Metabolism of 2,6-dichlorobenzamide in rats and mice

1. Oral doses of 2,6-dichlorobenzamide (DCB) were excreted by rats as DCB, two monohydroxy-DCBs, 2-chloro-5-hydroxy-6-(methylthio)benzamide and 2-chloro-5-hydroxy-6-[S-(N-acetyl)cysteinyl]benzamide (mercapturic acid).

2. Biliary excretion (33% of the dose), enterohepatic circulation and intestinal microfloral metabolism were involved in formation of 2-chloro-5-hydroxy-6-(methylthio)benzamide, and the mercapturic acid served as a precursor.

3. Whole body autoradiography and microautoradiography showed the accumulation of non-extractable. residues from DCB in the nasal mucosa and contents of the large intestines of rats and mice dosed with ¹⁴C-labelled DCB.     

Excretion of mercapturic acids in human urine after occupational exposure to 2-chloroprene

A pilot study was conducted for human biomonitoring of the suspected carcinogen 2-chloroprene. For this purpose, urine samples of 14 individuals occupationally exposed to 2-chloroprene (exposed group) and of 30 individuals without occupational exposure to alkylating substances (control group) were analysed for six potential mercapturic acids of 2-chloroprene: 4-chloro-3-oxobutyl mercapturic acid (Cl-MA-I), 4-chloro-3-hydroxybutyl mercapturic acid (Cl-MA-II), 3-chloro-2-hydroxy-3-butenyl mercapturic acid (Cl-MA-III), 4-hydroxy-3-oxobutyl mercapturic acid (HOBMA), 3,4-dihydroxybutyl mercapturic acid (DHBMA) and 2-hydroxy-3-butenyl mercapturic acid (MHBMA). In direct comparison with the control group, elevated levels of the mercapturic acids Cl-MA-III, MHBMA, HOBMA and DHBMA were found in the urine samples of the exposed group. Cl-MA-I and Cl-MA-II were not detected in any of the samples, whereas HOBMA and DHBMA were found in all analysed urine samples. Thus, for the first time, it was possible to detect HOBMA and Cl-MA-III in human urine. The mercapturic acid Cl-MA-III could be confirmed as a specific metabolite of 2-chloroprene in humans providing evidence for the intermediate formation of a reactive epoxide during biotransformation. The main metabolite, however, was found to be DHBMA showing a distinct and significant correlation with the urinary Cl-MA-III levels in the exposed group. The obtained results give new scientific insight into the course of biotransformation of 2-chloroprene in humans.     

Formation of methylthio metabolites of indene in the guinea pig and the rat

The formation of methylthio metabolites of epoxides has been shown to be a significant route of metabolism in some species. Several aspects of this metabolic conversion for indene were examined. Two isomers of hydroxy(methylthio)indane were found in the urine of guinea pigs administered indene (14.3 and 100 mg/kg, ip). The major isomer, 2-hydroxy-1-methylthioindane (I) was present as 6-9% of the administered dose after 24 hr, while lower amounts (0-0.6%) of a minor isomer (II) were observed. A significant amount of isomer I was found as a urinary metabolite of indene oxide (14% of 12.5 mg/kg, ip). To further elucidate the route of formation of I, the glutathione (I-GLU) and mercapturic acid (I-MER) conjugates of indene oxide were synthesized and administered to the guinea pig. The methylthio metabolite I was present as a significant urinary metabolite of both conjugates of indene oxide, comprising 9.6% and 5.7% of the dose of I-GLU (5 mg, ip) and I-MER (4 mg, ip), respectively. These results show that the formation of a hydroxy(methylthio)indane is a significant route of metabolism for indene and indene oxide in the guinea pig, and that this metabolite arises via further metabolism of conjugates in the glutathione pathway. In the rat, isomer I is a minor metabolite. Mechanistic aspects of the formation of these thioether metabolites are discussed.     

