2898
IIGXRYGIIXAN, LUDOVICO SANTUCCI, D. I-(phenylazo)-benzeneboronic acid anhydride was increased 20 to 2.7' by use of this longer coupling time. There was no increase in the amount of crude material. The longer coupling time increased the amount of crude 2-hydroxy-S-(p-bromophenylazo)-benzeneboronic acid anhydride by 22V0 and raised the melting point by approximately 45'. .Uthough this improvement probably carried through to the purified product, no such data were accumulated because of the difficulty involvetl in finding suitable purification procedures. Recrystallization1Jof these dyes irom an ethanolwater system, as was used by Snyder' for azoboronic acids derived from benzidine, failed to work satisfactorily. In most cases some purifica(lG) F. D . Ch.ittaa:i>- xnil I I
R . H i l l , J . C h c i i i . S u c . , 121, 2756
(1922).
(17) K . Illhs, 0 . IIirschel, 1'. Wagner. K , Himmlcr, !A.' T n r k , A lienrich :rnd Ii E , I,ehmann, J , prafct. C h e w , 108, 209 ( 1 9 2 I). 118) I n all cases ri-here piirification WR' accomplished I>?- recrvstallizatirin, the crystal f o r m , if a n y , \\-:IS rather indefinite. 'l'he f i r m ranged from a n apparently amoryhuus powder to an indefinite fibrous 1leedIP.
June 5, 1057
Azo DYESFROM 0-HYDROXYBENZEXEBORONIC ACIDANHYDRIDE
2899
tion resulted as noted by an increase in the melting had a melting point of 1S3-1S4° when the sample point over t h a t of the crude. However, the lack was placed in a previously heated bath and a meltof purity was well established by either the neutrali- ing point of 250-251' when the sample was imzation equivalent or quantitative boron analysis, mersed initially in the unheated bath. or both, on the dyes in this series. I n no instance was any effort made to prove the Dry ethylene chloride'g was found to be a good structure of the compounds other than from the recrystallizing solvent for the 2-hydroxy-s-(phenyl- information t h a t could be obtained from the inazo) -benzeneboronic acid anhydride, the 2-hy- frared spectra. This was believed to be adequate drox y-5- (9-bromophenylazo) -benzeneboron ic acid even though little is known about the exact posianhydride and the 2-hydroxy-5-(o-nitrophenyl- tion of the absorption bands for the azo group and azo) -benzeneboronic acid anhydride. Even though the boronic acid anhydride substituent. The first none of these compounds was exceptionally soluble of these possibly occurs a t approximately 6.35in the refluxing solvent, 1 g. to 1.5 g./liter, the 6.4 p,21,22although none of the spectra obtained material recrystallized readily and was far superior for these compounds showed any sharp absorption with respect to melting point and neutralization a t this wave length. Many of the boronic acid equivalent to material which could be obtained spectra show an absorption band a t 9.1-9.2 p Z o as from any other solvent or solvent system including do these azoboronic acids. This may be indicative benzene, benzene-petroleum ether (b.p. 77-1 Is'), of the carbon-boron bond, although this has not acetone-water and chloroform-carbon tetrachlo- been established definitely. The greatest use of ride. the infrared was to determine positions of substituBoth the m- and p-isomers of 2-hydroxy-5- tion on the aromatic rings as well as to prove the (nitropheny1azo)-benzeneboronic acid anhydride existence of the hydroxy, nitro and bromo groups were purified successfully using nitrobenzene as the whose absorption bands are well known. Acrecrystallizing solvent. The high boiling point of tually, the method of preparation should establish this solvent makes it difficult to remove from the the structure reasonably well, and the color should crystals, this being best accomplished by a washing give some evidence for the existence of the azo with petroleum ether (b.p. 28-38'), This partic- group while the neutralization equivalent should ular work was carried out before t h a t done with verify the presence of a boronic acid or anhydride ethylene chloride, and these particular compounds group. were never subjected to recrystallization from this Additional attempts were made to produce other solvent, although it is