Aspects of the regulation of long-chain fatty acid oxidation in bovine liver

Factors involved in regulation of bovine hepatic fatty acid oxidation were examined using liver slices. Fatty acid oxidation was measured as the conversion of 1-[14C] palmitate to 14CO2 and total [14C] acid-soluble metabolites. Extended (5 to 7 d) fasting of Holstein cows had relatively little effect on palmitate oxidation to acid-soluble metabolites by liver slices, although oxidation to CO2 was decreased. Feeding a restricted roughage, high concentrate ration to lactating cows resulted in inhibition of palmitate oxidation. Insulin, glucose, and acetate inhibited palmitate oxidation by bovine liver slices. We suggest the regulation of bovine hepatic fatty acid oxidation may be less dependent on hormonally induced alterations in enzyme activity as observed in rat liver and more dependent upon action of rumen fermentation products or their metabolites on enzyme systems involved in fatty acid oxidation.     

Role of diacylglycerol O-acyltransferase (DGAT) isoforms in bovine hepatic fatty acid metabolism

Fatty acid accumulation in hepatocytes induced by high concentrations of fatty acids due to lipolysis and the associated oxidative damage they cause occur most frequently after calving. Because of their role in esterification of fatty acids, diacylglycerol acyltransferase isoforms (DGAT1 and DGAT2) could play a role in the susceptibility of dairy cows to develop fatty liver. To gain mechanistic insights, we performed in vivo and in vitro analyses using liver biopsies or isolated primary hepatocytes. The in vivo study (n = 5 cows/group) involved healthy cows [average liver triacylglycerol (TAG) = 0.78%; 0.58 to 0.93%, ratio of triglyceride weight to wet liver weight] or cows diagnosed with fatty liver (average TAG = 7.60%; 5.31 to 10.54%). In vitro, hepatocytes isolated from 3 healthy female calves (1 d old, 44 to 53 kg) were challenged with (fatty acids) or without (control) a 1.2 mM mixture of fatty acids in an attempt to induce metabolic stress. Furthermore, hepatocytes were treated with DGAT1 inhibitor or DGAT2 inhibitor for 2 h followed by a challenge with (DGAT1 inhibitor + fatty acids or DGAT2 inhibitor + fatty acids) or without (DGAT1 inhibitor or DGAT2 inhibitor) the 1.2 mM mixture of fatty acids for 12 h. Data analysis of liver biopsies was compared using a 2-tailed unpaired Student's t-test. Data from calf hepatocyte treatment comparisons were assessed by one-way ANOVA, and multiplicity for each experiment was adjusted by the Holm's procedure. Data indicated that both fatty liver and in vitro challenge with fatty acids were associated with greater mRNA and protein abundance of SREBF1, FASN, DGAT1, and DGAT2. In contrast, mRNA and protein abundance of CPT1A and very low-density lipoprotein synthesis-related proteins MTTP and APOB were markedly lower. However, compared with fatty acid challenge alone, DGAT1 inhibitor + fatty acids led to greater mRNA and protein abundance of CPT1A and APOB, and greater mRNA abundance of SREBF1 and MTTP. Furthermore, this treatment led to lower mRNA abundance of FASN and DGAT2 and TAG concentrations. Compared with fatty acid challenge alone, DGAT2 inhibitor + fatty acids led to greater mRNA and protein abundance of CPT1A, MTTP, and APOB, and lower mRNA and protein abundance of SREBF1 and FASN. In addition, compared with control and fatty acids, there was greater protein abundance of GRP78 and PERK in both DGAT1 and DGAT2 inhibitor with or without fatty acids. Furthermore, compared with control and fatty acids, reactive oxygen species concentrations in the DGAT1 inhibitor with or without fatty acid group was greater. Overall, data suggested that DGAT1 is particularly relevant in the context of hepatocyte TAG synthesis from exogenous fatty acids. Disruption of both DGAT1 and DGAT2 altered lipid homeostasis, channeling fatty acids toward oxidation and generation of reactive oxygen species. Both DGAT isoforms play a role in promoting fatty acid storage into TAG and lipid droplets to protect hepatocytes from oxidative damage.     

