Identification of TAG and DAG and their FA Constituents in Lesquerella (Physaria fendleri) Oil by HPLC and MS

Castor oil has many industrial uses because of its high content (90 %) of the hydroxy fatty acid, ricinoleic acid (OH¹²18:1⁹). Lesquerella oil containing lesquerolic acid (Ls, OH¹⁴20:1¹¹) is potentially useful in industry. Ten molecular species of diacylglycerols and 74 molecular species of triacylglycerols in lesquerella (Physaria fendleri) oil were identified by electrospray ionization mass spectrometry as lithium adducts of acylglycerols in the HPLC fractions of lesquerella oil. Among them were: LsLsO, LsLsLn, LsLsL, LsLn–OH20:2, LsO–OH20:2 and LsL–OH20:2. The structures of the four new hydroxy fatty acid constituents of acylglycerols were proposed by the MS of the lithium adducts of fatty acids as (comparing to those in castor oil): OH¹²18:2^9,14 (OH¹²18:2^9,13 in castor oil), OH¹²18:3^9,14,16 (OH18:3 not detected in castor oil), diOH^12,1318:2^9,14 (diOH^11,1218:2^9,13 in castor oil) and diOH^13,1420:1¹¹ (diOH20:1 not detected in castor oil, diOH^11,1218:1⁹ in castor oil). Trihydroxy fatty acids were not detected in lesquerella oil. The differences in the structures of these C18 hydroxy fatty acids between lesquerella and castor oils indicated that the polyhydroxy fatty acids were biosynthesized and were not the result of autoxidation products.     

Ratios of Regioisomers of the Molecular Species of Triacylglycerols in Lesquerella (Physaria fendleri) Oil Estimated by Mass Spectrometry

The ratios of regioisomers of 72 molecular species of triacylglycerols (TAG) in lesquerella oil were estimated using the electrospray ionization mass spectrometry of the lithium adducts of TAG in the HPLC fractions of lesquerella oil. The ratios of ion signal intensities (or relative abundances) of the fragment ions from the neutral losses of fatty acids (FA) as α‐lactones at the sn‐2 position (MS³) of the molecular species of TAG were used as the ratios of the regioisomers. The order of the preference of FA incorporation at the sn‐2 position of the molecular species of TAG in lesquerella was as: normal FA > OH18 (monohydroxy FA with 18 carbon atoms) > diOH18 > OH20 > diOH20, while in castor was as: normal FA > OH18 > OH20 > diOH18 > triOH18. Elongation (from C₁₈ to C₂₀) was more effective than hydroxylation in lesquerella to incorporate hydroxy FA at the sn‐1/3 positions. The block of elongation in lesquerella may be used to increase the content of hydroxy FA, e.g., ricinoleate, at the sn‐2 position of TAG and to produce triricinolein (or castor oil) for industrial uses. The content of normal FA at the sn‐2 position was about 95 %, mainly oleate (38 %), linolenate (31 %) and linoleate (23 %). This high normal FA content (95 %) at the sn‐2 position was a big space for the replacement of ricinoleate to increase the hydroxy FA content in lesquerella oil. The content of hydroxy FA at the sn‐1/3 positions was 91 % mainly lesquerolic acid (85 %) and the content of normal FA was 6.7 % at the sn‐1/3 position in lesquerella oil.     

Identification of Tetraacylglycerols in Lesquerella Oil by Electrospray Ionization Mass Spectrometry of the Lithium Adducts

Tetraacylglycerol (an acylglycerol estolide) contains an acyl chain attached to the hydroxyl group of another acyl chain attached to the glycerol backbone. Lesquerolic acid (Ls, OH¹⁴20:1¹¹) is the main fatty acid in lesquerella oil and may be used industrially for the manufacture of biodegradable industrial products. Electrospray ionization mass spectrometry of the lithium adducts of acylglycerols in the high-performance liquid chromatography fractions from the seed oil of lesquerella (Physaria fendleri) was used to identify thirteen tetraacylglycerols. They were LsLsLsLn, LsLsLsL, LsLs-OH20:2-O, LsLsLsO, LsLsLnLn, LsLsLLn, LsLsOLn, LsLsLL, LsLsOL, LsLsOP, LsLsOO, LsLsLS and LsLsOS. The OH20:2 was auricolic acid (OH¹⁴20:2^11,17). For the four tetraacylglycerols containing one normal fatty acid (non-hydroxy fatty acid), LsLsLsLn, LsLsLsL, LsLs-OH20:2-O and LsLsLsO, the normal fatty acids were all directly attached to the glycerol backbone, not to the hydroxyl group of fatty acids. We propose that the biosynthetic precursors (triacylglycerol acyltransferase) of these four tetraacylglycerols were LsLsLn, LsLsL, LsLsO (Ls-OH20:2-O) and LsLsO individually. LsLsO and Ls-OH20:2-O were equally active as the biosynthetic precursors for LsLs-OH20:2-O. For LsLsLS, linoleate were all attached to the glycerol backbone and LsLsL was proposed to be the biosynthetic precursor. For LsLsOS, stearate were all attached to the glycerol backbone and LsLsS was proposed to be the biosynthetic precursor. For the other seven tetraacylglycerols containing two normal fatty acids, LsLsAB, the biosynthetic precursors could be both LsLsA and LsLsB.     

