June 5, 1956
COMMUNICATIONS TO THE EDITOR
2647
0 anhydride, a copolymer (V) was formed with an ab1 sorption spectrum containing the same peaks and x/x\O o.x\o an analysis indicating a ratio of 1.5 methacrylate units to one maleic anhydride. (Calculated for I1 I NX\ x\ \,o /x o / s (C2~H2306)~: C, 69.0; H, 6.01. Found: C, 68.94; x/o\&O H , 6.13.) The molar rotation per benzyl unit was I I1 I O // less than t h a t of the polymer, although little differ0 \\s o\x/o o/s\o ence would be expected if the rotation of the copolyI 0 mer was due solely to the methylbenzyl groupsI6: ' 0 \o 0 I ([&fIz5D- 1 2 8 O , a = -0.53', 2 = 0.5 dcm., c = S I o/s\o 2.1y0in dioxane). x/ \o-When the original asymmetric centers were reI i moved by phosphonium iodide, the reduced copolymer (VI), exhibited no characteristic absorption peaks a t the previously mentioned wave lengths and it showed optical rotation of the opposite sign: ([>f]25D +33.2, a = 1-0.22, = 0.5 dcm., c = C14-ergosterol,obtained from Saccharomyces cere2.2y0in dioxane). Although the analysis suggests more reduction visiae grown in the presence of carboxyl-labeled acethan expected (calcd. for (C10H1106)~: C, 52.9; tate, was converted to 3/3-acetoxy-l l-ketoergostane H , 4.85. Found: C, 54.32; H, 7.24), the only ex- (111)5-8and Cll and Clz obtained individually from planation for the observed rotations of the reduced I11 by the following series of reactions. For the and unreduced copolymer appears to be that asym- degradation leading to (211, I11 was converted to 3p, metry has been induced during the process of radi- 1l-diacetoxyergost-9(ll)-ene (IV)g which, in turn, was ozonized and saponified to yield 3P-hydroxy-9cal polymerization. keto-9,ll-secoergostane-11-oic acid (V) [m.p. 172.5(16) "Optical Activity and Chemical Composition," by H . Landolt, 173.5'; [a]"D -37' Chf.; c , 74.82; H, 10.75; translated by J. hIcCrae, B'hittaker 8r Co., London, 1899, pp, 131, 132. neut. equiv., 4491. The acid V was allowed t o react CHEMISTRY DEPARTMENT SICKY BEREDJICK with HN3 and Cll was obtained as COz. STATE USIVERSITY OF XEW YORK COLLEGE OF FORESTRY COSRAD SCHUERCH For the degradation leading to C12, I11 first was which was dehydrated SYRACUSE reduced to the 11/3-01 10, XEW YORK RECEIVED APRIL 19, 1936 t o yield 3/3-acetoxyergost-9(ll)-ene (VII).6 VI1 was oxidized with t-butyl chromate and the 3/3-acetoxy- 12-ketoergost-9(11)-ene (VIII) [m.p. 133.5T H E BIOSYNTHESIS O F STEROIDS AND TRITER134.5'; [a]"D f45' Chi.; 238 mK ( E 10,900); PENES. T H E ORIGIN OF CARBONS 11 AND 12 OF ERGOSTEROL C , 79.12; H , 10.621 then was hydrogenated over Sir: Pd-CaCOa to form 3/3-acetoxy-12-ketoergostane ~ Chf.; C, 78.31; The squalene hypothesis2 for the mechanism of (IX) [m.p. 174-175'; [ a ] 2 5+53 biosynthesis of steroids and triterpenes has been H, 10.831. IX was allowed t o react with perbeninvestigated widely and all the evidence obtained zoic acidlo and the 3/3-acetoxy-13-hydroxy-12,13has substantiated the ~ o n c e p t . ~Since the central secoergostane-12-oic acid 12,13 lactone (X) [m.p. two carbon atoms of squalene are derived from the 158-159'; [ c ~ ] * ~-D14' Chf.; c , 75.83; H , 10.331 carboxyl of a ~ e t a t e this , ~ symmetry presents a de- formed was saponified in aqueous methanolic KOH manding requirement for the concept of the utiliza- to yield 3~,13-dihydroxy-12,13-secoergostane-12tion of intact squalene (I) in