Recycling HPLC of p-Bromophenacyl Esters of Saturated C35–56 Fatty Acids from Mycobacterium Tuberculosis H37Ra on a Silica Column

Abstract

The p-bromophenacyl esters of saturated C_35–56 fatty acids from Mycobacterium tuberculosis H37Ra were separated according to structural classes on a silica column by high performance liquid chromatography (HPLC). The sample was cycled five times during HPLC. Highly purified C_35–38 esters were obtained by this method. Further HPLC fractionation on a reverse-phase column (C₁₈-bonded silica) gave complete separation of most of the remaining fractions. By combining mass spectrometry with HPLC separations, many of the fatty acid esters were tentatively identified.     

Separation of Fatty Acids as Their p-Bromophenacyl Esters on a C30-Bonded Silica Column by High Performance Liquid Chromatography

Abstract

A new C₃₀-bonded silica column was developed for high performance liquid chromatography. This column was tested for the fractionation of fatty acids as their p-bromophenacyl esters by the reverse-phase mode. Certain pairs of fatty acid esters that are very difficult to separate on a C₁₈-bonded silica column, i. e., arachidonic (C₂₀:4)-palmitoleic (C₁₆:1); elaidic (trans C₁₈:1)-vaccinic (cis C₁₈:1); behenic (C₂₂:0)-nervonic (C₂₄:1); and arachidonic (C₂₀:0)-erucic (C₂₂:1) esters, were completely resolved on the C₃₀-bonded column using solvent gradients of acetonitrile: water and acetonitrile: p-dioxane. A solvent system of methylene chloride: acetonitrile (2:1, v/v) was developed for this column to achieve good separation of a homologous series of extremely nonpolar C₇₆ to C₈₂ α-mycolic acid esters from Mycobacterium tuberculosis H37Ra.     

Sampling and quantitation of lipids in aerosols from the remote marine atmosphere

A procedure is described for the determination of organic compounds in aerosols and rain from the remote marine atmosphere. Five classes of naturally occuring lipids (n-alkanes, was esters, fatty alcohols, sterols, and fatty acids) were quantified in the samples. Air samples (4000–10 000 m³) were collected on glass-fiber filters under authomatic control. Rain samples (1–5 l) were collected on an event basis. Filter and rain samples were extracted with dichloromethane and the extracts were fractionated into discrete chemical classes by adsorption chromatography with silica gel. The fractions were derivatized (if necessary) and quantified by high-resolution glass-capillary gas chromatography (HRGC) and HRGC/mass spectrometry. A second filter extraction was required for quantitation of fatty acid salts. On-column injection of the fractions provided identification and quantification of a wide range of homologs within each compound class: C₁₅–C₄₄ for n-alkanes, C₃₆–C₆₂ for was esters, and C₁₃–C₃₆ for fatty alcohols and fatty acids. Internal standards were used to quantify recoveries and concentratins. Mean recoveries relative to the internal standares were 96.5% for C₁₅–C₃₆ n-alkanes, 96.4% for C₂₀–C₃₀ n-fatty acids, 92.5% for C₁₄–C₃₀ n-fatty alcohols and 93.3% for cholesterol. The procedural blanks for the remote marine aerosol samples allow detection limits of 0.1–1.0 pg m^?3 for most compounds. These values are lower than any other method used at coastal marine, rural or suburban sampling locations.     

Synthesis of 4‐(7‐diethylaminocoumarin‐3‐yl)benzeneisocyanate (DACB‐NCO): A highly sensitive fluorescent derivatization reagent for alcohols in high‐performance liquid chromatography

4‐(7‐Diethylaminocoumarin‐3‐yl)benzeneisocyanate (DACB‐NCO) was synthesized as a new fluorescent derivatization reagent for alcohols for use in high‐performance liquid chromatography (hplc). Saturated alcohols (C₆‐C₂₂) were derivatized in good yields into the corresponding fluorescent DACB‐carbamic esters by treating with DACB‐NCO. The DACB‐carbamic esters of these alcohols were clearly separated on a reversed‐phase hplc column (Inertsil ODS‐2, mobile phase: methanol‐water, excitation wavelength 402 nm; emission wavelength 488 nm). The detection limit (S/N = 3) of cetyl alcohol, as a test compound, was 5 fmol/10 μl.     

