May 20, 1963
COMMUNICATIONS TO THE EDITOR
tive Pauly, Sakaguchi and Ehrlich positive spot with
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identical with paromomycin and paromomycin 11,
respectively. Rfl 0.61; single component on paper electrophoresis a t pH 3.5; amino acid ratios in acid hydrolysateser~.~~tyr~.g~-
R’”NH7NHR-I‘
but1.~3glu~.~0hisl.~4phe1.0garg3.04glyl.sllys~.80~r0~.0~vall.s1) was exposed to the action of anhydrous trifluoroaceticacid for ‘‘OR!! RO‘ 20 min. a t room temperature to give crude I which was OR‘ purified by chromatography on carboxymethylcelluIa-m losez2; ( [CX]’~D-61.7’ in 10% v./v. acetic acid; single ninhydrin, Pauly, Sakaguchi and Ehrlich positive spot Ia (Deoxystreptamine): R = R’ = R ” = R”’ = H Ib: R = R’ = H ; R ” = CHI; R”’ = COCHa with Rf20.7 X his; single component on disk electroIC: R = CH3; R’ = R ” = H ; R”’ COCH, phoresis a t pH 4.6; amino acid ratios in acid hydrolysate I d : R = H ; R’ = R ” = CHI; R”’ = COCH3 serl.97tyr0.9lbut0.94glu1.0~his1.~~p he~.98argz.91gly1.891YS3.31- Ie (Neamine) : R = Neosamine C; R’ = R” = R”’ = H pr02.~~vall.8~). The observation that leucine aminopeptiR = Neosamine C; R’ = Neobiosamine C ; If (Neomycin C ) : RO = R”’ = H dase digests of I contain glutamic acid and not glutaminez3demonstrates that the glutamine amide group is R = Neosamine C ; R’ = Neobiosamine B; Ig (Neomycin B): R ” = R”’ = H susceptible to hydrolysis by trifluoroacetic acid. Schwyzer, et al.,2 4 exposed N-t-butyloxycarbonylseryltyrosylIh (Paromomycin): R = n-Glucosamine; R’ = Neobiosamine B; R” = R//’ = H serylmethionylglutaminylhistidylphenylalanylarginylIi(Paromomycin 11): R = D-Glucosamine; R’ = Xeobiotryptophylglycyl - NE- t - butyloxycarbonyllysylprolylsamine C; R” = R”‘ = H (suggested) valylglycyl - Ne- t - butyloxycarbon yllysyl-Nf-t-butyloxyR = D-Glucosamine; R = R” = R”’ = H I j (Paromamine): carbonyllysylarginylarginylproline-t-butylester to the Ik (Kanamycin C): R = D-Glucosamine; R’ = R”’ = H ; action of trifluoroacetic acid and stated, without experiR ” = Kanosamine mental support, that this treatment did not remove the I1 (Kanamycin A ) : R = 6-Aminoglucose; R‘ = R”’ = H ; amide group from the glutamine residue. Our own reR” = Kanosamine sults indicate that the glutamine amide group in seIm (Kanamycin B ) : R = Diaminohexose; R’ = R”’ = H ; quences related to the N-terminus of the corticotropins R ” = Kanosamine undergoes significant hydrolysis on exposure to trifluoroThe work of Reeves5 provides a tool for the determiacetic acid. nation of the stereochemical relationship between the Acknowledgment.