Comparative metabolism of bis(2-methoxyethyl)ether in isolated rat hepatocytes and in the intact rat: effects of ethanol on in vitro metabolism

The metabolism of the reproductive and developmental toxicant bis(2-methoxyethyl)ether (diglyme) was studied in isolated rat hepatocytes and in the intact rat. Male Sprague-Dawley rats (190-220 g) were used in both studies. Hepatocytes, isolated by a two-step in situ collagenase perfusion of the liver, were cultured as monolayers and incubated with [14C]diglyme at 1, 10, 30, and 50 microM for up to 48 h. For the in vivo study, rats were given single oral doses of [14C]diglyme at 5.1 mmol/kg body wt, and urine was collected for up to 96 h. Radioactive compounds in the culture medium or in the urine were separated by high performance liquid chromatography and quantified with an in-line radioactivity monitor. Metabolites were identified by comparison of their chromatographic retention times and their mass spectra with those of authentic compounds. The principal metabolite from hepatocytes and in the urine was (2-methoxyethoxy)acetic acid (MEAA). This metabolite accounted for approximately 36% of the radioactivity in the 48-h culture medium and about 67% of the administered dose in the 48-h urine. Other prominent metabolites common to both systems included 2-(2-methoxyethoxy)ethanol, methoxyacetic acid (MAA), 2-methoxyethanol, and diglycolic acid. The diglyme metabolite profiles from urine and from hepatocytes were qualitatively similar, demonstrating that, in the rat, hepatocytes serve as a good model system for predicting the urinary metabolites of diglyme. Moreover, MEAA was shown to be the metabolite best suited for use as a short-term biological marker of exposure to diglyme.(ABSTRACT TRUNCATED AT 250 WORDS)     

Biotransformation of irbesartan in man

The metabolism of irbesartan, a highly selective and potent nonpeptide angiotensin II receptor antagonist, has been investigated in humans. An aliquot of pooled urine from healthy subjects given a 50-mg oral dose of [14C]irbesartan was added as a tracer to urine from healthy subjects that received multiple, 900-mg nonradiolabeled doses of irbesartan. Urinary metabolites were isolated, and structures were elucidated by mass spectroscopy, proton NMR, and high-performance liquid chromatography (HPLC) retention times. Irbesartan and the following eight metabolites were identified in human urine: (1) a tetrazole N2-beta-glucuronide conjugate of irbesartan, (2) a monohydroxylated metabolite resulting from omega-1 oxidation of the butyl side chain, (3, 4) two different monohydroxylated metabolites resulting from oxidation of the spirocyclopentane ring, (5) a diol resulting from omega-1 oxidation of the butyl side chain and oxidation of the spirocyclopentane ring, (6) a keto metabolite resulting from further oxidation of the omega-1 monohydroxy metabolite, (7) a keto-alcohol resulting from further oxidation of the omega-1 hydroxyl of the diol, and (8) a carboxylic acid metabolite resulting from oxidation of the terminal methyl group of the butyl side chain. Biotransformation profiles of pooled urine, feces, and plasma samples from healthy male volunteers given doses of [14C]irbesartan were determined by HPLC. The predominant drug-related component in plasma was irbesartan (76-88% of the plasma radioactivity). None of the metabolites exceeded 9% of the plasma radioactivity. Radioactivity in urine accounted for about 20% of the radiolabeled dose. In urine, irbesartan and its glucuronide each accounted for about 5 to 10% of the urinary radioactivity. The predominant metabolite in urine was the omega-1 hydroxylated metabolite, which constituted about 25% of the urinary radioactivity. In feces, irbesartan was the predominant drug-related component (about 30% of the radioactivity), and the primary metabolites were monohydroxylated metabolites and the carboxylic acid metabolite. Irbesartan and these identified metabolites constituted 90% of the recovered urinary and fecal radioactivity from human subjects given oral doses of [14C]irbesartan.     

