CIHM Microfiche Series (Monographs) ICMH Collection de microfiches (monographies) Canadian Institute for Historical Microroproductions / Institut canadien de microreproducbons historiques ^iogo Technical and Bibli' .: ..phi:; Notes / Notes techniques et bibliographiques The Institute ' ^s attempted to or. cX, Va c^st original copy available ior filming. Feat; , ... cf th:c oopy whi„h may be bibliographically unique, vvri.^ ray a;ter any of the images in the reproduction, or A^^ich may significantly change the usual method of timing are checked below. n n □ Coloured covers / Couverture de couleur Covers damaged / Couverture endommagee Covers restored and/or laminated / Couverture restauree et/ou pelliculee Cover title missing / Le titre de couverture manque Coloured maps / Carles geographiques en couleur Coloured ink (i.e. other than blue or black) / Encre de cculeur (i.e. autre que bleue ou noire) Coloured plates and/or illustranons / Planches el^ou illustrations en couleur Bound with other material / Relie avec d'autres Documents Only edition available / Seule edition disponible Tight binding may cause shadow? or distortion along interior margin / La reliure serree peut causer de I'ombre ou de la distorsion le long de la marge inteneure. Blank leaves added during restorations may appear within the text. Whenever possible, these have been omitted from filming / II se peut que certaines pages blanches ajoutees lors d'une restauration apparaissent dans le te.xte, mais, lorsque cela e'ait possible, ces pages n'ont pas ete filmees. L'Institut a microfilme le meilleur exemplaire qu'il lui a ete possible de se procurer. Les details de cet exem- plaire qui sont peut-etre uniques du point de vue bibli- ographique, qui peuvent modifier une image reproduite, ou qui peuvent exiger une modification dans la metho- de normale de filmage sont indiques ci-dessous. I i Coloured pages / Pages de couleur I I Pages damaged / Pages endommagees n Pages restored and/or laminated / Pages restaurees et/ou pelliculees D □ Pages discoloured, stained or foxed / Pages decolorees, tachetees ou piquees Pages detached / Pages detachees Showlhrough / Transparence Quality of print varies / Qualite inegale de I'impression Includes supplementary material / Comprend du materiel supplementaire Pages wholly or partially obscured by errata slips, tissues, etc., have been refilmed to ensure the best possible image / Les pages totalement ou partiellement obscurcies par un feuillet d'errata, une peiure, etc., ont ete filmees a nouveau de fa^on a obtenir la meilieure image possible. Opposing pages with varying colouration or discolourations are filmed twice to ensure the best possible image / Les pages s'opposant ayant des colorations variables ou des decolorations sont filmees deux fois afin d'obtenir la meilieure image possible. n □ Additional comments / Commentaires supplemenlaires: Pagination is as follons: p. 239-29b. This Item IS filmed at the reduction ratio checked below / Ce document est filme au t.iuK de reduction indiquo cidessous. 10x 14x 18x 22x 26x 30x i 12x 16x 20x 24x 28x 32x The copy filmed here has been reproduced thanks to the generosity of: J.J. Talfivan Regional Collection, D.B. Weldon Library, University of Western Ontario The images appearing here are the best quality possible considering the condition and legibiliry of the original copy and in keeping with the filming contract specifications. Original copies in printed paper covers are filmed beginning with the front cover and ending on the last page with a printed or illustrated impre«- sion, or the back cover when appropriate. All other original copies are filmed beginning on the first page with a printed or illustrated impres- sion, and ending on the last page with a printed or illustrated impression. The last recorded frame on each microfiche shall contain the symbol — ^ (meaning "CON- TINUED"), or the symbol V (meaning "END"). whichever applies. L'exemplaire film* fut reproduit