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Salicylate-induced occult gastrointestinal blood loss: comparison between different oral and parenteral forms of acetylsalicylates and salicylates.

Gastrointestinal fecal blood loss, determined by injecting 51Cr-labelled autologous red blood cells, was measured in 191 orthopaedic patients after oral or parenteral intake of different forms of acetylsalicylates or salicylates. Oral or parenteral administration of non-acetylated salicylates caused nearly no gastrointestinal bleeding, but the anti-inflammatory activity of these products can be questioned since they cannot inhibit prostaglandin synthetase. Buffered, soluble forms of acetylsalicylates caused gastrointestinal bleeding in more than 50% of the patients. However, enteric-coated and intravenous forms resulted in significantly less gastrointestinal bleeding. For both preparations a relationship between serum salicylate level and amount of fecal blood loss was found in the group of "bleeders" (an upper limit of physiological blood loss could definitely be determined by this method). The findings suggested a similar mode of action of enteric-coated and intravenous acetylsalicylates on gastric mucosa through a systemic action. It was concluded that in long-term anti-inflammatory salicylate treatment, enteric-coated forms are probably the galenic form of first choice.

Administration, Oral↗

Crystal structures of Yersinia enterocolitica salicylate synthase and its complex with the reaction products salicylate and pyruvate.

The salicylate synthase, Irp9, from Yersinia enterocolitica is involved in the biosynthesis of the siderophore yersiniabactin. It is a bifunctional enzyme that forms salicylate and pyruvate from chorismate and water via the intermediate isochorismate. Here we report the first crystal structure of Irp9 and also of its complex with the reaction products salicylate and pyruvate at 1.85 A and 2.1 A resolution, respectively. Like other members of the chorismate-utilizing enzyme family, e.g. the TrpE subunit of anthranilate synthase and the PabB subunit of 4-amino-4-deoxychorismate synthase, Irp9 has a complex alpha/beta fold. The crystal structure of Irp9 contains one molecule each of phosphate and acetate derived from the crystallization buffer. The Irp9-products complex structure was obtained by soaking chorismate into Irp9, demonstrating that the enzyme is still catalytically active in the crystal. Both structures contain Mg(2+) in the active site. There is no evidence of the allosteric tryptophan binding site found in TrpE and PabB. Mutagenesis of Glu240, His321 and Tyr372 provided some insight into the mechanism of the two transformations catalyzed by Irp9. Knowledge of the structure of Irp9 will guide the search for potent inhibitors of salicylate formation, and hence of bacterial iron uptake, which is directly related to the virulence of Yersinia.

Amino Acid Sequence↗

Studies of electron-transfer properties of salicylate hydroxylase from Pseudomonas cepacia and effects of salicylate and benzoate binding.

The pH dependence of the redox behavior of salicylate hydroxylase from Pseudomonas cepacia as well as the effects of salicylate, benzoate, and chloride binding is described. At pH 7.6 in 0.02 M potassium phosphate buffer E1(0')(EFl ox/EFl.-) is -0.150 V and E2(0')(EFl.-/EFl red H-) is -0.040 V versus the standard hydrogen electrode (SHE). A maximum of 5% of FAD anion semiquinone is thermodynamically stabilized under these conditions. However, in coulometric and dithionite titrations more semiquinone is kinetically formed, indicating slow transfer of the second electron. The potential/pH dependence is consistent with a two-electron, one-proton transfer. Upon salicylate binding the midpoint potential is shifted 0.020 V negative from -0.094 to -0.114 V vs SHE at pH 7.6. A maximum of 7% of the neutral semiquinone is stabilized both in potentiometric and coulometric titrations. This small potential shift indicates that the substrate is bound nearly to the same extent to all three oxidation states of the enzyme. It is clear that the substrate binding does not make the reduction of the flavin thermodynamically more favorable. In contrast to salicylate, the potential shift caused by the effector, benzoate, is much more significant. (A maximum potential shift of -0.07 V is calculated.) Benzoate binds most tightly to the oxidized form and is least tightly bound to the two-electron-reduced form of the enzyme. For the reduction of the free enzyme the transfer of the second electron or the transfer of the proton is rate limiting, as is shown by the kinetic formation of the anionic semiquinone.(ABSTRACT TRUNCATED AT 250 WORDS)

Benzoates↗

Direct ring fission of salicylate by a salicylate 1,2-dioxygenase activity from Pseudaminobacter salicylatoxidans.

