Selectivity of potato tuber lipid acyl hydrolase toward long-chain unsaturated FA in esterification reactions with glycerol analogs in organic media

FA selectivity of a Celite-immobilized potato lipid acyl hydrolase (LAH) in esterification reactions with long-chain FA, including stearic acid (18∶0), oleic acid (18∶1), linoleic acid (18∶2), α-linolenic acid (18∶3), EPA (20∶5), and DHA (22∶6), and alcohol co-substrates (n-propanol, isopropanol, 1,3-propanediol, and glycerol) was studied in isooctane. Immobilized LAH was selective for FA of greater degrees of unsaturation (18∶3>18∶2>18∶1>18∶0) for all alcohol acceptors evaluated. Selectivity of LAH toward unsaturated C₁₈ FA increased with an increase in water activity (a _w ) from 0.19 to 0.90 for n-propanol, isopropanol, and 1,3-propanediol as alcohol co-substrates. In contrast, with glycerol as the alcohol cosubstrate, selectivity of LAH toward these unsaturated C₁₈ FA increased with a decrease in a _w from 0.90 to 0.19. In addition, immobilized LAH strongly discriminated against EPA and DHA for both 1,3-propanediol and glycerol as alcohol co-substrates.     

Plackett-burman design for determining the preference of Rhizomucor miehei lipase for FA in acidolysis reactions with coconut oil

The preference of lipase (EC 3.1.1.3) from Rhizomucor miehei in the incorporation of 11 FA, ranging from C10∶0 to C22∶6, into coconut oil TAG during acidolysis was studied by applying the Plackett-Burman experimental design. Enzymatic acidolysis reactions were carried out in hexane at 37°C for 48 h with coconut oil (0.1 M) and a mixture of 11 FA at a TAG to FA molar ratio of 1∶1. Lipase was used at the 5 wt% level. The incorporation of FA into coconut oil TAG was determined by GC. The lipase showed preference for long-chain saturated FA for incorporation into coconut oil TAG. The FA with 18 carbon atoms showed a high incorporation rate (18∶1>18∶1>18∶3). The lipase showed the least preference for the incorporation of 12∶0, which occurs in maximal concentration (46%), whereas the most preferred FA, 18∶0, occurs at a very low concentration (<2%) in coconut oil. The overall preference of lipase for the incorporation of different FA into coconut oil TAG was 18∶0>18∶2, 22∶0>18∶1, 18∶3, 14∶0, 20∶4, 22∶6>16∶0>12∶0≫10∶0.     

Growth inhibitory effects of liposome-associated 1-O-octadecyl-2-O-methyl-sn-glycero-3-phosphocholine

The growth inhibitory effects of 1-O-octadecyl-2-O-methyl-sn-glycero-3-phosphocholine (ET-18-OCH₃) and various liposome compositions of ET-18-OCH₃ were compared in a standardized growth inhibition assay utilizing a diverse tumor cell line panel including cell lines expressing multidrug resistance. ET-18-OCH₃ and ELL-12 (4∶3∶1∶2, dioleoylphosphatidylcholine/cholesterol/dioleoylphosphatidylethanolamine-glutaric acid/ET-18-OCH₃), an optimal liposomal ET-18-OCH₃ formulation, inhibited growth in the micromolar range in drug-sensitive and-resistant cells. In general, ET-18-OCH₃-liposomes were about twofold less growth inhibitory than ET-18-OCH₃. However, the known hemolytic effects of ET-18-OCH₃ were greatly reduced, up to 20 or more times, by liposome association. The effects of ET-18-OCH₃ and ELL-12 were compared in intracellular [Ca²⁺] modulation and DNA fragmentation assays. ET-18-OCH₃ elicited both concentration- and serum-dependent transient and permanent increases in intracellular [Ca²⁺]. In contrast, ELL-12 did not modulate intracellular [Ca²⁺]. ET-18-OCH₃ and ELL-12 similarly affected DNA fragmentation, which may be indicative of apoptosis. The results suggest that, although the specific growth inhibitory effects of ET-18-OCH₃ and ELL-12 are similar, associating ET-18-OCH₃ with stable well-characterized liposomes eliminates nonspecific cell membrane-associated lytic effects.     

