Process optimization of simultaneous enzymatic production of 1,3-dioleoyl-2-palmitoylglycerol and 1-oleoyl-2-palmitoyl-3-linoleoylglycerol

1,3-dioleoyl-2-palmitoylglycerol (OPO), a structured triacylglycerols (TAG) containing palmitic acid at sn-2 position, is widely used as breast milk fat substitutes in infant formula. Interestingly, the main component of breast milk fat in Western countries is OPO while Chinese breast milk contains both 1-oleoyl-2-palmitoyl-3-linoleoylglycerol (OPL) and OPO. Thus, OPL is additionally essential for Chinese infants. In the present study, we developed a process for simultaneous production of OPL and OPO. According to the fatty acid (FA) composition of Chinese breast milk, soy oil, palm kernel stearin, and palm stearin were selected as the base oils for random interesterification, and the appropriate ratio was determined as 2:3:15. Then interesterification at sn-1,3 position was conducted, four reaction parameters were examined, where 1.6 molar ratio of oleic acid to linoleic acid (OLA/LNA), 10% enzyme quantity, TAG to FA ratio of 1:8 and reaction temperature of 56°C were ultimately established as the optimal conditions to achieve the target of the present study, and the OPO and OPL contents in the final products were 23.1% and 26.2%, respectively. Finally, molecular distillation was applied to purify above achieved breast milk fat substrates. Higher oil dropping speed, oil temperature, and agitation speed were favorable conditions, and the lowest acid value that achieved in the present study was 3.46 mgKOH/g. Overall, we established an efficient preliminary process for the simultaneous production of OPO and OPL, which could provide an important reference for commercial production of breast milk fat substrates for Chinese infants.     

Production of Human Milk Fat Substitutes by Interesterification of Tripalmitin with Ethyl Oleate Catalyzed by Candida parapsilosis Lipase/Acyltransferase

In human milk fat, the saturated fatty acids, namely palmitic acid, are located at the sn-2 position of triacylglycerols (TAG) while unsaturated fatty acids (e.g. oleic acid) are esterified at position sn-1,3. Thus, sn-1,3-dioleoyl-2-palmitoylglycerol (OPO) is the target TAG to be used as human milk fat substitutes (HMFS) in infant formulas. In this study, the noncommercial recombinant lipase/acyltransferase from Candida parapsilosis (CpLIP2) was immobilized in Accurel MP1000, and used as a biocatalyst for the interesterification of tripalmitin with ethyl oleate in a solvent-free medium, to obtain structured lipids used as HMFS. Different molar ratios (MR) of ethyl oleate to tripalmitin (2:1–8:1) were used. After 4 h reaction at 60°C, about 30 mol% of oleic acid incorporation was already observed for all tested MR. An apparent equilibrium was reached after 8–24 h, with 32–51 mol% final incorporation, increasing with the MR. The incorporation of oleic acid into TAG was compared with the maximum predicted values when a random or a sn-1,3-regioselective biocatalyst was used. The obtained values are consistent with the maximum incorporation expected for a sn-1,3-regioselective enzyme. In fact, the amount of oleic acid at position sn-2 was approximately 15% for all the MR tested, which is explained by the acyl migration phenomenon. CpLIP2 exhibited higher activity than most commercial immobilized lipases (e.g. faster reaction in solvent-free media, low enzyme load, and low MR needed), and showed a recognized sn-1,3 regioselective behavior.     

Preparation of Infant Formula Fat Analog Containing Capric Acid and Enriched with DHA and ARA at the sn-2 Position

