September 1967
and after 24 hr to 2x0 trip. So ;ippreclable change for i i w and useful aiialogs. Displacement of t'he occurred iii the spectrum during ail additional 16 hr of acetoxy group by nucleophiles2 has led t o the preparareaction time. Chromatography of the product O I L t'ioii of a large number of iiew compounds. HydroYilicic acid Jielded :I crystalline compound which \vagenolysis of the acetoxy group to yield the correspondassigned structure T7 on the basis of bpectral data (bee ing deacetoxycephalosporiris has been accomplished in Experimental Section and Scheme 11). The Rldehyde several i~ist~ances.~ Removal of the acetyl group of cephalosporin C: hy means of citrus acetylestera~e~ to give the correspodirig 3-hydroxymethyl compound \vas report,ed b), .Jeffery, et al.: This method has been riseti to prepare other deaccit\.l cephalosporinP.fi .\c\.lation of deacet!.lcephalosl)c,rjris ivit h aroyl halides ha+ I heeii reported h!. \'an Heyningeii.' The purpose COyCH, COZCHS o f the present report is t o describe the transformation V 111 o f the :IIC:\vas :30 pg/ml (gradient' plate iss say). In the mouse protection test against S f r e p t o c o m i s pyoyenea 1000 0 N &y~COCHa L&), the ED,,, \v:is greater t h a n 41.5 mg/kg X 2 COzNa (oral : i t i d suhcut:ineous) . I 2
H*NB>cHo I
'
Experimental SectionI3
COzK
C,OzCHs
0
"P 0
IV
c t r i i t a i r i i i i a i i t I J thv ~ iueth).l ester \\.as the Iactc~tie(IV)'" since it is k i i o u m that the cephalosporadesic :Ic;ids uiidergc~lactonizatioii \\-ith facilit!..' The lactone 11' ma!. he Formed also h!. cyclization ( ~ f111. 111 the oxidatiori of 111," after 1s hr the iiltraviolei :ihsorptiori m:tximiim at 265 mp ha(1 GhiEtetl to 275 mp,
A likelj.
(2) ( a ) C. \V, Hale. G.(+,E'. N r u ton, and 1,;. 1'. \ b r a h a m , B w r h r m . ./..79, 408 (IY61); th) F:. Van Heyninsen and i'. N. K r o u n . J . M e d . C h e m . . 8 , I7 I ( IRfiA) : c ) .1. I). Cocker, H. R . ('nwley. J. 3. G. Cox, 53. Eardley, G . I Gregory, .J. K . Lazenby, \ , 0 . Lone. . I . C'. P. Sly, and C . \ . Somerfirld. .I. ChPm. S o r . , ;701,5 (1965). ( 8 ) ia) R . H. Morin, K . G . .Iackwii, K . .I, .\Iueller, E , K . l.avagnino, \I, 13. Scanlon, and 8. L. .\ndreur, J . .in. Chem. Soc., 86, 1896 (1963); f b j tedman, K. S a e r e d , and .J. K. P:. Hoover, J . M e d . Chem.. 7 , 117 (1964,. E. 1.'. JansPn. R . I.ansr. and I.. R . 1IarDonnell. A r c h . Riochem.. 16, l l 5 (1017). t h ) .I. I>'.\. .Ieffery. F,:. 1'. . \ l ~ u i t l a maurl , (;. i;. t'. Neu-ton. H i o c h r m . J . . 81, 5 Y l (1981). i t i ) R. R. Cliauvette, E. H. FIynn, K. U. Jackson, E. K. Lnvagnino, K . ,Mueller, R . P. Pioch, R. W. Roeske, C. \V. R y a n , J. I.. Spencer, nnd E. Van Heyningen, ".\ntimicrobial .igents and Chemotherapy - 1962," .imerican Society for Microbiolony, Ann .Irbor. Mirh., 1963, p 6 R i . ( 7 J E. Van Heyningen, J . .Veri. Chem., 8, 22 ilQfi.5). ( 8 1 KeHinB, cephalothin. f 9) Deacetylcephalosporin* have twrn asrienerl the trivial name of ~,.phalosj)oradesicacids.7 I IO) R . R . Chaiivrttr. H I I ~P:, t l , E'lyiui, ./. . I f ~ ( i .P h r m . , 9 , i.41 I l Y t i A , .