Low-palmitic,low-linolenic soybean development

Plant breeding research efforts are currently focused on developing breeding procedures to decrease the saturated FA palmitic acid (16∶0) and the PUFA linolenic acid (18∶3) in U.S. soybean cultivars. Soybean oil with lower 16∶0 may provide cardiovascular benefits to health-conscious consumers, and lower 18∶3 could contribute to better flavor and stability of the oil. The purpose of this study was to determine genetic parameters that indicate the potential for breeding success and to characterize the correlated effect of the incorporation of the modified oil traits on the agronomic and seed quality traits of a soybean breeding population formed from a cross between the soybean cultivar Anand (normal) and germplasm N97-3708-13 (low 16∶0, low 18∶3). Although lines with only one modified oil quality trait (low 16∶0 or 18∶3) are useful as parents, commercial utilization requires productive cultivars with the combination of both oil traits. This paper shows the ease with which they may be combined with seed yield and other traits. Measurements were obtained from 179 F₂ single plants grown in 1999 and 121 F_2∶4 lines grown in replicated plots in 2000. Modified FA lines were developed with ca. 4% 16∶0 and 18∶3, respectively. Very weak positive correlations were found between 16∶0 concentration and seed yield (r=0.12) and between 16∶0 and seed oil concentration (r=0.13). No correlation was found between 18∶3 levels and seed yield, or between 18∶3 levels and seed oil concentration. These results indicate that breeding for reduced 16∶0 and 18∶3 should not have a negative impact on seed yield or oil concentration. 16∶0 and 18∶3 had moderately high heritabilities of 0.65 and 0.73, respectively. This indicates that breeders using low 16∶0, low 18∶3 germplasm in crosses with normal, elite lines can expect to recover low 16∶0 and low 18∶3 in pure line progenies via selection and generation advancement of F₂ individuals that express low levels of these FA.     

Natural abundance stable carbon isotope evidence for the routing and <Emphasis Type="Italic">de novo</Emphasis> synthesis of bone FA and cholesterol

This research reported in this paper investigated the relationship between diet and bone FA and cholesterol in rats raised on a variety of isotopically controlled diets comprising 20% C₃ or C₄ protein (casein) and C₃ and/or C₄ nonprotein or energy (sucrose, starch, and oil) macronutrients. Compoundspecific stable carbon isotope analysis (δ¹³C) was performed on the FA (16∶0, 18∶0, 18∶1, and 18∶2) and cholesterol isolated from the diet (n=4) and bone (n=8) of these animals. The dietary signals reflected by the bone lipids were investigated using linear regression analysis. δ¹³C values of bone cholesterol and stearic (18∶0) acid were shown to reflect whole-diet δ¹³C values. whereas the δ¹³C values of bone palmitic (16∶0), oleic (18∶1), and linoleic (18∶2) acids reflected dietary FA δ¹³C values. Dietary signal differences are a result of the balance between direct incorporation (or routing) and de novo synthesis of each of these bone lipids. Estimates of the degree of routing of these bone lipids gleaned from correlations between Δ¹³C _dlipid-wdiet (δ¹³C_{diet lipid}-δ¹³C_{whole diet}) spacings and Δ¹³C _blipid-wdiet (δ¹³C_{bone lipid}-δ¹³C_{whole diet} fractionations demonstrated that the extent of routing, where 18∶2>16∶0>18∶1>18∶0>cholesterol, reflected the relative abundances of these lipids in the diet. These findings provide the basis for more accurate insights into diet when the δ¹³C analysis of bone fatty FA or cholesterol is employed.     

