Analysis of Headspace Volatile and Oxidized Volatile Compounds in DHA-enriched Fish Oil on Accelerated Oxidative Storage

ABSTRACT: Oxidative stability of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) and volatile and oxidized volatile compounds in 2 types of DHA-enriched fish oil, triacylglycerol (TG) and ethyl ester (EE), were studied during storage at 80 °C with aeration. The rate of DHA autoxidation was higher than that of EPA. DHA in EE form was more susceptible to autoxidation than in TG form. Thirty-one volatile compounds were identified in EE and 23 volatile compounds in TG. (E)-2-pentenal, 2-(1-pentenyl) furan, and (E,E)-2,4-heptadienal were commonly detected as oxidized volatile compounds from TG and EE fish oil. These volatile oxidized compounds might be formed mainly from the oxidation of DHA and EPA, the main fatty acids of the oil.     

近红外技术在南极磷虾油关键品质指标快速检测中的应用研究

采用偏最小二乘法（Partial least squares,PLS）作为建模方法,对磷虾油近红外光谱的一阶微分（First-order difference,FD）、FD+SG（Savitzky-Golay,SG）滤波、FD+N（Norris,N）滤波、二阶微分（Second-order difference,SD）、SD+SG和SD+N等6种单一或复合方法进行处理,通过对不同方式处理后预测模型的交互验证均方根误差（Root mean square error of cross validation,RMSECV）、外部验证残差均方根（Root mean square error of external prediction,RMSEP）和外部验证用样品真实值的标准差（SD）与RMSEP的比值（The ratio of the RMSEP to standard deviation of reference data in the prediction,RPDEV）、建模相关系数（Correlation coefficient in calibration,RC）、交互验证相关系数（Correlation coefficient in cross validation,RCV）和外部验证相关系数（Correlation coefficient in external validation,REV）等参数比较,确定了磷虾油磷脂、EPA和DHA的近红外预测模型最佳处理方式为FD、FD和SD+N,酸价指标模型不需处理。在最优条件下,四种成分近红外预测模型的RC、REV和RCV,除了酸价的RCV略小（0.917）其余均达到0.95以上,同时,四种成分的RPDEV和RPDCV值,除酸价的RPDCV为2.365,略小于2.5,其余均符合大于2.5的要求,说明磷虾油磷脂、EPA和DHA的近红外预测模型预测准确度良好;RMSEC和RMSECV相差不大,说明模型稳定性较好。由于含量低、组成复杂等原因,磷虾油虾青素近红外检测模型的RC、RCV和REV均在0.60以下,说明近红外检测不适用于磷虾油中虾青素成分的快速检测。本文证实了近红外光谱技术可作为磷虾油中磷脂、EPA、DHA和酸价等主要指标的快速检测方法,是传统化学检测方法的有效替代和补充。     

超临界CO_2萃取精馏EPA与DHA的实验研究

采取超临界 CO2萃取—精馏技术,对经尿素包合预处理的鱼油脂肪酸乙酯在超临界 CO2流体中的溶解情况进行了考察,探索了 EPA、 DHA的分离提纯工艺,并讨论了实验的影响因素。实验结果表明：采取温度梯度结合逐步升压法,能使鱼油脂肪酸乙酯按碳链长度依次分离, EPA＋ DHA提纯至 90％,两次分离后, EPA提纯至 67％, DHA提纯至 90％以上。     

Dietary krill oil increases docosahexaenoic acid and reduces 2-arachidonoylglycerol but not N-acylethanolamine levels in the brain of obese Zucker rats