Evaluation of MALDI-TOF/MS Technology in Olive Oil Adulteration

Adulteration of extra virgin olive oil (EVOO) by addition of other vegetable oils or lower-grade olive oils is a common problem of the oil market worldwide. Therefore, we developed a fast protocol for detection of EVOO adulteration by mass spectrometry fingerprinting of triacylglycerol (TAG) profiles based on MALDI-TOF/MS. For that purpose, EVOO TAG profiles were compared with those of edible sunflower oil and olive oil composed of refined olive oil and virgin olive oils. Adulteration of EVOO was simulated by addition of sunflower and mixture of refined olive oil and virgin olive oils at 1, 10 and 20% w/w. Results of mass spectrometry TAG profiling were compared with routinely assessed K values for identification of adulteration. MALDI-TOF/MS technology coupled with statistical analysis was proven as useful for detection of adulteration in EVOO at a rate down to 1%. In contrast, standard spectrophotometric methods failed to identify minor adulterations. In addition, the ability of MALDI-TOF/MS in detection of adulteration was tested on EVOO samples from different geographical regions. Results demonstrated that MALDI-TOF/MS technology coupled with statistical analysis is able to distinguish adulterated oils from other EVOO.     

HPLC/MS测定啶虫脒在蔬菜、水果中残留量

建立了用HPLC/MS测定蔬菜、水果中啶虫脒残留量的方法。试样用酸性乙腈提取,C18柱净化,HPLC/MS测定。方法的最低检测限为0.001mg/kg,添加回收率的范围为86.4%～102.4%,相对标准偏差为4.16%～6.02%。     

Evaluation of the Triglyceride Composition of Pomegranate Seed Oil by RP‐HPLC Followed by GC‐MS

Triglyceride composition and fatty acid profiles of pomegranate seed oil were evaluated by newly developed methods in reverse‐phase‐high performance liquid chromatography (RP‐HPLC) and gas chromatography (GC), respectively. Different compositions of the mobile phase (acetone and acetonitrile) and flow rates for the HPLC system were used to obtain better separation for accurate quantitative analysis. Triglycerides with conjugated fatty acids (CLnAs) were eluted in order of the polarity of their geometrical isomers (c, t, c < t, t, c < t, t, t). The dominant triglyceride was found to be PuPuPu (32.99 %) in pomegranate seed oil, followed by PuPuCa and PuCaCa containing punicic acid and catalpic acid with total triglyceridelevels of 27.72 and 10.11 %, respectively. For fatty acid composition analysis, triglyceride fractions were derivatized into their respective methylesters which were injected into gas chromatography‐mass spectrometry (GC‐MS) to identify and gas chromatography‐flame ionization detector (GC‐FID) to quantify the conjugated fatty acids of each fraction of triglycerides. Punicic acid was found to be dominant (76.57 %) followed by catalpic acid (6.47 %) and β‐eleotearic acid (1.45 %). Pomegranate seed contained greater amounts of conjugated linolenic acids. These results showed that the present study provides more information about the composition of the triglyceride and fatty acid profiles of pomegranate seed oil compared to the reported studies. Therefore, the developed methods in this study can be used for the identification of the triglyceride and fatty acid composition for pomegranate seed oils and some such specials edible oils including CLnA isomers.     