Water-retaining ability of diacylglycerol

The water-in-oil emulsification characteristics and the adsorption properties of DAG at the oil/water interface were investigated for DAG having different FA compositions. The water-retaining ability of DAG is dependent on the FA composition but is not dependent on the interfacial tension at the oil/water interface in a simple way. The water-retaining ability is very different between uni-chain DAG (two FA have the same chain length) and complex-chain DAG (one FA is oleic acid and the other has a shorter alkyl chain). Uni-chain DAG, having long FA chains (R=C₁₂ or C_18∶1) have the ability to emulsify water at the volume fraction of 80% (ϕ80%), but uni-chain DAG with short or medium chain-length FA (R=C₃, C₄, C₆, C₈) show little ability to retain water. For complex-chain DAG, all the DAG studied here (R₁=C_18∶1, R₂=C₂−C₁₂) have the ability to emulsify water at ϕ80%. The stability of the emulsions, however, varies with the chain length of the R₂ FA (R₂ stability order: C₂, C₃>C_18∶1, C₁₀>C₈>C₄, C₆). The relationship between the water-retaining ability and the molecular structure of DAG is discussed from the viewpoint of intra- and intermolecular interactions between the FA chains.     

Reaction selectivity of <Emphasis Type="Italic">rhizomucor miehei</Emphasis> lipase as influenced by monoacylation of <Emphasis Type="Italic">sn</Emphasis>-glycerol

Reaction selectivities were determined in multicompetitive reactions mediated by Rhizomucor miehei (RM) lipase at water activity of 0.19 in hexane. Saturated FA (C4–C18 even chain) and oleic acid (C18∶1) were reacted with a single alcohol, glycerol, or α-or β-MAG containing C4, C10, C16, or C18∶1 individually as alcohol cosubstrate. Similar patterns of broad FA selectivity toward C8–C18 FA were generally observed for esterification into specific acylglycerol (AG) pools with the different α/β-CX-MAG cosubstrates. Exceptions were enrichment of C18 in the MAG pool with α-C16-MAG substrate, and a general suppression of C4/C6 FA reactivity and a specific discrimination toward >C8 FA incorporation into the TAG pool, both for reactions with α-C10- and α-C16-MAG. RM lipase selectivity toward MAG was in descending order: β-C18∶1-MAG>α/β-C4-MAG∼β-C10-MAG∼β-C16-MAG>α-C18∶1-MAG >α-C10-MAG∼α-C16-MAG. Selectivity in channeling CX of the original CX-MAG substrates into higher AG species was in descending order: α-C10-MAG∼α-C16-MAG>β-C10-MAGβ-C16-MAG>α-C18∶1-MAG>β-C18∶1-MAG∼ α/β-C4-MAG. Aside from their characteristic FA selectivity, Burkholderia cepacia (PS-30) and RM lipases behaved similarly in terms of MAG selectivity as well as a general conservation of FA selectivity throughout the sequential steps of TAG assembly from FA and glycerol for processes designed to yield specifically structured TAG.     

Fatty acids and oil content in white lupin seed as affected by production practices

Characteristics of the seed oil of white lupin (Lupinus albus L.), a potential alternative winter crop in the mid-Atlantic region of the United States, are not well established. Replicated experiments were conducted during the 1998–1999 and 1999–2000 growing seasons with a determinate and an indeterminate cultivar to characterize oil and FA in lupin seed in relation to production practices. The experiments were planted in early October, late October, and mid-November using row spacings of 0.3, 0.6, and 0.9 m at each planting time. Seeds from the planting date of early October had significantly (P<0.05) higher oil content than the later plantings (late October and mid-November). A closer row spacing (0.3 m) also had significantly (P<0.05) higher oil content than the wider row spacing (0.9 m). Planting data effects on FA content were significant for some FA, but row spacing did not affect FA contents. Oil content in the seed varied from 7.2 to 8.2% (w/w). The oil from white lupin seed contained FA in the order of 18∶1>18∶2> 18∶3>16∶0>20∶1>22∶1>22∶0>18∶0>24∶0>20∶0. The saturated FA/unsaturated FA ratio in lupin oil was 0.14. White lupin seed contained higher contents of oil and FA than literature values for seed of navy, kidney, and pinto beans.     