quite possible t h a t the boron-azo compounds, but it has not yet been posethylene chloride would be quite adequate. sible to purify these to an acceptable degree. In studying the purification of these azoboronic These compounds include those formed by couacids, the possibility of using chromatographic pling o-hydroxybenzeneboronic acid anhydride with techniques was investigated, most of the work be- diazotized sulfanilic and p-aminobenzoic acids and ing carried out on 2-hydroxy-S-(phenylazo)-ben- tetrazotized benzidine. zeneboronic acid anhydride and a smaller amount Experimenta123 on the 2-hydroxy-5-(p-bromophenylazo)-benzene2-Hydroxy-5-(phenylazo)-benzeneboronic Acid Anhyboronic acid anhydride, the 2-hydroxy-5-(m-nitrodride.-The diazotization of 18.6 g. (0.2 mole) of aniline phenylazo) -benzeneboronic acid anhydride and the was carried out according t o German Dyestuff inf0rmation.2~ 2 - hydroxy - 5 - ( p - nitrophenylazo) - benzeneboronic T h e benzenediazonium chloride solution mas then added over acid anhydride. This method looked very promis- a period of 30 minutes t o a stirred solution composed of 22 g. ing in the early stages, but it was not possible to (0.55 mole) of sodium hydroxide, 24 g. (0.2 mole) of 2-hyacid anhydride and 200 ml. of water, get material of a quality equal to t h a t which could droxybenzeneboronic cooled t o 5' by means of an ice-salt-bath; 200 g. of ice was be obtained by recrystallization from either added t o t h e reaction flask immediately before t h e additior~ ethylene chloride or nitrobenzene. The best re- was started. Agitation was continued for 11 hr. a t 0-5' sults were obtained using chloroform or a chloro- following completion of t h e addition. T h e reaction mass then filtered, t h e filter cake was suspended in 2000 ml. form-carbon tetrachloride system as the solvent was of water a t 40" and was acidified by t h e addition of lOcT, and eluent and a 1:2 mixture of Celite and silicic hydrochloric acid. After cooling t o room temperature, the acid as the adsorbent. precipitated solid was removed by filtration and was dried Melting points have long been regarded as a in a vacuum oven at 45'; 45 g. ( l O O ~ o ) of crude reddishmaterial having a melting point range of 178-188" criterion of purity for most organic compounds. brown was obtained. Acidification of t h e filtrate from t h e reaction However, this does not seem to hold true for azo- mass gave less than 1 g. of additional crude material which boronic acids and in general for organoboronic was combined with t h e bulk of the crude. Materials having the same neutralizaacids. l 5 t z 0 (21) L. J. Bellamy. "The Infrared Spectra of Complex Molecules," tion equivalent may vary considerably in melting John Wiley and Sons, Inc., New York, N. Y., 1954, p. 229. point. The rate of heating appears to be very im( 2 2 ) H. 31. Randall, R . G. Fowler, PI'. Fuson and J. R . Dangl, portant, much more so for this class of compounds "Infrared Determination of Organic Structures,'' D . Van Nostrand Inc., New York, PI'. Y., 1949, p. 5 2 . than for the usual organic compound. Also, a wide Co., (23) All melting points are uncorrected. I n taking t h e melting difference in melting point is experienced depend- points, t h e sample generally was immersed in a previously unheated ing on whether the sample is immersed initially in bath. However, when i t was noted t h a t t h e rate of heating affected an unheated bath or in a previously heated bath. t h e melting point appreciably, a previously heated bath was used. T h e neutralization equivalents were obtained by complexing the comFor example, partially purified 2-hydroxy-5-(@- pound with D-mannitol in a n ethanol-water system and titrating with broinopheny1azo)-benzeneboronic acid anhydride standard sodium hydroxide using a Beckman model G p H meter t o (19) Eastman Kodak white label 1.2-dichloroethane distilled over phosphorus pentoxide was used for this purpose. (20) L. Smtucci and H. Gilman, THISJ O U R N A L , 79, in p r e s (1957).
detect t h e end-point. (24) I. G. Farbenindustrie, British Intelligence Objectives Subcommittee, Report N o . 1348, p. 6 5 (PBS5593).
