The right testis served as a non-injected control. Testicular tissues collected 'at' and 'distal to' the injection site and from the non-injected control testes were evaluated microscopically 24 hr and 1, 2 and 3 wk post-injection. Testicular tissues obtained at and distal to the injection site in all fluoride-injected groups resembled tissues collected from corresponding areas in the controls. Seminiferous tubule damage observed in both the vehicle-injected control testes and the fluoride-injected testes but not in the non-injected testes was attributed to injection trauma. Polymorphonuclear leucocyte infiltration was observed 24 hr post injection only at the injection site in the vehicle- and fluoride-injected groups. Leydig cells were unaffected. Leucocyte infiltration with seminiferous tubule damage was not considered to be a fluoride treatment-related effect because it was observed in both vehicle- and fluoride-injected testes. The results demonstrate that the rat is not adversely affected by direct exposure to fluoride at levels 200 times greater than those under normal conditions.

Teratogenic potential of purified pentachlorophenol and pentachloroanisole in subchronically exposed Sprague-Dawley rats.

Male and female Sprague-Dawley (Spartan) rats were exposed to dietary levels of 0, 60, 200 or 600 ppm purified pentachlorophenol (PCP) or pentachloroanisole (PCA) for 181 days, through mating and pregnancy. The daily intakes of PCP were 0, 4, 13 or 43 mg/kg body weight and of PCA were 0, 4, 12 or 41 mg/kg body weight. Animals exposed to PCP generally consumed more food than control animals during pregnancy. Dams at the high-dose level of both compounds showed evidence of toxicity, weighing less on day 0 of gestation and gaining less throughout pregnancy than did the controls. Dams exposed to the high dose of PCP gained less weight during pregnancy (exclusive of the gravid uterus) than control dams. At the 43 mg/kg/day dose level PCP was embryolethal. Foetuses at the lower dose levels of PCP exhibited dose-related decreases in body weights. A reduction in crown-rump length and an increase in foetal skeletal variations were seen at 13 mg/kg/day in PCP animals only. An intake of 41 mg PCA/kg/day was associated with a decrease in the number of corpora lutea and in embryolethality. PCA exposure also resulted in reductions in foetal body weight and crown-rump lengths of males at 4 and 41 mg/kg/day. Female foetuses were unaffected.

Mineral interactions in rats fed AIN-76A diets with excess calcium.

The effects of moderate increases in dietary calcium on maternal and foetal mineral interactions were studied in Charles River CD/VAF Plus rats. Female rats were given 0.50, 0.75, 1.00 or 1.25% dietary calcium as calcium carbonate in AIN-76A diets for 6 wk before mating, during mating and for 20 days of gestation. Inductively coupled argon plasma-atomic emission spectrometry was used to determine mineral levels in the tissues of non-pregnant rats after 42 days on the diets, in the tissues of pregnant rats on day 20 of gestation and in the whole body of day-20 foetuses. The femurs of the non-pregnant and pregnant rats had a dose-related linear increase in calcium content. In livers of the non-pregnant rats, dose-related linear increases in the phosphorus, zinc and magnesium content were observed, but there was a dose-related decrease in the iron content. There were dose-related linear decreases in the iron and copper contents of the kidneys from the non-pregnant rats. In pregnant rats dose-related linear decreases were observed in the iron content of the liver and in the zinc, iron and magnesium contents of the kidney. The foetuses from rats given a moderate increase in dietary calcium had dose-related decreases in the whole-body contents of phosphorus, iron, copper and magnesium.

Testing the potential of sodium fluoride to affect spermatogenesis in the rat.

