Rapid separation of neutral lipids,free fatty acids and polar lipids using prepacked silica sep-Pak columns

A method is described for the separation of neutral lipid, free fatty acid and polar lipid classes using small (600 mg), prepacked silica Sep-Pak columns. Combinations of hexane and methyltertiarybutylether were used to progressively elute cholesteryl ester first then triglyceride from the column. After column acidification, fatty acids were eluted followed by cholesterol. Recoveries of these lipids were 96% or greater. Polar lipids were eluted from the column using combinations of methyltertiarybutylether, methanol and ammonium acetate. Phospholipid classes could not be separated completely from each other. Phosphatidylethanolamine and phosphatidylinositol eluted together, whereas the more polar phosphatidylcholine, sphingomyelin and lysophosphatidylcholine were eluted as a second fraction. Recoveries of each phospholipid was greater than 98%.     

Analysis of neutral lipids and glycerolysis products from olive oil by liquid chromatography

A simple method was developed to separate model triglycerides, 1,3-diglycerides, 1,2-diglycerides, monoglycerides and free fatty acids and products generated in enzymatic glycerolysis of olive oil by high-performance liquid chromatography with a laser light-scattering detector. The separation was carried out with a silica column at 30°C. A gradient elution with mobile phase A (hex-ane/chloroform/formic acid) and mobile phase B (hex-ane/acetone/chloroform) can separate the compounds into distinct peaks in less than 20 min.     

Quantitative Papier-Chromatographie der Fettsäuren

Systematic Quantitative Analysis of Natural Waxes with the Help of Ion-Exchange, Column and Thin-Layer Chromatography A method was developed for the quantitative separation of natural waxes into substance classes. The hydrocarbons were separated by the adsorption chromatography on silica gel/CS₂ column and the separation of acids from alcohols was achieved on a two-phase ion-exchange column after saponification. The isolation of the individual homologues (C₁₆? C₃₂) from the alcohol and acid mixtures was carried out with the help of reversed phase column chromatography by stepwise increase in temperature and acetic acid concentration.     

Rapid separation of the major phospholipid classes on a single aminopropyl cartridge

A rapid method for the separation of the individual phospholipid classes phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS) and phosphatidylinositol (PI) by a single solid-phase extraction was developed. PC, PE, PS and PI were sequentially eluted from aminopropyl bonded silica with acetonitrile/n-propanol (2∶1, vol/vol), methanol, isopropanol/methanolic HCl (4∶1, vol/vol) and methanol/methanolic HCl (9∶1, vol/vol). Standard recoveries were over 95% for PC and PE and over 85% for PS and PI with undistorted fatty acid composition. The separation of complex lipid mixtures on aminopropyl minicolumns can be refined to the level of individual phospholipid classes.     

Gradient high-performance liquid chromatography of statistical and block copolymers of styrene and t-butyl methacrylate

Copolymers of styrene (S) and tert-butyl methacrylate (TBMA) containing 24–92 mass % of the latter monomer were investigated using a silica column with isooctane/tetrahydrofuran (THF) gradients and on a phenyl bonded-phase column by methanol (MeOH)/THF gradients using UV detection. In both cases, retention decreased with increasing TBMA content of the sample. This is in contrast to the behavior of copolymers of S and methyl methacrylate (MMA) whose retention in gradient elution on silica columns increases with MMA content due to the polarity of this unit. The inversion of elution order is a result of the bulky TBMA residues shielding the polar groups of the ester units. For copolymers of varying composition, the gradation in retention on retention on silica is quite sufficient for separating S/MMA but not for S/TBMA mixtures. The latter could be separated by composition on phenyl packings through MeOH/THF gradient elution. Block copolymers containing 80 or 92% TBMA could be baseline-separated. It was also possible to separate these samples from admixed PTBMA or PS homopolymers. Under the conditions used, the retention of a given block copolymer was somewhat higher than that of a statistical copolymer having the same composition. A block copolymer containing 80% TBMA was converted by transesterification into a S/MMA block copolymer. The elution of the latter fitted well in the sequence of S/MMA copolymers on silica column.     