Acetyl CoA carboxylase shares control of fatty acid synthesis with fatty acid synthase in bovine mammary homogenate

The objectives of this research were to determine the flux control coefficients for acetyl CoA carboxylase and fatty acid synthase using an in vitro preparation of bovine mammary homogenate. For an enzyme to be considered rate limiting with the use of metabolic control analysis, its control coefficient would be equal to unity. The hypothesis for this experiment was that the control coefficient for acetyl CoA carboxylase was not equal to unity, and that this enzyme was not, therefore, the rate-limiting step. Mammary tissue was isolated from lactating Holstein cows at slaughter and frozen in liquid nitrogen. Tissue was ground, homogenized, and centrifuged to obtain a postmitochondrial supernatant for use in in vitro incubations containing labeled acetate. Specific inhibitors for acetyl CoA carboxylase and fatty acid synthase were used to fractionally inhibit de novo synthesis for the calculation of flux control coefficients. The composition of fatty acids synthesized in the absence of enzyme inhibitors was similar to the composition of fatty acids in the presence of inhibitors. Calculations following avidin inhibition of acetyl CoA carboxylase determined the flux control coefficient was 0.63 ± 0.15, which means that 63% of the control of fatty acid synthesis is exerted by acetyl CoA carboxylase. The remaining control (37%) was from fatty acid synthase, which indicates a significant degree of control over the flux of acetate in de novo synthesis resides with this enzyme. The rate-limiting status ascribed to acetyl CoA carboxylase was not supported, because the flux control coefficient was less than unity. Metabolic control analysis, through its use of pathway product measurements, allows for potential interactions in the pathway such as feedback inhibition contribution to the flux control coefficients, which would not otherwise be considered in studies measuring enzyme kinetics with purified enzymes.     

A relationship between bovine fat colour and fatty acid composition

Subcutaneous adipose tissue was obtained from pasture-grazed (n = 13) and short-term (70 days) grain-fed (n = 13) cattle. The yellow colour of the adipose tissue was assessed by Minolta b(?) value readings and by determination of total carotenoids and the two measurements gave a correlation coefficient of 0·79 (P < 0·01). The fatty acid composition of the samples varied with fat colour. As the b(?) value and the carotenoid content of the fat increased, there was a significant increase in the total percentage of cis mono-unsaturated fatty acids and a decrease in saturated fatty acids (P < 0·01). Consequently, the ratio of cis mono-unsaturated to saturated fatty acids was found to be higher in those samples exhibiting a greater yellow colour.     

γ-辐射对不饱和脂肪酸氧化反应的影响

对亚油酸和共轭亚油酸,分别在空气气氛和氮气气氛下进行了一定剂量的60Co γ-辐射,每隔2 d测定其过氧化值.研究结果表明,辐射剂量和放置时间对不饱和脂肪酸的过氧化值有较大影响,共轭亚油酸的氧化稳定性好于亚油酸.并进一步探讨了不饱和脂肪酸在γ-辐射下过氧化值与反应机制之间的内在联系.     

Distribution and fatty acid content of phospholipids from bovine milk and bovine milk fat globule membranes

The phospholipid (PL) content was determined comparatively in the milk fat globule membrane (MFGM) and whole milk including their fatty acid profiles. The possible role of milk PLs in defence against pathogens was also addressed. The MFGM and whole milk showed a similar distribution of PL species; however, the fatty acid contents of the PL species were different. Total PL from MFGM showed a decrease in C18:0 content in parallel with an increase in C18:1 and C18:2 and very long-chain fatty acid (more than C20) content. No significant differences in the fatty acid content of phosphatidylcholine and sphingomyelin from either source were found. However, the phosphatidylethanolamine from MFGM had more C18:1 and C18:2 and less C14:0 and C16:0 than that from whole milk. A similar but less pronounced result was found for phosphatidylserine/phosphatidylinositol. Enterotoxigenic Escherichia coli strains failed to bind to PL, which had been previously separated by high-performance thin-layer chromatography.     

焙烤对核桃仁风味及其油脂氧化的影响

焙烤可以有效增加坚果的风味,但由于坚果中含有的大量不饱和脂肪酸极易受热氧化,因此要严格控制焙烤工艺.以带囊衣核桃仁作为研究材料,对不同温度和时间下焙烤的核桃仁进行了风味评定和油脂过氧化值的测定,得到125℃,4 h是油脂氧化程度最小而风味最佳的焙烤条件.     

