A radioimmunoassay method for the measurement of plasma 18-0H-DOC has been developed. The antiserum against 18-OH-DOC was produced in rabbits immunized with 18-OH-
DOC
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3
-oxime-porcine gammaglobulin. Both 1, 2-
3H-18-OH-DOC and Standard 18-OH-DOC were stored in 0.1% pyridine ethanol at -20°C. 1, 2-
3H-18-0H-DOC was added to all samples to compensate for procedural losses. Plasma (1-3ml) was extracted with dichloromethane and chromatographied on LH-20 column (1×50cm). The purified extracts were incubated with antiserum at a 1/1000 dilution for 30 minutes at 25°C, and then for 16 hours at 4°C. Saturated ammonium sulfate was used to separate free from bound 18-OH-DOC.
Recovery after extraction was 4.6 ± 7.5 (SD) %. The accuracy and precision of the method were acceptable, and a sensitivity of 5pg per sample enabled the measurement of very low levels of plasma 18-OH-DOC. High specificity of the method was obtained by a high specifity of the antiserum and a good separating ability of Sephadex LH-20 column chromatography.
The mean plasma 18-OH-DOC levels at 8 a.m. were 14.3 ± 5.4ng/100ml in 22 normal subjects. There was a marked increase of plasma 18-0H-DOC by ACTH and a decrease by dexamethasone. Upright position after furosemide administration, dietary sodium restriction and angiotensin II infusion increased moderately plasma 18-OH-DOC.
These results confirm that 18-0H-DOC secretion is regulated primarily by the anterior pituitary, and the renin-angiotensin system plays minor role in 18-OH-DOC secretion. When compared to mineralocorticoid activity of aldosterone or DOC, the 18-OH-DOC level of 14.3ng/100ml and its biological activity would seem that 18-0H-DOC plays a minor role in homeostasis in the normal human body.
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