Metabolic diversity in biohydrogenation of polyunsaturated fatty acids by lactic acid bacteria involving conjugated fatty acid production

Lactobacillus plantarum AKU 1009a effectively transforms linoleic acid to conjugated linoleic acids of cis-9,trans-11-octadecadienoic acid (18:2) and trans-9,trans-11–18:2. The transformation of various polyunsaturated fatty acids by washed cells of L. plantarum AKU 1009a was investigated. Besides linoleic acid, α-linolenic acid [cis-9,cis-12,cis-15-octadecatrienoic acid (18:3)], γ-linolenic acid (cis-6,cis-9,cis-12–18:3), columbinic acid (trans-5,cis-9,cis-12–18:3), and stearidonic acid [cis-6,cis-9,cis-12,cis-15-octadecatetraenoic acid (18:4)] were found to be transformed. The fatty acids transformed by the strain had the common structure of a C18 fatty acid with the cis-9,cis-12 diene system. Three major fatty acids were produced from α-linolenic acid, which were identified as cis-9,trans-11,cis-15–18:3, trans-9,trans-11,cis-15–18:3, and trans-10,cis-15–18:2. Four major fatty acids were produced from γ-linolenic acid, which were identified as cis-6,cis-9,trans-11–18:3, cis-6,trans-9,trans-11–18:3, cis-6,trans-10–18:2, and trans-10-octadecenoic acid. The strain transformed the cis-9,cis-12 diene system of C18 fatty acids into conjugated diene systems of cis-9,trans-11 and trans-9,trans-11. These conjugated dienes were further saturated into the trans-10 monoene system by the strain. The results provide valuable information for understanding the pathway of biohydrogenation by anaerobic bacteria and for establishing microbial processes for the practical production of conjugated fatty acids, especially those produced from α-linolenic acid and γ-linolenic acid. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Biotransformation of Unsaturated Long-Chain Fatty Acids by Eubacterium lentum

Eubacterium lentum (33 strains) isomerized the 12-cis double bond of C₁₈ fatty acids with cis double bonds at C-9 and C-12 into an 11-trans double bond before reduction of the 9-cis double bond. The 14-cis double bond of homo-γ-linolenic acid was isomerized by 29 strains into a 13-trans double bond. The same strains isomerized the 14-cis double bond of arachidonic acid into a 13-trans double bond and then isomerized the 8-cis double bond into a 7-trans double bond; the 13-cis double bond of 10-cis, 13-cis-nonadecadienoic acid was isomerized into a 12-trans double bond. None of these isomerization products was further reduced. Studies with resting cells showed optimal isomerization velocity at a linoleic acid concentration of 37.5 μM; higher concentrations were inhibitory. The pH optimum for isomerization was 7.5 to 8.5. The isomerase was inhibited by the sulfhydryl reagents iodoacetamide, bromoacetate, and N-ethylmaleimide and by the chelators EDTA and 1,10-phenanthroline.     

Fat source and dietary forage-to-concentrate ratio influences milk fatty-acid composition in lactating cows