An infant formula fat analog with capric acid mostly esterified at the sn‐1,3 positions, and substantial amounts of palmitic, docosahexaenoic (DHA), and arachidonic (ARA) acids at the sn‐2 position, was prepared by physically blending enzymatically synthesized structured lipids (SL) with vegetable oils. The components of the blend included high sn‐2 palmitic acid SL enriched with capric acid (SL_CA), canola oil (CAO), corn oil (CO), high sn‐2 DHA (DHAO_m), and high sn‐2 ARA (ARAO_m) enzymatically modified oils. Each component was proportionally blended to match the fatty acid profile of commercial fat blends used for infant formula. The infant formula fat analog (IFFA₁) was characterized for total and positional fatty acids (FA), triacylglycerol (TAG) molecular species, thermal behavior, and tocopherol content. IFFA₁ contained 17.37 mol% total palmitic acid of which nearly 35 % was located at the sn‐2 position. The total capric acid content was 13.93 mol%. The content of DHA and ARA were 0.49 mol% (48.18 % at sn‐2) and 0.57 mol% (35.80 % at sn‐2), respectively. The predominant TAG were OPO (24.09 %), POP (15.70 %), OOO (11.53 %), and CLC (7.79 %). The melting completion and crystallization onset temperatures were 18.65 and ?2.19 °C, respectively. The total tocopherol content was 566.45 μg/g. This product might be suitable for commercial production of infant formulas.     

Chylomicron and VLDL TAG structures and postprandial lipid response induced by lard and modified lard

Alterations in chylomicron and VLDL TAG and the magnitude of postprandial lipemia were studied in healthy volunteers after two meals of equal FA composition but different TAG-FA positional distribution. Molecular level information of individual lipoprotein TAG regioisomers was obtained with a tandem MS method. The incremental area under the response curve of VLDL TAG was large (P=0.021) after modified lard than after lard. In plasma TAG, the difference did not quite reach statistical significance (P=0.086). In general, there were less TAG with palmitic acid in the sn-2 position and more TAG with oleic acid in the sn-2 position in chylomicrons than in fat ingested. From 1.5 to 8 h postprandially, the proportion of individual chylomicron TAG was constant or influenced by TAG M.W. VLDL TAG regioisomerism was similar regardless of the positional distribution of fat ingested. Significant alterations were seen in VLDL TAG FA, in M.W. fractions, and in individual regioisomers with respect to time. The TAG sn-14∶0-18∶1-18∶1+sn-18∶1-18∶1-14∶0, sn-16∶0-16∶1-18∶1+sn-18∶1-16∶1-16∶0, and sn-16∶1-18∶1-18∶1+sn-18∶1-18∶1-16∶1 decreased (P<0.05); and sn-16∶0-16∶0-18∶2+sn-18∶2-16∶0-16∶0, sn-16∶0-16∶0-18∶1+sn-18∶1-16∶0-16∶0, sn-16∶0-18∶1-16∶0, and sn-16∶0-18∶1-18∶2+sn-18∶2-18∶1-16∶0 increased (P<0.05) after both meals. In conclusion, positional distribution of TAG FA was found to affect postprandial lipid metabolism in healthy normolipidemic subjects.     

Enzymatic Synthesis of Infant Formula Fat Analog Enriched with Capric Acid

A structured lipid (SL) with a substantial amount of palmitic acid at the sn‐2 position and enriched with capric acid (C), was produced in two enzymatic interesterification stages by using immobilized lipase, Lipozyme^® TL IM (Novozymes North America Inc., Franklinton, NC, USA). The substrates for the reactions were high melting point palm stearin, high oleic sunflower oil and tricaprin. The SL was characterized for total and positional fatty acid profiles, triacylglycerol (TAG) molecular species, free fatty acid content, melting and crystallization profiles. The final SL contained 20.13 mol% of total palmitic acid, of which nearly 40 % was located at the sn‐2 position. The total capric acid content was 21.22 mol%, mostly at the sn‐1 and sn‐3 positions. The predominant TAGs in the SL were oleic–palmitic–oleic, POP and CLC. The melting completion and crystallization onset temperatures of the SL were 27.7 and 6.1 °C, respectively. The yield for the overall reaction was 90 wt%. This SL might be totally or partially used in commercial fat blends for infant formula.     

Lipase-assisted acidolysis of high-laurate canola oil with eicosapentaenoic acid

The production of structured lipids via acidolysis of high-laurate canola oil (Laurical 15) with EPA in hexane was carried out using lipase from Pseudomonas sp. The optimal reaction conditions used 4% lipase, at a mole ratio of oil to EPA of 1∶3 at 45°C over 36 h. The positional distribution of FA on the glycerol backbone of unmodified oil indicated that lauric acid was mainly located at the sn-1,3 positions. Stereospecific analysis of the oil modified with EPA showed that lauric acid remained mostly esterified to the sn-1,3 positions of the TAG molecules and that EPA was also primarily in the sn-1,3 positions of the TAG molecules. Thus, the resultant structured lipids may have optimal value for use in applications where quick energy release and EPA supplementation are required.     

