Presence Of Acetylsalicylic Acid Research Articles

The PGI 2-analogue iloprost and the TXA 2-receptor antagonist sulotroban synergistically inhibit TXA 2-dependent platelet activation

The stable PGI 2-analogue iloprost and the TXA 2-receptor antagonist sulotroban (BM 2 13177) were investigated for possible synergistic effects on platelet aggregation in human platelet rich plasma in vitro. Iloprost and sulotroban synergistically inhibited U 46619, collagen, and the second wave of ADP-induced platelet aggregation. Iloprost and sulotroban at concentrations showing little or no inhibition alone resulted, in combination, in marked or complete inhibition of U 46619 or collagen induced aggregation. Combination of iloprost 10 −10M, which had no effect on the concentration-response curve (CRC)_to U 46619, with sulotroban 5 × 10 −6 M, which shifted the CRC to U 46619 by a factor of 3 to the right, resulted in a rightward shift of the U 46619 CRC by a factor of 4.5. To attain a 4.5-fold shift with either compound alone, a concentration of 5 × 10 −10 M iloprost or 10 −5 M sulotroban was required. A similar mutual enhancement of inhibitory effects was seen for combinations of the PGI 2-analogue cicaprost (ZK 96.480) with sulotroban or the TXA 2-receptor antagonist SQ 29548 with iloprost. When the TXA 2-dependent part of collagen-induced aggregation was fully inhibited by sulotroban, the concentrations of iloprost necessary for 90% inhibition were reduced by a factor of 2.5–3. In the presence of acetylsalicylic acid, the synergistic action of sulotroban and iloprost was reduced and merely additive effects against U 46619-induced platelet aggregation were found, suggesting that the release of endogenous TXA 2 plays an important role for the synergistic effect of the two compounds. The combination of a PGI 2-analogue and a TXA 2-antagonist may lead to a safer and more effective control of platelet activation than with either compound alone.

Sulfation in vitro of mucus glycoprotein by submandibular salivary gland: effects of prostaglandin and acetylsalicylic acid

Enzymatic sulfation of mucus glycoprotein by rat submandibular salivary and the effect of prostaglandin and acetylsalicylic acid on this process were investigated in vitro. The sulfotransferase enzyme which catalyzes the transfer of sulfate ester group from 3′-phosphoadenosine-5′-phosphosulfate to submandibular gland mucus glycoprotein has been located in the detergent extracts of Golgi-rich membrane fraction of the gland. Optimum enzyme activity was obtained at pH 6.8 with 0.5% Triton X-100, 25 mM NaF and 4 mM MgCl 2, using the desulfated glycoprotein. The enzyme was also capable of sulfation of the intact mucus glycoprotein, but the acceptor capacity of such glycoprotein was 68% lower. The apparent K m of the submandibular gland sulfotransferase for salivary mucus glycoprotein was 11.1 μM. The 35S-labeled glycoprotein product of the enzyme reaction gave in CsCl density gradient a 35S-labeled peak which coincided with that of the glycoprotein. This glycoprotein upon reductive β-elimination yielded several acidic 35S-labeled oligosaccharide alditols which accounted for 75% of the 35S-labeled glycoprotein label. Based on the analytical data, the two most abundant oligosaccharides were identified as sulfated tri- and pentasaccharides. The submandibular gland sulfotransferase activity was stimulated by 16,16-dimethyl prostaglandin E 2 and inhibited by acetylsalicylic acid. The rate of enhancement of the glycoprotein sulfation was proportional to the concentration of prostaglandin up to 2 · 10 −5 M at which point a 31% increase in sulfation was attained. The inhibition of the glycoprotein sulfation by acetylsalicylic acid was proportional to the drug concentration up to 2.5 · 10 −4 M at which concentration a 48% reduction in the sulfotransferase activity occurred. The apparent K i value for sulfation of salivary mucus glycoprotein in presence of acetylsalicylic acid was 58.9 μM. The results suggest that prostaglandins may play a role in salivary mucin sulfation and that this process is sensitive to such nonsteroidal anti-inflammatory agents as acetylsalicylic acid.

Rat kidney lysosomal membrane damage induced by suramin in vitro and in vivo.

The effect of suramin, an acid naphthylamine, on rat kidney lysosomal membrane integrity was studied. Lysosomal particles were incubated with suramin (0.1-0.4 mM) and light scattering behaviour of the mixtures were subsequently measured. There was a significant decrease (P less than 0.005) in the amount of light absorbed in the presence of suramin compared to lysosomal particle suspension alone. This was accompanied by release of acid phosphatase, a lysosomal 'marker' enzyme, into the suspending medium. These effects were reduced in the presence of acetylsalicylic acid, a known lysosomal membrane stabilizer. Administration of suramin to rats resulted in loss of kidney acid phosphatase and lysozyme activities from the tissue. These results indicate labilization of rat kidney lysosomal membrane by suramin molecules both in vitro and in vivo.