the biosynthesis of ster- oic acid (XI) [m.p. 188.5-189.3'; [a]"D 0" MeOH; oids. These two central carbon atoms of squalene C, 74.36; H, 11.18; N.E. 4311. The acid XI was must be in the juxtapositions, C11 and CIZ, of ring C allowed to react with HN3 and C12 obtained as COz. It was found that both C11 and C ~ were Z derived of the steroids. Degradation of ring C of such a compound would yield information with regard t o from the carboxyl of acetate and were equally lathis unique symmetry requirement. The sterol beled (GI,13.5 dis./min./mg. BaC03; C12, 12.8 dis./ chosen to study was ergosterol (11) since the pres- min./mg. BaC03). The values found for the speence of a homoannular diene in ring B facilitates the cific activities of these carbon atoms were less than preparation of compounds required for the degra- that calculated (17.3 dis./min./mg. BaC03) on the basis of the squalene hypothesis. However, this dation of ring C. could well be due t o the large counting error (20y6) (1) This work was supported, in part, by Grant No. AT (I].]),
b
Project No. IO, E. S. Atomic Energy Commission. ( 2 ) R. B. Woodward and K. Bloch, THISJ O U R N A L , 76, 2023 (1953); W. G. Dauben, S. Abraham, S. H o t t a , I. L. Chaikoff, H. L. Bradlow and A. H. Soloway, ibid., 75, 3038 (1953). (3) H. N. Little and K. Bloch, J . Bioi. Chem., 183, 33 (1950); J. Wuersch, R . L. Huang and K. Bloch, ibid., 196, 139 (1952); J. W. Cornforth, G. D. Hunter and G. Popjak, Biochem. J . , 64, 597 (1953); I(. Bloch, H e l v . Chim. Arfn,36, 1611 (1953); D . J. Hanahan and S. J. Wakil, THISJ O W R S A L , 7 6 , 2 7 3 (19531; W. G. Dauben a n d K. H. Takemura, ibid., 76, 0302 (1953); J . W.Cornforth, Revs. P u r e A p p l . Chem., 4 , 286 (1954); W.G . Dauben and J. H. Richards, Chemistry and I?zdnst v y , 94 (1955); R . B. Clayton and K. Bloch, J . Bioi. Chem., 218, 305, 319 (19.56); T. T. Tehen and K . Bloch, THISJOURNIL,77, 0085 (19.53). (4) J W Cornforth and G . Popiak. Biochein J . . 68, 403 (1954).
~
( 5 ) m'. V. Ruyle, E . M. Chamberlin, J. M. Chemerda, G. E. Sita. L. h l . Aliminosa and R. L. Erickson, THISJOURNAL, 74, 5929 (19.51). (0) R. C. Anderson, R. Stevenson and F. S. Spring, J . Chem. SOL.. 2901 (1952). (7) E. M. Chamberlin, W. V. Ruyle, A. E . Erickson, J. M . C h e m erda, L. M . Aliminosa, R . L. Erickson, G . E. Sita and M . Tishler, THIS JOURNAL, 76, 3477 (1953). (8) P. Bladon, H. B. Henbest, E . R. H . Jones, B. J. Lovell, G. F Woods, G. W. Wood, J. Elks, R . M. Evans, D. E. Hathway, J. F. Oughton and G. H. Thomas, J . Chem. Soc., 2921 (1953). (9) A. Crawshaw, H . B. Henbest and E. R. H. Jones, ibid., 731 (1954). (10) E. S . Rothman, M. E . \Val1 and C. R. Eddy, THISJ O U R N A L , 76, 527 (1954).
COMMUNICATIONS TO THE EDITOR
2648
VOl. 78
resulting from the very low activity of the ergosterol (7.5 dis./min./mg. BaCOJ. It can be concluded that steroids derived from carboxyl-labeled acetate are labeled in the juxtapositions, CI1and C12, as demanded by the squalene hypothesis and such a result strongly supports a concept of the intact utilization of the acyclic triterpene, squalene. We wish t o thank Professor D. J. Hanahan of the University of Washington for kindly supplying the C14-ergosterol,Merck and Co., Inc., for a generous gift of ergosterol derivatives, and Dr. E. 11. Baker of the Radiation Laboratory, University of California, for the CI4determinations.
