Study on the stereoselective excretion of tetrahydropalmatine enantiomers in rats and identification of in vivo metabolites by liquid chromatography‐tandem mass spectrometry

The objective of this work was to study the stereoselectivity in excretion of tetrahydropalmatine (THP) enantiomers by rats and identify the metabolites of racemic THP (rac‐THP) in rat urine. Urine and bile samples were collected at various time intervals after a single oral dose of rac‐THP. The concentrations of THP enantiomers in rat urine and bile were determined using a modification of an achiral–chiral high‐performance liquid chromatographic (HPLC) method that had been previously published. The cumulative urinary excretion over 96 h of (?)‐THP and (+)‐THP was found to be 55.49 ± 36.9 μg and 18.33 ± 9.7 μg, respectively. The cumulative biliary excretion over 24 h of (?)‐THP and (+)‐THP was 19.19 ± 14.6 μg and 12.53 ± 10.4 μg, respectively. The enantiomeric (?/+) concentration ratios of THP changed from 2.80 to 5.15 in urine, and from 1.36 to 1.80 in bile. The mean cumulative amount of (?)‐THP was significantly higher than that of (+)‐THP both in urine and bile samples. However, the enantiomeric (?/+) concentration ratios in rat urine and bile were significantly lower than those ratios in rat plasma. These findings suggested the excretion of THP enantiomers was stereoselective rather than a reflection of chiral pharmacokinetic aspects in plasma and (?)‐THP was preferentially excreted in rat urine and bile. Three O‐demethylation metabolites and the parent drug rac‐THP were detected by liquid chromatography‐tandem mass spectrometry in rat urine. One metabolite was obtained by preparative HPLC and identified as 10‐O‐demethyl‐THP. Chirality, 2010. © 2009 Wiley‐Liss, Inc.     

A comparison of sex steroid hormone excretion and metabolism by psittacine species

The metabolism, excretory rates, and excretory patterns of carbon 14 (¹⁴C) radiolabeled estradiol (E₂) and testosterone (T) were studied in female budgerigars (Melopsittacus undulatus) and orange‐winged Amazon parrots (Amazona amazonica). Radiolabeled E₂ and T were injected intramuscularly into six budgerigars and two orange‐winged Amazon parrots. Serial fecal/urine samples were collected for 168 h post‐radiolabel injection. Peak radiolabeled E₂ excretion was observed at 4 h post‐injection, and by 24 h, 93.3 ± 6.3 and 65.9% (range, 59.1–72.7%) of the injected radiolabel was recovered in the fecal/urine matter of budgerigars and orange‐winged Amazon parrots, respectively. Similarly, peak radiolabeled T excretion was observed at 4 h post‐injection with 92.7± 3.6 and 66.2% (range, 57.5–75.2%) of the injected radiolabel recovered in the fecal/urine matter by 24 h in the budgerigars and orange‐winged Amazon parrots, respectively. High‐performance liquid chromatography (HPLC) analysis of the fecal/urine material revealed that both parrot species excreted >80% of the radiolabel in the form of complex steroid conjugates. Immunoreactive E₂ and T metabolites were detected using estrone (E₁) and C‐21/C‐19 conjugate enzyme immunoassays, respectively. Hydrolysis of the E₂ metabolites and HPLC analysis of the ether extracts revealed that E₂ and E₁ were the major steroid moieties. Hydrolysis of the T metabolites and HPLC analysis of the ether extracts revealed two and three major unconjugated peaks for the budgerigars and the orange‐winged Amazon parrots, respectively. Zoo Biol 18:247–260, 1999. © 1999 Wiley‐Liss, Inc.     

