Toxicokinetics of phenolphthalein in male and female rats and mice.

Phenolphthalein (PTH), which has been used as the active ingredient in a number of prescription and over-the-counter laxative products, is a rodent chemical carcinogen in multiple organs in the NTP 2-year bioassay at doses of 291-2927 mg/kg. This paper describes the toxicokinetics and estimates the internal dose of PTH administered as a single iv or gavage dose, or ad libitum for 14 days in feed to F344 rats, B6C3F1 mice, p53 (+/-) mice, and C57BL mice at doses that bracketed those used in the bioassay. Plasma concentrations for free phenolphthalein (PTH-F) and phenolphthalein glucuronide (PTH-G) were obtained for each dose regimen. Total phenolphthalein (PTH-T) was calculated as the sum of the molar concentrations of PTH-F and PTH-G. Noncompartmental pharmacokinetic models were used to calculate the area under the curve (AUC) from 0 h to infinity (AUC(infinity)), clearance (Cl), and oral bioavailability (F) for PTH-F; and were used to calculate AUC(infinity), t((1/2)), and relative absorption (Q) for PTH-T. After iv administration, PTH-F rapidly declined in rats and mice; PTH-T rose rapidly to Cmax and slowly declined 6-8 h after dosing, with no sex-related differences for rats or mice. For feed studies, mean plasma concentration (f1.gif" BORDER="0">(infinity)) and 24-h area under the curve (AUC(24h)) values were calculated. Results from feed studies showed no dose response in rat plasma PTH-F above approximately 50 mg/kg. Rat PTH-T AUC(24h) and f1.gif" BORDER="0">(infinity) were linear with doses up to approximately 650 mg/kg. In B6C3F1 mice, PTH-F and PTH-T AUC(24h) increased nonlinearly with doses above approximately 165 mg/kg. PTH is well absorbed and readily converted to PTH-G when administered in feed to rats and mice, except at the highest bioassay doses, where PTH absorption may be saturated.

Safety assessment, in vitro and in vivo, and pharmacokinetics of emivirine, a potent and selective nonnucleoside reverse transcriptase inhibitor of human immunodeficiency virus type 1.

Emivirine (EMV), formerly known as MKC-442, is 6-benzyl-1-(ethoxymethyl)-5-isopropyl-uracil, a novel nonnucleoside reverse transcriptase inhibitor that displays potent and selective anti-human immunodeficiency virus type 1 (HIV-1) activity in vivo. EMV showed little or no toxicity towards human mitochondria or human bone marrow progenitor cells. Pharmacokinetics were linear for both rats and monkeys, and oral absorption was 68% in rats. Whole-body autoradiography showed widespread distribution in tissue 30 min after rats were given an oral dose of [(14)C]EMV at 10 mg/kg of body weight. In rats given an oral dose of 250 mg/kg, there were equal levels of EMV in the plasma and the brain. In vitro experiments using liver microsomes demonstrated that the metabolism of EMV by human microsomes is approximately a third of that encountered with rat and monkey microsomes. In 1-month, 3-month, and chronic toxicology experiments (6 months with rats and 1 year with cynomolgus monkeys), toxicity was limited to readily reversible effects on the kidney consisting of vacuolation of kidney tubular epithelial cells and mild increases in blood urea nitrogen. Liver weights increased at the higher doses in rats and monkeys and were attributed to the induction of drug-metabolizing enzymes. EMV tested negative for genotoxic activity, and except for decreased feed consumption at the high dose (160 mg/kg/day), with resultant decreases in maternal and fetal body weights, EMV produced no adverse effects in a complete range of reproductive toxicology experiments performed on rats and rabbits. These results support the clinical development of EMV as a treatment for HIV-1 infection in adult and pediatric patient populations.

Formamide and dimethylformamide: reproductive assessment by continuous breeding in mice.