Glutathione conjugation of 1,2-dibromo-1-phenylethane in rats in vivo

The metabolism of 1,2-dibromo-1-phenylethane (DBPE) was studied in rats. Administration of DBPE orally, in doses of 0.25-1.25 mmol/kg (66-330 mg/kg), to male Wistar rats resulted in the excretion of a single mercapturic acid in urine. The methyl esters of three potential mercapturic acid metabolites were synthesized: N-acetyl-S-(2-oxo-2-phenylethyl)-L-cysteine methyl ester (O),N-acetyl-S-(2-hydroxy-1-phenylethyl)-L-cysteine methyl ester (I), and N-acetyl-S-(2-hydroxy-2-phenylethyl)-L-cysteine methyl ester (II). GC/MS analysis showed that the methyl ester of the excreted mercapturic acid was identical with II. Quantitative measurement of II in urine by GLC showed that, after 24 hr, excretion of the mercapturic acid was almost complete and amounted to 41% of the administered dose. At doses higher than 1.00 mmol/kg, the excretion no longer increased. Inhibition of the oxidative pathways by ip injection of 1-phenylimidazole resulted in an excretion decrease of about 40%. (Pre)treatment with diethyl maleate lowered the excretion of mercapturic acid by 30-60%. Glutathione conjugates synthesized from DBPE and styrene oxide were separated by HPLC. Both compounds can produce the same two pairs of diastereomers, viz. (R)- and (S)-(2-hydroxy-1-phenyl-ethyl)glutathione ((R)-1 and (S)-1), and (R)- and (S)-(2-hydroxy-2-phenylethyl)glutathione ((R)-2 and (S)-2). These could be separated in the order (R)-2, (R)-1, (S)-1, and (S)-2 within 20 min. This method was also applied to examine glutathione conjugates excreted in bile after DBPE administration.(ABSTRACT TRUNCATED AT 250 WORDS)     

Bromo(monohydroxy)phenyl mercapturic acids. A new class of mercapturic acids from bromobenzene-treated rats.

Alkaline permethylation and GC/MS analysis of urinary mercapturic acids from rats given bromobenzene yielded several quinone-derived bromodimethoxythioanisole isomers as expected. Unexpectedly, seven bromomonomethoxythioanisole isomers were also observed, suggesting the presence of bromomonohydroxyphenyl mercapturic acids in the urine. Alkaline permethylation of synthetic 4- and 5-bromo-2-hydroxyphenyl mercapturic acid gave 4- and 5-bromo-2-methoxythioanisole, respectively, which were also observed after alkaline permethylation of urine from bromobenzene-treated rats, as was 2-bromo-4-methoxythioanisole. To explore the biosynthetic origin of the bromonohydroxyphenyl mercapturic acids, rats were separately dosed intraperitoneally with synthetic racemic 2-, 3-, or 4-bromophenyl mercapturic acid, or biosynthetic L-(-)-4-bromophenyl mercapturic acid, or a biosynthetic mixture of the 3,4- and 4,3-premercapturic acids from bromobenzene, and their urine (0-24 hr) analyzed by alkaline permethylation and GC/MS. The administered mercapturic acids and premercapturic acids were partly excreted unchanged (60-80% and 24%, respectively), but both gave rise to bromomonohydroxyphenyl mercapturic acids (0.1-5.2% of dose). Results indicated that the latter could be formed by 1) dehydrogenation of premercapturic acids and 2) hydroxylation of mercapturic acids (or their cysteine equivalents).     