Seasonal variation in fatty acid and triacylglycerol composition of bovine milk fat

The aim of this study was to assess the effects of seasonal variation on the changes of the fatty acid (FA) and triacylglycerol (TAG) composition of bovine milk fat (MF) in a nonseasonal milking system. Weekly milk samples were collected from 14 dairy factories and pooled per week as representative samples of the average Dutch bovine milk. The sample collection started in May 2017 and finished in April 2018, resulting in a total of 52 samples, corresponding to each week of the year. The samples were analyzed for MF content (%) and FA and TAG composition using gas chromatography with flame-ionization detection. The increased intake of C18:3 cis-9,12,15 through grass feeding in spring and summer was associated with major changes in MF FA composition, including reduced proportions of de novo synthesized FA and presence of several rumen biohydrogenation products and conjugated linoleic acid isomers in MF. These changes in seasonal FA composition had an effect on TAG seasonal variation. The TAG seasonal variation showed that all TAG groups were significantly different between months. The low molecular weight and the medium molecular weight TAG groups increased in winter and decreased in summer, whereas the high molecular weight TAG groups increased in summer and decreased in winter. Based on pooled monthly samples, MALDI-TOF-mass spectrometry allowed the analysis of even- and odd-chain TAG species in MF based on their total carbon number and number of double bonds. These analyses indicated saturated TAG species to be greatest in winter, whereas monounsaturated, polyunsaturated, and odd-chain TAG species were greatest in summer. Our study showed that TAG seasonal variation in a nonseasonal milking system is influenced by the variation in FA composition throughout the seasons.     

Facilitative effects of Eucommia ulmoides on fatty acid oxidation in hypertriglyceridaemic rats

BACKGROUND: Tea made from Eucommia ulmoides leaves is widely consumed as a health food, since recent studies have revealed various pharmacological effects of the tea, e.g. a hypotriglyceridaemic effect. This study was conducted to elucidate the mechanisms underlying the plasma triglyceride‐lowering effect of E. ulmoides leaves. RESULTS: Rats were divided into four groups: a normal group, a group fed a high‐fat/high‐fructose diet (untreated group) and two groups fed a high‐fat/high‐fructose diet and E. ulmoides tea (4 or 20 g L^?1 extract, treated groups). Plasma triglyceride concentrations were reduced in treated groups in a dose‐dependent manner compared with the untreated group. DNA microarray analysis revealed that genes involved in hepatic α‐, β‐ and ω‐oxidation, mainly related to the peroxisome proliferator‐activated receptor α and δ signalling pathway, were up‐regulated in the treated group. High‐performance liquid chromatography analysis showed that E. ulmoides leaves contain three phytochemicals that make up 60 mg g^?1 of the material and are likely to be the active components. CONCLUSION: This study indicates that the promotion of fatty acid oxidation, probably by the action of phytochemicals, participates in the ameliorative effect of E. ulmoides leaves on hypertriglyceridaemia. These findings provide the scientific evidence for the functionality of E. ulmoides. Copyright © 2011 Society of Chemical Industry     

高温处理对牛肉脂肪酸及脂肪氧化的影响

对新鲜牛背最长肌进行高温处理(110,115,121℃),并分别保持5,10,15,20min,采用气相色谱法研究高温对牛背最长肌肌内脂肪酸组成和脂肪氧化的影响。结果表明,121℃高温处理时,肌内脂肪氧化速度显著大于110,115℃的(P0.05);饱和脂肪酸(SFA)含量明显增加(P0.05),不饱和脂肪酸(UFA)含量降低(P0.05);SFA受处理温度(110,115℃)影响不显著(P0.05),受处理时间影响显著(P0.05),而热处理温度和时间对UFA含量均有显著影响(P0.05);长链饱和脂肪酸出现一定程度的断裂和降解,短链及中链脂肪酸含量显著上升(P0.05)。     

