Jerome C. Pekas

Naphthol metabolism: Glucuronide conjugation and transport by the rat intestine in vitro☆

Abstract [ 14 C]Naphthol was converted to water-soluble 14 C-labeled metabolites by everted sacs of rat small intestine. The extent of conversion to metabolites and release to the bathing medium was greater for sacs from cranial (80%) than for sacs from caudal (60%) small intestine. The concentration of metabolites was always greater in serosal than in mucosal fluids. This concentration gradient increased from the cranial to the caudal end of the intestine. Net 14 C transport to the serosal fluid was entirely as metabolites and was associated with the volume of water transported. The principal metabolites were glucuronide conjugates of naphthol which were hydrolyzed by β- d -glucuronidase. Naphthyl-β- d -glucuronide appeared to be the principal metabolite in the recovered mucosal and serosal fluids and tissues. The quantities of metabolites synthesized by sacs from the various regions of the intestine were not notably different when either pH 7.4 or pH 6.5 medium was used. The tissue concentration of 14 C and quantity of metabolite transferred to serosal fluid in the pH 7.4 and pH 6.5 media were slightly different. The mechanism for the apparent uphill transport of metabolites was not apparent. The results have been compared to those from previous investigations where [ 14 C-naphthyl] N -methylcarbamate was studied and provide greater insight into the intestinal metabolism of this naphthol-containing pesticide.

Intestinal hydrolysis, metabolism and transport of a pesticidal carbamate in pH 6.5 medium☆

Abstract The metabolism and transport of (1-naphthyl-1-14C) N-methylcarbamate by everted sacs of rat small intestine in pH 6.5 medium were remarkably different from those reported for a pH 7.4 medium. Active liberation of naphthol from the carbamate by the intestine was demonstrated, and the rate of hydrolysis was greater in cranial than in mid or caudal intestine. Water-soluble 14C-metabolites were also synthesized more rapidly in cranial than in mid or caudal intestine. The major 14C-metabolite cochromatographed with naphthyl glucuronide, previously found to be the major metabolite in pH 7.4 medium. Net mucosal-to-serosal transfer of 14C occurred throughout the intestine but was most rapid in the cranial section. The serosal: mucosal concentration gradient of the water-soluble 14C-metabolites was greatest in the caudal section.

A method for direct gastric feeding and the effect on voluntary ingestion in young swine

The purpose of the study was to determine the effect of direct administration of diet into the stomach on total alimentation (quantity consumed voluntarily plus the quantity administered by young pigs. Ten cross-bred pigs, 82 days of age, were assigned to five treatments (two replicates of five pigs) consisting of a control (C) and four gastric fistulated (GF) pigs. The GF pigs were fed daily via the fistula 0, 20, 40 and 60% of the dry matter consumed voluntarily by the control pigs (17h per day). Relative to the control pigs, the GF-0, GF-20, GF-40 and GF-60 pigs, respectively, ate 102.4, 99.9, 91.7 and 97.3% the quantity of diet and gained 116.7, 111.1, 113.0 and 114.8% body weight; thus, the surgery, device and procedures were not detrimental. Voluntary consumption was physiologically regulated rapidly with good precision to adjust for the gastric fistula administered diet. Correlation and regression analysis of daily voluntary ingestion (y) versus gastric administration (x) of the six GF-20, -40 and -60 pigs gave a correlation coefficient, r = -0.926 (P less than 0.01), and regression coefficient, b= -1.089 (0.10 greater than P greater than 0.05). The statistical coefficients indicate that voluntary ingestion was suppressed somewhat more than the quantity administered into the stomach.

Gastric absorption of a pesticide (1-naphthyl N-methylcarbamate) in the fasted rat☆

Abstract The gastric absorption of [1-naphthyl-1- 14 C] N -methylcarbamate (radiolabeled carbaryl) administered intragastrically (7.5 μmoles/kg body wt) to fasted, anesthetized female rats was investigated by measurement of 14 C absorption and by identification of the radiolabeled constituents in the portal blood. At 22 and 67 min after dosing, 52.6 ± 14.1% ( n = 3) and 81.7 ± 15.7% ( n = 3), respectively, of the 14 C was absorbed from the stomachs (ligated pylori) of rats with intact portal circulation. In another experiment, 69.1 ± 25.9% ( n = 3) of the 14 C was absorbed from the rat stomachs (ligated pylorus and esophagus) during 66.0 ± 1.0 min of total portal blood collection (via an in vivo perfusion technique). The radiolabeled material in the collected portal blood accounted for 97.2 ± 1.9% ( n = 3) of the 14 C absorbed from rat stomachs. On the basis of thin-layer chromatography, gas chromatography, and infrared spectroscopy, 89.3% of the radiolabeled material in the collected portal blood was [1-naphthyl-1- 14 C] N -methylcarbamate.

