Part 4. Trienoic Fatty Acids

Mass Spectra of DMOX Derivatives

Methylene-Interrupted Trienoic Fatty Acids

The mass spectra of DMOX derivatives of trienoic fatty acids permit location of the double bonds, but less easily than those of monoenes and dienes. The principles described in the earlier documents in this series apply (see the web-page on the mass spectra of DMOX derivatives of monoenes, for example). While identification from first principles can sometimes be problematic, different isomers tend to have very different spectra so that characterization is possible when spectra of authentic fatty acids can be compared. Unlike the dienes, few model compounds are available. Therefore, most of the following spectra have been gleaned from analyses of natural products from my own laboratory, and fatty acids with a variety of chain-lengths are described.

For example, the mass spectrum of the DMOX derivative of 6,9,12-octadecatrienoate (γ-linolenate or 18:3(n-6)) is illustrated below (Sayanova et al., 1997) -

Mass spectrum of the DMOX derivative of 6,9,12-octadecatrienoate

The double bonds in positions 9 and 12 are easily recognized from the gaps of 12 amu between m/z = 194 and 206, and between 234 and 246, respectively. That in position 6 must be identified by the fingerprint characteristic for an isomer with the first double bond in position 6, i.e. the odd-numbered ion at m/z = 167 (or the triplet at 167, 180 and 194).

DMOX derivative of 8,11,14-octadecatrienoate (18:3(n-4)) - a minor component of fish oils -

Mass spectrum of the DMOX derivative of 8,11,14-octadecatrienoate

DMOX derivative of 9,12,15-octadecatrienoate (α-linolenate or 18:3(n-3)) (Zhang et al., 1988) -

Mass spectrum of the DMOX derivative of 9,12,15-octadecatrienoate

In this and the previous example, all three double bonds are easily recognized by the gaps of 12 amu as indicated in the spectrum. Thus in the last, gaps of 12 amu between m/z = 196 and 208, 236 and 248, and 276 and 288, locate the double bonds in position 9, 12 and 15, respectively. Note that gaps of 40 amu between m/z = 196, 236 and 276, and between 208, 248 and 288 are also useful diagnostically. Comparable ions, 14 amu less, are present in the spectrum of the 8,11,14-isomer.

Trienes of the n-1 family of fatty acids have yet to be found in nature, but this spectrum (from a genetically modified plant) is illustrated next for the sake of completeness (Sayanova. et al., 2006). The first two double bonds are easily located but terminal double bonds give problems with all derivative types. Here again the gaps of 40/41 amu are useful indicators (m/z = 210 to 250 to 290 to 331). DMOX derivative of 11,14,17-octadecatrienoate (18:3(n-1)) -

Mass spectrum of the DMOX derivative of 11,14,17-octadecatrienoate

All three double bonds are easily recognized by the gaps of 12 amu, as indicated, in the spectra of the two 20:3 isomers that follow. Thus, the DMOX derivative of 8,11,14-eicosatrienoate (20:3(n-6)) -

Mass spectrum of the DMOX derivative of 8,11,14-eicosatrienoate

DMOX of 11,14,17-eicosatrienoate (11,14,17-20:3 or 20:3(n-3)) -

Mass spectrum of the DMOX derivative of 11,14,17-eicosatrienoate

 

Bis- and Polymethylene-Interrupted Trienoic Fatty Acids

As described elsewhere for dienes, it has become apparent that bis- and polymethylene-interrupted trienoic fatty acids are more common in nature than may have been supposed. In particular, fatty acids with a 5,9-double bond system or their chain elongation products are present in seed oils from Gymnosperms or in certain marine invertebrates such as sponges.

The spectrum of the DMOX derivative of 5,9,12-octadecatrienoate from a pine species is typical (Fay. and Richli, 1991).