Excretion and metabolic fate of radiolabeled estradiol and testosterone in the cockatiel (Nymphicus hollandicus)

The metabolism and time courses for clearance of radiolabeled estradiol and testosterone were studied in the female cockatiel (Nymphicus hollandicus) using a simple technique of solubilizing dried fecal/urine matter in an aqueous solution. Carbon ¹⁴ radiolabeled estradiol and testosterone were injected intramuscularly and the urine and fecal matter collected for the pursuant 168 hr. The predominant radiolabel peak was found associated with the aqueous residue of the ether extracted aliquot for both hormones. High-performance liquid chromatographic (HPLC) separation of solubilized fecal/urine material collected during the first sampling interval (0–4 hr after injection) demonstrated that the majority of the excreted radiolabel was in the form of conjugated steroid metabolites in both the estradiol and testosterone injected birds. Subsequent hydrolysis of the aqueous residue of the ether extracted aliquots and HPLC characterized the estrogen and testosterone metabolites as being estrone/estradiol and a variety of androgen based moieties, respectively. By 24 hr postinjection, 79.4 and 79.1% of the original radiolabel was recovered in birds injected with estradiol and testosterone, respectively. These findings demonstrate that steroid hormone excretion in the cockatiel is a rapid and efficient process that is 79% complete by 24 hr and that the primary excretion products are conjugated metabolites. This study also supports the concept that fecal/urine collection is a practical and efficient method of monitoring sex steroid excretion and provides additional evidence that simple solubilization of fecal matter is a sufficient and efficient method for processing feces for subsequent metabolite measurements. Zoo Biol 16:505–518, 1997. © 1997 Wiley-Liss, Inc.     

Cortisol metabolism in the domestic cat and implications for non-invasive monitoring of adrenocortical function in endangered felids

Three domestic cats were given i.m. injections of ³H-cortisol to determine the time course and relative proportion of excreted ³H-cortisol metabolites into urine and feces. Most urinary radioactivity was detected in the first sample collected at 3.9 ± 2.5 hr postinjection and accounted for 13.9 ± 2.1% of the total radioactivity recovered. High performance liquid chromatography (HPLC) detected four urinary metabolites, one of which (13.7% urinary radioactivity) eluted with the ³H-cortisol reference tracer and was quantifiable using a commercial cortisol radioimmunoassay (RIA). The majority of cortisol metabolites in feces (85.9 ± 2.1%) was excreted at 22.3 ± 6.2 hr. HPLC analysis detected several fecal metabolites consisting primarily of nonhydolyzable water-soluble forms, none of which eluted with ³H-cortisol or ³H-corticosterone reference tracers. No immunoreactivity was detected in HPLC-separated fecal eluates using the cortisol RIA; however, two of the more polar metabolites were quantifiable using a commerical cortisosterone RIA. The physiological relevance of the immunoreactive fecal metabolites was determined in four domestic cats given an adrenocorticotropin (ACTH) challenge. Increased serum cortisol concentrations were detected within 30 min of ACTH injection, which was maintained for at least 6 hr. A corresponding increase in fecal cortisol metabolite concentrations (ranging from 238% to 826% over individual baseline values) was observed 24–48 hr later. These data indicate that adrenocortical activity can be monitored nonivasively in the cat by measuring cortisol metabolites excreted in feces. This procedure is a potentially valuable tool for endangered felid management to help evaluate responses to physiological and psychological stressors associated with environmental conditions and husbandry practices. (This article is a US Government work and, as such, is in the public domain in the United States of America.) © 1996 Wiley-Liss, Inc.     

Comparison of the urinary excretion time courses of pyrene-1,6-dione,pyrene-1,8-dione and 1-hydroxypyrene in rats intravenously exposed to pyrene

