Salicylamide Glucuronide Research Articles

An assessment of indomethacin-induced mucosal damage in vivo by measuring the metabolism of salicylamide in rabbit intestine.

Indomethacin-induced mucosal damage was assessed in vivo by measuring salicylamide (SAM) metabolism in rabbit intestine. Intestinal mucosal damage 48 h after oral indomethacin (500 mg/kg) administration was examined using a scanning electron microscope. Duodenal, jejunal and ileal mucosal toxicity was compared with that in controls. Intestinal first-pass metabolism of SAM was studied using in situ intestinal sacs with intact mesenteric venous blood collection. The appearance of both SAM and its metabolites in the mesenteric venous blood was measured following cannulation of the mesenteric vein of the exposed intestine and collecting all venous blood draining from the absorbing region. Following oral pretreatment with indomethacin, the appearance of SAM and SAM glucuronide (SAMG) in the mesenteric venous blood was significantly increased. The concentrations of SAM and SAMG in the blood increased following intraduodenal administration of SAM in vivo in rabbits orally pretreated with indomethacin compared with controls. However, after intravenous administration of SAM, the blood concentration of SAM and SAMG was not increased compared with controls. These findings suggest that the differences in intestinal first-pass metabolism of SAM may be due to the intestinal mucosal damage induced by oral indomethacin pretreatment. The results indicate that the alteration of intestinal first-pass metabolism of a marker compound may be utilized to assess intestinal mucosal damage in vivo.

An assessment of mucosal damage in vivo: effect of oral pretreatment with 5-fluorouracil on the intestinal first-pass metabolism of salicylamide in rabbits.

An assessment of 5-fluorouracil (5-FU)-induced mucosal damage in vivo by measuring the metabolism of salicylamide (SAM) was investigated in rabbit intestine. The mucosal damage in the intestine 48 h after oral administration of 5-FU (30 mg/kg) was examined using a scanning electron microscope. By the oral pretreatment with 5-FU, the morphological changes of jejunal and ileal mucosa were recognized compared with the control. The intestinal first-pass metabolism of SAM was studied using in situ intestinal sacs with complete mesenteric venous blood collection. The appearance of both SAM and its metabolites into the mesenteric venous blood was measured directly by cannulating the mesenteric vein of exposed intestine and collecting all venous blood draining from the absorbing region. Following oral pretreatment with 5-FU, the appearance of SAM glucuronide (SAMG) in the mesenteric venous blood was significantly increased. The increased blood concentration of SAMG following intraduodenal administration of SAM in vivo was observed in rabbits pretreated with 5-FU orally. However, the blood concentration of SAMG after intravenous administration of SAM was not increased compared with the control. These findings suggest that the change in intestinal first-pass metabolism of SAM may be due to the intestinal mucosal damage by oral pretreatment with 5-FU. The alteration of intestinal first-pass metabolism of a marker compound may be utilized for the assessment of intestinal mucosal damage in vivo.

An Assessment of Salicylic Acid-Induced Mucosal Damage in Vivo by Measuring the Metabolism of Salicylamide in Rabbit Intestine.

An assessment of salicylic acid-induced mucosal damage in vivo by measuring the metabolism of salicylamide (SAM) was investigated in rabbit intestine. The intestinal first-pass metabolism of SAM was studied using in situ intestinal sacs with complete mesenteric venous blood collection. The appearance of both SAM and its metabolites into the mesenteric venous blood was measured directly by cannulating the mesenteric vein of exposed intestine and collecting all venous blood draining from the absorbing region. Following oral pretreatment with salicylic acid, the appearance of SAM glucuronide (SAMG) in the mesenteric venous blood was significantly increased compared with the control. The increased blood concentration of SAMG following intraduodenal administration of SAM in vivo was observed in rabbits pretreated with salicylic acid orally. The blood concentration of SAMG after the intravenous administration of SAM was not increased compared with the control. We suggest that the change in the intestinal first-pass metabolism of SAM may be due to the intestinal mucosal damage induced by oral pretreatment with salicylic acid. The measurement of SAM metabolites may be of value in the assessment of intestinal mucosal damage in vivo.