Synthesis and antiviral activity of certain 4-substituted and 2,4-disubstituted 7-[(2-hydroxyethoxy)methyl]pyrrolo[2,3-d]pyrimidines

Treatment of the sodium salt of 4-chloro-2-(methylthio)pyrrolo[2,3-d]pyrimidine (2) with (2-acetoxyethoxy)methyl bromide (3) has provided 4-chloro-2-(methylthio)-7[(2-acetoxyethoxy)methyl]pyrrolo[2,3- d]pyrimidine (4). Ammonolysis of 4 at room temperature gave 4-chloro-2-(methylthio)-7-[(2-hydroxyethoxy)methyl]pyrrolo[2,3- d]pyrimidine (5). However, ammonolysis of 5 at 130 degrees C furnished 4-amino-2-(methylthio)-7-[(2-hydroxyethoxy)methyl]-pyrrolo[2,3- d]pyrimidine (6), which on desulfurization with Raney Ni yielded 4-amino-7-[(2-hydroxyethoxy)-methyl]pyrrolo[2,3-d]pyrimidine (7) (acyclic analogue of tubercidin). The oxidation of 6 with m-chloroperbenzoic acid provided the sulfone derivative 8. A nucleophilic displacement of the 2-methylsulfonyl group from 8 with methoxide anion provided 4-amino-2-methoxy-7-[(2-hydroxyethoxy)methyl]pyrrolo[2,3-d]pyrimidine (9). Demethylation of 9 with iodotrimethylsilane gave 4-amino-2-hydroxy-7-[(2-hydroxyethoxy)methyl]pyrrolo[2,3-d]pyrimidine (10). Treatment of 2,4-dichloropyrrolo[2,3-d]pyrimidine (11) with 3 gave the protected acyclic compound 12, which on deacetylation and ammonolysis under controlled reaction conditions gave 2,4-dichloro-7-[(2-hydroxyethoxy)-methyl]pyrrolo[2,3-d]pyrimidine (13) and 4-amino-2-chloro-7-[(2-hydroxyethoxy)methyl]pyrrolo[2,3- d]pyrimidine (14), respectively. The condensation of 2-acetamido-4-chloropyrrolo[2,3-d]pyrimidine (15) with 3 gave the protected acyclic compound 16, which on concomitant deacetylation and ammonolysis with methanolic ammonia at an elevated temperature yielded 2,4-diamino-7-[(2-hydroxyethoxy)methyl]pyrrolo[2,3-d]pyrimidine (17) in moderate yield. In tests involving human cytomegalovirus (HCMV) and herpes simplex virus type 1 (HSV-1), only slight activity and cytotoxicity were observed. The most active compounds (12 and 13) were slightly more active against HCMV than acyclovir, but both compounds were inactive against HSV-1. The activity against HCMV, however, was not well separated from cytotoxicity leading to the conclusion that these compounds did not merit further study.     

Sulphur-containing metabolites of beta-bromostyrene formed by the marmoset, rabbit and rat.

1. The administration of beta-bromostyrene to the rat results in a fall in the level of hepatic glutathione. 2. Marmosets, rabbits and rats dosed with beta-bromostyrene excrete two mercapturic acids. One of these, N-acetyl-S-(2-hydroxy-2-phenyl-1-bromoethyl)-cysteine is readily converted into N-acetyl-S-(1-phenyl-2-bromo-2-ethenyl)-cysteine, the structure of which was established by mass spectrometry. 3. Mass spectrometric evidence suggests that the second mercapturic acid is N-acetyl-S-(1-hydroxy-2-phenylethyl)cysteine. 4. Mandelic acid was detected as a metabolite in all three species.     