Mass spectrum of the DMOX derivative of 5,9,12-octadecatrienoate

As described for dienes (Mass spectra of DMOX derivatives. Part 3. Dienoic fatty acids), the prominent ion at m/z = 180 represents cleavage at the centre of the bis-methylene-interrupted (5,9) double bond system. Analogous ions are present in the corresponding spectra of pyrrolidides and 3-pyridylcarbinol ('picolinyl') esters. The double bond in position 12 is located by a gap of 12 amu between m/z = 234 and 246. Note that the odd-numbered ion at m/z = 153 is diagnostic for a double bond in position 5, while the fact that the ion at m/z = 113 is so much larger than that at m/z = 126 is also characteristic (see the corresponding web page on monoenes).

The spectrum of the DMOX derivative of 5,9,13-eicosatrienoate (5,9,13-20:3) from a sponge may be unique in having two bis-methylene-interrupted double bond systems -

Mass spectrum of the DMOX derivative of 5,9,13-eicosatrienoate

The ion at m/z = 180 represents cleavage at the centre of the 5,9-double bond system, while that at m/z = 234 is diagnostic for the 9,13-system. However, ions for the individual double bonds are not easily recognized.

DMOX derivative of 5,9,19-hexacosatrienoate (5,9,19-26:3) -

Mass spectrum of the DMOX derivative of 5,9,19-hexacosatrienoate

This very-long chain triene is typical of the type of fatty acid component found in sponges, in this instance from Hymeniacidon cinerea. It has the ion at m/z = 180 for the 5,9-double bonds, while the gap of 12 amu later as indicated locates that in the 19-position.

The spectrum of the DMOX derivative of 7,11,14-eicosatrienoate (7,11,14-20:3) from seed oils of pine species.

Mass spectrum of the DMOX derivative of 7,11,14-eicosatrienoate

In this instance, the spectrum resembles that of 5,9,12-18:3 except that the diagnostic ions are shifted upwards by 28 amu (Wolff et al., 1997)). In particular, the ion representing cleavage at the centre of the bis-methylene-interrupted double bond system is now at m/z = 208.

A few trienoic fatty acids are known in which double bonds are separated by more than two methylene groups, for example from certain conifer species. The spectrum of the DMOX derivative 5,11,14-eicosatrienoate (5,11,14-20:3) from Pinus contorta seed oil follows (see also Zhang et al., 1988) -

Mass spectrum of the DMOX derivative of 5,11,14-eicosatrienoate

All the double bonds must be identified individually - that in position 5 by the diagnostic ion at m/z = 153, and the others by the gaps of 12 amu.

A few fatty acids with isolated double bonds in position 3 are found in plants, although these are often of the trans rather than the cis configuration as in the example here, the DMOX derivative of 3,9,12-octadecatrienoate from Tanacetum zawadskii seed oil (Tsevegsuren et al., 2003).

Mass spectrum of the DMOX derivative of 3,9,12-octadecatrienoate

The base peak at m/z = 152 is the important diagnostic feature, though this would also be the case if the double bond were in position 2 (see our web page on DMOX derivatives of monoenes). Unfortunately, it is possible that the double bond in position 3 has migrated to position 2 because of the harsh alkaline conditions during derivatization, and that this is in fact the spectrum of the 2,9,12-18:3 isomer.

 

Trienoic Fatty Acids with Conjugated Double Bonds

Trienoic fatty acids with only two of the double bonds in conjugation are not often encountered in nature, but 9-cis,11-trans,15-cis-octadecatrienoic acid is formed by biohydrogenation of α-linolenic acid in the rumen of cows and sheep and occurs as a minor component of milk fat, adipose tissue and bile of these species. The mass spectrum of its DMOX derivative is -

DMOX derivative of 9,11,15-18:3

The gap of 12 amu between m/z = 196 and 208 confirms the double bond in position 9, while that for the similar gap between 222 and 234 confirms the double bond in position 11. The outstanding feature of the spectrum is the base ion at m/z = 262, which represents cleavage at the centre of the bis-methylene-interrupted double bond system. The gap to m/z = 288 also confirms the double bond in position 15. The mass spectrum does not of course give information on the configurations of the double bonds (Destaillats, F. et al., 2005).