T h c crude material was washed with approximately SUO nil. of d r y ethylene chloride. This removed a large portion of t h e impurities, t h e washed material now having a melting point range of 210-220". This material was recrystallized three times from ethylene chloride t o give 14 g. (31%,) of bright yellow fibrous material melting a t 236-238". After each recrystallization t h e volume of t h e filtrate was reduced in order t o recover t h e maximum amount of material. T h e infrared spectrum supported t h e expected structure. A n d . Calcd. for C14H9BNPO2: B, 4.84; N, 12.51; neut. equiv., 224. Found: B, 5.08, 4.96; S,12.63, 12.42; iicut. equiv., 226, 227. 111 :iii experiment where a 3-1ir. cuupling time was employed, t h e crude yield was lO0':b and t h e material had a iiielting point range of 150-185'. A 9-hr. coupling time gave a 94i.h yield of material isolated in two portions; t h e first had a melting point range of 170-180° and represents 21.3$& of t h e yield while the second portion h a d a melting point range of 188-192" and represents 72.75(, of the 5-ield. Chromatographic purification of this dye was studied to :L rcasonable extent without any outstanding success being attaiiied. In one experiment, 2 . 2 5 g. of crude dye (n1.p range 188-19'7') was dissolved in 500 nil. of chloroform and \vas chromatographed on a column composed of 225 g. of a 1 : 2 misture of Celite aiid silicic acid. T h e column was cluted with additional chloroform. Kine fractions were collected. Fractions 1 through 3 were semi-solids and represetitetl 20';6 of t h e original weight. S o melting point was iibtaitied on this portion of t h e material. Fraction 4 which represented 11(,'' of t h e original weight had a melting point range of 140-150°. Fractions 5 through 8 all had melting points higher than t h e crude, fraction 6 having the maximum 1-angeof 204-209". This series represents 49'7;, of t h e origiiial weight. Fraction 9 was again a semi-solid, no melting point being obtained, and represents 4.5rzi of t h e original weight. Fraction 6 had a neutralization equivalent of 234 which indicates material of fair purity. T h e calculated value for t h e neutralization equivalent for t h e anhydridc is 224. T h e use of a 1 : 1 mixture of chloroforin aiid carbon tetrachloride solution a s t h e solvent and eluent was superior in one respect t o chloroform alone in t h a t sharper separations could be made b u t was inferior in another in t h a t tlie solubility o f t h e compound was less. 21 1: 1 mixture of Celite and silicic acid sliowed n o advantage while activated alumina was of no value whatsoever. 2-Hydroxy-5-(p-bromopheny1azo)-benzeneboronicAcid Anhydride.-Eight and six-tenths grams (0.05 mole) of phomoaniline mas diazotized according t o t h e method of Gomberg and Bachmann .25 This p-bromobenzenediazoiiium chloride solution was then added over a 10-minute period to a stirred solution composed of 5.5 g. (0.135 mole) of sodium hydroxide, 6.0 g. (0.05 mole) of %-hT.droxybenzeneborr,nic acid anhydride, 50 nil. of water and 50 g. of ice; and cooled in addition with an ice-salt-bath. iZgitatiori was continued for 9 hr. following completion of the addition, the temperature being maintained at 0-5'. T h e reaction mixture was filtered, t h e filter cake was suspended in 800 ml. of lvater a t 40' antl was then acidified by the addition of lOf;umber;: xnri W
1%;. Bachmann, 0i.c. .Syi:thr,i,~s, 8 ,
I