The disposition of allyl isothiocyanate in the rat and mouse

The urine was the major route of excretion of radioactivity (50-80% of dose) following the oral administration (2.5 and 25 mg/kg body weight) of allyl[14C]isothiocyanate (AITC) to male and female Fischer 344 rats and B6C3F1 mice. Smaller amounts were found in the faeces (6-12%) and expired air (3-7%). The major difference between the two species was the greater retention of radioactivity after 4 days within rats (18-24% of dose) when compared with mice (2-5% of dose). Three radioactive components were found in the urine of mice and two in rats. The three components were inorganic thiocyanate, allylthiocarbamoylmercapturic acid and allylthiocarbamoylcysteine in mice, but no cysteine conjugate was found in rat urine. In the mouse, approximately 80% of the 14C was present in the urine as the thiocyanate ion whereas in the rat some 75% was as the mercapturate. This indicates that in the mouse, hydrolysis of AITC was the major metabolic pathway whereas in the rat glutathione conjugation was the major route. A species difference was seen in the amount of [14C]AITC-derived radioactivity present in the whole blood of rats and mice; measurable levels of radioactivity remained within rat blood for a longer time period (up to 240 hr) when compared with mice (96 hr). Examination of the urinary bladders of male and female rats following oral dosing with [14C]AITC showed a sex difference with greater amounts of [14C]AITC and/or its metabolites within the bladder tissue of male rats. This data is discussed in terms of the known species- and sex-specificity of the urinary bladder tumours, which occurred after long-term administration to male rats, but not to female rats or mice of either sex, in a carcinogenicity study conducted by the National Toxicology Program in the USA.     

Biotransformation, excretion, and nephrotoxicity of the hexachlorobutadiene metabolite (E)-N-acetyl-S-(1,2,3,4, 4-pentachlorobutadienyl)-L-cysteine sulfoxide

The disposition of ethyl 4-(3,4-dimethoxyphenyl)-6,7-dimethoxy-2-(1,2,4- triazol-1-ylmethyl) quinoline-3-carboxylate (CAS 158146-85-1, TAK-603) after single oral dosing of 14C-labeled TAK-603 ([14C]TAK-603) at 10 mg/kg to rats and dogs was studied. In rats, the concentration of unchanged drug in plasma reached a peak (Cmax, 0.31 microgram/ml) 2 h (Tmax) after dosing of TAK-603 and declined biphasically with apparent half-lives (t 1/2 alpha, t 1/2 beta) of 1.5 and 3.6 h. In dogs, Tmax, Cmax, T 1/2 alpha, and t 1/2 beta were 1.7 h, 0.36 microgram/ml, 1.2, and 10.8 h, respectively. [14C]TAK-603 dosed orally was absorbed quantitatively in rats, while the extent of absorption in dogs was 54%. The bioavailability of TAK-603 was 53% and 42% in rats and dogs, respectively. In rats, 14C was distributed widely in various tissues, with relatively high concentrations in the liver, adrenal gland, and gut. The elimination of 14C from the thyroid was slower than that from other tissues. Unchanged TAK-603 and its pharmacologically active metabolite, M-I, which has the same potency as TAK-603, were distributed in articular soft tissues and synovial fluids, as target tissues, in rats and dogs, respectively. After oral administration of [14C]TAK-603, most of the 14C dosed was excreted within 48 h in rats and within 96 h in dogs. In both animals, a greater amount of the 14C dosed was excreted in feces than in urine. In biliary duct cannulated rats given [14C]TAK-603 intraduodenally, 69% of the dose was excreted in bile, and biliary 14C in part underwent enterohepatic circulation.     

