HPLC-ELSD与GC-MS法测定牛乳甘油三酯sn-2位脂肪酸组成

研究建立了快速、准确测定牛乳脂肪甘油三酯sn-2位脂肪酸组成的方法．利用胰脂酶专一水解甘油三酯sn-1和sn-3位置上的脂肪酸得到sn-2单甘油酯和游离脂肪酸,再通过蒸发光散射高效液相色谱分离出sn-2位单甘油酯,然后对其进行衍生,用气相色谱质谱联用仪对sn-2脂肪酸组成进行分析．结果显示用蒸发光散射高效液相色谱法分离sn-2位单甘油酯时方法的回收率达到83．3％～85．1％,该法省去了传统测定中费时费力的薄层色谱分离步骤．用气相色谱质谱联用法对产物进行分析,精密度高,结果可靠．分析结果表明,牛乳脂肪sn-2位脂肪酸由2．57％月桂酸、7．68％豆蔻酸、34．74％棕榈酸、11．56％亚油酸、22．53％油酸和15．21％硬酯酸组成．     

超高效液相色谱法测定生物柴油中的11种脂肪酸及脂肪酸甲酯

建立了采用超高效液相色谱(UPLC)-蒸发光散射检测器(ELSD)测定生物柴油中11种常见的脂肪酸及脂肪酸甲酯含量的方法。这11种常见的脂肪酸及脂肪酸甲酯为豆蔻酸、亚油酸、棕榈酸、油酸、亚麻酸甲酯、硬脂酸、亚油酸甲酯、棕榈酸甲酯、油酸甲酯、芥酸和硬脂酸甲酯。样品经提取后用甲醇溶解,采用Acquity UPLC BEH Phenyl C18柱(100 mm×2.1 mm,1.7 μm)分离,乙腈-水（体积比为3∶1)混合液为流动相进行等度洗脱,采用的ELSD条件为增益80,漂移管温度为45 ℃,载气压力为172 kPa,雾化器为冷却模式,并用外标法进行定量分析。结果表明,在一定的质量浓度范围内,峰面积的对数和质量浓度的对数线性关系良好。与其他检测生物柴油成分的方法相比,该方法简单,分离效果好,速度快,特别是此方法可以同时实现脂肪酸及脂肪酸甲酯的分离,并进行定量分析,能有效测定反应的进行程度,从而满足生物柴油工艺研究的需要。     

正相液相色谱法测定婴幼儿配方乳粉中1,3-二油酸-2-棕榈酸甘油三酯

建立并比较了正相色谱与反相色谱测定婴幼儿配方乳粉中1,3-二油酸-2-棕榈酸甘油三酯的方法。利用乙醚和石油醚提取婴幼儿配方乳粉中的油脂,经旋转蒸发浓缩后,分别采用银离子色谱柱和C18色谱柱,配合蒸发光散射检测器检测。其中采用银离子色谱柱以二氯甲烷-丙酮为流动相梯度洗脱的正相色谱法完全分离了1,3-二油酸-2-棕榈酸甘油三酯(OPO)和1,2-二油酸-3-棕榈酸甘油三酯(OOP)两种同分异构体,方法的重复性和线性关系良好,回收率为93.0%~101.5%,精密度(RSD)为2.89%~4.56%,检出限为20 mg/kg。该方法可准确测定婴幼儿配方乳粉中OPO的含量。     

痕量动物油和植物油的区分检验研究

采用碱催化甲酯化的前处理方法,应用气相色谱-质谱联用技术对动物油和植物油的区分检验进行了研究。实验检出了6种主要脂肪酸:肉豆蔻酸、棕榈油酸、棕榈酸、亚油酸、油酸、硬脂酸。分析了动物油与植物油在成分及相对含量上的差异,并且找出了动物油和植物油在不饱和度上的差别。根据动物油与植物油的差异点,可以对动物油和植物油进行区分。     

GC／MS法分析鳗鱼骨油的脂肪酸

用乙醚／已烷混合溶剂抽提鳗鱼骨油，用ＫＯＨ－ＣＨ_３ＯＨ酯交换法甲酯化，以毛细管色谱／质谱方法测定脂肪酸的组成。共检出２７种脂肪酸，其中以棕榈油酸、棕榈酸、油酸、硬脂酸、花生烯酸为主。     