Acute Hypercapnia/Ischemia Alters the Esterification of Arachidonic Acid and Docosahexaenoic Acid Epoxide Metabolites in Rat Brain Neutral Lipids

In the brain, approximately 90% of oxylipins are esterified to lipids. However, the significance of this esterification process is not known. In the present study, we (1) validated an aminopropyl solid phase extraction (SPE) method for separating esterified lipids using 100 and 500 mg columns and (2) applied the method to quantify the distribution of esterified oxylipins within phospholipids (PL) and neutral lipids (NL) (i.e. triacylglycerol and cholesteryl ester) in rats subjected to head-focused microwave fixation (controls) or CO₂-induced hypercapnia/ischemia. We hypothesized that oxylipin esterification into these lipid pools will be altered following CO₂-induced hypercapnia/ischemia. Lipids were extracted from control (n = 8) and CO₂-asphyxiated (n = 8) rat brains and separated on aminopropyl cartridges to yield PL and NL. The separated lipid fractions were hydrolyzed, purified with hydrophobic–lipophilic–balanced SPE columns, and analyzed with ultra-high-pressure liquid chromatography coupled to tandem mass spectrometry. Method validation showed that the 500 mg (vs 100 mg) aminopropyl columns yielded acceptable separation and recovery of esterified fatty acid epoxides but not other oxylipins. Two epoxides of arachidonic acid (ARA) were significantly increased, and three epoxides of docosahexaenoic acid (DHA) were significantly decreased in brain NL of CO₂-asphyxiated rats compared to controls subjected to head-focused microwave fixation. PL-bound fatty acid epoxides were highly variable and did not differ significantly between the groups. This study demonstrates that hypercapnia/ischemia alters the concentration of ARA and DHA epoxides within NL, reflecting an active turnover process regulating brain fatty acid epoxide concentrations.     

Effect of absorbent in solid‐phase extraction on quantification of phospholipids in palm‐pressed fiber

Palm‐pressed fiber (PPF) is a by‐product of palm oil milling and it has been found to contain a high percentage of phospholipids (PL). The aim of this work was to evaluate the best solid‐phase extraction (SPE) method to purify PL from PPF. The purified PL were analyzed using high‐performance liquid chromatography (HPLC) with an evaporative light‐scattering detector (ELSD). The oil was extracted from PPF using the Folch method and classes of PL were purified using SPE. Different solvent phases and normal‐phase SPE cartridges [silica (Si), aminopropyl‐bonded silica (NH₂) and diol‐bonded silica (2OH)] at the same ratio of oil to sorbent mass were used to study the separation of PL from the extracted oil. The recovery of each standard PL was performed in a model oil system with the same amount of standard PL being added, and the repeatability of the SPE method was investigated. The isolation of PL by SPE diol cartridge and subsequent analysis by HPLC/ELSD have shown to be an accurate and reproducible analytical method for the determination of phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine and lysophosphatidylcholine in PPF. This method provided a high yield and rapid separation of PL in PPF with less contamination from other lipid groups.     

Design and operation of a modified silica gel column chromatography

Liquid–solid chromatography (LSC) is the oldest of the various liquid chromatography methods. Despite the fact that high-performance liquid chromatography (HPLC) operation leads to better separation and analysis, classical column chromatography and thin-layer chromatography (TLC) are still widely practiced because of their convenience. In this study, a modified silica gel column chromatography was designed with the objective of reducing the amount of solvent required to achieve the same degree of separation as the classical silica gel column chromatography. The separation of squalene and fatty acid steryl esters (FASEs) from non-polar lipid fraction (NPLF) of soybean oil deodorizer distillate (SODD) was employed as a model system to test the effectiveness of this new design. Modified silica gel column chromatography process is feasible from economic point of view compare to classical silica gel column chromatography because it significantly reduces the amount of solvent and time required to achieve the same degree of separation. By employing modified silica gel column chromatography to obtain the squalene-rich fraction, the mobile phase volume and elution time required as fractions of those needed in classical silica gel column chromatography are 1/73 and 1/18, respectively. To obtain the FASEs-rich fraction, the corresponding mobile phase volume and elution time are 1/221 and 1/23, respectively of those needed in classical silica gel column chromatography.     

