Synthesis of sulpha drug based hydroxytriazene derivatives: Anti-diabetic, antioxidant, anti-inflammatory activity and their molecular docking studies.

Herein, we report synthesis, characterization, anti-diabetic, anti-inflammatory and anti-oxidant activities of hydroxytriazenes derived from sulpha drugs, namely sulphanilamide, sulphadiazine, sulphapyridine and sulphamethazine. Before biological screening of the compounds, theoretical prediction using PASS was done which indicates probable activities ranging from Pa (probable activity) values 65-98% for anti-inflammatory activity. As per the predication, experimental validation of some of the predicted activities particularly anti-diabetic, anti-inflammatory and anti-oxidant was done. Anti-diabetic activities have been screened using two methods namely α-amylase and α-glucosidase inhibition method and IC50 values were ranging from 66 to 260 and 148 to 401 µg/mL, while for standard drug acarbose the values were 12 µg/mL and 70 µg/mL, respectively. Docking studies have also been done for antidiabetic target pancreatic alpha amylase. The molecular docking studies in α-amylase enzyme reveal that the middle phenyl ring of all the compounds mainly occupies in the small hydrophobic pocket formed by the Ala198, Trp58, Leu162, Leu165 and Ile235 residues and sulphonamide moiety establish H-bond interaction by two water molecules. Further, anti-inflammatory activity has been evaluated using carrageenan induced paw-edema method and results indicate excellent anti-inflammatory activity by hydroxytriazenes (71 to 97%) and standard drug diclofenac 94% after 4 h of treatment. Moreover, antioxidant effect of the compounds was tested using DPPH and ABTS methods. All the compounds displayed good results (24-488 µg/mL) against ABTS radical and many compounds are more active than ascorbic acid (69 µg/mL) while all other compounds showed moderate activity against DPPH radical (292-774 µg/mL) and ascorbic acid (29 µg/mL). Thus, the studies reveal potential of sulfa drug based hydroxytriazenes as candidates for antidiabetic, anti-inflammatory and antioxidant activities which have been experimentally validated.

An Unusual Type of Kidney Stone.

A very rare case of acetylsulfapyridine nephrolithiasis is presented in a 54-year-old female patient who had been prescribed sulfasalazine (6 x 500 mg/day) because of psoriatic arthritis for the last 9 years. The patient's renal function was only slightly impaired. Reflectance infrared spectroscopy and gas chromatography-mass spectrometry allowed the identification of the chemical nature of the stone. As acetylsulfapyridine is a metabolite of sulfasalazine, administration of the latter drug was the cause of the nephrolithiasis.

Biodegradation studies of N4-acetylsulfapyridine and N4-acetylsulfamethazine in environmental water by applying mass spectrometry techniques.

This work evaluates the biodegradation of N(4)-acetylsulfapyridine (AcSPY) and N(4)-acetylsulfamethazine (AcSMZ), metabolites of two of the most commonly used sulfonamides (SAs) in human and veterinary medicine, respectively. Aerobic transformation in effluent wastewater was simulated using aerated fixed-bed bioreactors. No visible changes in concentration were observed in the AcSMZ reactor after 90 days, whereas AcSPY was fully degraded after 32 days of experiment. It was also demonstrated that AcSPY transformed back to its parent compound sulfapyridine (SPY). The environmental presence of these two metabolites in wastewater effluent had been previously investigated and confirmed, together with three more SA acetylated metabolites and their corresponding parent compounds, in 18 different wastewater treatment plants in Hesse (Germany). Sulfamethoxazole (SMX) and SPY were the two SAs detected most frequently (90% and 89% of the samples, respectively) and in the highest concentrations (682 ng L(-1) for SMX and 532 ng L(-1) for SPY). To conclude, hazard quotients were calculated whenever toxicity data were available. None of the SAs studied posed an environmental risk.

Phenotyping with sulfasalazine - time dependence and relation to NAT2 pharmacogenetics.