Abstract

The urinary excretion time courses of pyrene-1,6-dione (P16D), pyrene-1,8-dione (P18D) and 1-hydroxypyrene (1-OHP) were compared in Sprague–Dawley and Wistar rats. Groups of five male rats, of about 200 g of body weight, were injected intravenously with 0.05, 0.5, 5 and 50 µmol pyrene kg^?1 of body weight. Urine was collected at 2, 4, 6, 8, 10, 12, 18, 24, 30, 42 and 48 h post-dosing. Pyrene metabolites were measured by high-performance liquid chromatography (HPLC)/fluorescence after enzymatic hydrolysis of the glucurono- and sulfo-conjugates, extraction on Sep-Pak C18 cartridges and, for the analysis of dione metabolites, derivatization to stable diacetoxypyrene molecules. Over the 48-h sampling period, on average 17.4–25.6% of the injected pyrene was excreted overall as P16D, 6.4–8.8% as P18D and 0.6–0.8% as 1-OHP in the urine of Sprague–Dawley rats. By comparison, on average 10.3–14.7% of the intravenous pyrene dose was recovered as P16D, 4.8–6.4% as P18D and 0.3–0.4% as 1-OHP in the urine of Wistar rats. In both strains of rats there was no clear effect of the dose on the 0–48-h cumulative urinary excretion of P18D and 1-OHP over the entire dose range, while the percentage of dose recovered overall as P16D in urine at the highest dose (50 µmol kg^?1) was statistically lower than at the other doses. The 0–48-h cumulative percentage of pyrene dose excreted as metabolites in the urine of Sprague–Dawley rats was also significantly higher than in Wistar rats (p<0.01) exposed under identical conditions. As for the urinary excretion-time courses of the different metabolites, for a given dose and strain of rats, excretion curves of P16D, P18D and 1-OHP generally evolved in parallel. There was also no clear effect of the dose on the excretion rate, thus half-life, of pyrene metabolites, except for P16D in Sprague–Dawley rats at the highest dose where elimination tended to be slower compared with the other doses (p<0.01). The average first-order elimination half-life of P16D, P18D and 1-OHP was 4.0, 5.7 and 4.1 h, respectively, in Sprague–Dawley rats, and 5.1, 6.1 and 5.1 h, respectively, in Wistar rats (all doses combined but excluding the highest dose for P16D). This study showed the relative importance of metabolic pathways leading to diones compared with 1-OHP. These dioxygenated metabolites appear to be interesting biomarkers of pyrene exposure at environmentally and occupationally relevant doses. Their adequacy as biomarkers of human exposure has yet to be confirmed.     

Metabolism of N-(3′,5′-Dichlorophenyl)succinimide in Rats and Dogs

When N-(3′,5′-dichlorophenyl)succinimide (DSI)-carbonyI-¹⁴C and –pheny-³H were each orally administered to rats, regardless of the label site, most of the dose was readily eliminated. There was no difference in the excretion rate between male and female rats. No radioactive residues were detected in tissues and organs 24 hr after dosing. Urinary metabolites consisted of N-(3′,5′-dichlorophenyl) succinamic acid (DSA), N-(3′,5′-dichlorophe-nyl) malonamic acid (DMA), N-(3′5’-dichlorophenyl)-2-hydroxysuccinamic acid (2-OH-DSA) and 2-OH-DSA derivatives. In dogs, most of the administered dose was excreted in equal amounts in urine and feces. 2-OH-DSA derivatives were main urinary metabolites and most of fecal radiocarbon was due to intact DSL. The results of this study indicate that DSI is a biodegradable compound which is unlikely to leave any persistent residues in animals.

The degradation of DSI to DSA was mediated by an arylamidase-type hydrolase, which was present in the microsomal fraction of rabbit liver. The enzyme activity was found in livers and kidneys of four animal species tested. Depending on the animal species, the enzyme appears to be important for the metabolism of DSI.     

The biotransformation and urinary excretion of dexamethasone in equine male castrates

The pro-drugs of dexamethasone, a potent glucocorticoid, are frequently used as anti-inflammatory steroids in equine veterinary practice. In the present study the biotransformation and urinary excretion of tritium labelled dexamethasone were investigated in cross-bred castrated male horses after therapeutic doses. Between 40-50% of the administered radioactivity was excreted in the urine within 24 h; a further 10% being excreted over the next 3 days. The urinary radioactivity was largely excreted in the unconjugated steroid fraction. In the first 24 h urine sample, 26-36% of the total dose was recovered in the unconjugated fraction, 8-13% in the conjugated fraction and about 5% was unextractable from the urine. The metabolites identified by microchemical transformations and thin-layer chromatography were unchanged dexamethasone, 17-oxodexamethasone, 11-dehydrodexamethasone, 20-dihydrodexamethasone, 6-hydroxydexamethasone and 6-hydroxy-17-oxodexamethasone together accounting for approx 60% of the urinary activity. About 25% of the urinary radioactivity associated with polar metabolites still remains unidentified.     