In cell extracts of Pseudaminobacter salicylatoxidans strain BN12, an enzymatic activity was detected which converted salicylate in an oxygen-dependent but NAD(P)H-independent reaction to a product with an absorbance maximum at 283 nm. This metabolite was isolated, purified, and identified by mass spectrometry and (1)H and (13)C nuclear magnetic resonance spectroscopy as 2-oxohepta-3,5-dienedioic acid. This metabolite could be formed only by direct ring fission of salicylate by a 1,2-dioxygenase reaction. Cell extracts from P. salicylatoxidans also oxidized 5-aminosalicylate, 3-, 4-, and 5-chlorosalicylate, 3-, 4-, and 5-methylsalicylate, 3- and 5-hydroxysalicylate (gentisate), and 1-hydroxy-2-naphthoate. The dioxygenase was purified and shown to consist of four identical subunits with a molecular weight of about 45,000. The purified enzyme showed higher catalytic constants with gentisate or 1-hydroxy-2-naphthoate than with salicylate. It was therefore concluded that P. salicylatoxidans synthesized a gentisate 1,2-dioxygenase with an extraordinary substrate range, which also allowed the oxidation of salicylate.

Chromatography, High Pressure Liquid↗

Potentiometric membrane electrode for salicylate based on an organotin complex with a salicylal Schiff base of amino acid.

A novel salicylate-selective electrode based on an organotin complex with a salicylal Schiff base of amino acid salicylaldehydeaminoacid-di-n-butyl-Sn(IV) [Sn(IV)-SAADB] as ionophore is described, which exhibits high selectivity for salicylate over many other common anions with an anti-Hofmeister selectivity sequence: Sal- >> PhCOO- > SCN- > Cl04- > I- > NO3- > NO2- > Br- > Cl- > CH3COO-. The electrode, based on Sn(IV)-SAADB, with a 30.44 wt% PVC, a 65.45 wt% plasticizer (dioctyl phthalate, DOP), a 3.81 wt% ionophore and a 0.3 wt% anionic additive is linear in 6.0 x 10(-6) - 1.0 x 10(-1) mol l(-1) with a detection limit of 2.0 x 10(-6) mol l(-1) and a slope of 62.0 +/- 1.2 mV/decade of salicylate concentration in a phosphate buffer solution of pH 5.5 at 25 degrees C. The influence on the electrode performances by lipophilic charged additives was studied, and the possible response mechanism was investigated by UV spectra. The electrode was applied to medicine analysis and the result obtained has been satisfactory.

Amino Acids↗

Quantification of salicylate in serum by use of salicylate hydroxylase.

In this enzymatic method for salicylate in serum, Pseudomonas salicylate hydroxylase (EC 1.14.13.1) is used. This stable enzyme catalyzes the stoichiometric, unidirectional conversion of salicylate and NAD(P)H to catechol and NAD(P)+ in the presence of molecular oxygen. The concentration of salicylate in a clinical sample is determined by measuring the delta A at 340 nm as compared with a standard. This new method is rapid, highly specific, requires 40 microL or less of sample, and we saw no interference by any of 61 commonly used drugs. Lipemic, icteric, or hemolyzed samples can be used. Furthermore, this method does not involve extraction, deproteinization, or derivatization. Results are precise and agree well with those obtained by the Trinder test.

Catalysis↗

A novel salicylate-selective electrode based on a Sn(IV) complex of salicylal-imino acid Schiff base.