Biotransformation of styrene in mice. Stereochemical aspects

Biotransformation of styrene and its toxic metabolite, phenyloxirane (1), in mice in vivo was studied. Mice were treated with single intraperitoneal doses of styrene (400 mg/kg of body weight), and with (R)-, (S)-, or racemic styrene oxide (150 mg/kg of body weight). Profiles of neutral and acidic metabolites were determined by GC/MS. Mandelic acid (3) and two mercapturic acids, N-acetyl-S-(2-hydroxy-2-phenylethyl)cysteine (5) and N-acetyl-S-(2-hydroxy-1-phenylethyl)cysteine (6), were found to be major urinary metabolites of both styrene and phenyloxirane. 1-Phenylethane-1,2-diol (2) was the main neutral metabolite. The rate of excretion of this metabolite, as determined by GC, was 5-10 times lower than that of mandelic acid. Several minor acidic metabolites were also identified. Among them, novel phenolic metabolites, namely, 2-(4-hydroxyphenyl)ethanol (7), (4-hydroxyphenyl)acetic acid (11), and two isomeric hydroxymandelic acids (12), are of toxicological significance. Main stereogenic metabolites were isolated as methyl esters from extracts of pooled acidified urine treated with diazomethane. The mandelic acid that was obtained was converted to diastereomeric Mosher's derivatives prior to analysis by NMR. Mercapturic acids were analyzed directly by (13)C NMR. Pure enantiomers of 1 were metabolized predominantly but not exclusively to corresponding enantiomers of 3. Styrene yielded predominantly (S)-mandelic acid. Fractions of mercapturic acids 5 and 6 isolated from urine amounted to 12-15% of the dose for all compounds that were administered. Conversion to mercapturic acids was highly regio- and stereoselective, yielding predominantly regioisomer 5. Styrene, as compared to racemic phenyloxirane, yielded slightly more diastereomers arising from (S)-1 than from (R)-1. These data can be explained by formation of a moderate excess of the less mutagenic (S)-1 in the metabolic activation of styrene in mice in vivo.     

Identification of new sulfur-containing metabolites of naphthalene in mouse urine

Seven acidic sulfur-containing metabolites were isolated from mouse urine following administration of naphthalene. The metabolites have been identified as (1-hydroxy-1,2-dihydro-2-naphthalenylthio)acetic acid (I), 2-hydroxy-3-(1-hydroxy-1,2-dihydro-2-naphthalenylthio)propanoic acid (II), (1,2,3-trihydroxy-1,2,3,4-tetrahydro-4-naphthalenylthio)acetic acid (III), and N-acetyl-S-(1-hydroxy-1,2-dihydro-2-naphthalenyl)-L-cysteine (IV). The dehydration products of I, II, and IV, namely 1-(naphthalenylthio)acetic acid (V), 2-hydroxy-3-(1-naphthalenylthio)propanoic acid (VI), and N-acetyl-S-(1-naphthalenyl)-L-cysteine (VII), respectively, were also present in several urinary extracts. Nine methylthio derivatives were identified in the neutral extract of urine. These metabolites were the following: 1-methylthionaphthalene, trans-1-hydroxy-2-methylthio-1,2-dihydronaphthalene, two stereoisomeric 1,2,3-trihydroxy-4-methylthio-1,2,3,4-tetrahydronaphthalenes, 1,3-di(methylthio)-2,4-dihydroxy-1,2,3,4-tetrahydronaphthalene, 1,4-di(methylthio)-2,3-dihydroxy-1,2,3,4-tetrahydronaphthalene, two methylthiohydroxy-naphthalenes, and a methylthiodihydroxydihydronaphthalene. Following intraperitoneal administration of N-acetyl-S-(1-hydroxy-1,2-dihydro-2-naphthalenyl)-L-cysteine to mice, the acidic metabolites I, II, and unchanged IV were found. The gas-chromatographic and gas chromatographic-mass spectral properties of the methyl ester-trimethylsilyl derivatives of the acidic sulfur metabolites of naphthalene are presented.     

Identification of the metabolites of 9-nitro-20(S)-camptothecin in rats.

9-Nitro-20(S)-camptothecin is a novel anticancer drug. In this study, metabolites of 9-nitro-20(S)-camptothecin in rats were identified. Rats were dosed with the drug, and the metabolites in the bile were isolated and collected by high-performance liquid chromatography using a gradient elution. By LC/MSn(n =1-3), the biliary metabolites in addition to the unchanged drug were identified as 9-amino-20(S)-camptothecin (M3), 9-acetamido-20(S)-camptothecin (M4), and the glucuronide of 9-hydroxy-20(S)-camptothecin (M1) with the aid of the reference substances and the enzymatic hydrolysis. The accurate mass data for metabolites M2 and M5 were determined by a quadrupole time-of-flight mass spectrometer. Metabolite M5 was suggested to be the glutathione conjugate of 10-hydroxy-20(S)-campto-thecin, and M2 may arise from the loss of glutamic acid from M5. Metabolites M1 and M3 were also found in the urine, and M4 in the feces. Fecal metabolite M7 was confirmed to be 9-hydroxy-20(S)-camptothecin (the aglycon of M1) by being compared with the reference standard. The mercapturic acid conjugate of 10-hydroxy-20(S)-camptothecin (M6) was detected in the urine and feces by LC/MSn     

