PART 1. STRUCTURE AND COMPOSITION
Nearly all the commercially important fats and oils of animal and plant origin consist almost exclusively of the simple lipid class - triacylglycerols. This includes all the vegetable oils, such as those from corn (maize), olive, palm, and sunflower, and animal fats, such as tallow, lard and butter, as well as commercial products such as margarines. The more abundant animal triacylglycerols are depots fats (from adipose tissue) or milk fats, where their main function may be as a store of energy, but some triacylglycerols (e.g. those of plasma or liver) may have a more dynamic role. Similarly, seed oils serve as a source of energy and structural fatty acids for the developing embryo.
In chemical terms, triacylglycerols consist of the trihydric alcohol glycerol esterified, almost invariably, with long-chain fatty acids. When the two primary hydroxyl groups are esterified with different fatty acids, the resulting triacylglycerol can be asymmetric and thus can display "optical activity", although this is usually too low to be measured. The conventional D/L or R/S systems could designate such enantiomers without ambiguity with simple molecules, but problems arise in application to the complex mixtures of triacylglycerols found in nature. Such problems can be avoided if the stereochemistry of triacylglycerols and other glycerolipids is described by the "stereospecific numbering" (sn) system as recommended by a IUPAC-IUB commission (see also our web page on Nomenclature).
In a Fischer projection of a natural L-glycerol derivative as shown above, the secondary hydroxyl group is shown to the left of C-2; the carbon atom above this then becomes C-1 while that below becomes C-3, and the prefix sn is placed before the stem name of the compound. The term "triacyl-sn-glycerol" should then be used to designate the molecule rather than "triglyceride". The former is technically more accurate and is essential for the stereospecific numbering system, while the latter term may only be familiar now to older readers. When the detailed stereochemistry is not specified, the primary hydroxy groups are often termed the α- and α’-positions and the secondary, the β-position. As an example, the single molecular species 1,2-dihexadecanoyl-3-(9Z-octadecenoyl)-sn-glycerol is illustrated.
Differences in the distributions of fatty acids among the positions of the glycerol moiety in triacylglycerols from natural fats and oils were first demonstrated systematically by enzymatic hydrolysis procedures, specifically pancreatic lipase hydrolysis for the analysis of the fatty acids of position sn-2 (regiospecific analysis), before complex stereospecific hydrolysis procedures were developed that permitted the complete positional distributions of the fatty acids to be determined. Because of this historical development of the analytical procedures, there has been a tendency to assume that the composition of fatty acids esterified to the sole secondary hydroxyl group must have greater importance than those of the two primary positions. It is certainly true that the composition of position sn-2 is of great importance when triacylglycerols are consumed and digested by animals, since 2-monoacyl-sn-glycerols are then formed which can be absorbed by the intestines and utilised as such. On the other hand, the results of stereospecific analyses have shown that the compositions of all three positions in certain fats can be distinctive and can highlight important aspects of the biosynthetic processes.
The discussion that follows centres on the fatty acid compositions and the stereospecific distribution of fatty acids within triacyl-sn-glycerols with a few selected examples. Note that all data are presented as mol% not weight %. The second document here dealing with triacylglycerols (Triacylglycerols. Part 2) discusses the biochemistry of triacylglycerols.
2. Triacylglycerols from Seed Oils
While seed and fruit oils within the plant kingdom as a whole can contain a vast range of different fatty acids, those that are used most widely for food purposes tend to have relatively simple compositions in which C18 unsaturated fatty acids, especially oleate and linoleate are predominant. Thus, olive oil contains over 70% of oleic acid, while safflower oil can contain a similar proportion of linoleate, which is also the major component of maize (corn) and soybean oils. Linseed oil is used mainly for industrial purposes as it contains about 60% α-linolenic acid, which is not suitable for food purposes in such concentrations as it oxidizes too readily. Soybean oil, with 3 to 7% linolenic acid, is often hydrogenated before it is used as a food ingredient. Rapeseed oil is available in two forms, i.e. with high or low erucic acid contents. The former has many industrial uses, but is banned from foods because of fears about the safety of erucic acid for human consumption. Castor oil, with mainly industrial applications, contains up to 90% ricinoleic acid (see our web pages on hydroxy fatty acids). The so-called ‘tropical oils’, such as palm oil, contain higher amounts of saturated fatty acids than most other commercial oils. Palm kernel and coconut oils are rich sources of medium-chain fatty acids. Some representative analyses are listed in Table 1.
Table 1. Positional distributions of fatty acids (mol %) in triacyl-sn-glycerols of seed oils.
|tr = trace ( <0.5% ). * High erucic acid rapeseed oil.
TG = intact triacylglycerols
Data from - Brockerhoff, H. and Yurkowski, M., J. Lipid Res., 7, 62-64 (1966). Christie, W.W. et al., Lipids, 68, 695-701 (1991).