On the basis of the potential benefits to human health there is an increased interest in producing milk containing lower-saturated fatty acid (SFA) and higher unsaturated fatty acid (FA) concentrations, including cis-9 18:1 and cis-9, trans-11-conjugated linoleic acid (CLA). Twenty-four multiparous Holstein cows were used in two experiments according to a completely randomized block design, with 21-day periods to examine the effects of incremental replacement of prilled palm fat (PALM) with sunflower oil (SFO) in high-concentrate diets containing 30 g/kg dry matter (DM) of supplemental fat (Experiment 1) or increases in the forage-to-concentrate (F : C) ratio from 39 : 61 to 48 : 52 of diets containing 30 g/kg DM of SFO (Experiment 2) on milk production, digestibility and milk FA composition. Replacing PALM with SFO had no effect on DM intake, but tended to increase organic matter digestibility, yields of milk, protein and lactose, and decreased linearly milk fat content. Substituting SFO for PALM decreased linearly milk fat 8:0 to 16:0 and cis-9 16:1, and increased linearly 18:0, cis-9 18:1, trans-18:1 (Δ4 to 16), 18:2 and CLA concentrations. Increases in the F : C ratio of diets containing SFO had no effect on intake, yields of milk, milk protein or milk lactose, lowered milk protein content in a quadratic manner, and increased linearly NDF digestion and milk fat secretion. Replacing concentrates with forages in diets containing SFO increased milk fat 4:0 to 10:0 concentrations in a linear or quadratic manner, decreased linearly cis-9 16:1, trans-6 to -10 18:1, 18:2n-6, trans-7, cis-9 CLA, trans-9, cis-11 CLA and trans-10, cis-12 CLA, without altering milk fat 14:0 to 16:0, trans-11 18:1, cis-9, trans-11 CLA or 18:3n-3 concentrations. In conclusion, replacing prilled palm fat on with SFO in high-concentrate diets had no adverse effects on intake or milk production, other than decreasing milk fat content, but lowered milk fat medium-chain SFA and increased trans FA and polyunsaturated FA concentrations. Increases in the proportion of forage in diets containing SFO increased milk fat synthesis, elevated short-chain SFA and lowered trans FA concentrations, without altering milk polyunsaturated FA content. Changes in fat yield on high-concentrate diets containing SFO varied between experiments and individual animals, with decreases in milk fat secretion being associated with increases in milk fat trans-10 18:1, trans-10, cis-12 CLA and trans-9, cis-11 CLA concentrations.     

Biohydrogenation of C20 polyunsaturated fatty acids by anaerobic bacteria

The PUFAs include many bioactive lipids. The microbial metabolism of C₁₈ PUFAs is known to produce their bioactive isomers, such as conjugated FAs and hydroxy FAs, but there is little information on that of C₂₀ PUFAs. In this study, we aimed to obtain anaerobic bacteria with the ability to produce novel PUFAs from C₂₀ PUFAs. Through the screening of ∼100 strains of anaerobic bacteria, Clostridium bifermentans JCM 1386 was selected as a strain with the ability to saturate PUFAs during anaerobic cultivation. This strain converted arachidonic acid (cis-5,cis-8,cis-11,cis-14-eicosatetraenoic acid) and EPA (cis-5,cis-8,cis-11,cis-14,cis-17-EPA) into cis-5,cis-8,trans-13-eicosatrienoic acid and cis-5,cis-8,trans-13,cis-17-eicosatetraenoic acid, giving yields of 57% and 67% against the added PUFAs, respectively. This is the first report of the isolation of a bacterium transforming C₂₀ PUFAs into corresponding non-methylene-interrupted FAs. We further investigated the substrate specificity of the biohydrogenation by this strain and revealed that it can convert two cis double bonds at the ω6 and ω9 positions in various C₁₈ and C₂₀ PUFAs into a trans double bond at the ω7 position. This study should serve to open up the development of novel potentially bioactive PUFAs.     

Feeding heat-oxidized oil to dairy cows affects milk fat nutritional quality

Heating oil and oilseeds results in oxidation products that affect ruminal biohydrogenation of polyunsaturated fatty acids, altering milk fatty acids profile, and could be transferred to milk. An experiment was conducted to investigate the effects of oil heating on rumen and milk fatty acids profile and the transfer of oxidation products to milk. Sunflower oil was heated at 150°C for 15 h and given to lactating dairy cows in a 2×2 arrangement: two groups of two cows, equipped with a ruminal cannula and receiving two diets (containing either heated or unheated oil) during two experimental periods. Oil heating generated hydroperoxides and/or hydroxyacids and aldehydes, in particular trans-2,trans-4-decadienal. In milk, heated oil only significantly decreased trans-11-C18:1 and cis-9,trans-11-CLA percentage compared to non-heated oil, and slightly increased cis-9,cis-12-C18:2 percentage, which was probably linked to an inhibition of the ruminal Δ12 isomerase by oxidative products in the rumen. However, feeding highly oxidized oil did not result in the appearance of hydroperoxides or hydroxyacids in milk and did not increase milk aldehydes content.     

