Purification of sporidesmin by preparative high-pressure liquid chromatography

An acid/alkali treatment which effects a fourfold purification of sporidesmin, making the sample immediately amenable to purification by preparative high-pressure liquid chromatography (h.p.I.c.), is described. The alkali treatment also removes polyphenols which have been responsible for discolouration problems in previous purification methods.     

生 化 分 离 技 术──制备型高效液相色谱

制备型高效液相色谱（ＨＰＬＣ）是在传统的分析型ＨＰＬＣ的基础上发展起来的一种高效分离纯化技术，主要用于制备高纯度生物活性物质。本文介绍了制备型ＨＰＬＣ的装置组成、制备规模及应用实例。     

阻燃剂磷酸双酚A四苯酯（BAPP）的液相色谱分析

用高效液相色谱法（HPLC）分离阻燃剂磷酸双酚A四苯酯,液-质谱联用定性分析,面积归一化法定量。采用Spheri SorbC18／SiO2（粒径5μm,规格250mm×4．6mm）为色谱柱,以乙腈／四氢呋喃／0．04％磷酸水溶液（三者体积比为35：30：35）为流动相,流速0．8mL／min,乙腈的吸收波长为210nm,四氢呋喃吸收波长为280nm,样品BAPP分别在270nm、254nm波长下有两个吸收峰,通过对乙腈、四氢呋喃、BAPP紫外光谱的分析,选择测定波长254nm。该分析方法可以作为BAPP纯度的分析方法来表征所合成的产品质量,虽然双酚A组分相对偏差较大,基本能够满足控制分析方法的需要。     

Separation and purification of lecithins by high pressure liquid chromatography

Ten different synthetic lecithins have been analyzed by reverse-phase high pressure liquid chromatography. An empirical lecithin “carbon number” that depends on the total number of carbons and double bonds in the fatty acyl chains in a useful index in predicting retention volumes of lecithins on a nonpolar octadecyl fatty acid column. Commercial egg lecithin is separated into its components by this technique.     

Analysis of dolichol in human tissues by high pressure liquid chromatography

Dolichol from human liver was shown by reverse-phase high pressure liquid chromatography to consist of a series of homologues ranging in length from 17 to 23 isoprene units. The two major components, corresponding to 19 and 20 isoprene units, respectively, were isolated and identified by mass spectrometry. Dolichyl palmitate, synthesized from liver dolichol, showed an identical series of peaks with longer retention times. Attemps to chromatograph dolichyl phosphate under similar conditions were unsuccessful. Dolichol from uterine tissue and several other human tissues showed a shift toward shorter chain length, with a predominance of homologues containing 18 and 19 isoprene units.     

Separation ofCis andTrans isomers by reverse phase high pressure liquid chromatography

A reverse phase high pressure liquid chromatography procedure was devised for the analytical or preparative separation of geometric isomers of dodecenyl acetates, tetradecenyl acetates, hexadecenyl acetates, tridecadienyl acetates, and methyl oleate and elaidate. Use of μ Bondapak C-18 permitted the separation of these isomers. ARS, USDA.     

Jojoba oil analysis by high pressure liquid chromatography and gas chromatography/mass spectrometry

Two analytical procedures for determining com-positions of jojoba liquid wax esters are described and compared. One, the more tedious, involves separation of wax ester homologs by high pressure liquid chro-matography followed by determination of the acid and alcohol moieties from each homolog. The second allows rapid determination of wax ester composition by gas Chromatographic separation of hydrogenated jojoba wax esters according to chain length, followed immediately by ancillary mass spectrometric identifi-cation of the acid and alcohol moieties. Double bonds in the alkyl chains in jojoba liquid waxes were almost exclusively (98%) ω-9, when examined by gas chro-matography/mass spectrometry (GC/MS) and ozonolysis/GC/MS. Presented in part at the 2nd International Conference on Jojoba and Its Uses, Ensenada, Mexico, February, 1976.     

The purification of fatty acid methyl esters by high pressure liquid chromatography

Procedures are described for the rapid purification of gram quantities of methyl oleate, methyl linoleate, methyl α- and γ-linolenates and methyl ricinoleate from appropriate natural oils by high pressure liquid chromatography (HPLC).     

