4482
COMMUNICATIONS TO THE EDITOR
Vol. 83
structure B ; m.p. 269-273'; [ a ]+134' ~ (pyr.); E 25,900), and hydrogenated to yield 18-nor-D(m.p. 201Xz:P 241 mp, E 33,500; AXNuio' 3.00, 6.00, 6.10, homo-13(17a)-androstene-3~-ol-17-one 6.20 p ) by its infrared and ultraviolet spectra and 204'; [ a ] D -36'; 240 mp, E 16,200; Ax""'"' by chemical transformations. Compound I was 2.96, 6.05, 6.17 p ; reported13 m.p. 194-196'. Recently a free radical rearrangement which is acetylated readily with acetic anhydride in pyridine a t room temperature t o give the 11-acetate (m.p. formally of the Wagner-Meerwein type has been 239 mp, E 34,200; observedI4; we believe that the rearrangement 161-165'; [ a ] D +148'; AXNuio' 5.78, 6.00, 6.16, 8.00 p). Dehydration of I reported here is another instance of this reaction with perchloric acid in methanol gave 18-nor-D- type,. A plausible mechanism is indicated in homo-4,11,13(17a)-androstatriene-3,17-dione (par- partial structures E to H, the intermediate primary [ a ] -888'; ~ tial structure C ; m.p. 189-192'; AX",: 238 mp, E 15,200 and 280 mp, E 29,000; AANujol 6.00, 6.06, 6.20, 6.32 p ) which was hydrogenated to the known6 18-nor-~-homo-4,13(17a)androstadiene-3,17-dione(partial structure D ; m.p. A B 196-199'; [ a ] D +49; Xz;,.,"" 242 mp, E 33,600; XXNuiol 6.01, 6.18 p ; reporteda m.p. 196'; [a] -I-47'; ; : :A 243 mp, E 33,000) using palladium on calcium carbonate catalyst in benzene.? Analogously, 1,4-androstadiene-l1/3-ol-3,17-di' \ D c one,8 11P-hydroxyestrone 3-acetateg and andro~ t a n e - 3 6lP-diol-17-one ~1 3-acetate1° were converted to the corresponding 11-nitrites (respectively m.p. 109-111"; [a]D $179'; 239 mp, E 16,400; ) , ~ N u i o l 5.72, 5.76, 6.00, 6.10, 6 . 2 2 ~ ;m.p. 176-178'; [a]D +93; AX",: 268 my, E 1,340 and 275 mp, E 1,200; AANuiol 5.65, 5.72, 6.06, 6.20, 6.67, 8.3 p and m.p. 134-136'; [a]~ +69'; XXN"'"' 5.78, 6.08, 6.12, 8.12 p ) which were photolyzed to give, 18-nor-~-homo-1,4,13(17a)-androrespectively, statriene-llP-ol-3,17-dione(I1; m.p. 254-257' ; [ a ] D +35' (pyr.); "::A: 243 mp, e 32,600; AXNuio' 2.97, 6.02, 6.12, 6.18 p ) , 1S-nor-~-homo-1,3,5(10),13(17a)-estratetraene-3,ll/3-diol-17-one 3-acetate (111; m.p. 181-183'; [ a ] +117' ~ (pyr.); A":, H G 237 mp, E 19,500; AXNui0' 2.96, 5.70, 5.95, 6.15, radical F rearranging via the open chain inter6.28, 6.68, 8.1 p ) and 18-nor-~-homo-13(17a)- mediate G to the tertiary radical HI which then may androstene-3P,l l@-diol-17-one3-acetatell (IV; m.p. lose a hydrogen atom to a radical species in the 165-168'; [ a ] -3' ~ (pyr.); "::A: 241 mp, E system to give the final product B. 15,200; XXNuio'2.90, 5.75, 5.99, 6.12, 8.0~).Com(13) K.Miescher and H. Kggi, Helu. Chim. Acto, 32, 761 (1949). pound I1 was converted to the 11-acetate (m.p. (14) J. A. Berson, C. J. Olsen and J. S. Walia, J . Am. Chem. SOL., 199-201'; [ a ] D $80"; 240 mp, E 32,000; 82, 5000 (1960). AANujol 5.77, 6.00, 6.15, 6.24), and dehydrated to SCHERING CORPORATION HANSREIMANN give 18-nor-~-homo-1,4,11,13( 17a)-androstatetra- BLOOMFIELD, XEWJERSEY ALFREDS. CAPOMAGGI TOVA STRAUSS ene-3,17-dione (m.p. 200-202'; [&ID - 193'; P. OLIVETO EUGENE AX":", 240 mp, E 15,700 (shoulder) and 278 mp, RESEARCH INSTITUTE FOR MEDICINE AND CHEMISTRY E 34,000; AXNuio' 5.99, 6.16, 6.21, 6.29 p ) . D. H. R. BARTON 49 AMHERST STREET Further substantiation of partial structure B for CAMBRIDGE, MASSACHUSETTS the photolysis product is furnished by the following RECEIVED SEPTEMBER 29, 1961 reactions. Treatment of compound I V with perchloric acid or potassium hydroxide in methanol OF THE STRUCTURE OF A URINARY gave 11,13(17a)-androstadiene-3/3-01-17-0ne (m.p. CORRECTION METABOLITE O F ALDOSTERONE' 255-258"; [ a ] D -46'; 284 mp, e 27,800; Sir: AANujol 2.98, 6.08, 6.18, 6.32 p ) which was converted The major urinary metabolite of aldosterone, [a]D -22'; to the 3-benzoate (m.p. 220-223'; XX:;xOH 238 mp, E 14,200 and 283 mp, E 23,800; measured as the triacetoxy derivative, has been 283 mp, used to estimate the rate of secretion of the hormone reported12 m.p. 223'; [ a ] D -25'; in man.2,3 The structure 3aI18,21-trihydroxy(6) H. Heusser, J. Wohlfahrt, M. Muller and R. Anliker, Hela. 11,20-diketopregnane, previously4 assigned to the Chim. A&, 42,2140 (1959). (7) C f . R. B. Woodward, F. Sondheimer, D. Taub, K. Heusler and metabolite, however, was that of a contaminating
