Naohiro Gotoh

Interaction among vitamin C, vitamin E, and beta-carotene.

The effects of vitamin C (ascorbic acid), vitamin E (alpha-tocopherol), and beta-carotene as antioxidants and their cooperative action against the oxidation of lipid in solution, membranes, and lipoproteins have been studied and reviewed. Ascorbic acid and alpha-tocopherol act as potent, and probably the most important, hydrophilic and lipophilic antioxidants, respectively. They function at their own site individually and furthermore act synergistically. beta-Carotene has lower reactivity toward radicals than does alpha-tocopherol and acts as a weak antioxidant in solution. It is more lipophilic than alpha-tocopherol and is assumed to be present at the interior of membranes or lipoproteins, which enables it to scavenge radicals within the lipophilic compartment more efficiently than does alpha-tocopherol. The cooperative interaction between vitamin C and vitamin E may be quite probable, that of vitamin C and beta-carotene is improbable, whereas that between vitamin E and beta-carotene may be possible.

Double-Blind Controlled Study on the Effects of Dietary Diacylglycerol on Postprandial Serum and Chylomicron Triacylglycerol Responses in Healthy Humans

Objective: The effects of dietary diacylglycerol (DG) on postprandial lipemia in healthy humans were investigated. Methods: Forty normolipidemic male volunteers ingested fat emulsions containing either DG oil or triacylglycerol (TG) oil, at different doses: 10 g (n = 13), 20 g (n = 10) and 44 g (n = 17). Two test emulsions were given at seven-days intervals in random order. Fatty acid compositions of the test oils had been adjusted to be equal. Fasting and postprandial serum lipid concentrations in each group and plasma lipoprotein lipids in the 20 g-fat ingestion group were measured during the postprandial intervals. Results: When DG emulsion was ingested, serum TG concentrations were significantly lower (p < 0.05) in the late postprandial phase, i.e., 4 hours, 6 hours as compared to the TG emulsion. The magnitude of postprandial lipemia (the area bounded by the curve above the fasting concentration) after ingestion of 44 g-DG emulsion was significantly less than that of 44 g-TG emulsion (6.54 ± 5.12 and 8.45 ± 7.54 mmol · h/L, mean ± SD, respectively). Chylomicron TG, cholesterol, and phospholipid concentrations at 4 hours after ingestion of DG emulsion were significantly lower (p < 0.05) than those after the ingestion of TG emulsion at the same time point. No marked differences were observed for VLDL, LDL and HDL lipids between the test emulsions. Conclusion: In the usual range of fat intake (10–44 g), postprandial response after ingestion of DG emulsion was significantly less than that after ingestion of TG emulsion in healthy human subjects.

Rates of interactions of superoxide with vitamin E, vitamin C and related compounds as measured by chemiluminescence

The rate constants for the interactions of superoxide with vitamin E (alpha-tocopherol), vitamin C (ascorbic acid) and their related compounds have been measured by a chemiluminescence method. A strong chemiluminescence of a constant intensity was observed when xanthine oxidase was added to an aqueous solution of hypoxanthine and a Cypridina luciferin analog, 2-methyl-6-phenyl-3-7-dihydroimidazo[1,2-a]pyrazin-3-one (CLA). Vitamin E, vitamin C and their related compounds competed with CLA to react with superoxide and reduced the chemiluminescence intensity. From a kinetic analysis of the effect of addition of these compounds on the chemiluminescence intensity, the rate constants for their interactions with superoxide were measured at 25 degrees C and pH 7.8. The rate constants were obtained as 3.3 x 10(5) and 1.7 x 10(4) M-1 s-1 for ascorbate and 2-carboxy-2,5,7,8-tetramethyl-6-chromanol, respectively, and also as 4.9 x 10(3) and 4.5 x 10(3) M-1 s-1 for alpha-tocopherol incorporated into soybean and dimyristoyl phosphatidylcholine liposomal membranes, respectively. It has been shown that this method is a sensitive and a quick method which can be applied for measurement of the reactivities of various natural and synthetic compounds toward superoxide. In addition it has been shown that this method can also be applied to the heterogeneous system as well as homogeneous solution, which makes it more versatile and useful for the study in biochemistry.

