Enzymatic synthesis of medium‐chain triacylglycerols by alcoholysis and interesterification of copra oil using a crude papain lipase preparation

We have examined the possibility of producing analogs of medium‐chain triglycerides (MCT) from copra oil, i.e. a triacylglycerol mixture with a high content of medium‐chain fatty acid moieties (C₆–C₁₀). A two‐step enzymatic process was used in which copra triacylglycerols were first split with papain lipase by alcoholysis with an alkyl alcohol and then subjected to interesterification with the alkyl esters recovered using papain lipase. Effects of temperature, water activity content, substrate ratio, biocatalyst amount, and alcohol chain length were also investigated. On the one hand, the sn‐3 stereoselectivity of the lipase in the alcoholysis of copra oil with butanol has permitted a direct enrichment of caproic, caprylic and capric moieties in the synthesized butyl esters. Thus, in the batch reactor, the reaction led to about 31% conversion of the oil after 24 h, and the content of C₆–C₁₀ acids in the synthesized esters increased from about 16% in the starting oil to almost 42%. A similar enzymatic alcoholysis in a packed‐bed column bioreactor gave 31% conversion of the oil after 120 min of reactor residence time. The reaction was also very selective because the C₆–C₁₀ fatty acyl groups represented about half of the newly formed butyl esters, whereas they accounted for only 16% of total fatty acids in the starting oil. On the other hand, the transesterification of the alkyl esters recovered (highly enriched in C₆–C₁₀ fatty acyl groups) with native copra oil directly led to an increase in the content of MCT in the oil, from 18 mol‐% at the beginning of the reaction to 61 mol‐% of MCT after a time period of 72 h in the batch reactor.     

Galacto-oligosaccharide production with immobilized β-galactosidase in a packed-bed reactor vs. free β-galactosidase in a batch reactor

We report here that the usage of immobilized enzyme in a continuous packed bed reactor (PBR) can be a good alternative for GOS production instead of the traditional use of free enzyme in a batch reactor. The carbohydrate composition of the product of the PBR with immobilized enzyme was comparable to that of the batch reactor with free enzyme. The stability of the immobilized enzyme at a lactose concentration of 38% (w/v) and at 50 °C was very high: the half-life time of the immobilized enzyme was approximately 90 days. The enzymatic productivity of GOS production using immobilized enzyme in a PBR can be more than six times higher than that of GOS production with free enzyme in a batch reactor. Besides, when aiming for an equal volumetric productivity to the batch process in designing a PBR, the volume of the PBR can be much smaller than that of the batch reactor, depending on the enzyme dosage and the run time of a single batch.     

Micro‐reactor for transesterification of plant seed oils

The fatty acid compositions of vegetable or other plant seed oils are generally determined by gas chromatography (GC). Methyl esters (the most volatile derivatives) are the preferred derivatives for GC analysis. Esters of higher alcohols are good for the separation of volatile and positional isomers. All the esters of the C₁–C₈ alcohols of vegetable oils were silmilarly prepared by passing the reaction mixture containing the desired alcohol, oil and tetrahydrofuran through the micro‐reactor (a 3‐mL dispossible syringe packed with 0.5 g of NaOH powder). The reaction products were acidified with acetic acid and the mixture was analyzed by high‐performance size exclusion chromatography and GC. Transesterification was quantitative for primary alcohols, but an appreciable amount of free fatty acids was formed for secondary alcohols. Coriander seed oil was quantitatively esterified with 2‐ethyl 1‐hexanol with the micro‐reactor in less than 1 min. Oleic and petroselinic acid 2‐ethyl 1‐hexyl esters are baseline separated on an Rtx‐2330 capillary column (30 m×0.25 mm, 0.25 µm film thickness).     

Production of structured triglycerides rich in n-3 polyunsaturated fatty acids by the acidolysis of cod liver oil and caprylic acid in a packed-bed reactor: equilibrium and kinetics

