Effects of chronic internal alpha irradiation on physiology, growth and reproductive success of Daphnia magna

Daphnids were chronically exposed to waterborne Am-241, an alpha-emitting radionuclide, ranging in concentration from 0.4 to 40 Bq ml(-1). Am-241 amounts were monitored in the medium, daphnid tissues and cuticles. Corresponding average dose rates of 0.02, 0.11 and 0.99 mGy h(-1) were calculated for whole organisms with internal alpha-radiation contributing 99% of total dose rates. Effects of internal alpha irradiation on respiration and ingestion rates, adult, egg and neonate individual dry masses, fecundity and larval resistance to starvation were examined in 23-day experiments. Daphnids showed increased respiratory demand after 23 days at the highest dose rate, suggesting increased metabolic cost of maintenance due to coping with alpha radiological stress. Although no effect was detected on ingestion rates between contaminated and control daphnids, exposure to dose rates of 0.11 mGy h(-1) or higher, resulted in a significant 15% reduction in body mass. Fecundity remained unchanged over the 23-day period, but individual masses of eggs and neonates were significantly smaller compared to the control. This suggested that increased metabolic expenditure in chronically alpha-radiated daphnids came at the expense of their energy investment per offspring. As a consequence, neonates showed significantly reduced resistance to starvation at every dose rate compared to the control. Our observations are discussed in comparison with literature results reported for cadmium, a chemical toxicant which affects feeding activity and strongly reduces individual energy uptake.     

Gestational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) severely alters reproductive function of female hamster offspring.

Low doses of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), administered as a single dose to the dam during gestation, alter development of the fetal rodent reproductive system. In male rat and hamster offspring, dosing with TCDD during gestation reduces epididymal and ejaculated sperm counts and delays puberty. In female rats, in utero TCDD-exposure results in reduced ovarian weight and fecundity, and induces cleft phallus and a persistent thread of tissue across the vaginal orifice. Here, we demonstrate that 2-microgram TCDD/kg, administered as a single oral dose prior to sexual differentiation, alters reproductive function in female hamster offspring, a species relatively resistant to the lethal effects of TCDD. In the current study, pregnant hamsters (P0 generation) were dosed orally with vehicle (corn oil) or 2 micrograms TCDD/kg on gestational day (GD) 11.5. P0 maternal viability, body weight, fertility, and F1 litter size did not differ between control and treated groups. In the F1 generation, body weights were permanently reduced by about 30%, vaginal opening was delayed (p < 0.0001), and vaginal estrous cycles were altered by TCDD treatment. In contrast, most treated female offspring displayed regular 4-day behavioral estrous cycles, indicating that in utero TCDD treatment did not markedly disrupt hypothalamic-pituitary-gonadal hormonal cyclicity. Although both control and TCDD-treated F1 females mated successfully with a control male (estrous cyclicity was abolished by mating), 20% of the F1 treated females did not become not pregnant (no implants). In addition, 38% of pregnant F1 females from the TCDD group died near-term, and the numbers of implants in pregnant animals (treated 5.1 versus 11.3) and pups born live (2.7 treated vs. 8.7 control) were reduced by TCDD-treatment. In the F2, survival through weaning was drastically reduced (15% treated vs. 78% for control) by TCDD treatment of P0 dams. F1 female hamster offspring exposed in utero to TCDD displayed external urogenital malformations, with most females having complete clefting of the phallus, an effect previously reported in the rat. Unlike rats exposed to TCDD (0.2-1.0 microgram/kg) on GD 15 or GD 8, hamster offspring did not display vaginal threads. These results demonstrate that in utero administration of TCDD adversely affects growth, reproductive function, and anatomy in female hamster offspring given a dosage level nearly four orders of magnitude below the dosage level toxic to the adult animal. Adverse effects of TCDD persisted through two generations (F1 and F2), even though the F1 was only indirectly exposed during gestation and lactation.     

