Renal damage, metabolism and covalent binding following administration of the nephrotoxicant N-(3,5-dichlorophenyl)succinimide (NDPS) to male Fischer 344 rats.

In vivo metabolism, nephrotoxicity and covalent binding to proteins were evaluated in male Fischer 344 rats that received [2,3-¹⁴C]-N-(3,5-dichlorophenyl)succinimide (¹⁴C-NDPS). Some animals were pretreated with the enzyme inducer phenobarbital (PB, 80 mg/kg per day, for 3 days, i.p. in saline) prior to receiving a non-nephrotoxic dose of ¹⁴C-NDPS (0.2 mmol/kg, i.p. in corn oil). Other rats were pretreated with the cytochrome P450 inhibitor 1-aminobenzotriazole (ABT, 100 mg/kg, 1 h prior to NDPS, i.p. in saline) before administration of a non-toxic or a toxic dose (0.2 or 0.6 mmol/kg, respectively, i.p. in corn oil) of ¹⁴C-NDPS. Non-pretreated animals received either dose of ¹⁴C-NDPS, but did not receive PB or ABT. All rats were sacrificed 6 h after administration of ¹⁴C-NDPS. Nephrotoxicity was monitored by measuring urine volume, urine protein concentrations, blood urea nitrogen levels, and kidney weights. The NDPS metabolic profile in tissue, blood, and urine was analyzed by HPLC. Covalent binding of ¹⁴C-NDPS-derived radioactivity to tissue proteins was also measured. Compared with non-pretreated rats, PB-pretreatment potentiated the toxicity of the non-toxic dose of ¹⁴C-NDPS. In contrast, ABT-pretreatment protected the rats against NDPS nephrotoxicity. The amount of N-(3,5-dichlorophenyl)-2-hydroxysuccinamic acid (2-NDHSA), an oxidative, nephrotoxic metabolite of NDPS, was elevated in kidney homogenates and urine by PB-pretreatment (0.2 mmol/mg NDPS). ABT pretreatment inhibited NDPS metabolism at both doses. Covalent binding of ¹⁴C-NDPS (0.2 mmol/kg)-derived radioactivity to renal and plasma proteins was higher in the PB-pretreated rats than in the non-pretreated animals. In contrast, ABT-pretreatment partially inhibited covalent binding at both doses of ¹⁴C-NDPS. Our results suggest that there is a relationship between oxidative metabolism of NDPS, covalent binding of an NDPS metabolite to renal proteins, and NDPS-induced nephrotoxicity in rats.     

Studies on benzyl acetate. I. Effect of dose size and vehicle on the plasma pharmacokinetics and metabolism of [methylene-14C]benzyl acetate in the rat

Male Fischer 344 rats received [methylene-¹⁴C]benzyl acetate by gavage in a dose of 5, 250 or 500 mg/kg, as the neat substance, in corn oil or in propylene glycol. Urine and faeces were collected and urinary metabolites were assayed by radio-TLC and HPLC. Other animals were killed at various times and exsanguinated, and plasma levels of ¹⁴C in plasma occurred earliest and were highest when benzyl acetate was given neat. Peak levels were lower and absorption was delayed with the propylene glycol vehicle. The use of corn oil as the dose vehicle at the higher doses (250 and 500 mg/kg) led to the maintenance of plateau plasma levels, at about one half of the peak levels seen with the neat compound, for up to 8 hr after administration. At the 5 mg/kg dose, the plasma levels of ¹⁴C were essentially the same whether the dose was given in corn oil or propylene glycol. At the 250- and 500-mg/kg doses, at all time points, the major metabolite in plasma was benzoic acid, accompanied by smaller amounts of hippuric acid. Benzyl alcohol was also detected in some plasma samples. At the 5-mg/kg dose, the major plasma metabolite was hippuric acid, together with a smaller amount of benzoic acid. When propylene glycol was used as the vehicle at this dose level, benzylmercapturic acid was also present in the plasma. The major urinary metabolite was hippuric acid (c. 66% of the dose), with benzoic acid (2%) and benzylmercapturic acid (1%) also present. The elimination of benzoyl glucuronide increased with increasing dose, from c. 3 to 11% of the dose.     