The potential of sodium fluoride (NaF) to affect spermatogenesis and endocrine function was assessed in P and F1 generation male rats. Male and female experimental rats received sodium fluoride in their drinking water at one of four concentrations (25, 100, 175, 250 ppm). P generation male and female rats were exposed to sodium fluoride in their drinking water for 10 wk and then males were mated to females within the same treatment groups. Reproductive tissues were collected from P generation male rats after approximately 14 wk of treatment. Pregnant females (P) were exposed to sodium fluoride via their drinking water through gestation and lactation. F1 generation weanling male rats remained within the same treatment groups as their parents. F1 generation male rats were exposed to sodium fluoride in their drinking water for 14 wk, at which time reproductive tissues were collected. Dose-related effects were not observed within the P and F1 treatment groups in testis weights, prostate/seminal vesicle weights, non-reproductive organ weights, testicular spermatid counts, sperm production per gram of testis per day, sperm production per gram of testis, LH, FSH or serum testosterone concentrations. Histological changes were not observed in testicular tissues from either the P or F1 generation. We conclude that prolonged exposure to sodium fluoride in drinking water at the doses administered in this study does not adversely affect spermatogenesis or endocrine function in the P and F1 generation male rats.

The effect of maternal exposure to flaxseed on spermatogenesis in F(1) generation rats.

Pregnant Sprague-Dawley rats were exposed to a flaxseed (20 or 40%), flaxmeal (13 or 26%) or standard NIH AIN-93 (0% flaxseed control) diet throughout gestation and until their offspring were weaned. After weaning, F(1) generation males were placed in the same diet treatment groups as their mothers for 70 days. Statistically significant differences were not observed between either low-dose or high-dose flaxseed and flaxmeal-treated animals and the 0% flaxseed control animals for testis weights, homogenization resistant spermatid counts, daily sperm production rates, epididymal weights, seminal vesicle weights, seminiferous tubule fluid testosterone concentrations and the percentage of sperm abnormalities. The following statistically significant differences were observed when treated groups and the 0% flaxseed control groups were compared: (1) increases in serum LH in the 20% and 40% flaxseed treatment groups and in serum LH and testosterone in the 26% flaxmeal treatment group; (2) increases in the cauda epididymal weight from the 20% and 40% flaxseed groups; (3) increases in cauda epididymal sperm numbers/g epididymis from the 20% and 40% flaxseed and the 13% and 26% flaxmeal treatment groups; (4) a decrease in prostatic weight from the 20% flaxseed and 13% and 26% flaxmeal treatment groups. Prostate weight in the 40% flaxseed treatment group was lower but not statistically significantly different than the 0% flaxseed control group. Histological effects on spermatogenesis were not observed in either the control group, flaxmeal or the flaxseed treated groups.

Developmental toxicity of sodium fluoride measured during multiple generations.

Sodium fluoride (NaF) has been used to fluoridate drinking water in the United States since the mid 1940s. Because of the lack of reliable studies on the multigeneration effects of the compound, NaF (0, 25, 100, 175 or 250 ppm in drinking water) was given to rats continuously during three generations. Parental (F0) generation rats were treated for 10 weeks and mated within groups. At gestation day 20, caesarean sections were performed and eight F0 females per group and their litters (F1) were observed for implant status, fetal weight and length, sex and morphological development. The remaining F0 females (29-32 per group) were allowed to litter. F1 offspring (36 of each sex per group) were mated within groups, and caesarean sections were performed at gestation day 20. The F1 females and their litters (F2) were observed for implant status, fetal weight and length, sex and morphological development. In addition, F2 fetuses were evaluated for internal (soft-tissue) and skeletal development. Decreased fluid consumption for F0 and F1 dams at 175 and 250 ppm was attributed to decreased palatability of the solution. No dose-related effects in feed consumption or mean body weight gain were observed in either F0 or F1 females. Numbers of corpora lutea, implants, viable fetuses and fetal morphological development were similar in all groups. No dose-related anomalies in internal organs were observed in F2 fetuses. Ossification of the hyoid bone of F2 fetuses was significantly decreased at 250 ppm. Because of the decreased ossification of the hyoid bone, 250 ppm is considered the effect level.

Developmental effects of serum from flaxseed-fed rats on cultured rat embryos.

