Composition and activity of commercial triacylglycerol acylhydrolase preparations

Commercial lipase preparations were surveyed to determine gross composition, amounts of nonprotein impurities, and esterolytic and lipolytic activities. Most of the 34 commercial lipase preparations contained more than 80% non-proteinaceous material, with salt and carbohydrate being the most abundant materials. The tributyrin hydrolase activity of these commercial lipase preparations was determined and expressed as lipase/esterase forestomach units (LFU). Tributyrin hydrolase activity ranged from negligible (5.3 LFU/g) to very high (>1000,000 LFU/g). Aspergillus and Penicillium preparations were low in tributyrin hydrolase activity. Candida rugosa preparations were intermediate in activity. Preparations of porcine pancreas, Rhizomucor, Pseudomonas, and Rhizopus lipases exhibited a broad range of levels of activity. No relation between protein content and tributyrin hydrolase activity was observed. Isoelectric focusing of the proteins present in the preparations demonstrated the presence of between 2 and 27 isophoretically discrete bands in the isoelectric range of 3 to 9. Although there were many similarities of distribution of protein isoelectric points within genera and spcies, the preparations generally displayed unique patterns of isophoretically discrete protein bands. Lipase zymography demonstrated the presence of 0 to 7 isophoretically discrete lipase activities in each preparation, spanning the entire range of isoelectric points surveyed.     

Molecular weight distribution and regioisomeric structure of triacylglycerols in some common human milk substitutes

Triacylglycerol (TAG) molecular weight distribution and regioisomeric structure of selected molecular weight species in human milk and in 32 human milk substitutes was determined. Negative ion chemical ionization mass spectrometry was used to determine the molecular weight distribution and collisionally induced dissociation tandem mass spectrometry applied to identify the sn-2 and sn-1/3 positions of fatty acids in TAG. The main molecular weight species of human milk TAG in decreasing order of abundance were 52∶2, 52∶3, 52∶1, 54∶3, 50∶2, 50∶1, 54∶4, 48∶1, 54∶2, 48∶2, 46∶1, 52∶4, and 50∶3 (acyl carbon number/number of double bonds), constituting 83 mol% of total TAG molecular species. In human milk substitutes, the proportion of the corresponding molecular weight species varied from 33 to 87 mol%. The main TAG regioisomers within the molecular weight species 52∶2, 52∶3, and 50∶1 in human milk were 18∶1-16∶0-18∶1 (83 mol%), 18∶1-16∶0-18∶2 (83 mol%), and 18∶1-16∶0-16∶0 (80 mol%), respectively. In human milk substitutes, the corresponding proportions varied in a wide range of 0–82 mol%, 0–100 mol%, and 0–73 mol%, respectively. Although TAG structures in some human milk substitutes closely resembled those in human milk, the great variation among samples leads to the conclusion that it is still possible to improve the TAG composition in human milk substitutes by applying novel methods to synthesize structured TAG.     

Triacylglycerol composition of protected designation of origin cheeses during ripening. Authenticity of milk fat

Triacylglycerol (TAG) composition by carbon number in 2 protected designation of origin cheeses, Mahón (cheese from cow milk) and Manchego (cheese from ewe milk) that were manufactured by 3 different producers was analyzed during cheese ripening using gas chromatography with a short capillary column. The TAG composition at different times during cheese ripening was also analyzed in cheeses from different batches produced at the same plant. Lipolysis levels in the Mahón and Manchego cheeses during ripening were low; free fatty acid values ranged from 2,500 to 4,000 ppm at the end of ripening. The TAG composition did not change significantly during ripening. The TAG values obtained from each cheese sample were substituted into the multiple regression equations that have been proposed to detect foreign fats in milk fat. The values obtained using the equations for bovine (proposed by the European Union) and ovine milk (proposed by our laboratory) were within the normal range. Accordingly, these equations can be considered useful for detecting foreign fat in these cheeses during the ripening period contemplated during this study.     

Feeding increasing amounts of ruminally protected choline decreased fatty liver in nonlactating,pregnant Holstein cows in negative energy status