Effects of flaxseed,raw soybeans and calcium salts of fatty acids on apparent total tract digestibility,energy balance and milk fatty acid profile of transition cows

Oilseeds offer some protection to the access of ruminal microorganisms and may be an alternative to calcium salts of fatty acids (FA), which are not fully inert in the ruminal environment. This study aimed to evaluate the effects of different sources of FA supplementation on apparent total tract nutrient digestibility, milk yield and composition, and energy balance (EB) of cows during the transition period and early lactation. We compared diets rich in C18:2 and C18:3 FA. Multiparous Holstein cows were randomly assigned to receive one of the four diets: control (n=11); whole flaxseed (WF, n=10), 60 and 80 g/kg (diet dry matter (DM) basis) of WF during the prepartum and postpartum periods, respectively; whole raw soybeans (WS, n=10), 120 and 160 g/kg (diet DM basis) of WS during the prepartum and postpartum periods, respectively; and calcium salts of unsaturated fatty acids (CSFA, n=11), 24 and 32 g/kg (diet DM basis) of CSFA during the prepartum and postpartum periods, respectively. Dry cows fed WF had higher DM and net energy of lactation (NE_L) intake than those fed WS or CSFA. The FA supplementation did not alter DM and NDF apparent total tract digestibility, dry cows fed WF exhibited greater NDF total tract digestion than cows fed WS or CSFA. Feeding WS instead of CSFA did not alter NE_L intake and total tract digestion of nutrients, but increased milk fat yield and concentration. Calculated efficiency of milk yield was not altered by diets. FA supplementation increased EB during the postpartum period. Experimental diets increased long-chain FA (saturated and unsaturated FA) in milk. In addition, cows fed WS and CSFA had higher C18:1 trans-11 FA and C18:2 cis, and lower C18:3 FA in milk than those fed WF. Furthermore, cows fed CSFA had higher C18:1 trans-11 and cis-9, trans-11 FA than cows fed WS. Although supplemental C18:2 and C18:3 FA did not influence the milk yield of cows, they positively affected EB and increased unsaturated long-chain FA in milk fat.     

Combined effects of oleic,linoleic and linolenic acids on lactation performance and the milk fatty acid profile in lactating dairy cows

The potential combined effects of oleic, linoleic and linolenic acids supplementation on lactation performance and the milk fatty acid (FA) profile in dairy cows have not been well investigated. Our objective was to examine the effects of supplementation with a combination of these FA as well as the effects of removing each from the combination on lactation performance and the milk FA profile in dairy cows. Eight Holstein cows (101±11 days in milk) received four intravenously infused treatments in a 4×4 Latin square design, and each period lasted for 12 days which consisted of 5 days of infusion and 7 days of recovery. The control treatment (CTL) contained 58.30, 58.17 and 39.96 g/day of C18: 1 cis-9; C18: 2 cis-9, cis-12; and C18: 3 cis-9, cis-12, cis-15, respectively. The other three treatments were designated −−C18: 1 (20.68, 61.17 and 41.72 g/day of C18: 1 cis-9; C18: 2 cis-9, cis-12; and C18: 3 cis-9, cis-12, cis-15, respectively), −C18: 2 (61.49, 19.55 and 42.13 g/day of C18: 1 cis-9; C18: 2 cis-9, cis-12; and C18: 3 cis-9, cis-12, cis-15, respectively) and −C18: 3 (60.89, 60.16 and 1.53 g/day of C18: 1 cis-9; C18: 2 cis-9, cis-12; and C18: 3 cis-9, cis-12, cis-15, respectively). Dry matter intake and lactose content were not affected by the treatments, but the milk protein content was lower in cows treated with −C18: 2 than that in CTL-treated cows. Milk yield as well as milk fat, protein and lactose yields were higher in cows treated with −C18: 3 than the yields in CTL-treated cows, and these yields increased linearly as the unsaturation degree of the supplemental FA decreased. Compared with the CTL treatment, the −C18: 2 treatment decreased milk C18: 2 cis-9 content (by 2.80%) and yield (by 22.12 g/day), and the −C18: 3 treatment decreased milk C18: 3 cis-9, cis-12, cis-15 content (by 2.72%) and yield (by 22.33 g/day). In contrast, removing C18: 1 cis-9 did not affect the milk content or yield of C18: 1 cis-9. The −C18: 2-treated cows had a higher C18: 1 cis-9 content and tended to have a higher C18: 1 cis-9 yield than CTL-treated cows. The yields of C8: 0, C14: 0 and C16: 0 as well as <C16: 0 tended to increase linearly as the unsaturation degree of the supplemental FA decreased (P=0.06, 0.07, 0.07 and 0.09, respectively). These results indicated that supplementation with C18 unsaturated FA might not independently affect the lactation performance and the milk FA profile of dairy cows.     

