R. B. TURNER, V. R. MATTOX, W. F. MCGUCKIN AND E. C. ?LEND ILL 1 CONTRIBUTION
Yol. 74
FROM SECTION OF BIOCHEMISTRY, AIATO CLISIC]
Steroids Derived from Bile Acids. XIX. Barbier-Wieland Degradation in the 11-Keto Series'
c.
B Y RICHARD B. TURNER, VERNONR. MATTOX, WARRENF. MCGUCKIN AND E D W ~ R D KENDALL RECEIVED bfAP 19, 1952 The degradation of methyl 3a, Qa-epoxy-ll-keto-12~-bro1nocholanate (I) to 3a-acetouy-11-ketoetiocholanicacid ( X I I I ) by a combination of the Barbier-Wieland and Hoehn-Mason procedures is described. The etio acid mas obtained in an over-all yield of 22%.
Prior to publication by hleystre and his associ- stance6 was dissolved in a mixture of acetic anhyates$ of their superior procedures for the degrada- dride and chloroform and treated a t 0" in a sealed tion of the side chains of bile acids, the more container with a large excess of hydrogen bromide. derivlaborious Barbier-Wieland method3 was employed The resulting 3a-acetoxy-11-keto-12a-bromo extensively in this Laboratory. By various modi- ative was re-esterified with concomitant loss of the fications of published procedures, we were able to acetyl group a t C-3 and methyl Sa-hydroxy-11increase the yields of the intermediate compounds, keto-12a-bronionorcholanate(IV) was isolated in and, despite the large number of steps involved, to S7% yield by crystallization from chloroformachieve a satisfactory method for the preparation petroleum ether. of derivatives of pregnan-20-one and of etiocholanic Degradation to the bisnor acid was carried out acid. Certain of these modifications appear to under conditions essentially the same as those dehave general applicability, and we wish to report scribed in the preceding paragraph. Addition of a in the present paper our experience with bile acids solution of IV in benzene to an ethereal solution of of the 11-keto series. the Grignard reagent resulted in separation of an Methyl 3a,9a-epoxy- 11-keto-1"a-bromocholan- insoluble complex, which, however, rapidly disate (I) was available as starting material for this solved as the solution was stirred. -liter 42 hours investigation. Conversion of this substance into a t room temperature the Grignard coniplex was methyl 3a-hydroxy-11-keto-12a-bromocholanatedecomposed, and the diphenylcarbinol was esby cleavage of the epoxide ring has been described tracted and simultaneously dehydrated and acetylin a previous comm~nication.~In practice, how- ated (C-3) in hot acetic anhydride. Oxidation ever, a satisfactory cleavage of the 3,g-cyclic ether followed by alkaline hydrolysis gave 3a-hydroxj--l1was obtained a t the noracid stage, and methyl ketobisnorcholanic acid (Vi in 74% over-all yield 3a,9a-epoxy-ll-keto-l2a-bromocholanate (I) with from IV. An additional 2y0 of V could be obtained the oxide bridge intact was accordingly employed by reprocessing the neutral material which rein the first step of the degradation. Debromination mained after separation of the acidic product. a t C-12 in a separate step was not necessary, since Methyl Xa-hydroxy-11-ketobisnorcholanate (VI) formation of the Grignard addition product was was obtained in 96y0 yield by esterification of V in accompanied by complete removal of the bromine5 methanol which contained S70 of hydrogen chloride. The solution was boiled under a reflux for 7 hours. a t this position. Our experience with preparation of the diphenylWhen a solution of I in dry benzene was slowly added to an ice-cold ethereal solution which con- carbinol from VI indicated that this step does not tained an excess of phenylmagnesium bromide, a proceed satisfactorily below room temperature or if homogeneous reaction mixture was obtained, which an insoluble complex is formed. Preliminary was allowed to stand a t 0". The addition product, investigation of the addition of V1 to phenyl3a,9a - epoxy- 11- ketonorcholanyldiphenylcarbinol, magnesium bromide in benzene-ether, benzene, m-as not usually isolated, but was dehydrated, in anisole and other