Fully conjugated trienoic acids are well known constituents of certain seed oils and the mass spectrum of the DMOX derivative of punicic or 9-cis,11-trans,13-cis-octadecatrienoic acid follows.

DMOX of 9,11,13-18:3

In this instance, all the positions of all the double bonds can be recognized from the gaps of 12 amu as illustrated. The mass spectrum of the DMOX derivative of the isomeric triene α-eleostearic acid is identical to this and has been published by Spitzer (1997). There seems little doubt that DMOX derivatives are better than 3-pyridylcarbinol esters and pyrrolidides for locating double bonds in conjugated fatty acids. On the other hand, there is a danger of isomerization occurring if the derivatization conditions are too vigorous.

We have mass spectra of the DMOX derivatives of further trienoic fatty acids on file and these are illustrated in the Archive Section of these web pages, but without detailed interpretation. Most of these have not been formally published elsewhere. References are listed when we are aware of prior formal publication of spectra in the scientific literature.

 

References

  • Destaillats, F., Trottier, J.P., Galvez, J.M.G. and Angers, P. Analysis of α-linolenic acid biohydrogenation intermediates in milk fat with emphasis on conjugated linolenic acids. J. Dairy Sci., 88, 3231-3239 (2005) (DOI: 10.3168/jds.S0022-0302(05)73006-X).
  • Fay,L. and Richli,U. Location of double bonds in polyunsaturated fatty acids by gas chromatography-mass spectrometry after 4,4-dimethyloxazoline derivatization J. Chromatogr. A, 541, 89-98 (1991) (DOI: 10.1016/S0021-9673(01)95986-2).
  • Sayanova, O., Haslam, R., Guschina, I., Lloyd, D., Christie, W.W., Harwood, J.L. and Napier, J.A. A bifunctional Δ12, Δ15-desaturase from Acanthamoeba castellanii directs the synthesis of highly unusual n-1 series unsaturated fatty acids. J. Biol. Chem., 281, 36533-36541 (2006) (DOI: 10.1074/jbc.M605158200).
  • Sayanova, O., Smith, M.A., Lapinskas, P., Stobart, A.K., Dobson, G., Christie, W.W. and Shewry, P.R. Expression of a borage desaturase cDNA containing an N-terminal cytochrome b5 domain results in the accumulation of high levels of Δ6-desaturated fatty acids in transgenic tobacco. Proc. Natl. Acad. Sci. USA, 94, 4211-4216 (1997).
  • Spitzer, V. Structure analysis of fatty acids by gas chromatography - low resolution electron impact mass spectrometry of their 4,4-dimethyloxazoline derivatives - a review. Prog. Lipid Res., 35, 387-408 (1997) (DOI: 10.1016/S0163-7827(96)00011-2).
  • Tsevegsuren, N., Fujimoto, K., Christie, W.W. and Endo, Y. Occurrence of a novel cis, cis, cis-octadeca-3,9,12-trienoic (Z,Z,Z-octadeca-3,9,12-trienoic) acid in Chrysanthemum (Tanacetum) zawadskii Herb. (Compositae) seed oil. Lipids, 38, 573-578 (2003) (DOI: 10.1007/s11745-003-1498-6).
  • Wolff, R.L., Christie, W.W. and Coakley, D. Bishomopinolenic (7,11,14-20:3) acid in Pinaceae seed oils. J. Am. Oil Chem. Soc., 74, 1583-1586 (1997) (DOI: 10.1007/s11746-997-0081-5).
  • Zhang, J.Y., Yu, Q.T., Liu, B.N. and Huang, Z.H. Chemical modification in mass spectrometry IV. 2-Alkenyl-4,4-dimethyloxazolines as derivatives for double bond location of long-chain olefinic acids. Biomed. Environ. Mass Spectrom., 15, 33-44 (1988) (DOI: 10.1002/bms.1200150106).

 Updated November 6, 2013

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