Effect of triacylglycerol structures on the thermal oxidative stability of edible oil

Different molecular species of TAG were assessed to determine the influence of TAG structure on the thermal oxidative stability of edible oil. TAG containing palmitic acid (16∶0, P) as saturated FA (SFA) and oleic acid (18∶1, O), linoleic acid (18∶2, L), or linolenic acid (18∶3, Ln) as unsaturated FA (UFA) were chemically synthesized and then heated at 180 or 150°C. Thermal oxidative stability of TAG was determined by evaluating the resultant UFA, polar compound, FFA, carbonyl compound, polymerized compound, and tocopherol contents. When TAG containing 16∶0 and 18∶2 in the ratio of 2∶1 (mol/mol) were heated at 180°C, a 2∶1 (mol/mol) mixture of saturated TAG (PPP) and unsaturated TAG (LLL) was found to be more susceptible to thermal oxidation than PPP/PLL (1∶1) and PPL. Similarly, a 2∶1 mixture of PPP and OOO or LnLnLn was more unstable toward thermal oxidation than PPO or PPLn, respectively. Thermal oxidative stability of TAG containing SFA and UFA (2∶1) was negatively correlated with the moles of UFA in a single TAG molecule. This tendency was also observed at 150°C. From these results, it is suggested that the TAG structure could be one of the factors determining the thermal oxidative stability of edible oil.     

Triacylglycerols of Apiaceae seed oils: Composition and regiodistribution of fatty acids

Oils from the seeds of caraway (Carum carvi), carrot (Daucus carota), celery (Apium graveolens) and parsley (Petroselinum crispum), all from the Apiaceae family, were analyzed by gas chromatography for their triacylglycerol (TAG) composition and fatty acid (FA) distribution between the sn‐1(3) and sn‐2 positions of TAG. Twenty‐two TAG species were quantified. Glyceryl tripetroselinate was the major TAG species in seed oils of carrot, celery and parsley, with levels ranging from 38.7 to 55.3%. In caraway seed oil, dipetroselinoyllinoleoylglycerol was the major TAG species at 21.2%, while the glyceryl tripetroselinate content was 11.4%. Other TAG species were linoleoyloleoylpetroselinoylglycerol and dipetroselinoyloleoylglycerol. Predominantly, TAG were triunsaturated (72.2–84.0%) with diunsaturates at 14.4–25.9%, and small amounts of monounsaturated TAG. Results for regiospecific analysis showed a non‐random FA distribution in Apiaceae for palmitic, petroselinic, linoleic and oleic acids. Petroselinic acid was predominantly located at the sn‐1(3) position in carrot, celery and parsley seed oils, while it was mainly at the sn‐2 position in caraway seed oil. The distribution of linoleic acid was opposite to that of petroselinic acid. Oleic acid was mostly located at the sn‐2 position, except for caraway, where it was evenly distributed between the sn‐1(3) and sn‐2 positions. Both the saturated FA, palmitic and stearic acid, were located mainly at the sn‐1(3) position. The presence of a high level of tripetroselinin in parsley seed oil (55.3%) makes it a potential source for the production of petroselinic acid.     

Synthesis of Infant Formula Fat Analogs Enriched with DHA from Extra Virgin Olive Oil and Tripalmitin

Structured lipids (SLs) containing palmitic, oleic, and docosahexaenoic acids for possible use in infant formulas were synthesized by enzymatic acidolysis reactions. The substrates used were tripalmitin, extra virgin olive oil free fatty acids (EVOOFFA), and docosahexaenoic acid single cell oil free fatty acids (DHASCOFFA) in 1:1:1, 1:2:1, 1:3:2, 1:4:2, and 1:5:1 molar ratios. Reactions were carried out at 65 °C for 24 h using Lipozyme^® TL IM lipase. The products were analyzed for total and positional fatty acids by GC-FID, triacylglycerol (TAG) molecular species by HPLC-ELSD, and thermal behavior by DSC. The SLs, SL132, SL142, and SL151 had desirable fatty acid distribution for infant formula use with nearly 60 mol% palmitic acid at the sn-2 position and oleic acid predominantly at the sn-1,3 positions. The total DHA content of SL132, SL142, and SL151 were 7.54, 6.72, and 5.89 mol%, respectively. The major TAG molecular species in the SLs were PPP, OPO, and PPO. The melting completion temperature of SL132 was 37.1, 35.2 °C in SL142, and 32.9 °C in SL151. The SLs synthesized in this study have potential use in infant formulas.     