Gestation day 9.5 rat embryos were cultured for 45 h in serum obtained from pregnant rats that had been fed throughout gestation with either a control diet (based on the AIN-93 formulation), a diet supplemented with flaxseed (20% or 40%, w/w), or a diet supplemented with de-fatted flaxseed ("flaxseed meal", 13 or 26%, w/w). The embryos were fixed in neutral formalin at the end of culture. Overall growth and development was assessed, and the presence of abnormalities was noted. A significant inhibition of growth (as determined by crown-rump length) relative to control was observed in embryos cultured in serum from rats fed the 20% flaxseed diet. The incidence of spontaneous heart inversions was increased significantly in the embryos cultured in serum from the 20% flaxseed and 26% flaxseed meal fed rats. The incidence of flexion defects was increased significantly in embryos cultured in serum from 20% flaxseed-fed rats. The lack of an apparent dose response in any of the statistically significant effects suggests that the observed anomalies were chance occurrences unrelated to the treatment group from which serum was obtained. It is therefore concluded that diets high in flaxseed or flaxseed meal do not result in serum factors that are directly embryotoxic to organogenesis-staged rat embryos. This finding is consistent with the findings of a parallel in vivo rat teratology study where no significant embryotoxicity attributable to flaxseed exposure was observed.

Foetal development in rats fed AIN-76A diets supplemented with excess calcium.

This study was designed to evaluate the developmental effects of moderate dietary calcium increases in rats fed nutritionally adequate diets. Female Charles River CD/VAF Plus rats were given 0.50 (control), 0.75, 1.00 or 1.25% dietary calcium as calcium carbonate in AIN-76A diets for 6 wk before mating, during mating and for 20 days of gestation. On gestation day 20, the animals were killed and caesarean sections were performed. Both the non-pregnant and pregnant rats in the 0.75, 1.00 and 1.25% groups ate slightly more than did the control group during most of the intervals measured, but not all the increases were statistically significant. There was no consistent pattern of increase or decrease in weight gain. No dose-related changes were found in maternal clinical findings, the average number of implantations, resorptions and viable foetuses, or foetal length or weight. Under the conditions of the study, there were no statistically significant increases as compared with the control group in the litter incidence regarding specific external, visceral or skeletal variations of the foetuses. Dietary calcium was neither foetotoxic nor teratogenic at the concentrations used.

Reproduction study of caffeine administration to male Osborne-Mendel rats.

The potential for caffeine, administered twice daily by gavage in a total dose of 40 or 80 mg/kg/day, to adversely affect the reproductive performance of male rats was investigated. Treatment was continued through 3 wk of serial mating; mating and dosing were terminated concurrently, after which the sires were autopsied and their testes weighed. Controls were treated similarly with distilled water. Significant dose-related differences were detected for sire body weight, but not for testes weight. The majority of the significant effects on the offspring were for those born to the low-dose sires. Statistically significant differences were sporadically detected for the number of pups born and their body weights and survival; however, these differences were not consistently detected in either a dose- or temporal-related fashion. Thus, caffeine appears to have little potential to produce adverse reproductive effects when administered by gavage to male rats at the levels tested in this study.

Developmental toxicity of sodium fluoride in rats.

Despite the chronic exposure of the US population to fluoridated drinking water since the 1940s, existing studies have been judged inadequate to determine any potential reproductive or developmental hazard. This study was conducted to determine the effects of sodium fluoride (NaF) on foetal development. Sperm-positive female rats were given 0, 10, 25, 100, 175 or 250 ppm NaF daily throughout gestation. They were dosed by drinking water to mimic human exposure to fluoridated water. No dose-related behavioural changes or maternal clinical signs were noted. Fluid consumption by females in the 175- and 250-ppm groups was significantly less than that of the control females. Because of this decreased fluid consumption, the daily amount of NaF ingested (0, 1.4, 3.9, 15.6, 24.7 and 25.1 mg/kg body weight) was less than expected at the two high levels. Feed consumption decreased significantly at 250 ppm, and body weights of pregnant females reflected feed consumption trends. The mean number of viable foetuses per female in all treated groups was similar to that of the control group. The significant decrease in the mean number of implants per litter in the 250-ppm group is probably linked to the lower mean number of corpora lutea in this group. The occurrence of in utero deaths was similar in the control and treated groups. Foetal growth (in terms of foetal body weight and crown-rump length) was not affected by NaF, despite the fact that the dams in the 250-ppm group ate significantly less feed and drank significantly less fluid. There was no dose-related increase in the number of external anomalies in foetuses due to NaF ingestion. At the doses given, NaF had no effect on the development of specific bones, including sternebrae. A significant increase was seen in the average number of foetuses with three or more skeletal variations in the 250-ppm group; the number of litters with foetuses with three or more skeletal variations was increased in the 250-ppm group also, but the increase was not significant. There was no dose-related effect of NaF on the incidence of soft tissue variations.

