MASS SPECTRA OF PICOLINYL ESTERS
Part 1. Saturated and Branched-Chain Fatty Acids
Although I don’t wish
to be dogmatic on the subject, in my opinion, picolinyl esters are the best single derivative available
for identification of fatty acids by gas chromatography-mass spectrometry (GC-MS), at least in mass spectrometry terms. Their GC properties are far from ideal, but many of the modern
polar stationary phases are sufficiently thermally stable to afford good resolution. That said,
dimethyloxazoline (DMOX) and pyrrolidide derivatives are also extremely useful, and can be better
than picolinyl esters in some circumstances. For example, DMOX derivatives have much better GC properties
and are particularly suited to location of conjugated double bond systems. Rather than considering them
as competitors for the title 'ideal method', I prefer to treat them as providing useful complementary data.
Faced with a novel sample, it is always useful to prepared more than one type of derivative for analysis by GC-MS.
This and the next few web pages describe the use of picolinyl esters specifically for structure determination of fatty acids. In addition to analysis by GC, they can be separated by reversed-phase HPLC provided that a base-deactivated stationary phase is employed (Christie, 1998) (see our web page on concentration of minor components). This can be a useful technique to enrich minor components. Methods for preparing the derivatives are described elsewhere in our website in the section Preparation of derivatives.
D.J. Harvey was responsible for much of the early work on picolinyl esters, and his review article is still very valuable (Harvey, 1992).
Straight-Chain Saturated Fatty Acids
The mass spectrum of picolinyl palmitate (hexadecanoate or 16:0)(see Harvey, 1982) is illustrated -
It is typical in that it has prominent ions at m/z = 92, 108, 151 (the McLafferty ion) and 164, which are all fragments about the pyridine ring (if any of these ions is missing from a spectrum it may be indicative of a functional group adjacent to the carboxyl moiety). The molecular ion (m/z = 347) is easily distinguished and it is always odd-numbered, because of the presence of the nitrogen atom, but most other ions are even numbered. Ions below m/z = 92 can usually be ignored.
In interpreting such spectra, the simplest approach is to start with the molecular ion and progress downward, as if one were unzipping the molecule one methylene group at a time. Thus there is loss of a methyl group to m/z = 332, followed by a series of ions 14 amu apart for loss of successive methylene groups. There is little sign of the complex rearrangement ions that can be found with such fatty acid derivatives as methyl esters.
The main fragmentations are often portrayed simplistically as cleavages at the points shown, but in reality those close to the carboxyl group involve some rearrangement with specific hydrogen abstractions (Harvey, 1992; Hamilton and Christie, 2000; Yang et al., 2006). In particular, the ion at m/z = 151 is not formed simply by the well-known McLafferty rearrangement as is usually proposed (see our web pages on Methyl esters of Saturated Fatty Acids for a discussion of the mechanism). However, those ions further down the saturated chain probably represent simple radical-induced cleavage. As these web pages are not intended as detailed mechanistic accounts, fragmentations are generally represented in a simple manner here.
There follow spectra for picolinyl decanoate (10:0) -
- and picolinyl tetracosanoate (24:0), which show essentially the same features -
We have spectra on file for picolinyl esters of straight-chain fatty acids from 2:0 to 30:0, including nearly all the odd-chain ones, and others labelled with stable isotopes. These can be accessed (but with no interpretation) from our Archive page. Only a few of them have been published formally.
Monomethyl-Branched-Chain Saturated Fatty Acids
The majority of naturally occurring monomethyl-branched fatty acids have saturated alkyl chains, and the most common of these are the iso- and anteiso-methyl isomers. They are produced by bacteria mainly, but they can enter animal tissues via the food chain, for example. The mass spectrum of picolinyl iso-methyl-octadecanoate (17-methyl) follows (see Harvey, 1982) -
The spectrum resembles that of a straight-chain saturated fatty acid except for the very obvious gap of 28 amu between m/z = 346 and 374, which represents loss of carbon-17 and its attached methyl group.
In the mass spectrum of picolinyl anteiso-methyl-octadecanoate (16-methyl), this gap is shifted to between m/z = 332 and 360 as -
With that of picolinyl 15-methyloctadecanoate (not published elsewhere), the gap is shifted to m/z = 318 to 346 -
With that of picolinyl 11-methyloctadecanoate (not published elsewhere), the gap is between m/z = 262 and 290 -
With that of picolinyl 10-methyloctadecanoate (tuberculostearate) (not published elsewhere), the gap is between m/z = 248 and 276 -
The mass spectrum of picolinyl 3-methylpentadecanoate is interesting in that the usual ion at m/z = 164 is shifted to 178, and there is a gap of 27 amu between m/z = 151 and 178 to locate the branch point.