drp, a novel protein expressed at high cell density but not during growth arrest

In this study the disposition of 1,2-[14C]dibromoethane (1, 2-[14C]DBE) was investigated in male Wistar rats. 1,2-DBE is a cytotoxic and carcinogenic compound that has been used as an additive in leaded gasoline and as a fumigant. 1,2-[14C]DBE was administered orally or iv. Radioactivity was recovered (mostly within 48 hr after administration) in urine (75-82% of the dose), feces (3.2-4% of the dose), and expired air (0.53-7.2% of the dose). One hundred-sixty-eight hours after administration of 1,2-[14C]DBE, most of the radioactivity in tissues was found in the liver, lungs, and kidneys (<1% of the dose) and the red blood cells (0.3% of the dose). Identified urinary metabolites were S-(2-hydroxyethyl)mercapturic acid, thiodiacetic acid, and thiodiacetic acid sulfoxide, together accounting for, on average, 78% of the total amount of radioactivity in urine. In addition to S-(2-hydroxyethyl)mercapturic acid, thiodiacetic acid, and thiodiacetic acid sulfoxide, several compounds were anticipated as potential urinary metabolites of 1,2-DBE, i.e. S-(carboxymethyl)mercapturic acid, S-(2-hydroxyethyl)thioacetic acid, S-(2-hydroxyethyl)thiopyruvic acid, S-(carboxymethyl)thiopyruvic acid, S-(2-hydroxyethyl)thiolactic acid, and S-(carboxymethyl)thiolactic acid. All of the postulated urinary metabolites were synthesized and searched for in urine samples. None of these metabolites could be detected in urine, however. The data obtained in the present study might be useful for risk assessment and biomonitoring studies of 1,2-DBE and will also be used to further validate a physiologically based pharmacokinetic model for 1, 2-DBE in rats and humans that was recently developed.     

Metabolism of tetramethrin isomers in rat: II. Identification and quantitation of metabolites

1. To examine the metabolic fate of (1RS, trans)- or (1RS, cis)-tetramethrin [3, 4, 5, 6-tetrahydrophthalimidomethyl (1RS, trans)- or (1RS, cis)-chrysanthemate], rat was administered a single oral dose of trans- or cis-[alcohol-14C]tetramethrin at dose levels of 2 or 250 mg/kg. 2. The radiocarbon was almost completely eliminated from rat within 7 days after administration in all groups. 14C-recoveries (expressed as percentages relative to the dosed 14C) in faeces and urine were 38-58 and 42-58% respectively in rat administrated trans-[alcohol-14C]tetramethrin, and in faeces and urine were 66-91 and 9-31% respectively in rat administered cis-[alcohol-14C]tetramethrin. 3. Fourteen metabolites found in excreta were purified by using several chromatographic techniques and identified by spectroanalyses (nmr and MS). Five sulphonate derivatives and three dicarboxylic acid derivatives were found. 4. The main metabolites were sulphonate derivatives in the faeces, and in the urine, alcohols, dicarboxylic acid and reduced metabolites derived from the 3,4,5,6-tetrahydrophthalimide moiety.     

Metabolism of [1-(14)C] and [2-(14)C] leucine in cultured skin fibroblasts from patients with isovaleric acidemia. Characterization of metabolic defects

Leucine metabolism in cultured skin fibroblasts from patients with isovaleric acidemia was compared with that in normal fibroblasts and in cells from patients with maple syrup urine disease using [1-(14)C] and [2-(14)C] leucine as substrates. Inhibitory effects of methylenecyclopropylacetic acid on leucine metabolism in normal cells were also investigated. Production of 14CO2 from [2-(14)C] leucine was very reduced (96-99%) in both types of mutant cells. Radioactive isovaleric acid accumulated in assay media with isovaleric acidemia cells but not in those with maple syrup urine disease cells. Unexpectedly, 14CO2 production from [1-(14)C] leucine was partially depressed (80%) in isovaleric acidemia cells whereas in maple syrup urine disease cells it was strongly depressed (99%) as expected. These two mutant cells were clearly distinguished by detection of 14C-isovaleric acid accumulation after incubation with [2-(14)C] leucine. A pattern of inhibition of leucine oxidation similar to that seen in isovaleric acidemia cells was induced in normal cells by the addition of 0.7 mM methylenecyclopropylacetic acid to the assay medium. The partial inhibition of [1-(14)C] leucine oxidation seen in isovaleric acidemia cells and also in normal cells in the presence of the inhibitor appears to be, at least in part, due to an accumulation of isovalerate in the cells. Isovaleric acid (5-10) mM) inhibited [1-(14)C] leucine oxidation 32-68% when added to the assay medium with normal cells. Addition of flavin adenine dinucleoside to culture medium or assay medium or both did not restore oxidation of either leucine substrate in isovaleric acidemia cells.     