A novel poly(vinyl chloride) membrane electrode with high selectivity toward salicylate (Sal-), based on the use of the salicylal-imino acid Schiff base dibenyl complex of Sn(IV) [Sn(IV)-SIADBen] as ionophore is described. The influence of lipophilic charged additives on the performance of the electrode was studied. The results suggested that Sn(IV)-SIADBen according to a positively-charged carrier mechanism. The influence of several other variables was investigated in order to optimize the potentiometric response and selectivity of the electrode. The electrode based on Sn(IV)-SIADBen, with 30.44 wt% PVC, 64.55 wt% plasticizer [dioctyl phthalate (DOP)], 3.81 wt% ionophore, and 1.2 wt% anionic additive exhibited a linear response for the Sal- ion over the concentration range 1.0x10(-1) to 2.5x10(-6) mol l-1, and displayed an anti-Hofmeister selectivity sequence as follows: salicylate>>perchlorate>thiocyanate>benzoate>iodide>nitrate>chloride>nitrite approximately acetate>citrate>sulfate. UV-Visible absorption spectra were used to examine the specific interaction of salicylate with the ionophore. The electrode was applied to clinical medical analysis, and the results obtained were consistent with those obtained by conventional methods.

Journal Article↗

Studies on the reactivity of acyl glucuronides--VII. Salicyl acyl glucuronide reactivity in vitro and covalent binding of salicylic acid to plasma protein of humans taking aspirin.

Salicyl acyl glucuronide (SAG) is a significant metabolite of salicylic acid (SA) and aspirin. We have shown that, under physiological conditions in vitro, SAG undergoes rearrangement in a manner consistent with acyl migration to its 2-, 3- and 4-O-acyl positional isomers as the predominant pathway (T1/2 values were 1.4-1.7 hr in buffer at pH 7.4 and 37 degrees). Incubation of SAG or a mixture of its rearrangement isomers (iso-SAG) (each at approximately 50 micrograms SA equivalents/mL) with human serum albumin (HSA, at approximately 40 mg/mL) revealed the formation of covalent adducts with the protein, with peak concentrations of 1-2 micrograms SA equivalents/mL. The data support a role for the rearrangement/glycation mechanism of adduct formation. Covalent adducts of SA were also detected in the plasma of humans taking aspirin (at > or = 1200 mg/day), but the concentrations were low (<< 100 ng SA equivalents/mL). Reactivity of SAG thus provides a mechanism (though of uncertain quantitative importance) of covalent attachment of the salicyl moiety of aspirin to tissue macromolecules, which is in addition to its well-known acetylating capacity.

Animals↗

Fluorescence study on the interaction of salicylate with rat small intestinal epithelial cells: possible mechanism for the promoting effects of salicylate on drug absorption in vivo.

The water-soluble drug, salicylate, was rapidly taken up by rat small intestinal epithelial cells. Salicylate, known to enhance the absorption of poorly absorbable drugs by rectum and small intestine, caused a significant decrease in the fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene (DPH) and a slight increase in the fluorescence polarization of 8-anilino-1-naphthalene sulfonic acid (ANS) in the isolated rat small intestinal epithelial cell suspension. An increase in the membrane fluidity of epithelial cells may possibly contribute to the enhancement of drug absorption by salicylate.

Anilino Naphthalenesulfonates↗

Salicylate metabolite kinetics after several salicylates.

Single oral doses of aspirin (ASA, 1,500 mg), sodium salicylate (NaSA, 1,500 mg, 1,200 mg), and salicyluric acid (SUA, 500 mg) were given to five subjects. Serial plasma and urine samples were collected for 24 hr (plasma) and up to 48 hr (urine); salicylic acid (SA), SUA, and gentisic acid (GA) were measured by high-pressure liquid chromatography. The plasma concentration/time profiles for SUA after ASA and NaSA were fitted to the empirical equation CpSUA = A-Bt-Ce-alpha t -- (A-C)e-beta t. Michaelis constants (Vm and Km) for the conversion of SA to SUA were calculated from the equation (formula see text), where Cl is the renal clearance of SUA, ke is the rate constant of elimination of SUA, CpSA is the plasma concentration of salicylic acid. The term Cl (formula see text) is the estimated rate of formation of SUA from SA at any time (t). The calculated values (mean +/- SD) of Vm, Km, and Kmf (Km in terms of unbound SA) were 43.4 +/- 10.1 mg SA/hr, 14.3 +/- 3.4 mg SA/l plasma, and 0.75 +/- 0.15 mg unbound SA/l plasma. The Vm values were in accord with those reported, but the value for Km was considerably lower. Renal clearances of SUA and GA were 340 +/- 51 and 65 +/- 10 ml/min.