enipluycd, an 86;: yield of crude niaterial li:iviiig a melting point range of 130-141' was obtained. In the attempted chromatographic ~iurificatiui~ of illis compound, 12 g. of crude ( m . p . range 180-200°) \viis pmtially dissiilved in 500 ml. of benzene and was pouretl on :L column containing 4.50 g. o f a 1 : 2 mixture of Celite i t r i d silicic acid. The purest material which could be obtained OIL elution \Tit11 additional benzene had a melting point r m g e ( i f 210-220' which is far below t h a t of the recrystallized inaterial. Poorer separations were obtained when chloroform was substituted for benzene as tlie solvent antl eluent. mixture of chloroforin and carbon tetrachloritlc iI : I ) \vas superior t o chloroforni alone. 2-Hydroxy-5-( o-nitropheny1azo)-benzeneboronicAcid Anhydride.-Two anti eight-tenths grams (0.02 inole) of nnitroanilinc was diazotized according t o t h e method cuiployed by Smith antl Boyer.26 T h e filtered o-tiitrubeiizericdiazonium chloride scilutioii was added to a stirred solution composed of 2.76 g. (0.023 mole) of %-liytlrc onic acid anhydride, 2.2 g. (0.055 mole) ( i f X I ( mid 25 nil. of water cooled to 0--3" by IIIC';IIIS of an ice siltbath. Agitation was continuctl for 2 hr. follo\ring conipletioii of the addition, the tenipcrnturc beiug ~naint;tiiietl below 5". T h e reaction mass n w filtcred after xvhich the filter cake was suspended ill 200 nil. of water :inti acitlifietl by t h e addition of 10';;. hyclrcicliloric acid. The product was filtered, Tvashed and air-dried tu give 4 g. o f i i brov'n product having a melting point range of 1X7-li)8°. Thiy weight represents a i5':;,crude yield. .lcidification o f t h e filtrate from t h e reaction mixture gave a verJ- small atnotint of even poorer qualit!- material. T h e crude matcrinl \vas recrystallized tn.u times frolri benzene tu give 1 .3 g. i i f i i n orange colored solid melting a t 213-214". This weight rcpreients :I 24'~,; yield ( i f reasonably pure mitcrial. 'l'lle infr:iretl sliectrum supported t h e structure. i l n d . Caled. f i r CI?ll~13S.,01: S, 1 269. Found: iY,15.18, 15.24; neut. cqu i1fter i t was found tliat ethylene chlo crystallizing solvent for this t y p e of compriund, sonic o f tllr. benzene-recrystallized tnuterial, obtained in the expcrimcii t above, was also recrystallized from eth~-lenechloride. This gave orange colored material huviiig ;L melting poiiit uf 230 238" and a neutralizatioti equiv:tlc.nt closer t o the :in111 tlridc than tlie material which wis recrystallized friiin henzene. .Inn/. C:iletl. for C 1 ~ l f ~ B S d 013, 4 :4.02; S , 1.5.61; iteut. equiv., 269. Found: B, 4.46, 4,:ji'; S , 15.15, lt7.43; itelit. equiv., 280. I n aiiotlier cxperinictit wliere ;in 8-hr. coupling i i ~ n c\\:is usctl, a 78'J; yield of material having :I melting point r:iiigc o f 130-180° was obtaiiied. Recrystallization of this frijni ethylene chloride gave a 24' yield of materkil melting ovcr the range of 230-23.5'. XI) c1iri)iiiatographic purificntioii \v:ts attempted 011 this cvmpuund. 2-Hydroxy-5-(nz-nitrophenylazo)-benzeneboronic Acid Anhydride.-To a stirred solution of 6.00 g. (0,(J5 rncilc) o f o-hydrosybeiizeneboronic acid anliydritlc alitl 1l).O g. (0.2-7 inole) of sodium hydroxide in 50 nil. of Ivater, cuoled to 1) 5' in an ice-salt-bath, was added slowly a solution contaiiiing 6.9 g. (0.05 mole) of w-nitroaniline diazotized according to t h e method of Kaslow and Summers.28 The alkaline reaction mixture ( p H approximately 81 w:is stirred for 10 Iir. a t 0-5' and was then filtered. T h e orange filter cxke w a s suspended in 500 ml. o f water and was acidified b y the x l t l i tion of l0yo hydrochloric acid. -193': ( 1 2 . 3 i g.) yield ( i f material melting over t h e range of 168-180' was obtained. Acidification of t h e filtrate from t h e reaction mixture gavc no appreciable amount of additional material. T h e crude product was recrystallized twice from nitrobenzene, a Korit-A treatment being employed and care being taken not t o heat t h e solution above 95". Thorough washing of t h e recrystallized material with petroleum ether ( b . p . i 2 b ) P .4.S. Smith and J . 11. Buyer, i b i d , 31, 14 ( l % ? l ) .