Metabolism of linoleamides. II. Absorption, distribution, excretion and metabolism of N-[alpha-phenyl-beta-(p-tolyl)ethyl]linoleamide

1. Absorption, distribution, excretion and metabolism of (-)N-[alpha-phenyl-beta-(p-tolyl)ethyl][14C]linoleamide (14C-PTLA) were studied in rats and dogs. Faecal excretion of PTLA was studied in dogs and men by g.l.c. 2. 14C-PTLA (10 mg/kg) given orally to rats resulted in urinary and faecal excretion of radioactivity of 2 and 93%, respectively, by male rats and 8 and 87% by female rats in 48 h. Faecal excretion of PTLA in men was similar to that in rats. 3. Distribution of radioactivity in rats and dogs after oral administration of 14C-PTLA showed that a major part of the dose was not absorbed. 4. N-[alpha-phenyl-beta-(p-tolyl)ethyl]glutaric acid monoamide were detected in the urine of rats dosed orally with 14C-PTLA.     

Evaluation of the carcinogenic effect of ceramic fibers in experiments on rats and mice

The metabolism of toborinone, (+/-)-6-[3-(3,4-dimethoxybenzylamino)-2-hydroxypropoxy]-2(1H)-quin - olinone, a novel inotropic agent, was studied in rats and dogs after intravenous administration. Chemical structures of the 13 metabolites were characterized by direct-probe FAB/MS and field desorption/MS, LC/FAB/MS, and various NMR measurements. After intravenous dosing of 10 mg/kg [14C]toborinone, fecal and urinary recoveries of the 14C dose were approximately 70% and 26-30%, respectively, in both rats and dogs. The predominant component of radioactivity was the unchanged toborinone in every biological specimen in rats and dogs. Although unchanged toborinone was predominantly observed, toborinone underwent extensive conjugations with glucuronic acid, sulfate, and glutathione, either directly or following phase I reaction. Metabolites resulting from oxidative N-C cleavage were minor both in number and in quantity in every biological specimen in rats and dogs. In rats, toborinone underwent O-demethylation to form M-7 and successive phase it reaction to yield the glucuronide M-1 and the sulfoconjugate M-2, and deconjugation to yield M-7, which was a primary metabolite accounted for 35.67% of the radioactivity excreted in the feces by 48 hr. Conjugates M-1 and M-2 were the major metabolites in rat plasma. In dogs, toborinone was metabolized via mercapturic acid pathway to yield the primary metabolites, cysteine conjugates M-10 and M-11 that accounted for 19.10% and 6.70% of the radioactivity excreted in the feces by 48 hr and that were detected species specifically in dogs. The glutathione conjugate M-13, which was isolated from in vitro incubations using dog liver, led us to consider a possible mercapturic acid pathway from the parent compound to M-10. Metabolites in dog plasma and those in urine in both rats and dogs were minor in quantity. The metabolic pathways of toborinone in rats and dogs are proposed herein.     

Synthesis and excretion profile of 1,4-[14C]phenylenebis(methylene)selenocyanate in the rat

1,4-Phenylenebis(methylene)selenocyanate (p-XSC) inhibits chemically induced tumors in several laboratory animal models. To understand its mode of action, we synthesized p-[14C]XSC, examined its excretion pattern in female CD rats and also the nature of its metabolites. p-[14C]XSC was synthesized from alpha,alpha-dibromo-p-[ring-14C]xylene in 80% yield. The excretion profile of p-[14C]XSC (15.8 mg/kg body wt, 200 microCi/rat, oral administration, in 1 ml corn oil) in vivo was monitored by measuring radioactivity and selenium content. On the basis of radioactivity, approximately 20% of the dose was excreted in the urine and 68% in the feces over 3 days. The cumulative percentages of the dose excreted over 7 days were 24% in urine and 75% in feces, similar to excretion rates of selenium. According to selenium measurement, <1% of the dose was detected in exhaled air; radioactivity was not detected. Only 15% of the dose was extractable from the feces with EtOAc and was identified as tetraselenocyclophane (TSC). Most of the radioactivity remained tightly bound to the feces. Approximately 10% of this bound material converted to TSC on reduction with NaBH4. Organic soluble metabolites in urine did not exceed 2% of the dose; sulfate (9 % of urinary metabolites) and glucuronic acid (19.5% of urinary metabolites) conjugates were observed but their structural identification is still underway. Co-chromatography with a synthetic standard led to the detection of terephthalic acid (1,4-benzenedicarboxylic acid) as a minor metabolite. The major urinary conjugates contained selenium. Despite the low levels of selenium in the exhaled air, the reductive metabolism of p-XSC to H2Se cannot be ruled out. Identification of TSC in vivo indicates that a selenol may be a key intermediate responsible for the chemopreventive action of p-XSC.     

