Isolation, determination of structure and synthesis of the acid-labile conjugate of aldosterone

Abstract

1. After administration of 600mg of 3H-labelled aldosterone to human volunteers, 57 mg of homogeneous acid-labile conjugate was isolated from the urine and identified as aldosterone 18 beta-D-glucosiduronic acid. 2. Esterification and acetylation of the conjugate gave a tetra-acetate methyl ester, which, by measurement of the optical rotation and nuclear-magnetic-resonance spectrum, was shown to be a beta-glucosiduronate. This tetra-acetate methyl ester was synthesized in approx. 10% yield by the Koenigs-Knorr procedure. 3. Removal of the acetyl and methyl ester groups from the tetra-acetate methyl ester with alkali was accompanied by almost complete isomerization at C-17 to give 17-isoaldosterone 18 beta-D-glucosiduronic acid. 4. To prevent inversion at C-17 during removal of the acetate and ester groups of beta-glucosiduronate (a) the 3,20-disemicarbazone was prepared, (b) the acetate and ester groups were removed from the disemicarbazone by treatment with alkali, and (c) the semicarbazone groups were removed from the product at pH 2.0, and aldosterone 18 beta-D-glucosiduronic acid was obtained in 47% overall yield. 5. In the presence of components used to synthesize beta-glucosiduronate by the Koenigs-Knorr reaction this substance is converted slowly into the alpha-glucosiduronate; this conversion is responsible, in part, for the low yield of beta-glucosiduronate. 6. Two additional conjugates were obtained in the Koenigs-Knorr reaction; a provisional structure was assigned to one substrate. The other substance is a C-18 alpha-glucosiduronate. Removal of the acetyl and ester groups from C-18 alpha-glucosiduronate gave the alpha-glucosiduronic acid in 84% yield and the 17-isoaldosterone alpha-glucosiduronic acid in 12% yield. 7. The rate at which several types of beta-glucuronidase hydrolyse the foregoing steroidal alpha- and beta-glucosiduronic acids is given.

Aldosterone Glucuronide, an Important Biomarker: Synthesis and Structure Elucidation of Novel Isomers

“…Since the Koenig-Knorr reactiono f7 and 8 gave only al ow yield of 9b ( Figure 3) in ahardly reproducible way as described by Carpenter et al [10] we have investigated new approaches for the synthesis of aldosterone glucuronides. For this purpose, we used ap rocedure being developed by us for the preparation of iridoid glucosides using at rimethylsilyl tetraacetyl-1b-dglucoside.…”

“…Since the Koenig–Knorr reaction of 7 and 8 gave only a low yield of 9 b (Figure 3) in a hardly reproducible way as described by Carpenter et al [10] . we have investigated new approaches for the synthesis of aldosterone glucuronides.…”

“…The isomerization at C-17 was also am ajor dilemmai nt he synthesis of 3 by Carpenter et al [10] Thesea uthors solved the problem in ar ather complicated procedure by transforming the obtained glucuronide into ad ouble semicarbazone, fol-lowed by removal of the acetyl moieties and cleavageo ft he methyle ster as well as final acid catalyzed hydrolysis of the two semicarbazonem oieties. Unfortunately,n oN MR data of 3 prepared by this procedure have been reported;m oreover, NMR data of 3 do not exist in the literature at all.…”

“…Moreover,i ts chemical synthesis has also caused severe problems so far.T he first synthetic investigationsh ave been performed by Underwooda nd Frye [9] using aK oenigs-Knorr reaction of aldosterone 21-acetate 8,w hich however did not give the desired derivative of compound 3. Later,C arpenter andM attox [10] again used aK oenigs-Knorr reaction of 8 with at wentyfold excess of methyl acetobromoglucuronate 7 in the presence of at wentyfold excess of Ag 2 CO 3 via the formal carbocation 4 as ap roposed intermediate. They obtained am aximum of 10 %o ft he desired aldosterone-18-bd-glucuronate 9b and in addition the 18-a-d-glucuronate 9a as well as the 1-acetoxy-glucuronic acid methyl ester derivative 10.T he authors mentioned in their publication that the reproducibility of their process is low.W ehaverepeated the work of Carpenter and Mattox using as ynthetically acceptable approach with only at wofold access of 7 and were not able to isolate any of the desired aldosterone-18-b-d-glucuronate 9b.…”

“…The first synthetic investigations have been performed by Underwood and Frye [9] using a Koenigs‐Knorr reaction of aldosterone 21‐acetate 8 , which however did not give the desired derivative of compound 3 . Later, Carpenter and Mattox [10] again used a Koenigs‐Knorr reaction of 8 with a twentyfold excess of methyl acetobromoglucuronate 7 in the presence of a twentyfold excess of Ag 2 CO 3 via the formal carbocation 4 as a proposed intermediate. They obtained a maximum of 10 % of the desired aldosterone‐18‐β‐ d ‐glucuronate 9 b and in addition the 18‐α‐ d ‐glucuronate 9 a as well as the 1‐acetoxy‐glucuronic acid methyl ester derivative 10 .…”