Effect of temperature on the incorporation into phospholipid classes and metabolismvia desaturation and elongation of n−3 and n−6 polyunsaturated fatty acids in fish cells in culture

The incorporation of 18∶2n−6, 18∶3n−3, 20∶4n−6 and 20∶5n−3 was greater at 10°C than at 22°C in Atlantic salmon (AS), rainbow trout (RTG-2) and turbot (TF) cells. However, there were generally no significant differences between the amount of incorporation of all four polyunsaturated fatty acids (PUFA) into total lipid within a cell type at either 22°C or 10°C. The distributions of the PUFA between individual phospholipid classes at 22°C was essentially the same in AS and TF cells—with the C₁₈ PUFA the order of incorporation in these cells was phosphatidylcholine (PC) > phosphatidylethanolamine (PE) > phosphatidic acid/cardiolipin (PA/CL); with 20∶4n−6 the order was PE and phosphatidylinositol (PI)>PC; with 20∶5n−3, PE>PC. In RTG-2 cells at 22°C the distributions of the C₁₈ PUFA were similar to the other cell lines, but with 20∶4n−6 the order was PC>PI>PE, and with 20∶5n−3 it was PC>PE. At 10°C the incorporation of C₁₈ PUFA into PC increased and into PE and PA/CL decreased, in general, in all cell lines. Incorporation of 20∶5n−3 into PC and PE was increased and decreased at 10°C, respectively, in AS and TF cells, whereas in RTG-2 cells the changes at 10°C were opposite i.e., increased in PE and decreased in PC. With 20∶4n−6, incorporation into PC at 10°C was increased in all cell lines with decreased incorporation into PI in AS and RTG-2 cells and into PE in AS and TF cells, whereas incorporation of 20∶4n−6 into PE increased in RTG-2 cells. The metabolismvia desaturation and elongation of the n−3 PUFA was greater than that of the equivalent n−6 PUFA in all cell lines, irrespective of temperature. There was less conversion of the C₁₈ PUFA at 10°C than at 22°C in RTG-2 and TF cells, but the conversion of 18∶3n−3 by AS cells was increased at 10°C. Temperature had no effect on the conversion of the C₂₀ PUFA.     

Triglyceride selectivity of immobilized <Emphasis Type="Italic">Thermomyces lanuginosa</Emphasis> lipase in interesterification

The triglyceride (fatty acid) selectivity of an immobilized lipase from Thermomyces lanuginosa (Lipozyme TL IM) was investigated in lipase-catalyzed interesterification reactions between two nono-acid TG in n-hexane. Tristearin (tri-C18∶0) was used as a reference in a series of TG with saturated FA from tri-C4∶0 to tri-C20∶0, except for tri-C6∶0, and in a series of unsaturated FA from tri-C18∶1 to tri-C18∶3. The quantification was performed by HPLC, and different methods of selectivity evaluation were used. None of the methods used showed any significant differences between the performances of the lipase on the different TG, indicating that Lipozyme TL IM is nonselective toward FA or TG in the system used. A response surface design was used to investigate the influence of water activities (a _w ) and reaction temperatures on the reactivity of Lipozyme TL IM with a system of tripalmitin (tri-C16∶0) and trilaurin (tri-C12∶0) in n-hexane. An increase in temperature (40 to 60°C) was found to affect the reactivity of the lipase significantly. The reactivity of Lipozyme TL IM was unaffected by the change in a _w from 0.1130 to 0.5289. An increase in a _w only led to an increase in FFA formation.     

