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

1. [4-(14)C]Testosterone was administered intravenously to anaesthetized male cats as a single injection or as a 45-60min. infusion. 2. Most of the administered radioactivity was excreted in the bile (70-80%); only 2.9-5.5% of the dose was excreted in the urine. 3. Bile and urine samples were hydrolysed successively to yield glucuronide, ;cold-acid-hydrolysed' and ;hot-acid-hydrolysed' fractions. 4. The proportion of glucuronides in bile decreased in successive samples, but cold-and hot-acid-hydrolysed metabolites showed no consistent change. 5. After hydrolysis most of the radioactivity in both bile and urine could not be extracted by ether from neutral aqueous solution.     

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.     

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.     

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

1. [4-(14)C]Cortisone was administered to anaesthetized male and female New Zealand White rabbits as a single injection or as a 45-60min infusion. 2. The method of administration of the steroid did not significantly affect the total excretion of radioactivity in bile and urine [83.8+/-10.8%(s.d.)]. 3. The mean ratio of metabolites in urine to those in bile was 0.97+/-0.23% (range 0.64-1.3). 4. When bile and urine samples were hydrolysed successively by beta-glucuronidase, cold acid and hot acid, neutral metabolites extracted by ethyl acetate-ether were found mainly after hydrolysis by beta-glucuronidase. 5. An approximately equal proportion of the dose was converted into substances not extractable from alkaline aqueous solution after hydrolysis.     

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

1. [4-(14)C]Cortisone was administered to anaesthetized male cats as a single injection or as a 45-60min. infusion. 2. After the single dose a total of 69.6-89.6% of the radioactivity was excreted in bile, and 0.5-7.1% in urine. After infusion total recovery in bile was 73.4-92.1%, and 1.2-1.7% in urine. 3. When bile and urine samples were hydrolysed successively by beta-glucuronidase, cold acid and hot acid, most of the radioactivity was converted into substances not extractable from neutral aqueous solution by ethyl acetate-ether. 4. In bile, metabolites hydrolysable by beta-glucuronidase were found in only small proportions (3-4%) of the dose.     

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

1. [4-¹⁴C]Oestradiol was administered to seven male, seven female and two castrated male cats as a single intravenous injection. Bile and urine were collected for 6h. 2. The radioactivity was excreted mainly in the bile of all animals (53–60%); only approx. 1% of the dose appeared in the urine. 3. Bile and urine samples were hydrolysed successively by β-glucuronidase, cold acid and hot acid. There were significant differences (P<0.005) between the percentage of the dose present in the bile fractions hydrolysed by β-glucuronidase (male, 9.0±1.7%; female, 18.6±1.45%) and by cold acid (male, 18.9±1.44%; female 12.1±1.02%). The excretion of radioactivity in these fractions by the castrated male cats was closer to that of female cats. 4. Approx. 20–27% of the dose could not be extracted from aqueous solution (pH10.5) by ethyl acetate–ether after hydrolysis.     

Biliary excretion of barium in the rat

Biliary excretion of barium was studied in Sprague-Dawley bile-duct-cannulated rats injected intravenously with 1.8 micrograms Ba/rat as 133Ba-labeled barium chloride. Approximately 0.5% of the barium dose was excreted into bile within 2 h. The time-course profile of biliary excretion of the radiotracer closely reflected that of plasma concentrations. Biliary barium levels reached their peak in the first 15-min period after administration and rapidly declined thereafter. The plasma-to-bile barium-concentration ratio was approx 1 at 2 h after injection. There was no tendency of barium to concentrate in liver, and the 133Ba levels in stomach and small intestine largely exceeded hepatic levels. There is evidence indicating that barium is predominantly excreted with feces following parenteral administration in rats and humans. The results of this study suggest that biliary excretion is of little quantitative importance and that physiological routes other than bile contribute to elimination of barium by the digestive tract.     

Biliary metabolites of estriol in the rat

Following the subcutaneous administration of estriol-6,7-³H to rats, biliary metabolites were identified and quantitated. Approximately 70% of the metabolites were excreted in the form of “glucosiduronate” conjugates. 3, 17β-Dihydroxy-2-methoxy-1,3,5(10)-estratrien-16-one was the major metabolite in this conjugate fraction. Significant amounts of 3,17β-dihydroxy-1,3,5(10)-estratrien-16-one and 2,3,17β-trihydroxy-1,3,5(10)-estratrien-16-one, as well as smaller quantities of 1,3,5(10)-estratriene-2,3,16α,17β-tetrol and 2-methoxy-1,3,5(10)-estratriene-3,16α, 17β-triol, were also found. In 17α-ethinylestradiol - treated animals, the rate of excretion of radioactivity and the proportion of 16-oxo-17β-ol metabolites found in the “glucosiduronate” fraction were reduced.     

