Analysis of tocopherols by capillary supercritical fluid chromatography and mass spectrometry

Tocopherol-containing mixtures were analyzed by gas chromatography (GC) and capillary supercritical fluid chromatography (SFC). GC analysis of tocopherols required the formation of the silyl derivatives, while SFC analysis of the tocopherol-containing mixtures was accomplished on neat samples. SFC analysis conditions were optimized with respect to column type and density/pressure programming. Enhanced resolution of many components was achieved by using inverse temperature programming during the SFC analyses. Both SFC and GC analyses permitted the separation and quantitation of alpha-, beta-, gamma- and delta-tocopherols. In addition, SFC proved particularly applicable for characterizing the composition of a deodorizer distillate and commercial antioxidant formulation. Coupling of a quadrapole mass spectrometer with a supercritical fluid chromatograph was also achieved; the mass spectrometer provided electron impact mass spectra on the underivatized tocopherol and sterol moieties. Both SFC and SFC/mass spectrometry proved effective for the analysis of complex lipid-containing mixtures, requiring minimal sample preparation prior to analysis. Presented at the 82nd Annual Meeting of the American Oil Chemists’ Society, Chicago, IL, May 12–15, 1991.     

Comparison of high-temperature gas chromatography and CO2 supercritical fluid chromatography for the analysis of alcohol ethoxylates

This work compares capillary supercritical fluid chromatography (SFC) and capillary high-temperature gas chromatography (HTGC) for the quantitative characterization of nonionic alcohol ethoxylate surfactants. Supercritical fluid chromatographic separations of the alcohol ethoxylates were obtained with a density-programmed carbon dioxide mobile phase and a fused silica capillary column. High-temperature gas chromatographic separations were obtained with a high-temperature polyimide-coated fused silica capillary column. In addition, a procedure was developed for the quantitation of the capillary chromatographic data using flame ionization molar response factors based on the effective carbon theory. The alcohol and ethoxylate distributions, mean molecular weights and average moles of the ethylene oxide are rapidly calculated from the chromatographic data. Advantages and limitations of SFC and HTGC procedures are illustrated and discussed. Based on this work, the following conclusions can be drawn: i) For routine quality control analyses of known alcohol ethoxylates, SFC and HTGC appear to be equally applicable. ii) SFC has the advantage of time because derivatization is not required, although derivatization does improve resolution. iii) HTGC has the advantage of resolving C₁₂ through C₁₈ alcohol ethoxylate oligomers, avoiding ambiguous identification of components. iv)SFC and HTGC both have disadvantages. SFC has a resolution limitation and HTGC discriminates against high molecular-weight components. Presented in part at the 83rd Annual Meeting in Chicago, IL, May 1991.     

Modification of margarine fats by enzymatic interesterification: Evaluation of a solid-fat-content-based exponential model with two groups of oil blends

Lipozyme TL IM-catalyzed interesterification for the modification of margarine fats was carried out in a batch reactor at 70°C with a lipase dosage of 4%. Solid fat content (SFC) was used to monitor the reaction progress. Lipase-catalyzed interesterification, which led to changes in the SFC, was assumed to be a first-order reversible reaction. Accordingly, the change in SFC vs. reaction time was described by an exponential model. The model contained three parameters, each with a particular physical or chemical meaning: (i) the initial SFC (SFC₀), (ii) the change in SFC (ΔSFC) from the initial to the equilibrium state, and (iii) the reaction rate constant value (k). SFC_o and ΔSFC were related to only the types of blends and the blend ratios. The rate constant k was related to lipase activity on a given oil blend. Evaluation of the model was carried out with two groups of oil blends, i.e., palm stearin/coconut oil in weight ratios of 90∶10, 80∶20, and 70∶30, and soybean oil/fully hydrogenated soybean oil in weight ratios of 80∶20, 65∶35, and 50∶50. Correlation coefficients higher than 0.99 between the experimental and predicted values were observed for SFC at temperatures above 30°C. The model is useful for predicting changes in the SFC during lipase-catalyzed interesterification with a selected group of oil blends. It also can be used to control the process when particular SFC values are targeted.     

超临界流体色谱法制备EPA-EE和DHA-EE

以二氧化碳作流动相,C18柱为固定相,用超临界流体色谱法制备二十碳五烯酸乙酯(EPA-EE)和二十二碳六烯酸乙酯(DHA-EE).考察了流速、温度、压力、上载量以及进样浓度对制备分离的影响.结果表明,在实验范围内,流速对分离效果影响不大.对于5-C18,250mm×4.6mm色谱柱,其最适制备条件是柱前压12MPa,温度55℃,通常流速下的压降约为1MPa,鱼油的最大上载量大于20μl.而对于8-C18,250mm×10mm色谱柱,其相应的最适制备条件是柱前压11MPa,温度55℃,通常流速下的压降约为0.5MPa,鱼油的最大上载量约为100μl.实验表明,鱼油无需溶解于其他溶剂而可直接进样.     