(27) T h e calculated values shown in this analysis are for the anhydride At t h e time this material was prepared i t was believed t h a t the acid was being formed, b u t later work on boron-azo compounds of this type indicated t h a t i t a a s more likely t h e anhydride. T h c calculated values f o r t h e acid are: N, 14.64; neut. e q u i v . , 287. i 2 X ) C' I< Ii;islniv 2nd R 11. Summers, OI'R .Yyiilhcs?s, 3 3 , ,711 (lY.53).
June 5,-1057
STRUCTURE OF
CACTUS TRITERPENE MYRTILLOGENIC ACID
28-38') was done in order t o remove t h e nitrobenzene completely. -4 20.5% (2.74 9.) yield of yellow material melting a t 220-222" was obtained. T h e analytical sample was recrystallized a third time from nitrobenzene t o give material melting a t 221-222". T h e infrared spectrum supported t h e expected structure. Anal. Calcd. for C12HsBNtOa: C, 53.60; H , 3.00; N, 15.61; B, 4.02; neut. equiv., 269.06. Found: C, 53.73, 53.80; H, 3.27, 3.18; N, 15.39, 15.41; B, 4.00, 3.98; neut. equiv., 285.7. In another experiment where t h e reaction mass was stirred a t 0-5' for 8 hr., a n 82.5% yield of crude material melting over t h e range of 144-170" was obtained. This gave a 17.5yo yield of pure material. For best results, t h e optimum pH was found t o be about 8. Both sodium bicarbonate and sodium acetate were used a s buffers in some experiments, but they did not seem t o be particularly useful. Chromatographic purification was iried without a n y great success.
2-Hydroxy-5-(p-nitropheny1azo)-benzeneboronic Acid Anhydride.-To a stirred solution of 6.00 g. (0.05 mole) of o-hydrosybenzeneboronic acid anhydride and 10 g. (0.25 mole) of sodium hydroxide in 50 ml. of water, cooled in a n ice-salt-bath t o 0-5', was added slowly a solution of 6.9 g. (0.05 mole) of p-nitroaniline diazotized according t o t h e method of Ropp and C ~ y n e r .T~h~e alkaline reaction mixture (PH approximately 8) was stirred a t -10 t o 0' for 6 hr. and n a s then filtered. T h e brick-red filter cake was suspended in 500 ml. of water, and this was acidified by t h e addition of 10% hydrochloric acid. A 70% (9.35 g.) yield of material melting over the range of 184-200" was obtained. Acidification of t h e filtrate from the reaction mixture gave a n additional 0.51 g. (3.Sc;) of material having a melting point range of 205-225'. T h e combined crude materials could not be recrystallized directly from any solvent with much success. T h e crude was therefore extracted with a small amount of hot acetone and t h e residue was dissolved in 250 (29) G. A Ropp and E . C. Coyner. 01.n S w l h e s r s , 31, 80 (1931).
[CONTRIBUTION FROM
Terpenoids.
XXVII.
THE
2901
ml. of hot acetone. T h e insoluble portion was removed by filtration and t h e dye was reprecipitated from t h e filtrate by t h e addition of 900 ml. of warm water. T h e material, now having a melting point of 207-208", was recrystallized from nitrobenzene using t h e procedure described above for t h e m-isomer; 1.4 g. (10.4y0) of orange product melting sharply a t 241 3-242' was obtained. T h e analytical sample was recrystallized one additional time from nitrobenzene t o give material melting a t 242-242.2'. T h e infrared spectrum supported t h e expected structure. Anal. Calcd. for C12HsBN30a: N, 15.61; B, 4.02; ncut. equiv., 269.06. Found: N, 15.39, 15.16; B, 3.92, 4.22; neut. equiv., 290.1. In another experiment a n 84T0 yield of crude material was obtained when a n 8-hr. coupling time a t 0-5" was used. When t h e coupling time was increased t o 10.5 hr., t h e amount of crude material did not change, but t h e product melted a t 187' instead of over t h e range reported above. In this particular experiment 17y0 of t h e original o-hydroxybenzeneboronic acid anhydride was recovered. T h e pure yield was not determined exactly in these last two cases. T h e use of sodium bicarbonate a s a buffer did not show any advantages. All of t h e couplings were run a t a pH of 8. Chromatographic purification also was attempted on this compound, b u t i t did not work very satisfactorily.