The objectives were to determine the optimal feeding amount of choline in a ruminally protected form to reduce the triacylglycerol (TAG) concentration in liver and to increase TAG in blood plasma of dairy cows. Pregnant, nonlactating multiparous Holstein cows (n = 77) were blocked by body condition score (3.59 ± 0.33) and assigned to treatment at 64 ± 10 d before calculated calving date. Dietary treatments were top-dressing of 0, 30, 60, 90, or 120 g/d of ruminally protected choline (RPC; Balchem Corp., New Hampton, NY) ions to supply the equivalent of 0, 6.5, 12.9, 19.4, and 25.8 g/d of choline ions. Diets were formulated to exceed nutrient requirements for maintenance and pregnancy and fed in ad libitum amounts for the first 5 d. From d 6 to 15, cows were restricted to consume approximately 31% of their net energy requirements to simulate early lactating cows in negative energy balance. Methionine intake was maintained throughout each 15-d period. Liver was biopsied at 5 and 14 d and analyzed for TAG and glycogen. Blood was sampled on d 5 and 14 and plasma analyzed for glucose, insulin, cholesterol, β-hydroxybutyrate, long-chain fatty acids, and haptoglobin. On d 14, a mixture of saturated long-chain fatty acids, ground corn, and dried molasses (50:37:13) was offered (908 g, as-is basis) 10 h after the single daily feeding. Blood samples were collected for 19 h and plasma analyzed for TAG and cholesterol to assess apparent absorption of dietary fat. Mean dry matter intake and energy balance decreased from means of 9.5 to 3.3 kg/d and from 0.6 to ?9.2 Mcal of net energy for lactation/d during the ad libitum and restricted feeding periods, respectively. Plasma concentrations of the lipid-soluble choline biomolecules, namely total phosphatidylcholines, total lysophosphatidylcholines, and sphingomyelin, increased with choline supplementation. Feed restriction increased plasma concentrations of β-hydroxybutyrate and free long-chain fatty acids, whereas those of glucose, insulin, and total cholesterol decreased. During feed restriction, concentration of hepatic TAG and plasma haptoglobin decreased linearly, whereas concentration of hepatic glycogen tended to increase quadratically with increasing intake of RPC. After fat supplementation, mean plasma concentration of TAG increased by an average of 21% with intake of RPC ions, peaking at intakes of ≥6.5 g/d of RPC ion. In summary, feeding RPC ions to cows in negative energy balance had increasing lipotropic effects on the liver when consumed up to 25.8 g/d, whereas feeding only 6.5 g/d increased concentrations of hepatic glycogen and TAG in the blood.     

Structural investigations of β′ triacylglycerols: An x-ray diffraction and microscopic study of twinned β′ crystals

To reveal the structure ofβ′ triacylglycerols in detail, LML (C₁₂C₁₄C₁₂) was purified by a zone-melting procedure, and twinned crystals ofβ′ stable LML were obtained from a melt,β′ LML crystallizes in the monoclinic space group C₂, with eight molecules in the unit cell. A powder X-ray diffraction study of solid compounds of 1:1 mixtures of selected triacylglycerols led to the conclusion that the triacylglycerol molecules in theβ modification have a 1,2 chair-conformation (i.e., the fatty acid chains on glycerol positions 1 and 2 are adjacent, with the chain on the 3-position forming the back rest of the chair). Packing studies and the positions of two-fold axes and two-fold screw axes in the unit cell require that the molecules are bent at the glycerol site. The fatty acid chains make an angle of 25° with the long axis of the unit cell. Electron micrographs and precession photographs indicate that the twinning results from the stacking of a large number of thin crystalline platelets in two distinct orientations.     

A trimer plus a dimer-gallate reproduce the bioactivity described for an extract of grape seed procyanidins

The relationship between grape seed-derived procyanidin extract components and their bioactivity was explored. The monomeric and dimeric structures only acted as anti-inflammatory agents. Similarly, pure C1 trimer was highly effective on LPS-activated macrophages. To reproduce all of the bioactivities of the total extract, a fraction enriched with trimeric structures was needed. This trimeric-enriched fraction was divided into subfractions, the most bioactive of which contained two compounds with a molecular weight equal to a trimer (865) and a dimer-gallate (729), according to spectrometric analysis. Thus, it may be concluded that a mixture of both molecules reproduces the bioactivity in glucose metabolism (3T3-L1), lipid metabolism (HepG2) and macrophage functionality (RAW 264.6).     

Effects of dietary flaxseed oil on cholesterol metabolism of hamsters

High-fat/cholesterol diets (HFCD) formulated by addition of butter (BU), coconut oil (CO), or flaxseed oil (FX) enhanced (P < 0.05) serum lipids of hamsters compared to the low-fat/cholesterol diet (Control). However, FX groups showed a hypocholesterolaemic effect compared to CO and BU groups. Lower (P < 0.05) hepatic triacylglycerol and cholesterol contents were measured in FX groups than those of CO and BU groups; whereas, higher (P < 0.05) faecal triacylglycerol and cholesterol contents were observed in FX groups. HMG-CoA reductase mRNA expression was upregulated (P < 0.05) by HFCD, whilst FX groups showed no (P > 0.05) influence on LDL-receptor mRNA expression compared to that of Control groups; however, higher (P < 0.05) than those of CO and BU groups. Meanwhile, there was a tendency towards higher CYP7A1 expression in the CO or FX group than the BU group. Thus, the hypocholesterolaemic effect of FX might result from increases of LDL-receptor mRNA expression, and cholesterol catabolism/output.