Glutathione conjugation of chlorobenzylidene malononitriles in vitro and the biotransformation to mercapturic acids in rats

The glutathione conjugation of 2-chloro-, 3-chloro-, 4-chloro- and 2,6-dichlorobenzylidene malononitrile (chloroBMNs) was investigated in vitro. In incubation mixtures containing rat liver cytosol (9000 g), the decrease in the initial amount of glutathione due to the various chloroBMNs ranged from 40 to 60% and occurred both enzymatically and spontaneously at physiological conditions (37°C, pH7.4). 2,6-DichloroBMN, however, depleted glutathione largely spontaneously (38±3%). The steric hindrance of the two chlorosubstituents probably plays an important role during the glutathione-S-transferase catalyzed reaction.The hydrolysis of the chloroBMNs to the corresponding chlorobenzaldehydes and malononitrile was studied in a mixture of buffer pH 7.4 and ethanol. The rate of hydrolysis of 2,6-dichloroBMN was slower than those of the related chloroBMNs. This means that 2,6-dichloroBMN will be the most stable compound in the presence of water.Only IP administration of 2-chloroBMN (CS) to adult male Wistar rats gave enhancement of urinary thioether excretion. A thioether could be isolated and was identified as the N-acetyl-S-[2-chlorobenzyl]-L-cysteine. The quantity of this benzylmercapturic acid in the urine of rats amounted to 4.4% dose (0.07 mmol/kg, n=12).After IP administration of 2-chloro- and 3-chlorobenzaldehyde to rats benzylmercapturic acid excretion in the urine was found to be 7.6 and 1.1% of the dose, respectively. Administration of the related 4-chloro- and 2,6-dichlorobenzaldehyde, however, resulted in _no urinary mercapturic acid excretion.It is very likely that in rats the initial biotransformation of chloroBMNs is mainly hydrolysis to corresponding chlorobenzaldehydes, leading in the case of 3-chloro-, 4-chloro- and 2,6-dichloroBMN to no mercapturic acid excretion in the urine.Nevertheless, 2,6-dichloroBMN will be the most reactive compound with proteins and therefore the best haptene in comparison with the related chloroBMNs.This work was financially supported by a grant from the Dutch Foundation for Medical Research FUNGO, grant no. 13-28-57     

Mercapturic acid urinary metabolites of 3-butene-1,2-diol as in vivo evidence for the formation of hydroxymethylvinyl ketone in mice and rats

3-Butene-1,2-diol (BDD), a major metabolite of 1,3-butadiene (BD), can readily be oxidized to hydroxymethylvinyl ketone (HMVK), a Michael acceptor. In previous studies, 4-(N-acetyl-l-cystein-S-yl)-1,2-dihydroxybutane (DHB), a urinary metabolite of BD that was used to assess human BD exposure, was suggested to be a metabolite of HMVK, but DHB formation from BDD and the formation of the DHB precursor 4-(N-acetyl-l-cystein-S-yl)-1-hydroxy-2-butanone (HB) have not been previously investigated. In the current study, four HMVK-derived mercapturic acids [DHB, HB, 3-(N-acetyl-l-cystein-S-yl)propan-1-ol (POH), and 3-(N-acetyl-l-cystein-S-yl)propanoic acid (PA)] were identified in the urine of mice and rats given BDD (284-2272 micromol/kg, i.p.) based on GC/MS analyses and comparisons with synthetic standards after esterification and silylation of the carboxyl and hydroxyl groups, respectively. The combined amounts of the mercapturic acids excreted after BDD exposure were dose-dependent and were mostly similar between mice and rats given equivalent doses of BDD. The mercapturic acids accounted for a greater fraction of the administered BDD dose as the dose was lowered, suggesting that HMVK formation represents a prominent route for BDD metabolism in both mice and rats. The major mercapturic acid excreted by mice was DHB, whereas rats excreted equivalent amounts of DHB and HB. The levels of POH or PA were significantly lower in both species relative to DHB or HB. The observed species differences in the excretion of DHB and HB were thought to be due to differences in the capacity of mice and rats to reduce HB to DHB.     