solvents was not encouraging. boiling acetic acid to the corresponding diphenyl- In every instance a voluminous precipitate sepaethylene, which was then oxidized with chromic rated, and the yields of diphenylethylene obtained acid to give 3a,9a-epoxy-ll-ketonorcholanic acid after dehydration were poor, However, these (11) in an over-all yield of about 80% from I . difficulties were circumvented when it was disOxidation of the ethylene was carried out in a mix- covered that a mixture of benzene and N-ethylture of acetic acid, chloroform and water in the morpholine was a satisfactory solvent; the precipipresence of sulfuric acid. Esterification with tate which formed initially dissolved within 24 methanol which contained 1% of sulfuric acid hours. After 3 to 4 days a t room temperature, ice furnished the methyl ester (111), in 96% yield. and acid were added, the diphenylcarbinol was For cleavage of the oxide function the latter sub- extracted, extraneous volatile by-products jdiphenyl and so forth) were removed with steam and :1) The work reported in this paper was carried out in 1943-1944. :2) C. Meystre, H. Frey, A. Wettstein and K. Miescher, X e h . the diphenylethylene was formed in boiling acetic Chim. Acta., 27, 1815 (1944), and subsequent papers. acid. The product was treated with methanolic (3) (a) Heinrich Wieland, 0. Schlichting and R. Jacobi, 2. phrsiol. Chcm., 161, 80 (1926); (b) W. M. Hoehn and H. L. Mason, THIS alkali to hydrolyze acetylated material and to remove unchanged bisnor ester (IT) . CrystallizaJOURNAL. 60, 1493 (1938); (c) see also W. P. Long, C. W. Marshall and T. F . Gallagher, J . Biol. Chem., 166, 197 (1946). tion from methanol gave a 2% yield of 3cu-hy(4) R. B. Turner, V. R . Mattox, L. L. Engel, B. F. McKenrie and E. C. Kendall, ibid., 166, 345 (1946). ( 5 ) E.P. Kohler and If.Tishler, T ~ r JOURNAL, s 64, 1594 (1932).
(6) The ester is more soluble than the free acid and was preferred fr,r this reason.
Dec. 5 , 1952
DEGRADATION OF M E T H Y L 3 a,9 a-EPOXY- 11-KETO- 12a-BROMOCHOLANATE
droxy-ll-keto-22,22-diphenylA20(22)-bisnorcholene (VII). The acetyl derivative (VIII) was obtained in nearly quantitative yield by acetylation of VI1 with acetic anhydride and pyridine. Oxidation of the acetoxyethylene (17111) with chromic acid was unsatisfactory, but ozonizationSb proceeded smoothly when conducted in a mixture of ethyl acetate and methanol' a t about -10'. The ozonide was decomposed with zinc and acetic acid in the cold, and benzophenone was removed by prolonged distillation with steam. By the use of this procedure Sa-acetoxypregnane-1 1,20dione (IX) could be obtained consistentljin yields of more than 80%. Ozonization of the free hydroxy compound (VII) under the same conditions afforded 3ahydroxypregnane- 11,20-dione (X), but the quality of the product was poor and purification was more difficult than was that of the acetyl derivative (IX). Conversion of Sa-acetoxypregnane11,20-dione (IX) into Sa-acetoxy-1l-ketoetiocholanic acid (XIII) was accomplished by the procedure developed by Hoehn and Mason.3b A solution of IX in a minimal amount of methanol was treated with ap excess of benzaldehyde in the presence of sodium methoxide. The benzylidene derivative (XI), which began to crystallize in a few minutes, was isolated after 12 hours in 95% yield. This material was then reacetylated (95% yield), and, after ozonization in ethyl acetate-methanol followed by treatment with periodic acid, gave Sa-acetoxy-11-ketoetiocholanic acid (XIII). This yield was 86%, and the over-all yield of XIII from methyl 3a,9aepoxy- 11-keto- 1Pa-bromocholanate (I) was 22%. In the course of this investigation several derivatives of bile acids not hitherto described in the literature were obtained. The preparative methods employed are unexceptional and therefore are not described in detail. Data for these compounds are listed in Table I.
5815
DEGRADATIOK OF SIDECHAIX
1
C02CH3
3 steps
-+
I V I
J
H 11, R = H 111, R = CHI
I
I!