Inhibition of Oxidation of Low Density Lipoprotein by Vitamin E and Related Compounds

The oxidation of low density lipoprotein (LDL) was carried out aiming specifically at elucidating the anti-oxidant action of alpha-tocopherol. Lipophilic and hydrophilic azo compounds and copper induced the oxidation of LDL similarly to give cholesterol ester and phosphatidylcholine hydroperoxides as major products. The antioxidant potency of alpha-tocopherol in LDL was much poorer than in homogeneous solution. Doxyl stearic acids were used as spin probe and incorporated in LDL. The rate of reduction of doxyl nitroxide in LDL by ascorbate decreased with increasing distance from the LDL surface. From the competition between the spin probe and alpha-tocopherol in scavenging radical, it was found that the efficacy of radical scavenging by alpha-tocopherol became smaller as the radical went deeper into the interior of LDL. On the other hand, 2,2,5,7,8-pentamethyl-6-chromal spared the spin label regardless of the position of nitroxide. The antioxidant activity of chromanols against LDL oxidation increased with decreasing length of isoprenoid side chain at the 2-position. All these results were interpreted by location and low mobility of alpha-tocopherol in LDL. The tocopherol mediated propagation was observed notably at low rate of radical flux, but this was suppressed by reductant such as ascorbic acid and ubiquinol.

Effects of three different highly purified n-3 series highly unsaturated fatty acids on lipid metabolism in C57BL/KsJ-db/db mice.

Triglycerides (TG) consisting of highly purified (>97%) n-3 series highly unsaturated fatty acids, eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA), and docosahexaenoic acid (DHA), were administered to C57BL/KsJ-db/db mice for 4 weeks by pair-feeding to compare their effects on lipid metabolism and to evaluate the effects of DPA on lipid metabolism. The hepatic TG level and total amount was decreased by treatment with DHA and DPA compared to the control. The efficacy of DPA was greater than that of EPA, but less than that of DHA. In contrast, EPA had the greatest serum TG reducing effect. The hepatic cytosol fraction of the DHA-treated group contained the lowest fatty acid synthase (FAS) and malic enzyme (ME) activity levels. Furthermore, the DHA-treated group contained the highest serum adiponectin concentrations. These findings indicate that the strong hepatic TG-lowering effect of DHA is due to the suppression of TG synthesis. The same tendencies were observed in DPA-treated mice, and the effect was stronger than that observed in EPA-treated mice, but equivalent to that observed in DHA-treated mice. Based on these results, DPA possesses lipid metabolism-improving effects. The beneficial effects of DPA for lipid metabolism were not superior to those of EPA and DHA, and the effect was always intermediate between those of EPA and DHA.

Enantiomeric separation of asymmetric triacylglycerol by recycle high-performance liquid chromatography with chiral column

In our previous studies, we employed recycle HPLC for the separation of triacylglycerol (TAG)-positional isomers (PIs). In this study, a recycle HPLC system equipped with a polysaccharide-based chiral column was applied to the enantiomeric separation of some asymmetric TAGs having straight-chain C16-C18 acyl residues. As a result, 1,2-dipalmitoyl-3-oleoyl-rac-glycerol (rac-PPO), 1,2-dioleoyl-3-palmitoyl-rac-glycerol (rac-OOP), and 1,2-dipalmitoyl-3-linoleoyl-rac-glycerol (rac-PPL) were resolved into their respective enantiomers. However, neither 1,2-dioleoyl-3-linoleoyl-rac-glycerol (rac-OOL), consisting of only unsaturated fatty acids, nor 1,2-dipalmitoyl-3-stearoyl-rac-glycerol (rac-PPS), consisting of only saturated fatty acids, was resolved. These results suggest that the asymmetric TAGs, used in this study, having both a palmitic acid moiety and an oleic acid (or a linoleic acid) moiety at the sn-1 or sn-3 positions are resolved by the chiral column. This new chiral separation method can be used in combination with atmospheric pressure chemical ionization mass spectrometry to determine the sn-OOP/sn-POO ratio in palm oil. This method is applicable for the chiral separation of asymmetric TAGs in palm oil.

Profiling of regioisomeric triacylglycerols in edible oils by supercritical fluid chromatography/tandem mass spectrometry☆

In this study, supercritical fluid chromatography (SFC) coupled with triple quadrupole mass spectrometry was applied to the profiling of several regioisomeric triacylglycerols (TAGs). SFC conditions (column, flow rate, modifier) were optimized for the effective separation of TAGs. In the column test, a triacontyl (C30) silica gel reversed-phase column was selected to separate TAG regioisomers. Multiple reaction monitoring was used to selectively quantify each TAG. Then, the method was used to perform detailed characterization of a diverse array of TAGs in palm and canola oils. Seventy TAGs (C46:0-C60:2) of these oils were successfully analyzed as a result, and twenty isomeric TAG pairs were separated well. In particular, this method provided the fast and high resolution separation of six regioisomeric TAG pairs (PPLn/PLnP, PPL/PLP, PPO/POP, SPLn/SLnP, SPO/SOP, SSO/SOS-stearic acid (S, 18:0), oleic acid (O, 18:1), linoleic acid (L, 18:2), linolenic acid (Ln, 18:3), palmitic acid (P, 16:0)) in a short time (50min) as compared to high performance liquid chromatography. We were able to demonstrate the utility of this method for the analysis of regioisomeric TAGs in edible oils.