new compounds were prepared by condensing 3 to 5 parts of the appropriate amine3 with one part of 6-methylmercaptopurine in a sealed micro Carius tube heated t o 130 to 140' for 12 to 18 hours.' Excess solvent was removed under reduced pressure and the crystalline residue washed with cold alcohol and recrystallized from alcohol-water. Biological activity is expressed as the minimum concentration which will produce complete inhibition of visible tentacle formation after 18 hours a t 27". Relative activities are compared using adenine as a standard. All tests were conducted in a buffered (pH 7.4) solution containing all inorganic ions required for optimum rate of regeneration. The strong inhibitions obtained a t the very low CHEMICAL LABORATORY UXIVERSITY OF CALIFORSIA \VILLIAM G. DACBEK concentrations of the higher analogs suggest that BERKELEY 4, CALIFORNIA THOMAS LY.HUTTON they block a fundamental controlling process rather than the gross metabolism of the organism. CurRECEIVED -APRIL 9, 1956 rent investigations are directed both a t determining the structural specificity of the active compounds INHIBITION OF REGENERATION I N HYDRA BY CERand a t determining the system involved. full TAIN NEW 6-(PHENYLALKYL)-AMINOPURINES report of the synthesis and testing of these and other Sir 6-(substituted) purines is being submitted for publiMethods have been developed for quantitatively cation. studying the processes of regeneration in hydra, a BIOCHEMICAL IKSTITUTE AND THE RICHARD G. HAM5 primitive organism that may well serve as a model DEPARTMENT OF CHEMISTRY ROBERTE E A K I X system of development and cell differentiation in THEUSIVERSITPOF TEXAS,A N D CHARLES G. SKINKER higher animals.' Adenine and various adenine de- THE CLAYTON F O C S D i l T I O N FOR RESEARCH \VILLIAM SKIVE rivatives have been found to retard the formation of AUSTIS, TEXAS RECEIVED XPRIL2. 1956 new tentacles in hydra whose hypostome and tenta(3) 3-Phenylpropylamine, 4-phenylbutylamine and 5-phenylpentylcles have been cut away. In an attempt t o further characterize the nature of the effect, a variety of 6- amine were prepared by catalytic hydrogenation of t h e nitriles using Raney nickel. 5-Phen)-lvaleronitrile was prepared from 5-phenylva(substituted)-purines have been synthesized and leric acid kindly furnished by Dr. P . D. Cardner. 7-Phenylheptylatested. Most of the compounds are considerably mine also was furnished by Dr. Cardner, unpublished data. (4) G. B. Elion, E . Burgi and G. H. Hitchings, THISJ O U R N A L , 74, more active than adenine. One series in particular, the 6-(a-phenylalkyl) - 412 (1952). ( 3 ) National Science Foundation Predoctoral Fellow. aniinopurines, is extremely active, especially certain higher homologs (Table I). I n this animal system all members of the series are more effective A REARRANGEMENT INVOLVING A 1,5-PHENYL MIGRATION than the recently reported cell division factor, for plants, kinetin (6-(2-furfuryl)-aminopurine) ,z which Sir: has an activity only 20 times t h a t of adenine. The \Ve have observed t h a t S-benzhydryl-1 -naphthoic acid (Ia) isomerizes under Friedel-Crafts conditions TALBEI to a cyclic hemiketal (IIa). This reaction involves S Y N T H E S I S A N D A\CTIVITY O F 6-(w-PHEXYLALKYL)-AMINO- a 1,6-phenyl migration, and is the first example of a PURI\ES rearrangement of this type. 5linimum concn. Very few, if any, acid catalyzed reactions have for full Activity in been described in which an alkyl or aryl group is terms of Up., inhibition Compound Yield, adenine 70 OC. (dec.) ( p mole/ml.) 6-(R)-aminopurine transferred directly between carbon atoms that are not adjacently bound. -1case t h a t can be H-(Adenine) .. ... 5 .0 1 formulated conveniently as a 1,%methyl migration I-Phenylmethyl-" .. . . 0.18 30 has been reported' ; however, the possibility that 2 - P h e n y l e t h ~ l - ~ 69 239-240 .01 130 the product resulted from a sequence of conven3-Phenylpropyl-" 37 173-175 .02 250 tional 1,%migrations cannot be excluded. Re4-PhenylbutylLd 42 148-149 .003 1700 cently, .Ileinwald2 conclusively demonstrated that 40 145-147 ,003 1700 5-Phenylpentyl-" the isomerization of a-cinenic acid, a reaction for 54 112-113 ,001 5000 7-Phenylheptyl-' C. G. Skinner and \Y.Shive, THISJOURSAL, 77, 6692 which a 1,5-methyl migration had been proposed, (1955). Anal. Calcd. for CIIHI3X;: C, 65.25; H, 5.47. did not actually involve a methyl shift. Found: C, 65.11; H , 5.49. c .Anal. Calcd. for Cl4H1:S:: Compound I a 3 (1.00 g.) was converted to the acid dnill. chloride with thionyl chloride, then warmed with C, 66.38; E€, 5 9 7 . F(iund: C , 60.19; H, 5.74. Calcd. for ClaH1;Xj: C, 67.39; H , 0.41, Foulld: C, 67.13; H , 6.77. A n a l . Calcd. for C,sH19N5:C, 68.30; H , 6.81. 1.2 nil. of stannic chloride it1 20 1111. of carbon disulFound: C, 68.19; H, 7.15. J .Anal. Calcd. for CISH23S~: fide for ninety minutes. Upon hydrolysis and reC, 69.87; H, 7.49. Found: C, 69.99; H , 7.56. crystallization, 0.90 g. of IIa was obtained; 111.1). @
1:
11)
\V,A. \ i o - h e r n n d J .
(:'I
Ibor t h r im?i,ar )',I#! 58, 1 1 I
c'll,,
i:. C O Y l, ' r i i s J o i r x N , % i ,72,3701( 1 O i O ' l t h i i c o i n ~ ~ ~ ~ P~Ct \t' n < I1,:
I i . i v l i l I i i l t i l l itnil
