Sulfonamides tissue levels

The sulfonamides are a group of organic compounds with chemotherapeutic activity they are antimicrobial agents and not antibiotics. They have a common chemical nucleus that is closely related to PABA, an essential component in the folic acid pathway of nucleic acid synthesis. The sulfonamides are synergistic with the diaminopyrim-idines, which inhibit an essential step further along the folate pathway. The combination of a sulfonamide and a diaminopyrimidine is advantageous because it is relatively non-toxic to mammalian cells (less sulfonamide is administered) and is less likely to select for resistant bacteria. Only these so-called potentiated sulfonamides are used in equine medicine. These drugs are formulated in a ratio of one part diaminopyrimidine to five parts sulfonamide, but the optimal antimicrobial ratio at the tissue level is 1 20, which is achieved because the diaminopyrimidines are excreted more rapidly than the sulfonamides. 

Oral Therapy Sulfonamides Trimethoprim-sulfamethoxazole (TMP-SMX) These agents generally have been replaced by more agents due to resistance. This combination is highly effective against most aerobic enteric bacteria except Pseudomonas aeruginosa. High urinary tract tissue levels and urine levels are achieved, which may be important in complicated infection treatment. Also effective as prophylaxis for recurrent infections. 

Although the determination of the tissue levels gives important information about distribution of the sulfonamides in various organs (15), it furnishes no indication as to the local active concentration of the antibacterial. According to modem views, sulfonamides are bound reversibly to blood proteins, mostly the albumins. The degree of protein binding varies considerably among various... 

Rapid progress has been reported in the development of methods for sulfonamide residues in tissues, milk, and eggs since the subject was reviewed by Horwitz ( ) in 1981, The colorimetric method of Tishler et al (j ) has in the past been used to detect violative levels of sulfonamide residues in animal tissues. The lack of specificity and the variable background levels produced by this method have been discussed by Horwitz ( ), Matusik et al (15), and Lloyd et al (16), Recently, a number of specific chromatograpiic methods have been described for determination of residues of a variety of sulfonamides, These are summarized in Table 1 and suggest that HPLC is emerging as the method of choice followed by GLC and TLC methods. 

Sulfonamides can be divided into three major groups (1) oral, absorbable (2) oral, nonabsorbable and (3) topical. The oral, absorbable sulfonamides can be classified as short-, intermediate-, or long-acting on the basis of their half-lives (Table 46-1). They are absorbed from the stomach and small intestine and distributed widely to tissues and body fluids (including the central nervous system and cerebrospinal fluid), placenta, and fetus. Protein binding varies from 20% to over 90%. Therapeutic concentrations are in the range of 40-100 mcg/mL of blood. Blood levels generally peak 2-6 hours after oral administration. 

The kidney was the target tissue analyzed for antibiotics. When violative residues were found in the kidney, the liver of that animal was subsequently analyzed. When violative residues were also found in the liver, muscle tissue was then analyzed. For sulfonamides, the target tissue was liver. When the liver contained violative levels, muscle tissue was also analyzed. 

The applicability of the APCI interface is restricted to the analysis of compounds with lower polarity and lower molecular mass compared with ESP and ISP. An early demonstration of the potential of the APCI interface is the LC-APCI-MS-MS analysis of phenylbutazone and two of its metabolites in plasma and urine (128). Other applications include the LC-APCI-MS analysis of steroids in equine and human urine and plasma (129-131), the determination of six sulfonamides in milk samples after a simple solid-phase extraction and LC separation (132), of tetracyclines in muscle at the 100 ppb level (133), of fenbendazole, oxfendazole, and the sulfone metabolite in muscle at the 10 ppb level, and of five thyreostats in thyroid tissue at the 1 ppm level (134). 

The four-plate test was initially based on the German Hemmstoff-test with an additional plate of Sarcina lulea at pH 8.0, designed for the detection of lower levels of macrolides, and a fourth plate of Escherichia coli at pH 7.2 for the detection of sulfonamides (74,75). The modified version adopted by the European Community for screening carcasses is based on three plates with Bacillus subtilis BGA at pH values of 6.0, 8.0, and 7.2 with added trimethoprim, respectively, and a fourth plate with Micrococcus luteus NCTC 8340 at pH 8.0 (74). This test as described elsewhere (76) is intended to detect residues of -lactams, tetracyclines, aminoglycosides, sulfonamides, and macrolides in muscle tissue of slaughtered animals, without any prior extraction or cleanup. 

However, recent investigations on the effect of the tissue matrix on the detection limits attained by this test have indicated that ceftiofur, sulfonamides, streptomycin, and some macrolide antibiotics cannot be detected in intact meat with the plates and the bacterial strains prescribed in the European four-plate test (81, 82). Two plates of this system were not found suitable for screening sulfamethazine or streptomycin at levels far above the MRL the third plate detected tetracyclines and -lactams up to the MRL levels whereas the fourth was sensitive to -lactams and some but not all macrolides. Detection, on the other hand, of the fluoroquinolones enrofloxacin and ciprofloxacin could only be made possible by an additional Escherichia coli plate not included in the four-plate test. 

W. Haasnoot, M. Bienenmann-Ploum, U. Lamminmaki, M. Swanenburg and H. van Rhijn, Application of a multi-sulfonamide biosensor immunoassay for the detection of sulfadiazine and sulfamethoxazole residues in broiler serum and its use as a predictor of the levels in edible tissue, Anal. Chim. Acta, 552 (2005) 87-95. 