Chronic effects of cyanobacterial toxins on Daphnia magna and their offspring

The zooplankton grazer Daphnia magna endures living in water bodies up to moderate densities of cyanobacteria, such as Microcystis spp., known for producing toxic secondary metabolites. Although daphnids are affected via decreased food filtering, inhibition of digestive proteases and lethality, development of tolerance against cyanobacterial toxins has also been observed. Aim of our study was to investigate in detail chronic effects of cyanobacterial toxins, with emphasis on microcystin, on D. magna. The animals were exposed chronically for two generations to either microcystin-LR in 5 or 50 μg L⁻¹, or to cyanobacterial crude extract containing the same amount of total microcystin, starting at neonate stadium. Survival, growth, maturation and fecundity were observed for the first generation during two months. In the offspring survival, maturation, and growth were followed for the first week.Low concentration of microcystin-LR slightly affected the growth and reproduction of parent daphnids. Survivorship decreased during chronic exposure with increasing microcystin concentration. Age to maturity of the offspring increased and their survival decreased after parent generation was exposed to the toxin, even if the offspring were raised in control medium. Besides, cessation of the eggs/embryos was observed and malformation of neonates caused by cyanobacterial toxins was firstly recorded.     

Effects of tartrazine on exploratory behavior in a three-generation toxicity study in mice

Tartrazine was given to mice in the diet at levels of 0 (control), 0.05%, 0.15%, and 0.45% from 5 weeks of age of the F(0) generation to 9 weeks of age of the F(2) generation, and selected reproductive and neurobehavioral parameters were measured. In the F(1) generation, the development of swimming direction at postnatal day (PND) 7 was accelerated significantly in male offspring in a dose-related manner. Surface righting at PND 7 was affected significantly in female offspring in dose-related manner. Several variables in exploratory behavior showed significant tendencies to be affected in the treatment groups in male offspring at 3 weeks of age. In the F(2) generation, the development of swimming direction at PND 7 was accelerated significantly in the high-dosed group in male offspring. Time taken of olfactory orientation at PND 14 was accelerated significantly in male offspring in a dose-related manner. Several variables in exploratory behavior showed significant tendencies to be affected in the treatment groups in male offspring at 3 weeks of age, and in males at 8 weeks of age. The dose levels of tartrazine in the present study produced a few adverse effects on neurobehavioral parameters throughout generations in mice.     

The reproductive and developmental toxicity of the antifungal drug Nyotran (liposomal nystatin) in rats and rabbits.

Nyotran is a liposomally encapsulated i.v. formulation of the antifungal polyene nystatin. This drug was evaluated in a series of reproductive toxicity studies, according to the guidelines outlined by the International Conference on Harmonization (ICH). A fertility and early embryonic development study (SEG I) and a prenatal and postnatal development (SEG III) study were conducted in rats, and embryo-fetal development (SEG II) studies were conducted in rats and rabbits. Nyotran was administered iv in all studies. In SEG I and SEG III, rats were administered daily doses of 0.5, 1.5, or 3.0 mg/kg Nyotran. In both studies, parental mortality and toxicity in the 3.0 mg/kg dose group necessitated the lowering of the high dose to 2.0 mg/kg/day. Parental toxicity, in the form of decreased body weights, decreased food consumption, and piloerection were also observed at the 1.5 mg/kg/day dose level in the SEG I and SEG III studies. Despite the parentally toxic doses in the SEG I study, there was no effect of Nyotran on F0 male or female fertility or early embryonic development of F1 offspring. In the SEG III study, lactational body weights of the F1 generation were decreased at all Nyotran dose levels. There was no effect on pre-wean developmental landmarks, but post-wean development was affected by Nyotran administration at all dosage levels. Preputional separation was delayed in the 1.5 and 3.0/2.0 mg/kg/day F1 offspring, auditory startle function was decreased in F1 females at all dose levels, and motor activity was decreased in male F1 offspring at all dose levels. However, there were no treatment-related effects on the subsequent mating of the F1 generation and resulting F2 offspring. In SEG II studies, rats and rabbits were also administered 0.5, 1.5, or 3.0 mg/kg/day of Nyotran during gestation. The high dose in these SEG II studies was not lowered, as the maternal animals were able to tolerate the shorter duration of dosing. Maternal effects in rabbits were observed only in the high-dose group and were limited to decreased food consumption and decreased absolute and relative liver weight. Decreased food consumption in high-dose dams and clinical weight loss in some animals at the mid- and high-dose levels evidenced maternal toxicity in rats. Nyotran did not have any effect on Caesarian section parameters in either rats or rabbits and no effect on the incidence of fetal malformations in rabbits. A statistically significant increase in mild hydrocephaly, observed in 4 rat fetuses, was seen at the highest dose level of 3.0 mg/kg/day. The biological significance and relationship to Nyotran treatment of this finding is not clear. This finding may represent a change in the background incidence or a change in the pattern of responsiveness of this strain of rat fetus to the test chemical. Toxicokinetic data were also collected in the SEG II rabbit and rat studies for comparison to human exposures. In both species, systemic exposure to the nystatin at effective antifungal concentrations was demonstrated. The systemic exposures in rats and rabbits were, however, considerably less than have been reported in humans administered clinical doses of 2 or 4 mg/kg/day Nyotran. Thus, humans tolerate higher dosages and systemic exposures of Nyotran relative to rats and rabbits and there is no margin of safety in either dosage level or systemic exposure to drug. Given this lack of a margin of safety and the effects on postnatal development in F1 rats, caution should be exercised when using this drug in females of childbearing potential.     