Protein synthesis by HepG2 cells infected with influenza B virus.

Reye's syndrome is an acute hepatopathy and encephalopathy affecting children during the convalescent period of a viral infection, frequently influenza B. The role of influenza B in the pathogenesis of Reye's syndrome is unknown. To investigate this relationship, an in vitro system was designed to examine the interaction of influenza B virus with a cell line of human hepatocytes (HepG2) which retains many specific hepatic synthetic characteristics. HepG2 was capable of supporting a productive infection by influenza B, although greater virus inputs were required than in fully permissive Madin-Darby canine kidney cells. Because of the recent association of Reye's syndrome with aspirin use, the kinetics of influenza B growth were studied in the presence of acetylsalicylic acid (25 and 100 micrograms/ml) and were not found to be altered. Protein synthesis by HepG2 cells was decreased by 80% in influenza B-infected cells when compared to uninfected controls. Acetylsalicylic acid, 100 micrograms/ml, did not affect the rate of 35S-methionine incorporation by infected or uninfected HepG2 cells. The rates of synthesis of specific proteins (albumin, transferrin, and apoprotein B) by HepG2 cells were determined by immunoprecipitation of 35S-methionine labeled cell lysates. After 12 h of infection, synthesis of all three plasma proteins was decreased by 40-60%. These studies describe a useful system for delineation of mechanisms by which influenza B virus interacts with host hepatocytes and the host-cell protein synthetic machinery. The studies could be pertinent to the pathogenesis of Reye's syndrome.

Measurement of 6-keto-prostaglandin F 1α in human plasma with radioimmunoassay: Effect of prostacyclin infusion

A radioimmunoassay for 6-keto-prostaglandin F 1a (6-keto-PGF 1α), a stable hydration product of prostacyclin (PGI 2), is described. The method utilises plasma extraction with ethyl acetate and saturation analysis. Antiserum was produced in rabbits against 6-keto-PGF 1α-BSA conjugate, and its cross-reactions with 22 prostaglandins and related compounds were insignificant. The mean mass of added 6-keto-PGF 1α required to displace zero-point binding by 50 % was 55.6 ± 5.1 pg (mean ± SD, N = 10). The method blank was 7.8 ± 2.4 pg/ml (N = 10), resulting in a detection limit of 12.6 pg/ml (mean blank + 2 SD). The intro-assay variation was between 4.1 − 8.2 % at concentrations between 150.3 pg/ml and 560.7 pg/ml (N = 10). The interassay variation was 12.4 % (N = 15). The recoveries of 100 pg/ml or 200 pg/ml added to the plasma samples (N = 10) were 90.2% and 95.4 %, respectively. The 6-keto-PGF 1a concentrations in healthy subjects ranged from 92.7 to 394.5 pg/ml with a mean of 182.5 pg/ml in plasma. The presence of acetylsalicylic acid or indomethacin in the collection tubes caused no changes in plasma 6-keto-PGF a concentration. Intravenous infusion of PGI 2 (1–8 ng/kg/mini into 6 healthy women caused linear increases in 6-keto-PGF 1α. concentrations in plasma. After the discontinuation of PGI 2 infusion, the 6-keto-PGF levels decreased rapidly, the half-life being approximately 30 minutes.

Mucosal Response to Acetylsalicylic Acid

An ultrastructural study was undertaken to chronicle the morphologic aspects of the response of the alimentary tract to the presence of acetylsalicylic acid in the lumen. A clinical dose (10 mg/kg) of buffered acetylsalicylic acid was administered to male Wistar rats by (a) gastric intubation (feeding tube), and (b) gastrointestinal perfusion. The animals used in this study were sacrificed at various intervals 15, 30, 60 minutes and 4 hours after administration of the dose. Tissues from the fundus of the stomach and the jejunal region of the small intestine were examined.Tissues were examined by light microscopy as well as transmission and scanning electron microscopy. Significant deviations from the appearance of control tissue samples were observed in all animals. Surface gastric erosions were observed in both intestine and stomach, as well as localized areas of punctate hemorrhage. Conventional light micrographs revealed a separation of the epithelial lining from the basement membrane structure: the intercellular space was greatly distended compared to normal.

Consumption of kininogen, formation of kinin and activation of arginine ester hydrolase in rat plasma by rat peritoneal fluid cells in the presence of l-adrenaline