3-Hydroxylation of Salicylamide in Mice

Salicylamide is an important model compound for use in investigations concerning drug disposition. In this study the metabolic fate of salicylamide at high doses was evaluated in male mice using HPLC methodology. The concentrations of salicylamide and its metabolites were determined in urine and in blood at various times after the administration of 2 or 4 mmol kg−1 salicylamide. Salicylamide, gentisamide, and their glucuronide and sulfate conjugates were detected. 2,3‐Dihydroxybenzamide, the 3‐hydroxy metabolite of salicylamide, as well as its glucuronide and sulfate conjugates, were identified and quantitated for the first time by HPLC. 2,3‐Dihydroxybenzamide had previously been detected only as a minor metabolite of salicylamide by paper chromatography. However, in the present study, 18% of the salicylamide metabolites appearing in urine after either dosage of salicylamide were 3‐hydroxylation products. When a previously published HPLC method for salicylamide analysis was used, 2,3‐dihydroxybenzamide glucuronide coeluted with salicylamide glucuronide. The possible formation of 3‐hydroxy metabolites must be evaluated in any study of drug metabolism using salicylamide as a model compound.

Effect of oral pretreatment with indomethacin on the intestinal first-pass metabolism of salicylamide in rabbits.

The effect of oral pretreatment with indomethacin on the intestinal first-pass metabolism of salicylamide (SAM) was studied in rabbits using in situ intestinal sacs with complete mesenteric venous blood collection. The appearance of both SAM and its metabolites into the mesenteric venous blood was measured directly by cannulating the mesenteric vein of exposed rabbit intestine and collecting all venous blood draining from the absorbing region. By oral pretreatment with indomethacin, the total amounts of SAM absorbed in 20 and 120 min were significantly increased compared to the control. These results indicated the alteration of the permeability in the intestinal mucosa. In 20 min, indomethacin pretreatment resulted in increased appearance of SAM and SAM glucuronide in the mesenteric venous blood. In 120 min, increased appearance of SAM and decreased appearance of SAM sulfate were observed, compared to the control. These findings suggested that the change in the intestinal first-pass metabolism of SAM is probably due to the intestinal mucosal damage by oral pretreatment with indomethacin.

High-performance liquid chromatographic method for the quantitation of salicylamide and its metabolites in biological fluids

A high-performance liquid chromatographic (HPLC) assay is described for the determination of salicylamide (SAM) and its metabolites in biological fluids obtained from an in situ rat liver preparation and rat in vivo. SAM and its six metabolites, salicylamide sulphate, salicylamide glucuronide, gentisamide (GAM), gentisamide 2-sulphate, gentisamide 5-sulphate and gentisamide 5-glucuronide, were separated on a μ Bondapak C 18 column with 0.085 M potassium dihydrogenphosphate pH 3.35, as mobile phase and by using a flow gradient. Enriched recovery of SAM and GAM from blood was achieved by extraction followed by a second HPLC procedure with a mobile phase consisting of acetic acid and methanol. Furthermore, the reproducibility and the stability of the samples under assay conditions were investigated.

Depression of salicylamide glucuronidation by bacterial lipopolysaccharide.

The effect of lipopolysaccharide (LPS) on the urinary excretion of salicylamide (SAM) (10 mg/head) given orally to rats was examined 24 h after intraperitoneal administration of LPS (0.5 mg/kg) by the direct analysis of urine samples by high-performance liquid chromatography. The cumulative amount of SAM glucuronide excreted in urine in 4 h was decreased by 40-70% in the LPS-administered rats, while that of SAM sulfate was increased by 7-15% compared with the control in the same rats. The amounts of the conjugates of gentisamide (the hydroxylated metabolite of SAM) excreted in urine were decreased by approx. 70% compared with the control. LPS administration provoked a decrease in the activities of aniline hydroxylase of liver and uridine diphosphate (UDP) -glucuronyltransferase of liver, lung and kidney, but did not change the activities of sulfotransferase of liver or UDP-glucuronyltransferase obtained from the small intestine. The mechanism of the decreased urinary excretion of SAM glucuronide in LPS-administered rats is discussed.

Conjugation of salicylamide in the intestinal wall of dogs and rabbits.

