Isolation of Thymosin α1 from Thymosin Fraction 5 by High Performance Liquid Chromatography

Abstract

An HPLC system for quickly isolating individual peptides from thymus extract is described. A series of reversed phase separations followed by ion-exchange HPLC produces pure peptides with minimal chromatographic time. A 2.4 gram column load of crude extract was carried through to the isolation of 4 mg of pure thymosin α₁ within 6–8 hours of chromatography.     

Separating Ability of Some Polar Mobile Phases in Reverse Phase High Performance Liquid Chromatography

Abstract

Homologous series of n-alkanes, n-alcohols, ethers, esters, amides and ketones were studied as solutes. Methanol, acetonitrile, ethanol, dioxane, 2-propanol and tetrahydrofuran were mobile phases. Spherisorb ODS columns with light and heavy octadecyl coatings were used. Methanol and acetonitrile were found to be the most selective solvents for reverse phase separation. The polar amides, alcohols, and ketones behaved differently from nonpolar solutes in acetonitrile mobile phases. Results were interpreted with the solvophobic theory.     

Separation of T-MAZ ethoxylated sorbitan fatty acid esters by reverse phase chromatography

Summary The method for determination of T-MAZ ethoxylated sorbitan fatty acid esters is described. This work demonstrates that with a less retentive C₈ alkyl bonded phase packing, reverse phase chromatography can be used to analyze nonionic polymer mixtures with a molecular weight range of 900 to 1500. Using a gradient elution, a complete separation of T-MAZ oligomers was achieved, comparable to that obtained by Supercritical Fluid Chromatography (SFC). Isocratic elution is used to quantify T-MAZ and the detection limit is 321 ppm, which is acceptable for polymers with high molecular weights and no UV-absorbing chromophores. This work also shows the comparison of the separations of T-MAZ using gel permeation chromatography and reverse phase chromatography.     

Purification of Commercially Prepared Bovine Trypsin by Reverse Phase High Performance Liquid Chromatography

Abstract

A reverse phase high performance liquid chromatographic method for the purification of milligram amounts of commercially prepared bovine trypsin is described. Increased specific enzymatic activity is observed in the purified material. The advantages of using purified trypsin in protein digestions as well as the potential application of reverse phase high performance liquid chromatography for enzyme characterization are described.     

A Reverse Phase Technique for Separating the Linearc Furano Coumarins in Celery

Abstract

A reverse phase method for separating the four linear furanocoumarins psoralen, xanthotoxin, bergapten, and isopimpinellin in celery is described. The photosensitizing compounds, psoralen, xanthotoxin, and bergapten were also quantified in celery blades and leaves. The method utilizes a phenyl-bonded HPLC column with a methanol-water solvent system. This system allows the use of a detection wavelength (225 nm) more appropriate for these linear furanocoumarins than is currently used with normal phase systems.     

Derivatization of Xanthomegnin for Fluorometric Determination

Abstract

A derivatization procedure was developed for the fluorometric determination of purified xanthomegnin. A highly fluorescent product is formed by reaction of xanthomegnin with ammonia and hydrogen peroxide under defined conditions. The derivative is a light brown compound that can easily be dissolved in water or polar organic solvents. Its excitation and emission wavelengths in methanol are 340 and 445 nm, respectively. As little as 0.1 ng of xanthomegnin can be detected using high pressure liquid chromatography in a reverse phase system with fluorescence detection.     

Dynamic Fluorogenic Labelling of Pesticides for High Performance Liquid Chromatography: Detection of N-Methylcarbamates with o-Phthalaldehyde

Abstract

A post-column fluorogenic labelling reaction between o-phthalaldehyde (OPA) and methylamine, a hydrolysis product of the N-methylcarbamate and carbamoyl Florida Agricultural Experiment Station Journal Series No. 654. oxime pesticides, is described which provides a highly sensitive and selective detection mechanism for the high performance liquid chromatography of these pesticides.

Separations were achieved by reverse phase, with solvent programming of water-dioxane mixtures. As little as 1 ng of the pesticides could be detected, with the last eluting carbamate, mesurol, requiring from 22 to 54 minutes, depending upon column and program contour. Chromatography was performed with a modular instrument and with a commercially available fully cabineted instrument.     