OBJECTIVE: N-acetyltransferase 2 (NAT2) genotype-phenotype relation with sulfasalazine as probe drug by means of detailed genotype analysis and kinetic data evaluation. BACKGROUND: Though phenotype analysis of sulfasalazine metabolism has been described before, genotype investigations in this regard are scarce. The influence of different single point mutations on the metabolism of the sulfasalazine metabolite sulfapyridine (SP) should give more insight into the functionality of different alleles especially with those still under discussion. METHODS: In two bioavailability studies performed under comparable conditions with 24 healthy subjects of both genders equally distributed, plasma levels of SP and acetylsulfapyridine (Ac-SP) were determined after oral intake of enteric coated formulations of sulfasalazine (500 mg and 1,000 mg, respectively). The resulting metabolic ratios were calculated. NAT2 genotype was analyzed in parallel for all subjects deducing haplotype set as well as putative functional phenotype as (homozygous or heterozygous) rapid acetylator (RA) or slow acetylator (SA) and correlated with the PK results. RESULTS AND DISCUSSION: RA genotype in the overall study population was seen with 45.5% (including 6.8% homozygous wildtype *4/*4) and SA genotype with 54.5%. Compared to RA genotype, apparent terminal elimination half-life of SP as well as of Ac-SP was prolonged in the SA genotype population, C(max) and AUC values of SP were higher whereas average C(max) value of Ac-SP was lower (with AUC only some tendency to lower values). In general, phenotype-genotype correlation was good with only few exceptions. Strongest functional effect on enzyme activity was noticed in slow acetylators carrying the 341T > C mutation, followed by 590G > A mutation whereas the influence of 857G > A was considerably less pronounced. Homozygous 803A > G mutation (lysine > arginine shift) did not reveal enzyme activity reduction.

Effects of NAT2 polymorphism on SASP pharmacokinetics in Chinese population.

BACKGROUND: Sulfasalazine (SASP) pharmacologic actions are widely applied in clinical therapy. The role of N-Acetyltransferase 2 (NAT2) in the pharmacokinetics of SASP and its metabolites has not been clarified. We investigated the effects of genetic polymorphism of NAT2 on pharmacokinetic profiles of SASP and its two metabolites, sulfapyridine (SP) and N-acetylsufapyridine (AcSP). METHODS: Eighteen subjects were recruited and divided into 3 groups by NAT2 genotype: wild type (w/w), heterozygous variant (w/m), homozygous variant (m/m). After taking 1000mg SASP tablets, the plasma concentrations of SASP, SP and AcSP were measured with HPLC method and pharmacokinetic parameters were calculated by using the computing program 3P97. RESULTS: The AUC(0)(-)(72) and Cmax of SP in m/m subjects were significantly higher than those in w/m and w/w subjects, with the values of 172.57+/-49.42, 103.38+/-39.85, 71.37+/-17.52mg h/l, and 9.65+/-2.34, 6.10+/-1.79, 4.55+/-1.38mg/l, respectively. In contrast, the AUC(0)(-)(72) of AcSP was significantly lower in m/m subjects. The Cmax of AcSP in w/w, w/m and m/m subjects was 12.67+/-3.32, 9.07+/-2.29 and 4.22+/-0.93mg/l, respectively, with significant differences among groups. However, there was no significant difference in any pharmacokinetic parameter of SASP among groups. CONCLUSION: Different NAT2 genotypes, leading to functional heterogeneity of NAT2, may affect pharmacokinetics of SP and AcSP. Therefore, genotyping NAT2 gene before administration would be important in SASP therapy.

Sulphasalazine-induced leucopenia in a patient with renal dysfunction.

BACKGROUND: Sulphasalazine is used for the long-term maintenance therapy of ulcerative colitis to prevent the relapse of symptoms. However, its clinical use is often restricted by its serious adverse effects. OBJECTIVE: Leucopenia occurred in a patient with severe renal dysfunction after administration of sulphasalazine. The present study was designed to examine whether the adverse event was associated with a disability in the metabolism of sulphasalazine. SUBJECT: A 29-year-old male patient with ulcerative colitis, who underwent haemodialysis thrice a week because of severe renal dysfunction. The chief complaint was diarrhoea. METHODS: Serum concentrations of three major metabolites of sulphasalazine, (5-aminosalicylic acid, sulphapyridine and N-acetyl-sulphapyridine), were measured. The polymorphism of N-acetyltransferase 2, an enzyme that metabolizes sulphapyridine, was also determined by polymerase chain reaction. RESULTS: The trough levels of 5-aminosalicylic acid, sulphapyridine and N-acetyl-sulphapyridine were 0.77-1.45 microg/mL, 31.20-39.25 microg/mL and 14.19-15.03 microg/mL, respectively. The gene diagnosis of N-acetyltransferase 2 suggested that the type was classified as NAT2*6A/*7B, indicating that the patient was a slow acetylator. CONCLUSION: The patient was a slow acetylator, which might lead to a rise in the serum sulphapyridine concentration. Moreover, the decrease in protein binding of sulphasalazine as a result of severe renal dysfunction might have potentiated the effect because of the extremely high protein binding of this compound. Thus, it is most likely that these two factors contributed to the sulphasalazine-induced leucopenia.