茶树油的分子蒸馏精制及其GC/MS分析

利用新型、特殊分离技术——分子蒸馏对茶树油进行精制,并对精制后的茶树油进行GC/MS定性分析及其总离子流图的面积归一法定量分析。结果从粗茶树油中分离出了61种化合物,鉴定出36种组分,占总量的98.67%,其中4-松油醇高达49.56%,并且精制的茶树油符合国际标准(ISO 4730-1996)。     

生姜挥发油的GC/MS分析

采用气相色谱－质谱联用技术,对广西百色生姜挥发油进行了定性、定量分析,共分离２２种成分,鉴定了其中的１９种成分结构,其含量占挥发油总量的９７．２９％     

高效液相色谱法测定食用植物油中的游离棉酚

研究采用高效液相色谱法测定食用植物油中游离棉酚的方法,最佳色谱条件:色谱柱C18柱,流动相为甲醇-磷酸溶液(1%)=84+16,波长240 nm,流速为1.0 mL/min,柱温30℃。曲线方程y=3.1033×10-5x,相关系数r=0.999871,线性关系良好。检出限为2.5 mg/kg,回收率为98.0%～100.2%。该法简便,快速,准确性高,重现性好。     

Antioxidant Distributions and Triacylglycerol Molecular Species of Sesame Seeds (Sesamum indicum)

Extracted lipids from sesame (Sesamum indicum) seeds of three varieties were determined by high-performance liquid chromatography (HPLC) for endogenous antioxidants. The molecular species and fatty acid (FA) distribution of triacylglycerol (TAG) isolated from total lipids in sesame seeds were analyzed by a combination of argentation thin-layer chromatography (TLC) and gas chromatography (GC), and were investigated in relation to their antioxidant distribution. γ-Tocopherol was present in highest concentration, and δ-, and α-tocopherols were very small amounts. Sesamin and sesamolin were the main lignan components. A modified argentation-TLC procedure, developed to optimize the separation of the complex mixture of total TAG, provided 12 different groups of TAG, based on both the degree of unsaturation and the total acyl-chain length of FA groups. With a few exceptions, the major TAG components were SM₂ (6.5–6.7%), SMD (19.8–20.7%), M₂D (15.0–26.3%), MD₂ (23.6–35.0%), and D₃ (7.7–10.7%) (where S denotes a saturated FA, M denotes a monoene, D denotes a diene, and T denotes a triene). It seems that the three varieties were highly related to each other based on the FA composition of the TAG as well as the distribution pattern in the different TAG molecular species. These results suggest that there are no essential differences in the oil components among the three varieties.     

花椒挥发油化学成分的GC/MS分析

文章采用常规水蒸气蒸馏法提取花椒的挥发油,经气相色谱-质谱技术分离和鉴定,用归一化法测定其相对百分含量,可得花椒挥发油的主要成分为β-月桂烯（1.94192％）、柠檬烯（12.25932％）、沉香醇（14.96057％）,α,α,4-三甲基-3-环己烯-1-甲醇（13.3886％）,2-氨基苯甲酸-3,7-二甲基-1,6-辛二烯-3-醇酯（20.0481％）,乙酸松油酯（5.8074％）.     

白兰花油和白兰叶油的GC／MS成分分析及在香精中的应用研究

运用气相色谱／质谱联用法结合计算机谱图检索对白兰花油和白兰叶油进行了成分分析和鉴定,分别鉴定出83和72种化合物,其中相同的化合物有62种,得到鉴定的化合物分别占挥发油总量的96.07％和98.08％。比较分析白兰花油和白兰叶油在成分上的区别,并对由此导致的香气差异进行了阐释。对两种精油在日用香精中的应用进行了研究,比较在同一配方中的应用区别,对它们在香精中的应用提出一些见解。     

Quantification of Phorbol Esters in Jatropha curcas by HPLC‐UV and HPLC‐ToF‐MS with Standard Addition Method

