Metabolism and testicular toxicity of 1,3-dinitrobenzene in the rat: effect of route of administration.

Studies investigating the testicular toxicity of 1,3-dinitrobenzene (1,3-DNB) have utilized both the oral (po) and intraperitoneal (ip) routes of administration. These two administration routes could be expected to produce different pharmacokinetic profiles and, potentially, different degrees of toxicity. In the present work, the effect of route of administration upon 1,3-DNB disposition and susceptibility to testicular damage has been investigated. Male Sprague-Dawley rats were given 25 mg/kg 1,3-DNB either ip or po. Metabolites were quantitated in blood, urine, and feces, and methemoglobin levels were determined. Peak blood levels of 1,3-DNB and its major metabolite were three times higher in ip-dosed rats than in po-dosed rats. While the lower blood levels seen after po administration were maintained for greater than 6 hr, blood levels fell rapidly after ip dosing, reaching po levels at 6 hr postadministration. Peak methemoglobin levels in ip-dosed animals were twice that of po-dosed animals. Route of administration had a minor effect on the levels of urinary metabolites, while there was a significantly higher excretion of metabolites in the feces of po-dosed animals. Despite the markedly higher 1,3-DNB blood levels after ip administration, there were only subtle differences in testicular damage. The data raise the possibility that above a threshold level of 1,3-DNB in the blood, only the duration of testicular exposure to the toxicant may govern susceptibility to testicular toxicity.

Metabolism and testicular toxicity of 1,3-dinitrobenzene in rats of different ages.

Susceptibility to 1,3-dinitrobenzene (1,3-DNB)-induced testicular damage is known to increase with age. The present study investigated the possibility that age-dependent differences in metabolism and disposition could account for differences in toxicity. [14C]1,3-DNB (25 mg/kg, ip) was administered to Sprague-Dawley rats which were 31, 75, or 120 days of age. Levels of 1,3-DNB and 1,3-DNB metabolites were determined in blood and urine. As animal age increased, peak blood concentrations of 1,3-DNB were lower and declined more slowly indicating an age-dependent decrease in rate of metabolism and a possible increase in volume of distribution. In younger animals, faster elimination rates were associated with higher blood levels of metabolites. Urinary metabolites were generally similar for all age groups with the exception of the diacetamidobenzene metabolite which was significantly lower in the urine of 31 days old rats. There were clear differences in the toxicokinetic profile for 1,3-DNB between the 31 day old rats and the other two age groups. However, differences between the 75 and 120 day old animals were less marked. Testicular damage induced by 1,3-DNB (25 mg/kg, ip) was hardly detectable in the youngest animals, while the intermediate age group showed a moderate lesion particularly in later stages of spermatogenesis. For the oldest animals, testicular damage was more severe, particularly in the earlier stages of spermatogenesis. Overall, the rapid elimination rate could account for the lack of 1,3-DNB toxicity in very young animals. However, simple metabolic differences were less likely to adequately explain the increase in testicular damage found as animal age increased from 75 to 120 days.

Application of ELISA techniques to metabolic disposition studies for 1,3-dinitrobenzene: comparison with HPLC and radiochemical methods.

An enzyme-linked immunosorbent assay (ELISA) is reported for the detection and quantitation of 1,3-dinitrobenzene, a widely used chemical intermediate in industrial syntheses and potent testicular toxicant in rats. Radiolabeled [14C]-1,3-dinitrobenzene (25 mg/kg, ip) was administered to rats, and the blood disappearance curve generated with the ELISA assay was compared with HPLC separation and quantitation by using both UV and radiochemical methods. An excellent correlation was found between the two methods. Matrix effects from the blood samples were minimal. Cross reactivity with metabolites of 1,3-dinitrobenzene occurred only at levels that were 3 orders of magnitude greater than for the parent compound. The specificity of the ELISA for 1,3-dinitrobenzene was further demonstrated with a variety of other nitroaromatic compounds, of which only 2,4-dinitrotoluene showed significant cross reactivity. The sensitivity and specificity of the 1,3-dinitrobenzene ELISA demonstrate the usefulness of an approach which, if verified with conventional analytical methods, allows rapid and inexpensive measurement of xenobiotic levels in biological samples.

Metabolism and pharmacokinetics of 1,3-dinitrobenzene in the rat and the hamster.

Recent studies have documented that the Sprague-Dawley rat is markedly more sensitive than the Golden Syrian hamster to 1,3-dinitrobenzene (1,3-DNB)-induced testicular toxicity and methemoglobinemia. The present studies have investigated the possibility that differences in 1,3-DNB metabolism and pharmacokinetics could explain this species difference. [14C]1,3-DNB (25 mg/kg, ip) was administered to both species at a dose known to induce a testicular lesion in the rat and levels of 1,3-DNB and its metabolites were measured in blood and urine. Elimination of 1,3-DNB from the blood was initially rapid, followed by a second, much slower phase. Peak blood levels of 1,3-DNB were very different between the two species, with the hamster reaching levels only one-half those found in the rat (46.3 vs. 99.5 nmol/ml, respectively). Administration of a 50-mg/kg dose to the hamster resulted in 1,3-DNB blood levels which were similar to those found in the rat at 25 mg/kg. Since no obvious testicular toxicity is apparent in the hamster even at the higher dose level, a direct effect of 1,3-DNB on the testes seems unlikely, although it is possible that hamster cells inherently lack sensitivity to toxicity. Other major differences between the two species were a more rapid initial elimination rate and much higher blood levels of nitroaniline in the rat. Analysis of urinary metabolites revealed that the rat excreted more unconjugated metabolites and less phenolic metabolites compared with the hamster. Overall, the data indicate that the capacity to form reductive metabolites may play an important role in susceptibility to toxicity.

Acetylcholinesterase and neuropathy target esterase in chickens treated with acephate.

Reports that near-lethal doses of the pesticide methamidophos (O,S-dimethyl phosphoramidothioate) caused a delayed neurotoxicity (OPIDN) in humans and that another phosphoramidate, isofenphos, caused OPIDN in the hen at high doses, prompted a study of the abilities of acephate (O,S-dimethyl acetylphosphoramidothioate) to inhibit brain acetylcholinesterase (AChE) and neuropathy target esterase (NTE) in vivo. Hens were treated orally with 5-700 mg/kg of acephate, or im with 50-200 micrograms/kg of diisopropyl-fluorophosphate (DFP, positive control) and sacrificed 24 hr later. Brain homogenates were assayed for AChE as an estimate of acute toxicity, for NTE to indicate acephate's potential to cause OPIDN, and for residues of acephate and its metabolite methamidophos. A range finding study confirmed the LD50 level for acephate was approximately 800 mg/kg. Regression analyses indicated an ID50 (a dose that inhibits 50% of activity) for acephate inhibition of AChE of 10 mg/kg and an extrapolated ID50 for inhibition of NTE of 1300 mg/kg, almost twice the LD50. In contrast, ID50 values for DFP were similar for AChE (146 micrograms/kg) and NTE (132 micrograms/kg). Brain methamidophos levels were 10 to 16 percent of the total acephate plus methamidophos brain concentration. The lower the dose of acephate, the higher was the relative percentage of methamidophos. The results show acephate is a more potent inhibitor of AChE than it is of NTE in hens and suggest it would be difficult to administer a single dose of acephate sufficient to cause OPIDN without killing the animal.