钝顶螺旋藻中γ-亚麻酸甲酯的提取研究

用１∶２∶０８（Ｖ／Ｖ）的氯仿 甲醇 水溶剂系统提取螺旋藻中总类脂,总类脂在硅胶柱上经氯仿、丙酮、甲醇依次洗脱,分离得到中性脂、总脂肪酸半乳糖脂、磷脂。总脂肪酸半乳糖脂与５∶９５（Ｖ／Ｖ）的氯乙酰 甲醇作用转变为总脂肪酸甲酯,再通过多次与尿素形成复合物,可从总脂肪酸甲酯中分离、纯化出 亚麻酸甲酯,其纯度达到了９６５％。     

Gram-scale preparative HPLC of phospholipids from soybean lecithins

A preparative procedure has been developed to isolate gram quantities of phospholipid classes from soybean lecithins. Various steps taken to accomplish the isolation are described—an analytical method with silica column and light scattering detector, alcohol fractionation of deoiled lecithins, and columns with increasing internal diameters but packed with the same stationary phase. The loading study showed that it was possible to inject 20 mg on a 100 × 8 mm RadialμPorasil colume. The separation was scaled up to a 25-mm i.d. column and finally to a 50-mm i.d. column. With the larger column, 2.1 g of phospholipids were separated. The collected fractions (phosphatidylethanolamine, phosphatidylinositol, phosphatidic acid and phosphatidylcholine) were of high purity (>99%). The solvent consumption was 7.2 L (separation and column equilibration), and a minimum of 10 g of polar lipids can be separated daily. Presented at the 82nd American Oil Chemists’ Society Annual Meeting, May 13–15 1991, Chicago, IL.     

非抑制型离子色谱法检测矿泉水中硅酸盐

该文介绍了一种采用阴离子色谱交换柱进行分离,非抑制型电导离子色谱法检测,直接进样测定矿泉水中硅酸盐（偏硅酸）含量的方法。选择的色谱条件为：氢氧化物选择性的IonPac AS11-HC阴离子交换柱,淋洗液自动发生装置在线产生KOH梯度淋洗,非抑制型电导检测。样品无需任何前处理直接进样。此方法具有选择性好,操作简单,适用性广,节省时间,对环境友好等特点,用于实际样品测定,所得结果令人满意。     

Lipid Studies of Pimpinella acuminata

The seed oil of Pimpinella acuminata species of the family Umbelli-ferae was extracted with chloroform : methanol (2:1) for the separation of different classes of lipids as hydrocarbons (1.7%), sterol esters (traces), triglycerides (74.1%), free fatty acids (6.6%), 1,3-diglycerides (1.6%), 1,2-diglycerides (1.6%), sterols (1.8%), mono-glycerides (2.0%), phosphatidyl ethanolamine (0.8%), phosphatidyl choline (1.2 %), lyso phosphatidyl ethanolamine (0.4%), phosphatidyl inositol (1.2%) and unknown (7.0%). The fatty acids as determined by application of gas liquid chromatography is C₁₀-C₂₂, whereas C_16:0, C_18:0-C_18:2 and C_18:2 are predominantly present in all polar and non-polar lipid classes.     

Chromatographic Detection of Silicones in Vegetable Oils

The method is based upon separation of silicone in the oil by thin layer Chromatography on silica gel layer, using the solvent system petroleum, ether—diethyl ether = 98:2 (v/v) and visualisation of the spots developed by means of Rhodamin B reagent. Under the described condition separation of silicones seems to be free from interference from other non-silicone containing materials. The limit of detection was approximately 2 γ.     

高效液相色谱手性固定相法拆分O，O-二甲基-1-（苯氧乙酰氧基）乙基膦酸酯和O，O-二甲基-1-（4-甲基苯氧乙酰氧基）乙基膦酸酯对映体

在直链淀粉-三-(3,5-二甲基氨基甲酸酯)(Chiralpak AD-H)手性柱上对O,O-二甲基-1-(苯氧乙酰氧基)乙基膦酸酯和O,O-二甲基-1-(4-甲基苯氧乙酰氧基)乙基膦酸酯对映体的分离进行了研究。分别考察了流动相极性改良剂的类型和比例以及温度对拆分效果的影响,并初步讨论了拆分的热力学机理。实验结果显示:流动相为正已烷-异丙醇(体积比90∶10),流速为1.0mL/min,检测波长220nm,温度25℃,O,O-二甲基-1-(苯氧乙酰氧基)乙基膦酸酯和O,O-二甲基-1-(4-甲基苯氧乙酰氧基)乙基膦酸酯在Chiralpak AD-H柱上分离度Rs分别为2.63、1.56,对映体都可得到满意的基线分离。     

Complete separation of phospholipids from human heart combining two HPLC methods

The separation of phospholipid classes from human heart was achieved in two steps by high performance liquid chromatography (HPLC) using a silica column with an ultraviolet spectromonitor at 206 nm. A complete partitioning of phosphatidylcholines (PC), phosphatidylethanolamines (PE), phosphatidylinositols (PI), phosphatidylserines (PS), cardiolipins (CL), lysophosphatidylcholines (LPC) and sphingomyelins (Sph) was obtained for further analysis.     