Effect of extruded linseeds alone or in combination with fish oil on intake,milk production,plasma metabolite concentrations and milk fatty acid composition in lactating goats

Based on the potential benefits for long-term human health, there is interest in developing sustainable nutritional strategies for lowering medium-chain saturated fatty acids (FA) and increasing specific unsaturated FA in ruminant milk. Dietary supplements of extruded linseeds (EL), fish oil (FO) or a mixture of EL and FO increase cis-9,trans-11 CLA and long-chain n-3 polyunsaturated FA in bovine milk. Supplements of FO cause milk fat depression in lactating cows, but information for dairy goats is limited. A total of 14 Alpine goats were used in a replicated 3×3 Latin square with 28-days experimental periods to examine the effects of EL alone or in combination with FO on animal performance, milk fat synthesis and milk FA composition. Treatments comprised diets based on natural grassland hay supplemented with no additional oil (control), 530 of EL or 340 g/day of EL and 39 g/day of FO (ELFO). Compared with the control, ELFO tended (P=0.08) to lower milk fat yield, whereas EL increased (P<0.01) milk fat content and yield (15% and 10%, respectively). Relative to EL, ELFO decreased (P<0.01) milk fat content and yield (19% and 17%, respectively). Relative to the control and ELFO, EL decreased (P<0.05) milk 10:0 to 16:0 and odd- and branched-chain FA content and increased 18:0, cis-18:1, trans-13 18:1 (and their corresponding ∆-9 (desaturase products), trans-12,cis-14 CLA, cis-13,trans-15 CLA, cis-12,trans-14 CLA and trans-11,cis-13 CLA and 18:3n-3 concentrations. ELFO was more effective for enriching (P<0.05) milk cis-9, trans-11 CLA and trans-11 18:1 concentrations (up to 5.4- and 7.1-fold compared with the control) than EL (up to 1.7- and 2.5-fold increases). Furthermore, ELFO resulted in a substantial increase in milk trans-10 18:1 concentration (5.4% total FA), with considerable variation between individual animals. Relative to the control and EL, milk fat responses to ELFO were characterized by increases (P<0.05) in milk trans-16:1 (Δ9 to 11), trans-18:1 (Δ6 to 11), trans-18:2, CLA (cis-9,trans-11, trans-9,cis-11, trans-8,trans-10 and trans-7,trans-9) and 20- and 22-carbon FA concentrations. Overall, EL resulted in a relatively high cis-9 18:1 concentration and an increase in the 18:3n-3/18:2n-6 ratio, whereas combining EL and FO resulted in substantial increases in trans-FA, marginal enrichment in 20:5n-3 and 22:6n-3 and lower 16:0 concentration changes associated with a decrease in milk fat content. In conclusion, data provide further evidence of differential mammary lipogenic responses to diet in the goat compared with the cow and sheep.     

The biohydrogenation of α-linoleic acid and oleic acid by rumen micro-organisms

1. α-[U-¹⁴C]Linolenic acid was incubated with the rumen contents of sheep and the metabolic products were characterized by thin-layer chromatography, gas–liquid chromatography and absorption spectroscopy in the ultraviolet and infrared. 2. A tentative scheme for the biohydrogenation route to stearic acid is presented. The main pathway is through diconjugated cis–cis–cis-octadecatrienoic acid, non-conjugated trans–cis (cis–trans)-octadecadienoic acid and trans-octadecenoic acid, but other pathways are apparent. 3. Washed rumen micro-organisms possessed only a limited capacity to hydrogenate α-linolenic acid and oleic acid but the rate was greatly stimulated by a factor(s) present in the supernatant rumen liquor. 4. Pure cultures of Clostridium perfringens, Streptococcus faecalis, Escherichia coli and a coliform organism isolated from sheep faeces possessed negligible ability to hydrogenate unsaturated fatty acids compared with a mixed population of rumen micro-organisms. Butyrivibrio fibrisolvens slowly converted linoleic acid into octadecenoic acid.     