Microdetermination of Heavy Metal Chelates by High-Performance Liquid Chromatography

Abstract

Adsorption chromatography of diethyldithiocarbamate chelates of heavy metals is described. Determination of heavy metals including Hg (II), Cd (II), Pb (II), Cr (VI), Bi (III) and Cu (II) in micro to nano gram order is successfully carried out with good accuracy and reproducibility by a deactivated silica gel column. Separative microdetermination of these chelates is accomplished within 12 min, and selective determination is also possible by choosing the detection wavelength of a UV detector.     

Liquid Chromatographic Analysis of Pentazocine and Tripelennamine in Combination

Abstract

A reverse phase and normal phase liquid chromatographic procedure is described for the separation of pentazocine and tripelennamine. The isocratic methods use dual wavelength detection at 254, 280, and 313 nm.     

Modified silica gels as recyclable adsorbents of aqueous polycyclic aromatic hydrocarbons

ABSTRACT

Polycyclic aromatic hydrocarbons (PAHs), a class of toxic organic molecules containing multiple fused aromatic rings, are found in air, soil and water as a by-product from the incomplete combustion of organic matter. We report on the modification of silica gel using lipophilic molecules containing carboxylic acids, for anchoring to the surface via hydrogen bonds. The lipophilic component captures aqueous PAH, specifically phenanthrene, through the agency of π–π and π–sp³ interactions. The structure–sorption-relationships suggest optimum phenanthrene adsorption is achieved with unsaturated bonds in linear chains or two phenyl rings. Examples include linoleic acid or 2,3-diphenylpropionic acid with removal values of 281 and 283?ng PAH per gram of silica gel, respectively. Saturation adsorption is achieved within four hours. Proposed modes of binding of the new reverse phase silica gels with phenanthrene are described. Recycling of the silica gel is accomplished by washing with organic solvents to remove PAH.     

Separation of unmodified α-amino acid enantiomers by reverse phase HPLC

Summary A novel chiral phase system is presented for the resolution of unmodified α-amino acid racemates which is composed of a reverse phase packing coated with N-alkyl-L-hydroxyproline (where alkyl is n-C₇H₁₅-, n-C₁₀H₂₁- and n-C₁₆H₃₃-) and a hydro-organic eluent containing copper(II)acetate. The factors controlling retention and enantioselectivity such as concentration of Cu(II)ion and pH of the eluent, addition of NH₄Ac to the eluent, type and content of organic solvent of hydro-organic eluent and column temperature were examined. The extremely high enantioselectivity observed (α up to 16) is assumed to be caused by a three site sorbate-sorbent interaction involving bidentate coordination of two amino acids to a Cu(II)ion and hydrophobic attractions between hydrocarbon side chains of amino acids and the n-octadecyl groups of the support. The efficiency and selectivity of the system permits resolution of up to 7 racemic amino acids into enantiomers within 35 minutes on a conventional HPLC apparatus. 13th publication in the series dealing with the ligand-exchange chromatography. Previous publication: ref. [1].     

Microdetermination of Bile Acids by High-Performance Liquid Chromatography Equipped with a High Sensitive Conductance Detector

Abstract

Conductimetric detection of bile acids in reversed phase high-performance liquid chromatography is described. The solvent system of the mixture of water and organic solvent containing small amount of basic salts such as ammonium carbonate is found useable by removing the cation with the cation exchange column inserted between the ODS column and the conductance detector. Thus, a few ng of tauro-and glyco-conjugated bile acids can be detected without tedious derivatization and hydrolysis.     

The Preparative Scale Reverse Phase HPLC Separation of Epimeric Alkaloids Using Camphorsulfonic Acid as an Ion Pairing Reagent

Abstract

A reverse phase paired ion HPLC procedure is described for the separation of multigram quantities of epimeric alkaloids using camphorsulfonic acid as the ion-pairing reagent.     

Synthesis,structure and properties of new chiral liquid crystalline monomers and homopolysiloxanes containing menthyl groups