The metabolism of niacytin in the rat. Studies of the excretion of nicotinic acid metabolites

1. Nicotinic acid-deficient rats were given a dose of niacytin or an equivalent one of free nicotinic acid or hydrolysed niacytin. 2. The excretion of N'-methylnicotinamide and of tertiary nicotinoyl derivatives in urine showed that niacytin was not metabolized as free nicotinic acid, although hydrolysed niacytin was equivalent to free nicotinic acid. 3. Little or none of the niacytin dose was recovered as tertiary nicotinoyl derivatives in faeces. This result was not affected by fitting rats with tail-cups to prevent coprophagy. 4. At the high doses used niacytin restored the growth rate of the deficient animals because of a small degree of hydrolysis of the bound nicotinic acid.     

Biliary excretion of amphetamine and methamphetamine in the rat

1. (14)C-labelled amphetamine and methamphetamine were injected into rats cannulated at the bile duct under thiopentone anaesthesia and the output of their metabolites in urine and bile was determined. 2. With amphetamine, 69% of the (14)C was excreted in the urine and 16% in the bile in 24h. The main metabolite in bile was the glucuronide of 4-hydroxyamphetamine. The output of unchanged amphetamine was much greater in cannulated rats than in intact rats. 3. With methamphetamine, 54% of the (14)C appeared in the urine and 18% in the bile. The main metabolite in the bile was the glucuronide of 4-hydroxynorephedrine. The output of amphetamine, a metabolite of methamphetamine, was much greater in cannulated rats than in intact rats. 4. Evidence has been obtained for the enterohepatic circulation of certain amphetamine and methamphetamine metabolites in the rat. 5. Thiopentone anaesthesia appeared to inhibit the ring hydroxylation of amphetamine administered as such or formed as a metabolite of methamphetamine.     

Flavonoid metabolism: the interaction of metabolites and gut microbiota

Abstract

Several dietary flavonoids exhibit anti-oxidative, anti-inflammatory, and anti-osteoporotic activities relevant to prevention of chronic diseases, including lifestyle-related diseases. Dietary flavonoids (glycoside forms) are enzymatically hydrolyzed and absorbed in the intestine, and are conjugated to their glucuronide/sulfate forms by phase II enzymes in epithelial cells and the liver. The intestinal microbiota plays an important role in the metabolism of flavonoids found in foods. Some specific products of bacterial transformation, such as ring-fission products and reduced metabolites, exhibit enhanced properties. Studies on the metabolism of flavonoids by the intestinal microbiota are crucial for understanding the role of these compounds and their impact on our health. This review focused on the metabolic pathways, bioavailability, and physiological role of flavonoids, especially metabolites of quercetin and isoflavone produced by the intestinal microbiota.     

Biliary metabolites of all-trans-retinoic acid in the rat

Biliary metabolites from physiological doses of all-trans-[10-3H]retinoic acid were examined in normal and vitamin A-deficient rats. The bile from normal and vitamin A-deficient rats contained approximately 60% of the administered dose following a 24-h collection period. However, vitamin A-deficient rats show a 6-h delay in the excretion of radioactivity compared to normal rats. Retinoyl-beta-glucuronide excretion was particularly sensitive to the vitamin A status of the rats. In normal rats, retinoyl-beta-glucuronide reached a maximum concentration of 235 pmol/ml of bile 2 h following the dose and then rapidly declined. Vitamin A-deficient rats show a relatively constant concentration of this metabolite (100-150 pmol/ml of bile) over a 10-h collection period. Retinoic acid excretion was low in both normal and deficient rats. The concentration of retinotaurine, a recently identified biliary metabolite, was approximately equal to retinoyl-beta-glucuronide in normal rats and appeared in the bile 2 h later than the glucuronide.     

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

1. [4-(14)C]Testosterone was administered to anaesthetized male and female New Zealand White rabbits as a single injection or as a 45-60min. infusion. 2. After a single dose a total of approx. 56-80% of the radioactivity was excreted in bile and urine. After infusion total recovery of radioactivity was approx. 63-75%. 3. The mean ratio of metabolites in urine to those in bile was 0.77+/-0.41 (range 0.3-1.5). 4. Bile and urine samples were hydrolysed successively by beta-glucuronidase, cold acid and hot acid. In both bile and urine neutral metabolites extracted by ethyl acetate-ether were found mainly after beta-glucuronidase hydrolysis, but a considerable proportion of the dose was converted into substances not extractable from alkaline aqueous solution after all forms of hydrolysis used.     