-The skillful technical assistance remaining adjacent hydroxyl groups of Ib, by measuring of Mrs. Chizuko Yanaihara, Mrs. Maria Gunther, the change in optical rotation, A [ M l c u p r a B, when Miss Priscilla Holland and Mr. John Humes is gratecuprammonium hydroxide solution (“Cupra B is fully acknowledged. substituted for water as the solvent. The above compound (Ib) has +9.8’ (c 0.6, (22) E. A. Peterson and H. A. Sober, J. A m . Chem. Soc., 1 8 , 751 (1956). H20) and [a]% +620° (c 0.47, Cupra B), giving (23) The enzyme preparation used in these studies fails to show glutaminase activity. A [ M ] ~ u p r aB = +1590. The high positive increment (24) R. Schwyzer, W.Rittel, H . Kappeler and B. Iselin, A n g c v . Chem., is similar to that observed with methyl 2-O-methyl-P72, 915 (1960). D-glucoside, I1 ( A [ M ] ~ u p r aB = +2190) but of OPBIOCHEMISTRY DEPARTMENT KLAUSHOFMANN posite sign to that of methyl 4-O-methyl-P-~-g~ucoside, UNIVERSITY OF PITTSBURGH ROBERTD. WELLS I11 (A[hl]cupraB = -2020).6 Thus, the adjacent SCHOOL OP MEDICINE HARUAKI YAJIMA hydroxyl groups in 6-0-methyldeoxystreptamine are PITTSBURGH, PENNSYLVANIA JOACHIM ROSENTHALER related as those a t C-3 and C-4 of the glucopyranose RECEIVED MARCH7 , 1963 ring,7 i.e., 6-0-methyldeoxystreptamine has structure Ib, rather than that of its mirror image IC. CHEMISTRY OF THE NEOMYCINS. XII’ THE H ’I)
ABSOLUTE CONFIGURATION OF DEOXYSTREPTAMINE IN THE NEOMYCINS, PAROMOMYCINS AND KANAMYCINS
NHAc
H N O H -
Sir:
Ho
The final uncertainty in the stereochemistry‘ of neomycin C, the absolute configuration of the unsymmetrically substituted deoxystreptamine portion of the antibiotic, now has been resolved. N,N’-Diacetyl-6-O-methyldeoxystreptamine (Ib),2-4 m.p. 280-284’ dec., [ a I z 3+5.0° ~ (c 0.64, water), was prepared from poly-0-methyl-hexa-N-acetyl-neomycin B. This compound has been isolated previously from neomycin B (reported m.p. 282-284’dec., [cY]’~D+12°),2 frOm paromomycin (reported m.p. 280-282’, [aIz7o +15°),3a and from zygomycins A1 and Az (reported map. 280-283’, [ r u I z 4 ~ +4°),3b which from their properties and degradation products are probably (1) Paper XI in this series: K . L. Rinehart, Jr., W. S. Chilton, M. Hichens and W. v. Phillipsborn, J. A m . Chem. Soc., 84,3216 (1962). (2) K. L. Rinehart, Jr., M. Hichens, A. D. Argoudelis, W. S. Chilton, H. E. Carter, M. Georgiadis, C. P. Schaffner and R . T. Schillings, ibid., 84, 3218 (1962). (3) (a) T. H. Haskell, J. C. French and (2. R . Bartz, ibid., 81,3483 (1959); (b) S.Horii, J . Anfibiofics(Tokyo), Ser. A , 15, 187 (1962). (4) The numbering shown is that defined recently2 for deoxystreptamine in neamine in order to place neosamine C on the lower numbered carbon. I t is seen in the present report t o have the additional advantage that it gives the lower number to the hydroxyl-bearing carbon (C-4) of absolute stereochemistry R.
Ac HO
CHsO Ib
H
k
I1
H
H
I11
(5) R. E. Reeves, Advan. Carbohydrafe Chem., 6 , 107 (1951). mol. wt. A[M1cupraB = ( [ a l i a 6 Cupra B [ a I 4 s s water) X 100 (7) The possibility of unexpected interference by the acetamido groups in the cuprammonium complex is eliminated by the observation that crystalline N,N,-diacetyl-5,6-di-O-methyldeoxystreptamine (Id, sublimes above 260’ without melting; A n a l . Found: C, 52.70; H, 8.24; N, 10.68; OCHa, 22.6), prepared from poly-0-methyl-tetra-N-acetylneamine, exhibited no enhanced rotation in Cupra B solution. (6)
-
-.