Reproductive toxicity in Swiss mice, during chronic exposure to formamide (FORM) or dimethylformamide (DMF), was evaluated using the Reproductive Assessment by Continuous Breeding Protocols. FORM administered in drinking water at 0, 100, 350, and 750 ppm (approximately 20 to 200 mg/kg/d) reduced fertility and litter size in F0 animals without generalized toxicity at 750 ppm FORM. Crossover matings suggested that females were the affected sex. After F1 mating, FORM reduced F2 litter size, increased days to litter, reduced relative ovarian weight, and lengthened estrous cycles at 750 ppm. The No-Observed-Adverse-Effect-Level for generalized toxicity was 750 ppm for the F0 and 350 ppm for the F1 generation. Reproductive performance was normal at 350 ppm for both F0 and F1 mice. Chronic exposure to DMF in drinking water at 0, 1000, 4000, and 7000 ppm (approximately 200 to 1300 mg/kg/d) reduced fertility by the first litter at 4000 ppm, reduced body weight in F0 females at 7000 ppm, and increased liver weights at all doses in both sexes. A crossover mating at 7000 ppm identified F0 females as the affected sex. F1 postnatal survival was reduced at > or =4000 ppm DMF. F1 mating reduced F2 litter size and live pup weight at > or =1000 ppm. At necropsy, body weight of F1 males and females was reduced at > or =4000 ppm. DMF-treated pups (both F1 and F2) and F1 adults had cranial and sternebral skeletal malformations. Only DMF caused overt developmental toxicity. A No-Observed-Adverse-Effect-Level for DMF was not established.

Nitrofurazone: reproductive assessment by continuous breeding in Swiss mice.

Nitrofurazone (NTFZ), a nitrofuran antibiotic, was evaluated for reproductive toxicity in Swiss CD-1 mice using the Reproductive Assessment by Continuous Breeding protocol. Male and female mice were cohabited for 15 weeks and exposed to NTFZ in feed at concentrations of 0, 100, 375, and 750 ppm (14-102 mg/kg/day). Fzero 750-ppm breeding pairs had significantly reduced fertility after 7 days of exposure to NTFZ (17% fertile compared to 98% for control pairs) and were infertile after the second litter. Fzero mid-dose pairs had progressively decreasing fertility (47% by the fifth litter), reduced litter size, and reduced proportion of pups born alive. Crossover breeding of control and high-dose Fzero animals confirmed infertility in high-dose males and reduced litter size and pup weight in high-dose females when compared to the control x control group. At necropsy, there were no effects on body weight, but Fzero males had reduced testis weight at the high dose and reduced epididymal sperm concentration and abnormal sperm morphology at all doses of NTFZ. Increased liver as well as kidney and adrenal weights (combined) were observed at 375 and 750 ppm; hepatic hypertrophy was noted microscopically at 750 ppm. Fzero females had reduced body weight, hepatic hypertrophy, and altered estrous cycles at 750 ppm and reduced ovarian weight at all doses. In the second generation, F1 mice at 375 ppm had reduced postnatal survival and body weight and produced smaller F2 litters compared to control mice. At necropsy, F1 males had reduced testes weight and epididymal sperm concentration, abnormal sperm morphology, hepatic hypertrophy at 375 ppm, and borderline nephropathy at 100 and 375 ppm. F1 females had decreased body, liver, and ovarian weight at 375 ppm and altered estrous cycles at 100 and 375 ppm. Thus, NTFZ at > or = 100 ppm (> or = 14 mg/kg/day) caused adverse reproductive effects in Fzero male and female and F1 female mice in the presence of relatively mild systemic toxicity.

The reproductive and neural toxicities of acrylamide and three analogues in Swiss mice, evaluated using the continuous breeding protocol.

