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Mass Spectrometry of Methyl Esters


Monoenoic Fatty Acids



As cautioned in the 'Introduction' to these documents, mass spectra of methyl esters obtained with electron-impact ionization often afford limited information only concerning the structures of unsaturated fatty acids. The molecular weight is usually obtainable, and this is an important piece of information. If chromatographic retention data are added to this, it is often possible to be 90% certain of the identity of a fatty acid. Unfortunately, mass spectra of methyl esters of most monoenoic fatty acids contain no information that helps to locate the position or geometry of double bonds. While there have been suggestions that such information can be obtained from close examination of certain minor peaks in the spectrum, I am doubtful of the value of such techniques with real samples, as small changes in instrumental parameters, imperfect peak resolution and background noise could mask such effects. Methods of preparing methyl esters are described on another web page. In addition, there is a document on the chemistry and occurrence of monoenoic fatty acids on this site here... Many of the spectra that follow have not been illustrated elsewhere.


Straight-Chain Monoenoic Fatty Acids

The mass spectra of methyl esters of monoenoic fatty acids under electron-impact ionization tend to be rather uninformative. However, they do allow the molecular weight to be determined and this combined with GC retention data can be of practical value. The introduction of a double bond changes the spectrum appreciably from that of the corresponding saturated ester.

The mass spectrum of methyl oleate (cis-9-octadecenoate) is illustrated first (Hallgren et al., 1959) -

Mass spectrum of methyl oleate

The molecular ion (m/z = 296) is clearly seen, and ions representing loss of the elements of methanol (m/z = 264 or [M-32]+), i.e. a methoxyl group plus a hydrogen atom, and the loss of the McLafferty ion (m/z = 222) are abundant, as is the McLafferty ion per se (m/z = 74) (see the web page on mass spectra of methyl esters of saturated acids). Smaller ions equivalent to [M‑60/61]+ and [M‑49/50]+ at m/z = 235 and 246, respectively in this instance, are present here and in the spectra of most other isomers. A characteristic ion at [M-116]+ (m/z = 180 in this instance), together with homologous ions at 166, 152, etc, are also diagnostic. That at [M‑116]+ is formed by loss of a fragment containing the carboxyl group by cleavage between carbons 5 and 6 with addition of a rearranged hydrogen atom. Thus, in contrast to the spectra of methyl esters of saturated fatty acids, hydrocarbon ions (general formula [CnH2n‑1]+) dominate the spectrum, with m/z = 55 as the base peak usually. The relative abundances of many of these ions tend to be appreciably greater than in the mass spectra of dienes and polyenes.

However, there is no feature that permits location of the double bond, because this can migrate to any position when the alkyl chain is ionized in the mass spectrometer. Thus, other than when the double bond is adjacent to the carboxyl group, all the cis- and trans-18:1 isomers have very similar spectra. The same ions are present, and while the relative intensities can be variable, especially in the high mass region, the finger-prints are not sufficiently distinctive to be of value diagnostically.

The main exception is the spectrum of methyl 2-octadecenoate (below) in which the double bond and carboxyl group presumably form a relatively stable resonance structure. For example, there is a distinctive ion at m/z = 113, which is believed to occur via a cleavage between carbons 5 and 6 followed by formation of a stable 6-membered ring with the carboxyl group (Ryhage et al., 1961)). The base peak is at m/z = 87 in this instance.

Mass spectrum of methyl 2-octadecenoate 

This particular fatty acid does not occur naturally to my knowledge, but it can be formed artefactually by over vigorous trans-esterification of the 3-isomer. However, some natural fatty acids with α,β-unsaturation in addition to other functional groups are known.

Trans-3-octadecenoic acid (trans-3-18:1) and trans-3-16:1 occur naturally in photosynthetic tissues and some seed oils and the spectrum of methyl ester is of the former is -

Mass spectrum of methyl trans-3-octadecenoate

There are minor differences in the relative abundances of some ions relative to the spectrum of methyl oleate, and the author has found the fingerprint useful for diagnostic purposes in analyses of certain seed oils. In addition, the GC retention time relative to those of other 18:1 isomers is a further aid to identification.

As further examples, the mass spectrum of methyl 5-eicosenoate (from meadowfoam oil) is -

Mass spectrum of methyl 5-eicosenoate

This and the next spectrum have similar ions as those for methyl oleate, especially the distinctive ions for [M-32]+, [M-74]+ and [M-116]+ are now at m/z = 292, 250 and 208, respectively, in the first.