Structured triglycerides (ST) enriched in n-3 polyunsaturated fatty acids (PUFAs) (eicosapentaenoic acid, EPA, and docosahexaenoic acid, DHA) in position 2 of the triglyceride backbone were synthesised by acidolysis of cod liver oil (CLO) and caprylic acid (CA) catalysed by the 1,3-specific immobilised lipase Lipozyme IM. The reaction was carried out in three ways: (1) in a batch reactor (where the influence of temperature on the incorporation of CA into the CLO triglyceride was studied); (2) in an immobilised lipase packed-bed reactor (PBR) by recirculating the reaction mixture from the exit of the bed to the substrate reservoir (product recirculation) to determine the equilibrium composition; and (3) in a PBR without recirculation. A “lag” period of duration inversely proportional to the initial water amount of the lipase, was observed when new lipase was used. Apparently, during this “lag” period the hydro-enzymatic layer that surrounds the lipase surface reaches its water equilibrium content. A reaction scheme, where only the fatty acid in the positions 1 and 3 of the glycerol backbone were exchanged by CA, was proposed. The exchange equilibrium constants between CA and the native fatty acids of CLO were determined. The n-3 PUFAs (EPA and DHA) were the most resistant native fatty acids to exchange with exchange equilibrium constants of 1.32 and 0.28, respectively. Also, average reaction rates and kinetic constants of exchange of CA and native fatty acid of CLO were calculated. Low kinetic constants were observed for EPA, DHA and palmitic acid. For acidolysis reaction in the continuous mode PBR, the lipase amount/(flow rate × substrate concentration) ratio (m_L/q[TG]₀) could be considered as the intensive variable of the process for use in scale up of the PBR. A simple equation was proposed for the prediction of the fatty acid composition of the ST at the exit of the PBR as a function of the intensive variable m_L/q[TG]₀. At equilibrium, the ST produced had the following composition: CA 57%, EPA 5.1%, DHA 10.0% and palmitic acid 6.3% (only considering the major fatty acids). In addition, the proportion of EPA and DHA that esterified the position 2 of the ST was 13.5%, which represented 44% of the total fatty acids in the position 2 of the resultant ST.     

Dietary modulation of phospholipid fatty acid composition and lipoxygenase products in mouse lung homogenates

This study investigated the potential of dietary fats to modulate the arachidonic acid content of mouse lung phospholipids and the formation of lipoxygenase products from arachidonic and eicosapentaenoic acids. Prior to breeding, female mice were fed for five months diets with 10 wt% of either olive oil, safflower oil, fish oil, or linseed oil. The same diets were fed to the females during gestation and to the pups from day 18 to day 42 postpartum. On day 42, the phospholipids were extracted from fresh lung tissue and separated into classes [phosphatidylethanolamine (PE), phosphatidylserine (PS), phosphatidylcholine (PC), and phosphatidylinositol (Pl)] by thin-layer chromatography. Methyl esters of phospholipid fatty acids and unesterified fatty acids were analyzed by gas chromatography. At comparable dietary n-3/n-6 ratios, arachidonic acid was reduced 85 and 75% in lungs from mice fed linseed oil and fish oil, respectively, compared to lungs of safflower oil-fed mice. Dietary fats affected the proportion of arachidonic acid in phospholipids in the order: PE>PS>PS>Pl. Following incubation of homogenized lung tissue, the total amount of 12-lipoxygenase products was lowest in lungs from mice fed olive oil, and 12-hydroxyeicosatetraenoic acid was lowest in incubated lungs from mice fed linseed oil. Comparison of the amounts of lipoxygenase substrate fatty acids in the individual phospholipids with the lipoxygenase products suggested that the major substrate pool for the 12-lipoxygenase pathway in mouse lung homogenates was PC.     

Preparation and Characterization of Human Milk Fat Substitutes Based on Triacylglycerol Profiles

Human milk fat substitutes (HMFS) having similarity in (TAG) composition to human milk fat (HMF) were prepared by Lipozyme RM IM‐catalyzed interesterification of lard blending with selected oils in a packed bed reactor. Four oil blends with high similarity in fatty acid profiles to HMF were first obtained based on the blending model and then the blending ratios were screened based on TAG composition similarity by enzymatic interesterification in a batch reactor. The optimal ratio was determined as lard:sunflower oil:canola oil:palm kernel oil:palm oil:algal oil:microbial oil = 1.00:0.10:0.50:0.13:0.12:0.02:0.02. This blending ratio was used for a packed bed reactor and the conditions were then optimized as residence time, 1.5 h; reaction temperature, 50 °C. Under these conditions, the obtained product showed high degrees of similarity in fatty acid profile with 39.2 % palmitic acid at the sn‐2 position, 0.5 % arachidonic acid (n‐6) and 0.3 % docosahexaenoic acid (n‐3) and the scores for the degree of similarity in TAG composition was increased from 58.4 (the oil blend) to 72.3 (the final product). The packed bed reactor could be operated for 7 days without significant decrease in activity. The final product presented similar melting and crystallization profiles to those of HMF. However, due to the loss of tocopherols during deacidification process, the oxidative stability was lower than that of the oil blend. This process for the preparation of HMFS from lard with high similarity in TAG composition by physical blending and enzymatic interesterification, as optimized by mathematical models in a packed bed reactor, has a great potential for industrialization.     