Cells harvested from the peritoneal cavity of the rat, were able to lower kininogen, release kinin and activate benzoylarginine ester (BAEE) hydrolase following brief incubation with rat plasma in the presence of l-adrenaline. These effects were not reproduced when either cells or the catecholamine were incubated with plasma. Peritoneal cells incubated with l-adrenaline did not acquire detectable BAEE-esterase activity, but became able to induce this activity as well as lowering of kininogen, in adrenaline-free plasma. Phenoxybenzamine and, to a lesser extent, propranolol, blocked the action of adrenaline, which was also sensitive to Trasylol, the anti-kallikrein inhibitor from bovine tissues. l-Noradrenaline was less effective than adrenaline, isoprenaline, ineffective. Peritoneal cavity washings which contained no mast cells, but were normal in their lymphocyte and eosinophil content, were unable to lower plasma kininogen in the presence of l-adrenaline. Pretreatment with compound 48/80, leading to substantial release of histami did not decrease the cells' ability to lower plasma kininogen in the presence of adrenaline. The catecholamine failed to release histamine from peritoneal fluid cells. Acetylsalicylic acid (Aspirin®) blocked cell-linked lowering of kininogen and the activation of BAEE-esterase by l-adrenaline. This effect was obtained at a dose (2–10 μg/ml) of the analgesic, 200–500 times lower than that required to block histamine release by compound 48/80. While the action of acetylsalicyclic acid on this release was counteracted by glucose, its effect on kininogen breakdown and esterase activation occurred in the presence of physiological levels of the sugar. While peritoneal fluid cells pretreated with acetylsalicylic acid could not be activated by adrenaline, the action of adrenaline-activated cells on plasma kininogen and BAEE-esterase was not inhibited by the presence of acetylsalicylic acid in the incubation medium. Acetylsalicylic acid prevented the appearance of the discrete morphological alterations normally evoked by adrenaline in rat mesentery mast cells, but failed to affect the degranulation evoked in these cells by compound 48/80.

Studies on the Mechanism of Action of Salicylates IV: Effect of Salicylates on Oxidative Phosphorylation

The hydrolysis of acetylsalicylic acid and the oxidative-phosphorylation uncoupling activity of salicylic acid and acetylsalicylic acid in rat liver mitochondrial preparations at pH 7.4 and 25° have been studied. The rate of hydrolysis of acetylsalicylic acid in mitochondrial preparations under these conditions is fairly slow. The uncoupling activity observed in the presence of acetylsalicylic acid is due to the salicylic acid produced during the experiment. The results obtained in the present study support some recent findings on the effect of salicylates on several other biologic activities.

Presence of acetylsalicylic acid in plasma following oral ingestion of aspirin.

SummaryThe absorption and hydrolysis of aspirin has been examined under various experimental conditions. Aspirin appears to be absorbed intact from the gastrointestinal tract. High concentrations of acetylsalicylate in the plasma can be demonstrated following oral ingestion of soluble but not insoluble aspirin preparations. This is correlated with the increased rate of absorption of the soluble products. There appears to be a rapid hydrolysis of acetylsalicylate in vivo and it is suggested that the principal site of this hydrolysis is the tissues. Direct evidence for hydrolysis of acetylsalicylate to salicylate by the liver of the dog has been demonstrated.

放射性同位元素による医薬品の分析 (第2報)

Isotopic dilution analysis of caffeine was carried out by the use of caffeine[1 N-methyl-14C], prepared by methylation of theobromine with methyl[14C] iodide. Satisfactory results were obtained by determination of 30-80mg. of caffeine, either alone or in the presence of acetylsalicylic acid, theobromine, aminopyrine, or acetophenetidine.

A Method for the Quantitative Determination of Salicylic Acid in the Presence of Acetyl Salicylic Acid

DURING the past few months both state and federal laboratories have found many adulterated samples of acetyl salicylic acid (asperin), one of the most common adulterants of this product being salicylic acid. Among the qualitative tests for the differentiation of acetyl salicylic acid and salicylic acid is the test with bromine water. Bromine water gives a precipitate with salicylic acid. but does not give a precipitate with acetyl salicylic acid.' This fact and the method of Fr. Freyer2 for determination of salicylic acid by means of a N/10 bromine solution were utilized for the quantitative determination of salicylic acid in mixtures of the two drugs. The method of Freyer is practically that of the U. S. P. for the determination of phenol. The end product, tri-brom-phenol, is the same with both drugs. Two solutions were prepared, one containing 100 mg. of salicylic acid in 100 cc., and the other 100 mg.. salicylic acid and 100 mg. acetyl salicylic acid in 100 cc. Ten cc. of the solution, equivalent to 10 mg. of U. S. P. salicylic acid, were used for a determination. Twenty cc. of N/10 bromine, U. S. P., were added, followed by 75 cc. water and 5 cc. hydrochloric acid. The mixture was allowed to stand, with occasional shaking, in a glass-stoppered flask for ten minutes, when 5 cc. of potassium iodide, T. S., were added, and the liberated iodines titrated with N/10 sodium thiosulphate, V. S. The two solutions required the same number of cc. of N/10 thiosulphate and showed the presence of 9.9881 mg. of salicylic acid in 10 cc. samples of each solution. A 50 mg. sample of the same salicylic acid showed, by direct titration with N/10 KOH, a purity of 99.9 percent, which result checks very closely with the preceding. It must be remembered, however, that a cold aqueous solution of acetyl salicylic acid decomposes slowly, forming salicylic acid, and if heated the reaction is comparatively rapid.