Evidence for radiation hormesis after in vitro exposure of human lymphocytes to low doses of ionizing radiation

Previous research has demonstrated that adding a very small gamma-ray dose to a small alpha radiation dose can completely suppress lung cancer induction by alpha radiation (a gamma-ray hormetic effect). Here we investigated the possibility of gamma-ray hormesis during low-dose neutron irradiation, since a small contribution to the total radiation dose from neutrons involves gamma rays. Using binucleated cells with micronuclei (micronucleated cells) among in vitro monoenergetic-neutron-irradiated human lymphocytes as a measure of residual damage, we investigated the influence of the small gamma-ray contribution to the dose on suppressing residual damage. We used residual damage data from previous experiments that involved neutrons with five different energies (0.22-, 0.44-, 1.5-, 5.9-, and 13.7-million electron volts [MeV]). Corresponding gamma-ray contributions to the dose were approximately 1%, 1%, 2%, 6%, and 6%, respectively. Total absorbed radiation doses were 0, 10, 50, and 100 mGy for each neutron source. We demonstrate for the first time a protective effect (reduced residual damage) of the small gamma-ray contribution to the neutron dose. Using similar data for exposure to gamma rays only, we also demonstrate a protective effect of 10 mGy (but not 50 or 100 mGy) related to reducing the frequency of micronucleated cells to below the spontaneous level.     

Reproductive and neurobehavioural toxicity study of erythrosine administered to mice in the diet.

Erythrosine was given in the diet to provide levels of 0 (control), 0.005, 0.015 and 0.045% from 5 weeks of age of the F(0) generation to 9 weeks of age of the F(1) generation in mice, and selected reproductive and neurobehavioural parameters were measured. There were no adverse effects of erythrosine on either litter size, litter weight or sex ratio at birth. The average body weight of the offspring was significantly increased in the middle-dose group in both sexes during the lactation period. In behavioural developmental parameters, any variables showed no significant adverse effects in either sex in the lactation period. In movement activity of exploratory behaviour, several parameters were significantly changed in the high-dose group, and those effects were dose related in adult females in the F(0) and F(1) generations and in male offspring in the F(1) generation. The dose level of erythrosine in the present study produced few adverse effects in reproductive and neurobehavioural parameters in mice.     

Effects of tetradifon on Daphnia magna during chronic exposure and alterations in the toxicity to generations pre-exposed to the pesticide