Evidence suggests that dietary long chain polyunsaturated fatty acids (LCPUFAs), and particularly those belonging to the n-3 family, may influence the brain fatty acid profile and, thereby, the biosynthesis of endocannabinoids in rodents. However, the doses used are usually quite high and not comparable with human intake. Recently, we have shown that relatively low doses of dietary n-3 LCPUFAs (4 weeks), in the form of either fish or krill oil, balanced for EPA and DHA content, and against a control diet with no EPA and DHA and similar contents of oleic, linoleic and α-linolenic acids, lower the concentrations of the endocannabinoids, anandamide (AEA) and 2-arachidonoylglycerol (2-AG), in the visceral adipose tissue, and of AEA in the liver and heart, of obese Zucker rats. This, in turn, is associated with lower levels of arachidonic acid in membrane phospholipids and with amelioration of some metabolic syndrome parameters. We investigated here whether in Zucker rats, under the same conditions, fish and krill oil are also able to influence LCPUFA and endocannabinoid profiles in brain. Only krill oil was able to increase significantly DHA levels in brain phospholipids, with no changes in arachidonic acid. DHA increase was associated with lower levels of 2-AG in the brain, whereas AEA and its congeners, N-palmitoylethanolamine and N-oleoylethanolamine, were unchanged. We conclude that, despite the strong impact of dietary n-3 fatty acid on endocannabinoid levels previously observed in peripheral tissues, in the brain only 2-AG is affected by dietary krill oil, suggesting that the beneficial effect of the latter on the metabolic syndrome is mostly exerted by modifying peripheral endocannabinoids. Nevertheless, possible effects of dietary krill oil in the brain through modification of 2-AG levels deserve further investigation.     

NMR结合GC-MS技术分析鱿鱼肝脏油脂及其脂肪酸组成

利用NMR结合GC/MS技术对鱿鱼肝脏油脂及其脂肪酸组成进行分析。₁NMR结果表明,鱿鱼肝脏总脂脂肪酸中,多不饱和脂肪酸与饱和脂肪酸的含量比约为7:5,n-3型多不饱和脂肪酸占脂肪酸总量的35%;₁₃C NMR结果表明,鱿鱼肝脏总脂以甘油三酯(71.87%)为主,其次是游离脂肪酸(20.98%)和磷脂(5.29%),胆固醇(1.13%)和胆固醇酯(0.73%)含量较低,鱿鱼肝油中的DHA和EPA主要以游离形式存在,分别占DHA总量和EPA总量的61.24%和63.11%;GC/MS结果显示,鱿鱼肝脏总脂脂肪酸主要为C16:0、C18:1、C20:1、EPA和DHA,多不饱和脂肪酸的含量高达38.80%,且EPA和DHA占脂肪酸总量的31.50%,表明鱿鱼肝脏具有较高的营养价值及开发前景。     

Comparison of Infrared Spectroscopy and Nuclear Magnetic Resonance Techniques in Tandem with Multivariable Selection for Rapid Determination of ω-3 Polyunsaturated Fatty Acids in Fish Oil

This study investigated the potential of using four spectroscopic techniques including visible–short-wave near infrared, long-wave near infrared (LNIR), mid-infrared, and nuclear magnetic resonance (NMR) spectroscopy in tandem with multivariable selection and calibration for rapid determination of three important ω-3 polyunsaturated fatty acids (PUFA), namely eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and docosapentaenoic acid (DPA) in fish oil. Quantitative models were established between the spectral data and reference PUFA contents of samples based on partial least squares regression (PLSR) algorithm. Successive projections algorithm (SPA) and uninformative variable elimination (UVE) were used to select the most important variables for prediction. The average decrements of 23.20 % for root mean square error of cross-validation (RMSECV) and 64.90 % for the absolute value between root mean square error of calibration and RMSECV (AV_RMSE) in all 12 cases achieved after over 90 % variables were eliminated. UVE was also found to be helpful to improve the efficiency of SPA’s variable selection in 8/12 cases. The best predictions for EPA, DHA, and DPA were all achieved by NMR spectroscopy (determination coefficients of cross-validation (r _CV ² ) of 0.970, 0.982, and 0.983 and the RMSECV of 11.48, 4.73, and 0.77 mg/g for the EPA, DHA, and DPA predictions, respectively). LNIR spectra also did good predictions similar to NMR. The results demonstrated that the laborious and time-consuming gas chromatography method could be replaced by spectroscopic techniques in tandem with PLSR modeling and variable selection in order to provide a rapid and reliable inspection of PUFA in fish oil.     

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.     