1, HBr 2, CHaOH, H'
I
V,R = H VI, R CHI
I
2 steps
VII, R = H VIII, R = CHaCO-
CH=CHCsH:,
I
Experimental6
C=O
3a,9a-Epoxy-l1 -ketonorcholanic Acid (11).The conversion of methyl 3u,Qa-epoxy-ll-keto12a-bromocholanate into 3a,9a-epoxy-ll-ketonorcholanyldiphenylcarbinolg and 3a,ga-epoxy1l-keto-24,24-diphenyl-~~~-cholene9 has been deI scribed. In the following procedure the intermediate carbinol and ethylene were not isolated. A solution of methyl 3a,9a-eposy-ll-keto-l2a- Ro A/ H bromocholanate (101.5 g.) in 700 ml. of benXI, R = H (7) These conditions have since been successfully emXII, R CHaCO-
d
ployed by Long, Marshall and Gallagher.% (8) All melting points were determined on the Fisher-Johns apparatus. We are indebted to Merck & Co., Inc., Rahway, New Jersey, and to Mr. William Saschek. Department of Biochemistry, Columbia University, for microanalyses. The yields reported in this paper represent that material which was satisfactory for use in subsequent experiments without further purification. Optical rotations were taken in chloroform ( c , 1) at 27 + 2' unless otherwise noted. (9) V. R . Mattox and E. C.Kendall, J . BioZ. Chcm., 186,689 (1850).
I S , R = CH3COS , R = H
COcH
I
HI04
CHSCOO'
J SI11
zene was added with shaking to 650 ml. of 1.88 molar phenylmagnesium bromide in ether. During the addition the flask, which was provided with a calcium chloride tube, was immersed in an ice salt bath and the temperature was kept below 5'. The reaction mixture was then stored at 0' under nitrogen. After 4 days10 the solution was poured (IO) I t was later shown that formation of the Grignard reaction product proceeded very fast and was complete in 15 minutes.
5816
K.R. TURXER, v. R.3ZATTOX, w.F. M C c U C K I N TABLE
SELV
Compound
AND
Vol. 74
1
DERIVATIVES OF BILE A C I D S
Sdvent"
E. c. KENDALL
hI.p., 'C.
Methyl 3a-acetosy-11-keto-12 ~ - b r o n i o - Chf.-P.e. 233-233.5 norcholailate 3a-Acetoxy-11-ketoiiorcholai~ic acid E.-P.e. 162.5-163.5 3a-Hydroxy-11-ketoiiorcholanic acid Dilute hIeOH 2.53.5-255 3.11-Diketonorcholanic acid Chf.-P.e. 202-204 Methyl 3a-hydroxy-11-ketonorcholanateE.-P.e. 124-124.5 3,1l-Diketobisnorcholanicacid r.:.-r'.e. 308-209 * Chf. = chloroform; E. = ether; P. e. = petrolerun ether. * In
[a]~ "C. ,
-5f2
Carbon, % Calcd. Found
61.05 Br. 15.62 +78f-2 71.74 +64=!~2~ 73.36 f663~2 73.76 73.81 +39f3 73.*30 methanol.
60.88 15.30 71.62 73.10 73.49 74.05 73.18
Hydrogen, tr, Calcd. Found
7.69
7.87
9.15 9.64 9.15 9.81 8.95
9.11 9.58 8.83 10.10 8.84
on a mixture of ice and 160 ml. of 12 N hydrochloric acid, decomposition of the excess acetic anhydride with ice and and the aqueous phase was separated and extracted with ben- water, the chloroform layer was separated, and the aqueous phase was extracted with chloroform. The chloroform soluzene. Steam was passed through the organic phase until crystals formed and about 2.5 liters of water had condensed. tions were combined, washed with water, dried over anhyThe crystals were filtered off, dissolved in benzene, and the drous sodium sulfate and diluted with 250 ml. of methanol, solution was dried by filtration through a pad of sodium to which 5 ml. of acetyl chloride was added.'* After 18hours a t room temperature, the solution was diluted with a large sulfate and then concentrated to dryness. DehydrationThe crude carbinol was dissolved in 1375 ml. of hot acetic volume of water, the chloroform layer was separated, the acid and the solution VTXS concentrated until about 100 ml. aqueous phase was extracted with ether, and the combined organic phase was washed successively with dilute alkali, of distillate had been collected. The solution was then refluxed for 2.25 hours. Oxidation.