Stereospecific analysis of TAG from sunflower seed oil

Stereospecific analysis of TAG from a sunflower seed oil of Tunisian origin was performed. The TAG were first fractionated according to chain length and degree of unsaturation by RP-HPLC. The four major diacid- and triacid-TAG fractions were palmitoyldilinoleoyl-glycerol, dioleoyllinoleoylglycerol, oleoyldilinoleoylglycerol, and palmitoyloleoyl-linoleoyl-glycerol, amounting to 7.2, 16.6, 29.5, and 12 mol%, respectively. The TAG of the four fractions were individually submitted to stereospecific analysis, using a Grignard-based partial deacylation, separation of sn-1,2(2,3)-DAG from sn-1,3-DAG by boric acid-impregnated silica gel TLC plates, conversion of the sn-1,2(2,3)-DAG to their 3,5-dinitrophenylurethane (DNPU) derivatives, fractionation of DNPU derivatives by RP-HPLC, resolution of the DNPU-DAG by HPLC on a chiral column, transmethylation of each sn-DNPU-DAG fraction, and analysis of the resulting FAME by GC. The data obtained were used to determine the triacyl-sn-glycerol composition of the main TAG of the oil. Fifteen triacyl-sn-glycerols were identified and quantified, representing, along with the monoacid-TAG, trilinoleoylglycerol and trioleoylglycerol, more than 90% of the total oil TAG. The two major triacyl-sn-glycerols were trilinoleoyl-glycerol and 1-linoleoyl-2-linoleoyl-3-oleoyl-glycerol (18.6 and 18.5% of the total, respectively). Results clearly identified linoleic acid as the major FA at the sn-2 position, whereas oleic and palmitic acids were the major FA at the sn-3 position. The sn-1 position was occupied to nearly the same extent by linoleic and oleic acids, and to a greater extent by palmitic acid, which was practically absent at the sn-2 position.     

Structured lipids from high-laurate canola oil and long-chain omega-3 fatty acids

The lipase-assisted acidolysis of high-laurate canola oil (HLCO; Laurical 25) with long-chain n−3 FA (DHA and EPA) was studied. Response surface methodology was used to obtain a maximal incorporation of DHA or EPA into HLCO. The studied process variables were the amount of enzyme (2–6%), reaction temperature (35–55°C), and incubation time (12–36 h). The amount of water added and the mole ratio of substrates (oil to DHA or EPA) were kept at 2% and 1∶3, respectively. All experiments were conducted according to a face-centered cube design. Under optimal conditions (4.79% of enzyme; 46.1°C; 30.1 h), the incorporation of DHA into HLCO was 37.3%. The corresponding maximal incorporation of EPA (61.6%) into Laurical 25 was obtained using 4.6% enzyme, a reaction temperature of 39.9°C, and a reaction period of 26.2 h. Examination of the positional distribution of FA on the glycerol backbone of modified HLCO with DHA showed that the DHA was primarily located in the sn-1,3 positions of the TAG molecules. However, lauric acid also remained mainly in the sn-1,3 positions of the modified oil. For EPA-modified Laurical 25, lauric acid was present mainly in the sn-1,3 positions, whereas EPA was randomly distributed over the three positions.     