Metabolism of 3, 4-dihydroxyphenylalanine by isolated perfused rat liver

The metabolism of -3, 4-dihydroxyphenylalanine ( -dopa) and -dopa by the isolated perfused rat liver is described. After injection of 5 μc(192μg) -dopa-3-¹⁴C into the portal vein, the liver took up approximately one-third of the dose within 30 min. Blood cells of the perfusate (chiefly erythrocytes) took up one-fourth of the dose within 5 min; subsequently, most of this radioactivity was released back into plasma. -Dopa disappeared from plasma rapidly (half-life, approximately 30 min); after 2 hr of perfusion, less than 2 per cent of the dose could be recovered unmetabolized. Metabolites of dopa released from liver into bile accounted for up to 48 per cent of the dose and were chiefly glucuronides of N-acetyldopamine and N-acetyl-3-methoxydopamine. Major metabolites released into plasma were the glucuronides of 3-methoxy4-hydroxyphenylacetic acid and 3, 4-dihydroxyphenylacetic acid, the sulfate of N-acetyl-3-methoxydopamine, and 3-methoxy-4-hydroxyphenylalanine. Amines in plasma represented a minor fraction (less than 1 per cent of the dose) at all times during the perfusion. Norepinephrine and its metabolites were never detected. After injection of 5 μc -dopa-2-¹⁴C (32 or 192μg), less ¹⁴C was taken up by liver than when -dopa was used; similar amounts of ¹⁴C were taken up by the blood cells but were retained for the 5 hr of perfusion. -Dopa disappeared more slowly from plasma than -dopa; after 2 hr, 16 per cent of the dose was recovered unmetabolized. The metabolites of -dopa in bile (22 per cent of the dose) and in plasma were similar to those found when -dopa was substrate.     

Studies on benzyl acetate. II. Use of specific metabolic inhibitors to define the pathway leading to the formation of benzylmercapturic acid in the rat

Specific metabolic inhibitors were used to define the route of metabolism of benzyl acetate leading to the formation of benzylmercapturic acid. Male Fischer 344 rats were dosed by gavage with [methylene-¹⁴C]benzyl acetate (500 mg/kg) alone or together with pyrazole (200 mg/kg), pentachlorophenol (10 mg/kg) or both pentachlorophenol (10 mg/kg) and pyrazole (200 mg/kg), given in each case ip. Urine and faeces were collected and urinary metabolites were assayed by radio-TLC and HPLC. The excretion of ¹⁴C was rapid in all cases, with most of the dose excreted in the urine within 24 hr. Co-administration of pyrazole (an inhibitor of alcohol dehydrogenase) with benzyl acetate caused an 11-fold increase in the excretion of benzylmercapturic acid and halved the percentage of the dose excreted as benzoyl glucuronide. Pretreatment with pentachlorophenol, an inhibitor of sulphotransferase activity in vivo, abolished the excretion of benzylmercapturic acid, while excretion of the mercapturate following treatment with both pyrazole and pentachlorophenol was higher than in control or pentachlorophenol-treated rats, but much lower than in the animals given pyrazole alone. Taken together, these results suggest very strongly that the formation of benzylmercapturic acid involves the sulphate ester of benzyl alcohol as an obligatory intermediate and does not appear to involve a metabolic intermediate with sufficient reactivity to have toxicological relevance.     

The disposition and metabolic fate of 14C-cibenzoline in man

The disposition and metabolic fate of cibenzoline (CBZ) following single oral 153-mg doses of 14C-CBZ succinate were studied in five healthy adult males. The mean maximum plasma radioactivity of 386 ng eq/ml occurred at 2.4 hr after administration. The mean half-life, determined from the 14C plasma concentration and urinary excretion rate data, was 13.1 and 14.8 hr, respectively. The mean maximum CBZ concentration was 196 ng/ml at 1.2 hr post-dose. The mean half-life, determined from the plasma concentration and urinary excretion rate data, was 7.2 and 7.3 hr, respectively. The mean total clearance of radioactivity and CBZ was 300 ml/min and 1224 ml/min, respectively, due to elimination via both renal and nonrenal pathways. The only unconjugated metabolite in the plasma was 4,5-dehydrocibenzoline which, together with other unidentified metabolites, is presumed responsible for the longer observed half-life for total radioactivity. Approximately 75% of the dose was recovered in the urine in the first 24 hr after dosing, 80% of which was present at CBZ and known metabolites. After 6 days, a mean of 85.7% of the dose was excreted in urine and 13.2% in feces. The predominant excreted compound was CBZ (55.7% of the dose) in the 0-72 hr urine. Although several metabolites were identified in urine samples, none were found in substantial amounts relative to the parent drug. Two of these substances showed slight antiarrhythmic activity, whereas the 4,5-dehydro metabolite, representing approximately 4% of radioactivity in urine, was inactive.     

Pharmacokinetics of 2-ethyl-1,3-hexanediol. II. Nonsystemic disposition following single percutaneous or peroral doses in Fischer 344 rats.