The intestinal conjugation of salicylamide (SAM) the formation of SAM sulfate (SAMS) and SAM glucuronide (SAMG) were investigated in dogs and rabbits by using in situ intestinal sac preparation with mesenteric vein cannulation. SAM, SAMS and SAMG in the mesenteric venous blood, sampled at successive intervals after injection of SAM solutions into the sac, were determined. Total recoveries in 1 h were not significantly different in dogs (71%) and rabbits (64%). Both animals, however, showed a quantitative difference in intestinal conjugation. Sulfation predominated over glucuronidation in dog intestine while the formation of the glucuronide was exclusively high in rabbit intestine. Furthermore, the dose-dependent conjugation of SAM was investigated in dog intestine at four doses, 2, 3, 6 and 10 mg/one animal. Upon increasing the doses from 2 to 10 mg, the amounts of SAMS appearing in the blood remained unchanged, those of SAMG increased about two times and those of free SAM increased about six times, while the total recoveries increased parallel to the doses, suggesting saturation kinetics in the formation step of the two conjugates. Simultaneous computer-fitting of the experimental data for 2 and 10 mg doses to a simplified model, containing Michaelis-Menten kinetics in the formation of the conjugates, was examined and good agreement between the observed data and the theoretical values was obtained.

Determination of Salicylamide and Five Metabolites in Biological Fluids by High-Performance Liquid Chromatography

Two high-performance liquid chromatographic (HPLC) assay procedures were developed for the determination of salicylamide and its metabolites in serum, urine, and saliva. One method involves reverse-phase ion-pair chromatography and UV detection, and is used to determine salicylamide, salicylamide glucuronide, and salicylamide sulfate. The other method, with a different mobile phase and without the ion-pairing reagent, is used to determine gentisamide (the hydroxylated metabolite of salicylamide), gentisamide glucuronide, and gentisamide sulfate. The assays are performed by direct injection of the sample after protein precipitation with ethanol containing the internal standard. Increased sensitivity for the determination of low concentrations of salicylamide is obtained by organic extraction of this drug from serum or saliva. Calibration curves for the conjugates of salicylamide and gentisamide were obtained, in the absence of authentic standards, by partial enzymatic hydrolysis, using the decrease of the conjugate peaks and the concomitant increase of free salicylamide or gentisamide concentrations to determine peak area ratio–concentration relationships. Application of the HPLC assay procedures to the determination of salicylamide excretion products in the urine of three normal human subjects resulted in 98.6% (range: 97.1–100.1%) recovery of a 1-g oral dose of the drug. All five metabolites of salicylamide were found in urine, but only salicylamide glucuronide, salicylamide sulfate, and gentisamide glucuronide were found consistently and in appreciable quantities. Salicylamide and all of its metabolites except gentisamide sulfate were found in human and rat serum, and unconjugated salicylamide as well as gentisamide were found in human saliva.

Biopharmaceutical studies of drugs. III. Serum concentration level of o-ethoxybenzamide and N-(beta-hydroxybutyryl)-p-phenetidine after simultaneous administration to rabbit (author's transl)

The concentrations of o-ethoxybenzamide (EBA), N-(β-hydroxybutyryl)-p-phenetidine (HBP) and metabolites in the rabbit serum were investigated when EBA was administered orally with HBP and EBA or HBP administered alone. 1) The concentration of EBA in the serum increased when 100 mg/kg of EBA was administered concurrently with 28.5, 33.3 and 62.5 mg/kg of HBP as compared with the administration of 100 mg/kg of EBA alone. The co-administration of EBA and HBP also decreased the concentration of salicylamide (SAM) sulfate in the serum. 2) The concentration of HBP in the serum increased when 28.5, 33.3 and 62.5 mg/kg of HBP and 100 mg/kg of EBA were administered together, while that of N-(β-hydroxybutyryl)-p-aminophenol (HBA) glucuronide was decreased. 3) The deethylation of EBA was inhibited competitively by the addition of HBP, and also the deethylation of HBP inhibited competitively by the addition of EBA in 9000×g supernatant fraction of the rabbit liver. 4) The area under the serum concentration-time curve (AUC, 0-8 hr) of SAM sulfate approached a limiting value when 300 mg/kg of EBA was administered orally, while the AUC values of SAM glucuronide, EBA, SAM total and SAM total plus EBA increased with increasing doses of EBA. 5) The deethylation of HBP approached a limiting value when 300 mg/kg of HBP was administered orally, while the AUC values of HBP and HBA total plus HBP total (HBP plus HBP glucuronide) increased with increasing doses of HBP.