-The solution of the water and a saturated solution of sodium chloride. The crude diphenylethylene was cooled, 868 mi. of alcohol-free solution was dried over anhydrous sodium sulfate, concenchloroform and 217 ml. of'water were added and the flask trated and diluted with petroleum ether. The p r o d y t was was placed iu a bath a t 17 . While the mixture was being obtained in three crops: I, 17.96 g., m.p. 155-15: ; 11, stirred mechanically, a solution of 217 ml. of 10.2 N chromic 2.50 g., m.p. 151-154'; 111, 0.54 g., m.p. 150-153 ; total acid in 85% acetic acid was ad$d a t such a rate that the crude yield, 21 .OO g. (87%). Several recrystallizations temperature remained below 20 . The solution was stirred from chloroform-petroleum ether furnished the analytical for 2 hours a t 18-20', after which 217 ml. of 10 .2' sulfuric sample, m.p. 155-156', [LY]D -25 i S o . acid in 85% acetic acid was added in five equal portions xt Anal. Calcd. for CaHa704Br: C, 61.40; H, 7.95; Br, ten-minute intervals. The solution was stirred for an addili.02. Found: C, 61.46; FI, 7.71; Br, 17.04. tional 2 hours a t 18-20' and poured into about 7 liters of 3a-Hydroxy-11-ketobisnorcholanic Acid (V).-Methyl water. The orgaiiic phase was separated and the aqueous phase was extracted with two 500-ml. portions of chloroform. 3a-hydroxy-11-keto-12a-bromonorcholanate(I\') (60.63 g.) The chloroforin solutions were combined, washed with ivater was dissolved in 650 ml. of dry benzene and added slowly to 650 1111.of an ether solution which contained 1.30 moles of and concentrated to an oil. Crystals formed on addition phenylmagnesium bromide. During the addition the flask of benzene; yield 66.4 g., m.p. 138-159'. The filtrate was cooled in an ice-bath. The voluminous precipitate was concentrated to dryness, the residue was dissolved in LThich separated was dissolved by agitation. The flask ether and the ether solution was extracted three times with was closed and allowed to stand at room temperature for 0.5 N potassium hydroxide. The alkaline extracts were 42 hours. The contents were poured over a mixture of ice combined, aaidified with hydrochloric acid and the solution was extracted with chloroform. The chloroform solution aud 400 ml. of concentrated hydrochloric acid, the aqueous was extracted with ether and the combined organic was concentrated to dryness and crystals (3.9 g., m.p. 1,57- phase 1.58") were obtained from benzene. The nor-acid sepa- fractions were washed with dilute hydrochloric acid and The solvents were evaporateti, and the residue subr,ites from benzene with solvent of crystallization and usu- water. jected to a rapid curreut of steam. The solid phase was ally melts with effervescence at about llOo, recrystallizes dissolved in ether, filtered through anhydrous sodium sulxiid remelts at about 157-159". A sample of the acid which concentrated and refluxed for 3 hours with 250 ml. of had been crystallized from benzene and air-dried overnight fate, acetic anhydride. The material obtained after removal lost 9.18% a t 100" and 0.1 rnm.; calculated for 0.5 CsHs; of the acetic anhydride under diminished pressure was dis9 45%. The solvent-free product melted at 1.59.5-160 solved in a mixture of 570 nil. of acetic acid, 280 ml. of alnithout previous softening; [ N ] D +90 2". cohol-free chloroform, 85 ml. of water and 85 ml. of a 10 'V Anal. Calcd. for C2{lL4O4:C. i 4 . i G ; 1-1, 9.1.5. I ~ i ~ t i ~solution l: of sulfuric acid in acetic acid. Addition of 140 i d . C, 73.84; €1, 8.93. of a 9 iT solution of chromic acid in 85% acetic acid was Methyl Ja,Pa-Epoxy-l I-ketonorcholanate (111:. -A XJh- made a t such a rate that the temperature of the reaction tion of 70.3" g. of I1 in 1050 tnl. of methanol which con- mixture did not rise above 65' and, after addition vas cointained 10.5 ml. of concentrated sulfuric acid was refluxed for plete, the solution was stirred