Established analytic methods for the quantification of phorbol esters (PE), which are some toxic components in Jatropha curcas L., include HPLC with UV‐detection with the commercially available phorbol myristate acetate (PMA) as internal standard or HPLC coupled with MS detection with an external calibration, mostly also with PMA. The differences in the fatty acid side chains and connection to the base structure of PMA compared to PE leads to different UV absorption and MS ionization effects and cause problems for exact quantitative measurements. In this paper, a method is presented which combines both detection types and shows differences between both results. For this purpose, an extraction routine is performed on a PE‐containing seed oil to get a PE standard in high purity, which was used for a standard addition method on two real J. curcas oil samples, derived from Ghana and Mexico. Furthermore, a detection window of ±10 ppm for the high accurate ToF‐MS detection is set to eliminate isobaric interferences from co‐eluting material. Method evaluation of inter‐ and intra‐day variance as well as the recovery rate are performed and determined. With this method a limit of detection of 62 ng mL^?1 (UV) and 11 ng mL^?1 (MS) can be achieved. Practical Applications: Due to the good biological and technical properties of Jatropha curcas L., its seed oil seems perfect for the application as biodiesel feedstock. The toxicity on the other hand could cause problems when converting side products from the oil production to products of higher value. With the here described method an accurate and precise analysis procedure for the quantification of the toxic compounds namely, phorbol esters, could be applied for toxicity studies or routine checks in industry which is converting plant material from J. curcas, so that no toxic material is used for example as animal feed. In this paper, an exact and robust analysis method is described for the quantification of phorbol esters (PE) in Jatropha curcas L. seed oil. This method procedure includes the extraction of PE in methanol, chromatographic separation on a reverse phase C18 HPLC column and the quantification by standard addition method. For the standard addition method a highly pure PE standard is used, which is extracted and purified by semi preparative HPLC right before the measurements. The used detector for identification and quantification is UV set at 280 nm and ESI‐ToF‐MS with a ±10 ppm mass difference of the deprotonated and formate adduct pseudo molecular ion of PE.

     

Analysis of triacylglycerols in refined edible oils by isocratic HPLC‐ESI‐MS

A simple, fast and reproducible reversed‐phase high performance liquid chromatography (HPLC) method coupled to electrospray ionization mass spectrometry (ESI‐MS) for the analysis of triacylglycerols (TAGs) species in the commercial edible oils has been developed. The TAGs species were separated using isocratic 18% isopropanol in methanol and a Phenomenex C18 column. The ESI‐MS conditions were optimized using flow injection analysis of standard TAG. Fifteen, fourteen, and sixteen TAGs were separated and identified in corn oil, rapeseed oil, and sunflower oil, respectively. The presence of intense protonated molecular (M + H⁺), ammonium (M + ${\rm NH}_{4}^{ + } $ ), and sodium (M + Na⁺) adducts ions and their respective diacylglycerols ions in the ESI‐MS spectra showed correct identification of TAGs. Some minor potassium adducts (M + K⁺) were also found. In addition, the identity of the fatty acid, position of each fatty acid, and the location of the double bond in the fatty acid moiety were explained. It was found that this isocratic method is useful for fast screening and identification of triacylglycerols in lipids.     

UPLC MS/MS Quantification of Primary Metabolites of Benzo[a]pyrene and Fluoranthene Produced In Vitro by Sole (Solea solea) Liver Microsomal Activation

Biotransformation pathways of PAHs in a flat fish, Solea solea, living in nurseries close to coastal areas and estuaries constitute a great challenge for the environmental risk. Among PAHs, Fluoranthene (Fluo) has a different chemical structure than benzo[a]pyrene (BaP), and can influence its biotransformation pathways and genotoxic potential. The aim of this study was to bring some response elements about the in vitro metabolic activation of Fluo and BaP by sole liver microsomes. The quantification of several primary metabolites of BaP and Fluo produced in vitro, following a sole liver microsomal activation, was conducted using a sensitive analytical UPLC MS/MS method.

Four types of BaP metabolites (dihydrodiols, diones, OH-BaP, and 7,8-dihydro-epoxide), and three types of Fluo metabolites (2,3-dihydrodiol, 2,3-dione, and sum of OH-Fluo) were quantified. The concentration for which the metabolite production was maximal was of 40 μM and 100 μM for BaP and Fluo, respectively. The biotransformation rate was higher for Fluo (9.97%) than for BaP (6.69%).Whatever the PAH, dihydrodiols and phenols were the major metabolites produced accounting for, respectively, 50% and 30–40% for BaP, and 40 and 60% for Fluo. Despite some differences in conformation, both PAHs seem to be activated by similar pathways and the diol-epoxide appears as a predominant one.     