The spectrum of methyl 13-docosenoate (also from meadowfoam oil) is -

Mass spectrum of methyl 13-docosenoate

Again the ions representing [M-32]+, [M-74]+ and [M-116]+ at m/z = 320, 278 and 236, respectively, stand out.

The mass spectrum of the shorter-chain methyl 9-dodecenoate (from milk fat) is –

Mass spectrum of methyl 9-dodecenoate

Analogous ions to those in the previous spectra are present, although the relative intensities are somewhat different as might be expected.

Note that the presence of an alicyclic ring reduces the molecular weight of a fatty acid by 2 amu in comparison to the corresponding straight-chain saturated fatty acid, i.e. the same amount as a double bond. While this is not a problem when ring structures are the end of the chain, methyl esters of cyclopropyl fatty acids have mass spectra that are virtually identical to those of monoenes with the same total number of carbon atoms (see the web page on Mass spectra of methyl esters of cyclic fatty acids). Where there is doubt, GC retention data are an important aid to identification.


Branched-Chain Monoenoic Fatty Acids

Methyl branches in saturated fatty acids produce distinctive cleavages in the mass spectra that permit characterization (see the relevant web page). However, few authentic spectra are available from branched-chain monoenes to allow anything other than speculation in interpretation. Unsaturated branched-chain fatty acids are occasionally encountered as minor components of marine lipid samples. The mass spectrum of methyl 15-methyl-hexadec-9-enoate (from a sponge) is -

Mass spectrum of methyl 15-methyl-hexadec-9-enoate

The ions at m/z = 235 ([M-47]+) and 227 ([M-55]+) and presumably those at m/z = 177 and 195 may be related in an unspecified way to the presence of the iso-methyl group, perhaps influenced by the presence of the double bond.

The mass spectrum of methyl 7-methyl-hexadec-6-enoate (from a jellyfish in this instance, but often present as a minor component in fish from warm seas) is -

Mass spectrum of methyl 7-methyl-hexadec-7-enoate

Unlike other monoenes, ions representing [M-32]+, [M-74]+ and [M-116]+ are small. It is tempting to suggest that the ions at m/z = 115 and 167 result simply from fragmentations between carbons 5 and 6 as illustrated. Evidence supporting this interpretation comes from the spectrum of the ethyl ester, where the ion at m/z = 167 remains, but that at m/z = 115 is replaced by one at m/z = 129. After publication here, this spectrum was illustrated in a paper by Fardin-Kia, A.R et al. (2013), together with that of methyl 7-methyl-octadec-6-enoate in which the ion at m/z = 167 is replaced by one at m/z = 195. The distinctive ions at m/z = 138 and 151 are present in the spectra both of the ethyl ester and of the 7-methyl-18:1 analogue also, so are presumed to result from a rearrangement of the carboxyl end of the molecule (with loss of at least a methoxyl/ethoxyl moiety).

We also have the mass spectrum of the isoprenoid methyl 3,7,11,15-tetramethylhexadec-trans-2-enoate (phytenate) on file in the Archive Section.

Dimethyl disulfide adducts: To get round the problem of location of double bonds, it is possible to prepare specific derivatives of unsaturated fatty acids that 'fix' the double bond. Many have been described, but the only one to have stood the test of time is the dimethyl disulfide adduct, as this has excellent mass spectrometric properties and is prepared in a simple one-pot reaction (see the section on 'Mass spectra of methyl esters of fatty acids - further derivatization'). Alternatively, 3-pyridylcarbinol (‘picolinyl’) esters or DMOX or pyrrolidine derivatives can be utilized to locate double bonds, as described in some detail elsewhere in these web pages.

In addition, mass spectrometry with chemical ionization and acetonitrile as the reagent gas can enable both the position and geometry of a double bond to be determined, as reviewed by Brenna (2006). I have no personal experience of this methodology so cannot comment further.

Spectra of many more methyl esters of monoenoic fatty acids can be accessed from our Archive pages (without interpretation).


References


I also recommend - Christie, W.W. and Han, X. Lipid Analysis - Isolation, Separation, Identification and Lipidomic Analysis (4th edition), 446 pages (Oily Press, Woodhead Publishing and now Elsevier) (2010) - at Science Direct.

Credits/disclaimer Updated: March 23rd, 2017 Author: William W. Christie LipidWeb icon

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