A fluorometric determination of urinary 17-hydroxycorticosteroids using benzamidine

✍ Scribed by Yoshihisa Yamaguchi; Tokuichiro Seki

• Publisher: Elsevier Science
• Year: 1984
• Tongue: English
• Weight: 205 KB
• Volume: 142
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(84)90538-4

✦ Synopsis

A fluorometric determination of urinary 17-hydroxycorticosteroids using a reaction of benzamidine with compounds carrying the dihydroxyacetone side chain is described. The fluorescent compounds have excitation and emission maxima at 370 and 480 nm, respectively. The method includes enzymatic hydrolysis with 6-glucuronidase (EC 3.2. I .3 1, from Escherichia co/i) and extraction with methylene chloride and generation of fluorescence in alkaline solution (pH 13.4). The specificity of the reaction was examined and the results were compared with those of an accepted method based on the Potter-Silber reaction (C. C. Porter and R. H. Silber, 1950, J. Biol. Chem. 185, 201-207). The coefficient of correlation was 0.945 with regression line of y = 0.9 Ix + 0.7 mg/day (y, present method, x, Porter-Silber reaction method). Sensitivity of the reaction was 0.5 &ml of standard or sample, mean recovery of cortisol added to five urine samples (5qg addition) was 9590, and the coefficient of variation of the method (five repeated assays of sample with a value of 5.2 mg/liter) was 6.2%. Q 1984 Academic Rtss, Inc.