A novel technique for the preparation of secondary fatty amides

A technique for the synthesis of monosubstituted fatty amides at low temperature and ambient pressure was developed. This method involved the condensation of an amine with a triacylglycerol. The primary amine (ethyl,n-butyl,n-hexyl andn-octyl were tested) acted as reagent and solvent for the fatty substrates. No additional organic solvent or catalyst was added. Tallow, vegetable oils and fish oil all served well as substrates, as did pure tripalmitin. The rate of amidation was dependent upon temperature and the ratio of fat to amine. In a series of experiments conducted with tallow andn-butylamine at a fat:amine molar ratio of 1:16, amidation could be carried out at 20°C, producingn-butyltallowamide in 83% yield in 24 hr. When the fat:amine molar ratio was reduced to 1:8, and the temperature raised to 45°C, the amide yield was 87.6% in 24 hr. When the reaction was carried out at the boiling point ofn-butylamine (78°C) and at a fat:amine ratio of 1:8, the amide yield was 93.2% in 4 hr. The reaction progressed more rapidly with higher molecular weight amines. The identity and purity of the amides was assessed by thin-layer chromatography and confirmed by elemental analyses and infrared and C¹³ nuclear magnetic resonance spectroscopy.     

Effect of emulsion temperature on physical properties of palm oil-based margarine

Margarines made from refined, bleached, and deodorized palm oil at different emulsion temperatures showed no significant difference in their consistency, polymorphic behavior, and solid fat content (SFC) during storage, although differences were observed during processing. The emulsion temperatures studied were 40, 45, and 50°C, with other parameters such as emulsion flow rates, tube cooler temperature, and pin rotor speed kept constant. The SFC developed during processing and storage at 28°C was measured to evaluate the quality of margarine. The emulsion contained no SFC at any emulsion temperature studied. However, the amount of SFC in the perfector or tube cooler unit increased to 15.9, 13.9, and 15.6% in margarine produced at emulsion temperatures of 40, 45, and 50°C, respectively. At 40°C, the lowest SFC was developed during storage even though this margarine had the highest consistency. The softening point of this sample was moderately high and closely related to the type of crystal developed, which was a mixture of β′ and β crystals. Emulsion at 45°C gave the most stable margarine consistency and SFC with crystal in the β′ form even after the fourth week. At 50°C, moderately soft product was produced, which might be undesirable for some applications, although the crystals were in the β′ form.     

Stereoregular oligomers of methyl methacrylate

Summary Packed column SFC has been found suitable for the rapid and detailed analysis of the isotactic and syndiotactic oligomers of MMA, when the temperature gradient technique was applied and the modifier was employed. Oligomer components from trimer to 20-mer separated completely. The heptamer fraction collected three times by SFC gave the ¹H NMR spectrum of satisfactorily high S/N ratio; the spectrum agreed well with that of the standard sample. Separation by tacticity as well as by molecular weight was observed for the SFC of a mixture of the isotactic and syndiotactic oligomers. The isotactic oligomers had longer retention time than the syndiotactic oligomers of the corresponding degree of polymerization. Part 3: cf. Ute K. Nishimura T, Hatada K, Polym J (1989) 21: 1027     

Optimization of experimental parameters for separation of nonionic surfactants by supercritical fluid chromatography

A synthetic polymer generally consists of a mixture of many compounds that share the same repeating unit but have different degrees of polymerization. Because of its ability to cope with a wide range of molecular weights, supercritical fluid chromatography (SFC) is a powerful technique for separating the individual components of polymers. However, the separation conditions for SFC are more sensitive than those of other types of chromatography, and many experimental parameters need to be carefully selected to achieve high resolutions. Methods for optimizing the experimental parameters for SFC of a polymeric surfactant were investigated by chemometric techniques. Increases or decreases in peak resolution and elution times were measured for the separation by SFC of the nonionic surfactant poly(ethylene glycol) nonylphenyl ether for various column temperatures, flow-rate gradients of modifier, and modifier compositions. To maximize the separation efficiency, the experimental results were analyzed by using the chromatographic response function (CRF). This is a function of the desired peak resolution, the maximum acceptable elution time, and an arbitrary weighting factor that is adjusted to achieve an optimal balance between the peak separation and the elution time. Because the numerical value of the CRF changes in response to the chromatographic performance, it provides a means of identifying the conditions for achieving optimal separation for a particular SFC system.     