1
Po- \Po
W.M. McLamore, J . A m . Chem. S O L .74, , 4223 (1982). (1) This work was aided by Grant No. P-274 from the American (8) H. L. Herzog, C. C. Payne, M. A. Jevnik, D. Gould, E. L. Cancer Society, and by grants from the National Institutes of Health Shapiro, E. P. Oliveto and E. B. Hershberg, i b i d . , 77, 4781 (1955). and the John and Mary R. Markle Foundation. (9) B. J. Magerlein and J. A. Hogg, i b i d . . 80, 2220 (1958). (2) S. Ulick, J. H. Laragh, and S. Lieberman, Trans. Assoc. Amcr. (10) J. von EUWand T. Reichstein, Hcls. Chim. Acto, 26, 988 Phys., 71, 225 (1958). (1942). (11) Thh compound was isolated from the crude photolysfe p r e d ~ ~ d (3) C. Flood, D.S. Layne, 9. Ramcharan, E. Rossipal, J. F. Tait, and 5.A. TaIt, Acfa Endocvinolo~ica,86, 237 (1961). by partltion chromatography and fraetivael frystsllixntian~ Uliik ead 9. Liabarmsn, J , A m , Ch#nr, $99 , 19, 8617 (1967). (4) cia) s l w s P R W ~ 801,4w,
COJIMUNICATIOXS TO THE EDITOR
Nov. 5, 1961
substance, a metabolite of 18-hydroxycorticoster~ne.~ We wish to report the correct structure of the aldosterone metabolite as the 301,5p isomer of tetrahydroaldosterone (I). Tetrahydroaldosterone (301,5p) was prepared using a ~ o l u b l eenzyme ~~~ fraction. Male rat livers were homogenized in acetone a t -15O, the homogenate was filtered and the residue dried a t room temperature in vacuum. The acetone powder was suspended in buffer solution8 and the filtrate incubated with d-aldosterone-21 monoacetate9 in the presence of TPNH'O under nitrogen a t 37". Extraction and chromatography of the incubation mixture revealed only one substance (I), other than unchanged substrate, which reduced blue tetrazolium." Tetrahydroaldosterone (I) was isolated from extracts of combined incubates by chromatography on paper (ethylene dichloride-ethylene glycol, &ido = 0.312 and on Celite (toluene-ethyl acetate 9:1-methano1:water 1:1, R A l d o = 0.6) as a white amorphous solid, melting range 107-114,01a [ a ] " ~ 50' (CHC13 (C = 0.9), 2'c;:X: 2.78, 5.88 (weak) p ; C, 69.00; H, 8.88. Acetylation with acetic
+
OR CHZOR I I & o- . ,R o
@O
/O\
RO-'
I, R = H 11, R = AC
IV
111, R = H
V, R = A c
VI, R:H VII, R = AC
( 5 ) The separation of the metabolite of 18-hydroxycorticosterone from the aldosterone metabolite and the confirmation of its structure b y synthesis of the y-lactone obtained by periodate oxidation will be reported separately, S. TJlick and K. Kusch. In certain states of hyperaldosteronism such as cirrhosis of the liver and maligant hypertension, the secretion of 18-hydroxycorticosterone as measured by the excretion of this metabolite was greater than 4.0 mg. per day, exceeding that of aldosterone. (6) G. M. Tomkins, J . Bid. Chem., 225, 13 (1957). (7) J. T. August, Biochcm. Biophys. Acto, 48, 203 (1961). (8) p H 7.4 containing these concentrations of salts in m. moles per liter: NaCl 110, KCI 4.6, MgSOl 1.0, NaxHPOl 22. (9) We wish to express our appreciation to Dr. C. H. Sullivan, Ciba Pharmaceutical Products, for providing the d-aldosterone-21monoacetate used in this work. (10) Generated from T P N (triphosphopyridine nucleotide) by the coupled oxidation of glucose-6-phosphate in the presence of glucose-6phosphate dehydrogenase. (11) When cortisol was incubated with the same enzyme preparation, under the same conditions, the only product was 3a,17a,llp,21-tetrahydroxy-20-ketopregnane. (12) H values refer to the distance migrated by the sample on paper chromatogram relative t o the reference steroid. Abbreviations: Aldo 0 aldosterone, DOCA = desoxycorticosterone acetate, An
4-androstene-3,17-dione.