Chiral Separation of Racemic Tetrahedral Transition Metal Carbonyl Cluster SFeCoMo(CO)8(C2H5OCOCp) by Liquid Chromatography on Tribenzoylcellulose

Abstract

Direct separation of racemic tetrahedral transition metal carbonyl cluster SFeCoMo(CO)₈(C₂H₅OCOC_p) has been conducted by liquid chromatography using tribenzoylcellulose as chiral stationary phase. The effects of concentration of 2-propanol, various primary alcohols in the mobile phase and column temperature on the rentention and enantioselectivity of SFeCoMo(CO)₈(C₂H₅OCOC_p) cluster enantiomers were studied. The maximum resolution factor obtained was 2.306 when hexane/2-propanol(99.5/0.5,v/v) was used as solvent at 23°C.     

Isocratic Separation of Fatty Acid Derivatives by Reversed Phase Liquid Chromatography. Influence of the Solvent on Selectivity and Rules for Elution Order

Abstract

The p-bromophenacyl esters of 16 fatty acids (C₁₂-C₂₂) have been separated by isocratic chromatography on a Radial Pak A cartridge (Reverse phase C₁₈ material). The separation factors α were measured using two solvent mixtures of comparable strength and the superiority of methanol-water to acetonitrile-water becomes evident.

Five precise rules are established, which indicates the retention of every fatty acid. They explain the chromatographic process i.e. elution order, resolution and selectivity.     

HPLC of fatty acid esters of mono- and polyhydric alcohols. Part 1: Analytical separation

This paper shows how simply and yet very rapidly fatty acid esters of monohydric alcohols, but particularly partial and full fatty acid esters of fully hydric alcohols can be separated and determined by means of high-pressure liquid chromatography on LiChrosorb RP-8 with methanol/water. We have separated quantitatively the methylesters of the fatty acids C_8:0 to C_22:0 and C_24:0, the i-propyl-, i-butyl-, n-hexyl- and i-octyl-esters of the even-numbered fatty acids C_8:0 to C_18:0, mono- and difatty acid esters of the 1,3-bis-(2-hydroxyethyl)-5,5-dimethyl-hydantoin, mono-, di- and triesters of the trimethylolpropane as well as the tetraesters of the penta-erythrite.     

Thin-Layer Chromatography of Bile Alcohols,Bile Acids and Conjugated Bile Acids

Abstract

Thin-layer chromatography of bile alcohols, bile acids and bile acid conjugates has been reviewed. Particular emphasis has been placed on the separation of the glycine and taurine conjugated bile acids as a class and as individual compounds, and on the isolation of bile alcohols and C₂₇ bile acids diastereo-isomeric at C-25.     

A simple and mild derivatization method for hydroxyl compounds using condensation agent carbazole-9-N-(2-methyl)-acetyl-benzene-disulfonate and its application for the determination of volatile alcohols from plasma by HPLC with fluorescence detection

A simple and sensitive HPLC method for the determination of hydroxyl compounds including volatile alcohols from spiked plasma, using a fluorescent condensation agent, namely carbazole-9-N-(2-methyl)-acetyl-benzene-disulfonate (CMABS), has been developed. A mixture of alcohols and pyridine in dichloromethane was treated with CMABS to give a quantitative yield of esters. The maximum fluorescence emissions for the derivatized alcohols are at 365 nm (λ_ex 335 nm). Studies on derivatization conditions indicate that alcohols react very fast with CMABS in presence of pyridine in dichloromethane to give the corresponding fluorescent derivatives. The method, in conjunction with a multi-step gradient, offers a baseline resolution of the common alcohol derivatives on a reversed-phase C₁₈ column, which is more convenient and more efficient than previous methods which require the prior conversion of the carboxylic acids to the acyl chlorides. The application of this method to the analysis of volatile alcohols in plasma was also investigated. The LC separation shows good selectivity and reproducibility for alcohol derivatives. The relative standard deviations ¶(n = 5) for 100 pmol of each alcohol are < 4%. The detection limits are at the fmol level for C₁–C₃ alcohols and pmol level for C₄–C₉ alcohols.     