Acrylamide is a known genetic, reproductive, and neural toxicant, although it is not known if one effect is predominant. The toxicities of several structural analogues of acrylamide have been incompletely characterized, and the relative sensitivity of the second generation is not known. The present studies were designed to explore the relationship between neurotoxicity and reproductive toxicity, to further characterize the toxicities of three acrylamide analogues, and to evaluate the relative sensitivity of a second generation to these compounds. For the F0 generation, male and female Swiss CD-1 mice were provided drinking water containing acrylamide (ACR; 3, 10, 30 ppm), N,N'-methylenebisacrylamide (MBA; 10, 30, 60 ppm), N-(hydroxymethyl)acrylamide (HMA; 60, 180, 360 ppm), or methacrylamide (MACR; 24, 80, 240 ppm) during and after a 14-week cohabitation. The last litter was reared and dosed after weaning until mating at 74 +/- 10 days of age with the same level of compound given to the parents Neurotoxicity was assessed at several times in both generations by measuring forelimb and hindlimb grip strength. In the F0 generation, ACR caused an 11% decrease in pup number without measurable neurotoxicity; female fertility was not affected. Although both generations consumed the same amount of ACR, there were larger changes in the fertility-related endpoints in the F1 mice than in the F0's, with no concomitant change in organ weights or sperm parameters. In F0 mice, MBA reduced the number of live pups and their adjusted weight, with no neurotoxicity and no change in F0 female reproduction. MBA caused greater adverse effects in the second generation, concomitant with increased consumption. In the F0 generation, HMA caused the largest decrease in pup number during cohabitation (26%) together with a small effect on grip strength. Female reproduction was not affected. The second generation consumed more HMA and showed slightly greater toxic effects. In both generations, MACR was negative for both neurotoxicity and reproductive toxicity. Dominant-lethal studies showed that the fertility effects for ACR, MBA, and HMA could be explained by a male-mediated increase in postimplantation loss. These studies found that dominant lethality occurred without structural effects on the reproductive system in the presence of only minor effects on grip strength and without detectable neural histopathology. Female reproduction was not significantly affected by these compounds at the doses used. Thus, these data confirm the male as the affected gender and that the reproductive toxicity was greater than motoneuron toxicity when measured as grip strength.

Carisoprodol: reproductive assessment by continuous breeding in Swiss mice.

Carisoprodol (CARI), a commonly prescribed neuromuscular relaxant, was evaluated for reproductive toxicity in Swiss CD-1 mice using the Reproductive Assessment by Continuous Breeding (RACB) protocol. Male and female mice were given CARI in corn oil suspension by daily gavage at doses of 0, 300, 750, and 1200 mg/kg body wt/day. Clinical signs of general toxicity in F0 animals included sedation, primarily in the high-dose group during the first week of exposure, and reduced body weight in high-dose females. CARI administration for 14 weeks did not affect the ability of the F0 animals to produce litters. However, decreases in proportion of pups born alive (4%) and absolute (5%) and adjusted live pup weight (7%) were observed at 1200 mg/kg CARI when compared to controls. In a crossover mating trial to determine the affected sex, there were no significant differences in the measured reproductive parameters. CARI at the high dose increased the proportion of time spent in proestrus and estrus, but cycle length was unaffected. At F0 necropsy (Week 27 of treatment), all sperm parameters were normal. Right epididymis and liver weights, relative to body weight, were increased (12 and 23%, respectively) over the control group for high-dose males. A mating trial to determine the fertility and reproductive competence of the F1 generation showed no effect of CARI on indices of mating, pregnancy, or fertility, the proportion of F2 pups born alive, the sex ratio of live F2 pups, live F2 pup weight, or gestation length.(ABSTRACT TRUNCATED AT 250 WORDS)

Assessment of the reproductive and developmental toxicity of pesticide/fertilizer mixtures based on confirmed pesticide contamination in California and Iowa groundwater.