Adult↗

[Serum levels of salicylic acid in patients with psoriasis treated with 10% salicylic acid ointment].

Ten patients with psoriasis were treated for three days with salicylic acid ointment with vaseline vehicle. In all patients the serum concentration of salicylic acid increased systematically and the concentration depended on the amount of ointment used and on psoriasis form. Only in one case of exudative psoriasis the level of salicylic acid reached the value causing intoxication, and produced a diffuse inflammatory erythema with itching. The treatment caused no changes of biochemical indices.

Administration, Cutaneous↗

Structure and stability of salicylic acid-water complexes and the effect of molecular hydration on the spectral properties of salicylic acid.

Density functional theory with B3LYP parametrization and 6-311++G(d,p) basis set has been used to investigate the structure and stability of salicylic acid-water complexes. The vertical excitation energies for these complexes have been computed using time-dependent density functional theory (with B3LYP parametrization and a 6-311++G(d,p) basis set). It is shown that the hydrogen bond between the carboxylic hydrogen and the oxygen of water is the strongest among all possible hydrogen bonds in the system. The hydrogen bond strength in salicylic acid-water complexes seems to be nearly additive. The change in absorption maximum (lambda(max)) corresponding to the vertical excitation energy for the first three excited singlet and triplet states of the complex with 1-3 water molecules is nominal (approximately 1-3 nm). But with the addition of the fourth water molecule, the lambda(max) for S(1) and T(1) decreases by approximately 17 nm and it increases for S(2) and S(3) by about the same amount. The decrease in lambda(max) for transition to the T(2) state on the addition of the fourth water molecule is only approximately 9 nm. There seems to be an intersystem crossing between the S(1) and T(3) states that could account for the observed fluorescence quenching of salicylic acid in water.

Hydrogen Bonding↗

Salicylic acid carboxyl methyltransferase induced in hairy root cultures of Atropa belladonna after treatment with exogeneously added salicylic acid.

In Atropa belladonna hairy roots, exogeneously added salicylic acid (SA) is converted to methyl salicylate (MSA) through the reaction, which might be catalysed by S-adenosyl-L-methionine: salicylic acid carboxyl methyltransferase (SAMT). Here we cloned a cDNA for A. belladonna SAMT (AbSAMT1), which consisted of 357 aa residues. It was expressed in E. coli, and the recombinant AbSAMT1 showed SAMT activity. When A. belladonna hairy roots were exposed to a high concentration of SA, AbSAMT1 mRNA begins to be expressed 12 h after the exposure, and steady expression continued over 144 h.

Amino Acid Sequence↗

Overexpression of Pto induces a salicylate-independent cell death but inhibits necrotic lesions caused by salicylate-deficiency in tomato plants.

Tomato plants overexpressing the disease resistance gene Pto (35S::Pto) exhibit spontaneous cell death, accumulation of salicylic acid (SA), elevated expression of pathogenesis-related genes, and enhanced resistance to a broad range of pathogens. Because salicylate plays an important role in the cell death and defense activation in many lesion mimic mutants, we investigated the interaction of SA-mediated processes and the 35S::Pto-mediated defense pathway by introducing the nahG transgene that encodes salicylate hydroxylase. Here, we show that SA is not required for the 35S::Pto-activated microscopic cell death and plays a minor role in defense gene activation and general disease resistance in 35S::Pto plants. In contrast, temperature greatly affects the spontaneous cell death and general resistance in 35S::Pto plants, and high temperature inhibits the cell death. The NahG tomato plants develop spontaneous, unconstrained necrotic lesions on leaves. These lesions also are initiated by the inoculation of a virulent strain of Pseudomonas syringae pv. tomato. However, the NahG-dependent necrotic lesions are inhibited in the NahG/35S::Pto plants. This inhibition is most pronounced under conditions favoring the 35S::Pto-mediated spontaneous cell death development. These results indicate that the signaling pathways activated by Pto overexpression suppress the cellular damage that is caused by SA depletion. We also found that ethylene is dispensable for the 35S::Pto-mediated general defense.