Daphnia magna was exposed to some nominal sublethal tetradifon concentrations (0.10, 0.18, 0.22 and 0.44 mg/l) during 21 days. Chronic toxicity tests were carried out using neonates of F(1) first brood (F(1)-1st) and F(1) third brood (F(1)-3rd) offspring generations from parentals (F(0)) pre-exposed to the pesticide. The effect of tetradifon on survival, reproduction and growth was monitored for the selected daphnid generations. The parameters used to evaluate pesticide effect on reproduction were: mean total young per female, mean brood size, time to first reproduction, mean number broods per female and intrinsic rate of natural increase (r). Survival and growth (body length) were also determined after 21 days of exposure to the pesticide. Reproduction was significantly reduced when tetradifon concentration increased in the medium. This effect was higher in F(1)-1st and F(1)-3rd offsprings compared to the parental generation (F(0)) daphnids. The intrinsic rate of natural increase (r) decreased with increasing concentrations of tetradifon especially in those animals from F(1)-3rd. However, the growth of the exposed organisms decreased in the same order of magnitud in all the generations tested. Survival was not affected after exposure to the selected tetradifon concentrations. The maximum acceptable toxicant concentration (MATC) was calculated for F(0), F(1)-1st and F(1)-3rd generations exposed to the pesticide using as parameter of evaluation the intrinsic rate of natural increase (r). The interpolation of these results gave MATC values of 0.13 mg/l pesticide for F(0) and F(1). The EC50 values have been derived for some selected parameters on D. magna exposed to tetradifon. EC50 values decreased in F(1)-1st and F(1)-3rd generations compared to the parental generation F(0), these results mean that less amount of toxicant would be necessary to reduce the selected parameters to 50% in the first and third broods of D. magna. Expanding the reproduction tests to several broods of a parental generation revealed important information on chronic toxicity that could be used in the protection of aquatic environment.     

Two-generation reproduction toxicity studies of di-(C(7)-C(9) alkyl) phthalate and di-(C(9)-C(11) alkyl) phthalate in the rat

Di-(C(7)-C(9) alkyl) phthalate (D79P) and di-(C(9)-C(11) alkyl) phthalate (D911P), based on high-normality linear oxo-alcohols, have been assessed for their impact upon reproductive performance in Sprague-Dawley rats. Rats were continuously exposed to either D79P or D911P at dietary levels of 0%, 0.1%, 0.5%, or 1.0% over two generations. Selected F(0) offspring (F(1) generation) were exposed to the same dietary concentration of D79P or D911P as the respective F(0) animals, and were mated to produce F(1) offspring. Both D79P and D911P markedly reduced body weight gain in F(0) and F(1) adult males at the highest dose, but females were affected to a lesser extent. There was no impairment of fertility, fecundity, or development in either generation, but body weights of offspring in the 1.0% D79P and 1.0% D911P groups were slightly and transiently reduced over the weaning period. Although decreases in the weight of several organs were accounted for by depressed body weight, ovary weights were reduced in both generations exposed to 1.0% D79P, and epididymidal weights were slightly reduced in adults of both generations exposed to 1.0% D911P. However, ovarian function-assessed by the oestrus cycle and mating behaviour-and epididymidal sperm concentration, motility, and morphology were unaffected by either substance. Treatment resulted in liver changes, particularly in males, characterised by increased liver weight in young animals, histopathologic changes and reduced organ weight in mature animals, and an increase in palmitoyl CoA oxidase activity. In conclusion, neither D79P nor D911P impaired reproductive function in rats when administered in the diet at levels that induce systemic toxicity, and the NOAEL for effects on reproduction in the rat is 0.5% for both D79P and D911P.     

Reproductive and neurobehavioural toxicity study of bis(2-ethylhexyl) phthalate (DEHP) administered to mice in the diet.

Bis(2-ethylhexyl) phthalate (DEHP) was given in the diet to provide levels of 0 (control), 0.01, 0.03, and 0.09% from 5 weeks of age of the F0 generation to 9 weeks of age of the F1 generation in mice, and selected reproductive and neurobehavioural parameters were measured. There were no adverse effects of DEHP on either litter size, litter weight or sex ratio at birth. The average body weight of male offspring was significantly decreased in the low-dose group at birth. In behavioural developmental parameters, surface righting at PND 4 was significantly delayed in the low- and middle-dose group in female offspring, and those effects were slightly dose related (P < 0.05). Surface righting at PND 7 was significantly depressed in the high-dose group of male offspring, and those effects were significantly dose related (P < 0.001). That of female offspring was significantly depressed in the low-dose group. The dose level of DEHP in the present study produced few adverse effects in reproductive and neurobehavioural parameters in mice.     