Protein Recovery from Sweet Potato Starch Wastewater by Foam Separation

In this study, an inclined foam separation column was designed to effectively recover protein from sweet potato starch wastewater. The effects of the influent protein concentration, pH, air flow rate, influent volume, foaming time, and inclined column angle on foam separation performance were assessed. The optimum foam separation conditions consisted of influent protein concentration 4.51 mg/mL, pH 4, air flow rate 0.15 mL/min, influent volume 500 mL, foaming time 100 min, and inclined column angle 30°. In these conditions, protein recovery percentage and enrichment ratios were 84.1% and 1.3, respectively. The biochemical oxygen demand (BOD₅) and chemical oxygen demand (COD_Cr) of the residual solution (620 and 950 mg/mL, respectively) were lower than those of the original (influent) solution.     

An affinity adsorbent derived from aminopropyl silica for serine protease chromatography

A silica-based adsorbent for affinity chromatography of serine protease was prepared by bonding p-aminobenzamidine (pABZ) to aminopropyl silica. Silanization of silica with both γ-aminopropyltrimethoxysilane and γ-amino-propyltriethoxysilane under anhydrous conditions led to a monolayer density of primary amino groups. The aminopropyl silica was converted to primary hydroxylcontaining silica via diazotization, and consequently activated with p-nitrophenyl chloroformate and immobilized with ligand pABZ. The resultant silica–pABZ has a ligand density of 13.8 μmol g^?1. Pyridine was indicated by the data to increase the reactivity of the chloroformate toward the hydroxyl group more efficiently than 4-dimethylaminopyridine. Two stages of adsorption were found in batch adsorption of trypsin with an equilibrium adsorption isotherm of the Langmuir type. When the chromatographic column packed with this silica–p ABZ was operated under a higher flow rate (2·33 cm³ min^?1) and with 3 g dm^?3 of crude urokinase as the influence, the yield was 55%. Both the flow rate and the concentration of the crude protein were shown, by measuring the dynamic binding capacity from chromatographic experiments, to be the factors which influenced the chromatographic efficiency.     

High-pressure liquid chromatography fingerprinting of petroleums and petroleum products

Combined GC/HPLC fingerprinting of petroleums and process streams derived therefrom gives detailed information on their constituents. The silica gel-heptane system affords separation into classes of hydrocarbons which, together with dual u.v. and R.I. modes of detection, gives abundant information on the composition of such samples. GC on a medium-efficiency SCOT column is used as a complementary technique only. Interpretations of HPLC chromatograms for samples ranging from gasolines to bitumens are given.     

Pulse response test in medium pressure protein chromatography

An MPLC (Medium Pressure Liquid Chromatography) column was inhouse packed with an anion exchange silica gel (PAE 1000, Amicon) in order to carry out pulse response experiments to investigate MPLC column efficiency. Bovine serum albumin was isocratically eluted from the column with varying ionic strength of NaCl, and the chromatograms obtained were analyzed by the moment method. Broadening of the elution band was caused mostly by a longitudinal dispersion in our system, and no significant numerical differences in HETP with the velocity change were observed at the high ionic strength.     

Precipitation-redissolution liquid chromatography of styrene-ethyl acrylate copolymers

The technique of Precipitation-Redissolution Liquid Chromatography (PRLC) has been applied for fractionation of styrene-ethyl acrylate copolymers in the whole composition range, from one homopolymer to the other. Two different polar types of column packings were used, pure silica and CN modified silica. Samples dissolved in tetrahydrofuran were injected in the columns where a nonsolvent mixture is used as chromatographic mobile phase. Polymers precipitate in the columns, and their elution is obtained by applying a gradient of eluent with increasing polarity and solvent power. The different copolymers, and the homopolymers, could be separated as a function of acrylate unit content. The best gradient conditions were obtained with a compromise between resolution, efficiency and analysis time. It was found that the polarity of the packing influences the separation in term of retention and peak resolution. This means that a composite mechanism is involved, with adsorption effects adding to the precipitation in favoring the fractionation according to the sample polarity.