Gastrointestinal Metabolism and Transport of Pesticidal Carbamates

Published evidence pertaining to gastrointestinal absorption and metabolism of pesticidal carbamates will be reviewed. The major body of evidence will come form investigations conducted by the author on N-methylcarbamate pesticides with rats; few investigations directly involving the gastrointestinal system have been reported by other research groups. Results derived from in vitro and isolated in vivo preparations show that pesticidal carbamates are absorbed and metabolized by gastrointestinal tissues. Numerous gastrointestinal metabolites have been separated; some have been identified and their behavior in the gut characterized; these data will be included in the review to the extent they have been published. The absorptive and metabolic capacities of the gastrointestinal system vary from one region to another. The author draws from evidence obtained from in vitro and in vivo studies and proposes the events that occur between carbamate ingestion and the appearance of the compound and/or its metabolites in the portal blood or mesenteric lymph. In this endeavor, the author acknowledges that superimposition of the metabolic precursory substrate-product sequence on the sequential physiologic steps from ingestion to defecation produces a complex situation which is difficult if not impossible to study meaningfully by investigation of one tissue component at a time (i.e., epithelial uptake, epithelial metabolism, mucosal to serosal transport, etc.).Published evidence pertaining to gastrointestinal absorption and metabolism of pesticidal carbamates will be reviewed. The major body of evidence will come form investigations conducted by the author on N-methylcarbamate pesticides with rats; few investigations directly involving the gastrointestinal system have been reported by other research groups. Results derived from in vitro and isolated in vivo preparations show that pesticidal carbamates are absorbed and metabolized by gastrointestinal tissues. Numerous gastrointestinal metabolites have been separated; some have been identified and their behavior in the gut characterized; these data will be included in the review to the extent they have been published. The absorptive and metabolic capacities of the gastrointestinal system vary from one region to another. The author draws from evidence obtained from in vitro and in vivo studies and proposes the events that occur between carbamate ingestion and the appearance of the compound and/or its metabolites in the portal blood or mesenteric lymph. In this endeavor, the author acknowledges that superimposition of the metabolic precursory substrate-product sequence on the sequential physiologic steps from ingestion to defecation produces a complex situation which is difficult if not impossible to study meaningfully by investigation of one tissue component at a time (i.e., epithelial uptake, epithelial metabolism, mucosal to serosal transport, etc.).

Further metabolism of naphthyl N-methylcarbamate (carbaryl) by the intestine

Abstract Metabolism of the insecticide (14C-naphthyl) N-methylcarbamate by the rat small intestine was observed in vitro. Volatile derivatives of the intact 14C-metabolites or of the 14C-labeled hydrolysis products (14C-lipophiles) from metabolites were analyzed by gas-liquid chromatography (glc) interfaced to a mass spectrometer (glc/ms). Of the 1.75 mg of substrate equivalents of 14C-labeled material recovered, approximately 43% was the unmetabolized 14C-carbamate; [1-14C]naphthyl glucuronide and [14C]hydroxy-1-naphthyl glucuronide accounted for about 23%. The rest (about 34%; 0.59 mg-eq) was distributed among numerous identified and unidentified 14C-lipophiles derived from polar 14C-metabolites. The following 14C-lipophiles were identified: 1-naphthyl N-methylcarbamate, hydroxy-1-naphthyl N-methylcarbamate, 1-naphthol, hydroxy-1-naphthol, dihydroxy-1-naphthol (tentative), and dihydro-dihydroxy-1-naphthol. The evidence indicates that the same 14C-lipophile was released by hydrolysis of several 14C-metabolites; presumably, the hydrophilic groups were cleaved from the lipophile of these metabolites unintentionally during chromatgraphy and preparation of volatile derivatives. Glucuronic acid was the only hydrophile identified. Thus, the small intestine was capable of metabolism of this carbamate involving oxidations of the naphthol moiety, hydrolysis of the carbamate esters, and conjugations of the carbamate and of lipophilic products derived from the carbamate. Trimethylsilyl derivatives were found to be useful for glc and glc/ms analyses of the metabolites of this carbamate.

