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The following diagrams illustrate the method: Les cartes, planches, tableaux, etc.. peuvent etre filmis A des taux de reduction diffirents. Lorsque le document est trop grand pour etre reproduit en un seul clich*. il est film6 d partir de Tangle supirieur gauche, de gauche d droite, et de haut en bas. en prenant le nombre d'images nicessaire. Les diagrammes suivants illustrent la m^thode. 1 2 3 1 2 3 4 5 6 MICROCOPY RESOLUTION TEST CHART ANSI and ISO TEST CHART No 2 1.0 ^^ I.I 1 - 9 32 iiiji 2 2 2.0 1.8 1.25 1.4 i 1.6 js APPLIED IN/MGE Inc ^ ON THE GUANYLIC ACID OF THE SPEEEN 47996 W M.ll.K J0M;S as;, r., G. KOWNIKI !, FR'IM 1H» I.\EOI!\r"RV -1 PllV^lnioolCM ClUMlvTBV INTHF [•HIS' H..PKI\« V' !»..■ I HI \r.l. IV. Nc « H'^ 'LOGIC \I. CIIKMISIRV • ON THE GUANYLIC ACID OF THE SPLEEN. Bv WALTER JOXES and L. G. ROWXTKKE. (From the Laboratory oj Phviioloi^ical Chemistry in the Johns Hopkins V ni: :rsity.) (Receivcil for publication, February ni, n)o>.l Ten years ago, Ivar Ban,re]'arati"n emplovc.1 1)V V. 1-urih (propose.l by Bang and Raaschou-J leads to a miclcic'acid cf the ordinary type, but that by followmg the original method n{ Bang it is ].MSsible fi <.l.'ain fnan thox pan- creas a true guanylic aci.l, i. e., an organic ranjiound containing phosphorus which yields guanin but not adenin. Unfortunately Steudel agrees with v. Furth that guanylic acid contains no glvceritic group. ' It is the purpose of this eimimunication to show that not only is a true guanylic acid obtainable from <.x pancreas but iliat sub- stances (.f this class are conlined ncitherto a single animal species nor t'.> a single gland. Tin- prkp.'lR.xtion- of the xt'ci.HOPROTHiN- OF Pifi's splei;n-. Fourteen kilos of carefully trimmed and hnely grcnind fresh tissue were thoroughly mixed in portions with 21 liters of cdd distilled water, heated quickly to boiling and the solution liltereil off. It is immaterial whether the lluid be filtered hot or after cooling; in either case a fairly clear filtrate is obtained which by repeated filtration through the same hlter becomes almost as clear as water but ])ossesscs a very pale reddish tinge. If the residue be jiresscd through linen a milky fluid results which we have not been able to render clearer, either by repeated tiltration or bv long continued centrifugation; it is therefore advisable to avoid disturbing the residue in any way and to use n(i greater amount of water for the extraction than that stated. This part of the process was exceedingly more satisfactory than was the case with beef pancreas (see below), where we were never able to obtain anything better than a highly opalescent filtrate, although the solution obtained from this gland is described by various writers as perfectly v lear. The neutral fluid when cool was trcateil with acetic acid up to 5 to 10 per mille and the prcciT.itated gelatinous nucleojircleid was allowed to subside over niglit. Tins nu(le< .profcid is so 'StciK-k-l: /ritschr. j. physi,'t. Ch, it. liii. ]'■ 53'l. : Han^' and Raastliou: llofiiinstrr's !