Substrate selectivity of various lipases in the esterification of cis- and trans-9-octadecenoic acid

The substrate selectivity of numerous commercially available lipases from microorganisms, plants and animal tissue towards 9-octadecenoic acids with respect to the cis/trans configuration of the CC double bond was examined by the esterification of cis- and trans-9-octadecanoic acid (oleic and elaidic acid respectively) with n-butanol in n-hexane. A great number of lipases studied, e.g. those from Pseudomonas sp., porcine pancreas or Carica papaya, were unable to discriminate between the isomeric 9-octadecenoic acids. However, lipases from Candida cylindracea and Mucor miehei catalysed the esterification of oleic acid 3–4 times faster than the corresponding reaction of elaidic acid and therefore have a high preference for the cis isomer. Of all biocatalysts examined, only recombinant lipases from Candidaantarctica favoured elaidic acid as substrate. While the preference of Candida antarctica lipase B for the trans isomer was quite low, Candida antarctica lipase A had an extraordinary substrate selectivity and its immobilized enzyme preparation [Chirazyme L-5 (3) from Boehringer] esterified elaidic acid about 15 times faster than oleic acid. Received: 29 October 1998 / Received revision: 18 December 1998 / Accepted: 21 December 1998     

Effects of starch-rich or lipid-supplemented diets that induce milk fat depression on rumen biohydrogenation of fatty acids and methanogenesis in lactating dairy cows

Optimizing milk production efficiency implies diets allowing low methane (CH₄) emissions and high dairy performance. We hypothesize that nature of energy (starch v. lipids) and lipid supplement types (monounsaturated fatty acid (MUFA) v. polyunsaturated fatty acid (PUFA) mitigate CH₄ emissions and can induce low milk fat content via different pathways. The main objective of this experiment was to study the effects of starch-rich or lipid-supplemented diets that induce milk fat depression (MFD) on rumen biohydrogenation (RBH) of unsaturated fatty acids (FA) and enteric CH₄ emissions in dairy cows. Four multiparous lactating Holstein cows (days in milk=61±11 days) were used in a 4×4 Latin square design with four periods of 28 days. Four dietary treatments, three of which are likely to induce MFD, were based (dry matter basis) on 56% maize silage, 4% hay and 40% concentrates rich in: (1) saturated fatty acid (SFA) from Ca salts of palm oil (PALM); (2) starch from maize grain and wheat (MFD-Starch); (3) MUFA (cis-9 C18:1) from extruded rapeseeds (MFD-RS); and (4) PUFA (C18:2n-6) from extruded sunflower seeds (MFD-SF). Intake and milk production were measured daily. Milk composition and FA profile, CH₄ emissions and total-tract digestibility were measured simultaneously when animals were in open-circuit respiration chambers. Fermentation parameters were analysed from rumen fluid samples taken before feeding. Dry matter intake, milk production, fat and protein contents, and CH₄ emissions were similar among the four diets. We observed a higher milk SFA concentration with PALM and MFD-Starch, and lower milk MUFA and trans-10 C18:1 concentrations in comparison to MFD-RS and MFD-SF diets, while trans-11 C18:1 remained unchanged among diets. Milk total trans FA concentration was greater for MFD-SF than for PALM and MFD-Starch, with the value for MFD-RS being intermediate. Milk C18:3n-3 content was higher for MFD-RS than MFD-SF. The MFD seems more severe with MFD-SF and MFD-RS than PALM and MFD-Starch diets, because of a decrease in milk SFA concentration and a stronger shift from trans-11 C18:1 to trans-10 C18:1 in milk. The MFD-SF diet increased milk trans FA (+60%), trans-10 C18:1 (+31%), trans-10,cis-12 CLA (+27%) and PUFA (+36%) concentrations more than MFD-RS, which explains the numerically lowest milk fat yield and indicates that RBH pathways of PUFA differ between these two diets. Maize silage-based diets rich in starch or different unsaturated FA induced MFD with changes in milk FA profiles, but did not modify CH₄ emissions.     