Evaluation of liver function by co-administration methodology using 13C-labelled benzoic acid and hippuric acid coupled with nuclear magnetic resonance spectroscopy

The amount of hippuric acid synthesized and excreted in the urine after benzoic acid loading (hippuric acid test) is a useful index of liver function. However, the hippuric acid test gives erroneous results in the event of failure of renal excretory function. A new stable isotope co-administration methodology using nuclear magnetic resonance (NMR) spectroscopy has been developed to overcome this defect. [7-(13)C]Benzoic acid and [glycine carbonyl-13C]hippuric acid ([gly-13C]hippuric acid), each 0.4-0.6 mmol kg(-1) were simultaneously administered intravenously as probes to normal or liver-injured rats and the urine was analysed by 100 MHz 13C NMR spectroscopy. Consequently, urinary excretion of [7-(13)C]hippuric acid formed from [7-(13)C]benzoic acid and [gly-13C]hippuric acid was successfully traced with very simple and convenient procedures. The urinary excretion of [7-(13)C]hippuric acid indicated the combined functions of hippuric acid synthesis and renal excretion, whereas that of [gly-13C]hippuric acid was indicative of renal excretion of hippuric acid only. The heights of resonances for C7 of [7-(13)C]hippuric acid and the glycine carbonyl carbon of [gly-13C]hippuric acid were used to calculate the concentrations of labelled hippuric acids. [7-(13)C]Hippuric acid was excreted more slowly than [gly-13C]hippuric acid by both normal and liver-injured rats. The liver-injured rats excreted the labelled hippuric acids more slowly than the normal rats. The kinetic parameters were computed for the individual rats on the basis of Michaelis-Menten elimination for benzoic acid and first-order elimination for hippuric acid. The maximum rates of metabolism (Vmax) (4.8-5.8 micromol min(-1) kg(-1)) and the renal elimination rate constants of hippuric acid (Kre) (0.010-0.021 min(-1)) in the liver-injured rats were lower than those (Vmax 6.7-11.8 micromol min(-1) kg(-1); Kre 0.026-0.045 min(-1)) in the normal rats. These results have demonstrated that liver function can be evaluated from the Vmax value even though the renal function of hippuric acid excretion (Kre) is impaired. Thus the co-administration methodology is feasible and can remove the defect of the previous hippuric acid test. These results could form the basis for a more convenient and reliable hippuric acid test in man.     

Metabolism of linoleamides. I. Absorption, excretion and metabolism of N-(alpha-methylbenzyl)linoleamide in rat and man

1. In urine of rats dosed with N-(alpha-methylbenzyl)linoleamide (MBLA), three dicarboxylic acid monoamides, N-(alpha-methylbenzyl)succinic acid monoamide, N-(alpha-methylbenzyl)glutaric acid monoamide and N-(alpha-methylbenzyl)adipic acid monoamide, were identified. Conjugated alpha-methylbenzylamine, hippuric acid and conjugates of the dicarboxylic acid monoamides were also found in the urine. N-(alpha-Methylbenzyl)succinic acid monoamide was the main metabolite in rats. 2. Biliary excretion of radioactivity was studied in rats, cannulated for collection of bile and duodenal infusion, after oral administration of N-(alpha-methylbenzyl)[1-14C]linoleamide. With constant duodenal infusion of bile, about 7% of the dose was excreted in the bile, while excretion of radioactivity was negligible without bile infusion. 3. The g.l.c. analysis of human urine after oral administration of MBLA revealed that two dicarboxylic acid monoamides were present and N-(alpha-methylbenzyl)succinic acid monoamide was the main metabolite. 4. MBLA was excreted unchanged in the faeces of men who received MBLA to the extent of about 53% dose in 3 days. 5. MBLA was not detected (less than 1 mug/ml) in the serum of a volunteer who had been taking an oral daily dose of 1500 mg of MBLA for 3 months.     