The pharmacokinetics of [1,3-14C]-2-ethyl-1,3-hexanediol (EHD) were investigated following single percutaneous doses of 150 mg/kg, applied to male and female Fischer 344 rats, or single peroral doses of 1.5 or 150 mg EHD/kg given by gavage to male Fischer 344 rats. EHD-derived radioactivity was slowly absorbed through skin and relatively rapidly excreted through the urine in a first-order manner over 48 hr postdosing. Skin penetration of 14C was sufficiently slow that the terminal rate constant for the plasma concentration data had to be derived from the absorption phase of this curve, based on the terminal rate constant for a comparable intravenous dose plasma curve [Frantz et al.: Drug Metab. Dispos. 19, 881 (1991)]. Plasma data from perorally doses rats exhibited dose-linearity over a 1.5-150 mg/kg range, with plasma 14C concentration vs. time plots for oral doses of EHD resembling the iv time-course data. This resulted from a very rapid absorption phase (5.5 min t1/2), with plasma 14C levels for both dose levels decreasing in a biexponential manner. The major route of excretion after peroral doses was in urine, making this mode of excretion consistent for both routes of administration evaluated in this study and including the doses given in previous iv work. Kinetic analysis confirmed that this route of excretion was first-order. HPLC analysis of urine from both routes demonstrated that EHD was metabolized and excreted as at least two major, water-soluble urinary metabolites; these metabolites were not identified in this investigation. No unmetabolized EHD was detected in urine, indicating that EHD may be completely metabolized in the rat. Overall, EHD was absorbed, distributed, metabolized, and eliminated from the Fischer rat in a first-order manner following either cutaneous or peroral doses. The results of this study indicate that the kinetic patterns observed experimentally will be dose-proportional for doses administered in the range of 1.5-150 mg/kg.     

Disposition and metabolism of [2-14C]epichlorohydrin after oral administration to rats

A comprehensive disposition and metabolism study of epichlorohydrin (ECH) has not been previously reported. In this study, male Fischer 344 rats were dosed (6 mg/kg) orally with [2-14C]ECH (98% radiochemically pure) as an aqueous solution and killed after 3 days. Approximately 38% of the radioactive dose was exhaled as CO2, 50% was excreted as metabolites in the urine, and 3% was present in the feces. Radioactivity in tissues accounted for the remainder of the administered dose. When expressed per gram of tissue, radioactivity was highest in liver, kidney, and forestomach. The half-life of initial elimination of radioactivity in both the urine and exhaled air was about 2 hr, indicating that ECH was rapidly absorbed and metabolized. The major metabolites in the urine were identified as N-acetyl-S-(3-chloro-2-hydroxypropyl)-L-cysteine and alpha-chlorohydrin, about 36 and 4% of the administered dose, respectively. Finding these metabolites, which have not been previously reported, is consistent with the initial metabolic reactions being conjugation of the epoxide with glutathione and hydration of the epoxide.     

Absorption, distribution, metabolism and excretion of intravenously and dermally administered triethanolamine in mice.

Triethanolamine (TEA) is an amino alcohol having widespread applications in consumer goods and as an industrial chemical. A number of relatively high-dose dermal toxicity studies have been conducted in rats and mice reflecting the principal route of human exposure to TEA. The absorption, distribution, metabolism and excretion (ADME) of (14)C-TEA derived radioactivity were determined in male C3H/HeJ mice following dermal application of 2000 mg/kg (neat) or, to characterize blood kinetics, intravenous (iv) injection of 1 mg/kg (14)C-TEA. Balance and excretion data were also collected in mice utilizing several dermal dosing scenarios (1000 mg/kg in acetone, 2000 mg/kg neat, 2000 mg/kg in water) and, for comparative purposes, in male Fischer 344 rats dosed dermally with 1000 mg/kg neat (14)C-TEA. Urine, feces, expired CO(2) (iv) and, where appropriate, blood were collected over a 24- or 48-hour period post-dosing. The half-life for dermal absorption of radioactivity was estimated to be 1.3 hours. Intravenously administered radioactivity was eliminated in a biphasic manner with a prominent initial phase (half-life of 0.3 hr) followed by a slower terminal phase (half-life of 10 hr). Radioactivity was excreted primarily via the urine (49-69%) as unmetabolized TEA, regardless of dosage, route or vehicle used. Fecal excretion of radioactivity comprised 16-28% of dose administered. The body burden at sacrifice (sum of liver, kidney, carcass and non-application site skin) ranged from 3 to 6% of the dose. It was concluded that TEA is absorbed extensively following dermal application to mice at dosages relevant to toxicity testing and that acetone or water vehicles do not appear to significantly alter total uptake. Significantly, the blood kinetics and ADME of TEA in mice and/or rats differs from that of a related chemical, diethanolamine, which appears to be more toxic to rodents than TEA.     

