The Lipid Library

MASS SPECTRA OF OXYGENATED FATTY ACIDS

Part 1B. 10- to ω-Hydroxy Acids

As in the previous section, mass spectra are arranged and discussed according to the positions of hydroxyl groups in the fatty acyl chains. Spectra of methyl esters, picolinyl esters, DMOX derivatives and pyrrolidides are all described in this document (when available). As with my other documents on mass spectrometry, this is a subjective account that details only those oxygenated fatty acids encountered during our research activities here and for which we have spectra available for illustration purposes. However, I trust that we have a sufficiently wide range of spectra to give the flavour of the topic. My colleagues and I have never worked with eicosanoids, so no spectra of such compounds can be discussed here. Many of the spectra have not been published elsewhere.

A common feature of mass spectra of hydroxy acids is fragmentation leading to the loss of the elements of water, especially corresponding to [M-17]⁺ or [M-18]⁺. Molecular ions may not be seen without amplification of the appropriate part of the spectrum. Preparation of trimethylsilyl ethers and related derivatives is often helpful.

10-Hydroxy Fatty Acids

10-Hydroxy-octadecanoic acid is produced by several microorganisms, and is also a minor component of cow's milk fat, which was the origin of the following mass spectra. Methyl 10-hydroxy-octadecanoate has the spectrum -

In common with the other hydroxy acids discussed in the previous web page on this topic, the molecular ion is virtually absent. The main fragmentations occur alpha to the carbon carrying the hydroxyl group, with a further rearrangement leading to the loss of the elements of methanol, as illustrated.

The trimethylsilyl (TMS) ether derivative of methyl 10-hydroxy-octadecanoate also gives a distinctive and relatively simple fragmentation, with the two main ions formed by cleavage alpha to the carbon carrying the TMS group standing out clearly.

12-Hydroxy Fatty Acids

The mass spectrum of methyl 12-hydroxystearate (produced by hydrogenation of ricinoleate) is -

Cleavage occurs beta to the oxygen atoms to give the ions at m/z = 229 and 200/199. The base peak at m/z = 197 is formed by loss of methanol (32 amu) from the fragment at m/z = 229.

The mass spectrum of picolinyl 12-hydroxystearate is -

The molecular ion is not seen, and the first significant ion at m/z = 373 represents the loss of the elements of water. The base peak at m/z = 306 is for cleavage alpha to the carbon carrying the hydroxyl group, i.e. between carbons 12 and 13.

12-Hydroxy-octadec-9-enoic (ricinoleic acid) is the main component of castor oil (Ricinus communis), which is a major agricultural and industrial product. The mass spectrum of methyl ricinoleate, which follows, is not especially exciting. The molecular ion is not seen, and the only significant ion in the high mass region is for loss of water, at m/z = 294. However, the ion at m/z = 198 probably represents cleavage between carbons 11 and 12, and the ion at m/z = 166 a further loss of methanol from the carboxyl group.

The trimethylsilyl ether derivative has a more distinctive spectrum (Kleiman, R. and Spencer, G.F. J. Am. Oil Chem. Soc., 50, 31-38 (1973)) -

The ions at m/z = 187 and 299 represent cleavage on either side of the hydroxyl group. That at m/z = 270 is formed by a complex rearrangement involving transfer of the -TMS group to the carbonyl group. Similarly, I suspect that a small ion at m/z = 146 in the spectra of trimethylsilyl ethers of methyl esters of other hydroxy acids is formed by a transfer of the trimethylsilyl group to the McLafferty ion.

The mass spectrum of picolinyl ricinoleate is more informative and the various fragmentations can be interpreted in a straightforward manner.

Ions at m/z = 275 (presumably carrying an extra proton) and 304 are produced by cleavage on either side of the carbon carrying the hydroxyl group. The double bond in position 9 is most easily recognized by the gap of 40 amu between m/z = 220 and 260. It must have an influence on the fragmentation at the hydroxyl-carbon to enhance the size of the ion at m/z = 275.

The spectrum of the DMOX derivative of ricinoleate -

In this instance, the double bond is recognized as in other monoenes by the gap of 12 amu between m/z = 196 and 208. The fragments at m/z = 237 (presumably protonated and unusual in being odd-numbered) and at 266 are formed by cleavage alpha to the carbon carrying the oxygen atom. The mass spectrum of the pyrrolide derivative is very similar to this.

The trimethylsilyl ether of the DMOX derivative of ricinoleate has a simple but much less interesting spectrum -

There are fragments that clearly locate the hydroxyl group, as illustrated, but none that fix the double bond.

In contrast, the pyrrolidide derivative has a more complex though no more informative spectrum (see the Archive page for an illustration). It contains the same ions that locate the hydroxyl group as for the DMOX derivative, but there is an abundant ion at m/z = 185, which I suspect is formed by transfer of the trimethylsilyl group to the McLafferty ion. An ion at m/z = 198 may have a related origin, while one at m/z = 309 ([M−114]⁺) may represent the loss of the McLafferty ion from the parent molecule. There are analogous ions in the spectra of the corresponding C₂₀ fatty acid (again, see the archive page). Of course, detailed mechanistic studies would be necessary to confirm these suggestions.

13- and 14-Hydroxy Fatty Acids

The following spectra were obtained from minor components (differing in chain-length) of cow's milk fat.

Methyl 13-hydroxy-octadecanoate - TMS ether derivative -

Methyl 14-hydroxy-hexadecanoate - TMS ether derivative -

Fragments adjacent to the TMS ether are as expected. Note the increase in the abundance of the ion from the distal portion of the molecule relative to that from the carboxyl end.