The effect of substrate concentrations on the production of biodiesel by lipase‐catalysed transesterification of vegetable oils

All the kinetic studies, found in the literature, on the production of biodiesel (fatty acids methyl esters) considered the esterifications of free fatty acids rather than the transesterification of the vegetable oil itself. The main industrial interest, however, is for the production of biodiesel with the triglyceride (oil) being the substrate. A mathematical model taking into account the mechanism of the methanolysis reaction starting from the vegetable oil as substrate, rather than the free fatty acids, has been developed. From the proposed model equation, the regions where the effect of alcohol inhibition fades, at different substrate concentrations, were identified. The proposed model equation can be used to predict the rate of methanolysis of vegetable oils in a batch or a continuous reactor and to determine the optimal conditions for biodiesel production. Copyright © 2005 Society of Chemical Industry     

Increasing n-3 polyunsaturated fatty acid content of fish oil by temperature control of lipase-catalyzed acidolysis

An attempt was made to further increase the content of n-3 polyunsaturated fatty acid (n-3 PUFA) of fish oil by lipase-catalyzed acidolysis (reaction between fish oil and n-3 PUFA-enriched free fatty acid) without solvent. A bioreactor system was constructed composed of a water-jacketed packed-bed column and a substrate reservoir with a circulation pipeline between the packed-bed column and the reservoir. By keeping the temperature of the reservoir at −10°C (for the first 20 h), followed by −20°C (for the subsequent 40 h) during the batch acidolysis, crystals of free fatty acid appeared, which were removed intermittently by a cotton plug packed in the tip of the outlet pipe in the reservoir. The n-3 PUFA content of the triacylglycerol fraction increased a further 10% by the reduced temperature of the reservoir. Bioreactors for Enzymatic Reaction of Fats and Fatty Acid derivatives, Part XV.     

Strategies for lipase‐catalyzed production and the purification of structured phospholipids

This work provides different strategies for the enzymatic modification of the fatty acid composition in soybean phosphatidylcholine (PC) and the subsequent purification. Enzymatic transesterification reactions with caprylic acid as acyl donor were carried out in continuous enzyme bed reactors with a commercial immobilized lipase (Lipozyme RM IM) as catalyst. Operative stability of the immobilized lipase was examined under solvent and solvent‐free conditions. The long reaction time required to have a high incorporation, combined with rapid deactivation of the enzyme, makes the solvent‐free transesterification reaction unfavorable. Performing the reaction in the presence of solvent (hexane) makes it possible to have high incorporation into PC and deactivation of the lipase is less pronounced as compared to solvent‐free operations. For solvent‐free operation, it is suggested to recycle the reaction mixture through the packed bed reactor, as this would increase incorporation of the desired fatty acids, due to increased contact time between substrate and enzyme in the column. Removal of free fatty acids from the reaction mixture can be done by ultrafiltration; however, parameters need to be selected with care in order to have a feasible process. No changes are observed in the phospholipid (PL) distribution during ultrafiltration, and other techniques as column chromatography may be required if high purity of individual PL species is desired. LC/MS analysis of transesterified PC revealed the presence of 8:0/8:0‐PC, showing that acyl migration takes place during the acidolysis reaction.     

Identification of C18 Intermediates Formed During Stearidonic Acid Biohydrogenation by Rumen Microorganisms In Vitro