Fecal androgen metabolite analysis for noninvasive monitoring of testicular steroidogenic activity in felids

Limited data are available on long-term, seasonal changes in testicular steroidogenic activity in nondomestic felids, primarily because of the difficulties associated with longitudinal blood sampling (e.g., handling, restraint, anesthesia). Therefore, a noninvasive approach for assessing testicular androgen production was developed using the domestic cat (Felis catus) as a model. Two adult males were injected i.m. with 4 μCi¹⁴-testosterone to determine the time course and relative proportions of androgen metabolites excreted in urine and feces. Peak urinary radioactivity was detected 13 and 19 hr postinjection and accounted for ∼8% of the total radioactivity recovered. High performance liquid chromatography (HPLC) analysis detected multiple polar urinary metabolites, none of which eluted with the ³H-testosterone reference tracer. The majority of urinary testosterone metabolites consisted of nonenzyme-hydrolyzable, water-soluble (presumably conjugated) forms. In feces, radioactivity was detected in the first sample collected at 22 hr postinjection for both males, although peak metabolite excretion in one male was not observed until 61 hr postinjection. HPLC analysis detected several fecal metabolites consisting primarily of nonhydrolyzable, water-soluble forms (84.4 ± 0.9%) with some ether-soluble forms (15.6 ± 0.9%). None of the fecal androgen metabolites were associated with free testosterone. However, one or more of the water-soluble fecal metabolites was quantifiable using a commercially available testosterone radioimmunoassay. The biological relevance of this immunoactivity was confirmed in the domestic cat; concentrations were high in adult, intact males and nondetectable in intact females and castrated males and females. In addition, fecal androgen concentrations in a male Pallas' cat (Felis manul) exhibited seasonal fluctuations that corresponded with parallel changes in serum testosterone and ejaculate quality. These data indicate that testicular steroidogenic activity can be monitored non invasively in felids, providing a potentially valuable tool for endangered felid management to: (1) assess pubertal status, (2) determine the influence of season on reproduction, and (3) diagnose possible causes of sub- or infertility. © 1996 Wiley-Liss, Inc.     

Stereoselectivity of the aliphatic hydroxylation of 6-n-propylchromone-2-carboxylic acid in rat and guinea pig

Following administration of 6-n-propylchromone-2-carboxylic acid (6-n-PCCA) (500 μmol/kg) to male rats, three metabolic products were detected and isolated from the 0–24 h urine. All were identified as resulting from oxidation exclusively along the 6-n-propyl moiety. Some 66% of the dose was excreted in the 0–24 h urine, 55% of which was 6-PCCA, with 15% as (6-1′-hydroxypropyl)chromone-2-carboxylic acid (6-1′-HPCCA), 22% as 6-(2′-hydroxypropyl)chromone-2-carboxylic acid (6-2′-HPCCA), and 4% as (6-3′-carboxypropyl)chromone-2-carboxylic acid (6-3′-CPCCA). Derivatization of the methyl esters of the hydroxylated metabolities with S-α-methoxy-α-(trifuloromethyl)-phenylacetyl chloride (Mosher's reagent) allowed the evaluation of urinary enantiomeric composition by HPLC and assignment of their absolute configurations by NMR. This was found to be 90:10 (R/S) for 6-2′-HPCCA, and 7:93 (R/S) for 6-1′-HPCCA. When rats were dosed with the racemic 1′- and 2-hydroxy metabolites; no stereoselective metabolism or excretion was observed. Administration of 6-n-PCCA to male guinea pigs revealed that this species was unable to metabolise this compound. © 1993 Wiley-Liss, Inc.     

Effects of dehydration,rehydration, and hyperhydration in the lactating and non-lactating black Moroccan goat

The effects of water deprivation, rehydration and hyperhydration were investigated in the black Moroccan goat (Capra hircus). Mean daily water intake was 46 ± 5 ml/kg in lactating and 36 ± 4 ml/kg in non-lactating black Moroccan goats, and milk production 21 ± 1 ml/kg. Mean urine excretion was 8 ± 2 ml/kg body weight in both groups, and the daily water losses via evaporation and feces were estimated at 23 ± 3 ml/kg during lactation and 28 ± 4 ml/kg during non-lactation. Forty-eight hours of water deprivation caused a body weight loss of 9% and 6% in lactating and non-lactating goats, respectively, and a drop of 28% in milk production with only a slight decrease in food intake. After rehydration, the elevated plasma osmolality as well as Na and total protein concentrations returned to basal values within 2–3 hr, indicating a rapid absorption of the ingested water, but urine excretion did not increase. After hyperhydration (10% of body weight), 46% of the load was excreted by the kidneys within 6 hr. In conclusion, black Moroccan goats have a low water turnover, and they can retain water upon rehydration but not store excess water after hyperhydration.     