Of course, it is easy to predict what the diagnostic features in the spectra of other isomers or homologues will be.
We have spectra of many more methyl esters of straight-chain saturated fatty acids on file, and they can be accessed (but with no interpretation) from our Archive page. Only a few of these have been published elsewhere.
Di- and Polymethyl-Branched-Chain Saturated Fatty Acids
Similar principles to the above apply in the interpretation of mass spectra of picolinyl esters of di- and polymethyl-branched-chain fatty acids. For example, the picolinyl ester of an unusual dimethyl-branched fatty acid from a sponge, i.e. 8,10-dimethyl-hexadecanoate follows (Nechev et al., 2002).
Gaps of 28 amu between m/z = 220 and 248, and between m/z = 262 and 290, locate the branch points.
Polymethyl-branched fatty acids derived from isoprene units via phytol are more common in nature, especially as minor components of fish oils. For example, the spectrum of the picolinyl ester of 4,8,12-trimethyltridecanoate is -
There are clear gaps of 28 amu between ions at m/z = 164 and 192, 234 and 262, and 304 and 332, which locate the methyl groups on carbons 4, 8 and 12, respectively (Christie, 1997).
In the spectrum of the picolinyl ester of the homologous fatty acid 5,9,13-trimethyl-tetradecanoate (not published elsewhere), the diagnostic gaps are all 14 amu higher as anticipated -
The mass spectrum of picolinyl 2,6,10,14-pentadecanoate (pristanate) has some similar features (not published elsewhere) -
- for example, the methyl branches in positions 6, 10 and 14 are located by gaps of 28 amu between m/z = 206 and 234, 276 and 304, and 346 and 374 respectively. However, the presence of the 2-methyl group is recognized as the McLafferty ion is shifted from m/z = 151 to 165, while the ion normally at m/z = 164 is shifted to 178.
In comparison, in the spectrum of the homologous 3,7,11,15-tetramethylhexadecanoate (phytanate), the gaps for the methyl branches on carbons 7, 11 and 15 are shifted up by 14 amu as expected (Christie, 1989).
The methyl group on carbon 3 is located by the fact that the McLafferty ion is again at m/z = 151, and the ion at m/z = 164 has shifted. to 178.
References
- Christie, W.W. Gas Chromatography and Lipids (Oily Press, Dundee) (1989).
- Christie, W.W. Structural analysis of fatty acids. In: Advances in Lipid Methodology - Four, pp. 119-169 (edited by W.W. Christie, Oily Press, Dundee) (1997).
- Christie, W.W. Gas chromatography-mass spectrometry methods for structural analysis of fatty acids. Lipids, 33, 343-353 (1998).
- Hamilton, J.T.G. and Christie, W.W. Mechanisms for ion formation during the electron impact-mass spectrometry of picolinyl ester and 4,4-dimethyloxazoline derivatives of fatty acids. Chem. Phys. Lipids, 105, 93-104 (2000).
- Harvey, D.J. Picolinyl esters as derivatives for the structural determination of long chain branched and unsaturated fatty acids. Biomed. Mass Spectrom., 9, 33-38 (1982).
- Harvey, D.J. Mass spectrometry of picolinyl and other nitrogen-containing derivatives of lipids. In: Advances in Lipid Methodology - One, pp. 19-80 (edited by W.W. Christie, Oily Press, Ayr) (1992).
- Nechev, J., Christie, W.W., Robaina, R., de Diego, F.M., Ivanova, A., Popov, S. and Stefanov, K. Chemical composition of the sponge Chondrosia reniformis from the Canary Islands. Hydrobiologia, 489, 91-98 (2002).
- Yang, S., Minkler, P., Hoppel, C. and Tserng, K.-Y. Picolinyl ester fragmentation mechanism studies with application to the identification of acylcarnitine acyl groups following transesterification. J. Am. Soc. Mass Spectrom., 17, 1620-1628 (2006).
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Updated: 22/4/2008 |
Scottish Crop Research Institute (and MRS Lipid Analysis Unit), Invergowrie, Dundee (DD2 5DA), Scotland
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