Chemtstry and Phystcs of Ltptds 14 (1975) 336-342 © North-Holland PubhshmgCompany

13C N U C L E A R M A G N E T I C R E S O N A N C E S P E C T R A O F 1,2-DIOCTADEC-CIS-ENOYL-SN-GLYCERO-3-PHOSPHORYLCHOLINES Peter G BARTON Department of Btochemtstry, Umverstty of Alberta, Edmonton, Alberta T6G 2E1, Canada Recewed January 13, 1975, acceptedMarch 7, 1975 The 13C NMR spectra of all sixteen 1,2-dmctadec-cts.enoyl-sn-glycero-3-phosphorylcholmes have been obtained Resonance hnes of the olefimc, methylene, methyl and caxboxylcarbon nuclei are sufficiently characteristic to permit uneqmvocaldeslgnaUon of double bond position for each isomer Two resonances of the sn-glycero-3-phosphorylchohnestructure have been reassigned

I Introduction We have recently completed the synthesis of all sixteen positional isomers of 1,2-dloctadec-cls-enoyl-sn-glycero-3-phosphorylchohne [1] These compounds are not resolved from one another by conventional chromatographac methods, such as than layer chromatography on sdlca gel, and it is therefore difficult to dlstlngmsh and identify the various Isomers Identification of double bond poslUon can be achieved by oxadatwe degradation of the derived fatty acids to shorter chain monoand dlcarboxyhc acids, conversion of the fragments to methyl esters and Identification of the esters by gas hquld chromatography (g 1 c ) [2] However, this procedure suffers several disadvantages It mvolves destruction of the analytical material, it is a fairly tedious procedure and it requires the avadabdlty of a wide range of mono- and dlcarboxyhc esters as standards for g 1 c In recent years, the development and avallabihty of 13C nuclear magnetic resonance (13C NMR), have permitted an Increasing utlhzatlon of thas technique for the analysis and identification of hplds The large chemical shafts of the carbon nuclei and the absence of C-C couphng m natural abundance spectra, together with the use of pulse Fourier transform and selectwe proton decouplmg methods, have improved the resolution of resonance lines to such an extent that most of the individual carbon atoms m lecithin mole~,ules can be separate/), observed [3 -6] This prompted us to investigate the posslbdlty that 13C NMR might be used to dlstmgmsh and identify the various isomers of 1,2-dloctadec-cts-enoyl-sn-glycero-3-phosphorylchohne Such a method would have the advantage of bemg non-destructwe

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Dunng the course o f our mvestxgatlon we became aware o f an extenswe 13C NM'R study o f the methyl esters o f long chain monounsaturated carboxyhc acids being carried out elsewhere [7, 8] These data have proved a valuable reference source m our own study

II Experimental The spectra were obtained with either a Bruker HFX-IO Pulse Fourier Transform NMR Spectrometer operating at 22 6 MHz and a pulse width o f 8/asec or with a modified Vanan HA-100 Spectrometer operating at 25 07 MHz and a pulse width o f 25/asec Normally, the sweep width was 5000 Hz and up to 8 K scans were collected As reqmred, a sweep width of 400 Hz or 800 Hz was used to obtain chemical shafts to an accuracy of-+O O1 p p m All the spectra were proton decoupled In most cases CDCI 3 was used as solvent and used as an internal d e u t e r m m lock with TMS as the internal reference Chemical shafts are then gwen as/i-values downfield from TMS The concentration o f hpld used was m the range from 200 to 300 mg m1-1 Spectra of all compounds were measured at 35°C and some spectra were measured additionally at 52°C The spectra o f 1,2-dloctadec-3'-cls-enoyl- and 1,2-dloctadec-15'cts-enoyl-sn-glycero.3.phosphorylchohne were also measured on somcated suspensions m D 2 0 with dloxan as internal reference Spectra o f chohne iodide, phosphorylchohne CaC12, (sn-glycero-3-phosphorylchohne)2 (CaC12) 3 and methylphosphorylchohne were taken as solutions m D 2 0 at concentrations o f 300 to 700 mg m1-1 with dxoxan as internal reference Chemical shifts m these cases were expressed as/i-values downfield from dloxan +67 00 p p m m order to permit direct comparison wRh TMS-referenced spectra

III Results and Discussion The spectrum o f 1,2.dloctadec-4'-cts-enoyl-sn-glycero-3-phosphorylchohne is shown m fig 1 A Resonances o f o l e f i n w carbons

Two olefimc resonances are seen m the spectrum o f each o f the sixteen isomers except the A13c roomer and the difference m shaeldmgs of the two nuclei m each case is characteristic o f the double b o n d poslUon * Thas is dlustrated m fig 2 With the cts double b o n d at the 17 position m the chain the/i-values for C17 and C18 are 139 15 p p m and 114 24 p p m respectively The separation then decreases to * Abbrevmtlons used are - e g A9c, cts unsaturatmn at the 9 10 posltlon, AC=C, separation of olefmlc absorptions m Hz

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