On the specificity of a phospholipase A2 purified from the 106,000×g pellet of bovine brain

Assessment has been made of the specificity of a purified phospholipase A₂ from the 106,000×g pellet (microsomal fraction) of bovine grey matter which shows strong activity toward phosphatidylinositol (PI). In the first series of experiments involving the utilization as substrates of PI with different¹⁴C- or³H-labeled fatty acids in the 2-position, the purified phospholipase A₂ most readily removed 16∶0 palmitic acid, followed by 18∶0 stearic acid, 18∶1 oleic acid and 20∶4 arachidonic acid. In the second series of experiments, the purified phospholipase A₂ showed preferential action toward PI (100%) compared to phosphatidylcholine (PC, 62.5%), phosphatidic acid (PA, 32.6%), phosphatidylethanolamine (PE, 25.1%) and phosphatidylserine (PS, 21.5%), where each phosphoglyceride was labeled in the 2-position with [1-¹⁴C] oleic acid. In the third series of experiments, fatty acids were shown to cause inhibition of action of the purified phospholipase A₂ on 1-acyl, 2-[1-¹⁴C] oleoyl PI in the order 20∶4>18∶1>18∶0>16∶0 which is the reverse order to that just noted. In the final series of experiments, the addition of the phosphoglycerides PC, PE, PS and PA in amounts of 5 or 10 μM caused either no inhibition (PE, 2%), slight inhibition (PC, 15%) or reasonably significant inhibition (PA, 20% and PS, 40%) of action of the purified phospholipase A₂ on 1-acyl, 2-[1-¹⁴C]-oleoyl PI. The pattern of specificity observed for the purified phospholipase A₂ combined with its microsomal location are the expected properties of a phospholipase A₂ that might function in a deacylation-reacylation cycle for modifying the fatty acid distribution in PI.     

Effect of dietary conjugated linoleic acid (CLA) on metabolism of isotope-labeled oleic,linoleic, and CLA isomers in women

The purpose of this study was to investigate the effect of dietary CLA on accretion of 9c-18∶1, 9c, 12c-18∶2, 10t, 12c-18∶2, and 9c, 11t-18∶2 and conversion of these FA to their desaturated, elongated, and chain-shortened metabolites. The subjects were six healthy adult women who had consumed normal diets supplemented with 6 g/d of sunflower oil or 3.9 g/d of CLA for 63 d. A mixture of 10t, 12c-18∶2-d ₄, 9c, 11t-18∶2-d ₆, 9c-18∶1-d ₈, and 9c, 12c-18∶2-d ₂, as their ethyl esters, was fed to each subject, and nine blood samples were drawn over a 48-h period. The results show that dietary CLA supplementation had no effect on the metabolism of the deuterium-labeled FA. These metabolic results were consistent with the general lack of a CLA diet effect on a variety of physiological responses previously reported for these women. The ²H-CLA isomers were metabolically different. The relative percent differences between the accumulation of 9c, 11t-18∶2-d ₆ and 10t, 12c-18∶2-d ₄ in plasma lipid classes ranged from 9 to 73%. The largest differences were a fourfold higher incorporation of 10t, 12c-18∶2-d ₄ than 9c, 11t-18∶2-d ₆ in 1-acyl PC and a two- to threefold higher incorporation of 9c, 11t-18∶2-d ₆ than 10t, 12c-18∶2-d ₄ in cholesterol esters. Compared to 9c-18∶1-d ₈ and 9c, 12c-18∶2-d ₂, the 10t, 12c-18∶2-d ₄ and 9c, 11t-18∶2-d ₆ isomers were 20–25% less well absorbed. Relative to 9c-18∶1, incorporation of the CLA isomers into 2-acyl PC and cholesterol ester was 39–84% lower and incorporation of 10t, 12c-18∶2 was 50% higher in 1-acyl PC. This pattern of selective incorporation and discrimination is similar to the pattern generally observed for trans and cis 18∶1 positional isomers. Elongated and desaturated CLA metabolites were detected. The concentration of 6c, 10t, 12c-18∶3-d ₄ in plasma TG was equal to 6.8% of the 10t, 12c-18∶2-d ₄ present, and TG was the only lipid fraction that contained a CLA metabolite present at concentrations sufficient for reliable quantification. In conclusion, no effect of dietary CLA was observed, absorption of CLA was less than that of 9c-18∶1, CLA positional isomers were metabolically different, and conversion of CLA isomers to desaturated and elongated metabolites was low.     

