The successive steps of an integrated analytical procedure aimed at the accurate determination of butterfat fatty acid composition, including trans-18:1 acid content and profile, have been carefully checked. This sequential procedure includes: dispersion of a portion of butter in hexane/isopropanol (2:1, vol/vol) with anhydrous Na2SO4, filtration of aliquots of the suspension through a microfiltration unit, subsequent preparation of fatty acid isopropyl esters (FAIPE) with H2SO 4 as a catalyst, and analysis of total FAIPE by capillary gas-liquid chromatography (GLC). Isolation of trans-18:1 isomers was by silver-ion thin-layer chromatography (Ag-TLC), followed by extraction from the gel of combined saturated and trans-monoenoic acids with a biphasic solvent system. Analysis of these fractions by GLC allows the accurate quantitation of trans-18:1 acids with saturated acids (14:0, 16:0, and 18:0) as internal standards. A partial insight in the distribution of trans-18:1 isomers can be obtained by GLC on a CP Sil 88 capillary column (Chrompack, Middelburg, The Netherlands). All steps of the procedure are quite reproducible, part of the coefficients of variation (generally less than 3%, mainly limited to butyric and stearic acids) being associated with GLC analysis (injection, integration of peaks) and, to a lesser extent, to FAIPE preparation. FA1PE appear to be of greater practical interest than any other fatty acid esters, including fatty acid methyl esters (FAME), for the quantitation of short-chain fatty acids, because peak area percentages, calculated by the integrator coupled to the flame-ionization detector, are almost equal (theoretically and experimentally) to fatty acid weight percentages and do not require correction factors. With this set of procedures, we have followed in detail the seasonal variations of fatty acids in butterfat, with sixty commercial samples of French butter collected at five different periods of the year. Important variations occur around mid-April, when cows are shifted from forage and concentrates during winters spent in their stalls to fresh grass in pastures. At this period, there is a decrease of 4:0-14:0 acids and of 16:0 (-2 and -6%, respectively), while 18:0 and cis-plus trans-18:1 acids rise suddenly (2 and 5%, respectively). These modifications then progressively disappear until late fall or early winter. Other variations are of minor quantitative importance. Although influenced by the season, the

The fatty acid composition of seeds from seven species of the genusPinus (P. pinaster, P. griffithii, P. pinea, P. koraiensis, P. sylvestris, P. mughus, andP. nigra) was established. Pine seeds are rich in oil (31–68% by weight) and contain several unusual polymethylene‐interrupted unsaturated fatty acids with acis‐5 ethylenic bond. These are thecis‐5,cis‐9 18:2,cis‐5,cis‐9,cis‐12 18:3,cis‐5,cis‐11 20:2, andcis‐5,cis‐11,cis‐14 20:3 acids, with a trace ofcis‐5,cis‐9,cis‐12,cis‐15 18:4 acid. Their percentage relative to total fatty acids varies from a low of 3.1% (P. pinea) to a high of 30.3% (P. sylvestris), depending on the species. The majorcis‐5 double bond‐containing acid is generally thecis‐5,cis‐9,cis‐12 18:3 acid (pinolenic acid). In all species, linoleic acid represents approximately one‐half the total fatty acids, whereas the content of oleic acid varies in the range 14–36% inversely to the sum of fatty acids containing acis‐5 ethylenic bond. The easily available seeds fromP. koraiensis appear to be a good source of pinolenic acid: their oil content isca. 65%, and pinolenic represents about 15% of total fatty acids. These values appear to be rather constant.Pinus pinaster, which is grown on several thousand acres in the southwest of France, is an interesting source ofcis‐5,cis‐11,cis‐14 20:3 acid (7% in the oil, which isca. 35% of the dehulled seed weight), an acid sharing in common three double bonds with arachidonic acid. Apparently,P. sylvestris seed oil contains the highest level ofcis‐5 double bond‐containing acids among pine seed oils that have ever been analyzed.

Doubling the length of a CP-Sil 88 capillary column (Chrompack, Middelburg, The Netherlands) from 50 to 100 m remarkably improves the resolution of individual trans-18:1 isomers from either ruminant fats or partially hydrogenated oils. Although the use of a 50-m column gives interesting results, it does not allow sufficient resolution of the trans-10 and trans-11 18:1 isomers. Moreover, the trans-6 to trans-9 18:1 isomers emerge as a single group of peaks, whereas the trans-12 isomer is only partly resolved from the adjacent trans-11 and trans-13 plus trans-14 isomers. With the 100-m column, the trans-9, trans-lO, and trans-12 18:1 isomers are almost baseline resolved from other isomers. However, with both columns, it is not possible to separate the critical pair of trans-13 and trans-14 18:1 acids which co-elute under a single peak. Despite this minor drawback, the 100-m CP-Sil 88 column appears to be of great interest for the separation and the quantitation of most individual trans-18:1 acids. Except for the use of argentation thin-layer chromatography, there is no need for complementary techniques, such as ozonolysis. This simple and powerful tool may be applied to ruminant fats, partially hydrogenated oils, and human tissue lipids. JAOCS 72, 1197-1201 (1 995).