Synthesis of deoxyglucose-1-phosphate, deoxyglucose-1,6-bisphosphate, and other metabolites of 2-deoxy-D-[14C]glucose in rat brain in vivo: influence of time and tissue glucose level

When the kinetics of interconversion of deoxy[14C]glucose ([14C]DG) and [14C]DG-6-phosphate ([14C]DG-6-P) in brain in vivo are estimated by direct chemical measurement of precursor and products in acid extracts of brain, the predicted rate of product formation exceeds the experimentally measured rate. This discrepancy is due, in part, to the fact that acid extraction regenerates [14C]DG from unidentified labeled metabolites in vitro. In the present study, we have attempted to identify the 14C-labeled compounds in ethanol extracts of brains of rats given [14C]DG. Six 14C-labeled metabolites, in addition to [14C]DG-6-P, were detected and separated. The major acid-labile derivatives, DG-1-phosphate (DG-1-P) and DG-1,6-bisphosphate (DG-1,6-P2), comprised approximately 5 and approximately 10-15%, respectively, of the total 14C in the brain 45 min after a pulse or square-wave infusion of [14C]DG, and their levels were influenced by tissue glucose concentration. Both of these acid-labile compounds could be synthesized from DG-6-P by phosphoglucomutase in vitro. DG-6-P, DG-1-P, DG-1,6-P2, and ethanol-insoluble compounds were rapidly labeled after a pulse of [14C]DG, whereas there was a 10-30-min lag before there was significant labeling of minor labeled derivatives. During the time when there was net loss of [14C]DG-6-P from the brain (i.e., between 60 and 180 min after the pulse), there was also further metabolism of [14C]DG-6-P into other ethanol-soluble and ethanol-insoluble 14C-labeled compounds. These results demonstrate that DG is more extensively metabolized in rat brain than commonly recognized and that hydrolysis of [14C]DG-1-P can explain the overestimation of the [14C]DG content and underestimation of the metabolite pools of acid extracts of brain. Further metabolism of DG does not interfere with the autoradiographic DG method.     

Urinary metabolites of p-hydroxyamphetamine in man, rat and guinea-pig

1. The metabolism of (+/-)-p-hydroxy[14C]amphetamine has been studied in the rat, guinea-pig and man. 2. Most of the administered 14C was excreted in the urine within the first 24 h (64-92%), and was present mainly as free and conjugated p-hydroxy[14C]-amphetamine. In the female rat and female guinea-pig the conjugate was a glucuronide, but in man, who received a much smaller dose, the conjugate was a sulphate ester. A sex difference in conjugation was found in the rat, the female partly conjugating the drug but not the male. 3. Small quantities (1-6% of dose) of p-hydroxynorephedrine, a putative false neurotransmitter, were found in the urine of the three species. 4. Some oxidative degradation of the side chain of p-hydroxyamphetamine occurred in rat and guinea-pig since small amounts of p-hydroxybenzoic acid (1-3%) were detected in the urine.     

Hospital administration--the new order; the medical administrator''s point of view

Urine from rats and sheep given single doses of [14C]propachlor contained 14C metabolites in which the chlorine of propachlor was replaced by a methylsulfonyl group. Methylsulfonyl-containing metabolites were also isolated from the urine of rats given an intraperitoneal dose of the cysteine conjugate of [14C]propachlor; this indicated that the methylsulfonyl-containing metabolites resulted from metabolic reactions subsequent to the mercapturic acid pathway.     

