Testosterone Binding and Free Plasma Androgen Concentrations under Physiological Conditions: Characterization by Flow Dialysis Technique*

Abstract

Testosterone [T] binding to plasma was analyzed under physiological conditions. Flow dialysis technique (FDT) permitted precise studies of whole 37 C plasma with negligible disturbance of the equilibrium state. The information obtained by FDT was compared with that obtained by a charcoal adsorption assay (CA), which primarily assesses T binding to testosterone-estradiol-binding globulin (TEBG). The results of the 2 methods correlated closely (r [correlation coefficient] = 0.98). Consequently, the percentage of T free in plasma under in vivo conditions could be readily calculated from data generated by the much easier CA technique. Plasma free androgen concentrations were then computed for a large variety of sera. For example, the mean (.+-. SD) plasma free T concentrations of normal men, hirsute women, normal women and pregnant women were found to be 128 .+-. 57, 16.1 .+-. 11.7, 5.4 .+-. 2.7, and 4.6 .+-. 1.7 pg/ml, respectively. These levels are in close agreement with previous estimates. The plasma free T level of women on estrogen-containing oral contraceptives was normal (6.6 .+-. 0.4 pg/ml) in the face of frequently elevated plasma total androgen levels. The FDT data are shown by mathematical modeling and competition studies to fit a general model for the binding of the TEBG ligands in plasma (plasma 17.beta.-hydroxysteroids) to TEBG and albumin. This model permitted the calculation of the association constants of these testosterone-binding species under physiological conditions. This made is possible to define the relative role of TEBG and albumin as determinants of the percentage of T free in plasma and to compute TEBG capacity directly from the total plasma 17.beta.-hydroxysteroid level and the percentage of free 17.beta.-hydroxysteroid obtained by CA. TEBG plays the central role in plasma T binding because of its high association constant (8.4 .times. 108 M-1) and the wide variation in its plasma level among individuals. Changes in serum albumin concentration within the normal range exert little influence on the percentage of free T at any level of TEBG. Hypoalbuminemia does result in a significant increase in the percentage of free T at any low TEBG titer, however. Means of correcting CA data for this effect are presented.