for 35 minutes during which tinic the trmperature fell to about rid". A large volume of 2 hours, cooled and diluted with 192 ml. of water. The product which separated was filtered off and washed with rvnrer was then added, and the aqueous phase was extrltctcd ivater; yield: 62.16 g . , n1.p. 124..7-125.5". The filtrate twice with chloroform and once with ether. The orgaiiic was concentrated to about 4011 nil, and the crystals which fractions were combined, washed several times with rate1 separated were recrystallized from dilute methanol; yield and extracted with 1 liter of 10% potassium hydroxide ioltltion in 5 portions. The alkaline liquors aere a1lou.i.d to 3.02 g., m.p. 123-124'. T h i s represents a yield of 96% in the esterification and an over-all yield of 77.5% from I. The stand overnight a t room temperature in order to remove the analytical sample, m.p. 125-125.5", [a]D +87 =k 2", ITRS acetyl group a t C-3, and acidified with dilute hydrochloric acid. The precipitate which formed was extracted with prepared by several recrystallizations from methanol. Anal. Calcd. for C?,H&: C, 74.19; IT, 9.34. Found: chloroform, the chloroform solution was dried, concentrated, and after dilution with petroleum ether the product C, 74.52; H, 9.51. was separated in 74'% yield in 3 crops: I , 32.61 g., m.p. Methyl da-Hydroxy-1 l-keto-12~-bromonorcholanate 231.,5-233'; 11, 1.79 g., m.p. 226-228"; 111, 0.21 g., 224(IV).-Twenty grams of methyl 3a,ga-epoxy-ll-ketonor- 2Zti". .4 saiiipk for analysis was purified bv several recholanate (111), 25 ml. of chloroform and 25 ml. of acetic crystallizations from acetone; m . ~ 231-232' . (rep~rted'~ anhydride were placed in a bomb tube and cooled in a Dry 195" and 223-225'), [ a ]+41 ~ i2 " . Ice-bath. Dry hydrogen bromide (95 g.) was condensed in Anal. Calcd. for C2ZH~*0,: C, 72.89; H, 9.34. Found: the reaction mixture and the tube was sealed. After 24 ' the tube was opened a t Dry Ice temperature C, 7239; H , 9.70. hours a t 0 and the bulk of the hydrogen bromide was allowed to evapo(12) Karl Freudenberg and Wiihelm Jakob, Ber., 74 [III, 1001 rate as the solution x-nrmed to room temperature. After (11) The crystals Contained 6.7% benzene.
(1041) ( 1 8 ) I,. H. Sarett,
J. B i d . Chrm.. 16'2.
601 (1946).
Dec. 5 , 1952
DEGRADATION OF h 1 E T H Y L ~CX,~CY-EPOXY11-KETO- 12CY-BROMOCHOLANATE
5817
The neutral material which remained after extraction of ing bath until the starch-iodide test was negative (about 10 the acidic oxidation product mentioned in the preceding minutes), the excess zinc was filtered off and the solvents paragraph was treated with methanol and acetyl chloride, together with the by-product, benzophenone, were reand the benzophenone was removed by steam distillation. moved by steam distillation. The residue was dissolved in The residue (8.1 g.) was re-treated with phenylmagnesium ether, and this solution was washed with dilute hydrochloric bromide, dehydrated and oxidized by the procedure de- acid, water, dilute sodium hydroxide solution, dried, conscribed in the preceding section. This afforded an addi- centrated and diluted with petroleum ether. The product tional 2.33 g. of 3a-hydroxy-11-ketobisnorcholanicacid, weighed 2.90 g. (81%) and melted a t 127-128'. The analytm.p. 227-229', and increased the yield to 76.6%. ical sample was recrystallized from ether-petroleum ether, m.p. 129.5-130' (reportedI4 132-133"), [ a ]+I30 ~ i 2'. Methyl 3a-Hydroxy-11-ketobisnorcholanate(VI).-3aHydroxy-11-ketobisnorcholanic acid (V) (7.55 g.) was disAnal. Calcd. for C23H3104: C, 73.76; H, 9.15. Found: solved in 100 ml. of 8% methanolic hydrogen chloride and C, 74.06; H, 9.37. the solution was refluxed for 7 hours. After dilution with 3a-Hydroxypregnane-11 JO-dione (X).