The impact of ethylene glycol monomethyl ether on ovarian function may extend to the next generation in female mice: A preliminary study

Ethylene glycol monomethyl ether (EGME) is known to alter the reproductive function of exposed animals and their offspring; however, its influence on cumulus–oocyte complexes (COCs) remains unclear. This study assessed the toxicity of EGME on oocytes and COCs by analyzing the number of oocytes in the F0 and F1 generations and evaluating apoptosis in oocytes and COCs after treating the F0 generation with EGME. There was a dose-dependent increase in the apoptosis ratios in the COCs from F1 mice, which were not directly exposed to EGME, with apoptosis ratios of 0.065, 0.102, 0.184, and 0.212 for the 0%, 0.05%, 0.1%, and 0.2% EGME dose groups, respectively. The increased apoptosis of cumulus cells may play a role in the toxicity of EGME toward ovarian function. EGME toxicity seems to affect female offspring in future generation(s), but further studies with a large number of animals are necessary to verify these conclusions.     

Two-generation reproduction study by dosing with glutaraldehyde in the drinking water of CD rats

Adult male and female CD rats (F0) were dosed with glutaraldehyde (GA; CAS number 111-30-8) in drinking water at concentrations of 0 (controls), 50, 250, or 1000 ppm for a 10-wk prebreed period and through mating, gestation, and lactation. Resultant F1 offspring, selected to be parents of the next generation, were continued on the same regime from prebreed through lactation. Twenty-eight parental animals per sex per generation for each dose group were evaluated for clinical signs, body weight (absolute and gain), and water and food consumption. The offspring were evaluated for survival and body weight to weaning. Necropsy and light microscopic examination of removed tissues were conducted in all F0 and F1 parents and in 10 offspring/sex/group/generation. Average daily consumptions of GA (as mean +/- SD) for the low, intermediate, and high concentrations were respectively 4.25 +/- 0.87, 17.50 +/- 4.16, and 69.07 +/- 14.58 mg/kg/d for F0 parental males, and 6.68 +/- 0.78, 28.28 +/- 4.09, and 98.37 +/- 11.71 mg/kg/d for F0 parental females. The corresponding values for the F1 parents were 4.53 +/- 1.02, 21.95 +/- 4.88, and 71.08 +/- 16.21 mg/kg/d for males and 6.72 +/- 0.84, 29.57 +/- 5.41, and 99.56 +/- 16.72 mg/kg/d for females. There were no effects on parental fertility and mating performance or on pup viability and litter size in any generation. No apparent treatment-related histopathology was seen in parents or offspring. Parental body weights and body weight gains were significantly reduced at 1000 ppm at a few isolated time periods, particularly during prebreed. Food consumption was significantly reduced at 1000 ppm for F0 and F1 parents during the prebreed and gestation periods, and at 250 ppm for F0 males during prebreed and gestation and F1 females during gestation and lactation. Water consumption by the F0 and F1 parents of the 250 at 1000 ppm groups was reduced throughout the prebreed period. At 1000 ppm, average litter weights were reduced over lactation d 21-28 for the F1 and F2 offspring. The no-observed-effect level (NOEL) for adult toxicity was 50 ppm and for offspring 250 ppm. There were no indications of reproductive toxicity, and the NOEL for this study was therefore > 1000 ppm.     

Reproductive and neurobehavioural toxicity study of tartrazine administered to mice in the diet.

Tartrazine was given in the diet to provide levels of 0% (control), 0.05%, 0.15%, and 0.45% (approximately 83, 259, 773 mg/kg/day, respectively) from five weeks of age of the F0 generation to nine weeks of age of the F1 generation in mice, and selected reproductive and neurobehavioural parameters were measured. In movement activity of exploratory behaviour in the F0 generation, number of vertical activity was significantly increased in the middle-dose group in males. There were no adverse effects of tartrazine on either litter size, litter weight and sex ratio at birth. The average body weight of male offspring was significantly increased in the high-dose group and that of female offspring was significantly increased in the middle-dose group at birth. In behavioural developmental parameters, surface righting at PND 4 was significantly accelerated in the high-dose group in male offspring, and those effects were significantly dose-related in a trend test (P<0.01). Cliff avoidance at PND 7 was significantly accelerated in the middle-dose group in male offspring. Negative geotaxis at PND 4 was significantly delayed in the high-dose group in female offspring. Other variables measured showed no significant adverse effects in either sex in the lactation period. In movement activity of exploratory behaviour in the F1 generation, number of movement showed a significant tendency to be affected in the treatment groups in male offspring in a trend test (P<0.05). The dose level of tartrazine in the present study produced a few adverse effects in neurobehavioural parameters during the lactation period in mice. Nevertheless, the high-dose level were in excess of the ADI of tartrazine (0-7.5 mg/kgbw), and the actual dietary intake of tartrazine is presumed to be much lower. It would therefore appear that the levels of actual dietary intake of tartrazine is unlikely to produce any adverse effects in humans.     