扁桃油中脂肪酸组成的GC-MS法分析

以正己烷为提取剂,采用超声法提取油脂,经KOH-甲醇甲酯化处理后,以气相色谱-质谱(GC-MS)联用技术鉴定出陕西蒲城3个品种扁桃油中的脂肪酸主要组成为:油酸、亚油酸、9-十六碳烯酸、硬脂酸、棕榈酸,其中不饱和脂肪酸总量占91.4%以上,主要成分油酸占73.3%以上。     

柱前衍生HPLC-MS法测定黑果枸杞果实中脂肪酸

采用柱前衍生荧光检测高效液相色谱-大气压化学电离源/质谱联用技术对黑果枸杞果实中的脂肪酸成分进行分析。经2-(11-H-苯-a-咔唑)乙基对甲苯磺酸酯荧光衍生试剂对脂肪酸进行柱前衍生,梯度洗脱分离,荧光检测,外标法定量和在线质谱鉴定,建立了同时测定20种脂肪酸含量的方法,并运用此方法对黑果枸杞果实中的脂肪酸进行了分析。结果表明。检测限可以达到0.42~1.84 ng/mL。黑果枸杞果实中主要含有不饱和脂肪酸,其含量约占整个脂肪酸总量的64%,其中不饱和脂肪酸种类主要是C18∶1(油酸),C18∶2(亚油酸)和少量的C18∶3(亚麻酸);饱和脂肪酸主要是以C16(棕榈酸),C18(硬脂酸),C20(花生酸)和C22(山嵛酸)为主,另外还含有少量的C12(月桂酸),C14(肉豆蔻酸),C17(珠光脂酸),C23(二十三酸)和C24(木蜡酸)。本研究可以为黑果枸杞在食品、医药和保健方面的进一步开发应用提供可靠的科学依据。     

甲鱼油脂肪酸组成的研究

用热提取方法抽取出甲鱼油，用ＫＯＨ－ＣＨ３ＯＨ酯交换法甲酯化处理，以色谱／质谱方法测定脂肪酸的组成。共鉴定出２１种脂肪酸，其中以棕榈酸、棕榈油酸、硬脂酸、油酸、亚油酸、二十碳五烯酸（ＥＰＡ）、二十二碳五烯酸（ＤＨＡ）为主     

气相色谱测定牦牛骨中脂肪酸的甲酯化方法比较

利用气相色谱(GC)技术,采用酸水解提取脂质,比较了6种甲酯化法(乙酰氯-甲醇法、H2SO4-甲醇法、HCl-甲醇法、KOH-甲醇法、KOH-甲醇+H2SO4-甲醇法和KOH-甲醇+HCl-甲醇法)对脂肪酸测定的影响,优选牦牛骨中脂肪酸测定的最佳方法。37种脂肪酸标准样品在0.28~250.00 mg/L范围内线性关系良好,相关系数均大于0.99(除C4∶0外)。碱酯化法和酸碱结合法几乎无法测出牦牛骨中的脂肪酸,其测得的总脂肪酸含量小于0.20 g/100g。乙酰氯-甲醇法测得的总脂肪酸含量(13.61 g/100 g)显著高于H2SO4-甲醇法(总脂肪酸含量为11.68 g/100 g)和HCl-甲醇法(总脂肪酸含量为3.18 g/100 g)测得的结果。乙酰氯-甲醇法和H2SO4-甲醇法的日内和日间精密度分别为0.27%~8.60%和0.34%~2.64%,两种方法中脂肪酸的回收率为83.06%~105.54%。结果表明,酸水解-乙酰氯-甲醇法是牦牛骨中脂肪酸测定的最佳方法。C18∶1n9c、C16∶0、C18∶0和共轭亚油酸(CLA)是牦牛骨的主要脂肪酸,其总和达脂肪酸总量的85%以上,饱和脂肪酸与不饱和脂肪酸含量比值约为1∶2。牦牛骨中脂肪酸的研究为骨资源脂质的有效利用提供了重要依据。     