Physical analyses of gel-like behavior of binary mixtures of high- and low-melting fats

Gel-like fat mixtures of high-melting (HM) and low-melting (LM) fats were formed by means of rapid cooling and subsequent heating. No “non-fat” ingredients such as emulsifiers, water, or waxes were added to the mixtures. The gel-like fats having solid fat content (SFC) values below 2.0 wt% formed crystal networks of HM-fats that entrapped the liquid oil fraction of LM-fats. In a search for optimal fat combinations exhibiting gel-like behavior, fully hydrogenated rapeseed oil with a high amount of behenic acid (FHR-B), fully hydrogenated rapeseed oil with a high amount of stearic acid (FHR-S), tristearoylglycerol (SSS), triarachidonoyl-glycerol (AAA), and tribehenoylglycerol (BBB) were examined as the HM-fats. For LM-fats, sal fat olein (SFO), cocoa butter (CB), palm super olein (PSO), and olive oil were examined. The following results were obtained: (i) the gel-like behavior was observed in mixtures of FHR-B/SFO and FHR-B/CB with initial concentrations of FHR-B of 1.5–4.0 wt%. (ii) Rapid cooling to T _c (crystallization temperature) from 70°C and subsequent heating to T _f (final temperature) were necessary to reveal the gel-like behavior, whereas simple cooling without a cooling/heating procedure did not form the gel-like fat mixture. (iii) Optimal values of T _c and T _f were related to the m.p. of the LM-fat and HM-fat, respectively. (iv) Temperature variations of SFC as well as X-ray diffraction spectra showed that the melt-mediated transformation from α to β of the HM-fat crystals was a prerequisite to reveal the gel-like behavior. Consequently, the fat mixture revealing the gel-like behavior might be called β-fat gel.     

Effects of hydrogenation parameters on trans isomer formation,selectivity and melting properties of fat

Effects of hydrogenation conditions (temperature, hydrogen pressure, stirring rate) on trans fatty acid formation, selectivity and melting behavior of fat were investigated. To this aim, soybean oil was hydrogenated under various conditions and fatty acid composition, trans isomer formation, slip melting point (SMP), solid fat content (SFC) and iodine number (IV) of the samples withdrawn at certain intervals of the reactions were monitored. A constant ratio (0.03%) of Nysosel 222 was used in the various combinations of temperature (150, 165 and 180 °C), stirring speed (500, 750 and 1000 rpm) and hydrogen pressure (1, 2 and 3 bar). Raising the temperature increased the formation of fatty acid isomers, whereas higher stirring rates decreased this formation, while changes in hydrogen pressure had no effect or slightly reduced it, depending on other parameters. Results also indicated that the trans fatty acid ratio increased with IV reduction, reached the highest value when the IV was about 70 and decreased at IV < 70 due to saturation. Selectivity values (S₂₁) at that point ranged between 5.78 and 11.59. Lower temperatures and higher stirring rates decreased not only the trans isomer content but also the S₂₁ values at significant levels. However, same effects were not observed with the changes in hydrogen pressure. It was determined that a high SMP does not necessarily mean a high SFC. Selective conditions produced samples with higher SFC but lower SMP, which is possibly because of higher trans isomer formation as well as lower saturation.     

Relationship between oil binding capacity and physical properties of interesterified soybean oil

The objective of this study was to identify the physical properties of an interesterified soybean oil (EIESOY), containing 45% saturated fatty acids (SFA), that correlates with high oil binding capacity (OBC) and low oil loss (OL). In this study, three EIESOY samples were analyzed; a 100% sample, a 50% sample diluted with 50% soybean oil, and a 20% sample diluted with 80% soybean oil. All samples were crystallized using fast (7.78°C/min) and slow (0.1°C/min) cooling rates as well as with and without high-intensity ultrasound (HIU, 20 kHz). The 100%, 50%, and 20% samples were crystallized at 38.5, 27.0, and 22.0°C, respectively. HIU was applied at the onset of crystallization and all samples were allowed to crystallize isothermally for 90 min. After 90 min, physical properties such as crystal microstructure, hardness, solid fat content (SFC), elasticity, and melting behavior were evaluated. Physical properties were also measured after storage for 48 h at 22 and 5°C. Results show that OBC was positively correlated with hardness, G′, and SFC after 48 h (r = 0.738, p = 0.006; r = 0.639, p = 0.025; r = 0.695, p = 0.012; respectively), OL was negatively correlated with hardness after 48 h (r = −0.696, p < 0.001), G′ after 90 min and 48 h (r = −0.704, p < 0.001; r = −0.590, p = 0.002), and SFC after 90 min and 48 h (r = −0.722, p < 0.001; r = −0.788, p < 0.001). Neither OBC nor OL were correlated with crystal diameter or the number of crystals.     