Effects of dietary sources of vegetable fats on performance of dairy ewes and conjugated linoleic acid (CLA) in milk

Two experiments were carried out to study the effects of supplementing the ration of lactating ewes with vegetable fats (sunflower oil, SO or hydrogenated palm oil, HPO; HIDROPALM^®) on diet digestibility, milk yield and milk composition, and on the concentration of the conjugated linoleic acid (CLA) C18:2 cis-9 trans-11 and C18:1 trans-11 (vaccenic acid, VA) and other main fatty acids in milk fat. Treatments involved a control diet, without added oil, and 2 diets supplemented with either 12 g/kg SO or 12 g/kg HPO on a dry matter (DM) basis. In the first experiment, 6 non-pregnant, non-lactating Lacaune ewes were used following a 3 × 3 replicated Latin Square design. Addition of vegetable fat supplement to the diet increased digestibility of DM, organic matter (OM) and crude protein (CP), but did not affect that of the ether extract (EE), neutral detergent fibre (NDF) or acid detergent fibre (ADF). In the second experiment, 60 Lacaune dairy ewes mid-way through lactation (120 ± 12 days in milk, 0.98 ± 0.03 kg/day average milk yield) were divided into three equal-sized groups each of which was assigned to one of the three experimental diets for 4 weeks. Compared with the control treatment, supplementation with HPO increased milk yield and energy-corrected milk. But neither vegetable fat supplement modified percentages of fat and protein in milk. Supplementation with HPO increased C14:1, C16:1 and C16:0 content and reduced C18:0 and C18:1 cis-9 content in milk fat. Supplementation with SO increased the VA content in milk fat by 36% and that of cis-9 trans-11 CLA by 29% in comparison with the control diet. Supplementation with HPO led to milk fat with 15% more cis-9 trans-11 CLA than control milk. In conclusion, adding a moderate dose of HPO or SO to the diets increased CLA concentration in milk fat. Nevertheless, supplementation with SO was more effective than HPO in increasing CLA concentration in milk fat and reducing the atherogenicity index, improving milk quality from the human health standpoint.     

Effect of double bond geometry in sphingosine base on the antioxidant function of sphingomyelin

We previously showed that sphingomyelin (SM) inhibits peroxidation of phosphatidylcholine (PC) and cholesterol. Since SM uniquely has a trans unsaturation in its sphingosine base, we investigated whether this feature is important for its antioxidant function. Substitution of the natural trans Δ⁴-double bond with a cis double bond (cis-SM), however, increased SM’s ability to inhibit Cu²⁺-mediated 16:0-18:2 PC oxidation by up to eightfold. Dihydro-SM, which lacks the double bond, was equally effective as trans-SM. In contrast to its effect in the sphingosine base, the presence of a cis double bond in the N-acyl group of trans-SM was not protective. cis-SM also inhibited the oxidation of cholesterol by FeSO₄/ascorbate more efficiently than the trans isomer. The enhanced protective effect of cis-SM is selective for metal ion-promoted oxidation, and appears to arise from a decrease in the effective concentration of metal ions. These studies show that the trans double bond of SM is not essential for its antioxidant effects.     

On the Specificity of a Fatty Acid Epoxygenase in Broad Bean (Vicia faba L.)