Pesticides and fertilizers, as used in modern agriculture, contribute to the overall low-level contamination of groundwater sources. In order to determine the potential of pesticide and fertilizer mixtures to produce reproductive or developmental toxicity at concentrations up to 100 x the median level found in groundwater, we prepared and studied two mixtures of pesticides and a fertilizer (ammonium nitrate). One mixture containing aldicarb, atrazine, dibromochloropropane, 1,2-dichloropropane, ethylene dibromide, and simazine plus ammonium nitrate was considered to be a representative of groundwater contamination in California (CAL). The other, containing alachlor, atrazine, cyanazine, metolachlor, metribuzin, and ammonium nitrate, simulated groundwater contamination in Iowa (IOWA). Each mixture was administered in the drinking water of either Swiss CD-1 mice during a Reproductive Assessment by Continuous Breeding study or pregnant Sprague-Dawley rats (gd 6-20) at three dose levels (1x, 10x, and 100x) where 1x was the median concentration of each pesticide component as determined in the groundwater surveys in California or Iowa. Unlike conventional toxicology studies, the purpose of this study was to evaluate the health effects of realistic human concentrations. Thus, the testing concentrations are probably well below the maximally tolerated dose. Propylene glycol was used as the solubilizer for the pesticides in drinking water formulations in both studies. In the reproductive study, neither mixture caused any clinical signs of toxicity, changes in food or water consumption, or body weight in either F0 or F1 mice at doses up to 100x the median groundwater concentrations. There were no treatment-related effects on fertility or any measures of reproductive performance of either the F0 or the F1 generation mice exposed to either CAL or IOWA at up to 100x. Similarly, measures of spermatogenesis, epididymal sperm concentration, percentage motile sperm, percentage abnormal sperm, and testicular and epididymal histology were normal. In the developmental study, CAL- or IOWA-exposed females did not exhibit any significant treatment-related clinical signs of toxicity. No adverse effects of CAL or IOWA were observed for measures of embryo/fetal toxicity, including resorptions per litter, live litter size, or fetal body weight. CAL or IOWA did not cause an increased incidence of fetal malformations or variations. In summary, administration of these pesticide/fertilizer mixtures at levels up to 100-fold greater than the median concentrations in groundwater supplies in California or Iowa did not cause any detectable reproductive (mice), general, or developmental toxicity (rats).

Reproductive effects of 4-vinylcyclohexene in Swiss mice assessed by a continuous breeding protocol.

4-Vinylcyclohexene (VCH), a dimer of 1,3-butadiene, was evaluated for reproductive toxicity in Swiss (CD-1) mice using the continuous breeding protocol (NTP, 1989). VCH in corn oil was administered by gavage at doses of 0, 100, 250, and 500 mg/kg/day to animals that were housed in same sex pairs for 1 week and then cohabited in breeding pairs for 14 weeks. During cohabitation, newborn litters were euthanized immediately after evaluation on postnatal Day (PND) 0. Litters born after Week 15 were reared until PND 21, when all F0 animals and low- and mid-dose F1 weanlings were humanely killed without a necropsy. At PND 74 +/- 10, control and high-dose F1 animals were cohabited within groups for 1 week and necropsied after delivery of the litters. In F0 breeding pairs, VCH did not affect measures of reproductive competence, including initial fertility, litters per pair, live litter size, or the proportion of pups born alive. Pup weight was decreased (4%) in the high-dose group relative to controls. High-dose F0 females exhibited slight general toxicity, manifested as an 8% difference in body weight compared to controls. VCH did not adversely affect preweaning growth or survival in the F1 generation. VCH had no effect on the reproductive competence of the F1 generation. High-dose F1 adult males and females had decreased body weight. At necropsy, increased relative liver weight (males 9% and females 8%) and sperm motility (although not thought to be biologically significant) were observed in the 500 mg/kg VCH group.(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of feed restriction on reproductive function in Swiss CD-1 mice.

Many test compounds used in toxicity studies produce a "systemic" toxicity manifested as reduced body weight gain. While it is known that reduced weight gain during juvenile growth reduces or delays reproductive competence, the effects of adult-onset feed restriction (FR) and inhibited weight gain on the reproductive system of mice are poorly known. To gain some information on the effects of graded body weight reduction, or reduced body weight gain, on commonly used reproductive endpoints, the studies reported below were conducted at two laboratories, using adult mice that were maintained at 90, 80, and 70% of concurrent control body weight (CBW) for up to 21 weeks. Estrous cyclicity and fertility in the females were significantly affected. While male fertility was variably affected, there was a significant decrease in the number of epididymal sperm and in the number of testicular spermatids in the 70% CBW groups. Testis weight was conserved in both studies; relative testis weight increased in all FR groups. These data can improve the interpretation of future studies by helping to separate chemically induced changes from those produced by reduced body weight gain.

Reproductive toxicity of boric acid in Swiss (CD-1) mice: assessment using the continuous breeding protocol.