Studies on the pharmacological bases of fetal toxicity of drugs. III. Fetal toxicity of potassium nitrate in 2 generations of rats

The fetal toxicity of potassium nitrate (KNO3) used widely as a food additive was studied in Wistar rats. The pregnant rats were fed a diet containing 2.5, 0.5 or 0.1% of KNO3 from day 7 to 14 of pregnancy. Neither maternal nor fetal toxicity including external malformations were observed at term in any group. After spontaneous delivery, the offspring were reared until 13 weeks after birth. No harmful effects were detected in any group. The female offspring (F1) of all groups were mated with the male (F1) of the same group. Good reproductive performances were shown in all groups. The pregnant rats were fed the same diet, which their mothers (F0) had been fed, from day 7 to 14 of pregnancy. Various types of malformations such as exencephaly, cleft lip and palate, polydactyly and micro- or anophthalmia were observed in 6 of 133 fetuses and 7 of 63 newborns from dams treated with 2.5% KNO3, but no external malformations were observed in other groups. The male offspring (F2) of the treated groups showed slow growth until 13 weeks after birth. These results suggest that a dose of 2.5% KNO3 is toxic to the F2 generation, but not to the F1 generation.     

Repeated in utero and lactational 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure affects male gonads in offspring, leading to sex ratio changes in F2 progeny

The effects of in utero and lactational 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the reproductive system of male rat offspring (F1) and the sex ratio of the subsequent generation (F2) were examined. Female Holtzman rats were gavaged with an initial loading dose of 400 ng/kg TCDD prior to mating, followed by weekly maintenance doses of 80 ng/kg during mating, pregnancy, and the lactation period. Maternal exposure to TCDD had no significant effects on fetus/pup (F1) mortality, litter size, or sex ratio on gestation day (GD) 20 or postnatal day (PND) 2. The TCDD concentration in maternal livers and adipose tissue on GD20 was 1.21 and 1.81 ng/kg, respectively, and decreased at weaning to 0.72 in the liver and 0.84 in the adipose tissue. In contrast, the TCDD concentration in pup livers was 1.32 ng/kg on PND2 and increased to 1.80 ng/kg at weaning. Ventral prostate weight of male offspring was significantly decreased by TCDD exposure on PND28 and 120 compared with that of controls. Weight of the testes, cauda epididymides, and seminal vesicle, and sperm number in the cauda epididymis were not changed by TCDD exposure at PND120. TCDD- or vehicle-exposed male offspring were mated with unexposed females. The sex ratio (percentage of male pups) of F2 offspring was significantly reduced in the TCDD-exposed group compared with controls. These results suggest that in utero and lactational TCDD exposures affect the development of male gonads in offspring (F1), leading to changes in the sex ratio of the subsequent generation (F2).     

Safety evaluation of phytosterol esters. Part 3. Two-generation reproduction study in rats with phytosterol esters--a novel functional food.