Use of <Emphasis Type="Italic">Rhizopus delemar</Emphasis> lipase as compared with other lipases for determination of <Emphasis Type="Italic">sn</Emphasis>-2 fatty acids in triacylglycerol

The FA composition in the sn-2 position of TAG is routinely determined after porcine pancreatic lipase hydrolysis. However, the content of saturated FA increased when a pancreatic lipase preparation with higher specific activity was used. Lipase from Rhizopus delemar was selected as a potential replacement lipase for the following reasons: (i) The FA specificity is nearly equivalent in hydrolysis activity toward FA such as lauric, myristic, palmitic, palmitoleic, stearic, oleic, linoleic, and α-linolenic acids; and (ii) lipase from R. delemar hydrolyzes fatty acyl residues at the sn-1,3 positions of TAG. Acyl migration products were present at less than 0.8% in lipase hydrolysates containing 6–14% of sn-2 MAG. A reproducibility CV of less than 5% was obtained in a collaborative study in which the compositions of the main FA at the sn-2 position in olive oil were determined using lipase from R. delemar. This article was presented in part at the Biocatalysis Symposium, 94th AOCS Annual Meeting & Expo, Kansas City, Missouri, May 2003.     

Triacylglycerol structure of human colostrum and mature milk

Because triacylglycerol (TAG) structure influences the metabolic fate of its component fatty acids, we have examined human colostrum and mature milk TAG with particular attention to the location of the very long chain polyunsaturated fatty acid on the glycerol backbone. The analysis was based on the formation of various diacylglycerol species from human milk TAG upon chemical (Grignard degradation) or enzymatic degradation. The structure of the TAG was subsequently deduced from data obtained by gas chromatographic analysis of the fatty acid methyl esters in the diacylglycerol subfractions. The highly specific TAG structure observed was identical in mature milk and colostrum. The three major fatty acids (oleic, palmitic and linoleic acids) each showed a specific preference for a particular position within milk TAG: oleic acid for thesn-1 position, palmitic acid for thesn-2 position and linoleic acid for thesn-3 position. Linoleic and α-linolenic acids exhibited the same pattern of distribution and they were both found primarily in thesn-3 (50%) andsn-1 (30%) positions. Their longer chain analogs, arachidonic and docosahexaenoic acids, were located in thesn-2 andsn-3 positions. These results show that polyunsaturated fatty acids are distributed within the TAG molecule of human milk in a highly specific fashion, and that in the first month of lactation the maturation of the mammary gland does not affect the milk TAG structure.     

Absorption of isomeric,palmitic acid-containing triacylglycerols resembling human milk fat in the adult rat

The effect of the positional distribution of palmitic acid (16∶0) in triacylglycerols (TAG) on 16∶0 apparent absorption in adult rats was investigated. The rats were fed two diets which contained 30 energy % as fat with identical total fatty acid compositions, both containing 30% 16∶0. The Betapol diet contained TAG with 73% of total 16∶0 in thesn-2 position, the control diet contained TAG with 6% of total 16∶0 in thesn-2 position. After six weeks on these diets, the rats were killed two or six hours after the last meal, and the small intestine was removed, cut into 10-cm segments, and the fatty acid composition of the segment's contents was determined. At both time points the amount of 16∶0 in the intestinal segments starting at 40 cm from the stomach was much lower in the animals fed Betapol than in the animals fed the control diet. Overall absorption of 16∶0 and stearic acid was significantly greater in the Betapol group. Absorption of oleic and linoleic acid from the small intestine was similar in both groups, although the overall absorption was significantly greater in the animals fed Betapol. Total fat absorption was significantly higher in the Betapol-fed rats than in the control-fed rats. No effect on calcium and nitrogen absorption, on plasma total cholesterol and TAG levels, and on bodyweights (growth) was seen. The data demonstrate that the positional distribution of the fatty acids in the TAG molecule affects the site of absorption in the small intestine and particularly the net absorption of saturated fatty acids.     

Aspergillus oryzae Lipase-Catalyzed Synthesis of Dioleoyl; Palmitoyl-Rich Triacylglycerols in Two Reactors