The potential reproductive toxicity of boric acid (BORA) in CD-1 mice (Swiss) was evaluated using the Reproductive Assessment by Continuous Breeding (RACB) Protocol. BORA was administered in the feed for 27 weeks to male and female Swiss (CD-1) mice at concentrations of 0, 1000, 4500, or 9000 ppm. Estimated doses, based on feed consumption and body weight, averaged 152, 636, and 1262 mg/kg body wt during Week 1 for males for 1000, 4500, and 9000 ppm, respectively. During 14 weeks of cohabitation, fertility of F0 mice was partially reduced at 4500 ppm and totally eliminated at 9000 ppm. No litters, dead or alive, were produced by 9000 ppm cohabited pairs. Among the litters born at 4500 ppm, live litter size and body weight were significantly reduced. A crossover mating trial of control and 4500 ppm groups confirmed the male as the affected sex, with fertility rates and the mating index significantly lower in the 4500 male x 0 ppm female group. At necropsy, after 27 weeks of BORA exposure, dose-related changes were present in F0 males for reduced body and reproductive organ weights, increased incidence of abnormal sperm, decreased sperm concentration and motility, and seminiferous tubule degeneration. In the 4500 ppm females, dietary BORA for 27 weeks caused significantly decreased weights of kidney/adrenals and livers; kidney/adrenal weight was also reduced in 4500 ppm males. The last litters of the control and 1000 ppm females, born in the 14-week breeding phase, were reared to 74 days of age and then mated in nonsibling pairs within treatment groups. These F1 mice had normal fertility, but the adjusted mean body weight of F2 pups was decreased. These data establish the reproductive toxicity of BORA in CD-1 mice and demonstrate that the male is the most sensitive sex.

Active immunization of boars against gonadotrop in releasing hormone. I. Effects on reproductive parameters.

Sexually mature boars were actively immunized against gonadotropin releasing hormone (GnRH) to characterize endocrine and gametogenic changes associated with immunoneutralization of endogenous GnRH. Injections of GnRH conjugated to bovine serum albumin (BSA) given five times over 24 wk induced production of antibodies against GnRH in all animals (n=5). Active immunization against GnRH reduced serum concentrations of testosterone (P<0.05) and luteinizing hormone (LH) (P<0.05), testes volume (P<0.01), paired testis weights (P<0.05), paired epididymis weights (P<0.05), sperm per testis (P<0.01) and seminiferous tubule diameters (P<0.001) when compared with controls (n=4). These results indicate that both steroidogenic and spermatogenic functions are impaired in testes of mature boars actively immunized against GnRH.

Triphenyl phosphite: in vivo and in vitro inhibition of rat neurotoxic esterase.

Organophosphorus compounds which, after acute administration, inhibit neurotoxic esterase (NTE) by greater than or equal to 65% and undergo a subsequent "aging" reaction, produce a delayed neuropathy characterized by degeneration of large and long nerve fibers (OPIDN). The present studies examine in detail the NTE-inhibiting properties of triphenyl phosphite (TPP), a plasticizer which produces ataxia and degeneration of the spinal cord in animals. A neurotoxic dosing regimen (1184 mg/kg/week, sc, for 2 weeks) inhibited both brain and spinal cord NTE (less than or equal to 40%) only marginally 4 and 48 hr postdosing. By contrast, TPP was shown in vitro to be a potent (150 = 0.98 microM) inhibitor of rat brain NTE relative to Mipafox or diisopropyl phosphorofluoridate. Compounds structurally related to TPP (i.e., triphenyl phosphate, triphenyl phosphine, trimethyl phosphite, and phenol) failed to inhibit NTE in vitro at less than 10 microM concentrations. Close examination of the TPP inhibition of NTE showed a nonlinear relationship between the duration of incubation time and loss of log(NTE activity). Preincubation of 10 microM TPP in buffer (37 degrees C) resulted in a time-dependent loss of TPP's ability to inhibit NTE. In summary, TPP is a powerful NTE inhibitor in vitro, but only a marginal NTE inhibitor after in vivo administration. These results raise questions as to the causal events mediating TPP-induced neuropathy in the rat.