Seeds of broad bean (Vicia faba L.) contain a hydroperoxide-dependent fatty acid epoxygenase. Hydrogen peroxide served as an effective oxygen donor in the epoxygenase reaction. Fifteen unsaturated fatty acids were incubated with V. faba epoxygenase in the presence of hydrogen peroxide and the epoxy fatty acids produced were identified. Examination of the substrate specificity of the epoxygenase using a series of monounsaturated fatty acids demonstrated that (Z)-fatty acids were rapidly epoxidized into the corresponding cis-epoxy acids, whereas (E)-fatty acids were converted into their trans-epoxides at a very slow rate. In the series of (Z)-monoenoic acids, the double bond position as well as the chain length influenced the rate of epoxidation. The best substrates were found to be palmitoleic, oleic, and myristoleic acids. Steric analysis showed that most of the epoxy acids produced from monounsaturated fatty acids as well as from linoleic and α-linolenic acids had mainly the (R),(S) configuration. Exceptions were C₁₈ acids having the epoxide group located at C-12/13, in which cases the (S),(R) enantiomers dominated. 13(S)-Hydroxy-9(Z),11(E)-octadecadienoic acid incubated with epoxygenase afforded the epoxy alcohol 9(S),10(R)-epoxy-13(S)-hydroxy-11(E)-octadecenoic acid as the major product. Smaller amounts of the diastereomeric epoxy alcohol 9(R),10(S)-epoxy-13(S)-hydroxy-11(E)-octadecenoic acid as well as the α,β-epoxy alcohol 11(R),12(R)-epoxy-13(S)-hydroxy-9(Z)-octadecenoic acid were also obtained. The soluble fraction of homogenate of V. faba seeds contained an epoxide hydrolase activity that catalyzed the conversion of cis-9,10-epoxyoctadecanoic acid into threo-9,10-dihydroxyoctadecanoic acid.     

Butters rich either in trans-10-C18:1 or in trans-11-C18:1 plus cis-9, trans-11 CLA differentially affect plasma lipids and aortic fatty streak in experimental atherosclerosis in rabbits

Dairy fat contains high amounts of saturated fatty acids (FA), which are associated with cardiovascular disease (CVD) risk. Manipulation of dairy cows nutrition allows to decrease the saturated FA content of milk fat, and is associated with increases either in conjugated linoleic acid (CLA) and trans-11-C18:1 contents, or in trans-10-C18:1 content. CLA putatively exhibits beneficial properties on CVD risk, whereas trans FA are suspected to be detrimental. The present study compared the effects of a trans-10-C18:1-rich butter (T10 butter), a trans-11-C18:1+CLA-rich butter (T11-CLA butter) and a standard butter (S butter) on lipid parameters linked to the CVD risk and fatty streaks. Thirty-six White New Zealand rabbits were fed one of the three butters (12% of the diet, plus 0.2% cholesterol) for 6 (experiment 1) or 12 (experiment 2) weeks. Liver lipids, plasma lipids and lipoprotein concentrations (experiments 1 and 2) and aortic lipid deposition (experiment 2) were determined. The T10 butter increased VLDL-cholesterol compared with the two others, and total and LDL-cholesterol compared with the T11-CLA butter ( P < 0.05). The T10 butter also increased non-HDL/HDL ratio and aortic lipid deposition compared with the T11-CLA butter ( P < 0.05). The T11-CLA butter non-significantly reduced aortic lipid deposition compared with the S butter, and decreased HDL-cholesterol and increased liver triacyglycerols compared with the two other butters (< 0.05). These results suggest that, compared with the S butter, the T10 butter had detrimental effects on plasma lipid and lipoprotein metabolism in rabbits, whereas the T11-CLA butter was neutral or tended to reduce the aortic lipid deposition.     

Rubber seed oil and flaxseed oil supplementation alter digestion,ruminal fermentation and rumen fatty acid profile of dairy cows