Simple and simultaneous determination of sulphapyridine and acetylsulphapyridine in human serum by column-switching high-performance liquid chromatography.

OBJECTIVE: A high-performance liquid chromatography (HPLC) with an automated on-line column-switching system was used for the simultaneous determination of sulphapyridine and acetylsulphapyridine, two major active metabolites related to the adverse effects of sulphasalazine, in human serum. METHODS: Serum samples were directly injected into the HPLC, with the valve automatically switched on to remove serum proteins and other hydrophilic components remaining in the pre-column after elution of sulphapyridine and acetylsulphapyridine to the analytical column. RESULTS: Serum proteins did not interfere with the analysis of either compound. The recoveries of SLP and Ac-SLP from drug-free human serum were 93.03-99.18% and CV were 2.88-4.34%. The within-run reproducibility of assays was excellent with relative standard deviations (RSD) of 1.01-3.90% (SLP) and 0.77-5.56% (Ac-SLP). The limit of quantification of sulphapyridine and acetylsulphapyridine was 3.13 microg/mL and 0.50 microg/mL, respectively. The serum concentrations in a patient with ulcerative colitis, who took 1.0 g sulphasalazine twice daily, were 31.20 microg/mL for sulphapyridine and 14.64 microg/mL for acetylsulphapyridine at 7 h after ingestion. CONCLUSION: The present simple and reproducible assay was useful for the monitoring of serum sulphapyridine and acetylsulphapyridine.

[A case of drug induced urolithiasis composed of acetyl sulphapyridine associated with ulcerative colitis].

A 26-year-old female visited our hospital complaining left flank pain and macroscopic hematuria. She had been suffering ulcerative colitis and administered salazosulphapyridine and predonisolone from 17-year-old. Intravenous urography showed radiolucent multiple stones in the left renal pelvis. Three sessions of extracorporeal shock wave lithotripsy were performed after ureteral stenting. Although disintegration and discharge of the stones were satisfactory, bladder stone induced by ureteral stent was complicated. The extracted bladder stone showed a yellowish brown color and the surface was granular shape. Composition of the stone was acetyl sulphapyridine which was a metabolite of salazosulphapyridine. After maintenance of the urinary pH ranges between 6.5 and 7.5 by medication of sodium bicarbonate, the patient remains free of stone for 3 years. Drug induced urolithiasis originated from salazosulphapyridine is extremely rare. Satisfactory oral fluid intake and urinary alkalization are important for prevention of sulpha drugs calculi of urinary tract.

The influence of multiple oral doses of zileuton on the steady-state pharmacokinetics of sulfasalazine and its metabolites, sulfapyridine and N-acetylsulfapyridine.

The effects of zileuton (Abbott-64077) on the pharmacokinetics of sulfasalazine (SASP) and its metabolites, sulfapyridine (SP) and N-acetylsulfapyridine (ASP), were studied in a randomised double-blind placebo-controlled study enrolling 14 healthy male volunteers. All subjects received SASP 1 g every 12 hours for 8 days and zileuton 800mg or placebo administered twice daily from day 4 to day 8 inclusive. Coadministration of zileuton did not significantly affect the area under the plasma concentration-time curve, the maximum (Cmax) or minimum (Cmin) plasma concentration and the time to Cmax of SASP, SP or ASP. Likewise, zileuton did not modify the terminal elimination half-life of SASP. It is concluded that coadministration of zileuton 1.6 g/day has no significant effects on the pharmacokinetics of SASP 2 g/day or its metabolites, SP and ASP.