13C NMR study of peach oil

The detailed analysis of peach seed oil was performed using ¹H and ¹³C NMR spectroscopy. From the ¹³C NMR solution spectra it was possible to detect the content of saturated, mostly palmitic fatty acids (∼100 g kg⁻¹), the oleic–linoleic ratio in sn‐1,3 and sn‐2 positions was (2:1 and 1:2) and the positional distribution of glycerol esters on triacylglycerol chains, which is a mixture of OPO and OOP. With these characteristics this oil can be used in food applications. Copyright © 2005 Society of Chemical Industry     

Properties of South African fish oils: A review

Abstract

This review records original research done at the Fishing Industry Research Institute (FIRI) Programme on crude fish oils since 1985. This work has led to a better understanding of all the variables that determine fish oil quality. It has been shown that while high free fatty acid (FFA) contents in fish oils are due to spoilage of the fish prior to processing, escalation of FFA levels during oil storage is due to ineffective polishing of the oil. Unpolished or poorly polished oils have high phospholipid (P) and protein (N) contents and as a result also high water levels. These three variables cause FFA escalation during oil storage. Statistically derived expressions have been developed which allow prediction of the daily increase in FFA content during storage of oil from the original FFA content of the oil and its water, P and N levels. These expressions have been used to implement a more equitable pricing system for locally produced fish oils. The primary cause, however, for FFA escalation of fish oil is contamination of the oil by bacteria belonging to the genus Alcaligenes. These aerobic bacteria thrive at the oil‐water interface at the bottom of oil storage tanks, and convert mainly phospholipids into oil‐soluble FFA and water‐soluble phosphate esters. The oil‐soluble FFA, which have a considerable lower density than the oil from which they are derived, then diffuse upwards and so contaminate the oil.

Total cholesterol levels in fish oil show a seasonal variation: low levels are found in summer months when the pelagic fish is fat, and high levels in winter when the fish is lean. The cholesterol consists of free and esterified cholesterol. The latter, which is difficult to remove in the refining process, increases during spoilage of the fish.

The levels of chlorophyll in anchovy (Engraulis japonicus) and pilchard (Sarditiops ocellata) oils vary widely and are unpredictable. High chlorophyll levels in these oils may give rise to “green margarine” if insufficient bleaching is carried out on the oil during production of margarine.

South African fish oils, especially anchovy and pilchard oils, are very rich in the two poly‐unsaturated fatty acids eicosapentaenoic (EPA) and docosahexaenoic acid (DHA), which are thought to be beneficial for various aspects of human and animal health. It was found that a high level of EPA corresponded to a low level of DHA and vice versa. This appears to be due to the fact that EPA and DHA are in competition for position two on the glycerol molecule. EPA and DHA levels of anchovy oils ranged from 14.3‐23.3% and 4.6‐12.5% respectively.     

POSITIONAL DISTRIBUTION OF FATTY ACIDS IN TRIACYLGLYCEROLS OF SEAL BLUBBER OIL

The positional distribution of long-chain ω3 polyunsaturated fatty acids namely eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA), and docosahexaenoic acid (DHA) in the triacylglycerol (TAG) molecules of seal blubber oil was determined and compared with that of commercially available menhaden oil via stereospecific analysis. In seal blubber oil, EPA, DPA and DHA occurred mainly in the sn-1 and sn-3 positions of the TAG molecules. The amounts were EPA, 8.36 and 11.2; DPA, 3.99 and 8.21; and DHA, 10.5 and 17.9%, respectively. In menhaden oil, DPA and DHA occurred mainly in the sn-2 position of the TAG at 3.11 and 17.2%, respectively. However, EPA was equally distributed in the sn-2 and sn-3 positions and was present only in minute amounts in the sn-1 position.     

降低高度不饱和脂肪酸乙酯中的过氧化值方法的研究

以LiAlH_4为还原剂,试图降低富含高度不饱和脂肪酸乙酯(主要是5、8、11、14、17、——廿碳五烯酸乙酯,5、8、11、14、17、20——廿二碳六烯酸乙酯。分别简称为EPA乙酯,DHA乙酯)的鱼油酸乙酯的过氧化值。其中,以乙醚为溶剂,LiAlH_4的用量为3％,反应时间为60～90min,去过氧化值的效果显著,方法较简便。     