Purification and partial characterization of antifreeze proteins from leaves of Ligustrum lucidum Ait

Three AFPs (AFP-1, AFP-2, AFP-3) of Ligustrum lucidum Ait leaves were purified by using ice-affinity, chromatography separation on a DEAE-cellulose-32 column and a Sephadex G100 column. The ice-affinity proteins were about 1.2 mg/100 g leaves. Their molecular weight was 66.1, 26.3, and 20.2 kDa, respectively. Their thermal hysteresis activity was 0.379, 0.678, and 0.460 °C respectively. Asx was very abundant in these AFPs; its molar ratio was 22.2, 24.9, and 27.8%, respectively. AFP-2 was effective to protect peroxidase, β-glucosidase, and trehalose synthase from freeze–thawing process. Cryoprotection of AFP-2 is better than trehalose.     

In vitro hydrolytic degradation of poly(para-dioxanone) with high molecular weight

The in vitro hydrolytic degradation of high molecular weight poly (para-dioxanone) was studied by examining the changes of weight retention, water absorption, pH value, tensile strength, break elongation, thermal properties, and morphology of high molecular weight PPDO in phosphate buffered saline (PBS) (pH 7.44) at 37°C for 8 weeks. During the degradation, all samples’ weight retention decreased and water absorption increased significantly, whereas hydrolysis rate of PPDO bars varied with molecular weight. Compared with lower molecular weight samples, higher molecular weight PPDO samples exhibited higher hydrolysis rate. The samples’ glass transition temperature (Tg) decreased notably, while the degrees of crystallinity (Dc) increased. The samples almost totally lost their tensile strengths and breaking elongation after 4 weeks of degradation. The results suggested that the stability of PPDO in vitro hydrolytic degradation increased with the increase of molecular weight.     

Effect of scraped-surface tube cooler temperatures on the physical properties of palm oil margarine

The effects of scraped-surface tube cooler temperatures on the isothermal solid fat content (SFC) of palm oil margarine during processing and on margarine consistency (yield value, g/cm²), SFC, and polymorphic changes in storage were studied. SFC was measured in the mixing tank after leaving the tube cooler and the pin worker. The SFC at the tube cooler exit was proportional to the amount of cooling; a higher SFC was produced by more extreme cooling treatment. The SFC of all margarines were reduced in the pin worker, and the reduction was related to the initial SFC profile of palm oil. Margarine samples were stored at 28°C for 28 d and tested daily. Margarine processed at 25°C in the tube cooler had the highest consistency and the least change in SFC, but by the second week crystals had transformed into the β form. Uniform product consistency and SFC were observed in margarines processed at 20 and 15°C. These margarines retained the β′ crystal form for 3 and 4 wk, respectively. The best palm oil margarine was obtained with a tube cooler temperature of 15°C and a residence time of 1.8 min.     

Recovery,fractionation and study of oils from a lignite

Oils from a lignite (Teruel, Spain) were studied. They were obtained by catalytic depolymerization, by reacting an aromatic substrate (tetralin), Lewis acid catalyst (BF₃) and the lignite (oil A) or asphaltenes (oil B) at atmospheric pressure and temperatures <220 °C. The A and B oils were fractionated on an alumina-silica gel dual column into their saturated, mono- and diaromatic, polyaromatic and polar components by elution with hexane, hexane-benzene, benzene and methanol-diethyl ether-benzene respectively. The elemental analysis, average molecular weight, i.r. and ¹H n.m.r. spectroscopy and aromaticity data of these subfractions were studied.     

Accelerated Fat Bloom in Chocolate Model Systems: Solid Fat Content and Temperature Fluctuation Frequency

Chocolate model systems were designed with high‐melting fat (cocoa butter stearin, CB‐S) mixed with low‐melting fats (sunflower oil, canola oil, cottonseed oil, peanut oil) to give 45, 55, and 65 % solid fat content (SFC). Defatted cocoa powder provided a 50 % particulate level in the model chocolates. The effects of SFC, low‐melting fat type and storage temperature fluctuation frequency on bloom whiteness were investigated. Both SFC and storage condition had significant influences on bloom whiteness (p < 0.0001), although the type of low‐melting fat did not (p = 0.1223). Increasing the SFC significantly reduced bloom formation, as did more rapid temperature fluctuations.     