Rubber seed oil (RO) that is rich in polyunsaturated fatty acids (FA) can improve milk production and milk FA profiles of dairy cows; however, the responses of digestion and ruminal fermentation to RO supplementation in vivo are still unknown. This experiment was conducted to investigate the effect of RO and flaxseed oil (FO) supplementation on nutrients digestibility, rumen fermentation parameters and rumen FA profile of dairy cows. Forty-eight mid-lactation Holstein dairy cows were randomly assigned to one of four treatments for 8 weeks, including basal diet (CON) or the basal dietary supplemented with 4% RO, 4% FO or 2% RO plus 2% FO on a DM basis. Compared with CON, dietary oil supplementation improved the total tract apparent digestibility of DM, neutral detergent fibre and ether extracts ( P < 0.05). Oil treatment groups had no effects on ruminal digesta pH value, ammonia N and microbial crude protein ( P > 0.05), whereas oil groups significantly changed the volatile fatty acid (VFA) profile by increasing the proportion of propionate whilst decreasing total VFA concentration, the proportion of acetate and the ratio of acetate to propionate ( P < 0.05). However, there were no differences in VFA proportions between the three oil groups (P > 0.05). In addition, dietary oil supplementation increased the total unsaturated FA proportion in the rumen by enhancing the proportion of trans-11 C18:1 vaccenic acid (VA), cis-9, trans-11 conjugated linoleic acid (CLA) and α-linolenic acid (ALA) ( P < 0.05). These results indicate that dietary supplementation with RO and FO could improve nutrients digestibility, ruminal fermentation and ruminal FA profile by enhancing the VA, cis-9, trans-11 CLA and ALA composition of lactating dairy cows. These findings provide a theoretical basis for the application of RO in livestock production.     

Lipoxygenase-catalyzed transformation of epoxy fatty acids to hydroxy-endoperoxides: a potential P450 and lipoxygenase interaction

Herein, we characterize a generally applicable transformation of fatty acid epoxides by lipoxygenase (LOX) enzymes that results in the formation of a five-membered endoperoxide ring in the end product. We demonstrated this transformation using soybean LOX-1 in the metabolism of 15,16-epoxy-α-linolenic acid, and murine platelet-type 12-LOX and human 15-LOX-1 in the metabolism of 14,15-epoxyeicosatrienoic acid (14,15-EET). A detailed examination of the transformation of the two enantiomers of 15,16-epoxy-α-linolenic acid by soybean LOX-1 revealed that the expected primary product, a 13S-hydroperoxy-15,16-epoxide, underwent a nonenzymatic transformation in buffer into a new derivative that was purified by HPLC and identified by UV, LC-MS, and ¹H-NMR as a 13,15-endoperoxy-16-hydroxy-octadeca-9,11-dienoic acid. The configuration of the endoperoxide (cis or trans side chains) depended on the steric relationship of the new hydroperoxy moiety to the enantiomeric configuration of the fatty acid epoxide. The reaction mechanism involves intramolecular nucleophilic substitution (S_Ni) between the hydroperoxy (nucleophile) and epoxy group (electrophile). Equivalent transformations were documented in metabolism of the enantiomers of 14,15-EET by the two mammalian LOX enzymes, 15-LOX-1 and platelet-type 12-LOX. We conclude that this type of transformation could occur naturally with the co-occurrence of LOX and cytochrome P450 or peroxygenase enzymes, and it could also contribute to the complexity of products formed in the autoxidation reactions of polyunsaturated fatty acids.     

Production of trans-10, cis-12 Conjugated Linoleic Acid in Rice

Conjugated linoleic acid (CLA) has anti-carcinogenic and anti-atherosclerosis activity, and modulatory effects on the immune system and lipid metabolism. To produce a transgenic rice plant that can accumulate CLA, a linoleate isomerase gene that can convert linoleic acid to trans-10, cis-12 CLA was introduced and expressed under the control of seed-specific promoters from the oleosin and globulin genes. The fatty acid composition of the transgenic rice grain was analyzed by gas chromatography. Although there was no clear difference in the fatty acid composition between seeds from transformed versus untransformed plants, a peak of trans-10, cis-12 CLA methyl ester, which was not present in seeds from untransformed plants, was found in transformed plants. The trans-10, cis-12 CLA comprised an average of 1.3% (w/w) of the total fatty acids in seeds carrying the oleosin promoter in comparison to 0.01% (w/w) in seeds carrying the globulin promoter. In addition, approximately 70 and 28% of the total amount of the CLA isomer were present in the triacylglycerol and free fatty acid fractions, respectively. These results demonstrate the ability to produce fatty acid components of vegetable oils with novel physiological activities in crops.