Comparison of 5-aminosalicylic acid and N-acetylaminosalicylic acid uptake by the isolated human colonic epithelial cell.

Abstract

Isolated human colonic epithelial cell suspensions were incubated with either 0.1 mM 5-aminosalicylic acid (5-ASA) or 0.1 mM acetylaminosalicylic acid (Ac-ASA) for up to two hours. Intra-and extracellular 5-ASA and Ac-ASA were measured by high performance liquid chromatography. Mean 5-ASA uptake in one hour was 160-5 nmollg dry weight, compared with an Ac-ASA uptake of only 5.75 nmollg dry weight. No unchanged 5-ASA was detected inside the cell. Repeated washing had no effect on the intracellular Ac-ASA concentration. This discrepancy in drug uptake may explain why Ac-ASA seems to be ineffective when given to patients with ulcerative colitis. (Gut 1992; 33: 1343-1347 5-Aminosalicylic acid (5-ASA) is the active ingredient of sulphasalazine in ulcerative colitis,' and is thought to work by a local mechanism. The absorption of 5-ASA by the colon is poor, resulting in very low serum and urine concentrations2 and high faecal concentrations.3 The main metabolite of 5-ASA is N-acetyl ASA (Ac-ASA), and the drug is found predominantly in this form in the body. The site of acetylation of 5-ASA was widely assumed to be the liver, but it has recently been shown that the human colonic epithelial cell is capable of acetylating 5-ASA, and that N-acetyltransferase activity is present in the cytosol.4 In addition, it has been shown that Ac-ASA is the form of the drug predominantly found in the rectal mucosa of patients taking oral sulphasalazine.5It has been suggested that this acetylation process may inactivate the drug, and the results of clinical treatment trials of Ac-ASA have been conflicting,'-with the drug being ineffective in two out of three trials. One possible explanation for this discrepancy might be that Ac-ASA is less well taken up by the colonic mucosa, and there is indirect evidence to suggest that this so.' 1" The aim of this study was to compare the uptake of 5-ASA with that of Ac-ASA into the isolated human colonic epithelial cell. Aliquots (2 ml) ofcell suspension were incubated, in duplicate or triplicate, with 01 mM 5-ASA and 5 mM glucose for one hour at 37°C in a shaking water bath (at 70-80 oscillations/ minute). The incubation was terminated by rapid centrifugation, following which the supernatant was analysed for the presence of Ac-ASA by high performance liquid chromatography (HPLC). Intracellular 5-ASA and Ac-ASA were measured by resuspending the cell pellet in 1 ml 0.05 M phosphate buffer, and disrupting it ultrasonically by six, 5 second bursts using an ultrasonicator (MSE). Control experiments were performed to assess the ability of the KrebsHenseleit saline, and also the supernatant of a cell suspension, to acetylate 5-ASA. It has previously been shown that there is no acetyltransferase enzyme activity in the epithelial cell membrane.4 The intracellular and extracellular production of Ac-ASA was therefore deemed to be the result of 5-ASA uptake and subsequent metabolism -that is, equivalent to 5-ASA uptake.

Identification and Functional Characterization of Arylamine N -Acetyltransferases in Eubacteria: Evidence for Highly Selective Acetylation of 5-Aminosalicylic Acid

“…The aminosalicylate 5-ASA is one of the most effective treatments for inflammatory bowel diseases in humans (50). It is mainly metabolized into N-acetyl-5-ASA, which is thought to be therapeutically inactive (35). We detected N-acetylation activity with 5-ASA in human feces, with a 1-to 10-fold interindividual variability in enzyme velocities (see Results).…”

“…N-Acetyl-5-ASA may be unstable in the intestine and/or hydrolyzed by nonspecific amidases to regenerate 5-ASA. However, administered acetyl-5-ASA was reported not to be deacetylated, and acetyl-5-ASA is considered more stable than 5-ASA (35). Thus, we suggest that the bacterial acetylation of 5-ASA could modulate the concentration equilibrium between the two forms, promoting the acetylated form in the gut.…”

Acute murine colitis reduces colonic 5-aminosalicylic acid metabolism by regulation of N-acetyltransferase-2

“…In both cell-free and intact-cell assays, 5-ASA is more potent as an antioxidant than Ac-5-ASA (1,4,26,58), but the two compounds have been reported to have similar potency at inhibiting lipoxygenase (5), eicosanoid production (21), interferon-␥ action (13,14), lipid peroxidation (67), and scavenging HOCl (63). While topical application of Ac-5-ASA to the colon may be of limited or no therapeutic value (61,66), negative findings may be explained by the slow uptake of Ac-5-ASA vs. 5-ASA by colonocytes (37). Conceptually, unless both 5-ASA and Ac-5-ASA are equally effective as anti-inflammatory drugs, changes in NAT activity are predicted to change the efficacy of 5-ASA therapy.…”

Salicylates used in inflammatory bowel disease and colchicine impair interferon-gamma induced HLA-DR expression.

“…46 There are no studies of the therapeutic effect of N-acetyl 5ASA in maintaining remission. N-acetyl 4ASA is the major metabolite of 4ASA47 but has not been used in ulcerative colitis or Crohn's disease.…”