Piroxicam binding to HSA was studied using equilibrium dialysis and fluorescence methods. It was shown that this drug, like its analogs isoxicam and tenoxicam, binds to the apazone locus (site I area) and to a lesser extent to the diazepam site (site II). The piroxicam binding to HSA can be modulated by various specific ligands--apazone, warfarin, diazepam, ibuprofen--and these drug interactions have to be considered not only as potential displacement from the HSA binding sites but also in terms of induced allosteric effects.

No abstract

Blood binding of tenoxicam was studied in vitro by equilibrium dialysis. Isolated human plasma proteins and blood cells were checked, and the distribution of the bound form was then calculated. The results showed that tenoxicam is mainly bound to HSA and that binding percentages are not different when measured in plasma (98.4%) and in an HSA solution at physiological concentration (704 microM, 98.15%). In these conditions, within the range of 1-150 microM, the tenoxicam binding percentage remained constant, evidence of a nonsaturable process. When a lower HSA concentration (10 microM) was used, the binding parameters of the tenoxicam interaction were calculated by using the same equilibrium dialysis data, by 3 methods of analysis- a stoichiometric method and site-oriented methods, fixing or not the number of HSA binding sites (n) as integer values. The best fit was observed with the first method, suggesting that two main interactions occurred. The site-oriented method gave lesser fits, the better being observed when n was not fixed. Its value, 1.77, suggest the possibility of two binding sites, one of them not preformed. The effects of known markers of site I, warfarin and apazone, of site II, diazepam and ibuprofen and of palmitic acid showed that tenoxicam is bound simultaneously to both sites I and II. The binding capacity of site I for tenoxicam is enhanced by diazepam: as this compound alone is bound to site II, this result suggests that the two HSA binding sites are not independent.

The free drug hypothesis, which states that only the unbound moiety of drug in blood is available for tissue diffusion, is discussed according to recent investigations. In some experimental conditions, it must be assumed that part of the protein-bound drug in plasma is extracted during a single passage through the organ studied. The mechanisms underlying these observations are not unequivocal and remain hypothetical. In the liver, high-affinity binding sites for serum albumin have been demonstrated, and they would explain the high extraction by liver of endogenous and exogenous compounds. However, these experiments measure the unidirectional transfer of a drug from the vascular to the extravascular space in non-steady-state conditions. Hence, in steady-state conditions, the free drug hypothesis cannot be ruled out because it is supported by numerous pharmacokinetic studies.

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Many diseases appear to lead to a decrease of drug plasma binding due either to hypoalbuminaemia or to a modification of albumin structure. In other diseases, the binding of a drug may increase due to elevated concentrations of alpha 1-acid glycoprotein or lipoproteins. However that may be, the free fraction of a drug may vary in different pathologies. But an increase or decrease of the drug free fraction does not automatically mean an increase or decrease of the free drug concentration. Whatever the drug, a variation in the volume of distribution more or less proportional to the variation in the plasma free fraction can be expected. With respect to the clearance, the problem is much more complex and depends on the hepatic extraction ratio of drug. If the extraction is related to the free fraction (fu) of drug, a variation in fu will lead to a variation in the total drug concentration but no variation in the free drug concentration and no change in the pharmacological effect. If the extraction of a drug is dependent on hepatic flow, a variation in fu will lead to a change in the free drug concentration (with no change in the total drug concentration) and hence changes in the pharmacological effect. The aim of this article is to review the literature concerning disease-induced variations in plasma protein levels during the past 10 years. Finally, possible implications for drug dosage regimens are discussed generally from examples studies in the literature.

Brain extraction of a tricyclic antidepressant, imipramine, was investigated using the carotid injection technique in the rat. The extent to which drug binding to plasma proteins and erythrocytes could inhibit the brain extraction was measured. Equilibrium dialysis showed that imipramine is highly bound to human serum albumin (HSA), alpha 1-acid glycoprotein (AAG), lipoproteins, and erythrocytes. The free dialyzable drug fraction was inversely related to the protein concentration. Despite this degree of binding, no significant reduction in the brain extraction of the drug was observed in the presence of HSA, lipoprotein, or erythrocytes. Only AAG reduced the brain transport of this drug in a ratio related to the protein concentration. However, the rat brain extraction was higher than expected from the in vitro measurement of the dialyzable fraction. These data indicate that the amount of circulating imipramine available for penetration in brain exceeds widely the dialyzable fraction of the drug as measured in vitro.

The aim of our study was to answer the following questions: (1) is thiamine deficient in chronic excessive drinkers; and (2) is peripheral neuropathy associated with thiamine deficiency or with alcohol intake itself? We performed direct assays of blood concentrations of free thiamine and thiamine phosphate in excessive drinkers with or without peripheral neuropathy and in control subjects. We found no difference in free thiamine concentrations between excessive drinkers with and without neuropathy, and no difference in free thiamine concentrations between the two groups of excessive drinkers and the control group. By contrast, a deficiency in thiamine phosphate was observed in each group of excessive drinkers compared to the control group. This was reflected in blood concentrations of total thiamine which were also lower in excessive drinkers than in controls. Finally, the thiamine phosphate: free thiamine ratio was slightly but significantly lower in the two groups of excessive drinkers than in the control group. Both groups of excessive drinkers showed typical moderate liver disease of alcoholic origin. In conclusion, the free thiamine fraction was not diminished in this group of alcoholic hospital inpatients. Thiamine deficiency would not therefore appear to play a determining role in the onset of peripheral neuropathy. In contrast, the phosphorylated fraction was slightly reduced, probably owing to the liver disease in these subjects. Contrary to studies using indirect assay techniques, our results suggest that thiamine deficiency is either slight or absent in chronic drinkers.