Dioleoyl; palmitoyl‐rich triacylglycerols (OPO‐rich TAG) were synthesized through Aspergillus oryzae lipase (AOL)‐catalyzed acidolysis of palm stearin with commercial oleic acid by a one‐step process in a stirred tank reactor and continuous packed bed reactor to evaluate the feasibility of using immobilized AOL. AOL was found to be valuable for the synthesis of OPO‐rich TAG when compared with commercial lipase from Thermomyces lanuginose (Lipozyme^® TL IM; Novozymes A/S, Bagsvaerd, Denmark). The C52 (triglycerides with a carbon number of 52, stands for OPO, OPL, LPL and their isomers) content of AOL was higher (45.65 %), and the intensity of treatment (IOT: lipase amount × reaction time/TAG amount) of AOL was just 6.25 % of that of Lipozyme^® TL IM under similar reaction conditions in the stirred tank reactor. Response surface methodology were used to optimize the reaction conditions of the AOL‐catalyzed acidolysis is reaction in the packed bed reactor. The optimal point for the set of experimental conditions generated were as follows: residence time 3.0 h; temperature 62.09 °C; substrate molar ratio 7.13 mol/mol. The highest C52 content obtained was 48.60 ± 2.36 %, with 57.46 ± 1.72 % total palmitic acid at the sn‐2 position and 74.21 ± 2.45 % oleic acid at the sn‐1,3 positions. The half‐life of AOL was 24 h in the stirred tank reactor and 140 h in the packed bed reactor. The immobilized AOL achieved similar conversion and selectivity to commercial lipases for the catalyzed synthesis of OPO‐rich TAG and may offer a cheaper alternative.     

Distribution of medium-chain FA in different lipid classes after administration of specific structured TAG in rats

Structured TAG (STAG) containing medium-chain FA (MCFA) in the sn-1,3 positions and essential FA in the sn-2 position were synthesized by lipase-catalyzed acidolysis. In our previous studies we found that part of the MCFA from STAG could be absorbed in the small intestine; however, it was unclear how they were absorbed. In order to get a better understanding of the metabolism of STAG to improve future design and application of STAG, in the present study lymph lipids collected after feeding STAG were fractionated into different classes and the FA composition of each lipid class was studied by GC after methylation to FAME. Caprylic acid was detected in the fraction of TAG only after administration of 1,3-dioctanoyl-2-linoleyl-sn-glycerol (8∶0/18∶2/8∶0), whereas lauric acid was detected in TAG, DAG, and FFA as well as phospholipids after administration of 1,3-didodecanoyl-2-linoleyl-sn-glycerol (12∶0/18∶2/12∶0). We conclude that the enterocyte has the ability to reacylate the MCFA into TAG and that the intestinal absorption of MCFA from STAG mainly occurs by resynthesis of TAG. Caprylic acid from STAG is not incorporated into phospholipids, whereas lauric acid from STAG can be incorporated into phospholipids.     

Lipase-catalyzed acidolysis of tristearin with oleic or caprylic acids to produce structured lipids

Two different structured lipids (SL) were synthesized by transesterifying tristearin with caprylic acid (C8∶0) or oleic acid (C18∶1). The objective was to synthesize SL containing stearic acid (C18∶0) at the sn-2 position as possible nutritional and low-calorie fats. The reaction was catalyzed by IM60 lipase from Rhizomucor miehei in the presence of n-hexane. The effects of reaction parameters affecting the incorporation of caprylic acid into tristearin were compared with those for incorporating oleic acid into tristearin. For all parameters studied, oleic acid incorporation was higher than caprylic acid. The range of conditions favorable for synthesizing high yields of C8∶0-containing SL was narrower than for oleic acid. An incubation time of 12–24 h and an enzyme content of 5% (w/w total substrates) favored C8∶0 incorporation. The mole percentage of incorporated C18∶1 did not increase further at enzyme additions greater than 10%. C18∶1 incorporation decreased with the addition of more than 10% water (w/w total substrates) to the tristearin-oleic acid reaction mixture. Increasing the mole ratio of fatty acid (FA) to triacylglycerol increased oleic acid incorporation. The highest C8∶0 incorporation was obtained at a 1∶6 mole ratio of tristearin to FA. Positional analysis confirmed that C18∶0 remained at the sn-2 position of the synthesized SL. The melting profiles of tristearin-caprylic acid and tristearin-oleic acid SL displayed peaks between −20 to 30°C and −20 to 40°C, respectively. Their solid fat contents (∼25%) at 25°C suggest possible use in spreads or for inclusion with other fats in specialized blends.     