Biotransformation of 17-alkylsteroids in the equine: gas chromatographic-mass spectral identification of ten intermediate metabolites of methyltestosterone

Parenterally administered domoic acid, a structural analog of the excitatory amino acids glutamic acid and kainic acid, has specific effects on brain histology in rats, as measured using different anatomic markers. Domoic acid-induced convulsions affects limbic structures such as hippocampus and entorhinal cortex, and different anatomic markers can detect these neurotoxic effects to varying degrees. Here we report effects of domoic acid administration on quantitative indicators of brain metabolism and gliosis. Domoic acid, 2.25 mg/kg i.p., caused stereotyped behavior and convulsions in approximately 60% of rats which received it. Six to eight days after domoic acid or vehicle administration, the animals were processed to measure regional brain incorporation of the long-chain fatty acids [1-(14)C]arachidonic acid ([14C]AA) and [9,10-(3)H]palmitic acid ([3H]PA), or regional cerebral glucose utilization (rCMRglc) using 2-[1-(14)C]deoxy-D-glucose, by quantitative autoradiography. Others rats were processed to measure brain glial fibrillary acidic protein (GFAP) by enzyme-linked immunosorbent assay. Domoic acid increased GFAP in the anterior portion of cerebral cortex, the caudate putamen and thalamus compared with vehicle. However, in rats that convulsed after domoic acid GFAP was significantly increased throughout the cerebral cortex, as well as in the hippocampus, septum, caudate putamen, and thalamus. Domoic acid, in the absence of convulsions, decreased relative [14C]AA incorporation in the claustrum and pyramidal cell layer of the hippocampus compared with vehicle-injected controls. In the presence of convulsions, relative [14C]AA incorporation was decreased in hippocampus regions CA1 and CA2. Uptake of [3H]PA into brain was unaffected. Relative rCMRglc decreased in entorhinal cortex following domoic acid administration with or without convulsions. These results suggest that acute domoic acid exposure affects discrete brain circuits by inducing convulsions, and that domoic acid-induced convulsions cause chronic effects on brain function that are reflected in altered fatty acid metabolism and gliosis.     

Urinary metabolites of 3,3-dimethyl-1-phenyltriazene

Urinary metabolites excreted after a subcutaneous injection of 3,3-dimethyl-1[14C] phenyltriazene (DM[1-14C]PT) to rats accounted for 82% of the applied radioactivity. We have isolated aniline (1-2%), 2-hydroxyaniline (5-7%), 3-hydroxyaniline (about 1%) and 4-hydroxyaniline (31-37%) from ethyl acetate extracts of acid-hydrolysed urine, UV spectrometric determination of 4-hydroxyaniline, using the indophenol reaction, showed that the most abundant metabolite accounted for 56 to 61% of the applied dose. We have also demonstrated the excretion of metabolites containing the intact triazene structure (0.9-1.1%) by cold acid cleavage of these compounds, followed by coupling of the released arenediazonium cations with N-ethyl-1-naphthylamine (EN). The coloured derivatives of these metabolites, 4-benzeneazo-N-ethyl-1-naphthylamine (BAEN) (0.6-0.7%), 4-(2-hydroxybenzeneazo)-N-ethyl-1-napthylamine (2-HO-BAEN) (0.02%) and 4-(4-hydroxybenzeneazo)-N-ethyl-1-naphthylamine (4-HO-BAEN) (0.3-0.4%) were isolated. The identification of BAEN as the principal azo derivative of the excreted triazene metabolites is in full agreement with the proposed in vivo activation of 3,3-dimethyl-1-phenyltriazene (DMPT) to a carcinogenic methylating agent. The hydroxylation of the methyl group at N-3 yields the corresponding aminol, some of which is covalently bonded to a water-soluble compound.     