<< ilr.n'r. iv. y. 17;. Walter Jones and L. G. Rowntree 291 sirikm-lv dilTcrcnt in ,)hysira! vrnpcrtics fn.m the M,rrfsi,. ^ivlm- liravv'tliieeulcnt sul>^;anfe i>i <>x pannvas that wc can scarcely belicvc the tv/o substaiue^ i.lenii.Ml. The ch.u.ly lluid \vrs partly (lecantc.l ami the remainder shari.ly reiuovecl after (e:itnfut;a- tu.n. The micle.ipn-tei.l wliich in tliis c.mi.a. i form resetiibles a preparation of staivli paste, was freed fmrn soluble impurities by alternate solution in a minimal amount of caustic so.la and pre- cipitation with the requisite aiiKiUnt of acetic aci^l. Suspended matter was removed as far as possible from each alkaline solution by Ions continued cent rifugat ion, and in the same manner the precipi'tate.l nucleopr<,teid v/as sharply separated fr^-m the super- natant lluid. After this operation had been repeate id to anv of these tiltrates causes tm iiiimeditiie precipitation of -uanvlic acid. This dif- ference m liehavior of spleen ...uaiiylic aci-l ffoiil ptmcreas siKiny- lie acid, nii-ht he explained by d.ilierences in the solvent J.ower of the impurities in the two cases hut si-leen -uanylic acid retains this i.ropertv after purification lo >.ucli an extent that we are in. line 1 to t'hc opinion that tlio 1 wo nucleic acid^ are not iden- liiaP The ori-inal neutral solution .u'enenilly tillers Mowly hut eontmuouslv and as a deposition of -uaiiyhc acid is not likely, the slov.ncNS of the hltnition is with.out con e eow.ler soluble in hot water f..rming a transiKirent liquid whieh has an arid reaction to litmus. It responds neither to the buiret nfir to Millon's reaction bu' contains phosphorus and exhibi.s .ueneial ].roperties and reactions '.vlii( h closely accord with those whim Bang describes for the tjuanylic acid of the i)ancreas. THE PfKIX liASKS PRODfC K I) HV HYDROLYSIS or Tlli-: GUAN'YLIC ACIU Ol' I'IC.'s SPLKHN. Owing to the misf.irtune of not knowin- in the earlier part of our work that larf,'e (juantities of -uanylic acid may be recovered from mother liquors by thcadditi^ai of acetic acid, the amount of material at our disposal was insulficient for an exhaustive exam- ination such as we ^vould otherwise have made and as we intend to make in the immc^iiate future; so that we decided to devoie all of our material to a tinal decision of the very important (|ues- tion, whether or nc^t we are here dealing with a true guanylic aciil. A uram and a half ..f the material was heated for three hours ^vitlil^s cc. of 5 per ( ent sulphuri..' acid in a vessel submerged in boiling water. (Jn stan.ling over niglit the lluid deposited in ])rofusion macroscopic needles of guanin sulphate. These were dissolved by warming and the S(rlution was first neutralized and then treated with such an excess of ammonia that the fluid con- tained 2 per cent of the reagent. The jiroducl after digestion in the warm for an hour was allowed to cool and the p..-ecipitated guanin liltered off. After thoroughly washing in turn with i I)er cent ammonia and water the base was dissolved in i per cent caustic s(jda and again prcciidtated by the addition of acetic acid. The precipitate was filtered off, washed, dried ami weighed. For tlie separation (if guanin from small quantities of adenin there are two properties of the liases whiJi can be used ami can be thoroughly depended ujion. First, gu.anin is almost insoluble in 2 per cent ammonia while adenin diss.ilves in this reagent with comparative case. Second, both bases dissolve easily m dilute caustic soda but while guanin is quantitatively precipitated Guanylir Acid of the Spleen fn,m '^uJ^ a s,.lut,nn bv ac-cii-- a.ul, adcnin remain, mvhr th.