Cell Death↗

Synthesis of anacardic acids, 6-[8(Z),11(Z)-pentadecadienyl]salicylic acid and 6-[8(Z),11(Z),14-pentadecatrienyl]salicylic acid.

11-Chloro-3-methoxy-2-undecenal was synthesized from 8-bromooctanol, and an annelation reaction with this aldehyde and ethyl acetoacetate proceeded to give the ethyl 6-(8-chlorooctyl)salicylate. Ethyl 6-(8-chlorooctyl)salicylate was converted to ethyl 6-(7-formylheptyl)-2-methoxybenzoate through the iodide after protection of the phenolic hydroxyl group. Finally, the Wittig reaction with the aldehyde and triphenylphosphonium iodides in the presence of BuLi gave the methoxybenzoates, and then treatments of these methoxybenzoates with BBr3 in CH2Cl2 and 10% NaOH in ethanol gave 6-18(Z),11(Z)-pentadecadienyllsalicylic acid (anacardic acid 3) and 6-[8(Z),11(Z),14-pentadecatrienyl]salicylic acid (anacardic acid 4) which were isolated from plants of the anacardiaceae.

Alkenes↗

Identification and determination of salicylic acid and salicyluric acid in urine of people not taking salicylate drugs.

BACKGROUND: Salicylic acid (SA) is present in the serum of people who have not taken salicylate drugs. Now we have examined the urine of these subjects and found that it contains SA and salicyluric acid (SU). We have established the identities of these phenolic acids and determined their concentrations. METHODS AND RESULTS: The acidic hydrophobic compounds of urine were separated using high-performance liquid chromatography (HPLC) and were detected and quantified electrochemically. Two approaches were used to establish the identity of SA and SU. First, the retention times (Rt) of the substances extracted and those of SA and SU were compared under two sets of chromatographic conditions; the Rt of the compounds suspected to be SA and SU and those of the authentic substances were very similar under both sets of conditions. Second, the unknown substances, isolated by HPLC, were treated with acetyl chloride in methanol and compared with the methyl esters of SA and SU by using gas chromatography-mass spectrometry; the unknown compounds after esterification had very similar mass spectra and gas chromatographic R, to those of methyl salicylate and methyl salicylurate. The median (n = 10) urinary concentration of SA was 0.56 micromol/L (range 0.07-0.89 micromol/L) and that of SU was 3.20 micromol/L (range 1.32-6.54 micromol/L). SA and its major urinary metabolite, SU, were found in the urine of all of the 10 people examined.

Adolescent↗

Potentiation of noise-induced audiogenic seizure risk by salicylate in mice as a function of salicylate-noise exposure interval.

Audiogenic seizure risk can be induced in genetically seizure-resistant BALB/c mice by exposure to an intense noise. Results of this experiment showed that combined exposure to noise and sodium salicylate could produce a greater priming effect than exposure to the noise alone, and the greatest potentiation effect was obtained when animals were exposed to the noise 6 hr after the intake of salicylate. The findings were taken as indirect evidence suggesting that the ototoxic action of sodium salicylate could potentiate vulnerability of the mouse cochlea to noise damage.

Acoustic Stimulation↗

Comparison of serum salicylate levels and gastro-intestinal blood loss between salsalate (Disalcid)and other forms of salicylates.

In a first stage the effect of a single dose of 3 g of salsalate on serum salicylate level was compared with a single intake of 3 g of soluble or enteric-coated acetylsalicylates in 12 healthy subjects. Salsalate seems to resorb faster than the enteric-coated forms but more slowly than the soluble forms of acetylsalicylate. However, in comparison with these latter forms, salsalate activity is more protracted. In the second part of the study, 42 patients were admitted to a trial in which 3 to 5 g of salsalate was given daily and in which serum salicylate levels and blood loss in stools were measured using the method of labelling red blood cells with Cr51. The ideal dosage to obtain a serum salicylate level of 20 mg/100 ml seems to lie between 3 and 4 g of salsalate a day. Salsalate caused abnormal gastro-intestinal blood loss in only 2 of the 42 patients studied, which is significantly fewer compared with the soluble, intravenous or enteric-coated forms of acetylsalicylates.

Aspirin↗