We also have spectra of various derivatives of 14-hydroxy-eicos-11-enoic (lesquerolic acid), the C₂₀ analogue of ricinoleic acid, available in our Archive pages, i.e. for methyl esters -- DMOX derivatives -- pyrrolidides.

15-Hydroxy Fatty Acids

We encountered 15-hydroxy-octadecanoic acid as a minor component of cow's milk fat, and the spectrum of methyl 15-hydroxy-octadecanoate - TMS ether is -

Fragmentations adjacent to the TMS ether moiety are as expected.

15-Hydroxy-octadeca-9,12-dienoic acid (15-hydroxy-linoleate or 'avenoleate') is found in the monogalactosyldiacylglycerol fraction of oat seed lipids. In nature, the hydroxyl group is esterified with a further molecule of linoleate, i.e. it is an estolide. The methyl ester of 15-hydroxy-linoleate has the spectrum illustrated next -

Again the molecular ion is not apparent, but the ion at m/z = 238 for cleavage between carbons 14 and 15 (containing the carboxyl group) is diagnostic, together with an ion for further loss of the elements of methanol (m/z = 206). Interestingly, the ion at m/z = 150, typical of an n-6 fatty acid, is still present.

The trimethylsilyl ether derivative methyl 15-hydroxy-linoleate has a rather distinctive spectrum -

The molecular ion (m/z = 382) can just be detected, but the outstanding feature is the base ion at m/z = 145 which represents cleavage between carbons 14 and 15, i.e. the terminal part of the molecule. The ion from the carboxyl end of the molecule (m/z = 339) is just detectable.

Picolinyl 15-hydroxy-linoleate -

In this instance, there is a distinct molecular ion (m/z = 387). Ions at m/z = 315 and 344 are produced by cleavage on either side of the carbon linked to the hydroxyl group, while the gaps of 26 amu between m/z = 234 and 260, and 274 and 300, serve to locate the double bonds in positions 9 and 12, respectively (see the web page on Picolinyl esters of dienoic fatty acids).

The spectrum of the DMOX derivative of 15-hydroxy-linoleate -

Again the molecular ion is obvious (m/z = 349), and there are ions diagnostic for the position of both the hydroxyl group and the double bonds. Thus, ions at m/z = 276 and 306 reflect cleavage on either side of the carbon linked to the hydroxyl group, while the gaps of 12 amu between m/z = 196 and 208, and 236 and 248, serve to locate the double bonds in positions 9 and 12, respectively (see the section on DMOX derivatives of dienoic fatty acids).

The spectrum of the trimethylsilyl ether derivative of the latter has a distinctive feature similar to that of the methyl ester in that the base peak is the ion at m/z = 145, which represents cleavage between carbons 14 and 15, and is the terminal part of the molecule (very unusual for a DMOX derivative). The ions expected for the double bonds are not easily seen, however.

(ω-1)- and ω-Hydroxy Fatty Acids

In animal tissues, a family of enzymes termed cytochromes P450s are involved in fatty acid oxidation, hydroxylating with high specificity at the energetically unfavorable terminal (omega) or omega-1 carbons. Similar fatty acids are found in waxes and plant cutins.

Methyl 15-hydroxy-hexadecanoate (from beeswax)-

The ion at m/z = 242 can be considered simplistically as representing cleavage as illustrated and serves to locate the hydroxyl group (Ryhage,R. and Stenhagen, E. Arkiv Kemi, 15, 545-574 (1960)). . On the other hand, the mass spectrum of the TMS ether (not shown) is dominated by an ion at m/z = 117 for cleavage at the same point but containing the terminal portion of the molecule.

The pyrrolidide of 15-hydroxy-hexadecanoate -

Interpretation of the spectrum is made more difficult by the presence of ions simply reflecting a loss of water. However, ions at m/z = 281 and 310 serve to locate the hydroxyl group. An ion at m/z = 280/281, and at m/z = 117 (terminal portion of the molecule) are diagnostic in the spectrum of the TMS ether.

Methyl 16-hydroxy-hexadecanoate -

The main diagnostic ion is that for the loss of the terminal carbon and its hydroxyl group. With methyl esters of saturated ω-hydroxy acids, the most significant ion in the high mass range represents a loss of 30 amu (not 31 or 32), m/z = 256 in this instance, presumably reflecting cleavage of the terminal -CH₂OH group.

The trimethylsilyl ether of this fatty acid on the other hand has a rather simple mass spectrum. The molecular ion is not seen, and the first significant ion at m/z = 343, represent the loss of a methyl group from the TMS moiety. The base peak at m/z = 311 is for a further loss of the methoxyl group from the carboxyl end of the molecule.

The pyrrolidide of 16-hydroxy-hexadecanoate -

Again, the main diagnostic ions is that for the loss of the terminal carbon and its hydroxyl group. The DMOX derivative is of little value with this isomer, as ions from loss of methyl groups from the oxazoline ring confound the spectrum.

Spectra of further hydroxy fatty acids ( and their trimethylsilyl ether derivatives) are available, but without interpretation, in the Archive Sections of these web pages, i.e. for methyl esters -- picolinyl esters -- DMOX derivatives -- pyrrolidides.

Updated: 19/12/2007 Credits/disclaimer	Scottish Crop Research Institute (and MRS Lipid Analysis Unit), Invergowrie, Dundee (DD2 5DA), Scotland