^se f these methois were ar:«Hcd u, n.rn to tl,e case wh.ch w.. a descnbin,. The original amnv.n.aal f.Urate from .uan.n an. the acetic aci.l tlui-l ubtame-l in its puriiicaii..n were urn: e.l an,l treateortion failed to give a distinct pie- ciphate wi,h sih-er nitrate and ammonia whil.. in an^ 'her portion pi,ric acid did not even produce even an opalescence. I lu pu,v vUc nad of the sf^lccu gives no trace of ador.n. The main vield <,f guanin together with the small amou, obtained from the mother liqu-.rs weighed ;,„o ,m hgrams. 1 Zl issolved in hot 5 per cent hydrochloru- acid and decolorized with a small an.ount of animal charcoal. The solution on cool ng deposited the characteristic centimeter-long feathery nee. e guanin hvdrochloratc. Thesal, w,.s all-.wcd .-, drv ,n the an and analyzed' with the f..llowing results. , o ■ - w Rrani lost ocSo gram at , 00^ and rcuirc.l 706 cc. of stand- ard sulphuric acid >> cc. « 0.0077 gram ol ">\^"«^";. , ^^ ^,,^ ,. o.'u.M pram lost o. 0,^0.. gram at loc and rciu.red ,.H,, cc. ot same sulphuric acid. ,■ II s ., ,. IICI. .'11.. o: I. !, ir p; 11-- tiiT cent. \'.. .:a.:n " • ' -' ' It mav appear that our eonclusum. viz: that we are here deal- ing uith a guanvlic acid, is base.l ..n an experiment with rath r a ntall amount 'of material. In atiswer to such an objection .e Walter jcno and L. C Rcwntree 295 w,mM state that ^,c sh.,uM uso rm m,,re if wo wvr. tn n^eat the ,vnrk wuh an unUnnted stn-ply "( '"^"^"='1 ^'1 V^"" '■.'^l'"';'; , ''^ arc n,.v cn.auc.' in an inve^n.ati.n of nude.c acMs M .n- volves a lar,e't.un.bcr of ju.t such an-Oyses as tha, ,losonbcd uvl where the amount of material is of no consi.lcrai. on to us. Kx,;t- ri en e in these cases has tau.ht us tha, the best quantuat.v results can bo obtah^ed by us,n, no n.oro nuokno ao.l than w,ll pnuluoe 300 to 400 milligrams of the base sou-ht. ON THE msTRiiu-nox or (U-anvlic Ann in i.n: okoan.sm. We arc now ocoupicl with the oxanunatiou of . nundK," ot .lands for substances of this typo and hav. "";'"-;>,";;';; Substances whose ,hys.cal propert,cs curres,K:.nd ^'l'- -; those of KuanyUc acid. Thepig's pancreas yields ^ I-^'-"> ;^; aqueous extract from which acetic acid prccpuates a tloccu ont ,^ ,,U„,,,.,.,cid wlhch closely resembles the nucleoprotcd o ox ancrcls and from which a ,uanvlic a, id can be prepared .huh !,.„„„ be distinguished by anv apparent dhference from ox ,,,„,,vas ^uanvhc a.id. Wuh ox pancreas we exponencod oi- lid:rable dhhculty. The .lands used wore perfoctlv trosh and to „,ethod ,nvcn bv Ban. cdosely foU-.-od bu, m spue o, ex c eilort w;were unable to prepare any Mnn, approach.n, a d. u aqueous extra,,. However, the cloudv lUud ,u>ves a nucleop,o- tcid which in turn yu'lds a true Kuanvhc acid. Our results show conclusively , ha, ,uanv.. ac,d f. ,r t ho ,>,.>- H, ands') are considc,-ablv ,n.rc w.lolv d,s,nbu,cd , ,n a f,.rmerlv supposed and lend m -Mva, n,oasu,-c ,0 'l:o bohot h.a ,lu.se substances are conmu.n nuclear cons,„uen,s. As a . - - ,,,,n< >,f orduKU-v nu, Ice aod ln,herto prepa,-cd have boon lound ,„ ,,,,,1 „,,n,n as .oil as ,nan,n „ seems cor,a,n ,hat ,he .lands winch veld ,uanyhca.>d nu>s, also con,a,n e.thcr adcn,hc arid" ,',rnaclc.oa,,ds whuh pn.iuccbo,h bases. S,ncewri,m. the above ar„.]o our a„cnt,.m has been called t,, ,he work of Odcn.us^ who prepared .uanyhc a. .d Iron. ,he ,ni> loopnitiid .if ,he mammary .land. difrlnccs amen, the nucleic acids of this class, thus cstahhshu., a s.rus of jjuanvlic acids. 'Sec Mulys Jahn-sbrrulU. sss:s. u)oo. niEK!" or WII.I.IAMK * W1I.«IN» (•.Mr»VT BAI.TIMORR