In vitro comparison of hepatic metabolism of 9cis-11 trans and 10trans-12cis isomers of CLA in the rat

Hepatic metabolism of the two main isomers of CLA (9cis-11 trans, 10trans-12cisC_18∶2) was compared to that of oleic acid (representative of the main plasma FA) in 16 rats by using the in vitro method of incubated liver slices. Liver tissue samples were incubated at 37°C for 17h under an atmosphere of 95% O₂/5%CO₂ in a medium supplemented with 0.75 mM of FA mixture (representative of circulating nonesterified FA) and with 55 μM [1-¹⁴C]9cis-11 trans C_18∶2, [1-¹⁴C]10trans-12cis C_18∶2, or [1-¹⁴C]oleate. The uptake of CLA by hepatocytes was similar for both isomers (9%) and was three times higher (P<0.01) than for oleate (2.6%). The rate of CLA isomer oxidation was two times higher (49 and 40% of incorporated amounts of 9cis-11 trans and 10trans-12 cis, respectively) than that of oleate (P<0.01). Total oxidation of oleate and CLA isomers into [¹⁴CO₂] was low (2 to 7% of total oxidized FA) compared to the partial oxidation (93 to 98%) leading to the production of [¹⁴C] acid-soluble products. CLA isoemrs escaping from catabolism were both highly desaturated (26.7 and 26.8%) into conjugated 18∶3. Oleate and CLA isomers were mainly esterified into neutral lipids (30%). They were slowly secreted as parts of VLDL particles (<0.4% of FA incorporated into cells), the extent of secretion of oleate and of 10trans-12 cis being 2.2-fold higher than that of 9cis-11 trans (P<0.02). In conclusion, this study clearly showed that both CLA isomers were highly catabolized by hepatocytes, reducing their availability for peripheral tissues. Moreover, more than 25% of CLA escaping from catabolism was converted into conjugated 18∶3, the biological properties of which remain to be elucidated.     

FA selectivity of lipases in acyl-transfer reactions with acetate esters of polyols in organic media

FA reaction selectivity of Burkholderia cepacia, Rhizomucor miehei, and Candida antarctica fraction B lipases was compared between acyl-transfer and esterification reactions. Multicompetitive reaction mixtures containing a series of n-chain FA (a C₄–C₁₈ series; and a C_18∶x series, where X=0-3 double bonds) and a single acetate ester co-substrate [triacetin, 1,2-propanediol (1,2-PD)diacetate, and 1,3-PD diacetate] were studied in tert-butyl methyl ether at an a _w of 0.69. For B. cepacia lipase, FA optima for C₈, C₁₆, and C_18∶2 were observed in all reactions with 1.0- to 5.9-fold differences in FA selectivity. For R. miehei lipase, an optimum for C₈ FA was observed in all reactions with 1.2- to 6.7-fold differences in FA selectivity. For C. antarctica lipase, FA optima for C₈/C₁₀ were observed in all reactions with 1.0- to 2.8-fold differences in FA selectivity. FA selectivities were broadly modulated upon changing from free polyol to acetate ester co-substrates for B. cepacia and R miehei lipases, whereas FA selectivity modulations were more specific upon this change in reaction configuration for C. antarctica B lipase. For all lipases, reactivity toward unsaturated C_18∶x FA was enhanced in acyl-transfer relative to esterification reactions with these polyol co-substrates.     