Excretion of DNA adducts of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline, PhIP-dG, PhIP-DNA and DiMeIQx-DNA from the rat

The heterocyclic aromatic amines, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx) are formed during frying of meat. PhIP and 4,8-DiMeIQx have, after metabolic activation, been shown to form adducts with DNA at the C8 of guanine both in vitro and in vivo. In order to investigate possible urinary biomarkers for estimation of the genotoxic dose of PhIP and 4,8-DiMeIQx, [3H]PhIP-dG, [3H]PhIP-DNA and [14C]4,8-DiMeIQx-DNA were injected i.p. to rats and the excretion of radioactivity in urine and faeces were measured. For all three [3H]PhIP-dG, [3H]PhIP-DNA and [14C]4,8-DiMeIQx-DNA 15-20% of the dose were excreted in the urine and 80-85% of the dose were excreted in the faeces. Urinary excretion showed maximum to 24 h (90%) with a rapid decline, 10% to 48 h and 0% to 72 h. Faecal excretion also showed maximum to 24 h (60%) with a slower decline, 30% to 48 h and 10% to 72 h. HPLC analysis of samples of urine and extracts from faeces, from rats dosed with [3H]PhIP-dG, showed that approximately 90% of the radioactivity co-eluted with PhIP-dG, indicating that PhIP-dG is excreted unmetabolized. HPLC analysis of samples of urine and extracts from faeces, from rats dosed with [3H]PhIP-DNA, showed that approximately 85% of the radioactivity co-eluted with PhIP-dG, indicating that PhIP-DNA adducts is mainly excreted as nucleoside adducts. Approximately 5% of the radioactivity excreted in the urine co-eluted with PhIP-G, indicating loss of deoxyribose. HPLC analysis of samples of urine and extracts from faeces, from rats dosed with [14C]4,8-DiMeIQx-DNA, showed that approximately 90% of the radioactivity co-eluted with 4,8-DiMeIQx-dG, indicating that 4,8-DiMeIQx-DNA adducts is mainly excreted as nucleoside adducts. Man is able to eliminate compounds of a higher mol. wt in the urine than the rat, the percentage of PhIP-dG and 4,8-DiMeIQx eliminated in the urine of man would therefore be expected to be higher than in the rat. Measurement of urinary nucleoside adducts of PhIP and 4,8-DiMeIQx could therefore provide a basis for the development of a biomonitoring strategy for the genotoxic dose of these food derived HAA.     

Integrated physical and transcript map of 5q31.3-qter

The disposition of S-2-[4-(3-methyl-2-thienyl)phenyl]propionic acid (CAS 155680-07-2, S-MTPPA, code: M-5011) was studied after oral administration to rats, dogs and monkeys using the 14C-labeled drug. After oral dosing, S-MTPPA was well absorbed from the gastrointestinal tract, to the extent of 97.7% in rats. The concentration of S-MTPPA in rat plasma reached a peak (Cmax: 13.07 micrograms/ml) at 15 min (tmax) after dosing and declined with a half-life (t1/2) of 2.5 h. The values of the parameters tmax, Cmax and t1/2 for dogs were 30 min, 26.2 micrograms/ml and 7.0 h, and those for monkeys were 15 min, 12.8 micrograms/ml and 3.0 h, respectively. The radioactivity was widely distributed in tissues and almost completely excreted in urine and feces within 48 h after oral administration to rats. The excretion of radioactivity in bile, urine and feces within 48 h after oral administration of 14C-S-MTPPA to bile duct-cannulated rats amounted to 75.0, 18.6 and 1.4% of the dose, respectively. The drug was metabolized mainly by oxidation of the thiophenyl moiety and by glucuronidation of the carboxyl group in rats and monkeys. The major urinary and fecal metabolite in dogs was identified as the taurine conjugate of MTPPA.     

The biodegradation of the surfactant undecyl sulphate

1. The metabolism of the odd-numbered carbon chain surfactant, potassium undecyl [35S]sulphate in the rat was investigated. 2. The major route for elimination of radioactivity was the urine, regardless of the route of administration. 3. The surfactant was extensively degraded in vivo to yield propionic acid 3-[35S]sulphate, the major radioactive component in urine. A second urinary metabolite was identified tentatively as pentanoic acid 5-[35S]sulphate. 4. Whole-body autoradiography revealed the liver as the major site of metabolism. 5. The nature of the metabolic products of undecyl sulphate suggest that it is bio-degraded by initial omega-oxidation followed by beta-oxidation.