As was well known from studies involving hydrolysis with pancreatic lipase, position sn-2 of the triacylglycerols of seed oils is greatly enriched in polyunsaturated fatty acids (specifically linoleic and linolenic acids), while saturated fatty acids are concentrated in the primary positions, and monoenoic acids are relatively evenly distributed. There are exceptions to these rules and in cacao butter, for example, oleic acid is present largely in position sn-2. Minor differences only in the distributions of saturated and monoenoic fatty acids between positions sn-1 and sn-3 have been observed and too few samples have been analysed for definitive comment. Longer-chain fatty acids (C20-C24) are apparently concentrated in the primary positions with some small preference for position sn-3.
In those seed oils containing unusual fatty acids that have been subjected to stereospecific analysis, an allenic estolide was found entirely in position sn-3 in the triacyl-sn-glycerols of Sapium sebiferum, acetic acid appeared to be linked entirely to position sn-3 in Euonymus verrucosus, and much of the coriolic acid was in position sn-3 in Monnina emarginata.
In summary, seed oils containing the usual range of saturated and unsaturated fatty acids tend to have the polyunsaturated components in position sn-2, but relatively little difference between the primary positions; less-common fatty acids tend to be concentrated in position sn-3.
3. Triacylglycerols from Animal Fats - Adipose Tissue
Much of the triacylglycerols in animal tissues, including some of the commercially important fats such as lard or tallow, are contained within various adipose tissue sites, where they serve mainly as an energy store. The subcutaneous fats also help to insulate animals, while fat stores in fish and marine animals help to maintain buoyancy. Fatty acids are synthesised de novo in tissues (saturated and monoenoic acids), but the composition of the triacylglycerols also reflects the diet. In terrestrial animals, the composition tends to be quite simple, with C16 (mainly 16:0) and C18 fatty acids predominating. All the essential fatty acids, such as linoleic and α-linolenic acids, must come from the diet of course. Ruminant animals, such as the cow and sheep, have relatively saturated fats because the dietary unsaturated fatty acids are subjected to biohydrogenation in the rumen, a process that also generates trans fatty acids as by-products. These animals also tend to have relatively higher concentrations of odd- and branched-chain fatty acids from the rumen microflora. Non-ruminant herbivores, such as the horse, can have appreciable amounts of linolenic acid from grass in their adipose tissue, while that of marine mammals is characterized by high concentrations of long-chain mono- and polyenoic fatty acids because of their diet of fish.
Marked differences have been observed in the distributions of fatty acids among the three positions of the glycerol moiety in most species examined. There are also appreciable inter-species differences. A few representative results are listed in Table 2.
Table 2. Positional distributions of fatty acids (mol %) in triacyl-sn-glycerols of animal depot fats.
|a Results are listed for
cis-18:1 isomers only; trans-18:1 was present in positions
sn-1, sn-2 and sn-3 as 5,
2 and 6 %, respectively.
tr = trace (<0.5%). TG = intact triacylglycerols
Data from – Christie, W.W. et al., Lipids, 6, 854-856 (1971); Brockerhoff, H. et al., Biochim. Biophys. Acta, 116, 67-72 (1966); Christie, W.W. and Moore, J.H. J. Sci. Food. Agric., 22, 120-124(1971), Biochim. Biophys. Acta, 210, 46-56 (1970), J. Sci. Food. Agric., 23, 73-77 (1972).
For most species, saturated fatty acids are found predominantly in position sn-1, although appreciable amounts of oleic acid are usually present also. Position sn-2 tends to contain mainly unsaturated fatty acids, especially linoleic acid, although some of the shorter-chain fatty acids also accumulate here in some instances. There is some preference for the longer-chain fatty acids to be located in position sn-3. The main exception to these rules is in the pig and related species where it has been known for many years that palmitic acid can comprise more than 70% of the fatty acids in position sn-2. However, it is less well known though perhaps of equal interest that most of the stearic acid (75% of that in the tissue) is in position sn-1, while position sn-3 contains more than 70% of oleic acid. Although the absolute fatty acid compositions of adipose tissue at various sites in an animal can vary somewhat (subcutaneous fats tend to contain more unsaturated fatty acids than internal depot fats), the proportionate distributions of each fatty acid among the three positions does not vary significantly in any of these tissues in either in the pig or the sheep.
A variety of dietary factors can influence the fatty acid compositions and thence the structures of depot fat triacylglycerols, not least the composition of the diet. Few systematic studies of the relationship between triacylglycerol composition and structure have been performed, and in the example below the pig was the experimental animal. An example of the kind of result obtained is shown in Figure 1.