α-吡咯烷酮乙酸甲酯的H P LC测定

建立了测定α -吡咯烷酮乙酸甲酯的反相高效液相色谱法。采用的流动相为甲醇 -水 -四氢呋喃 (体积比30∶68∶2) ,流动相pH3.0,柱温45℃ ,流量梯度洗脱,检测波长208nm。在该条件下α -吡咯烷酮乙酸甲酯与相关物质的色谱峰分离完全,测定α -吡咯烷酮乙酸甲酯 ,含量为90 %～94 %时SD为0.59 %～0.72 % ,RSD为0.65 %～0.77 % ,与化学法相比 ,测定结果一致。     

Lipids of the fruit of Ficus carica

By chromatographic methods, about 30 groups of various lipid compounds belonging to the classes of neutral lipids, glycolipids, and phospholipids have been identified from the fruit of the fig tree. The main groups are triacylglycerols, free and esterified sterols, mono- and digalactosyldiglycerides, ceramide oligosides, cerebrosides, esterified sterol glycosides, and phosphatidylglycerols. In the fatty acid composition, linoleic, linolenic, oleic, and palmitic acids predominated (>90%).     

Production of Human Milk Fat Replacement Rich of 1,3-dioleoyl-2-palmitoilglycerol From Enzymatic Interesterification Tripalmitin,Ethyl Oleate And Mixture of VCO,Soybean Oil And Fish Oil

Human Milk is naturally the only source of food for infant in their early life. It contains 2-6% lipid which provides about 50% of the total energy needed by the infant. Human milk fat (HMF) mainly as TAG with the specific fatty acid composition, palmitic acid (C16:0) (20-25%), which is primary located at sn-2 of glycerol bonds (70%) and oleic acid (C18:1), located at sn-1,3 (35%). HMF also provide fatty acids such as linoleic acid, linolenic acid, EPA, DHA and lauric acid that are very important for infant. The purposes of this research are to synthesize of 1,3-dioleoyl-2-palmitoilglycerol (OPO) and to determine the best composition of OPO, VCO, soybean oil and fish oil for HMFS production for infant formula. Interesterification of tripalmitin and ethyl oleate using immobilized lipase from Rhizomucor miehei (Lipozym RM IM) were used to synthesize of OPO. Interesterification product of mixed VCO, soybean oil and fish oil that are source of lauric acid, linoleic acid, α-linolenic, EPA and DHA, were formulated in mass ratio (58:20:20:2) and (70:18:10:2) for obtaining HMFS which have fatty acids composition similar or close to HMF. Composition of fatty acids from product were analyzed by GCMS. From this research, were obtained HMFS containing palmitic acid as much as 28.89% where 84.49% of that are located at sn-2 while sn-1,3 position are dominated by oleic acid as much as 55.11% from the total 38.7% and 70:18:10:2 w/w is the best composition of interesterification product, VCO, soybean oil and fish oil to obtain HMFS similar to HMF.     

Quick regiospecific analysis of fatty acids in triacylglycerols with GC using 1,3-specific lipase in butanol

A new micro-procedure for the quick regiospecific analysis of triacylglycerols (TAG) with a 1,3-specific lipase, Lipozyme IM 20 (produced from strains of Muccor miehei) is described. After dissolution of triacylglycerols in butan-1-ol or butan-2-ol 250 mg of Lipozyme IM 20 were added and the whole mixture was agitated. This operation was repeated 5-10 times and the esterified fatty acids in the form of butyl esters (FABE) were dissolved in pentane and washed with water. The pentane layer was dried over sodium sulfate and the solvent was removed under vacuum to a constant weight. The residue was analysed using GC-MS. The same triacylglycerols were converted to fatty acid methyl esters (FAME). The FAME were analysed by gas chromatography (GC).     

Lipids of the fruit of Ficus carica

By chromatographic methods, about 30 groups of various lipid compounds belonging to the classes of neutral lipids, glycolipids, and phospholipids have been identified from the fruit of the fig tree. The main groups are triacylglycerols, free and esterified sterols, mono- and digalactosyldiglycerides, ceramide oligosides, cerebrosides, esterified sterol glycosides, and phosphatidylglycerols. In the fatty acid composition, linoleic, linolenic, oleic, and palmitic acids predominated (>90%).A. V. Bogatskii Physicochemical Institute, Academy of Sciences of the Ukrainian SSR, Odessa. Lomonosov Technological Institute of the Food Industry, Odessa. Translated from Khimiya Prirodnykh Soedinenii, No. 4, pp. 423–427, July–August, 1986.     