In vitro dermal absorption of pesticides: IV. In vivo and in vitro comparison with the organophosphorus insecticide diazinon in rat, guinea pig, pig, human and tissue-cultured skin

In vitro skin absorption tests are currently being developed as an alternative to in vivo animal tests for predicting the degree of occupational exposure to pesticides. In the study reported here, in vitro percutaneous absorption tests were conducted with the ¹⁴C-ring-labelled pesticide, diazinon, dissolved in acetone and applied to the dermatomed skin (0.5 mm) of a number of species at a dose rate of 9.5–16.7 μg/cm². Skin permeation was determined for 48 hr after exposure using an in vitro flow-through system. Skin permeation was calculated from the sum of the percentage recovery of ¹⁴C activity in the receiver solution and the percentage recovery obtained in methanol washes of the skin at 48 hr and in skin digests. Listed in decreasing order, the total percentage in vitro dermal absorptions (mean ± SD) obtained by 48 hr after exposure for the five skin types were: 47 ± 3.4% (rat), 36 ± 0.9% (tissue cultured Testskin), 33 ± 2.8% (hairless guinea pig), 20 ± 3.1% (human) and 15 ± 13.1% (pig). The percentage recoveries in soapy water skin washes at 24 hr, in methanol washes and skin digests at 48 hr and of ¹⁴C-labelled volatiles collected in air traps at 48 hr after exposure are reported. Comparative in vivo studies demonstrated 37 ± 0.8 and 24 ± 5.7% recovery of ¹⁴C in the urine of rats (dose rate, 6 μg/cm²) and hairless guinea pigs (dose rate, 5 μg/cm²), respectively, by 14 days after exposure. Total faecal recovery 14 days after exposure was 18 ± 0.4 and 4 ± 0.9% for rats and guinea pigs, respectively. Analysis of tissue taken at autopsy 14 days after exposure demonstrated a total tissue recovery of 0.6 ± 0.1% [¹⁴C]diazinon in rats and 1 ± 0.2% in hairless guinea pigs. The total recovery in skin removed from the dose site at 14 days after exposure was 0.2 ± 0.02% and 0.1 ± 0.05% in rats and hairless guinea pigs, respectively. Recovery of radioactivity from soapy water skin washes conducted at 24 hr after exposure was 21 ± 3.8% for rats and 2 ± 0.1% for hairless guinea pigs. Recovery in skin patches was 23 ± 5.4% and 73 ± 2.9% in rats and hairless guinea pigs, respectively. The in vitro data for dermal absorption of [¹⁴C]diazinon for rats (47 ± 3.4%) and hairless guinea pigs (33 ± 2.8%) were in good agreement with the data observed for rats (56 ± 1.03%) and hairless guinea pigs (28 ± 6.0%) in vivo. This study supported the use of in vitro skin absorption tests as an alternative to in vivo animal testing.     

In vitro dermal absorption of pesticides: V. In vivo and in vitro comparison of the herbicide 2,4-dichlorophenoxyacetic acid in rat, guinea pig, pig, human and tissue-cultured skin

In vitro dermal absorption tests were conducted with the ¹⁴C-ring-labelled herbicide 2,4-dichlorophenoxyacetic acid (2,4-D), dissolved in acetone and applied to dermatomed skin (0.5 mm) of a number of species at dose rates of 7–8 μg/cm². Skin absorption was determined for 48 hr after exposure using an in vitro flow-through system. Skin absorption was calculated from the sum of the percentage recovery of ¹⁴C activity in the receiver solution and the percentage recovery in the methanol washes of the skin at 48 hr and the skin digest samples. Two receiver solutions, Ringer's saline, and Hanks' HEPES buffered saline with 4% serum albumin were used. Listed in decreasing order, the total percentage in vitro dermal absorptions obtained by 48 hr after exposure for the five skin types were: 47 ± 4.3% [tissue cultured Testskin; Hanks' receiver (HR)], 40 ± 4.5% (rat; HR), 19 ± 1.8% (human; HR), 14 ± 2.3% (hairless guinea pig; HR), 14 ± 8.8% (pig; Ringer's receiver). The percentage recovery of the radiolabel in soapy water skin washes at 24 hr, methanol washes and skin digests at 48 hr, and of ¹⁴C-labelled volatiles collected in air traps at 48 hr after exposure are reported. Comparative in vivo studies were conducted for 14 days after exposure and demonstrated 32 ± 3.9 and 28 ± 7.8% recovery of ¹⁴C in the urine of rats (dose rate, 3 μg/cm²) and guinea pigs (dose rate, 4 μg/cm²), respectively. Total faecal recovery was 2 ± 0.3 and 9 ± 3.5% for rats and guinea pigs, respectively. Analysis of tissue taken at autopsy 14 day after dosing demonstrated a total tissue recovery of ¹⁴C activity of 1 ± 0.1 and 2 ± 0.5% in rats and guinea pigs, respectively. Including the ¹⁴C activity extracted from the skin removed from the dose site at 14 days after exposure, the total recovery of dermally absorbed residues was 49 ± 10.4 and 40 ± 9.9% in rats and guinea pigs, respectively. Recovery of ¹⁴C activity from soapy water skin washes conducted at 24 hr after exposure was 28 ± 8.1 and 43 ± 9.0% for rats and guinea pigs, respectively. Recovery in skin patches was 18% (guinea pigs) and 26% (rats). In summary, the in vitro/in vivo concordance for the rat dermal absorption data was good but the in vitro data for hairless guinea pigs underestimated the in vivo absorption, and therefore for 2,4-D, rat skin may provide a better model of percutaneous absorption.     