A simple and mild derivatization method for hydroxyl compounds using condensation agent carbazole-9-N-(2-methyl)-acetyl-benzene-disulfonate and its application for the determination of volatile alcohols from plasma by HPLC with fluorescence detection

A simple and sensitive HPLC method for the determination of hydroxyl compounds including volatile alcohols from spiked plasma, using a fluorescent condensation agent, namely carbazole-9-N-(2-methyl)-acetyl-benzene-disulfonate (CMABS), has been developed. A mixture of alcohols and pyridine in dichloromethane was treated with CMABS to give a quantitative yield of esters. The maximum fluorescence emissions for the derivatized alcohols are at 365 nm (λ_ex 335 nm). Studies on derivatization conditions indicate that alcohols react very fast with CMABS in presence of pyridine in dichloromethane to give the corresponding fluorescent derivatives. The method, in conjunction with a multi-step gradient, offers a baseline resolution of the common alcohol derivatives on a reversed-phase C₁₈ column, which is more convenient and more efficient than previous methods which require the prior conversion of the carboxylic acids to the acyl chlorides. The application of this method to the analysis of volatile alcohols in plasma was also investigated. The LC separation shows good selectivity and reproducibility for alcohol derivatives. The relative standard deviations ?(n = 5) for 100 pmol of each alcohol are < 4%. The detection limits are at the fmol level for C₁–C₃ alcohols and pmol level for C₄–C₉ alcohols. Received: 9 November 1998 / Revised: 3 May 1999 / Accepted: 6 May 1999     

Alcohol separation by thin-layer chromatography. II

Summary Thin-layer chromatography on rice starch, maize starch, cellulose and talc makes possible the separation of the homologous series of 3,5-dinitrobenzoates of aliphatic alcohols C₁ to C₂₀. Six different solvent systems have been employed. Dependence of R _f-values on the number of C-atoms on various supports is discussed.Part I: Fresenius Z. Anal. Chem. 264, 415 (1973)     

Liquid-Chromatographie von Alkoholen auf mikrokristalliner Cellulose

In “liquid chromatography” (LC), a stationary phase of microcrystalline cellulose is suitable for the separation of polyhydric alcohols such as glycerine, ethylene glycol, propane diol and butane diol when using a solvent mixture of ethyl acetate and n-propanol, saturated with approx. 15% of water. When using benzene as solvent, the monohydric alcohols C₁-C₅ can also be separated on the same column. Differential refractometers and micro adsorption detectors are suitable for detection.     

Analysis of high molecular weight esters and ketones in thymus

A crystalline mixture of esters from Thymus chamaedris is analyzed by mass spectrometry, and after hydrolysis, by gas chromatography. Two series of straight chain even aliphatic acids from C₁₆ to C₂₆, and from C₄₀ to C₄₈, and a series of straight chain even saturated primary alcohols, from C₈ to C₃₀, are its main constituents. Another crystalline mixture, isolated from Thymus caespititius, is shown by mass spectrometry and gas chromatography to be composed of a straight chain series of aliphatic ketones, from C₂₅ to C₃₇.     

A novel pillar[3]trianglimine macrocycle with a deep cavity used as a chiral selector to prepare a chiral stationary phase by thiol-ene click reaction for enantioseparation in high-performance liquid chromatography

A chiral pillar[3]trianglimine (C₆₀H₇₂N₆O₆) with a deep cavity has been developed as a chiral selector and bonded to thiolated silica by thiol-ene click reaction to fabricate a novel chiral stationary phase for enantioseparation in high-performance liquid chromatography. The enantioseparation performance of the fabricated chiral stationary phase has been evaluated by separating various racemic compounds, including alcohols, esters, amines, ketones, amino acids, and epoxides, in both normal-phase and reversed-phase elution modes. In total, 14 and 17 racemates have been effectively separated in these two separation modes, respectively. In comparison with two widely used chiral columns (Chiralcel OD-H and Chiralpak AD-H), our novel chiral stationary phase offered good chiral separation complementarity, separating some of the tested racemates that could not be separated or were only partially separated on these two commercial columns. The influences of analyte mass, mobile phase composition, and column temperature on chiral separation have been investigated. Good repeatability, stability, and column-to-column reproducibility of the chiral stationary phase for enantioseparation have been observed. After the fabricated column had been eluted up to 400 times, the relative standard deviations (n = 5) of resolution (Rs) and retention time of the separated analytes were < 0.39% and < 0.20%, respectively. The relative standard deviations (n = 3) of Rs and retention time for column-to-column reproducibility were < 4.6% and < 5.2%, respectively. This study demonstrated that the new chiral stationary phase has great prospects for chiral separation in high-performance liquid chromatography.     

