Determination of Urinary Unconjugated Cortisol by Glass Fiber Chromatography in the Diagnosis of Cushing's Syndrome1

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Abstract
Measurement of unconjugated cortisol in the urine affords a reliable index of the biologically active fraction of circulating cortisol useful in the diagnosis of Cushing's syndrome. A simple and accurate technique using glass fiber chromatography was developed to determine unconjugated urinary cortisol. One hundred ml of urine was extracted with dichloromethane without previous hydrolysis, applied to glass fiber sheets and run in 1 or 2 different chromatographic systems for 20 min. Quantitation was performed by the Porter-Silber reaction, and values were corrected for losses by means of added tracer amounts of ortisol-4-C14. Mean recovery was 80.5%, and replicate determinations had an average deviation from the mean of 5.5%. Thirty-eight normal subjects studied with this technique had a mean value of urinary unconjugated cortisol of 71.4 μg/day, with arange of 0–181. Nine patients with Cushing's syndrome had values of 297 to 3,605 μg/day. In thyrotoxic and obese patients with elevated urinary 17-hydroxycorticoid levels, normal excretion of unconjugated cortisol was found. The highest value was observed in a case of carcinoma of the adrenal gland. The decrease in cortisol excretion was greater than the fall of 17-hydroxycorticoids during dexamethasone suppression in normal subjects and in 3 patients with Cushing's syndrome due to adrenal tumor; there was no significant fall in urinary unconjugated cortisol upon suppression. This technique is deemed a superior method for the diagnosis of Cushing's syndrome, as it reflects, albeit indirectly, the circulating, biologically active fraction of excessive cortisol which is responsible for the pathologic state.

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