-Five grams of water and extraction with chloroform-ether, the organic 3a-hydroxy-11- keto - 22,22 diphenyl Am(,*) bisnorcholene phase was concentrated and the product was crystallized (VII) was ozonized as described in the preparation of I X . from chloroform-petroleum ether. Two crops, I , 7.24 g., The Droduct was obtained bv crvstallization from etherm.p. 189-191°, and 11, 0.27 g., n1.p. 188.5-190°, were ob- petrdeum ether in two crop; 1,*2.60 g., m.p. 157-162'; tained (96%). The analytical sample, prepared by re- 11, 0.20 g., m.p. 155-158' (81%). A pure sample, m.p. crystallization from chloroform-petroleum ether, melted a t 174-174.5' (reported" 172-174'1, could be obtained only .. 191.5-192° (reported13m.p. 193O), [ a ]+38 ~ f 2'. after several recrystallizations. The substance was iden-Anal. Calcd. for c231i1604: C , 73.36; H, 9.64. Found: tical with that obtained by hydrolysis of the acetyl derivaC, 73.46; H, 9.82. tive (IX). 3a-Hydroxy-11-keto-22 ,22-diphenyl-A2°(22)-bisnorcholene Anal. Calcd. for C21H3203: C, 75.86; H, 9.70. Found: (VII).-A solution of 30.89 g. of methyl 3a-hydroxy-11C,75.56; H, 9.43. ketobisnorcholanate (VI) in 75 ml. of dry benzene and 150 3a-Hydroxy-? l-benzylidenepregnane-ll,20-dione (XI) ml. of dry N-ethylmorpholine was added slowly to an ice- Ten grams of 3a-acetoxypregnane-1 I ,ZO-dione (IX) was cold slurry of Grignard reagent prepared by dilution of 325 dissolved in 30 ml. of absolute methanol with warming. ml. of 2.48 molar phenylmagnesium bromide in ether with The solution was cooled t o room temperature, and 6.8 ml. 150 ml. of benzene and 150 ml. of N-ethylmorpholine and of freshly distilled benzaldehyde and 14.2 ml. of a 4.35 N removal of the ether under diminished pressure. A large solution of sodium methoxide in methanol were added. amount of insoluble material was present a t the end of the The container was sealed, allowed to stand overnight at 5' addition. After 3 hours a t about 5' and 21 hours a t room and the product which separated was filtered and washed temperature the precipitate dissolved. During this interval with methanol; yield 10.29 g., map. 216.5-218.5'. Diluthe flask was shaken occasionally. At the end of 84 hours tion of the mother liquor with water, extraction with ether a mixture of ice and 250 ml. of concentrated hydrochloric and removal of the excess benzaldehyde in a current of acid was added, the aqueous phase was extracted with ben- steam gave an additional 0.35 g. of material, m.p. 212-215' zene, the benzene solution was washed with dilute hydro- (total crude product, 10.64 g., 95%). A sample prepared chloric acid and water, concentrated and volatile by-prodby recrystallization from benzene-petroleum ether melted ucts were removed with a jet of steam. The residue was a t 22&221', [a]D $57 f 2'. dried, dissolved in acetic acid and the solution was refluxed. Anal. Calcd. for C~sHaeOs: C, 79.96; H, 8.63. Found: After 3 hours the acetic acid was concentrated under re- C, 80.33; H, 8.89. duced pressure, diluted with ether and washed with water. ~21-Benzylidenepregnane-3,ll ,ZO-trione.-This compound The ether was removed, the residue was taken up in 100 ml. of methanol and treated with a solution of 50 g. of potassium was prepared as a derivative by oxidation of X I with a hydroxide in 200 ml. of methanol. After two days a t room slight excess of chromic acid in chloroform-acetic acid at 0'. The trione melted a t 190.5-192.5', [ a ] +67 ~ i 2', after tempetature, the mixture was diluted with a large volume of water extracted with ether. The ether extracts were washed recrystallization from benzene-Detroleum ether. Anal. Calcd. for C2&L403: C , 80.34; H, 8.19. Found: with water, dried over anhydrous sodium sulfate and the ether was replaced with methanol on the steam-bath. The C. 80.20:. H.. 7.90. product