Metabolic disposition of C-labelled Brown HT in the rat, mouse and guinea-pig

The absorption, metabolism, tissue distribution and excretion of ¹⁴C-labelled Brown HT has been studied in the rat, mouse and guinea-pig. Following administration of a single oral dose of either 50 or 250 mg Brown HT/kg, substantially all of the dose was excreted in the urine and faeces within 72 hr, with the majority (more than 80%) being accounted for in the faeces. A significant difference in urinary excretion of radioactivity was seen between male and female rats, as well as clear species differences at the two dose levels used. In all species studied, naphthionic acid was the major urinary metabolite, whereas in the faeces naphthionic acid, trace quantities of unchanged dye and at least two unidentified metabolites were found. Pregnant rats eliminated a single oral dose of ¹⁴C-labelled colouring at a rate similar to that in non-pregnant females, but some retention of radioactivity was found in the foetuses. Radioactivity was present in all tissues of male rats 24 hr after an oral dose of 250 mg ¹⁴C-labelled Brown HT/kg, with the highest concentrations in the gastro-intestinal tract, kidney and lymph nodes. Clearance from the gastro-intestinal tract was more rapid than from other tissues, but by day 7, the concentration of radioactivity (less than 0.001% of the dose/g) was similar in all tissues except the kidney and mesenteric lymph nodes. Similar results were obtained with animals pretreated for 21 days with either unlabelled or ¹⁴C-labelled Brown HT (250 mg/kg/day) prior to a radioactive dose. For most tissues examined, the concentration of radioactivity was greater with pretreatment than without. These results suggest that despite the rapid reduction and elimination of the major part of an oral dose of Brown HT, some colouring and/or metabolites accumulate in most tissues of male rats during repeated daily administration, but that only in the kidney and mesenteric lymph nodes is the accumulation tissue-specific. The accumulated radioactivity is cleared rapidly from most tissues on cessation of treatment. No significant absorption of either Brown HT, metabolites or subsidiary dyes was detected using isolated loops of small intestine.     

Allyl isothiocyanate: comparative disposition in rats and mice

Allyl isothiocyanate (AITC), the major component of volatile oil of mustard, was recently reported to induce transitional-cell papillomas in the urinary bladder of male Fischer 344 rats, but not in the bladders of female rats or B6C3F1 mice. The present investigation of comparative disposition in both sexes of each species was designed to detect sex or species differences in disposition which might explain susceptibility to AITC toxicity. AITC was readily cleared from all rat and mouse tissues so that within 24 hr after administration less than 5% of the total dose was retained in tissues. The highest concentration of AITC-derived radioactivity was observed in male rat bladder. Clearance of AITC-derived radioactivity by each species was primarily in urine (70 to 80%) and in exhaled air (13 to 15%) with lesser amounts in feces (3 to 5%). Rats excreted one major and four minor metabolites in urine. The major metabolite from rat urine was identified by NMR spectroscopy to be the mercapturic acid N-acetyl-S-(N-allylthiocarbamoyl)-L-cysteine. Mice excreted in urine the same major metabolite identified in rat urine as well as three other major and two minor metabolites. Sex-related variations were observed in the relative amounts of these metabolites. Both species excreted a single metabolite in feces. Metabolism of AITC by male and female rats was similar, but female rats excreted over twice the urine volume of male rats. Results of the present study indicate that excretion of a more concentrated solution of AITC metabolite(s) in urine may account for the toxic effects of AITC on the bladder of male rats.     

Disposition and metabolism of isoeugenol in the male Fischer 344 rat

The primary objective of these studies was to determine the absorption, distribution, metabolism and excretion of isoeugenol following oral and intravenous administration to male Fischer-344 rats. Following a single oral dose of [¹⁴C]isoeugenol (156 mg/kg, 50 μCi/kg), greater than 85% of the administered dose was excreted in the urine predominantly as sulfate or glucuronide metabolites by 72 h. Approximately 10% was recovered in the feces, and less than 0.1% was recovered as CO₂ or expired organics. No parent isoeugenol was detected in the blood at any of the time points analyzed. Following iv administration (15.6 mg/kg, 100 μCi/kg), isoeugenol disappeared rapidly from the blood. The t_1/2 was 12 min and the Cl_s was 1.9 l/min/kg. Excretion characteristics were similar to those of oral administration. The total amount of radioactivity remaining in selected tissues by 72 h was less than 0.25% of the dose following either oral or intravenous administration. Results of these studies show that isoeugenol is rapidly metabolized and is excreted predominantly in the urine as phase II conjugates of the parent compound.     