Bioavailability of (-)-epicatechin upon intake of chocolate and cocoa in human volunteers

We evaluated the levels of (-)-epicatechin (EC) and its metabolites in plasma and urine after intake of chocolate or cocoa by male volunteers. EC metabolites were analyzed by HPLC and LC/MS after glucuronidase and/or sulfatase treatment. The maximum levels of total EC metabolites in plasma were reached 2 hours after either chocolate or cocoa intake. Sulfate, glucuronide, and sulfoglucuronide (mixture of sulfate and glucuronide) conjugates of nonmethylated EC were the main metabolites present in plasma rather than methylated forms. Urinary excretion of total EC metabolites within 24 hours after chocolate or cocoa intake was 29.8 ± 5.3% and 25.3 ± 8.1% of total EC intake. EC in chocolate and cocoa was partly absorbed and was found to be present as a component of various conjugates in plasma, and these were rapidly excreted in urine.     

Smoking of phencyclidine: Disposition in man and stability to pyrolytic conditions

Two human volunteers who smoked parsley cigarettes to which 0.1 mg of tritium-labeled phencyclidine (PCP) hydrochloride had been added received ca. 59% of the applied dose, of which ca. 56% was excreted in the urine within 72 hr. PCP in smoke appeared to be absorbed well. Unchanged PCP and conjugated metabolites were found in plasma and urine. 1-Phenylcyclohexene and its metabolites were also present but did not interfere with analysis of PCP. PCP hydrochloride was extensively degraded (≥93%) at 300–800°C in a quartz pyrolysis tube, but was much less degraded under simulated smoking conditions on parsley cigarettes. Major products of pyrolysis were 1-phenylcyclohexene and piperidine hydrochloride.     

Stereoselective disposition of tiaprofenic acid enantiomers in rats

The pharmacokinetics and metabolic chiral inversion of the S(+)‐ and R(−)‐enantiomers of tiaprofenic acid (S‐TIA, R‐TIA) were assessed in vivo in rats, and in addition the biochemistry of inversion was investigated in vitro in rat liver homogenates. Drug enantiomer concentrations in plasma were investigated following administration of S‐TIA and R‐TIA (i.p. 3 and 9 mg/kg) over 24 hr. Plasma concentrations of TIA enantiomers were determined by stereospecific HPLC analysis. After administration of R‐TIA it was found that 1) there was a time delay of peak S‐TIA plasma concentrations, 2) S‐TIA concentrations exceeded R‐TIA concentrations from ∼2 hr after dosing, 3) C_max and AUC_(0‐∞) for S‐TIA were greater than for R‐TIA following administration of S‐TIA, and 4) inversion was bidirectional but favored inversion of R‐TIA to S‐TIA. Bidirectional inversion was also observed when TIA enantiomers were incubated with liver homogenates up to 24 hr. However, the rate of inversion favored transformation of the R‐enantiomer to the S‐enantiomer. In conclusion, stereoselective pharmacokinetics of R‐ and S‐TIA were observed in rats and bidirectional inversion in rat liver homogenates has been demonstrated for the first time. Chiral inversion of TIA may involve metabolic routes different from those associated with inversion of other 2‐arylpropionic acids such as ibuprofen. Chirality 11:103–108, 1999. © 1999 Wiley‐Liss, Inc.     

Steroid metabolism in the rabbit. Biliary and urinary excretion of metabolites of [4-14C]progesterone

1. [4-(14)C]Progesterone was administered intravenously to anaesthetized male and female New Zealand White rabbits as a single injection or as a 45-60min. infusion. 2. After a single dose about 60% of the radioactivity was recovered in 6hr., and twice as much radioactivity was present in bile as in urine. After infusion total recovery of radioactivity was only about 40% in 6hr., but the relative proportions of metabolites in bile and urine were about the same as after a single dose. 3. Bile and urine samples were hydrolysed successively by beta-glucuronidase, cold acid and hot acid. 4. In bile the major proportion of metabolites appeared in the glucuronide fraction; in urine beta-glucuronidase hydrolysis yielded the greatest amounts of ether-extractable radioactivity, but the greatest proportion of radioactivity could not be extracted by ether from an alkaline solution of the hydrolysed urine. 5. There was no apparent difference in the quantity or distribution of metabolites excreted by male and female animals.     