In vivo metabolism of isomeric naphthalene oxide glutathione conjugates

We have examined the fate of glutathione conjugates derived from naphthalene metabolism at various dose levels (5-80 mg/kg) in an effort to explore the potential use of urinary mercapturic acids as biomarkers of exposure to naphthalene and as indicators of the activity and stereoselectivity of cytochrome P-450-dependent naphthalene epoxidation. This approach extends previous studies which demonstrated a high degree of stereoselectivity in the formation of (+)-1R,2S-naphthalene oxide from naphthalene in target tissue microsomes (mouse lung), but not in microsomal preparations isolated from nontarget tissues such as mouse liver. To validate the use of mercapturic acids as indicators of epoxide formation in vivo, individual naphthalene oxide glutathione adduct isomers were administered iv to mice, and urinary metabolites were identified and quantified. Mercapturates accounted for 69-75% of the administered dose in the 8-hr urines of animals treated with trans-1-(S)-hydroxy-2-(S)-glutathionyl-1,2-dihydronaphthalene (adduct 1) and 76-84% for trans-1-(R)-hydroxy-2-(R)-glutathionyl-1,2-dihydronaphthalene (adduct 2). Only 39-57% of the dose of trans-1-(R)-glutathionyl-2-(R)-hydroxy-1,2-dihydronaphthalene (adduct 3) administered to mice was excreted as the mercapturic acid derivative; however, two additional metabolites were detected which were not present in the urine of animals treated with adducts 1 or 2. The first metabolite, accounting for 2-4% of the dose of adduct 3, was not identified. The second metabolite, isolated by HPLC and identified by mass spectrometry as (hydroxy-1,2-dihydronaphthalenylthio)pyruvic acid, accounted for 14-25% of the administered dose of adduct 3.(ABSTRACT TRUNCATED AT 250 WORDS)     

Biliary excretion and intestinal metabolism in the intermediary metabolism of pentachlorothioanisole

1. Biliary metabolites from rats dosed with pentachlorothioanisole (PCTA) were characterized by fast atom bombardment mass spectrometry and electron impact mass spectrometry.

2. Most of the biliary metabolites from PCTA were mercapturic acid pathway metabolites of methylsulphinyltetrachlorobenzene (51% of the dose); the remaining characterized biliary metabolites (20%) were mainly methylsulphinyltetrachlorothio-phenols excreted as unknown conjugates.

3. Pathways are proposed for the intermediary metabolism of PCTA to bis-(methylthio)tetrachlorobenzene (bis-MTTCB) involving glutathione conjugation, biliary excretion, intestinal metabolism, and enterohepatic circulation.     

Biliary excretion and intestinal metabolism in the intermediary metabolism of pentachlorothioanisole

1. Biliary metabolites from rats dosed with pentachlorothioanisole (PCTA) were characterized by fast atom bombardment mass spectrometry and electron impact mass spectrometry. 2. Most of the biliary metabolites from PCTA were mercapturic acid pathway metabolites of methylsulphinyltetrachlorobenzene (51% of the dose); the remaining characterized biliary metabolites (20%) were mainly methylsulphinyltetrachlorothiophenols excreted as unknown conjugates. 3. Pathways are proposed for the intermediary metabolism of PCTA to bis-(methylthio)tetrachlorobenzene (bis-MTTCB) involving glutathione conjugation, biliary excretion, intestinal metabolism, and enterohepatic circulation.     