Liver enzyme induction caused by antipyrine

Among the substances which have an inducing effect on the enzymatic activity of the liver, antipyrine has been lately used therapeutically, in pregnant women in order to prevent neonatal hyperbilirubinemia. In the present paper we determined liver microsomal enzyme activity caused by antipyrine, by studying various hapatic enzyme systems responsible for bilirubin as well as drug metabolism. Twelve (12) volunteers participated in the study, aged 29-36 years. Three hundred (300) mg of antipyrine per day were administered p.o., to each one of them for a period of 10 days. Urinary D-glucaric acid (D-GA) excretion, serum gamma-glutamyl-transpertidase (γ -GT) urinary salicylamide glucuronide formation rate and saliva half-life of antipyrine. No statistically significant difference was observed in serum γ -GT and saliva half-life of antipyrine. On the contrary, a very significant statistical difference (P < 0, 001) was found in urinary D-GA excretion and salicylamide glucuronide formation. The above results show that the administration of antipyrine caused an increase of the hepatic enzyme activity, as far as the enzymes involved in the metabolism of glucuronic acid, as well as those involved in the formation of salicylamide glucuronide are concerned. It is well known these enzyme systems take part in the metabolism of bilirubin.

医薬品の生物薬剤学的研究(第1報)経口投与時におけるPentazocineの血清中濃度におよぼすSalicylamideの併用効果

The serum concentration of unchanged pentazocine (PA) was investigated in rabbits when PA was administered concurrently with acetaminophen, salicylamide (SAM), salicylic acid, acetylsalicylic acid, or p-aminosalicylic acid. The serum concentration of unchanged PA increased when 20 mg/kg PA was administered concurrently with 80 mg/kg SAM as compared with the administration of 20 mg/kg PA alone, but no similar results were obtained with the other drugs. The co-administration of PA and SAM also decreased serum and urinary PA glucuronide. On the other hand, when 80 mg/kg SAM and 375.6 mg/kg PA were administered together, the serum and urinary concentration of unchanged SAM was increased, while SAM glucuronide was decreased. We posit that the effect noted in the simultaneous 20 mg/kg PA and 80 mg/kg SAM administration experiments is due to competitive inhibition of glucuronidation.

Age-related differences in salicylamide and acetaminophen conjugation in man

Following a concomitant oral dose of salicylamide and acetaminophen (5 mg/kg of each) the urinary excretion of glucuronide and sulfate conjugates of the drugs were followed in children (ages seven to ten years) and adults. No significant difference were observed between the two age groups in the half-lives for appearance of salicylamide conjugates in urine. Age-related changes in the metabolic pathways, however, were observed. The mean percentage of dose excreted as salicylamide sulfate was significantly higher in children (78%) than in adults (36%). In contrast, salicylamide glucuronide was the major excretory product in adults. Similar age-related differences were observed for acetaminophen conjugation. Pharmacokinetic analysis indicated that the deficiency in glucuronide conjugation of these drugs in children is accompanied by a higher rate of sulfate formation.

Salycylamide glucuronide formation in liver disease and its change by drugs.

Salicylamide glucuronide (SAMG) in 0-6 and 6-12 hours-urine specimens was determined after oral administration of salicylamide in 7 normal volunteers (NV), in 51 cases of various liver diseases and hyperbilirubinemias, and in 19 cases after drug administration, to predict the in vivo drug metabolism in man and its change by drugs. Maximal glucuronide formation was obtained by 1.0 g of salicylamide administered to NV; thus, this dosage was used in the present study. SAMG as percent of total salicylamide, the percent of SAMG, from 0-6 hours-urine specimens was high and constant in NV (71.3 +/- 8.3 (Mean +/- S.D.)). 0-0.08% of the total salicylamide was confirmed as free salicylamide in 0-12 hours-urine specimens of NV. The percent of SAMG of 0-6 hours-urine specimens was 57.2 +/- 8.6 in acute hepatitis, 66.6 +/- 10.9 in chronic hepatitis, and 48.6 +/- 10.7 in liver cirrhosis (mean +/- S.D.). Free salicylamide increased slightly in liver diseases. Serum bilirubin levels tended to be inversely correlated with the percent of SAMG. In most cases of Gilbert's syndrome, the percent of SAMG remained at a normal level. The percent of SAMG in cases with unconjugated hyperbilirubinemias of other geneses were almost within normal limits. Bucolome and phenobarbital increased the percent of SAMG in patients with various liver diseases. After rifampicin or phenytoin administration, the percent of SAMG of the patients with lung tuberculosis or epilepsy did not surpass that of NV.

