Gy. Vigh

Influence of temperature on the retention behaviour of members of homologous series in reversed-phase high-performance liquid chromatography

Abstract The capacity factors of C 6 C 16 n -alkanols, C 6 C 12 n -alkanal dinitrophenylhydrazones and C 6 C 12 2- n -alkanone dinitrophenylhydrazones have been determined at various temperatures on MicroPak CH-10 and Nucleosil C-18 octadecyl reversed-phase packings with aqueous methanol eluents. Linear log k′ vs. C n log k′ vs. I/ T , −Δ H ° vs. C n and log k′ vs. −Δ H ° relationships have been obtained. Equations are presented that describe the capacity factors as functions of both temperature and carbon number. The compensation temperatures determined according to Melander et al. indicate that the retention mechanisms underlying these reversed-phase separations are identical.

Studies in reversed-phase ion-pair chromatography. IV: The role of the chain length of the pairing ion

Abstract Adsorption isotherms of straight chain sodium alkanesulphonates (from butane to octane) were determined by the breakthrough method in aqueous phosphate buffer on ODS-Hypersil, along with retention data of similarly and oppositely charged ionic and non-ionic solutes. The effects on the solute retention of the chain length of the pairing ion were examined both at varying and constant counter ion (sodium) concentrations, in terms of both the mobile phase and stationary phase pairing ion concentrations. A practical consequence of the use of systems with constant counter ion concentration is that pairing ions with different chain lengths (members of a homologous series) are interchangeable in terms of their surface concentrations.

Studies in reversed-phase ion-pair chromatography. I: Adsorption isotherms of tetraalkylammonium ion-pair reagents on LiChrosorb RP-18 in methanol-water eluents

Abstract The adsorption isotherms of symmetrical tetraalkylammonium ion-pair reagents (tetramethyl, tetraethyl, tetrapropyl and tetrabutyl) on LiChrosorb RP-18 in 25 mM H3PO4NaH2PO4 aqueous buffer were determined from breakthrough curves at a constant Br− counter ion concentration of 200 mM, maintained by the addition of sodium bromide. The Langmuir and Freundlich isotherms were tested with the data and a poor fit was observed at higher mobile phase concentrations. The adsorption isotherms of tetrabutylammonium bromide were measured in the same system, in 0–70% (v/v) methanol—aqueous buffer eluents, at seven eluent compositions. Stationary phase ion-pair reagent concentration versus methanol concentration (at constant mobile phase pairing ion concentrations), and stationary phase versus mobile phase pairing ion concentration (at constant methanol concentrations) plots were obtained. With these plots, pre-selected stationary phase ion-pair reagent concentrations can be set (either held constant or varied according to a programme) under widely varying chromatographic conditions.

Semi-preparative high-performance reversed-phase displacement chromatography of insulins

A reversed-phase displacement chromatographic method has been developed for the purification of bovine and porcine insulin samples. Up to 500 mg of raw insulin could be purified on a Nucleosil C8 analytical column using a methanol-containing phosphate buffer carrier and a cetrimide-containing displacer. A proinsulin contamination level as low as 100 ppm in the collected fractions could be achieved.

Studies in ion-pair chromatography : II. Retention of positive and negative ions and neutral solutes in tetrabutylammonium bromide-containing methanol—water eluents on lichrosorb rp-18

Abstract The retention of positively and negatively charged ions, as well as of neutral solutes, was studied in 0–70% methanol in 25 mM aqueous phosphate buffer (pH 2.1–3.4) eluents as a function of the tetrabutylammonium bromide ion pair reagent concentration. The log k′ vs. mobile phase pairing ion concentration (Pm) plots (which showed Pm- and [methanol]-dependent maxima) could be transformed into log k′ vs. stationary phase pairing ion concentration (Ps) curves by using the simultaneously determined adsorption isotherms of tetrabutylammonium bromide. While the shape of the log k′ vs. Pm curves of oppositely charged solutes changed considerably with the concentration of methanol in the eluent, the log k′ vs. Ps curves were more similar, and their maxima are in the 80–120 μmole/g range. The log k′ vs. Pm curves of similarly charged solutes were more alike. The methanol concentration of the eluent influences the retention of oppositely charged solutes more sensitively than that of the similarly charged solutes. At constant Pm the log k′ vs. [methanol] curves of oppositely charged solutes were S-shaped, while the log k′ vs. [methanol] curves at constant Ps values ran similar to the log k′ vs [methanol] curves obtained in the absence of ion-pairing reagents.