Effect of ethanol and vitamin A excess on vitamin A status in the liver, plasma and foetuses of pregnant rats.

The effect of maternal consumption of dietary ethanol and high doses of vitamin A by gavage was investigated by evaluating plasma, liver and foetal vitamin A in Osborne-Mendel pregnant rats with a view to assessing whether ethanol modulated the potential toxicity of excess vitamin A. All groups received 4000 IU vitamin A/litre in a liquid diet. Ethanol-exposed groups also received 6.4% (v/v) ethanol in the liquid diet. Vitamin A was administered by gavage once per day in corn oil in doses ranging from 10,000 to 160,000 IU/kg body weight. Plasma vitamin A levels in ethanol-exposed groups were similar to levels in a pair-fed group. Plasma vitamin A levels were similar in the group given ethanol plus 40,000 IU vitamin A/kg and the group given 40,000 IU vitamin A/kg only, but were higher in the group receiving ethanol plus 80,000 IU vitamin A/kg than in the group given 80,000 IU vitamin A/kg only. Retinyl esters were present in the plasma of animals receiving 160,000 IU vitamin A/kg only, indicating possible saturation of the liver with vitamin A. Retinyl palmitate levels in female foetuses of the group administered ethanol plus 80,000 IU vitamin A/kg were significantly higher than those of the group administered 80,000 IU vitamin A/kg only; no significant differences in levels of retinyl palmitate in male foetuses were observed between these two groups. This observation suggests a possible sex difference in the modulation of vitamin A toxicity by ethanol in the foetus.

Developmental effects of combined exposure to ethanol and vitamin A.

The potential for ethanol (EtOH) to influence the developmental toxicity of vitamin A was investigated. 11 groups of approximately 31 FDA-bred Osborne-Mendel rats received either a control or isocaloric 6.4% EtOH liquid diet (containing 4000 IU vitamin A/litre) ad lib. The vehicle control, EtOH and pair-fed (pair-fed against the EtOH group) groups received corn oil (the vehicle) by gavage. Vitamin A was administered by gavage without EtOH at 40,000, 80,000, 120,000 or 160,000 IU/kg daily. Vitamin A was administered by gavage at 10,000, 20,000, 40,000 or 80,000 IU/kg with EtOH ad lib., daily throughout the study. Combined EtOH and vitamin A resulted in significant reductions in maternal diet consumption and body weight when doses of vitamin A were as low as 10,000 IU/kg. The most severe effects on overall (days 0-20) maternal body weight gain were observed in the groups receiving 120,000 or 160,000 IU vitamin A/kg alone or EtOH in combination with 80,000 IU vitamin A/kg. The overall diet consumption (days 0-20) paralleled the overall weight gain. In general, pups exposed to ethanol and vitamin A had a tendency to weigh less than those exposed to vitamin A alone, but to weigh more than those exposed to EtOH alone. EtOH combined with vitamin A at 80,000 IU/kg resulted in an increased incidence of cleft palate relative to the vehicle control or either treatment alone. The incidence of exencephaly and protruding tongue was significantly greater in the group given vitamin A at 160,000 IU/kg, compared with the vehicle control group. The most consistent statistically significant skeletal finding in the groups receiving combined treatment was a treatment-related increased incidence of supernumerary ribs [14th rib (C7), 14th rib bud (L1) and 15 ribs]. In addition, the incidence of misshapen zygomatic arch was also significantly increased in the group exposed to EtOH and vitamin A at 80,000 IU/kg. The incidence of moderately enlarged renal pelvis and severely enlarged ureter proximal to the kidney was increased in the group exposed to EtOH and vitamin A at 80,000 IU/kg relative to the vehicle control, or either treatment alone. Therefore, for some of the endpoints examined in this investigation, it would appear that ethanol potentiates the developmental effects of vitamin A.