Methyl 7 4Thiophene-2-acetamido)cephalosporadesate (111i. -To a wlutioti of 1.60 g of the potassium salt (11)prepared froni I as described hy \.a11 Heytiitigeri,' i i i 7,5 nil i i f HzO was added .io ml of ethyl acettite. The mixture was cooled in ice, qtirred vigoroilsly, i t n d the pH was adjiiGted t o 2.0 h>- the dropwise ;idditioti of I - \ HCI. The ethyl :icel:ite layer war qepamted After standing for i d treated a t oiice with ethereal CH,?;,. *everal mitirites, the soliitioii was evaporated to drynes:F irtider reduced pressure. The aolid residue amoruited to 1 . 0 i g. As d from the iimr spectrum of this material, it coiitaiiied I I ~111. The lactoile (IT.) appeared to be an impurity (see timi' data below). Attempts to purify 111 by recrystalliza1 ion or cnhromatography were tiot sLiccessfiil, and therefore the mide product was used for the iiext step. The crude prodiict exhibited A,,,;,, 236 mp ( 6 13,500) and 260 Inp ( e 7180); vmnx 2.75 (OH), 3.00 (NH), 5.60 (6-lactam C=O), 3.82 (ester C=Oj, 6.00 (amide I), and 6.51 p (amide 11): irmr (DJIF-d;) peaks at 6 ::.70, :S.X'2, :3.92 (C-2 CHz, ester methyl, and *ide-chaiii CH,j, 4.38 (doublet, J = 5.0 cps, C-3' CH,), 5.17 (douhlet, J = 4.5 cps, H-6: superimposed on this doublet were peaks assigiied to H-6 and the CH2 of the lactone ring of IT), 5.81 (quartet. J = 4..5 and 8.5 cps, H-i), and 6.95, i . 3 7 (aromatic protonsj. Attempts to purify the methyl ester (111)by recrystallimtiuii from aqueous acetone yielded a crystalline product which wa.: identified as the lactone IV: mp 23.5-238O dec 230-2:32'): hmxx 236 mp i e 12,800) and 260 mp ( 6 70'20); vmnl 3.10, 3.25 (NIT),5.56 (@-lactamC=O), 5.69 (lactuiie C=O), 6.00 (amide I ) xiid 6.X.i p (amide 11); iimr (DXIF-tl~)peaks at 6 3.88 (partially resolved doublet, C-2 CH, arid side-chain CH,), 3.12 (singlet, (11) 0. \lancerit. G. Rorenkranz, and F. Sondlieimer. J. Chem. Joc.. 2189 11953). ' 1 2 ) \ \ . \\-. M'ick, ".intiinicroliial .\gents and Cliemotlrerap~-lyH6,"
for hlicroliiology, .inn .\rbor, Micli., 1966, p 870. Melting points were determined on a Fisher-Johns apparatus and are uncorrected. T h e ultraviolet spectra were obtained in ethanol polution on a Car). M o d e l 14 recording spectrometer. The nmr spectra. tierc raken on a Varian .issociates Model HR-60 spectrometer. T h e chemical ;zliiftc a r e reported as 6 values in parts per million relative t o TYIH = 0. Tlie infrared rlw('tra ncrr drtermined in Niijol. .\ inerican Society f 13)
SOTES
9 tih
lactuiie CHz), 5.21 (doublet, J = 5.U cps, H-6), ti.UU (quartet, .I = 3.0 and 9.0 cps, H-7), and 7.00, 7.40 (aromatic protons). Methyl 3-FormyI-7-( thiophene-2-acetamido)-3-cephem-4carboxylate (V).-A mixture of 0.300 g of impure 111, 0.900 g of :tctivated MnOo,ll mid CHCl, ( 3 0 ml) was stirred at room tem: perature for 20 hr. Aii additional 0.450 g of N n 0 2 w a ~ added a r ~ dthe stirring wiis continued for ariothei, 20 hr. Aliquot haniples were taken after 18, 24, and 40 hi. The haniplex eshihited 1iv ahsorptioii i i i i CHCI,) at 27:, 280, iiiid 280 mu, i,e>pectively. due t o the cephalosporiii iiiiialeiia! and :it 241 nip due t o the thiophene side chain. The mixtiwe wah filtered a i d the filtrate was evapornted to dryi1ea.s. The residue was dissolved i i i CHCI, arid chrornatographed on Pi02 ~lIalliiickrodl,13 g ) . I t l u t i o r i with EtOAc-CHClr (1 : 3 ) afforded 0.050 g 11f \., inp from 2-propanol gave material 5 nip ( e 10,530). 2!)6 i10,730~. : ~ i t l :%IO ( ~ h ( i ~ 1 d e r ) : ~ ' (SIT)!3.39 (p-Iac*talll (I=O I, .-).SI(ester C=O), 6.OU (,amide I), 6 . 2 3 tdoirlile hoiicl), mid 6.23 p (amide 11); nnir (I)!dF-c/i) peaks at 6 3.03. 4.03 (C-2 CIf?. t,,.ier CIf8,, side-chaiii ( = € I , ) , 5.44 (doiihlet, ,I = (i.0 cpe, H-61, (\,I.-) (quartet, J = 6.0 aiid !).0 cps, H-7), 7.07, T.47 (aromatic protons), !I20 (doriblet, J = !J.O cps, S H ) , :uid 9.88 (singlet. ('I10 prcitoti). A >atisfactory mass spectriim w a , ~i r o t r~btained t'oi. this curnporuid diie l o thermal decrimpo4tiiiri of the 3:implr