Production of biodiesel by immobilized Candida sp. lipase at high water content

A new process for enzymatic synthesis of biodiesel at high water content (10–20%) with 96% conversion by lipase from Candida sp. 99–125 was studied. The lipase, a no-position-specific lipase, was immobilized by a cheap cotton membrane and the membrane-immobilized lipase could be used at least six times with high conversion. The immobilized lipase could be used for different oil conversion and preferred unsaturated fatty acids such as oleic acid to staturated fatty acids such as palmitic acid. The changes in concentration of fatty acids, diglycerides, and methyl esters in the reaction were studied and a mechanism of synthesis of biodiesel was suggested: the triglycerides are first enzymatically hydrolyzed into fatty acids, and then these fatty acids are further converted into methyl esters.     

Comparative quantitative fatty acid analysis of triacylglycerols using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and gas chromatography.

Quantitative analyses of fatty acids from five triacylglycerol products, coconut oil, palm kernel oil, palm oil, lard and cocoa butter, were carried out using two analytical methods: matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) and gas chromatography (GC), in an effort to validate the application of MALDI-TOFMS in quantitative fatty acid analysis. For the GC analysis, transmethylated products were used, whereas, for the MALDI-TOF analysis, saponified products were used. Under MALDI-TOF conditions, the acids were detected as sodiated sodium carboxylates [RCOONa + Na](+) consistent with the mode of ionization that was previously reported. Thus, the MALDI-TOF mass spectrum of saponified coconut oil showed the presence of sodiated sodium salts of caprylic acid (7.5 +/- 0.67, m/z 189), capric acid (6.9 +/- 0.83, m/z 217), lauric acid (47.8 +/- 0.67, m/z 245), myristic acid (20.4 +/- 0.51, m/z 273), palmitic acid (9.8 +/- 0.47, m/z 301), linoleic acid (0.9 +/- 0.07, m/z 325), oleic acid (4.8 +/- 0.42, m/z 327) and stearic acid (2.0 +/- 0.13, m/z 329). Saponified palm kernel oil had a fatty acid profile that included caprylic acid (3.5 +/- 0.59), capric acid (4.7 +/- 0.82), lauric acid (58.6 +/- 2.3), myristic acid (20.9 +/- 1.5), palmitic acid (7.2 +/- 1.1), oleic acid (3.8 +/- 0.62) and stearic acid (1.2 +/- 0.15). Saponified palm oil gave myristic acid (0.83 +/- 0.18), palmitic acid (55.8 +/- 1.7), linoleic acid (4.2 +/- 0.51), oleic acid (34.5 +/- 1.5), stearic acid (3.8 +/- 0.26) and arachidic acid (0.80 +/- 0.22). Saponified lard showed the presence of myristic acid (1.5 +/- 0.24), palmitic acid (28.9 +/- 1.3), linoleic acid (13.7 +/- 0.67), oleic acid (38.7 +/- 1.4), stearic acid (12.8 +/- 0.64) and arachidic acid (2.4 +/- 0.35). Finally, for saponified cocoa butter, the fatty acid distribution was: palmitic acid (32.3 +/- 1.0), linoleic acid (2.6 +/- 0.35), oleic acid (34.9 +/- 1.7) and stearic acid (30.3 +/- 1.6). Quantitative gas chromatographic analysis of the corresponding methyl esters from these triacylglycerol products yielded data that were mostly in agreement with the MALDI-TOFMS data. The MALDI-TOF experiment, however, proved to be superior to the GC experiment, particularly with regard to baseline resolution of unsaturated acids. Furthermore, the ability of MALDI-TOFMS to detect low concentrations of fatty acids rendered it more sensitive than the GC methodology.     

Analytical monitoring of the production of biodiesel by high-performance liquid chromatography with various detection methods.