Metabolic disposition of [14C] metanil yellow in rats

The absorption, metabolism and excretion of [14C] metanil yellow was studied in rats. Following administration of a single oral dose of 5 mg dye (7.6 microCi)/kg body weight, 80.5% of the dose was excreted in the urine and faeces within 96 hr, with the majority being accounted for in the faeces. Liver, kidney, spleen and testis retained no count whereas 13.6% of the radioactivity was retained by gastrointestinal tract. Analysis of urine and faeces detected two azo-reduction metabolites of metanil yellow which were characterized by TLC and IR, NMR and mass spectroscopic studies as metanilic acid and p-aminodiphenylamine.     

Steroid metabolism and validation of noninvasive endocrine monitoring in the African wild dog (Lycaon pictus)

The purpose of this study was to validate noninvasive endocrine monitoring techniques for African wild dogs (Lycaon pictus) and to establish physiological validity of these methods by evaluating longitudinal reproductive-endocrine profiles in captive individuals. To determine the primary excretory by-products of ovarian steroid metabolism, [¹⁴C]-progesterone and [³H]-estradiol were co-administered to a female and all excreta were collected for 80 hr postinjection. Radiolabel excretion peaked ≤ 18 hr postinfusion, and progesterone and estradiol metabolites were excreted in almost equivalent proportions in urine (39.7 and 41.1%, respectively) and feces (60.3 and 58.9%, respectively). Most of the urinary metabolites were conjugated (estradiol, 94.3 ± 0.3%; progesterone, 90.4 ± 0.5%), so that immunoassays for pregnanediol-3α-glucuronide (PdG) and estrogen conjugates (EC) were effective for assessing steroid metabolites. Two immunoreactive estrogens (estradiol and estrone) and at least one immunoreactive progesterone metabolite (3α-hydroxy-5α, pregnan-20-one) were detected in feces. Urine and fecal samples were collected (1–3 times per week) for 1.5 yr from one adult female and two adult males to assess longitudinal steroid metabolite excretion. Overall correlation of urinary PdG to matched, same-day fecal progesterone metabolites immunoreactivity was 0.38 (n = 71, P < 0.05). Similarly, urinary EC was correlated (P < 0.05) with same-day fecal estrogen immunoreactivity (r = 0.49, n = 71). During pregnancy and nonpregnant cycles, copulation occurred at the time of peak (or declining) estrogen metabolites and increasing progesterone metabolites concentrations. Estrus duration was 6–9 days and gestation lasted 69 days with parturition occurring coincident with a drop in progesterone metabolites. Males exhibited seasonal trends in fecal testosterone excretion with maximal concentrations from July to September coincident with peak mating activity. Although these limited longitudinal hormone profiles should be interpreted cautiously, noninvasive gonadal steroid monitoring suggests that: (1) both female and male wild dogs may exhibit reproductive seasonality in North America, (2) females are monoestrous, and (3) peak testicular activity occurs between August and October coincident with mating behavior. From a conservation perspective, noninvasive endocrine monitoring techniques should be useful for augmenting captive breeding programs, as well as for developing an improved understanding of the physiological mechanisms underlying reproductive suppression in response to social and ecological pressures. Zoo Biol 16:533–548, 1997. © 1997 Wiley-Liss, Inc.     

Absorption of cadmium after a long-term oral administration of cadmium to dogs

A long-term experiment using beagle dogs to investigate the absorption of cadmium was conducted. The dogs in the experimental groups were given a commercial diet and pelleted food containing 1, 3, 10, 50, and 100 mg of cadmium per day. The cadmium concentration in the blood increased continuously, gradually reaching a steady state following the administration of cadmium. The cadmium excreted daily in urine increased continuously. The cumulative excreted amount of cadmium in urine was calculated by using the trapezoidal rule based on the data of excretion of cadmium in urine. Then the absorbed fraction of administered cadmium was estimated on the basis of the relationship between the cumulative excreted amount of cadmium in urine and the cumulative administered dose of cadmium after the cadmium concentration in blood reached a steady state. The absorbed fraction of cadmium decreased with an increase in the administered dose of cadmium. A dose-dependent increase between the absorbed amount and the administered dose was observed.     