-
(13) Melting points (uncorrected) were determined on a micro hot stage. Only the hydroxyl and carbonyl regions of the Infrared are reported here. Elemental a n a l y s ~ swere performed by Sfhnsrzkopf Microandytical Lsboretoriar.
4483
anhydride in pyridine gave a t r i a ~ e t o x yderivative '~ (11) RnocA= 1.1in methylcyclohexane-formamide, = 1.3 in methylcyclohexane-methanol: water 4 : 1, 5.71, 5.77p, which like aldosterone15 readily lost one acetoxy group when treated with dilute acetic acid. Oxidation of tetrahydroaldosterone (I) with periodic acid yielded the theoretical amount of formaldehyde16 and a monohydroxy y-lactone (111), m.p. 252-254', 2.88, 5.64 M , C, 72.46; H , 8.46. This lactone (111) also was obtained from aldosterone etiolactone (IV) l7 using the enzyme preparation described above. Acetylation yielded the monoacetoxy dervative14 (V), RAn = 0.9 in methylcyclohexane-formamide, 5.60, 5.78 p . There was a single band of simple contour a t 8.08 p which was characteristic of equatorial 3-acetoxy steroids.I8 The 301,5/3 configuration was assigned to lacetone I11 by excluding the other possible 3-equatorial isomer. Hydrogenation of aldosterone etiolactone (IV) over platinum oxide in acetic acid gave the known 3P,501, lactone VI,1791g m.p. 241243O, which depressed the melting point of lactone 111. The infrared spectra of the pair of isomeric lactones (111 and VII) and of their acetoxy derivatives (V and VII) differed in the fingerprint region. Only lactone VI1 formed an insoluble digitonide. The metabolite of aldosterone (3.8 mg.), isolated as described4 from the urine (17 day pool) of a patient with cirrhosis of the liver, was compared with tetrahydroaldosterone (3cr,5p). Three derivatives (11, 111, V) of the isolated and the synthetic steroid were prepared. Both ketols as well as their corresponding derivatives had identical infrared spectra and chromatographic running rates. Both ketols also were shown to be identical by double isotope techniques. Following the administration of d-aldosterone-7-H3, the major radioactive moiety was isolated from the glucuronide fraction of urine and mixed with tetrahydroaldosterone (3a,5P)-4-C14,and derivatives 11, I11 and V of the doubly labeled mixture were prepared. The H3: C1* ratios of the keto1 and its derivatives agreed within 5%. (14) The acetate number was determined with "-labeled acetic anhydride. The number of moles of steroid present in the sample was determined from its Cl4 content. The c.p.m. per mole for tetrahydroaldosterone and its derivative was determined from the specific activity of aldosterone-4-Cl4 used as a substrate in the incubation. (15) E. A. Ham, R. E. Harman, N. G. Brink, and L. H. Sarett, J . A m . Chem. Soc., 77, 1637 (1955). (16) Measured colorimetrically with chromotropic acid: D. A. MacFayden, J . B i d . Ckem., 158, 107 (1945). (17) S. A. Simpson, J. F. Tait, A. Wettstein, R. Neher, J. v. Euw, 0. Schindler, and T. Reichstein, Helu. Chim. Acta, S7, 1200 (1954). (18) R. N. Jones and F. Herling, J . A m . Chum. SOL,78, 1152 (1956). (19) M. M. Pechet, R. H. Hesse and H. Kohler, ibid., 83, 5251 (1960). VETERANS L4DMINISTRATIONHOSPITAL
STANLEY ULICK
BRONX68, NEW YORK KATHRYN KUSCH DEPARTMENT OF MEDICINE J. T. AUGUST STANFORD UNIVERSITY SCHOOL OF MEDICINE PALO ALTO, CALIFORNIA RECEIVED SEPTEMBER 26, 1961
VINYL AZENE CHEMISTRY: FORMATION OF AZACYCLOPROPENE
Sir: As a continuation of our studies of the chemistry of the monovalent nitrogen speciea, ozenei,llathe