Mechanism of butylated hydroxytoluene-associated modification of diethylnitrosamine-induced squamous stomach carcinoma

The metabolism of butylated hydroxytoluene (BHT) and the effect of BHT on the metabolism of diethylnitrosamine (DEN) was studied in male and female BALB/c mice to further understanding of the selective protection of BHT on the incidence of DEN-induced squamous-stomach carcinomas in female (but not in male) mice. Following intragastric administration of [¹⁴C]BHT, the antioxidant was covalently bound to tissue macromolecules. The relative distribution of this bound BHT varied with time; 8 hr after [¹⁴C]BHT administration, most of the covalently bound BHT was associated with the protein components; at 96 hr the nucleic acid components bound more BHT than did the protein components. Animals pretreated with BHT and given [¹⁴C]DEN intragastrically had lower blood levels of radioactivity and eliminated a larger percentage of DEN and/or its metabolites in the urine and as carbon dioxide than animals given [¹⁴C]DEN alone. The binding of DEN and/or its metabolites to cellular macromolecules of the squamous stomach of female animals was decreased following pretreatment with BHT. However, the BHT-associated decrease in DEN binding was also observed in the squamous stomach of male animals and in the liver of both sexes, although the tumour incidence in these target organs for DEN carcinogenesis is not modified by BHT. These results suggest that the BHT-associated decrease in the binding of DEN to DNA is of a generalized rather than a selective nature, and may be insufficient to account for the protective effect of BHT. Two parameters that were found to parallel the susceptibility of DEN target tissues to the anticarcinogenic effects of BHT were the relative degree of inhibition of DEN bound to RNA species and the relative amount of BHT bound to DNA. Thus the anticarcinogenic properties of BHT may be more complex than an induction of enzymes that detoxify the carcinogen and/or an inhibition of enzymes that activate the carcinogen with a resulting decrease in the quantity of carcinogen available for electrophilic reactions.     

2,2,4-Trimethylpentane-induced nephrotoxicity. I. Metabolic disposition of TMP in male and female Fischer 344 rats

2,2,4-Trimethylpentane (TMP), a component of unleaded gasoline, causes nephrotoxicity in male, but not in female, rats. In the present study, male and female Fischer 344 rats were treated with a single oral dose of [14C]TMP (4.4 mmol/kg; 2 microCi/mmol). Radiolabeled material in kidney, liver, and plasma was determined at 4, 8, 12, 24, and 48 hr after dosing. Maximum concentration of TMP-derived radioactivity in kidney, liver, and plasma of male rats was found after 12 hr (1252, 1000, and 403 nmol eq/g, respectively), whereas those measured in females were found after 8 hr (577, 1163, and 317 nmol eq/g, respectively). A selective retention of the TMP-derived radiolabel in the kidneys of male rats was noted when peak tissue concentration was expressed as a percentage of administered dose. Kidney concentrations of TMP-derived radiolabel increased in a nonlinear, but dose-dependent, manner; the kidney to plasma ratio was greater at low doses than at higher doses. Increased retention of radiolabel material in the kidney was associated with a significant increase in renal concentration of the male-rat-specific protein, alpha 2u-globulin, 24 and 48 hr after TMP administration. Total radioactivity collected in urine 48 hr after TMP administration was similar in males and females (32 and 31% of dose). Identification and quantitation of the urinary metabolites of TMP showed that both male and female rats metabolize TMP via the same pathway and at a similar rate. Female rats, however, excreted more conjugates of 2,4,4-trimethyl-2-pentanol in urine than males. 2,4,4-Trimethyl-2-pentanol was the major metabolite present in the male rat kidney, but was absent in the female rat kidney. The renal retention of 2,4,4-trimethyl-2-pentanol appears to account for the delayed clearance observed in the disposition of [14C]TMP-derived radiolabel. Based on the concomitant accumulations in renal alpha 2u-globulin concentration and renal 2,4,4-trimethyl-2-pentanol concentration, an association is speculated between these two components. The male-rat-specific accumulation of 2,4,4-trimethyl-2-pentanol may therefore reflect the accumulation of a "metabolite-alpha 2u-globulin" complex. This may be relevant to the male-rat-specific nephrotoxicity produced by TMP.     