In vitro batch incubations were used to study the rumen biohydrogenation of unsaturated fatty acids. An earlier study using increasing supplementation levels of stearidonic acid (18:4n-3), revealed that the rumen microbial population extensively biohydrogenates 18:4n-3 after 72 h of in vitro incubation, though several intermediates formed were not completely characterized. Therefore, in the present study, samples were reanalyzed in order to identify the 18:2, 18:3 and 18:4 biohydrogenation intermediates of 18:4n-3. Gas–liquid chromatography coupled to mass spectrometry was used to characterize these intermediates. The acetonitrile chemical ionization mass spectrometry of the fatty acid methyl esters derivatives enabled the discrimination of fatty acids as non-conjugated or conjugated biohydrogenation intermediates. In addition, the acetonitrile covalent adduct chemical ionization tandem mass spectrometry yielded prominent ions indicative of the double bond position of the major 18:3 isomers, i.e. Δ5,11,15 18:3. Furthermore, the 4,4-dimethyloxazoline derivatives prepared from the fatty acid methyl esters enabled the structure of novel 18:2, 18:3 and 18:4 biohydrogenation intermediates to be elucidated. The intermediates accumulated in the fermentation media after 72 h of incubation of 18:4n-3 suggest that similar to the biohydrogenation pathways of linoleic (18:2n-6) and α-linolenic (18:3n-3) acids, the pathway of the 18:4n-3 also proceeds with the formation of conjugated fatty acids followed by hydrogenation, although no conjugated dienes were found. The formation of the novel biohydrogenation intermediates of 18:4n-3 seems to follow an uncommon isomerization pattern with distinct double bond migrations.     

Production of Structured Triacylglycerol via Enzymatic Interesterification of Medium-Chain Triacylglycerol and Soybean Oil Using a Pilot-Scale Solvent-Free Packed Bed Reactor

Oils rich in medium- and long-chain triacylglycerols (MLCT) serve as functional oils to help reduce body fat accumulation and weight gain. However, most of the MLCT-rich products on the market are physical blends of medium- and long-chain triacylglycerols (MCT and LCT, respectively) that are not structured triacylglycerols (TAG). In this study, an efficient pilot-scale packed bed reactor (PBR) of immobilized lipase from Thermomyces lanuginosus (Lipozyme® TL IM, Novozymes, Bagsvaerd, Denmark) was employed for producing structured MLCT via 1,3-specific interesterification of TAG enriched in caprylic and capric acyl groups and soybean oil (SBO). The PBR was operated under continuous recirculation mode in the absence of solvent. Optimal reaction conditions were determined to be: caprylic/capric TAG: SBO ratio (45:55 w/w), reaction temperature (75 °C) and residence time (16.0 min) on MLCT production were studied. When employing a pilot-scale PBR (100 kg day⁻¹) under optimal conditions, a product containing 76.61 wt% MLCT was produced. Lipozyme TL IM was reused for 25 successive batch reactions (125 kg substrates) with no significant reduction in catalytic efficiency. The light yellow MLCT-enriched product had a high level of saturated fatty acids (SFA, 82.74 wt%) in its sn-2 position as a result of the enzyme's 1,3-positional specificity. One-stage molecular distillation and methanol extraction were used to remove the free fatty acids, mono-, and diacylglycerols generated from hydrolysis. With distillation temperature of 150 °C and oil-to-methanol ratio of 1:3 v/v, MLCT content was further increased to 80.07 wt%. The enzymatic PBR was therefore effective in producing structured MLCT at a pilot-scale under solvent-free conditions.     

Enzymatic conversion of waste edible oil to biodiesel fuel in a fixed-bed bioreactor

The conversion of waste edible oil to biodiesel fuel in a fixed-bed bioreactor was investigated. Three-step methanolysis of waste oil was conducted using three columns packed with 3 g of immobilized Candida antarctica lipase. A mixture of waste oil and 1/3 molar equivalent of methanol against total fatty acids in the oil was used as substrate for the first-step reaction, and mixtures of the first- and second-step eluates and 1/3 molar equivalent of methanol were used for the second- and third-step reactions, respectively. Ninety percent of waste oil was converted to the corresponding methyl esters (ME) by feeding substrate mixtures into the first, second, and third reactors at flow rates of 6, 6 and 4 mL/h, respectively. We also attempted one-step methanolysis of waste oil. When a mixture of waste oil and 90% ME-containing eluate (1∶3, wt/wt) and an equimolar amount of methanol against total fatty acids in the waste oil was fed into a reactor packed with 3 g of immobilized C. antarctica lipase at a flow rate of 4 mL/h, the ME content in the eluate reached 90%. The immobilized biocatalyst could be used for 100 d in the two reaction systems without significant decrease in its activity. Waste oil contained 1980 ppm water and 2.5% free fatty acids, but these contaminants had little influence on enzymatic production of biodiesel fuel.     