Unusual minor nonmethylene-interrupted di-, tri-, and tetraenoic fatty acids in limpet gonads

Unusual minor nonmethylene-interrupted (NMI) FA have been identified in the lipids of gonads from the limpets Cellana grata and Collisella dorsuosa by using GC-MS of the combination of their 4,4-dimethyloxazoline derivatives and picolinyl esters. Among 23 NMI unsaturated FA from C₁₈ to C₂₂ and C₂₄ identified in this study, 5,11-nonadecadienoic (5,11-19∶2), 7,16-heneicosadienoic (7,16–21∶2), 9,15-tetracosadienoic (9,15–24∶2), 5,9,15-docosatrienoic (5,9,15–22∶3), and 5,9,15-tetracosatrienoic (5,9,15–24∶3) acids may not have been reported previously from living organisms. The presence of 5,11,14,17-eicosatetraenonoic (5,11,14,17–20∶4) and 7,13,16,19-docosatetraenenoic (7,13,16,19–22∶4) acids as FA components in marine mollusks may be reported here for the first time. In this study, the male and female gonads of both species showed distinct differences in both their composition and proportions of NMI FA. Most NMI FA identified were mainly present in the female gonads of both species, especially in TAG that contained 21 NMI FA.     

Selective hydrogenation with tris(triphenylphosphine) chlororhodium (I) catalyst: Preparation of octadecenoate isomers

Fractionation of products obtained from partial catalytic hydrogenation of methylcis-9,cis-12-octadecadienoate (9c,12c-18:2) with tris(triphenylphosphine) chlororhodium [RhCl(Ph₃P)₃] provided a facile method for preparation of a nearly equal molar mixture of methylcis-9- andcis-12-octadecenoate (9c-18∶1 and 12c-18∶1). Isolation of products was achieved by silver resin and C18 reverse phase liquid chromatography. Catalytic deuteration of 9c,12c-18∶2 yields a mixture of 9c-18∶1-12,13-d₂ and 12c-18∶1-9,10-d₂ with an isotopic purity of 85%. Final isolated yield of the mixture of 9c- and 12c-18∶1 products was 30%. Isolation of products from partial hydrogenation of conjugated octadecadienoates (9c,11t-18∶2 or 10t,12c-18∶2) provided a convenient method for synthesis of an almost equal molar mixture of methyltrans-10 andtrans-11-octadecenoate (10t-18∶1 and 11t-18∶1). Characterization of the reaction products from hydrogenation of 9c,12c-28∶2 indicates that the 9c- and 12c-18∶1 products are formed by the expected 1,2-hydride addition. The presence of small amounts of 10t- and 11t-18∶1 and conjugated octadecadienoates was evidence for a secondary isomerization-1,4-hydride addition pathway. Isolation and characterization of products from RhCl(Ph₃P)₃-catalyzed hydrogenation of 9c,11t-18∶2 and 10t,12c-18∶2 indicate that both 1,2- and 1,4-hydride addition to the conjugated diene isomers occurs at about equal rates, but only thecis bond is reduced by the 1,2-hydride addition pathway and the 1,4-hydride addition pathway yields only atrans-18∶1. Because of this unusual selectivity for acis bond conjugated with atrans bond, hydrogenation of both 9c,11t-18∶2 and 10t,12c-18∶2 yields the same mixture of t-18∶1 isomers.     