Lipid oxidation in n-3 fatty acid enriched Dutch style fermented sausages

Dutch style fermented sausages were manufactured with a substitution of 10%, 15% and 20% of pork backfat by flaxseed oil and canola oil, pre-emulsified with soy protein isolate. The 15% and 20% substitution were also reached by adding encapsulated flaxseed oil and encapsulated fish oil and by adding flaxseed oil, pre-emulsified with sodium caseinate, respectively. The products were sliced, packaged in an oxygen-enriched atmosphere and stored in the dark for 12 weeks at 7°C. No differences were detected in moisture, protein and fat content between control and modified sausages, with the exception of the formulation with sodium caseinate. The PUFA/SFA ratio increased from 0.30 in the control to 0.42-0.48 in the sausages with canola oil and to 0.49-0.71 in the sausages with flaxseed oil. The n-6/n-3 ratio decreased from 11.20 in the control to 6.94-5.12 in the sausages with canola oil and to 1.93-1.05 in the sausages with flaxseed oil. The addition of canola oil and encapsulated flaxseed oil resulted in a comparable shelf life as the control in terms of lipid oxidation. In the samples with addition of pre-emulsified flaxseed oil, especially with sodium caseinate, lipid oxidation clearly increased during storage. Physical and sensory analysis showed that the sausages with encapsulated fish oil and flaxseed oil resembled the control most.     

Interactions between gluconeogenesis and fatty acid oxidation in isolated sheep hepatocytes.

The interaction of gluconeogenesis and fatty acid oxidation in isolated sheep hepatocytes was studied. Addition of tetradecylglycidic acid, an inhibitor of carnitine palmitoyltransferase I (EC 2.3.1.21), to isolated hepatocytes inhibited gluconeogenesis from a mixture of pyruvate plus lactate and from propionate alone. Inhibition constants for tetradecylglycidic acid on gluconeogenesis were 4.77 +/- 1.00 microM and 7.25 +/- 1.52 microM, respectively, for pyruvate plus lactate and for propionate as gluconeogenic substrates. The inhibition constants were not different. At the highest substrate concentrations examined, gluconeogenesis from pyruvate plus lactate and from propionate in the presence of 10 microM tetradecylglycidic acid was 47.3 and 41.4% of their respective controls. Similar to previous observations with butyrate, caproate addition inhibited gluconeogenesis from propionate by isolated hepatocytes and was unable to prevent inhibition of gluconeogenesis induced by tetradecylglycidic acid. Carnitine palmitoyltransferase I activity was lower in mitochondria isolated from hepatocytes preincubated with insulin than in control hepatocytes. The data suggest 1) that maximum rates of gluconeogenesis in isolated sheep hepatocytes from either pyruvate plus lactate or from propionate as gluconeogenic substrates require beta-oxidation, 2) that intermediates common to the metabolism of butyrate and caproate may be involved in the inhibition of propionate conversion to glucose by isolated sheep hepatocytes, and 3) that carnitine palmitoyltransferase I activity in isolated sheep hepatocytes can be modulated by insulin treatment.     

脂肪氧化初期脂肪酸与氧化指标变化相关性研究

以牦牛肾周脂为研究对象,研究了其在储藏期(15±1)℃下脂肪酸与氧化指标(TBARS和PV)在动态变化中的相关性。结果表明:牦牛肾脂脂肪氧化初期发生于储藏期第0～25天。比较脂肪酸与氧化指标变化后发现,SFA与TBARS(和PV)呈较高正相关(≥0.971),MUFA和PUFA与TBARS(和PV)具有较低负相关系数(≤0.926)。在MUFA中,C16:1和C18:1与氧化指标的负相关系数较低,分别为-0.932和-0.981;在PUFA中,除CLA和LA外,其他多不饱和脂肪酸与氧化指标的相关性均较好(≥0.891)。因此,TBARS和PV可以被用于间接评估脂肪在工业开发前储存期间不饱和脂肪酸的含量分布状态。     

Inhibition of fatty acid synthesis in bovine mammary homogenate by palmitic acid is not a detergent effect