Retention of lipophilic compounds on laboratory tubing

Abstract 14 C-Labeled pesticidal carbamates were retained in polyvinyl chloride and latex components of intestinal perfusion apparatus approximately proportionate to their rate of disappearance from the perfusate during absorption experiments. Simulated control perfusions, equilibration of the perfusion system with the perfusate and analysis of the recovered substrate established that such a problem had been encountered. Precautions should be taken when-ever compounds with a high lipid partition coefficient are exposed to polymeric materials in an aqueous medium at very low concentrations.

Biliary secretion of 14C and identification of 5,6-dihydro-5,6-dihydroxycarbaryl glucuronide as a biliary metabolite of [14C]carbaryl in the rat☆☆☆

Abstract The biliary secretion of 14 C was observed in conscious, bile-fistulated rats given single oral doses of [ 14 C]carbaryl (1.5, 30, and 300 mg/kg). Over 94% of the 14 C was absorbed after 12 hr. From 15 to 46% of the 14 C was secreted in bile, 10–40% in urine, and less than 1% in feces 12 hr after dosing. Three metabolites were isolated from bile and identified by mass and/or NMR spectrometric methods. These metabolites were: 5,6-dihydro-5,6-dihydroxycarbaryl glucuronide (12–18% of the biliary 14 C), a conjugate(s) of carbaryl (12% of the biliary 14 C), and conjugated isomers of hydroxy-carbaryl (2% of the biliary 14 C). The majority of the biliary 14 C remains to be identified.

Effect of gastric-feeding on feed consumption, growth, organ size, and body composition of swine.

The effect of direct administration to the stomach (via gastric fistula) of 50% (GF-50) or 100% (GF-100) of the dry matter ingested voluntarily by control pigs (C and GF-0) on voluntary feed ingestion of young rapidly growing pigs was measured. Fistula administrations were intermittent (four consecutive days of each seven-day cycle) for a sustained period (eight cycles; 56 days). Total feed intake and body weight data were collected on an additional two days (58 days total) from the end of the eighth cycle to killing of the animals. Total alimentation (GF plus voluntary intake) by the GF-50 and GF-100 pigs was not affected for the entire 58-day feeding period or for each seven-day cycle but was perturbed profoundly within each cycle (three days of no GF vs. four days of GF administrations). Daily deviations from expected total alimentation (average daily ingestion of four control pigs) on a body weight basis (g·kg Bw−1.day−1) were analysed. The results clearly demonstrated that total alimentation of the GF100 pigs surpassed expected values during the four days of GF administration but fell below expected during the other three days of each cycle. Data from the GF-50 pigs were similar but of a lower magnitude. The deviations are the result of inadequate adjustment of voluntary intake for the quantity of GF administered feed; total intake was excessive during GF days and insufficient during days of only voluntary intake. It is remarkable that total alimentation over the seven days of each cycle and over the eight cycles was not significantly different than the controls, however, The rate and efficiency of growth, gastrointestinal and other organ size, and carcass components were not significantly affected by the repetitious perturbances of total alimentation caused by the GF-50 and GF-100 treatments.

A total-continuous portal vein fistula in gastrointestinal and toxicology research☆

Abstract A method for the total collection of portal vein blood over extended periods has been developed. The method has been applied in gastrointestinal toxicology research involving rats. The method facilitated quantitative recovery of portal vein blood for 120 min for measurement and identification of absorbed radiolabeled components derived from a toxic lipphilic pesticide placed in the lumen of the small intestine. The method can be applied to any substance—nutrient, anutrient, xenobiotic chemical, or endocrine factor—absorbed into the portal vein from the gastrointestinal tract. The method (a) provides for collection of compounds absorbed from the gastrointestinal tract before distribution to nondigestive tissues (i.e., liver), (b) allows measurement of the compounds absorbed without the need to monitor flow rate, and (c) facilitates accumulation of sufficient material from the total portal circulation for analyses when submicrogram quantities are absorbed. The latter two features are important but impossible to attain with small samples of portal blood and are crucial in research with minute quantities of substrate. These features become requirements in metabolism and toxicology research. The method involves replacement of the portal blood with a suspension of perfluorohydrocarbons to substitute for the major functions of natural blood. The total-continuous portal vein fistula makes it possible to investigate metabolic and transport phenomena in live animals which previously could be explored only by in vitro methods. The method may be applied to conscious animals with further refinements.