Temperature-dependent deacylation of molecular species of phosphatidylcholine by microsomal phospholipase A2 of thermally acclimated rainbow trout,Salmo gairdneri 1

Using the ratios of kinetic parameters, V/Km, the deacylation of different molecular species of 1-palmitoyl,2-acyl phosphatidylcholine via microsomal phospholipase A₂ (PLA₂) was studied in liver tissue of thermally acclimated rainbow trout (Salmo gairdneri). In general, PLA₂ from fish acclimated to cold temperatures showed an order of preference for the acyl moieties of 18∶1>18∶1>18∶2>18∶0. Trout acclimated to warm temperatures generally preferred 18∶0 PC, but the actual order of preference depended on the temperature of the assays and the presence of endogenous lipids in the enzyme preparation. At 5 C, the particulate (microsomal) enzyme preferred 18∶0>18∶2>18∶1, but a lipid-free preparation of the enzyme preferred 18∶2>18∶0>18∶1. At 20 C, particulate enzyme preferred 18∶1>18∶0>18∶2 but purified enzyme preferred 18∶0>18∶2>18∶1. Thus, assay temperature and the presence of microsomal lipids had a greater effect on PLA₂ from fish acclimated to warm temperatures than fish acclimated to cold temperatures. The substrate preference of PLA₂ is discussed with reference to the previously observed changes in membrane fatty acid composition that occur with thermal acclimation in rainbow trout.     

Fatty acids bound to <Emphasis Type="Italic">Fasciola hepatica</Emphasis> 12 kDa fatty acid-binding protein,a candidate vaccine,differ from fatty acids in extracts of adult flukes

The FA composition of Fasciola hepatica 12 kDA purified native FA-binding protein (nFh12), a candidate vaccine against fascioliasis, is described. The FA chain lengths ranged between 12 and 24 carbons. The principal FA were 16∶0 18∶1n−9, 18∶0, 20∶4n−6, and 20∶1n−9. The acids 16∶0, 18∶1n−9, and 18∶0 comprised over half the FA that were bound to the whole FA-binding protein. Small amounts (1.0–2.8%) of isoanteiso methyl-branched FA also were characterized. Forty-one different FA were identified in extracts of the adult flukes, with the three most abundant FA also being 16∶0, 18∶1n−9, and 18∶0. A similar proportion of saturated vs. unsaturated FA was observed between the whole extract from F. hepatica and the nFh12 protein. However, the n−3/n−6 ratio of PUFA was significantly different, being 1.2 in the whole extract vs. 9.6 in the nFh12 protein complex. The nFh12 protein binds more n−5, n−6, and n−7 PUFA and less n−3 and n−9 PUFA than the whole extract. In addition, cholesterol (56%), sitosterol (36%), and fucosterol (8%) also were bound to the nFh12 protein complex.     

Reaction selectivity of <Emphasis Type="Italic">Burkholderia cepacia</Emphasis> (PS-30) lipase as influenced by monoacylation of <Emphasis Type="Italic">sn</Emphasis>-glycerol

Reaction selectivities were determined in multicompetitive reactions mediated by Burkholderia cepacia lipase (Amano PS-30) at a water activity of 0.19 in hexane. Saturated FA (C4–C18 even chain) and oleic acid (C18∶1) were reacted with a single alcohol, glycerol, α-or β-MAG, containing C4, C10, C16, or C18∶1 individually as alcohol cosubstrate. Similar odrinal patterns of FA selectivity, with C8, C10, and C16 preferred over others, were generally observed for incorporation of FA into specific acylglycerol (AG) pools of the 24 specific cases evaluated. The three exceptions were enrichment of C14 and C18 in the MAG pool with α-C16-MAG, substrate, and a general suppression of >C8 incorporation into the TAG pool for reactions with α-C10- and α-C16-MAG. PS-30 lipase selectivity toward MAG was in descending order: α/β-C4-MAG>β-C10-MAG>β-C16-MAG>α/β-C18∶1-MAG>α-C10-MAG>α-C16-MAG. Selectivity in channeling CX of the original CX-MAG substrates into higher AG species was in descending order: α-C10-MAG∼α-C16-MAG>α-C18∶1-MAG>β-C10-MAG∼β-C16-MAG∼β-C18∶1-MAG >α/β-C4-MAG. Generally, MAG were better acyl donors than FA for esterification reactions leading to DAG formation. These observations are relevant to the design of biocatalytic processes intended to yield specifically structured TAG.     

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.