grace A la g^n^rosit^ de: J.J. Talman Regional Collection, D.B. Weldon Library, University of Western Ontario Les images suivantes ont ixi reproduites avec le plus grand soin. compte tenu de la condition et de la nertet* de l'exemplaire film*, et en conformit* avec les conditions du contrat de filmage. Lea exemplaires originaux dont la couverture en papier est imprimie sont fiimis er- commencant par le premier plat et en termina- soit par la derniAre page qui compone une empreinte d'impression ou d'illustration, soit par le second plat, selon le cas. Tous les autres exemplaires originaux sont filmis en commencant par la premiere page qui comporte une empreinte d'impression ou d'illustration et en terminant par la derniAre page qui comporte una teile empreinte. Un dee symboles suivants apparartra sur la darniire image de cheque microfiche, selon le cas: le symbole — ♦■ signifie "A SUIVRE ', le symbole V signifie "FIN". Maps, plates, charts, etc., may be filmed at different reduction ratios. Those too largd to be entirely included in one exposure are filmed beginning in the upper left hand corner, left to right and top to bottom, as many frames as required. 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,<j'- siu^'ccdeil in isolatinL; frnin nx pan- creas a nuelcic acid uiiich diflcrcil in a remarkable wax frmn all kn'iwn substances nf this idass. According to Bang the coin- pound is characterized by the following pro])ertics. (i) It contains a pentose group. (2) It coniaiiis a glycerine grnuj) and yields gh'cerine phos- phoric acid. The nucleic acid thus stamling as a connecting link between the pentosans and the lecithins estrdjlishcs a scries of jihysiul'igical I'heniieal rclatiuns wlii*.!) can scarceh' \)v found elsewhere in the science. (3) It contains an aniount of nitrogen and iihosphorus rela- ti\-ely gri'ater ihan is the ease with other nutleic acids. (4) It _\iclds (111 hydrolysis an excessiveaniount of guanin but no trace of cither adenin orthyiiiin. On account of this striking difference from other nucleic acids, the substance has rieei'.'ed the name lif "guanylic J' id." This woj-k which if true \\'ould constitute an unmistakable advance in the scieiice has unfortunately been the subject of ad\'ersc critiiisin of a kind from which one would scarcelv e.\pcct a rci o\-(.r_\'. Thus \\ Furih,^ who has gi\'cu the subject most careful attention, ^\as able to find among the split products neither glycerine iior a rediuing earboliyilrate; he claims that the nucleic acid }'iclds adenin as well as guanin ami that there is no reason for making any dist inci ion lietween guanvlic acid and thymonucleic acid. In consideration of a number of objections ail leading to the same coiulusion one might reasonabh- supjiose ' liang: Zcitschr. /. physiol. Client., .\xvi, p. i^^; xx\i, p. jii. ' V, Ftirtl; and Jerusalem; Hofmeister's Heitrui^e, \, ji. 174. 2.S() 290 Guanvlic Acid of the Spleen that V. Furth's w, .rk wt niM finally dispose of the matter of Ruany- lic aci'l. Rut SteiKlcl' in a very recent communication, takes an entirelv«litt'ercni view. He notes that themeihod .,f y>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<l several thnes a pro.luct was obtained which dissolved in a trace of alkali and on precipitati(.n from the alkaline solution by acetic acid left a perfectly clear fluid. The exceedingly gelatinous nucleo- protei<l thus puritic.l was d.ehydratcd with alcohol and ether, but unless this is done with the greatest care one will tmally obtain a bnAvn sticky mass which is unsuitable for the work that t.jUows. It IS necessary to begin with dilute alcohol (50 per cent) and to replace this gradually until absolute alcohol is linally readied. Th.' latter slvuld be repeatedly used and the material allowed to stan.l for several <lays in well cooked flasks with frequent and violent agitation. We mention the difficulty of dehydrating this nucleoproteid because we encountered no such ditficuliy m dealing with the nucleoproteid of ox pancreas and we regard this very sinking diflerence as sufficient ground for assuming that the tw.; nucleoproteids r.re not identical. This is however entirely aside from the question of the identity of the two nucleic acids. From 14 kilos of moist tissue after the sacrifice of relatively large quantities of material in the interest of a pure product, we finally obtained 64 grams of a perfectly dry pale yellow powder. THE l'REPAR.