Separation of Long And Short Chain Fatty Acids as Naphthacyl and Substituted Phenacyl Esters by High Performance Liquid Chromatography

Abstract

High performance liquid chromatography of various C₂ - C₂₄ fatty acids was performed with their p-bromophenacyl, p-nitrophenacyl, p-chlorophenacyl, and 2-naphthacyl esters. All separations were accomplished with two types of reversed phase columns with the eluent consisting of an acetonitrile: water gradient. For all derivatives tested, the separations were well defined and analogous, although certain esters eluted together as single peaks. Quantitative results indicate that the limit of detection in the present study is two picograms of n-caproic acid and 10 picograms of arachidonic acid as their p-bromophenacyl esters.     

A Rapid Isolation of Human Chorionic Gonadotropin and Its Subunits by Reversed-Phase High Performance Liquid Chromatography

Abstract

A rapid isolation of human chorionic gonadotropin and its subunits from a commercially available concentrate of human urine has been achieved using reversed-phase high performance liquid chromatography. With μBondapak C₁₈ columns and a gradient employing aqueous trifluoroacetic acid as one solvent and dilute trifluoroacetic acid in acetonitrile as the other, complete separation can be accomplished in one day whereas standard column chromatographic procedures take about two weeks. Specific radioimmunoassays, polyacrylamide gel electrophoresis, and amino acid analyses were used to identify and characterize chromatographic peaks.     

High Performance Liquid Chromatography of Cholesteryl Esters

Abstract

The separation of cholesteryl esters by high performance liquid chromatography was facilitated by cooling of the column, followed by spontaneous return to ambient conditions. the resolution of the esters was increased. the method was applied to detection of the cholesteryl esters in plasma.     

Separation of Xanthine Derivates by High Pressure Liquid Chromatography and Application to Plasma Analysis

Abstract

High pressure liquid chromatography (HPLC) methods were developed for separation and plasma analysis of ten xanthine derivates. Separation was evaluated on silica column and on three different reverse phase columns, with optimum conditions obtained on C₆ spherisorb column using isocratic elution with phosphate buffer 10^?2 M, pH 2.7 - acetonitrile mixture (80/20 V/V). Determination of these xanthine derivates in plasma for therapeutic control was studied.     

Chromatographic Separation on Silica of Polar Aromatic Compounds II) Research of the Best Chromatographic System by Different Preloading in Thin Layer for a Transposition on Column

Abstract

A high performance thin layer chromatographic method for aromatic compounds separation with absorbant deactivation has been given in the precedent work. Our aim was to transpose the results obtain from HPTLC to HPLC. The two types of results were then related by a transposition coefficient, K_tR which interprets the geometrical variations of the two chromatographic methods. The values obtained for K_tR represent fairly well the linear relation between the retentions in thin layer chromatography and those in column chromatography. We have thus desmontrated the possibility of modifying adsorbent activity in the same manner in thin layer as in column chromatography by the fixation of chemical compounds, thus allowing the separation of relatively polar compounds.     

The Purification of Xanthene Dyes by Reverse Phase High Performance Liquid Chromatography

Abstract

A reliable method for the separation of fluorescein dyes from their impurities was developed using high performance liquid chromatography and involved a μBondapak C₁₈ reverse phase column and mixtures of methanol and ammonium acetate buffer. This technique was used to verify the purity of commercial products as well as to aid in the development of an empirical theory related to retention of halogenated fluorescein dyes by reverse phase columns.