Effect of the type of ion-pairing reagent in reversed-phase ion-pair chromatography

SummaryThe retention of positively and negatively charged and neutral solutes was studied in an aqueous phosphate buffer eluent, with constant inorganic-counterion concentration, on ODS-Hypersil stationary phase in the presence of various sulfonic acid ion-pairing reagents. The adsorption isotherms of the ion-pairing reagents d-camphor-10-sulfonic acid, sodium cyclohexylsulfamic acid and sodium alkylsulfonates (butyl-, hexyl- and octyl-) were determined by the breakthrough method.

Role of buffer cations in the reversed-phase high-performance liquid chromatography of aromatic amines.

Abstract The role of buffer cations in the retention of aromatic amines in reversed-phase high-performance liquid chromatography was investigated. A methanol-rich eluent was used with a LiChrosorb RP-18 column. The eluent contained varying amounts of H3PO4, NaH2PO4, NaClO4, KClO4 tetramethylammonium (TMA+) chloride. Log k′ of aromatic amines at constant pH decreases linearly with increasing log (cation concentration). The retention-decreasing effect of inorganic buffer cations increases in the order H+

Artifact-free matrix-assisted laser desorption ionization time-of-flight mass spectra of tert.-butyldimethylsilyl ether derivatives of cyclodextrins used for the synthesis of single-isomer, chiral resolving agents for capillary electrophoresis

Artifact-free, high-resolution matrix-assisted laser desorption ionization (MALDI) time-of-flight mass spectra have been obtained for the labile, single-isomer, tert.-butyldimethylsilyl ether derivatives of alpha-, beta- and gamma-cyclodextrins by optimizing the MALDI sample preparation method. 2,5-Dihydroxybenzoic acid, a 3:1 mixture of 2,5-dihydroxybenzoic acid and 1-hydroxyisoquinoline, and 2,4,6-trihydroxyacetophenone were investigated as MALDI matrices with methanol and acetonitrile as matrix solvents. Partial-to-complete loss of the tert.-butyldimethylsilyl groups was observed when the commonly used 2,5-dihydroxybenzoic acid was the MALDI matrix and/or methanol was the solvent, both with and without trifluoroacetic acid as additive. Loss of the labile tert.-butyldimethylsilyl groups was avoided with 2,4,6-trihydroxyacetophenone as MALDI matrix and acetonitrile as matrix solvent. Good ion intensities were achieved for the (M+Na)+ and (M+K)+ quasimolecular ions in the positive-ion mode. Minor byproducts were observed in some of the samples and the information was used to aid the optimization of the synthetic work.

Determination of chlorophacinone in formulations by reversed-phase ion-pair chromatography

Abstract The retention behaviour of chlorophacinone [2-(p-chlorphenyl)phenylacetyl-1,3-indandione] has been studied using an octadecylsilica column and different eluents, e.g., methanol-water, methanol-aqueous phosphoric acid, methanol-aqueous McIlvaine buffer-tetramethylammonium bromide and tetrahydrofuranaqueous McIlvaine buffer-tetrabutylammonium bromide. The dependence of log k′ on the organic modifier content and the apparent pH of the eluents was determined. The retention behaviour could be interpreted by assuming that chlorophacinone existed mostly as a weakly acidic enol that could dissociate depending on the pH of the eluent. Chlorophacinone could be separated and determined quantitatively in 0.25–25% (w/w) oily formulations and in 10–100 ppm crushed grain bait using diphacinone [2-(diphenylacetyl)-1,3-indandione] as an internal standard.

Preparation of high-performance liquid chromatographic columns from glass-lined stainless-steel tubing

Abstract High-performance liquid chromatographic columns were prepared by packing nominally 10/2-20-μm silica into glass-lined stainless-steel columns, employing the so-called high-viscosity packing method. Cyclohexanol was used to prepare the silica slurry, containing 11% (w/w) of solid material. The slurry was pumped into the columns at 6000 p.s.i. A similar process was used to prepare columns made of precision-bore stainless-steel tubing. The columns were tested, and those made of glass-lined tubing were found to be about 1.5 times more efficient than those prepared under identical conditions but made of precision-bore stainless-steel tubing. Scanning-electron microscopic photographs revealed that the inner wall of the former columns was smoother than that of the latter and was free of deep axial gloves, and this is believed to be the main factor that gives more efficient columns.