Figure 1. Variation of the composition of palmitic acid in positions sn-1, sn-2 and sn-3 of the triacylglycerols of pig adipose tissue with changes in overall composition (Christie, W.W. and Moore, J.H. Lipids, 5, 921-928 (1970)).
As the proportion of palmitic acid in the triacylglycerols varied between 18 and 33%, the amount in position sn-1 remained constant at about 10%, that in position sn-3 increased linearly but relatively slowly from 2 to 4%, while that in position sn-2 was affected most and increased rapidly from less than 50% to more than 70%. Over the range of compositions studied, the amounts of all of the fatty acids varied in a characteristic manner that was linear in each of the three positions, though only data for 16:0 are illustrated.
4. Triacylglycerols from Animal Fats – Milk Fat
Milk fats are the only animal fat designed by nature to serve as a food. The range of fatty acids is more extensive than in adipose tissue with often a higher proportion of short- and medium-chain fatty acids, which are not used as such for structural purposes but provide a rapid source of energy. Ruminants, such as the cow, have a range of saturated fatty acids from butyric upwards (indeed even acetate has been found in esterified form), and there are relatively low amounts only of polyunsaturated fatty acids because of biohydrogenation in the rumen. In other species, the compositions are less extreme. In human milk, for example, 12:0 and 14:0 fatty acids are more abundant than in other tissues, but the linoleic acid concentration varies between 10 and 20% depending on diet.
Severe technical problems were encountered in the stereospecific analysis of milk triacyl-sn-glycerols from ruminants, because of the presence of short-chain fatty acids, which give rise to difficulties in the isolation of the required partially hydrolysed intermediates. Breckenridge, Kuksis et al. (see Table 3) overcame the problem by isolating fractions enriched in either long-chain or short-chain components by means of molecular distillation or thin-layer chromatography, and subjecting these separately to stereospecific analysis, recombining the results at the end of the procedure. These showed unequivocally that cows' milk is one of the most asymmetric of animal fats, containing all the butyric acid and most of the hexanoic acid in position sn-3. Some representative results for the cow and other species are listed in Table 3.
Table 3. The composition of the fatty acids esterified to each position of the triacyl-sn-glycerols in the milk fats of various species.
|tr = trace (< 0.5 %). TG = intact triacylglycerols.
Data from - Christie, W.W. and Moore, J.H. Biochim. Biophys. Acta, 210, 46-56 (1970); Christie, W.W. J. Dairy Res., 52, 219-222 (1985); Christie, W.W. and Clapperton, J.L. J. Soc. Dairy Technol., 35, 22-24 (1982); Breckenridge, W.C. et al., Canad. J. Biochem., 47, 761-769 (1969).
As the overall fatty acid compositions of the triacylglycerols are very different for each species, depending both on the diet and the nature of the fatty acids synthesised in the mammary gland, similarities are not immediately apparent, but a close perusal can reveal certain common features, especially for the longer-chain fatty acids. For example for nearly all species, much of the palmitic acid is concentrated in position sn-2, the only known exception being the primitive monotreme, the echidna. Other than in the pig (see above), a structure with a high content of palmitic acid in position sn-2 is not common in animal triacylglycerols. Myristic acid and the medium-chain fatty acids, like palmitic acid, are found in the greatest concentration in position sn-2, but stearic acid is concentrated in the primary positions, especially position sn-1. The unsaturated fatty acids also tend to be present in the greatest concentrations in positions sn-1 and sn-3.
5. Triacylglycerols from Animal Fats – Other Tissues
The fatty acid compositions of triacyl-sn-glycerols animal tissues other than adipose tissue tend to resemble those of the latter, but the stereospecific distributions can differ. Adrenal tissues can contain longer-chain polyunsaturated fatty acids (10% or more). Such data are often of most value to the biochemist. Some representative analyses are listed in Table 4.
Table 4. Positional distributions of fatty acids (mol %) in triacyl-sn-glycerols from animal tissues other than depot fats and milk.
|a sheep tissues contain appreciable amounts of minor components not listed here
(e.g. odd-and branched-chain, etc.).
TG = intact triacylglycerols.
Data from - Christie, W.W. and Noble, R.C. J. Sci. Food. Agric., 35, 617-624 (1984); Christie, W.W. and Moore, J.H. J. Sci. Food. Agric., 22, 120-124 (1971), Comp. Biochem. Physiol., 41B, 287-295 (1972), Biochim. Biophys. Acta, 218, 83-88 (1970).