\TION OF Gf-^NVLIC ACID FROM THE NUCLEO- PROTEID Ol" PIO'S SPLEEN. The nucleoproteid was treated in portions of 12 grams each with 150 cc. of 2 per cent caustic potash and heated for half an hour in a vessel submerged in boiling water. The red fluid was neutralized with acetic acid end whale hot filtered from a small 2q: Guanvlir Acid ot tlit' Spleen (luantitv -f i-erkTilv lihuk inatuKil; l.ni wen after stan.liriK' 12 lin.irs iheiv uas nn ,W\n,^hinu -f •^uin\yVu- acid alth<.u-b the snlutinii was iiiurh in., re o .nrciitralcd tlian that wliidi Han;,' pivparc.l fnim y.imvcufi luulc .vr-uM. i Wc used nnly i5o_y- „f J iKT .cut caustic ]H,!ash v.-i-on- Han- used 400 ce.i The perte^lh- dear vell.Av lluid was then made faintly aeid with aectie aeid and allowed Im stand <.ver ni-ht '."it there uas still nn deposition of Kuanylic aeid. Several days later when we had c.mehidedthat thisnuelenpn.teid yiel Is n-. sul.stanee eorn-pc.nd- ini,' to paiui-eas truanyhe arid and after v. e had praetieally alian- d/med the suhjeet, a vt^ry small deposit was noticed in ilie thud and on tlie ad'dution of a few drops of aeetie aeul there was an iinmed.iate and eojnous preeipitati. .n of while lloeeiilenl material wliile i.raetieidlv all the eol,,nn- mattiM' remained in solution. Tlie preeipilate'was tillered oil, dissohv ' in hot water and tlie solution til'ered from a small amount of ins.'luMe -ramilar maie- ,.,;,p On lin-. the lole yellow ihiid prompt ly deposited , uuany- li^. a,i,l l,ut ..nrei.eatm- the process the yield soon became notiec- al.lv siiudier ;is llie acetic acid was removed with the mother li,p'i.,i-s. Tite addition of acetic a> 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,pience. Hut it mav hapi^cn th.at the neutral fluid cannot he iiltere.l tit ;dl. In the one such ease which we met the fluid was markedly aciditied with .acetic acid, and co,,1ed in ice water when cletir and rapid nitration could he made leavin- the -uaiiyli.- acid on the filter. This dtirk l.rown residue was l.oiled with water and a product obtained whi,'h wtis easilv tiltertible and which deposited K"anv- lie acid on . oolin-. All specimens of what we considered the best products were cllected and dissolved in hot water and the guanvlic acid which was deposited .,n coojin- the fluid was dried in tlie ordinarv wav, witli alcohol and eth.er. From 5J -rtiiiis of \ I Waller Jones and L. Ci. Rnwntree 293 \ I K mu'lc'.prnteia after -cvere lussrs m the inuthcr l:(iuors f^r the reason state.] ue imally ..btaiiie^ i.;S 1,'ram of pure jjuanylie aeid. The suhstan> e e<msists of a perfeetly white dry j>ow.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 treate<l with silver nitrate and ammcmia. The small silver pie- cipitate was th..rou,l-.lv washed, suspended m boding watc^ and Xosed with hvdrochlonc acid. Th.e a. id lUiid was .dtere, from silver chloride, evaporated carefully just to dryness and the last traces ..f hydrochloric acid driven ot. by moistenmg ^Mtl ■ater and n,ain caa-fully evaporating. The insignihcant amount of esidue w;s dissolved in water at 40° and treated withammon.a. A v" stnall precipitate of guanin was forntcd which showed no inclination to dissolve in ammonia even afuT the ,M...... • - ,.reat excess of the reagent. The lluid was nltered off an. l-dcd tint pcrtectlv neutral to litmus. I. will be observe.l that any ad nirori.inallv present must now be foutid in this .lu,d ts volume w;; <.nlv .0 cc. yet a T>ortion 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