溶剂蒸馏气相色谱-质谱法研究菜籽油挥发性成分

溶剂蒸馏气相色谱-质谱法是一种有效的检测挥发性物质的研究方法,在本研究中对菜籽油中挥发性成分进行分析：将菜籽油和乙酸乙酯（V／V=2：1）混合溶解后蒸馏出溶剂,同时将蒸馏出来的挥发性组分进行富集,然后加热并用N2吹扫蒸馏液浓缩得到待测液,最后用GC／MS分析检测待测液成分。结果检测到挥发性成分46种,通过NIST 2006标准谱库检索、Wiley数据库和文献比较确定挥发性成分28种,分别是酮类、醛类、烷烃类、烯烃类以及其他少量化合物。     

Mass Spectrometry of the Lithium Adducts of Diacylglycerols Containing Hydroxy FA in Castor Oil and Two Normal FA

Castor oil can be used in industry. The molecular species of triacylglycerols containing hydroxy fatty acids (FA) in castor oil have been identified. We report here the identification of twelve diacylglycerols (DAG) containing hydroxy FA in castor oil using positive ion electrospray ionization mass spectrometry of the lithium adducts. They were RR (diricinolein, R is ricinoleate), RL, RS, R‐diOH18:0, R‐diOH18:1, R‐diOH18:2, R‐triOH18:0, R‐triOH18:1, R‐triOH18:2, diOH18:0‐diOH18:1, diOH18:1‐diOH18:1 and diOH18:1‐diOH18:2. The MS² fragment ions, [M + Li ? FA]⁺ and [FA + Li]⁺, from the lithium adducts of DAG containing hydroxy FA (one or two hydroxy FA), were used for the identification. The additional fragment ions from the neutral losses of FA lithium salts [M + Li ? FALi]⁺ were used for the identification of eleven DAG containing two normal FA in a soybean oil bioconversion product. The MS² fragment ions from the neutral losses of FA lithium salts [M + Li ? FALi]⁺ were not detected from the DAG containing hydroxy FA. The DAG containing FA with more hydroxyl groups than the other FA on the same DAG molecule tended to have a prominent fragment ion [FA + Li]⁺ and an undetectable fragment ion [M + Li ? FA]⁺ while the FA was the more hydroxylated FA. Also the less hydroxylated FA of a DAG tended to have a prominent fragment ion [M + Li ? FA]⁺ and an undetectable fragment ion [FA + Li]⁺ while the FA was the less hydroxylated FA.     

Identification of Acylglycerols Containing Dihydroxy Fatty Acids in Castor Oil by Mass Spectrometry

Ricinoleate, a monohydroxy fatty acid, in castor oil has many industrial uses. Dihydroxy fatty acids can also be used in industry. The C₁₈ HPLC fractions of castor oil were analyzed by electrospray ionization mass spectrometry of lithium adducts to identify the acylglycerols containing dihydroxy fatty acids and the dihydroxy fatty acids. Four diacylglycerols identified were diOH18:1-diOH18:1, diOH18:2-OH18:1, diOH18:1-OH18:1 and diOH18:0-OH18:1. Eight triacylglycerols identified were diOH18:1-diOH18:1-diOH18:1, diOH18:1-diOH18:1-diOH18:0, diOH18:2-diOH18:1-OH18:1, diOH18:1-diOH18:1-OH18:1, diOH18:1-diOH18:0-OH18:1, diOH18:2-OH18:1-OH18:1, diOH18:1-OH18:1-OH18:1 and diOH18:0-OH18:1-OH18:1. The locations of fatty acids on the glycerol backbone were not determined. The structures of these three newly identified dihydroxy fatty acids were proposed as 11,12-dihydroxy-9-octadecenoic acid, 11,12-dihydroxy-9,13-octadecadienoic acid and 11,12-dihydroxyoctadecanoic acid. These individual acylglycerols were at the levels of about 0.5% or less in castor oil and can be isolated from castor oil or overproduced in a transgenic oil seed plant for future industrial uses.     

高效液相色谱法测定甲氧苄啶氯化钠注射液中甲氧苄啶的含量

建立了甲氧苄啶氯化钠注射液中甲氧苄啶含量的HPLC测定方法。本法采用C18色谱柱,流动相为0.1mol/L磷酸二氢钾溶液(用磷酸调节pH值至2.5)-甲醇的混合溶液(按v∶v=70∶30配制),检测波长为280nm,流速为1.0mL/min。实验数据表明:甲氧苄啶在0.38～1.37μg的范围内,峰面积与进样量呈良好线性关系(r=0.9995),回收率RSD=0.69%,最低检测限为1ng,分析结果准确,可用于测定甲氧苄啶氯化钠注射液中甲氧苄啶含量。