Phytosterol esters (PE) are intended for use as a novel food ingredient with plasma cholesterol lowering activity which works by inhibiting the absorption of cholesterol from the gut. Although PE are naturally present in the normal diet, the levels are insufficiently large to ensure lowering of plasma cholesterol levels. Therefore PE may be added to spreads to achieve the desired cholesterol lowering activity. As part of an extensive programme of safety evaluation studies a two-generation reproduction study has been conducted in Wistar rats, in which the possible effect of PE on male and female reproductive performance and on the growth and development of the offspring was studied. Rats were fed diets containing PE at levels of 0, 1.6, 3.2 and 8.1% (w/w) PE over two successive generations, and a wide range of reproductive and developmental parameters, including sexual maturation parameters and oestrous cycle length, were determined. Macroscopic and microscopic examinations were conducted including a histological examination of selected organs from F1- and F2-weanlings and from F0- and F1-parental animals. Daily clinical observations did not reveal any unusual findings. In both generations, no effects of PE were observed on pup mortality (calculated on litter basis), precoital time, mating index, male and female fertility index, female fecundity index, gestation index, duration of gestation, number of females with stillborn pups, post-implantation loss and pup development. Furthermore, PE had no effect on sexual maturation parameters (preputial separation and vaginal opening) and oestrous cycle length. In addition, there were no dose-related effects on selected organs following histological examination. In conclusion, dietary administration of up to 8.1% PE (equivalent to a dose of 2.5 to 9.1 g PE/kg body weight/day, dependent on the period of the study) during two generations had no effect on reproduction of parental F0- and F1-generation Wistar rats, nor on the development of the F1- and F2-pups, nor on the sexual maturation of the F1-weanlings. Therefore, a nominal dietary PE concentration of 8.1% (equivalent to a dose of 2.5-9.1 g PE/kg body weight/day or 1.54-5.62 g phytosterol/kg body weight/day dependent on the period of the study) was considered to be the no-observed-adverse-effect level following daily oral administration of PE for two successive generations.     

Three-generation reproductive toxicity study of dietary bisphenol A in CD Sprague-Dawley rats.

Bisphenol A (BPA) was evaluated at concentrations of 0, 0.015, 0.3, 4.5, 75, 750, and 7500 ppm ( approximately 0.001, 0.02, 0.3, 5, 50, and 500 mg/kg/day of BPA) administered in the diet ad libitum to 30 CD((R)) Sprague-Dawley rats/sex/dose for 3 offspring generations, 1 litter/generation, through F3 adults. Adult systemic toxicity at 750 and 7500 ppm in all generations included: reduced body weights and body weight gains, reduced absolute and increased relative weanling and adult organ weights (liver, kidneys, adrenals, spleen, pituitary, and brain), and female slight/mild renal and hepatic pathology at 7500 ppm. Reproductive organ histopathology and function were unaffected. Ovarian weights as well as total pups and live pups/litter on postnatal day (PND) 0 were decreased at 7500 ppm, which exceeded the adult maximum tolerated dose (MTD). Mating, fertility, gestational indices; ovarian primordial follicle counts; estrous cyclicity; precoital interval; gestational length; offspring sex ratios; postnatal survival; nipple/areolae retention in preweanling males; epididymal sperm number, motility, morphology; daily sperm production (DSP), and efficiency of DSP were all unaffected. At 7500 ppm, vaginal patency (VP) and preputial separation (PPS) were delayed in F1, F2, and F3 offspring, associated with reduced body weights. Anogenital distance (AGD) on PND 0 was unaffected for F2 and F3 males and F3 females (F2 female AGD was increased at some doses, not at 7500 ppm, and was considered not biologically or toxicologically relevant). Adult systemic no observed adverse effect level (NOAEL) = 75 ppm (5 mg/kg/day); reproductive and postnatal NOAELs = 750 ppm (50 mg/kg/day). There were no treatment-related effects in the low-dose region (0.001-5 mg/kg/day) on any parameters and no evidence of nonmonotonic dose-response curves across generations for either sex. BPA should not be considered a selective reproductive toxicant, based on the results of this study.     

Peri- and postnatal developmental toxicity of the fluoroquinolone antibacterial DW-116 in rats.

DW-116 is a fluoroquinolone antibacterial developed by Dong-Wha Pharmaceutical Industry Co. The aim of this study is to determine the potential adverse effects of this chemical on pregnancy, delivery and lactation of dams and on peri- and postnatal development of F1 offspring. The test chemical was orally administered to pregnant rats from day 16 of pregnancy, through parturition and throughout the period of lactation up to weaning (postnatal day 21) at dose levels of 0, 10, 50, or 250 mg/kg/day. The progeny were examined at birth and subsequently to weaning. Mortality, body weight change, physical signs of postnatal development (pinna detachment, incisor eruption, fur development, eye opening, testis descent and vaginal opening) and behavioral function (righting reflex, negative geotaxis, grip-strength, pupillary reflex, acoustic startle response, rotating rod test, open field test and water-filled T-maze test) were evaluated. When the exposed offspring reached maturity (11 weeks old) their reproductive capacity was assessed. Maternal toxicity was observed only in the highest dose group and was limited to decreased food consumption during the late stage of pregnancy. However, this change was not observed during the lactation period. There were no adverse effects on mortality, clinical signs, body weight, necropsy findings, organ weight of dams in any treatment group. No adverse effects on the offspring were seen with the low and middle doses tested, but the highest dose increased postnatal mortality. The number of stillborn was also increased at the highest dose but the difference was not statistically significant. Meanwhile, no treatment-related effects were observed in clinical sign, developmental and behavioral landmarks and necropsy findings at any dose levels tested. There were no treatment-related effects on the mating of the F1 generation and resulting F2 offspring. The results of this study indicate that the peri- and postnatal administration of DW-116 to female rats results in an increase in postnatal mortality at a minimally maternotoxic dose, i.e., 250 mg/kg/day. Under the experimental conditions, the no-observed-adverse-effect level for peri- and postnatal developmental toxicity was considered to be 50 mg/kg/day.     