Supplemental fat fed to dairy cows affects the fat composition of milk by reducing the yield of mammary synthesized fatty acids. The effect has been attributed to a potential allosteric inhibition of acetyl coenzyme-A, a key enzyme in fatty acid synthesis. In vitro experiments have demonstrated an inhibition of fatty acid synthesis when long-chain fatty acids are added to incubations. However, in vitro inhibition can result from a nonspecific detergent effect arising from an inherent physical property of fatty acids. An allosteric role for palmitic acid has not been tested in bovine mammary tissue. The objective of this experiment was to test the hypothesis that palmitic acid is an allosteric inhibitor of fatty acid synthesis in mammary tissue. We tested for a detergent effect by including a synthetic detergent, sodium dodecyl sulfate, under identical incubation conditions. A subcellular supernatant fraction of mammary tissue was used for incubations in the present experiment. The incubation system produced free fatty acids in a linear fashion for time and protein content. Results indicated that fatty acid synthesis was affected by the addition of palmitic acid to the incubations but not by caprylic acid, a short-chain fatty acid. Sodium dodecyl sulfate did not affect fatty acid synthesis at the concentrations used. The results of the present experiment indicate that palmitic acid inhibited fatty acid synthesis, and the effect was not the result of a detergent effect.     

Regulation of volatile fatty acid uptake by mitochondrial acyl CoA synthetases of bovine heart

Purification of components of heart mitochondria activating short chain fatty acids prepared from tissue of lactating Holstein cows demonstrated predominantly one acyl CoA synthetase, acetyl CoA synthetase activating acetate, and propionate. Activity of butyryl CoA synthetase was low. Propionyl CoA synthetase characteristically in bovine liver and kidney tissue could not be demonstrated in heart mitochondria. Thus, of the ruminally derived volatile fatty acids only acetate can be used by heart mitochondria as a primary energy source because of small quantities of propionate in peripheral blood. Acetyl CoA synthetase was a glycoprotein composed of a single polypeptide chain of apparent molecular weight 67,500. The Michaelis-Menten constant for acetate was 1.8 x 10(-4)M. By comparison with literature for blood acetate concentration we concluded that enzyme is saturated with substrate at all physiological concentrations of acetate. These kinetic properties ensure a constant supply of acetate as an energy source for maintaining heart function in ruminants.     

Impact of free fatty acid concentration and structure on lipid oxidation in oil-in-water emulsions

Free fatty acids are strong prooxidants in both bulk and emulsified oils. Addition of oleic acid to an oil-in-water emulsions increased lipid hydroperoxide and hexanal formation at free fatty acid concentrations as low as 0.1% of the lipid. The prooxidant effect of free fatty acids was dependent on fatty acid type with lipid oxidation rates being in the order of linolenic < linoleic < oleic. There were no significant differences in lipid oxidation rates when free fatty acid isomers with cis or trans double bonds were compared. The prooxidant activity of the free fatty acids was postulated to be due to their ability to attract prooxidant metals as well as co-oxidise the triacylglycerol in the oil. Overall, these results show that the oxidative stability of oil-in-water emulsions is strongly linked to both the concentration and type of free fatty acids present.     

杀菌温度对乳化肠中脂肪酸组成和脂肪氧化的影响

为明确脂肪对不同熟化温度下（80 ℃,30 min、90 ℃,30 min、100 ℃,30 min和121 ℃,20 min）猪肉乳化香肠挥发性物质的影响,采用感官评价、电子鼻（Electronic nose,E-nose）和固相微萃取-气相色谱质谱联用技术（Solid-phase microextraction coupled-gas chromatography-mass spectrometry,SPME-GC-MS）对添加或不添加脂肪的乳化肠在不同熟化温度下挥发性物质进行分析。结果表明,添加脂肪的乳化肠,熟化条件为100 ℃,30 min的样品风味最好,添加脂肪可提高乳化肠脂香味、硫磺味、哈喇味和青草味的感官强度,抑制高温乳化肠肉香味和蘑菇味的感官强度;电子鼻能有效区分添加脂肪或不添加脂肪的样品,但80 ℃,30 min组的样品无法有效区分;添加脂肪的样品中,除100 ℃,30 min组和121 ℃,20 min组外,电子鼻能很好的将不同熟化温度的样品区分开。SPME-GC-MS结果显示,8个处理组共检测出56种挥发性化合物,己醛、庚醛、辛醛、戊醛、壬醛、1-辛烯、1-辛烯-3-醇和甲硫醇等关键化合物的含量随熟化温度的升高而增加。偏最小二乘判别分析（Partial least squares discriminant analysis,PLS-DA）筛选出己醛、戊醛和n-己酸乙烯酯是不同熟化温度乳化肠气味差异的潜在标志物。采用正交偏最小二乘判别分析（Orthogonal partial least squares discriminant analysis,OPLS-DA）筛选出添加与不添加脂肪的样品在4种熟化温度下的差异化合物分别为17种、17种、22种和25种。以上结果表明,熟化条件相同,添加脂肪的样品挥发性物质含量显著高于不添加脂肪的样品,100 ℃,30 min的熟化条件更有利于乳化肠风味的形成。