Gradient elution reversed-phase high-performance liquid chromatography (RP-HPLC) was used for the determination of compounds occurring during the production of biodiesel from rapeseed oil. Individual triacylglycerols (TGs), diacylglycerols, monoacylglycerols and methyl esters of oleic, linoleic and linolenic acids and free fatty acids were separated in 25 min using a combined linear gradient with aqueous-organic and non-aqueous mobile phase steps: 70% acetonitrile+30% water in 0 min, 100% acetonitrile in 10 min, 50% acetonitrile+50% 2-propanol-hexane (5:4, v/v) in 20 min and 5 min final hold-up. Another method with a non-aqueous linear mobile phase gradient [from 100% methanol to 50% methanol+50% 2-propanol-hexane (5:4, v/v) in 15 min] was used for fast monitoring of conversion of rapeseed oil triacylglycerols to fatty acid methyl esters and for quantitation of residual TGs in the final biodiesel product. Sensitivity and linearity of various detection modes (UV detection at 205 nm, evaporative light scattering detection and mass spectrometric detection) were compared. The individual sample compounds were identified using coupled HPLC-atmospheric pressure chemical ionization mass spectrometry in the positive-ion mode.     

Triacylglycerol lipase from rape (Brassica napus L.) suitable for biotechnological purposes

Triacylglycerol lipase (EC 3.1.1.3) from rape (Brassica napus L. cv Ceres) is quite easily prepared from the 100,000 x g supernatant of cotyledon homogenates. The lipase is present in a high-molecular-mass fraction ( > 1.5 x 10⁶ dalton by gel filtration), but it can be rapidly extracted from the 100,000 xg supernatant by precipitation with polyethyleneglycol 8000 (4%, w/v) and MgCl₂ (40 mM) giving about a 10-fold purification. After delipidation, the lipase has an M_r of about 300,000. It hydrolyzes triacylglycerols to fatty acids and glycerol, although the fatty acids from the sn-1 or -3 positions are hydrolyzed first to yield 1,2(2,3)-diaclyglycerols. Lipase immobilized onto Celite by precipitation with acetone at-20°C catalyzes the esterification of oleic acid with butanol dissolved in hexane.     

Enzymatic Synthesizing of Phytosterol Oleic Esters

A method of synthesizing the phytosterol esters from oleic acid and sterols was studied, using immobilized lipase Candida sp. 99-125 as catalyst. Molar ratio (oleic acid/phytosterols), temperature, reaction period, organic solvents, catalyst, and silica-gel drier were optimized, and the result showed that 93.4% of the sterols had been esterified under the optimal synthetic condition: the molar ratio of oleic acid/phytosterol is 1:1 in 10 mL iso-octane, immobilized lipase (w, 140% of the sterols), incubated in an orbital shaker (200 rpm) at a temperature of 45 °C for 24 h. The immobilized lipase could be reused for at least 13 times with limited loss of esterification activity. The conversion still maintained up to 86.6%. Hence, this developed process for synthesizing phytosterol esters could be considered as simple and low-energy consumption compared to existing chemical processes.     

Analysis of triacylglycerols with non-aqueous reversed-phase liquid chromatography and positive ion electrospray tandem mass spectrometry

A non-aqueous reversed-phase liquid chromatographic method coupled to electrospray ionisation (ESI) tandem mass spectrometry was developed for the analysis of triacylglycerols (TGs). The synthetic TGs studied were separated according to their equivalent carbon number with a gradient of methanol (containing 0.01% (v/v) formate adjusted to pH 5.3 with ammonia) and chloroform. ESI mass spectra of TGs yielded positive ion current signals for [M + NH(4)](+) and [M + NH(4)-RCOONH(4)](+). The mass spectra also showed signals believed to arise from [M + K](+). Collision-induced dissociation (CID) of the [M + NH(4)](+) precursor ion yielded [M + NH(4) - RCOONH(4)](+), [RCO + 74](+) and [RCO](+) product ions as aids for the structural elucidation of the TGs. In addition, [RCO - 18](+) and small amounts of [RCO - 2](+) product ions were also found. The latter ions were observed only for TGs containing unsaturated fatty acids. CID of ammoniated 1-stearoyl-2-oleoyl-3-linoleoyl-glycerol (18:0/18:1/18:2) indicated that neutral loss of the sn-2 fatty acid was energetically less favourable than loss of the fatty acid from the sn-1 or sn-3 position.