Regiospecific Analysis of Shark Liver Triacylglycerols

The liver oils of six shallow-water shark species, silky (Carcharhinus falciformis), thresher (Alopias superciliosus), oceanic whitetip (Carcharhinus longimanus), blue (Prionace glauca), hammerhead (Sphyrna lewini) and salmon (Lamna ditropis) were analyzed with particular attention to the regioisomeric composition of triacylglycerols (TAG). The TAG compositions were analyzed by using an HPLC-evaporative light scattering detector and each molecular species identified by HPLC-atmospheric pressure chemical ionization/mass spectrometry. Major lipid components of all sharks’ oils were TAG (~80 %) made up of omega-3 polyunsaturated fatty acids at 26–40 % and 20–25 % docosahexaenoic acid (DHA). Forty different molecular species were detected in the TAG fractions. TAG consisting of one palmitic acid, one DHA and one oleic acid (12.5–19.9 %) and TAG consisting of two palmitic acids and one DHA (8.4–15.4 %) were the predominant form while 30–50 % TAG molecular species were bound to one or more DHA. Distribution of fatty acids in the primary (sn-1 and sn-3) and secondary (sn-2) position of the glycerol backbones was examined by regiospecific analysis by using pancreatic lipase and it was found that DHA was preferentially positioned at sn-2. These findings greatly extend the utilization of shark liver oils in food productions and may have a significant impact on the future development of the fish oil industry.     

Re-esterified Palm Oils,Compared to Native Palm Oil,do not Alter Fat Absorption,Postprandial Lipemia or Growth Performance in Broiler Chicks

Re-esterified palm oils are obtained from the chemical esterification of palm acid oils (rich in free fatty acids) with glycerol, both economically interesting by-products from oil refining and biodiesel industries, respectively. Thus, re-esterified palm oils could be an economically interesting alternative to native palm oil in broiler chick diets. However, because they may have different physicochemical properties than have their corresponding native oil, we assessed the effect of fatty acid (FA) positional distribution within acylglycerol molecules and the effect of acylglycerol composition on FA apparent absorption, and their possible consequences on the evolution of postprandial lipemia and growth performance in broiler chicks. Seventy-two 1-day-old female broiler chicks were randomly distributed into 18 cages. The three treatments used were the result of a basal diet supplemented with 6 wt% of native palm oil (N-TAG), re-esterified palm oil (E-TAG), or re-esterified palm oil high in mono- and diacylglycerols (E-MDAG). Chemical esterification raised the fraction of palmitic acid at the sn-2 position from 9.63 mol% in N-TAG oil to 17.9 mol% in E-TAG oil. Furthermore, E-MDAG oil presented a high proportion of mono- (23.1 wt%) and diacylglycerols (51.2 wt%), with FA mainly located at the sn-1,3 positions, which resulted in a lower gross-energy content and an increased solid-fat index at the chicken’s body temperature. However, re-esterified palm oils did not alter fat absorption, postprandial lipemia, or growth performance, compared to native palm oil, so they can be used as alternative fat sources in broiler chick diets.     

Enzymatic preparation of enantiomerically pure sn-2,3-diacylglycerols: A stereoselective ethanolysis approach

Stereoselective ethanolysis of monoacid TAG by immobilized Rhizomucor miehei lipase (RML) was studied for preparation of optically pure sn-2,3-DAG. Trioctanoylglycerol (TO) was used as a model substrate. The enantiomeric purity of the product, sn-2,3-dioctanoylglycerol (sn-2,3-DO), was very high (percent enantiomeric excess >99%) when an excess of ethanol was used. The result indicated that RML was highly stereoselective toward the sn-1 position of TO under conditions of excess ethanol. The stereoselectivity of RML depended on the amount of ethanol. The larger the amount of ethanol was, the higher the stereoselectivity became. After optimizing the parameters such as reactant molar ratio, water content, and temperature, (ethanol/TO molar ratio =31∶1 and water content =7.5 wt% of the reactants at 25°C), optically pure sn-2,3-DO was obtained at 61.1 mol% in the glyceride fraction in 20 min. The above conditions were further applied for ethanolysis of monoacid TAG with different acyl groups such as tridecanoylglycerol (C10∶0), tridodecanoylglycerol (C12∶0), tritetradecanoylglycerol (C14∶0) and trioctadecenoylglycerol [triolein, (C18∶1)]. The yields and enantiomeric purities of 1,2(2,3)-DAG were dramatically reduced when TAG with FA longer than decanoic acid were used.