     

不同类型膳食脂肪酸对肥胖小鼠肝脏及其血液中脂肪酸组成和代谢相关基因影响

目的探讨不同类型膳食脂肪酸对肥胖小鼠肝脏及其血液中脂肪酸组成及代谢相关基因的影响。方法 8周龄雄性C57BL/6小鼠随机分为7组,即对照组(喂基础饲料)、长链饱和脂肪酸(LCSFA)组(喂猪油高脂饲料)、中链饱和脂肪酸(MCSFA)组(喂椰子油高脂饲料)、n-3多不饱和脂肪酸(n-3 PUFA)组(喂亚麻籽油高脂饲料)、n-6多不饱和脂肪酸(n-6 PUFA)组(喂大豆油高脂饲料)、单不饱和脂肪酸(MUFA)组(喂橄榄油高脂饲料)和反式脂肪酸(TFA)组(喂8%氢化大豆油高脂饲料),每组10只,共干预16周,所有种类饲料总能量均相同,基础饲料脂肪供能比为10%,各高脂饲料脂肪供能比均为45%,喂养周期结束后,禁食12 h,麻醉后立刻解剖取出肝脏。采用气相色谱法分析肝脏及其血液中脂肪酸组成的变化,实时荧光定量聚合酶链式反应(PCR)检测肝脏脂肪酸代谢相关基因的表达,肝脏脂质沉积采用油红O染色法检测。结果与对照组比较,LCSFA组、MCSFA组、n-6 PUFA组、MUFA组和TFA组小鼠肝脏中均出现明显的脂质沉积,n-3 PUFA组小鼠肝脏未出现明显的脂质沉积。与对照组比较,LCSFA组小鼠肝脏及血液中总n-6 PUFA和总PUFA含量升高; n-3 PUFA组小鼠肝脏及血液中总n-3 PUFA和总PUFA含量增加,但总MUFA含量减少; n-6 PUFA组小鼠肝脏及血液中总n-6PUFA、总n-3 PUFA和总PUFA含量升高,但总饱和脂肪酸(SFA)和总MUFA含量降低; MUFA组小鼠肝脏及血液中总SFA含量减少; TFA组小鼠肝脏及血液中C18∶1 n-9t(TFA)含量升高;以上差异均有统计学意义(P0. 05)。LCSFA组和MCSFA组小鼠肝脏脂肪酸代谢基因固醇调节元件结合蛋白1(SREBP-1c) mRNA水平高于对照组和n-6 PUFA组,差异均有统计学意义(P0. 05)。结论小鼠肝脏及其血液中脂肪酸构成与其对应的膳食脂肪酸模式一致。不同类型脂肪酸高脂饲料可通过对相关基因的表达影响肥胖状态下肝脏的脂质代谢及脂质沉积。     

Validation of the sensitive and accurate quantitation of the fatty acid distribution in bovine milk

A method for the precise analysis of the complex mixture of fatty acids in milk has been developed and validated. The triacylglycerol of nonanoic acid was applied as the internal standard (ISTD) for absolute quantification. Milk lipids were extracted by miniaturised ultrasonication and methylated with trimethylsulfonium hydroxide. Resulting fatty acid methyl esters were determined by gas chromatography with flame ionisation detection giving excellent resolution, including separation of several 18:1 isomers. The low quantitation limit (0.01 mg mL⁻¹ milk) indicates that the sensitivity of the method is sufficient to quantify up to 50 fatty acids, from 4:0 to 23:0. Measurements of precision provided excellent results for different bovine milk samples of different fat content (coefficient of variance: 1.9% and 9.8% for intra- and interday precision, respectively). Recovery averaged 108 ± 3.5%. Evaluation of methods for determining the total fat content showed that gravimetry is no longer needed when using the ISTD.