The synthesis is described of four new chiral liquid crystalline monomers (M₂–M₅ ) and their corresponding side‐chain homopolysiloxanes (P₂–P₅ ) containing menthyl groups. Chemical structures were characterised using FT‐IR or ¹H NMR spectra, and specific optical rotations were evaluated with a polarimeter. The phase behaviour and mesomorphic properties of the new compounds were investigated by differential scanning calorimetry, thermogravimetric analysis, polarising optical microscopy, UV/visible/NIR spectrocopy and X‐ray diffraction. The monomers and homopolymers with more aryl segments showed noticeably lower specific optical rotation value. The monomers M₂–M₅ formed a cholesteric or blue phase when a flexible spacer was inserted between the rigid mesogenic core and the terminal menthyl groups by reducing the steric effect. M₂–M₅ revealed enantiotropic cholesteric phase. Moreover, M₂ also exhibited a monotropic smectic A (SmA) phase, and M₄ also exhibited a cubic blue phase on cooling. The selective reflection of light shifted to the long wavelength region with increasing rigidity of the mesogenic core for M₂–M₅ . P₂–P₅ exhibited SmA phases, and the mesogenic moieties were ordered in smectic orientation with their centres of gravity in planes. Melting or glass transition temperature and the clearing temperature increased, and the mesophase temperature range widened with increasing rigidity of the mesogenic core.     

A Reverse Phase HPLC Method for the Determination of Chloramphenicol and Its Hydrolytic Product in Ophthalmic Solutions

Abstract

A reverse phase HPLC procedure is described for the determination of chloramphenicol and its hydrolytic product, 2-amino-1-(4-nitrophenyl)-propane-1,3-diol. The method was validated for reproducibility, linearity and recovery for both components. It was successfully applied to monitor post-marketing stability of ten batches of chloramphenicol ophthalmic solutions stored at room temperature. As compared to the BP method using solvent extraction and UV measurements, for the determination of chloramphenicol and its hydrolytic product in chloramphenicol eye drops, the suggested procedure was found to be relatively more accurate, precise and less time-consuming.     

Separation of Hydrocarbon Mixtures in the Microgram Range by Inclusion in a Urea-packed Mini Column

Abstract

The use of classical urea inclusion techniques for the separation of straight chain hydrocarbons from branched and cyclic compounds is satisfactory when applied to mixtures in the milligram to gram ranges, but leads to low separation efficiencies when quantities in the microgram to milligram range are involved. In this study, a modified inclusion technique using an urea-packed milli-bore column and a catalytic eluent is described. Examples of its use for the separation of mixtures of linear and cyclic hydrocarbons from 30μg up to a few milligrams are given. The versatility of this technique for the analyses of low amounts of environmental samples is described, and an application to the hydrocarbon fraction of surface sediment from lake Leman (Switzerland) is presented.     

Quantitative Analysts of 5-Hydroxytryptamtne in Biological Material by High Perfornance Liquid Chromatography and Electrochmical Detection

Abstract

A quantitative method for the analysis of 5-hydroxytryptamine in biological material is described. The method is based on high performance liquid chromatography (HPLC) with electrochemical detection. A simple purification on a weakly acidic ion exchange resin prior to the analysis gives quite clean samples and permits concentration of diluted samples. The chromatographic separation is performed on a reverse phase column with organic modifier added to an aqueous eluent. With this analytical system 25 pg of 5-hydroxytryptamine can be detected.     

Optimization of the isocratic non-aqueous reverse phase (NARP) HPLC separation of trans/cis-α-and β-carotenes

Non-aqueous reverse phase chromatography was used to optimize the separation of all-trans-α- and β-carotenes and their cis isomers extracted from dried carrots. The technique can easily be applied to routine assays. Optimization was carried out by modifying the concentrations of methylene chloride and acetonitrile, and by studying the influence of temperature on the capacity factors and selectivity. The effect of the type of C 18 stationary phase bonding (mono- and polyfunctional) on separation was also examined. The need for polymeric bonding on the stationary phase is stressed for the realization of the trans/cis isomer separation.     

Liquid Chromatographic Determination of Cationic Surfactants in Environmental Samples using a Continuous Post-Column Ion-Pair Extraction Detector with a Sandwich Phase Separator

Abstract

A new detection technique is described for the quantitative analysis of cationic surfactants by HPLC via post-column ion-pair formation. A new sandwich type phase separator, as part of the extraction detector, was successfully introduced. The method was used to determine ditallowdimethylammonium chloride (DTDMAC) in various environmental samples. Detection limits of DTDMAC in river water were about 2 μg/1 (60 ng absolute; S/N = 5) and 10 ng/1 (260 pg absolute; S/N = 5), using methyl orange and 9,10-dimethoxyanthracene-2-sulphonate (DAS) as ion-pairing reagents, respectively. The environmental concentration of DTDMAC found on random samples from two Belgian rivers range from 30 to 40 μg/1. The reproducibility of the determination of DTDMAC in river water was 4.2% (RSD) (n = 20).