Pro-oxidant/antioxidant behaviours of ascorbic acid, tocopherol, and plant extracts in n-3 highly unsaturated fatty acid rich oil-in-water emulsions.

This study investigated the oxidative stability of n-3 highly unsaturated fatty acid (n-3 HUFA) rich (35% n-3 HUFA) oil-in-water emulsions (10 wt% oil) with commercial antioxidants and natural plant extracts. Ascorbic acid, α-tocopherol, and the extracts of Indian gooseberry fruit (Emblica officinalis) (IGFE) and sweet basil leaves (Ocimum basilicum L.) (SBLE) were used for the study as antioxidants. The progress of oxidation in the systems was evaluated at 35 °C over 120 h against a control (without antioxidant) by monitoring the formation of primary (conjugated dienes) and secondary (volatile carbonyl compounds) oxidation products. Volatile carbonyl compounds were trapped as derivatives of pentafluorophenyl hydrazine and quantified by headspace solid-phase microextraction analysis. About 40 volatile carbonyls were successfully identified by this method. trans,trans-2,4-Heptadienal, trans,cis-2,4-heptadienal, 3,5-octadien-2-one, and 1-penten-3-ol were predominant. The volatile carbonyl compounds and conjugated dienes were formed at low rates in emulsion systems in which α-tocopherol and natural plant extracts had been introduced, compared to the control. Emulsion systems containing ascorbic acid showed low stability, as indicated by the oxidation products that were formed at high rates compared to the control. These results indicated that ascorbic acid activated the oxidation reactions in n-3 HUFA rich water emulsions, while natural plant extracts that were rich in polyphenols and α-tocopherol were active as antioxidants. The present study further demonstrated the applicability of the polar paradox theory in the determination of stability for n-3 HUFA rich water emulsions with commercial antioxidants and natural plant extracts.

Actual ratios of triacylglycerol positional isomers consisting of saturated and highly unsaturated fatty acids in fishes and marine mammals.

The distribution of fatty acid species at the (sn-1, 3) position or the (sn-2) position of triacylglycerol (TAG) in natural fats and oils has already been analysed by many researchers and several interesting results have been reported. However, most of these reports only focused on the distribution of fatty acids at the or positions in TAG, and did not take account of the combination of fatty acids in the TAG, i.e., the TAG positional isomers. In this study, the actual ratios of TAG positional isomer pairs, consisting of palmitic acid and highly unsaturated fatty acid (HUFA) such as DHA or EPA, in fish and marine mammals were investigated using a high-performance liquid chromatography/atmospheric pressure chemical ionisation-mass spectrometry (HPLC/APCI-MS) system equipped with tandem jointed non-endcapped polymeric ODS columns. The results show that for combinations of DHA or EPA with two palmitic acids in the TAG of marine mammals, binding was almost all at the α position. In contrast, binding of DHA or EPA was mainly at the β position in fish. The preferred DHA and EPA positions in TAG were the same in the same marine mammal or fish. The binding position tendency of HUFA in TAG positional isomers consisting of two HUFAs and one palmitic acid was the same as that for combinations of one HUFA and two palmitic acids. These results were interpreted as showing that the preferred fatty acid species of sn-glycerol-3-phosphate acyltransferase and 1-acyl-sn-glycerol-3-phosphate acyltransferase in marine mammals are different to those in fish and other animals, or that diacylglycerol acyltransferase in marine mammals favours 1,2-dipalmitoyl-sn-glycerol formed from 1,2-dipalmitoyl-sn-glycerol-3-phosphatidate if HUFA is the reaction substrate.

Variation in lipid classes and fatty acid composition of salmon shark (Lamna ditropis) liver with season and gender

The influence of season and gender on lipid content, lipid classes, and fatty acid compositions was assessed in livers of salmon shark (Lamna ditropis), caught in the Pacific Ocean. No significant difference in the hepatosomatic index was noted with season, though the lipid content was significantly higher (P<0.05) in winter. Triacylglycerol (TAG) was identified as the predominant lipid class (78.5-82.0%), followed by sterol esters (5.7-9.1%) and hydrocarbons (3.4-5.4%). No significant differences were observed in TAG composition with respect to the season or gender. However, diacylglyceryl ether contents were significantly higher (P<0.05) in winter (3.8-5.3%) than those obtained in summer (1.3-1.1%). Polyunsaturated fatty acids constituted the major fatty acid class of salmon shark total liver lipid and docosahexaenoic acid (C22:6n-3) (22.7-28.4%) was the most abundant fatty acid which was significantly lower (P<0.05) in winter. These results suggested that lipid characteristics of salmon shark liver were influenced by season, but not by gender.