\
to our hilowledge these derivatives have iiot tieeri
11SrO:>S:!: 0, 49.16: 1-1, 3. Found: C, 49.15: 11, 4.03; X, 7.18: S, lS.lX3, Ethylene Acetal of V (VI).--A mixture uf 0.090 g of Y,0.20 g of' c,thgleiie carbonate, (1.5 nil of ethyleiie glycol, 0.002 g of p-toluene,rilfoiiic acid moiiohyt1r:it e, :uid 2.,> ml of tetrahydi,ofuran was ;rllowed to starid a t room leinperatiire for 19.5 hr.l5 T h e mixtiire was t heii poiired into water a i d rstracted twice (Et70 Irt0.k~). The cL,mhiiied extracts were washed ( 1 .If S ~ I I C O B , twice with 1l20! oiice with >attirated XaCl), and lheii drietl Oc). Evaporatioii of the - C J ~ V ~ gave I ~ L 0.071 g of . d i d re,idire which was < tallized ftoni 2-propaiiol t o give 0.036 g ot' the ethylene acetal (\.I): mp 200-21).io dec: A,,,.,, 2XG nip L2!600) and 260 nip ( e ;:.BO i@-lactam ('~:-O), .j.77 (ester C'=-O), 6.04 (amide I ) , 6.20 (doiible boiitl'l, ;iiid fi.30 p (amide 11): iinir (CI)Cls) pe:ik,. at 6 3.43 ((*-2(:II!J. 17 (ncaetal Cl12,e v t n ' CII, i i i i d .ide-chaiii C H ~ I4.O.i , (diiiibIct, */ = 3.0 cp.., If-6); 5.74 (ciiiglet, 11-3'), >.SO (qiiartet, ./ und 8.0 cps, Il-7)> 6.41 (d(iiildel,.I = S . 0 cps, STT),a i i t l = ,i,(~
i)i
\ ' I were deterniiiiecl 1))- 11 . ~ ~ caicd ~ ~ ~fill,i r,7ir,,x20, . c , 410,f)6064.~"
Acknowledgment. -The author? wish I O cs1)ress t h e i r gratitude to H. Hoiiz. W. Hargrove. (;. 1laci:ik. 1). 0 . TYoolf, 1,. ( i . Ho\v:irdq and their associates for S ] J N ' t i ' d data; to 1). I,. Cline, e. Ashbrook, :iii(1 IY. 1,. R ~ ~ T Vfor I I mic.ro:tiiwlys~~s;to IV. E. Wipk ; i l l ( ] c'. \T. (;idzeski for iiiic~iobiological data: i i n ( l t r i ('. 1'. IT7:tltersfor thc eiizym:itica de:icetylatioii*. I i , .rile A?-aldeli>-iie~ Y I I \\a, l e p u r r e i 1 t u i i a \ e k:;;;'' P I ) % 111w i t l : ~ , ~ i o U ~ . I(. I],. \Vood\\ard, I.cinc. (111)
\V:L> prepared h\. tlic. S-carbos!,aiiti?,di,id~, procedu1.c. 111 our initial prep;iratioris, 111 \vas isolated : m I pwificd. It i w s then allorvcd t o rwet with CBZ-i)i,-u-nicthylglutamic anhydride5 i i i ; ~ j u e o u sdioxane :it pH S. Ho\T-ever, t h e purifiration \VU h t w found t o he iiiiiiece L ~ , I ' . 0 1 1 :djustirig the p H to 2 ;iiitl c?str:ictirig with e .I acetiitc, tallirie solid havi a trace of CBZ-a-meth\.lgliitamic acid :is impurity :i howl by thin lkiyer chronxitography. The blockin roup> of \' were remi)wd hy sodium i i i licliiid amnionin and the resulting thiol YI TW? purified t hr(>ughthe copperjl) d t . The g of \'I \vas c*t:ihlished bJ- its elemental anal metric :md base titratiori. The S-ethylm vative moved :is :1 single component b). elcctrophoicsi? bjp paper chromatography i i i
The nature of the glutamyl peptide link ( a os. is an important one. The use of glutamic anhydride generally leads to the formation of a n a-peptide as thr
K LRI. P F ~ ~ P 1E11K , i, \ i R ~ ~ L PHIRSCHMANN H
Merck Sha, p ik Dohnae Kesearch I,aboratories, Rahwcry, u! Jersey Rereizerl .lpril 6,1867
In recent years a-meth) Iamino acids have proven t o be of considerable interest in the study of enzyme
mechanisms' and in medicinal chemistry.2t2 However,
(1) 1'. b'adia, .\. Giariosio, and G. c f . tlammes, Rioch?niislr),, 5, Ill; (1966). (2) J. A. Oateb. L. Gillespie. S. Ldenfriend, a n d .