气相色谱法测定鱼油中的二十碳五烯酸、 二十二碳六烯酸和二十二碳五烯酸

目的通过优化鱼油中二十碳五烯酸(eicosapentaenoic acid,EPA)、二十二碳六烯酸(docosahexaenoic acid,DHA)和二十二碳五烯酸(docosapentaenoic acid,DPA)的测定条件,建立鱼油中EPA、DHA和DPA气相色谱定量检测分析方法。方法采用正己烷处理样品,经色谱柱(Agilent,Elite-WAX,30 m×0.25 mm,0.25μm)分离鱼油中的EPA、DHA、DPA甲酯标准品,并进行定量检测。结果 EPA浓度在0.36~3.6 mg/m L、DHA浓度在0.37~3.7 mg/m L、DPA浓度在0.16~1.62 mg/m L的范围内与峰面积的线性良好,相关系数r0.999。在80%、100%、120%添加水平下,EPA、DHA和DPA的检出限分别为0.01%、0.03%、0.009%,EPA、DHA和DPA的回收率分别为96.2%、96.4%、95.7%。结论气相色谱法灵敏度高、准确、重现性好,适用于鱼油中EPA、DHA和DPA的含量测定。     

利用气相色谱法分析不同萃取方法对鱼油中EPA、DHA含量的影响

本文利用气相色谱法(GC)对鱼油中的二十碳五烯酸(EPA)和二十二碳六烯酸(DHA)进行了定量分析,探究了快速检测EPA、DHA的分析条件,并以黄鲛鱼鱼胃为原料,利用建立的色谱分析方法分别测定了超声强化亚临界水萃取法、超声波辅助溶剂萃取法、酶解提取法和超临界CO₂萃取法获得的鱼油样品中的EPA和DHA的含量。结果显示:EPA的出峰时间是10.015 min,DHA的出峰时间是16.500 min。升温程序和分流比会显著影响目标峰(EPA、DHA)与其相邻峰的分离度R₁和R₂(p<0.01)。该法与国标GB5009.168-2016中的脂肪酸色谱分析方法相比,具有针对性强(可快速检测EPA、DHA)、准确性高、重现性好和快速高效的特点,样品分析时间由82 min缩短至18 min。在最佳分析条件下,上前述四种萃取方法获得鱼油中含有的EPA的检出量依次是54.67、35.23、40.13、33.40 mg/g,DHA的检出量依次是134.01、77.50、102.30、67.31 mg/g。     

Selective concentration of EPA and DHA using Thermomyces lanuginosus lipase is due to fatty acid selectivity and not regioselectivity

The selectivity of anchovy oil hydrolysis was optimised for Thermomyces lanuginosus lipase, so that docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) were concentrated and partially separated from each other. Enzyme concentration and pH control were important factors for effective hydrolysis. Monitoring percent hydrolysis using capillary chromatography with flame ionisation detector (Iatroscan) and fatty acid selectivity using gas chromatography (GC) indicated that during hydrolysis DHA primarily remained on the glycerol backbone, while EPA was progressively removed. ¹³C nuclear magnetic resonance (NMR) data showed that selectivity of hydrolysis was primarily due to fatty acid selectivity and not regioselectivity, with hydrolysis from both sn-1,3 and sn-2 sites being equally favoured.     

Separation and characterization of acetone-soluble phosphatidylcholine from Antarctic krill (Euphausia superba) oil

Antarctic krill is the biggest fishery resource in the world with high content of phosphatidylcholine (PC) as a valuable natural product. A new method was developed to extract and separate Antarctic krill PC. The PC was extracted only by acetone from Antarctic krill oil and separated by silica gel column chromatography mixtures of chloroform and methanol as eluting solvents. Thin-layer chromatography, high-performance liquid chromatography, and Fourier-transform infrared spectroscopy analyses were used to characterize the acetone-soluble Antarctic krill PC. The purity of the purified PC was able to reach 97.77 ± 0.65 %, which provided a reference basis for the preparation of standard Antarctic krill PC. Moreover, it was found in the fatty acid analysis that the purified PC contained mainly five fatty acids, including palmitic acid (41.25 %), oleic acid (8.62 %), 7-hexadecenoic acid (4.60 %), EPA (31.34 %), and DHA (14.52 %). The result is helpful for the analysis of Antarctic krill PC’s molecular structures and the evaluation of its nutritional value.     