The role of the gut flora in the metabolism of cyclamate

1. [(14)C]Cyclamate was not metabolized when incubated with the liver, spleen, kidney or blood of rats of rabbits kept on a cyclamate-containing diet, and that had become converters of cyclamate into cyclohexylamine. 2. [(14)C]Cyclamate was converted into cyclohexylamine when incubated under anaerobic conditions with the contents of the caecum, colon or rectum or with the faeces of cyclamate-pretreated rats. Similar results were obtained with cyclamate-pretreated rabbits. With cyclamate-pretreated guinea pigs, which did not readily convert cyclamate into cyclohexylamine, the colon contents showed only low activity in this respect. 3. The faeces of a human converter of [(14)C]cyclamate into cyclohexylamine were also very active, but became less active when cyclamate was removed from his diet. 4. On subculturing the organisms from the contents of the colon and rectum of rats, the ability to convert cyclamate into cyclohexylamine was lost during three subcultures, but the loss of the activity was considerably decreased by subculturing in the presence of cyclamate. 5. Incubation of rat faeces in broths containing cyclamate increased their ability to metabolize cyclamate, but similar treatment of rabbit and human faeces suppressed this activity. 6. When rats are kept on a cyclamate diet the number of clostridia in the faeces increased considerably. In human dietary cyclamate did not appear to alter the counts of various faecal micro-organisms. 7. The gut organisms that appear to develop the ability to convert cyclamate into cyclohexylamine are clostridia in rats, enterobacteria in rabbits and enterococci in man. 8. [(14)C]Cyclohexylamine injected into the caecum or colon of rats is readily absorbed and excreted in the urine. 9. It appears that on continued intake of cyclamate the gut flora develop the ability to convert cyclamate into cyclohexylamine, which is then absorbed and excreted mainly in the urine, although a small proportion is metabolized to other compounds.     

Biliary excretion of foreign compounds. Biphenyl, stilboestrol and phenolphthalein in the rat: molecular weight, polarity and metabolism as factors in biliary excretion

1. The extent of biliary excretion of biphenyl, tetralin, stilboestrol and phenolphthalein was studied in the rat. 2. Biphenyl and its 4-hydroxy and 4,4′-dihydroxy derivatives are extensively excreted in the bile as glucuronides in amounts increasing in order of molecular weight. 3. Stilboestrol and its glucuronide are excreted almost quantitatively in the bile mainly as the monoglucuronide, as are also phenolphthalein and its glucuronide. 4. Tetralin is excreted to the extent of about 13% of the dose, mainly as ac-tetralyl glucuronides. 5. The results and those of Abou-El-Makarem, Millburn, Smith & Williams (1967) are discussed and it is concluded that the extent of biliary excretion of foreign compounds in rats depends on their molecular weight and their possessing a strongly polar anionic group. There appears to be a minimum value of this molecular weight below which little biliary excretion (i.e. not more than 5–10% of the dose) occurs. There is some latitude in the choice of this molecular weight, which is about 325±50. The necessary molecular weight and polar group can be acquired by metabolism. Above this minimum value biliary excretion increases with molecular weight. It is suggested that the mechanism of the biliary excretion of foreign compounds may be similar to that of conjugated bile acids, which are highly polar and whose molecular weights exceed 400.     

Biliary metabolites of testosterone and testosterone glucosiduronate in the rat

A mixture of testosterone-4-¹⁴C and testosterone-1,2-³H-17-glucosiduronate was intraperitoneally administered into male and female rats with bile fistulas. Biliary metabolites were separated and purififd by a combination of column chromatography, enzymic hydrolysis or solvolysis of the conjugate fractions and identification of the liberated aglycones. The injected steroids were extensively metabolized and excreted predominantly in the blue. 5β-Androstane-3α, 17β-diol was found principally in monoglucosiduronate fraction and was produced preferentially from the injected conjugate in both sexes. Very marked sex differences from the injected conjugate in both sexes. Very marked sex differences were observed in the following metabolites: Androsterone was present only in the female as monoglucosidironate, which was preferentially derived from testosterone. 5α-Androstane-3α,17β-diol was identified in both monoglucosiduronate and diconjugate fractions of the female, which was formed significanrly more from the conjugate than testosterone. These findings provide evidence that testosterone glucosiduronate could be converted directly into 5α-steroids as well as 5β-ones $\text{in}$$\text{vivo}$. In marked contrast, the major portion of testosterone was metabolized to polar steroids in the male.     

Biliary excretion of tauroursodeoxycholate-3-sulfate in the rat

Biliary organic anion excretion is mediated by an ATP-dependent primary active transporter, multidrug resistance protein 2. On the other hand, a multiplicity of canalicular organic anion transport has been suggested. Ursodeoxycholic acid, the 7beta-epimer of chenodeoxycholic acid, is clinically used for various hepatobiliary diseases. In our previous study, the contribution of multidrug resistance protein 2 for biliary excretion of taurine-conjugated bile acid sulfates depended on the numbers of hydroxyl residue. Therefore, to further examine the effect of hydrophobicity on the substrate specificity of multidrug resistance protein 2, we examined the effect of bile acid conjugates and organic anions on biliary excretion of tauroursodeoxycholate-3-sulfate, taurine and sulfonate-conjugated ursodeoxycholic acid, in rats. Biliary tauroursodeoxycholate-3-sulfate excretions was markedly delayed in Eisai hyperbilirubinemic rats. Taurolithocholate-3-sulfate inhibited but ursodeoxycholate-3,7-disulfate did not affect biliary tauroursodeoxycholate-3-sulfate excretion. Biliary tauroursodeoxycholate-3-sulfate excretion was inhibited by sulfobromophthalein, but was not inhibited by dibromosulfophthalein and cefpiramide. These findings indicate that tauroursodeoxycholate-3-sulfate is very specific for multidrug resistance protein 2.