Bilateral acetylsulfapyridine nephrolithiasis associated with chronic sulfasalazine therapy.

We report on formation of bilateral renal calculi secondary to sulfasalazine therapy for juvenile rheumatoid arthritis. The condition was successfully treated with extracorporeal shock wave lithotripsy. Analysis of the fragments with thin layer chromatography and nuclear magnetic resonance revealed acetylsulfapyridine, a metabolite of sulfasalazine.

Indicaciones quirúrgicas en colitis ulcerosa crónica inespecífica / Surgical indications for nonspecific ulcerative chronic colitis

Objetivo: Evaluar los resultados del tratamiento quirúrgico en un grupo de pacientes con colitis ulcerativa crónica inespecífica (CUCI) . Sede: Servicios de Cirugía Médico Nacional Siglo XXI del IMSS en México, D.F. Diseño: Estudio retrospectivo, observacional, longitudinal en el que se revisaron datos demográficos, clínicos y quirúrgicos de 10 pacientes operados por colitis ulcerativa crónica insepecífica en el lapso de 7 años (1985-1992). Resultados: Seis pacientes fueron hombres y 4 mujeres, con promedio de edad de 34.5 años, los síntomas principales fueron: diarrea en el 100 por ciento, hemorragia de tubo digestivo en 90 por ciento y dolor abdominal en 80 por ciento. La indicación quirúrgica más frecuente fue falla del tratamiento médico en 6 enfermos. La Técnica quirúrgica más empleada fue colectomía subtotal con ileostomía y bolsa de Hartman en 7 pacientes. La mortalidad fue del 30 por ciento.

Structural requirements of some sulphonamides that possess an antifertility activity in male rats.

Sulphonamides with different chemical structures were synthesized and these 13 compounds together with 7 commercially available sulpha drugs were tested for antifertility activity by natural mating in male rats. All compounds were given daily by gastric intubation at doses of 125, 150, 250 or 450 mg/kg for 6 weeks. Sulphapyridine caused a dose-related and reversible reduction in fertility at doses between 125 and 450 mg/kg. At the high dose, fertility was reduced to 25.9% of control at 5 weeks after treatment, and complete recovery occurred by 3 weeks after drug withdrawal. This activity was abolished when the pyridine ring was substituted by other heterocyclic rings, except sulphachloropyridazine which had only weak activity. Replacement of the pyridine ring by a hydrogen atom or short aliphatic chains preserved or even enhanced the potency. Thus, sulphanilamide, N1-methylsulphanilamide or N1-diethylsulphanilamide produced a marked but reversible reduction in fertility. Removal or substitution of the N4-amino group on the benzene ring of sulphapyridine with a methyl group destroyed the activity. However, the bromo or nitro analogue (at the para- but not the meta-position of the benzene ring) still possessed some activity. N4-Acetyl derivatives of sulphapyridine, sulphanilamide, and N1-diethylsulphanilamide were as potent as their parent compounds. These results suggest that the presence of pyridine or other heterocyclic rings is not necessary for the antifertility activity of sulphonamide compounds. However, the N4-amino group is indispensable. In addition, acetylation of this amino group does not change the potency. The prototype of the antifertility sulphonamides therefore seems to be sulphanilamide.

Simultaneous determination of sulfasalazine and its metabolites sulfapyridine and N-acetylsulfapyridine in human serum by ion-pair high-performance liquid chromatography using a polymer-based column.

An HPLC method is described for the simultaneous and rapid determination of sulfasalazine (salicylazosulfapyridine) and two of its metabolites, sulfapyridine and N-acetylsulfapyridine, in human serum. The range of quantitation is 0.1 to 12 micrograms/mL for sulfasalazine and sulfapyridine and 0.25 to 12 micrograms/mL for N-acetylsulfapyridine. Serum is mixed with acetonitrile containing the internal standard sulfamethazine and the ion-pairing agent tetraethylammonium chloride. The acetonitrile extract is concentrated and analyzed by HPLC, using a new polymer-based column, and detected by UV spectroscopy at 270 nm. This paper is the first both to describe the simultaneous analysis of all three of the compounds from serum and to present sulfasalazine concentration-time data following oral administration to humans.