Reproductive and neurobehavioural toxicity study of Ponceau 4R administered to mice in the diet.

Ponceau 4R was given to mice in the diet at levels of 0 (control), 0.12%, 0.24%, and 0.48% from 5 weeks of age of the F(0) generation to 9 weeks of age of the F(1) generation, and selected reproductive and neurobehavioural parameters were measured. There was no adverse effect of Ponceau 4R on litter size, litter weight or sex ratio at birth. The average body weight of male and female offspring was increased significantly in the high-dose group at postnatal days (PNDs) 0, 4 and 21. In behavioural developmental parameters, surface righting at PND 4 was affected significantly in the high-dose group in male offspring. Other variables measured showed no consistently significant adverse effect on either sex in the lactation period. In multiple water T-maze performance in the F(1) generation, the time taken was significantly longer than the control in the middle-dose and high-dose groups in males, and those effects were significantly dose-related (P<0.01). The dose level of Ponceau 4R in the present study produced no adverse effect on reproduction, and a few adverse effects on neurobehavioural parameters in mice. The non-observed adverse effect level (NOAEL) was presumed to be 0.12% in the diet (approximately 205mg/kg per day) for maze learning by males in the F(1) generation. Nevertheless, the middle-dose and high-dose levels were in excess of the acceptable daily intake (ADI) of Ponceau 4R (0-4.0mg/kg body weight), and the actual dietary intake of Ponceau 4R in humans is presumed to be much lower. It would appear, therefore, that the level of dietary intake of Ponceau 4R is unlikely to produce any adverse reproductive or neurobehavioural effect in humans.     

Developmental and reproductive toxicity evaluation of toluene vapor in the rat. I. Reproductive toxicity

The reproductive toxicity of toluene was evaluated in a 2-generation test in which male and female Sprague–Dawley rats, parental (F0) and first generation (F1), were exposed to toluene via whole body inhalation, 6 h/day, 7 days/week for 80 days premating and 15 days of mating at concentrations of 0, 100, 500 and 2000 ppm (0, 375, 1875 and 7500 mg/m³). Toluene was administered at 2000 ppm to both sexes, or to females or males only to be mated with untreated partners. Pregnant females at all dose levels were exposed from gestation day (GD) 1–20 and lactation day (LD) 5–21. At LD5, females were removed from their litters for daily exposure and returned when 6 h of exposure was completed. F1 pups selected to produce the F2 generation were treated for 80 days beginning immediately after weaning (LD21) and initially mated at a minimum of 100 days of age. F2 pups were not exposed to toluene by inhalation.

Toluene exposure did not induce adverse effects on fertility, reproductive performance, or maternal/pup behaviors during the lactation period in males and females of the parental or first generation, but did inhibit growth in F1 and F2 offspring in the 2000 ppm (both sexes treated) and 2000 ppm (females only treated) groups. Caesarean section of selected 2000 ppm (both sexes treated) dams at GD20 showed reduced fetal body weight and skeletal variations. Exposure to toluene caused decreased pup weights throughout lactation in F1 and F2 2000 ppm (both sexes treated), and 2000 ppm (females only treated) groups. Exposure at 2000 ppm to male parents only did not induce similar weight inhibition in offspring. The toluene offspring NOAEL is 500 ppm in groups in which maternal animals were exposed, and 2000 ppm for male only treated groups.