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COMMUNICATIONS TO THE EDITOR
Vol. 85
From this assignment, it follows directly that the Acknowledgments.-This investigation was supabsolute stereochemistry of deoxystreptamine in neported in part by research grant No. E-1278 from the amine is as shown in Ie. The neamine component of National Institute of Allergy and Infectious Diseases, neomycins B and C is identical and substituted deoxyPublic Health Service. We also wish t o express our streptamine structures If and Ig may be written for thanks to the Upjohn Company for neomycin samples, the antibiotics themselves. Since Ib is also obtained to Mr. W. S . Chilton for assistance in carrying out some from poly-0-methyl-penta-N-acetylparomomycin, 3a and of the experiments and, particularly, to Professor Gabor from the poly-0-methyl-N-acetyl derivatives of zygoFodor, who, during discussions in Budapest, raised the mycins AI and A23b(probably identical with paromomypossibility of application of Reeves' method to this cins I and 11), the substituted deoxystreptamines in problem. these antibiotics are I h for paromomycin-zygomycin AI, DEPARTMENT OF CHEMISTRY ASD CHEMICAL EKGISEERING OF ILLINOIS MARTINHICHENS UNIVERSITY and probably Ii for paromomycin 11-zygomycin A2. URBANA, ILLINOIS KENNETH L. RINEHART, JR. Moreover, the very recent isolation of paromamine8 RECEIVED OCTOBER 24, 1962 [Ij; 4-(2-amino-2-deoxy-a-~-glucosy~)-deoxystreptamine] from partial hydrolysis of kanamycin C9 estab1,1,1,4,4,4-HEXAFLUORO-2,3-DIPHENYL-2,3-BUTANEDIOL lishes the absolute configuration of deoxystreptamine in that antibiotic'" as in Ik. Also, since kanamycins Sir : A" and B,12 like kanamycin C,l0 contain kanosamine We wish to report the observation of a remarkable re(3-amino-3-deoxy-~-glucose) and deoxystreptamine, but sistance to pinacol-pinacolone rearrangement exhibited differ from kanamycin C in the replacement of its by a symmetrical alkyl aryl ethanediol. glucosamine fragment by 6-aminoglucose in kanamycin 1,1,1,4,4,4-Hexafluoro-2,3-diphenyl-2,3-butanediol (I) A and by an unidentified diaminohexo~e'~(on biowas prepared in 17y0 yield by the photopinacoligenetic grounds this should be neosamine C) in kanazation reaction between 13 g. of trifluoromethyl mycin B the absolute stereochemistry of deoxystrepphenyl ketone' and 75 ml. of 2-propanol in a quartz tamine in these antibiotics is almost certainly as in tube a t 5 cm. from a Hanovia 16A-13 mercury vapor 11 and Im, respectively. lamp. Distillation of the solvent and recrystallization With the assignment of this last stereochemical of the residue from 80 : 20 hexane-ether produced 2.16 point the structure of neomycin C is completed and g. of white needles. These were further purified by submay be written as shown. limation in vacuo; m.p. 155-156O. ( A n d . Calcd. for Cl6Hl2O2F6:C, 54.86; H, 3.45. Found: C, 55.10; H , CHzNHz 3.38.) HO The compound possessed an infrared spectrum, in KBr disk, completely consistent with its assignment ps Deoxystreptamine the pinacol: -OH, 3440-3600 cm.-'; -CF3,2 11651210 cm.