基于脂肪酸特征指标的鳕鱼肝油掺假鉴定

通过分析鳕鱼肝油中各脂肪酸的相对含量及特征指标变化规律,建立鳕鱼肝油掺假鱼油的鉴定方法。鳕鱼肝油中所含脂肪酸经柱前衍生转化为甲酯后进行气相色谱测定,采用面积归一化法计算各脂肪酸相对总脂肪酸的相对含量。根据多样本脂肪酸相对含量分析结果,找出能有效识别掺假的特征性指标二十二碳六烯酸（docosahexaenoic acid,DHA）/二十碳五烯酸（eicosapentaenoic acid,EPA）值和鲸蜡烯酸相对含量,并确定其限度范围,DHA/EPA值为1.40～1.68,鲸蜡烯酸相对含量为7.0%～9.4%。当鳕鱼肝油样品中测得的DHA/EPA值和（或）鲸蜡烯酸相对含量超过此限度时,可初步判定该样品掺假鱼油。为进一步有效识别掺假水平,本研究探索建立一种以DHA/EPA值和鲸蜡烯酸相对含量为坐标的掺假模型,并绘制不同掺假水平的识别分析图,46 批样品经鉴定,有2 批样品疑似掺假25%的鱼油。该鉴定方法准确可靠、直观简便,可用于鳕鱼肝油中掺假鱼油的快速鉴定。     

Analysis of polychlorinated biphenyls (PCBs), heavy metals and omega‐3 fatty acids in commercially available Korean functional fish oil supplements

In this study, the levels of polychlorinated biphenyls (PCBs), heavy metals and omega‐3 polyunsaturated fatty acids (docosahexaenoic acid; DHA and eicosapentaenoic acid; EPA), as well as chemical indices including acid and peroxide values were determined among twenty‐four commercially available functional fish oil supplements. The sum of indicator PCB congeners (congeners 28, 52, 101, 118, 138, 153 and 180) was below the limit of quantification of 0.043 mg kg^?1. Metallic compounds (Cd, Pb, Hg and As) were not detected in all fish oil supplement samples. The relative percentages of EPA and DHA in fish oil supplements ranged from 25% to 80%. The acid values ranged from 0.1 to 1.4 mg KOH per g oil, and peroxide values ranged from 2.7 to 28.8 meq per kg oil. All fish oil supplement samples tested in current study met the safety standard for fish oil supplements set by Korean government as well as International standard.     

Optimization of omega-3 enriched-diacylglycerol production by enzymatic esterification using a response surface methodology

Incorporation of an eicosapentaenoic acid/docosahexaenoic acid (EPA/DHA) moiety into diacylglycerol (DAG) oil using lipase-catalyzed esterification was optimized using an ethyl ester form of EPA/DHA. A response surface methodology (RSM) was used to optimize reaction parameters (time, temperature, and substrate mole ratio) for incorporation of DHA and EPA into DAG oil. Predictive models for DHA+EPA contents of DAG and the amount of DAG produced after esterification were adequate and reproducible. DHA+EPA contents of DAG significantly increased with reaction time and substrate mole ratio (p<0.05). In contrast, the reaction temperature negatively affected the amount of DAG after esterification. Synthesis of DHA+EPA-enriched DAG was optimized for a maximum DAG content with the highest DHA+EPA content, in which 630.0 mg of DAG containing 34.8% DHA and EPA was predicted using the RSM model. The optimal reaction conditions were predicted at 20.6 h, 57.9 and a DHA/EPAenriched ethyl ester: DAG oil ratio of 2.5:1.     

n-3 Dietary supplementation and lipid metabolism: Differences between vegetable- and fish-derived oils

The effect of flaxseed oil rich in linolenic acid (ALA), and a mixed oil (flaxseed oil and fish oil) rich in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on the lipid clearance and peroxisome proliferator activated receptors (PPARs) in liver and adipose tissue of rats fed for 30 days with the two oils was evaluated. The results showed that after treatment with the mixed oil the hematic triacylglycerol content was significantly decreased compared to control animals. Regarding the tissue distribution of the major omega-3 fatty acids, both oils were able to increase ALA, EPA, docosapentaenoic acid (DPA) in liver and adipose tissue; and DHA solely in the adipose tissue. Finally the treatment with either flaxseed or mixed oil increased hepatic PPAR-γ expression but only the mixed oil enhanced the hepatic expression of PPAR-α. No effect on adipose tissue PPAR-γ expression was observed with both oils’ treatment.     

尿素包合法富集鱼油中的EPA和DHA的研究

用尿素包合法（脲包法）对大连鱼油中的ＥＰＡ和ＤＨＡ进行富集，得到较佳操作条件为温度０℃，脲酯比１３，鱼油甲酯为脲包客体，以甲醇作溶剂。一次富集产品中（ＥＰＡ＋ＤＨＡ）浓度和收率分别达到６０％和９０％以上