was isolated in two crops, I, 24.93 g., m.p. 239.53a-Acetoxy-2 l-benzylidenepregnane-ll,2O-dione(XII).241', and 11, 3.45 g., m.p. 235-238' (72%). From the The 3a-hydroxybenzylidene derivative (XI) (4.48 9.) was fraction obtained by acidification of the potassium hydroxide acetylated by treatment with acetic anhydride and pyridine solution 1.20 g. of 3a-hydroxy-11-ketobisnorcholanicacid a t room temperature. The product was isolated in the (V) was recovered. The diphenylethylene (VII), recrystal- usual manner and was obtained by crystallization from lized from methanol and from benzene-petroleum ether, chloroform-petroleum ether in two crops, I , 4.36 g., m.p. melted a t 24&241', . [. a. l +270 ~ & 4'. 205-207', and 11, 0.29 g., m.p. 197-199' (95%). The Anal. Calcd. for C34H4202: C, 84.60; H, 8.77. Found: analytical sample melted at 205-206.5'. [a]D +93 f 2'. C, 84.42; H , 8.99. Anal. Calcd. for CJOH~SO~: C, 77.89;-H, 8.28. Found: 3a-Acetoxy-11-keto-22,22-diphenyl-Am(**)-bisnorcholene C, 77.94; H, 8.18. - 3a-Acetoxy-11-ketoetiocholanicAcid (XIII).-Two molar (VIII).-A sample of 24.28 g. of VI1 was acetylated at room temperature in 75 ml. of acetic anhydride and 75 ml. of pyri- equivalents of ozone was passed into a solution of 4.35 g. of (XII), disdine. After 18 hours the material was separated in the 3a-acetoxy-21-benzylidenepregnane-11,2O-dione usual way and crystallized from ether-petroleum ether. solved in 75 ml. of ethyl acetate and 75 ml. of methanol cooled to -7" in a salt-ice mixture. The ozonide was deThe product which separated (25.6 g., 97%) melted a t 202203.5'. Recrystallization from methanol gave crystals composed in the cold with 4.0 g. of zinc dust, 20 ml. of acewhich were solvated, m.p. 132-135' (effervescence). The tic acid and 5 ml. of water, and after the starch-iodide test sample for analysis, m.p. 204-205', [ a ] +266 ~ i 2', was was negative, the mixture was filtered, the organic phase was washed with water, dried and concentrated under reobtained by recrystallization from chloroform-petroleum duced pressure. The oily yellow residue was dissolved in ether. Anal. Calcd. for C36H4403: C, 82.40; H, 8.45. Found: 75 ml. of acetic acid and treated with 11.00 ml. of 7.35 N aqueous periodic acid a t room temperature. After 3 hours C, 82.63; H, 8.97. the solution was diluted with a large volume of water, ex3~-Acetoxypregnane-11,2O-dione(IX).-A solution of tracted with ether, the ether solution was washed with 5.0 g. of 3a-acetoxy-ll-keto-22,2Z-diphenyl-A~~**)-bisnorwater and the steroid acid was extracted with ice-cold 1% cholene (VIII) in ,io ml. of ethyl acetate and 50 ml. of meth- aqueous sodium hydroxide solution. Acidification of the anol was cooled to -10' in a salt-ice-bath and treated with alkaline aqueous solution and extraction with ether afforded 3 molar equivalents of ozone. During the course of the re- XIII, which was obtained from ether-petroleum ether in action, when some of the starting materials crystallized two crops: I, 2.42 g., m.p. 215-218'; 11, 0.62 g., m.p. out, the flow of ozone was stopped and the material was re- 212.5-216' (86%). An analytical sample, m.p. 220.5dissolved by stirring a t about 0'. When the addition of ozone was complete, 3.0 g. of zinc dust and 20 ml. of 75% (14) J. von Euw, A. Lardon and T. Reichstefn, Helr. Chim. Acta, acetic acid were added, the mixture was stirred in the cool- 27, 821 (1944).
-
-
-
.-
5818
V. R.
hfATTOX,
R. B. TURNER, W. F. MCGUCKIN, E. J. H. CHU
222' (reportedla m.p. 219-22l0, reported's m.p. 223225O), was obtained by crystallization from chloroform-.
AND
E. C.KEXDALL VOI. 74
petroleum ether.
for C Z 2 ~ 3 2 0c, 5 :70.18; H , 8.57. Found: H' 8'52' ROCHESTER, MINNESOTA
-4nal.
" 70'34;
(15) J. von Eum, A. Lardon and T. Reichstein, ibid., 97, 1287 ( 1944).