Tissue disposition and excretion of 14C-labelled aflatoxin B1 after oral administration in channel catfish.

The pharmacokinetics, tissue distribution and excretion of ¹⁴C-labelled aflatoxin B₁ (AFB₁) were examined after oral administration (250 μg/kg body weight) in channel catfish (Ictalurus punctatus). Plasma concentrations of parent AFB₁ were best described by a one-compartment pharmacokinetic model, in which peak plasma concentration (503 ppb) occurred at 4.1 hr after dosing. The absorption and elimination half-lives were 1.5 and 3.7 hr, respectively. AFB₁ was highly bound (95%) to plasma proteins. Concentrations of ¹⁴C (in AFB₁ equivalents) measured in the tissues were highest at 4 hr, ranging from 596 ppb in the plasma to 40 ppb in the muscle. AFB₁ residues were rapidly depleted; at 24 hr the concentrations in the plasma and muscle were 32 and <5 ppb, respectively. Concentrations in the bile exceeded 2000 ppb (at 24 hr), whereas the highest concentration in the urine was 51 ppb (4–6-hr collection interval). Renal and biliary excretion accounted for <5% of the administered dose, indicating incomplete absorption. Pharmacokinetic modelling and tissue data demonstrate a very low potential for the accumulation of AFB₁ and its metabolites in the edible flesh of channel catfish through the consumption of AFB₁-contaminated feed.     

Percutaneous penetration of octyl salicylate from representative sunscreen formulations through human skin in vitro

The human skin penetration of [¹⁴C]octyl salicylate from two representative sunscreen vehicles was determined in vitro. ³H-sucrose was incorporated into all formulations and provided a marker for membrane integrity. When applied as a finite dose in an oil-in-water emulsion vehicle containing 5% (w/w) octyl salicylate, the average total absorption of ¹⁴C over 48 hr was 0.65 ± 0.16% of the applied dose (representing a total amount permeated of 1.58 ± 0.36 μg/cm²). When applied as an infinite dose in the oil-in-water emulsion vehicle the average total absorption of ¹⁴C over 48 hr was 0.47 ± 0.22% of the applied dose (representing a total amount permeated of 27.54 ± 13.91 μg/cm²). When applied as a finite dose in a representative hydroalcoholic formulation containing 5% (w/w) octyl salicylate, the average total absorption of ¹⁴C over 48 hr was 0.59 ± 0.09% of the applied dose (representing a total amount permeated of 1.58 ± 0.25 μg/cm²). When applied as an infinite dose in the hydroalcoholic formulation the average total absorption of ¹⁴C over 48 hr was 0.23 ± 0.05% of the applied dose (representing a total amount permeated of 11.28 ± 2.55 μg/cm²). The penetration of [¹⁴C]salicylic acid [applied at a concentration of 2.7% (w/w), in the oil-in-water emulsion] was also determined. When applied as a finite dose the average total absorption of ¹⁴C over 48 hr was 1.14 ± 0.23% of the applied dose (representing a total amount permeated of 1.65 ± 0.39 μg/cm²). These results suggest that the in vitro human skin permeation of octyl salicylate is relatively low. The amounts of octyl salicylate and salicylic acid permeated when applied in similar vehicles were remarkably similar over 48 hr (1.58 μg/cm² and 1.65 μg/cm², respectively). This suggests the possibility that the ¹⁴C label appearing in the receptor fluid may, in both cases, represent salicylic acid. If this is the case, then it is possible that the amount of octyl salicylate permeating through the skin is much less than that suggested by the data obtained here. This supposition is, however, entirely speculative and has yet to be confirmed experimentally.     

Age-related changes in the disposition of benzyl acetate. A model compound for glycine conjugation

The in vivo metabolism and excretion of benzyl acetate (BA), a model compound for glycine conjugation, was examined in male Fischer 344 rats and C57BL/6N mice. Rats aged 3-4, 9, and 25 months received a single oral dose of either 5 or 500 mg/kg 14C-BA, while male mice aged 2, 13, and 25 months received a single oral dose of 10 mg/kg 14C-BA. Urine and feces were collected for 96 hr. Biliary excretion and plasma elimination were also examined in male Fischer rats after iv administration of 5 mg/kg 14C-BA. In both young and old rats and mice, hippuric acid (HA) was the major urinary metabolite after oral dosing of BA. No significant age-related difference was observed in rats in the urinary elimination of BA-derived radioactivity or in the percentage of the total dose excreted as hippuric acid (approximately 95%). Twenty-five-month old rats excreted a significantly higher percentage of the total dose as benzyl mercapturic acid (approximately 2%) than did 3- to 4-month-old rats (approximately 1%) at the 5 mg dose. Benzyl mercapturic acid excretion in 3- to 4-month-old rats was also increased significantly at 500 mg/kg BA vs. 5 mg/kg BA. Fecal excretion of BA-derived radioactivity declined significantly in 25-month-old rats at both the 5 and 500 mg dose. This decrease was reflected by an age-related decline in biliary excretion and higher plasma levels of BA-derived radioactivity. Examination of plasma metabolites revealed a significantly higher level of HA and benzoyl glucuronide in 25-month rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Metabolism of 3-deoxy-4-sulphohexosulose, a reaction product of sulphite in foods, by rat and mouse