The pig and sheep have probably been most studied. With the latter in the liver triacylglycerols, for example, the stearic acid is distributed approximately equally between the three positions, and the oleic acid is present in high concentrations in positions sn-2 and sn-3, thus differing from the depot fats (Table 2). The distinctive feature of the sheep plasma triacylglycerols is a high concentration of palmitic acid in position sn-2, and in this they resemble those of the lymph, from which they are derived biosynthetically. In most other tissues of the sheep, the triacylglycerols resemble those of adipose tissue. However, in adrenal glands from which adhering adipose tissue had been carefully removed, triacylglycerols containing a high proportion of palmitic acid in position sn-2 were again found, and with long-chain polyunsaturated fatty acids (not listed) in positions sn-2 and sn-3. It is possible that triacylglycerols of this kind are more widespread than has been thought and that further research will reveal more examples.
Other distinctive triacylglycerols of animal origin are found in the tissues of the chicken. In the plasma triacylglycerols, for example, palmitic acid comprises over 70% of the fatty acids of position sn-1 with relatively small amounts in positions sn-2 and sn-3, while oleic acid comprised 60% of the fatty acids in position sn-1 and more than 70% of that in position sn-3. Virtually identical structures were found in the ovarian follicles and in the egg, suggesting a common biosynthetic origin. It is important to recognize that lipid structural analyses can rarely be used to prove the existence of particular biosynthetic pathways, but they can provide valuable pointers to the biochemist to potentially productive experimental approaches.
6. Triacylglycerols from Fish Oils
The fatty acid compositions of triacylglycerols of fish oils reflect their diet and usually comprise high concentrations of long chain monoenoic and polyunsaturated fatty acids, especially those of the omega-3 biosynthetic family. The triacyl-sn-glycerols in depot fats from a number of fish and other marine animals have been subjected to stereospecific analysis, and some typical results are listed in Table 5.
Table 5. Positional distributions of fatty acids (mol %) in triacyl-sn-glycerols of fish oils.
|TG = intact triacylglycerols.
Data from Brockerhoff, H., Hoyle, R.J., Hwang, P.C. and Litchfeld, C. Lipids, 3, 24-29 (1968).
Myristic, palmitic and palmitoleic acids are preferentially esterified to positions sn-1 and sn-3. Oleic and longer-chain monoenoic fatty acids are located mainly in the primary positions, with a tendency for a higher proportion to be in position sn-3 as the chain-length increases. Polyunsaturated fatty acids are in greatest concentration in position sn-2 with substantial amounts also being found in position sn-3. There are significant differences in triacylglycerol structure between those of fish and of marine animals that consume fish.
7. Triacylglycerols of Bacteria and Yeast
Triacylglycerols are fairly widespread in eukaryotic organisms such as yeasts, moulds and fungi, but among the prokaryotes only species from the actinomycetes group, including some of the human mycobacterial pathogens, accumulate triacylglycerols to a significant extent. They occur as cytoplasmic inclusions or lipid droplets within the organisms. As in eukaryotes, triacylglycerols appear to function as a reserve of fatty acyl groups. Those from a few species only have been subjected to stereospecific analyses, and the fatty acid compositions of each are very different, so no general conclusions can be drawn. Three sets of data are listed in Table 6.
Table 6. Positional distributions of fatty acids (mol %) in triacyl-sn-glycerols of bacteria and yeast.
|tr = trace (<0.5%). TG = intact triacylglycerols.
Other = branched, odd- or longer-chain.
Data from - Walker, R.W. et al. Lipids, 5, 684-691 (1970); Wältermann, M. et al. Microbiology, 146, 1143-1149 (2000); Haley, J.E. and Jack, R.C. Lipids, 9, 679-681 (1974).
In the triacyl-sn-glycerols of Mycobacterium smegmatis, oleic acid is the main component of position sn-1, palmitic acid is the main component of position sn-2, while C18 and longer-chain fatty acids are the principal constituents of position sn-3; indeed 90% of the 24:0 is in position sn-3. Although the fatty acid composition of Rhodococcus opacus is very different from this, the distribution of fatty acids is somewhat similar in that saturated and shorter-chain fatty acids are concentrated in position sn-2. In the triacylglycerols of the yeast Lipomyces lipoferus, oleic acid is the main fatty acid in positions sn-1 and sn-2, especially, while much of the palmitic, palmitoleic and stearic acids are found in position sn-3.
A variety of chromatographic and spectrometric methods are available for the analysis of molecular species of triacylglycerols, but stereospecific determination of positional distributions of fatty acids remains a substantial technical challenge. Such methods are described elsewhere on this site, for example in the sections Selected Topics in Lipid Analysis and Gas Chromatography and Lipids. Modern mass spectrometric methods are being used increasingly for determination of molecular species compositions in triacylglycerols, but they do not distinguish the primary positions. Also, data from such analyses are not easily presented in tabulated form for comparison purposes.
A further document on this site dealing with triacylglycerols (Triacylglycerols. Part 2) discusses the biochemistry and metabolism of triacylglycerols.
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James Hutton Institute (and Mylnefield Lipid Analysis), Invergowrie, Dundee (DD2 5DA), Scotland.
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