:COSTRIBUTION FROM SECTION OF BIOCHEMISTRY, MAUOCLINIC]
Steroids Derived from Bile Acids. XX. Degradation of 3a,9a-epoxy-ll-KetonorAcid' cholanic Acid to 3~~,9a-Epoxy-ll-ketoetiocholanic BY VERNONR.
JfATTOX,
RICHARD B. TURNER, WARRENF. MCGUCKIN, EDITHJ. H. EDWARD C. KENDALL
C H U AND
RECEIVEDMay 16, 1952 Methyl 3a,9a-epoxy-ll-ketonorcholanatehas been degraded to 3a,ga-epoxp-l I-ketoetiocholanic acid by a combination of the Barbier-Wieland and Hoehn-Mason procedures in a yield of 36%.
Cleavage of the oxide ring in methyl 3a19a- ether was inert toward oxidation it was advanepoxy-11-ketocholanate with hydrogen bromide tageous to degrade the side chain to the etio acid and degradation of the 3a-hydroxy steroid thus before restoration of the reactive hydroxyl group obtained to give 3a-acetoxypregnane-11,20-dione a t C-3. However, in a study of the cleavage of the and 3a-acetoxy-11-ketoetiocholanicacid have been oxide ring a t the nor, bisnor and etio ester stages reported.2 Degradation of the side chain was it was found that there was a progressive decrease achieved by a modification of the Barbier-Wieland in the reactivity of the oxide ring toward hydrogen method, and the pregnane was converted into the b r ~ m i d e . ~The conditions which were satisfactory acid etio acid by the Hoehn-Mason scheme. This for the ester of 3a,9a-epoxy-ll-ketonorcholanic paper describes the use of these procedures for con- failed to open the oxide completely when the ester version of methyl 3a,9a-epoxy-ll-ketonorcholanate of the etiocholanic acid was used; more severe conditions3 led to formation of the 3,12a-dibromointo 3C Y , 9a-epoxy- 11-keto etiocholanic acid. 11-keto derivative of etiocholanic acid. OH
Experimental4 3~,9~u-Epoxy-l I-keto-23,23-diphenylnorcholan-23-01 (II).-A solution of 19.49 g. of methyl 3a,9a-epoxy-l1-ketonorcholanatein 100 ml. of benzene was added slowly with mechanical stirring t o 200 ml. of an ethereal solution of 1.5 M phenylmagnesium bromide in a flask immersed in a bath a t -20'. The solution of steroid was added at such a ra!e that the temThe reaction perature in the flask was 0 to 3 mixture was stirred for 3 hours at 0", the flask was sealed and stored in an ice-bath. After 24 hours the salution was poured over ice and ammonium chloride, the organic phase was washed and concentrated to dryness and the residue was crystallized from acetone-methanol. The volatile products were removed from the filtrate by steam distillation and more material was seDarated from acetone-methanol: total yield 81%: The analqical sample melted at 152-153', [a]D+61 1. .4nal. Calcd. for C86H4403: C, 81.99; H , 8.65. Found: C, 82.22; H, 8.55. Ja,Qa-Epoxy-23 ,23-diphenyl-A2*--norcholen11-one (III).-A solution of 5.39 g. of 3a,9cuepoxy 11-keto- 23,23-diphenylnorcholan-23-01 in 10 ml. of acetic acid was refluxed for 1 hour and poured into water. The precipitate was collected, dried and recrystallized from acetonemethanol; wt. 5.02 g. (97%), m.p. 136-139:. The analytical soample melted a t 136.5-137.5 . [ol:D 1-172 i 2 . Anal. Calcd. for CzhH1202: C, 84.97; H , 8.56. Found: C, 85.12: H, 8.61.
.
OH
*
XVA, R = H IYB, R = CH,
VI
$€I3
CH=CHCeHs
c-=o
c=o
' '-\ ,-
VI1
I
I
I
--+
' A /--
\TI1
-
COIR
4
\\
+
I
i /'-
IXA, R = H IXB. R = CHs
(1) The work reported in this paper was carried out in 1943-1946. (2) R. B. Turner, V. R. Mattox, W. F. McGuckin and E. C. K e n dall, TEISJOURNAL, 74, 5814 (1932).
to Mr. William Saschek, Department of Biochemistry, Columbia University, for microanalyses. Optical rotations were taken in chloreform (c
-
1) a t 27
2O
i