The excretion of single intragastric doses of 14C-labelled 3-deoxy-4-sulphohexosulose (DSH) was studied in male CF1 mice and male and female Wistar albino rats. Urine and faeces were collected 6, 12, 24, (36), 48 and 72 hr after administration of 2100 mg [14C]DSH/kg body weight (to mice), 1700 mg/kg (to male rats) and 100 and 500 mg/kg (to male and female rats). After 72 hr, plasma and total carcass levels were determined in some experiments. In mice 29% of the administered radioactivity was excreted in the urine, 50% in the faeces and some 13% in cage washings. In rats, faecal excretion varied between 58.5 and 73%. Urinary excretion varied between 16.5 and 31% and was slightly higher in male than in female rats. No radioactivity was detected in expired air of rats, and carcass levels in rats and mice after 72 hr were less than 0.1% of the dose. TLC analysis of urine extracts revealed only unchanged [14C]DSH. In similar studies, male rats and mice were given 35S-labelled DSH in a dose of 6500 mg/kg or 10,700 mg/kg, respectively. Urinary activity accounted for 19.5% of the dose in rats and 27.5% in mice by 72 hr and no 35S-labelled sulphate was detectable in the urine. Organ analyses at nine intervals from 0.25 to 24 hr after intragastric administration of 1600 and 1800 mg [14C]DSH/kg to male rats and mice, respectively, showed that at all times most of the 14C activity was associated with the gastro-intestinal tract in both species. Maximum tissue levels were 2.16% of the dose in the rat liver 0.5 hr after dosing and 1.57% in the mouse kidney after 0.25 hr. Significant amounts of activity (greater than 0.25% of the dose) occurred transiently also in the pancreas and lungs of both species, in the rat testes and in the mouse bladder. Maximum plasma levels were 0.09% of the dose/ml in rats 0.5 and 1 hr after dosing and 0.34%/ml in mice at 0.25 hr.     

Effects of age and dose on disposition and metabolism of salicylic acid in male Fischer 344 rats

Salicylic acid (SAL)-induced nephrotoxicity has been reported to be greater in older rats. To examine age- and dose-related changes in disposition and metabolism, male Fischer 344 rats aged 3, 12, and 25 months were administered single doses of 14C-SAL at 5,50, and 500 mg/kg po. At 5 mg 14C-SAL/kg, urinary excretion of 14C-SAL derived radioactivity (RA) followed first-order kinetics and was complete by 24 hr in 3- and 25-month-old rats, but not until 48 hr in 12-month-old rats. The percentage of administered 14C-SAL excreted as the oxidative metabolites 2,3- and 2,5-dihydroxybenzoic acid (2,3- and 2,5-diOH), unmetabolized SAL, or salicyl ester glucuronide (SA-AG) was unchanged with age. The percentage excreted as the ether glucuronide (SA-PC) was significantly decreased in 25-month-old rats, while the percentage excreted as the glycine conjugate, salicyluric acid (SUA) was significantly increased in 12- and 25-month-old rats. At 50 mg SAL/kg, urinary elimination shifted toward zero-order kinetics and was not complete until 48 hr in all age groups. The percentage of an administered dose of 14C-SAL found in urine as 2,3- and 2,5-diOH and SA-AG increased significantly in all age groups, while the percentage excreted as SUA decreased significantly. Twelve- and 25-month-old rats excreted a significantly greater percentage of the total dose as 2,3- and 2,5-diOH than 3-month-old rats at this dose. No SA-PG was detected at this dose in any age group. At 500 mg SAL/kg, mortality was observed in both 3- and 25-month-old rats and excretion of SAL-derived RA in urine was incomplete at 48 hr. However, data indicated a further shift in biotransformation toward increased production of oxidative metabolites and a decrease in SUA production. No significant overall differences were observed between 3- and 25-month-old rats in plasma levels of 14C-SAL following iv administration of 5 and 50 mg SAL/kg. However, elimination half-life (t1/2) was significantly increased in 25-month-old rats at 5 mg SAL/kg vs. 3-month-old rats. These results indicate that the age-related increase in acute nephrotoxicity of SAL may result from increased production of oxidative metabolites in older rats at higher doses of SAL.