Optimization of supercritical CO2 extraction of Anastatica hierochuntica

Response surface methodology (RSM) was applied to optimize the variables affecting the supercritical carbon dioxide (SC-CO₂) extraction of non-polar compounds from Anastatica hierochuntica using the Central Composite Design technique (CCD). Independent variables were temperature (32–46 °C) and pressure (22–46 MPa). Dependent variables were the percentage of the content of hexadecanoic acid, 9,12-octadecadienoic acid, heneicosane and heptacosane. Pressure was the most significant parameter that affected the content of the compounds. The hexadecanoic and 9,12-octadecadienoic content decreased while heneicosane and heptacosane increased with pressure. A number of choices can be run either at low pressure and low temperature or at low pressure and high temperature in order to optimize extraction of the selected compounds. Extraction either at low temperature (33 °C) and low pressure (25.6 MPa), or at high temperature (42 °C) and low pressure (22.0 MPa) maximized the yield of hexadecanoic, 9,12-octedecanoic, heneicosane and heptacosane.     

Physical properties of Pseudomonas and Rhizomucor miehei lipase-catalyzed transesterified blends of palm stearin:palm kernel olein

The physical properties of Pseudomonas and Rhizomucor miehei lipase-catalyzed transesterified blends of palm stearin:palm kernel olein (PS:PKO), ranging from 40% palm stearin to 80% palm stearin in 10% increments, were analyzed for their slip melting points (SMP), solid fat content (SFC), melting thermograms, and polymorphic forms. The Pseudomonas lipase caused a greater decrease in SMP (15°C) in the PS:PKO (40:60) blend than the R. miehei lipase (10.5°C). Generally, all transesterified blends had lower SMP than their unreacted blends. Pseudomonas lipase-catalyzed blends at 40:60 and 50:50 ratio also showed complete melting at 37°C and 40°C, respectively, whereas for the R. miehei lipase-catalyzed 40:60 blend, a residual SFC of 3.9% was observed at 40°C. Randomization of fatty acids by Pseudomonas lipase also led to a greater decrease in SFC than the rearrangement of fatty acids by R. miehei lipase. Differential scanning calorimetry results confirmed this observation. Pseudomonas lipase also successfully changed the polymorphic forms of the unreacted blends from a predominantly β form to that of an exclusively β′ form. Both β and β′ forms existed in the R. miehei lipase-catalyzed reaction blends, with β′ being the dominant form.     

Optimization of enzymatic methanolysis of soybean oil by response surface methodology

Enzymatic methanolysis of refined soybean oil with methanol was investigated using Rhizomucor miehei lipase, Lipozyme RM IM, in n-hexane for reaction times of 30 min. Response surface methodology (RSM) based on three-level, three-factor (variable) face-centered cube design was used for the optimization of methanolysis. The independent variables that affect the methanolysis reaction conducted in n-hexane are temperature (°C), enzyme/oil weight ratio, and oil/methanol molar ratio. A good quadratic model was obtained for the methyl ester production by multiple regression and backward elimination. A linear relationship was observed between the observed and predicted values (R²−0.9635). The effects of temperature and enzyme amount, which affected methyl ester content of the product (response) positively, were significant (P<0.01). The quadratic term of temperature and the interaction term of enzyme amount with temperature affected the response negatively (P<0.01). The interaction term of enzyme amount with substrate mole ratio had a positive effect on the response (P<0.05). Critical conditions for the response at which methyl ester content of the product was 76.9% were determined to be 50°C, 2.37 methanol/oil mole ratio, and 0.09 enzyme/oil weight ratio.     

Chemical and Enzymatic Interesterification of a Blend of Palm Stearin: Soybean Oil for Low trans-Margarine Formulation

A blend of palm stearin and soybean oil (70/30, wt%) was modified by chemical interesterification (CIE) and enzymatic interesterification (EIE), the latter batch-wise (B-EIE) and in continuous (C-EIE). Better oil quality, mainly in terms of acidity, free tocopherol and partial acylglycerol content, was obtained after EIE. The clear melting point after any interesterification process was similar and about 9 °C lower as result of the modification in the TAG profile, which approaches the calculated random distribution. Interesterification changed the SFC profile significantly. For the fully refined interesterified blends, the SFC profile was similar and clearly different from the starting blend. Interesterification decreased the content of solids at temperatures >15 °C and increased the content of solids at temperatures <15 °C. This increase was less remarkable after C-EIE, suggesting that full randomization was not achieved in the used conditions, probably caused by a too short residence time of the oil in the enzymatic bed. During B-EIE, variations in SFC with time, principally at low temperatures, were still observed although the TAG composition was stable. At low temperatures, the reaction rate calculated from SFC was very low, confirming an important effect of the acyl migration on this parameter.