Variations in salicylamide glucuronide formation in normal and in G-6-PD-deficient children

Variations in salicylamide glucuronide formation in normal and in G-6-PD-deficient children

The effect of gentamicin on liver glucuronyl transferase

Summary The effect of gentamicin on the liverenzyme glucuronyl transferase activity in vitro and on salicylamide glucuronide excretion in vivo was investigated. Little or no inhibition of the enzyme activity was observed after the addition of gentamicin to the substrate in concentrations similar to those observed in blood after the therapeutic administration of this agent. Only in concentrations greater than those produced by therapeutic doses was significant inhibition of the enzyme glucuronyl transferase activity observed. A significant decrease in urinary excretion of salicylamide glucuronide was observed after the administration of gentamicin. No reduction in the binding capacity of a 4 Gm. per cent crystalline human albumin solution for HBABA was observed in any of the drug dilutions.

The influence of pyrogen-induced fever on salicylamide metabolism in man.

Salicylamide is metabolized in man by biotransformation to salicylamide glucuronide, salicylamide sulfate, and gentisamide glucuronide. The metabolites are quantitatively and rapidly excreted in urine. Study of the metabolism of this drug in volunteers during episodes of pyrogen-induced fever shows a significant reduction in the half-life (t(1/2)) of the excretion of the drug metabolites. The proportion of the drug transformed to its major metabolite, salicylamide glucuronide, is significantly reduced by fever, with concomitant increase in the proportion of one or both of the other metabolites. Thus, the pattern of urinary metabolites of salicylamide is altered. The shortened t(1/2) of the metabolite excretion is probably due to increased hepatic and renal blood flow known to accompany pyrogen-induced fever. This concept was supported by the observation that when two subjects were placed in a high-temperature environmental chamber, a condition in which hepatic and renal blood flows are known to diminish, the t(1/2) of salicylamide metabolite excretion actually increased. No simple explanation exists to explain the changed metabolite pattern noted during febrile periods. It is most likely to be due to complex interactions between the direct or indirect effects of the pyrogens and the factors affecting the hepatic biotransformation of drugs.

Drug Biotransformation Interactions in Man III: Acetaminophen and Salicylamide

Acetaminophen (1 and 2 g.), salicylamide (1 g.), and 1 g. acetaminophen followed 1.5 hr. later by 1 or 2 g. salicylamide were administered in aqueous solutions to healthy human subjects. Urine was collected every 15-30 min. for the first 4-5 hr. and then at longer intervals. The urine samples were analyzed for acetaminophen, acetaminophen sulfate, acetaminophen glucuronide, salicylamide, salicylamide sulfate, and salicylamide glucuronide. The excretion rates of acetaminophen sulfate and acetaminophen glucuronide decreased immediately after administration of salicylamide. Acetaminophen decreased the formation of salicylamide sulfate. Coadministration of L-cysteine (a source of sulfate) prevented this effect and also increased the excretion of acetaminophen sulfate. The results indicate a competitive inhibition in the formation of acetaminophen sulfate and salicylamide sulfate and suggest also a competitive inhibition in the formation of acetaminophen glucuronide and salicylamide glucuronide.

Intestinal Drug Absorption and Metabolism II: Kinetic Aspects of Intestinal Glucuronide Conjugation

Mechanisms of intestinal salicylamide glucuronide formation and transport were studied in the rabbit with in vitro, cannulated everted intestines and in vivo perfused and closed loops with complete mesenteric venous blood collection. Both experimental techniques indicate that glucuronide formation is capacity limited when the lumen concentration exceeds 10−3M. The glucuronide‐to‐free drug ratio appearing in mesenteric blood (in vivo) and serosal fluid (in vitro) decreases with increasing lumen concentrations. Appearance of glucuronide across the basal barrier is limited by the transport step rather than the rate of glucuronide synthesis leading to accumulation of glucuronide in the tissue compartment. Transport of glucuronide appears to be a simple first‐order diffusion process into the lumen contents as well as the mesenteric blood. Some implications of these findings in pharmacokinetic studies and dosage form design are discussed.

Drug Biotransformation Interactions in Man I: Mutual Inhibition in Glucuronide Formation of Salicylic Acid and Salicylamide in Man

Co-administration of salicylamide (SAM) and sodium salicylate to healthy adult humans resulted in a pronounced decrease in the formation of the glucuronides of these two drugs. This is thought to be due to a limited capacity for glucuronide conjugation in man and/or to the uncoupling action of salicylate on oxidative phosphorylation. Salicylate may inhibit also the formation of SAM sulfate; it has no effect on the excretion rate constants of SAM glucuronide and SAM sulfate in the doses used in the investigation. The results of this study indicate that concomitant use of certain drugs which are readily available to the public may lead to appreciable interactions in biotransformation and to enhanced pharmacologic effects.