Cyclopropenoid Fatty Acids in Malagasy baobab: Adansonia grandidieri (Bombacaceae) Seed Oil

Adansonia grandidieri (bombacaceae family) seed oil gives a positive Halphen test. Composition analysis of derivatized fatty acid methyl esters, in presence of silver nitrate in anhydrous methanol, after chromatography fractionation on silicagel column, were made by gas-liquid chromatography (GLC) using a packed DEGS column. Presence of malvalic and sterculic acids were detected. GLC analysis using glass capillary columns coated with Carbowax 20 M and BDS shows that A. grandidieri seed oil contains mainly palmitic (41%), oleic (22%) and linoleic (12%) acids. Cyclopropenic fatty acid concentration was 14% with 6% for malvalic and 8% for sterculic acids. A slight proportion of dihydrosterculic acid (1.5%) was observed. GLC fatty acid methyl esters analysis, without derivatization, on the two glass capillary columns coated with Carbowax 20 M and BDS gave the same results for cyclopropenic acids content.     

Characterization of wax esters in the roe oil of amber fish,Seriola aureovittata

The wax esters of the roe oil of the amber fish,Seriola aureovittata, have been resolved by high-performance liquid chromatography (HPLC) in the silver-ion mode. Each of the fractions collected was transmethylated, and the fatty acids and alcohols were identified by gas chromatography/mass spectrometry (GC/MS) as the picolinyl esters and nicotinates, respectively. Their compositions were determined by GC. The fatty acid composition is complex, and the main components are C18:1n-9 (35.5 mol%), C22:6n-3 (20.3 mol%), and C16:1n-7 (10.7 mol%), while fatty alcohols are limited to saturated (C16:0, 60.3 mol%; C18:0, 15.3 mol%; C14:0, 5.1 mol%) and monoenoic alcohols (C18:1n-9, 6.5 mol%; C16:1n-7, 4.5 mol%) with traces (<0.1 mol%) of polyunsaturated fatty alcohols such as C20:3n-3, C20:4n-6, C20:5n-3, and C22:5n-3. Silver-ion HPLC exhibited excellent resolution in which fractions were resolved on the basis of the number and configuration of double bonds as well as the distribution pattern between the acid and alcohol moieties of the molecules with a given number of double bonds. The main wax ester fraction are those of monoenoic acid-saturated alcohol species, hexaenoic acid/saturated alcohol species, and pentaenoic acid/saturated alcohol species. Appreciable specificity was observed in the esterification of fatty acids with alcohols, and surprisingly, no saturated acid-monoenoic alcohol species were detected.     

Column types for the chromatographic analysis of oleochemicals

Chromatography has developed into one of the principle methods of analysis of oleochemicals. Gas chromatography has been used extensively for the analysis of long-chain fatty acids as well as for the analysis of triglycerides and plant sterols. In recent years, high pressure liquid chromatography (HPLC) has been used for the analysis of triglycerides as well as for other related materials. Specialized gas chromatography columns have been developed for the separation of long-chain fatty acids such as the methyl esters. These columns have generally used high polarity stationary phases which separate fatty acids by degree of unsaturation. A specialized use of these high polarity stationary phases is separation ofcis-trans isomers as well ascis-cis andtrans-trans isomers. In this paper, packed and capillary columns are compared for the separation of thecis-trans isomers of fatty acid methyl esters prepared from a hydrogenated vegetable oil. For HPLC separations, the presence of a double bond is approximately equivalent chromatographically to shortening the alkyl chain by two carbons. The long-chain polyenic acids or ethyl esters thus elute near but are resolved from the short-chain saturated fatty acids or esters. HPLC is the method of choice for relatively complex, high molecular weight, or labile esters, such as those of retinyl or cholesterol. Glyceryl esters are particularly well resolved by HPLC in terms of both total chain length and degree of unsaturation. This technique is also useful for lipid class separations and for the analysis of modified fatty acid products, such as prostaglandins and related materials. In general, these analyses are conducted with octadecyl bonded phase column packings.     

Identification and Ruminal Outflow of Long-Chain Fatty Acid Biohydrogenation Intermediates in Cows Fed Diets Containing Fish Oil