Acknowledgment.--We wish to thank Mr. Robert McCord and Mr. E. Miller Layton of the Ames Laboratory of the Atomic Energy Commission for the infrared spectra. We also wish to acknowledge the support of the Division of Biology and hledicine of the United States Atomic Energy Commission who have made this work possible. The results of the biological testing of these compounds will be reported by Dr. Otho D. Easterday of the Brookhaven National Laboratory. AMES,IOWA
DEPARTMENT O F CHEMISTRY O F WAYPiE STATE UNIVERSITY]
The Structure of the Cactus Triterpene Myrtillogenic Acid' B Y CARL DJERASSIAND H. G. hfONSIMER3 RECEIVED JANUARY 9, 1957
Mj-rtillogenic acid has been shown t o be 3~,16~,28-trihydro~y-A~~-oleanen-29-0ic acid ( I a ) by various degradation reactions and by conversion t o longispinogenin triacetate ( X I I b ) .
During an investigation4 of the triterpene composition of the genus Myrtillocactus, there was isolated from several species in small amounts a new triterpene acid. Since this substance has not been encountered in any other genus of the cactus family, we have named it "myrtillogenic acid" and the present paper is concerned with its structure elucidation. Myrtillogenic acid was separated from the other triterpenes as the methyl ester (C31HS005) and then saponified to the free acid (C30H480b). Mild acetylation with acetic anhydride-pyridine of either the ester or the acid led to the corresponding triacetate, thus accounting for all oxygen functions. The ease of acetylation suggested t h a t the three hy(1) Paper XXVI, "Cactus Triterpenes," by C. Djerassi in "Festschrift Arthur Stoll " Birkhauser, Basel, 1957, pp. 330-352. ( 2 ) This investigation was supported by a research grant (No. RG3863) from the Division of Research Grants of t h e National Institutes of Health, U. S. Public Health Service.. (3) Taken from p a r t of t h e P h . D . dissertation of H. G. Monsimer. (4) C . Djerassi, S. Burstein, H. Estrada, A. J. Lemin, A. E. Lippman, A. Manjarrez and H. G. Monsimer, THISJOURNAL, in press
droxyl groups were either equatorially oriented or that one or more were primary. Membership in the P-amyrin class of triterpenes was indicated5 by the course of the selenium dioxide oxidation of triacetyl methyl myrtillogenate which furnished a Al1*l3(l8)-diene (subsequently shown to be 11) with the typical6 triple ultraviolet absorption maxima a t 240,248 and 258 mp. All of the triterpene acids which have been encounteredl so far among the Cactaceae contained the carboxyl group a t C-17, but two different lines of evidence could be presented t h a t this was not the case with myrtillogenic acid. Oleanolic acid (IIIa) (5) T h a t this is not a n unambiguous criterion. as had been believed earlier, was demonstrated recently in the case of the cactus triterpene dumortierigenin (C. Djerassi, C . H. Robinson and D. B. Thomas, ibid., 78, 5685 (1950)) which belongs to the p-amyrin series b u t does not react t o any appreciable extent with selenium dioxide. However. a positive reaction, as was observed with myrtillogenic acid, appears to be conclusive. (6) Cf. L. Ruzicka, G. Muller and H. Schellenberg, Helu. Chim. Acta, %a, 767 (1939); D. H. R. Barton and C. J. W. Brooks, J. Chem. Soc., 257 (1951).