The metabolism of arylazoisoxazolones by rats and dogs

1. When rats were given a single oral dose of the lipid-soluble fungicide 4-(2-chlorophenylhydrazono)-3-methyl[4-(14)C]isoxazol-5-one ([(14)C]drazoxolon), about 75% of the label was excreted in the urine and 13% in the faeces in 96hr. An additional 7% of the radioactivity was recovered as (14)CO(2) in 48hr. 2. About 8% of the label was excreted by rats in the bile in 0-24hr. and an additional 6% was excreted by the same route in 24-48hr. 3. When dogs were given a single oral dose of [(14)C]drazoxolon about 35% of the label was excreted in the urine and a similar amount was excreted in the faeces in 96hr. 4. The major metabolites in the urine of the rat and the dog were identified as 2-(2-chloro-4-hydroxyphenylhydrazono)acetoacetic acid (dog, 14%), the corresponding ether glucosiduronic acid (dog, 12%; rat, 13%) and ester sulphate (rat, 65%). 5. When rats were given a single oral dose of 3-methyl-4-([U-(14)C]phenylhydrazono)isoxazol-5-one about 75% of the label was excreted in the urine and 15% in the faeces in 96hr. The major metabolite in the urine was identified as the ester sulphate conjugate of 2-(4-hydroxyphenylhydrazono)-acetoacetic acid. 6. Reduction of the azo link was of minor quantitative significance. 7. These results are discussed in their relation to species differences in the toxicity of these compounds.     

Human urine: Epicatechin metabolites and antioxidant activity after cocoa beverage intake

Associations between cocoa consumption in humans, excreted metabolites and total antioxidant capacity (TAC) have been scarcely investigated. The aims of the study were to investigate the epicatechin (( ? )-Ec) metabolites excreted in urine samples after an intake of 40 g of cocoa powder along with the TAC of these urine samples and the relation between both the analyses. Each of the 21 volunteers received two interventions, one with a polyphenol-rich food (PRF) and one with a polyphenol-free food (PFF) in a randomized cross-over study. Urine samples were taken before and during 24 h at 0–6, 6–12 and 12–24 h periods after test intake. The excreted ( ? )-Ec metabolites and the TAC were determined in urine samples by LC-MS/MS and TEAC assay, respectively. The maximum excretion of ( ? )-Ec metabolites and the maximum TAC value were observed in urine samples excreted between 6 and 12 h after PRF consumption. Significance of TAC increase was found in urine samples excreted during 0–6 and 6–12 h (66.6 and 72.67%, respectively, with respect to the 0 h).     

Studies on flavonoid metabolism. Biliary and urinary excretion of metabolites of (+)-[U-14C]catechin

1. The biliary and urinary excretion of (+)-[U-(14)C]catechin was studied in normal male rats after a single injection of the flavonoid. 2. In rats large amounts of radioactivity (33.6-44.3% of the dose in 24h) were excreted in the bile as two glucuronide conjugates [one of which was a (+)-catechin conjugate] and three other unconjugated metabolites. 3. Excretion of radioactivity in the urine when the bile duct was not cannulated amounted to 44.5% of the dose. 4. In both the urine and bile the new metabolites showed maximum excretion in the (1/2)-1(1/2)h after intravenous injection of [(14)C]catechin. 5. The metabolites m-hydroxyphenylpropionic acid, p-hydroxyphenylpropionic acid, delta-(3-hydroxyphenyl)-gamma-valerolactone and delta-(3,4-dihydroxyphenyl)-gamma-valerolactione originate from the action of the intestinal micro-organisms on the biliary-excreted metabolites of (+)-catechin. These phenolic acid and lactone metabolites are then reabsorped and excreted in the urine. 6. It is proposed that, depending on the route of administration of (+)-catechin, there exists an alternative pathway, involving biliary excretion, for the metabolism of (+)-catechin.