-'; and the complete absence of carbonyl absorptions. O+NH~HO Oxidative cleavage of I with lead tetraacetate in boiling acetic acid gave only trifluoromethyl phenyl ketone, identified as its 2,4-dinitrophenylhydrazone : m.p. and HO mixture m.p. 108-109°; literature m.p. 106-107°.3 The pinacol was found to be completely resistant to Neosamine C >Ribose several concentrations of sulfuric acid in acetic acid a t rieornycin C steam bath temperatures. It could be recovered unchanged from 38y0 sulfuric acid after 4 hr. heating and Complete structures of neomycin B and paromomycin from 65% sulfuric after 6 hr. heating. I t resisted the await further evidence for the stereochemistry of neoaction of boiling acetic acid-0-naphthalenesulfonic acid samine B. The cuprammonium method provides for 30 hr. further evidence on this point, as well. As expected, The non-fluorinated analog of I , 2,3-diphenyl-2,3a high positive increment is observed in the rotation of b ~ t a n e d i o lprepared ,~ by the addition of excess methylmethyl N,N'-diacetyl-a-neosaminide C (a 2,6-diaminolithium to benzil, could be transformed into 3,3-diphenD-glucose derivative, with free hydroxyl groups a t C-3 yl-2-butanone by much less vigorous conditions: 0.6% and C-4) in cuprammonium B solution; A [ M ] ~ u p r aB p-toluenesulfonic acid or O.670 iodine in acetic acid = +2280". Of greater significance is the strong positive after 0.5 hr. reflux. increment of methyl N,N'-diacetyl-a-neosaminide B ; The resistance of I to acidic conditions substantially A [ M l ~ u p r aB = 1880". Though this observation does in excess of those required to rearrange similar diols may not of itself exclude L-talose or L-altrose stereochemistry be interpreted as a destabilizing influence of the powerfor neosamine B, it does exclude the five other isomers fully electron attracting trifluoromethyl group to the allowed by neosamine B's stereochemistry a t C-214,l 3 development of carbonium character on the hydroxyl(with the amino group on the right in the Fischer probearing carbons. Such carbonium character is generally jection formula) and is most consistent with the previously suggested2 2,6-diamino-2,6-dideoxy-~-idoseagreed to be a prerequisite to the rearrangement step.5 DEPARTMEXT OF CHEMISTRY WILLIAM X MOSHER stereochemistry. ~
HoGyH +
(8) T . H . Haskell, J. C. French and Q. K. Bartz, J . A m . Chem. S o r . , 81, 3480 (1959). (9) T. Wakazawa and S. Fukatsu, J. Anlibiolics (Tokyo), S e r . A , 16, 225 (1962). (10) M. Murase, ibid., 14, 367 (1961). (11) M. J. Cron, D. L. Evans, F. M. Palermiti, L). F. Whitehead, I . R . Hooper, P. Chu and R . U. Lemieux, J . A m . Chem. SOL., 80, 1742 (1968). (12) H . Schmitz, 0. B . Fardig, F. A. O'Herron, M. A. Rousche and I. R . Hooper, ibid., 80, 2912 (1958). (13) I. R. Hooper, personal communication. (14) K . L. Rinehart, Jr., A. L). Argoudelis, T . P. Culbertson, W. S. Chilton and K . Striegler, J . A m . Chem. Soc., 82, 2979 (19fiO). (15) T. H. Haskell, J. C. French and Q.R . Bartz, ibid., 81, 3181 (1959).
UNIVERSITY OF DELAWARE NEDD . HEINDEL~ XEWARK, DELAWARE RECEIVED MARCH 21, 1963
(1) The ketone was synthesized according t o the procedure of A . Sykes, J . C. Tatlow and C . R . Thomas, Chem. I n d . (London), 630 (1955). (2) I n agreement with previous spectral investigations of the -CFa group, an intense absorption band consisting of a series of multiple peaks is observed: 1.. J . Bellamy, "The Infrared Spectra of Complex Molecules," 2nd. Ed., John Wiley and Sons, Inc., New York, N. Y . , 1958, p. 330. (3) A . Sykes, J . C. Tatlow and C . R . Thomas, J . Chem. Soc., 835 (1956). (4) I). Cram and K . Kopecky, J . A m . Chem. SOL.,81, 2748 (1959). (5) J . F . 1)uncan and K. R . Lynn, Auslralian J . Chem., 10, 7 (1957). ( 6 ) N . S . F . Coiiperative Graduate Fellow, 1959-63