Two-way chromatography: Flow reversal in non-linear preparative liquid chromatography

A liquid chromatographic process is described in which the mixture to be separates and the eluant are introduced alternately at either end of the colum The products are also withdrawn alternately at either end or in side-streams. Difrerent operating schemes are shown to be possible, according as the el is more, or less strongly retained by the adsorbent than the components to be separated.Experiments are presented on the ion-exchange chromatography of Na⁺ = K⁺ mixtures with H⁺ as eluant. A theoretical analysis is given using the non-linear multicomponent equilibrium theory and performance criteria are determined. Both experiments and theory lead to the following salient fea of the process, which permit to distinguish it from existing, analytical or preparative chromatographic processes: the process is preparative: it treats large injections of highly concentrated components; the process is separative: it does not merely remove solutes from a solvant, it separates solutes from each other; the process is a binary split: it gives two products; thus multicomponent mixtures require several steps if complete separation is desired; the process is non-linear, as a result of the high concentrations used; the tailing of the peaks induced by the non-linearity is counteracted by recompression due to flow-reversal; the process reduces dilution and saves eluant: one of the components may be collected highly concentrated, even pure, avoiding thus secondary separation; the process requires slightly more adsorbent capacity than conven chromatography, for the same production; the process works best when the eluant is more, or less strongly retained than the components to be separated, but not intermediately; the process uses intermediate storage of part of the effluents, and their “reflux” in subsequent steps.     

Direct estimation of dolichyl phosphate in rat liver by high pressure liquid chromatography

A method involving reverse-phase high pressure liquid chromatography has been developed for determining the concentration of dolichyl phosphate (Dol-P) in tissues. Individual Dol-P homologs are resolved and amounts as small as 50 ng can be detected. Rat liver was found to contain 2.4 μg Dol-P/g wet weight, or ca. 4% of total liver dolichol. In contrast, rat liver microsomes contained 64 ng Dol-P/mg protein, which is about 40% of total microsomal dolichol. This enrichment in Dol-P is consistent with the role of microsomes as the major site of Dol-P-mediated, glycoprotein biosynthesis in liver     

Quantitative determination of polyethylene glycols in nonionic surfactants by high pressure liquid chromatography

A method employing high pressure liquid chromatography has been developed for the quantitative determination of polyethylene glycols in ethoxylated fatty alcohols and alkylphenols. This technique overcomes many of the limitations encountered in previously-reported methods. The polyethylene glycols are separated from the ethoxylated product and other sample components using a 65/35 acetonitrile/water mobile phase and a Bondapak C₁₈/Corasil reversephase column system. The response factor of the differential refractometer detector is determined using Carbowax standards of appropriate molecular weights. The molecular weight of the polyethylene glycols in each sample is approximated using thin layer chromatography prior to the high pressure liquid chromatography calibration and analysis. The precision of this method for the determination of polyethylene glycols is ± 4% relative or better, and the recovery of added polyethylene glycols is quantitative. Application of this method to a wide variety of commercial ethoxylated fatty alcohols and alkylphenols is presented. Presented at the Meeting of the American Oil Chemists’ Society, April 1976, New Orleans.     

New application of high pressure reversed-phase liquid chromatography in lipids

High pressure reverse-phase liquid chromatography has been used to separate saturated fatty acids, their methyl esters, polyunsaturated fatty acids, and triglycerides. Rapid separation of fatty acids differing in chain length and number of double bonds has been accomplished. Analysis time was less than 10 min in most cases. The high pressure reverse chromatography resulted in better separations of polyenoic acids than can be accomplished by conventional argentation silicic acid column chromatography. The analyses were carried out on a chemically bonded reverse phase packing, VYDAC reverse phase.     

Separation of oxidized and unoxidized molecular species of phosphatidylcholine by high pressure liquid chromatography

Soy phosphatidylcholine (PC) has been separated into its major molecular species by reversephase high pressure liquid chromatography (HPLC). An aqueous methanol gradient was used that allowed detection of the various species by their absorbance at 206 nm. Oxidized species were detected by their absorbance at 234 nm and were resolved from the unoxidized species. This technique has been used to separate and purify unoxidized dilinoleyl phosphatidylcholine (di 18∶2 PC) from other species of soy PC and to monitor the autoxidation of an aqueous suspension of the purified di 18∶2 PC. Two oxidized products were formed from di 18∶2 PC. Further analysis showed that they were PC, but one of the products contained an oxidized and an unoxidized fatty acid; in the other product, both fatty acids were oxidized. Present in part at FASEB 63rd Annual Meeting, Dallas, Texas, April 1979.     