The abundance of 20- to 24-carbon fatty acids in omasal digesta of cows fed grass silage-based diets supplemented with 0 (Control) and 250 g/day of fish oil (FO) was examined to investigate the fate of long-chain unsaturated fatty acids in the rumen. Complimentary argentation thin-layer chromatography and gas-chromatography mass-spectrometry analysis of fatty acid methyl esters and corresponding 4,4-dimethyloxazoline derivatives prepared from fish oil and omasal digesta enabled the structure of novel 20- to 22-carbon fatty acids to be elucidated. Compared with the Control, the FO treatment resulted in the formation and accumulation of 27 novel 20- and 22-carbon biohydrogenation intermediates containing at least one trans double bond and the appearance of cis-14 20:1, 20:2n-3, 21:4n-3 and 22:3n-6 not contained in fish oil. No conjugated ≥20-carbon fatty acids were detected in Control or FO digesta. In conclusion, fish oil in the diet results in the formation of numerous long-chain biohydrogenation intermediates in the rumen of lactating cows. Comparison of the intake and flow of 20-, 21- and 22-carbon fatty acids at the omasum in cows fed the Control and FO treatments suggests that the first committed steps of 20:5n-3, 21:5n-3 and 22:6n-3 hydrogenation in the rumen involve the reduction and/or isomerisation of double bonds closest to the carboxyl group.     

Interesterification of sesame oil and a fully hydrogenated fat using an immobilized lipase catalyst in both batch and continuous‐flow processes

Lipase‐mediated interesterification of sesame oil and a fully hydrogenated soybean oil was studied at 70 °C in both a batch reactor (BR) and a continuous‐flow packed‐bed reactor (PBR) using four different initial weight ratios of substrates (90 : 10, 80 : 20, 70 : 30 and 60 : 40) with Lipozyme TL IM (Thermomyces lanuginosa) as the biocatalyst. Reaction rates were determined by following the dependence of the profile of the product triacylglycerols (TAG) on the reaction time (BR) or the space time (PBR) via RP‐HPLC‐ELSD. Product TAG identities were confirmed by HPLC‐APCI‐MS. Primary differences between the performances of the two reactors were the maximum level of net hydrolysis (ca. 3 and 10 wt‐% lower acylglycerols at equilibrium for the PBR and BR, respectively), the time or space time required to approach quasi‐equilibrium conditions, and less migration of acyl groups in the PBR trials. For the BR trials, quasi‐equilibrium conditions were approached in 4–6 h, while for the PBR trials short space times (15 min to 2 h) were sufficient to produce effluent compositions similar to equilibrium BR compositions. The predominant TAG families formed by interesterification were LLS, PSO, PSL, SSL, and SSO (L = linoleic; S = stearic; P = palmitic; O = oleic). Oxidative stabilities, melting profiles and solid fat contents were determined for selected reaction products.     

Synthesis of linear alkylbenzene in a liquid–solid circulating fluidized bed reactor

The alkylation reaction of benzene with 1-dodecene was investigated in a liquid–solid circulating fluidized bed reactor (LSCFBR) using HY zeolite as catalyst. The conversion of 1-dodecene, the apparent reaction rate constant, the activity of catalyst and the product distribution in the LSCFBR were analysed and compared with the values obtained using a stirred tank batch reactor (STBR).     

Curupira tefeensis (Olacaceae) — A Rich Source of Very Long Chain Fatty Acids

The fatty acid composition of the seed oil of Curupira tefeensis was analysed by capillary GC of their methyl esters. The gaschromatographic assignments were ensured by corresponding mass spectra. The oil is composed to more than 62% of very long chain fatty acids (>C18). Erucic acid is found to be the main component (35%). The position of the double bonds of the monounsaturated fatty acids (MUFAME) was verified after derivatization with dimethyl disulfide and subsequent GC/MS analysis. All identified MUFAME belong to the (n-9-)type. The UV-spectroscopical data show that approx. 1.2% conjugated acetylenic fatty acids occur in the oil. Furthermore IR- and NMR-spectroscopical investigations and the basic analyses of the seed were carried out.     

n-3 Polyunsaturated fatty acid-induced changes in the molecular species composition of diradylglycerol in murine peritoneal macrophages remain stable during incubationex vivo

Three- to four-week-old C57BL/6 mice were maintained for four weeks on diets in which the 10% lipid component was the ethyl esters of either corn oil or n-3 polyunsaturated fatty acids (n-3 PUFA). Incubation of macrophagesex vivo for 14 h, a normal incubation time used for macrophage studies, in the absence of fetal calf serum, did not diminish the extent of incorporation of 20∶5n-3 (eicosapentaenoic acid) and 22∶6n-3 (docosahexaenoic acid) moieties into membrane phospholipids and into diradylglycerol (DG) relative to that after a very abbreviated incubation time. We conclude that studies examining the effects of dietary n-3 PUFA on DG formation and related physiological effects in macrophages can be performed after a normalex vivo incubation of at least 14 h without experiencing a significant loss of incorporated n-3 PUFA.