Metabolism of styrene oxide in the rat and guinea pig

The metabolism of styrene oxide has been studied in the rat and guinea pig, with emphasis upon bivalent sulfur metabolites. Methylthio analogs of phenylethylene glycol, with the methylthio group in both possible positions, were found as urinary metabolites in both species. These compounds were present in more than trace amounts. The excretion of 2-hydroxy-1-methylthio-1-phenylethane amounted to about 7% of the administered dose in the guinea pig, and about 2% in the rat, in o-24 hr urine samples. The positional isomer 1-hydroxy-2-methylthio-1-phenylethane was excreted in lesser amounts in both species. Acidic urinary metabolites derived from glutathione conjugates are species dependent. In this study, the only products observed in the rat were the mercapturic acids expected as a result of reaction of the oxide with glutathione. In the guinea pig, the major bivalent sulfur acids were the corresponding mercaptoacetic acids. Other related metabolites included a mercaptolactic and a mercaptopyruvic acid, together with one of the mercapturic acids. These metabolites result from partial acetylation or acetylation/deacetylation of cysteine or cysteinylglycine adducts. The hitherto unobserved dihydrodiol formed via an arene oxide was found as a minor metabolite for both styrene and styrene oxide.     

Total synthesis of norneolignans from<Emphasis Type="Italic">Krameria</Emphasis> species

The total synthesis of nomeolignans isolated from Krameria species, 2-aryl-5-(E)-propenylbenzofurans (5, 11), is described. The key step involves the one-pot reaction for 2-arylbenzofurans (2, 7) from 4-hydroxyphenylacetone with 4'-acetoxy-2-chloro-2-(methylthio)acetophenone (1) and 2-chloro-2-methylthio-(2',4',6'-trimethoxy)acetophenone (6) under Friedel-Crafts reactionconditions.     

Synthesis of 5-chloro-8-hydroxy-6-methoxy-3-pentylisocoumarin

The synthesis of title isocoumarin, the 5-chloro analog of naturally occurring 7-chloro-8-hydroxy-6-methoxy-3-pentylisocoumarin, isolated from Tessmannia densiflora is described. Chlorination of ethyl 2-(2-ethoxy-2-oxoethyl)-4,6-dimethoxybenzoate (2) afforded 3-chloro ester (3) followed by hydrolysis to furnish the 2-(carboxymethyl)-3-chloro-4,6-dimethoxybenzoic acid (4) that was converted to corresponding anhydride (5). Condensation of the latter with hexanoyl chloride in the presence of tetramethylguanidine and triethyl amine afforded 5-chloro-6,8-dimethoxy-3-pentylisocoumarin (6) which upon regioselective demethylation yielded the title isocoumarin (1).     

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)     

Synthese und Eigenschaften des 3-Hydroxy-1,10-dioxo-5,10-dihydro-1H-pyrido[2,1-b]chinazolin-2-carbonitrils

Synthesis and Properties of 3-Hydroxy-1,10-dioxo-5,10-dihydro-1H-pyrido[2,1-b]quinazoline-2-carbonitrile Anthranilic acid reacts with 2-chloro-5-cyano-4-hydroxypyrid-6-one (3) in glacial acetic acid to yield 3-hydroxy-1,10-dioxo-5,10-dihydro-1H-pyrido[2,1-b]quinazoline-2-carbonitrile (4) . When the reaction is carried out in DMF under Ullmann conditions, 2-(dimethylamino)-5-cyano-4-hydroxypyrid-6-one (5) forms as a by-product. The methylation of 3 with diazomethane affords 2-chloro-5-cyano-2-methoxy-N-methylpyrid-6-one (9) and 2-chloro-5-cynao-4,6-dimethoxypyridine (10) . Under similar conditions compound 4 undergoes an esterifying ring cleavage to furnish methyl 2-(5-cyano-4,6-dimethoxypyrid-2-ylamino)benzoate (7) .