The fatty acid composition of twelve French tub margarines and three industrial shortenings was established with particular attention to their trans-18:l acid content. Four of the twelve margarines (including two major brands, with 60% of market share) were devoid of trans isomers, one contained less than 2% trans-18:1 acids, whereas the seven others had a mean content of 13.5 _+ 3.6% trans isomers. Four years ago, no margarines with 0% trans-18:l acids could be found. It is deduced that the recent Dutch and American studies on possible effects of trans acids on human health (serum cholesterol, heart disease risks) may have had some influence on French margarine manufacturers. Presently, an average French tub margarine contains only 3.8% of trans-18:1 acids instead of 13% four years ago. To protect brand names, some manufacturers have replaced partially hydrogenated oils with tropical fats or fully hydrogenated oils. On the other hand, two of the three shortenings had high levels of trans-18:1 acids: 53.5 and 62.5%. This last value, obtained for a sample of hydrogenated arachis oil, seems to be one of the highest values ever reported for edible hydrogenated oils. In this sample, trans-18:1 plus saturated acids accounted for 85% of total fatty acids. This would indicate that shortening producers and users are not yet aware of recent dietary recommendations, probably because these products are not easily identifiable by consumers in food items, in contrast to margarines. JAOCS 72, 1485-1489 (1995).cholesterol among plasma lipoproteins, in such a way that they might increase heart disease risks. This potentiality was further supported by epidemiological studies from Harvard University (4,5), in which a significant positive correlation was found between margarine consumption and increased heart disease risks. In the latter studies, the range of trans acid consumed was in the range of that found in European countries (1,6), and far less than in the Dutch studies.Although these results are still controversial, they are a cause for concern, and it is useful to determine whether the trans studies have had any impact on margarine producers. Four years ago, we analyzed the trans-18:1 acid content of French margarines and related products. Although we did not fully publish our results, we were unable to find O%-trans margarines. The mean content at that time was around 13% (7). We know that French margarine producers are aware of and concerned about the trans-acid problem. Thus, we decided in this study to conduct a second investigation of the trans-acid content of present commercial margarines, after the publication of the Dutch and American studies. Our results clearly show that the major French margarine producers (subsidiaries of two multinational companies) have considerably lowered the trans-acid content in their products, and that a trend toward O%-trans margarines is evident.

To understand the cis-trans isomerization reaction of ethylenic bonds in heated octadecatrienoic acids (occurring during industrial deodorization of oils), we have prepared a mixture of cis-9,cis-12, cis-12,cis-15, and cis-9,cis-15 18:2 acids by partial hydrazine reduction of cis-9,cis-12,cis-15 18:3 acid present in linseed oil. This mixture (as fatty acid methyl esters) was heated under vacuum at 270~ for 2.25 h. The two methylene-interrupted acids isomerize at a similar rate under such conditions, but the nonmethylene-interrupted cis-9,cis-15 18:2 acid remains unchanged. This means that the mechanism of isomerization does not involve a direct interaction between the two external ethylenic bonds as previously hypothesized. The central cis-12 ethylenic bond is apparently necessary for the isomerization of the two external cis-9 and cis-15 ethylenic bonds. However, this bond is itself rather protected against isomerization in the original cis-9,cis-12,cis-15 18:3 acid which is mainly isomerized to trans-9, cis-12,trans-15, cis-9,cis-12,trans-15, and trans-9,cis-12,cis-15 18:3 acids. The cis-9,trans-12,cis-15 18:3 isomer is less than 10% of total trans isomers of (z-linolenic acid. As a general rule, only one of the two double bonds in a methylene-interrupted diethylenic system can undergo cis-trans isomerization when submitted to heat treatment, at least for temperatures equal to or less than 270~ JAOCS 73, 327-332 (1996). KEY WORDS: Geometrical isomerization, heat treatment, (zlinolenic acid, octadecadienoic acids, trans fatty acids. EXPERIMENTAL PROCEDURESHydrazine reduction of fatty acid methyl esters (FAME) prepared with linseed oil. Linseed oil (technical grade) was saponified, and the resulting free fatty acids were transformed into FAME by reacting them with 12% (wt/vol) methanolic BF 3. FAME were then submitted to hydrazine reduction according to Ratnayake (11) and Conway et al. (12). Briefly, FAME (3.5 g) were dissolved in 200 mL of 95% ethanol, maintained at 40~ Hydrazine (2 mL) was added, and a stream of oxygen was applied to the surface of the solution,