Pharmacokinetics of sulfasalazine metabolites in rats following concomitant oral administration of riboflavin.

Sulfasalazine, 60 mg/kg, was administered orally to groups of rats (n = 4) along with 1, 5, or 10 mg/kg of riboflavin. Plasma and urine were assayed for 5-aminosalicylic acid, acetyl-5-aminosalicylic acid, sulfapyridine, and acetyl-sulfapyridine using an HPLC method. The mean percent of dose recovered as total metabolites in urine was significantly greater (alpha = 0.01) for the group receiving 10 mg/kg riboflavin compared to the controls or the group receiving 1 mg/kg riboflavin. Plasma AUC and Cmax values were also significantly greater (alpha = 0.05) for the 10 mg/kg riboflavin group. These results suggest that at higher doses, a significant fraction of riboflavin reaches the colon intact and stimulates more efficient reduction of the azo bond in sulfasalazine. Since the concentrations of 5-ASA achieved in the colon may be directly related to the efficacy of sulfasalazine in treating inflammatory bowel disease, concomitant administration of riboflavin may enhance sulfasalazine's efficacy in humans.

Sulfasalazine treatment and lymphocyte function in patients with rheumatoid arthritis.

Sulfasalazine is now an established 2nd line agent in the treatment of rheumatoid arthritis (RA) but its mode of action is unknown. Two separate studies have investigated the possibility that it works in RA by influencing lymphocyte function. After 12 weeks of treatment with sulfasalazine, elevated levels of circulating activated lymphocytes and abnormal ex vivo mitogen response to concanavalin A (Con-A) in 11 patients with RA reverted to normal. An in vitro study investigated the effect of sulfasalazine and its metabolites on mitogen response by healthy and RA peripheral blood mononuclear cells (PBMC). Sulfapyridine (SP) and 5-hydroxy SP suppressed the response of RA PBMC to Con-A. Sulfasalazine, SP and N-acetyl SP suppressed the response of healthy PBMC to pokeweed mitogen. 5-aminosalicylic acid also affected mitogen response and cell viability, which may be relevant to actions of this metabolite within the gut.

Sulphasalazine in the treatment of rheumatoid arthritis: relationship of dose and serum levels to efficacy.

Previous studies of sulphasalazine in rheumatoid arthritis have chosen an empirical dose based upon its use in ulcerative colitis. In this study we compare the efficacy and toxicity of two doses (1.5 g/day and 3 g/day, 30 patients per group), and attempt to relate efficacy to serum levels of sulphasalazine and its metabolites. After six months 24 of the low-dose group and 20 of the high-dose group remained on treatment. Greater improvement was seen in the high-dose group. When dose was expressed as mg/kg, the dose efficacy ratio became more apparent and a dose of greater than 40 mg/kg/day appears to confer greater benefit. No relationship was demonstrated between serum levels of sulphasalazine or its measured metabolites, and efficacy. We conclude that response to sulphasalazine in rheumatoid arthritis is dose dependent but does not relate to serum levels of sulphasalazine, sulphapyridine, or acetyl sulphapyridine.

Plasma and synovial fluid concentrations of sulphasalazine and two of its metabolites in rheumatoid arthritis.

A study was made of plasma and synovial fluid levels of sulphasalazine, one of its dissociation products--sulphapyridine and a metabolite of the latter--acetyl sulphapyridine in patients with rheumatoid arthritis (RA) who were in a steady state on sulphasalazine therapy. Combined sulphapyridine levels were significantly higher than those of sulphasalazine both in plasma and synovial fluid. Synovial fluid levels of both drugs correlated with their plasma levels and were generally slightly lower. Some patients accumulated sulphasalazine and sulphapyridine in the synovial fluid and the mean concentration of sulphasalazine was higher in the fluid than in the plasma. The explanation for this is uncertain. The concentration of combined sulphapyridine in synovial fluid was related to local joint inflammation and more active systemic disease. No consistent association was found between sulphasalazine levels and local or systemic activity. The higher sulphapyridine levels in synovial fluid found in this study suggest the possibility that this moiety could play a more active role in RA than it does in inflammatory bowel disease.