Characterization of low molecular compounds in dashboard foils by extraction,preparative high performance liquid chromatography,and IR-spectroscopy

A procedure to obtain information concerning the low molecular weight compounds of dashboard foils is described. Solvent extracts of the foils were fractionated by preparative reversed or normal phase HPLC, respectively. The fractions were characterized by their infrared spectra. The benefits and the shortcomings of the two methods are discussed.     

Analysis of trichlorocarbanilide and triclosan in soaps by reverse phase high pressure liquid chromatography

A method has been developed for the rapid and direct analysis of bacteriostats (3,4,4′-trichlorocarbanilide and 2,4,4′-trichIoro-2′ -hydroxydiphenyl ether) in soaps with the aid of reverse phase high pressure liquid chromatography (HPLC) The bacteriostats were conveniently analyzed by UV absorption detection at 280 nm. A typical analysis of bacteriostats by this method requires 15 min for sample preparation and 15 min for the HPLC run. As the system needs no equilibration time (isocratic elution), it is immediately and reproducibility.     

Isolation and purification of deoxynivalenol and a new trichothecene by high pressure liquid chromatography

Deoxynivalenol (3,7,15-trihydroxy-12,l3-epoxytrichothec-9-ene-8-one) was extracted from corn with methanol/water (80:20, v/v) and purified by liquid:liquid partitioning and by preparative high pressure liquid chromatography (HPLC). This procedure was used to prepare mg quantities of toxin from field-inoculated corn for reference standards. Analysis of the isolated deoxynivalenol by analytical HPLC, gas liquid chromatography (GLC) and gas liquid chromatography/mass spectroscopy (GLC/MS) indicated the presence of a second compound similar to deoxynivalenol. This compound comigrates with deoxynivalenol on thin layer chromatography plates in chloroform/methanol (90:10, v/v), but can be separated by HPLC on a reverse-phase C₈ column with methanol/water (10:90, v/v). GC/MS of the compound and the trimethylsilyl ether derivative gave parent ions of m/e 280 and 424, respectively. These data and NMR data indicate that the compound is 3,15-dihydroxy-12,13-epoxytrichothec-9-ene-8-one, a previously unreported trichothecene.     

Determination of oligosaccharides in soybeans by high pressure liquid chromatography using an internal standard

A method was developed for the quantitative analysis of oligosaccharides in soybeans by high pressure liquid chromatography (HPLC). The sugars were extracted from the soy flour using an ethanol-water solution. Separation of the oligosaccharides was effected by injecting a sample extract onto an HPLC equipped with a μBONDAPAK/carbohydrate® column. Three quantitative techniques were investigated for determining the separate sugars: (a) repetitive injection (i.e., alternately injecting equal volumes of standards and samples); (b) extract analyzed before and after spiking with raffinose (one of the unknowns); (c) the addition of a pure inexpensive internal standard, β-cyclodextrin, which was separated completely from the other oligosaccharides. These methods were successfully applied to the quantitative analyses of two varieties of soybeans and other soybean products such as soy milk and soy protein concentrate.     

Correlation of HETP and experimental variables in preparative liquid chromatography

HETP (Height Equivalent to a Theoretical Plate) is widely designated as the column efficiency in Chromatographic separations. The effects of experimental variables such as concentration, injection volume, flow rate and composition of the mobile phase, column diameter, and column length on HETP were investigated by preparative liquid chromatography. Water and methanol as an organic modifier were used as the mobile phase. A sample of thymidine was injected into C₁₈ columns with different dimensions. From fifty experimental runs, it was shown that the larger amounts of sample and higher flow rates increased HETP and an optimum HETP existed at a column diameter of 7.8 mm for preparative packings used in this experiment. The experimental values of HETP were correlated into a quadratic equation with the interaction terms based on the logarithmic experimental values of the experimental variables ; its regression coefficient was 0.952. The experimental variables were simulated from the correlation equation, and their effects on HETP were discussed.     

Sorbic acid containing triglycerides in aphids and their fractionation by high pressure liquid chromatography

The sorbic acid containing triglycerides found in various aphid species have been fractionated by high pressure liquid chromatography. Mass spectral analysis afforded identification of three isolates as 2-trans, trans-sorbo-1, 3 dimyristin; 2-trans, trans-sorbo-1,3 myristopalmitin; and 2-trans, trans-sorbo-1,3 dipalmitin. No evidence of asymmetry was found by circular dichroism. The composition and proportions of these triglycerides was found to be species variable.