Liquid chromatography-ionspray-mass spectrometry has been shown to be an attractive approach for the determination of semduramicin in chicken liver. Tandem MS using the CID of the molecular ions further enhanced the specificity providing strucmre elucidation and selective detection down to 30 ppb. Liquid chromatography-ionspray-mass spectrometry has also been successfully applied for the assay of 21 sulfonamides in salmon flesh. Coupling of LC with either ISP-MS or ISP-MS-MS has also been investigated as an attractive alternative for the determination of erythromycin A and its metabolites in salmon tissue. The combination of these methods permitted the identification of a number of degradation products and metabolites of erythromycin at the 10-50 ppb level. Tandem MS with CID has also been... 

Sulfonamides are weak acids. They distribute well but relatively slowly (compared with trimethoprim), and tissue concentrations are lower than plasma concentrations. Some sulfonamides reach significant concentrations in the CSF. The highest drug concentrations are in the liver, kidneys and lungs lower levels are achieved in muscle and bone. Sulfonamides cross the placenta and some may achieve therapeutic concentrations in milk. Some are highly protein bound protein binding varies with both the species and the drug. The Vj values in horses for sulfamethazine, sulfadox-ine and sulfadiazine are 0.63, 0.39 and 0.581/kg, respectively. 

Diaminopyrimidines are weak bases. Peak plasma concentrations are reached early and diaminopyrimidines are soon found in high concentrations in tissues. In fact, the tissue concentrations are often higher than the concentrations in serum. When inflammation is present, trimethoprim levels in the CSF may reach 50% of the plasma concentrations. CSF concentrations of pyrimethamine are 25-50% of the plasma concentrations. The Vd for trimethoprim and pyrimethamine is 1.51/kg in horses. The protein binding of trimethoprim is moderate (50%). There is no protein-binding interaction between the sulfonamides and the diaminopyrimidines. 

Glutamylcysteine synthetase, cysteine, or methionine was 100 times more reactive to hypochlorous acid in comparison with amino acids that did not contain thiol groups (Folkes et al., 1995). Sublethal exposures to HOCl decreased GSH levels in several cell types (Vissers and Winterboum, 1995 Pullar et al., 1999). In a study by Pullar et al. (1999) using human umbilical vein endothelial cells, doses of 25 nmol of HOCl and less were sublethal when the exposure was done over 10 min, there was a concentration-dependent loss of intracellular GSH. Tissue exposure to HOCl resulted in a reduction of GSH. The metabolite of the HOCl interaction with GSH was an unexpected cyclic sulfonamide that was exported from the cell. The expected metabolites of glutathione disulfide (GSSH) and GSH sulfonic acid were actually minimal (Pullar et al., 2001). Inactivation of acetylcholinesterase by HOCl could be a contributory cause of airway hyperreactivity (den Hartog et al., 2002). 

An important property of sulfonamides is their ability to penetrate into the tissues and extra vascular fluids where they can exert their bacteriostatic function. This can best be demonstrated in animal experiments. If the sulfonamide is injected intravenously into an animal whose excretory functions have been markedly reduced by extirpation of the kidneys and ligation of the bile duct, one finds much lower blood levels than are calculated from the total blood volume and the total amount of drug administered. This proves that a substantial part of the drug has disappeared into the tissues (13,21). 

As an example of how the chemical modification can influence the affinity of sulfonamides to certain tissues, the results of another model experiment are briefly reviewed. Groups of four rabbits received a single oral dose of 500 mg. per kg. of five different sulfonamides and were sacrificed 3 hours later. Free and total sulfonamide were determined in the cortex and medulla of the kidneys as well as in the blood plasma. A summary of the results limited to the free sulfonamide is presented in Figure 9. The most pronounced differences are seen between sulfisoxazole and sulfamethoxazole, two agents which are chemically similar. With sulfisoxazole, the kidney levels are much higher than the plasma levels and the sulfonamide content in the medulla of the kidney markedly surpasses the one in the cortex (index medulla/cortex = 1.6). With sulfamethoxazole, almost identical sulfonamide levels are found in kidney cortex, medulla, and blood plasma. 

The data supporting this relationship have been reviewed in detail for pharmaceutical preparations at concentration levels of approximately 0.1 to 100 (1 ), for pesticide residues at about 1 ppm (] ), and for aflatoxins at about 10 ppb (Ijl). We have recently reviewed the collaboratively studied methods for sulfonamides in feeds at about 100 ppm (0.01 ), which shows a CV of about 4, and various drugs as tissue residues at about 1 ppm with a CV of about 16. We have also spot checked individual studies of major nutrients at the 0.1-10 levels, minor nutrients and drugs at the 10-100 ppm levels, and trace elements by atomic absorption... 

In a great many cases the level of free drug in the tissue fluids eventually becomes the same as that in the plasma. This relationship is convenient because the plasma is more accessible for analysis. Among examples investigated are the anti-bacterial sulfonamide, sulfadoxine ( Fanasil ) in the intraperitoneal cavity of rats after oral injection (McQueen, 1968), also various penicillins in lymph of dogs. The literature of this correlation has been reviewed by Robinson (1966). 

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