Syntheses of spherical silica and titania from alkoxides on a laboratory scale

Summary A process to synthesize silica and titania as spherical packing materials has been investigated on the laboratory scale by the sol-gel method. The silica and titania obtained were tested under normal-phase separation conditions for comparison of their retention characteristics silica witha a commercial. The silica was found to be similar in its retention behaviour to the commercial silica. The titania showed basic properties and strongly retained acidic compounds.     

A novel silica-based column packing for reversed-phase HPLC of basic compounds

Summary A novel bonded phase for reversed-phase HPLC was synthesized in two steps. Octylamine was first reacted with β-(3,4-epoxycyclohexyl)ethyltrimethoxysilane (β-ECTS) and then the intermediate product was coupled onto porous silica. The prepared packing was characterized by elemental analysis, solid-state¹³C NMR and Fourier transform infrared (FT-IR). Chromatographic evaluations were carried out by using a mixture of organic compounds including acidic, basic and neutral analytes and methanol-water as binary mobile phase. The results showed that the stationary phase has excellent chromatographic properties and is resistant to hydrolysis between pH=2≈8. It can be used efficiently for the separation of basic compounds.     

New polymer coated anion-exchange HPLC-phases: Immobilization of poly (2-hydroxy, 3N-ethylenediamino) butadiene on silica and alumina

Summary PHEB=POLY(2-hydroxy, 3N-ethylenediamino)butadiene has been synthesized. This unsaturated pre-polymer can be immobilized on different, preferably inorganic supports of suitable porosity such as silica or alumina by cross-linking. Weak anion-exchange phase are obtained. Such phases do not have the disadvantages of organic polymeric phases, regarding pressure stability and swelling by solvent influence. The ion-exchange capacity and the retentivity of IE-separations can deliberately be varied via the thickness of the cross-linked polymer layer. Alumina can also be coated with PHEB applying the same cross-linking procedure. Such phases exhibit an exceptional chemical stability even when operated with mobile phases at very high pH-values. Separations of excellent selectivity are achieved for inorganic anions, as well as with mixtures of aliphatic and aromatic free acids also including hydroxy- (mono- or poly)carboxylic acids.     

HPLC-separation of enantiomers using quinine,covalently bonded to silica as stationary phase

Summary Stationary phases for chiral separations have been synthesized by chemical modification of porous small particle silica using new procedures of fixation of the chiral moiety. So called pre-polymers of the methylpolysiloxane polysiloxane type are immobilized on silica surfaces by different procedures. These pre-polymers are substituted by chiral groups. Their synthesis is done externally i.e. not in-situ on the support surface. The immobilization on the silica surface is achieved by crosslinking and/or by chemical bonding. Anchor groups within the pre-polymer (e.g. SiH) as well as on the silica surface (SiOH) give rise to the chemical bonding therewith. Chiral phases with quinine as the chiral moiety were obtained which show high separation efficiency as well as chemical stability, in addition to the enantiomeric selectivity required for the separation of certain types of aliphatic and aromatic alcohols. Presented at the 16th International Symposium on Chromatography, Paris, September 1986     

The evaluation of “reversed phase” high-performance liquid chromatography packing materials

Summary For the evaluation of reversed phase packing materials a mixture of acetylacetone, I-nitronaphthalene and naphthalene is proposed. This will reveal the usual optimum kinetic chromatographic parameters (the naphthalene peak), the degree of activity or endcapping status of the column (the ratio of the I-nitronaphthalene and naphthalene retention times) and trace metal activity (the shape and intensity of the acetylacetone peak).     

Retention characteristics of octadecylsilyl silica,trimethylsilyl silica and phenyldimethylsilyl silica in reversed-phase liquid chromatography

Summary The hydrophobic retention characteristics of stationary ligands for reversed-phase high-performance liquid chromatography have been evaluated from the slope (r-value) of the plots relating the capacity factors (log k) of selected aliphatic and aromatic compounds with the reciprocal of methanol concentration (log (1/[MeOH])) in aqueous mobile phase. Octadecylsilyl (ODS), trimethylsilyl (TMS) and phenyldimethylsilyl (phenyl) groups were selected as the stationary ligands bonded to silica support.On ODS or TMS silicas, unlike on phenyl silica, aliphatic compounds gave slightly larger r-values than aromatic compounds, indicating that the shape of the ligand recognizes the hydrophobic surfaces of aliphatic and aromatic solute molecules. On TMS and phenyl silicas, the degree of solute hydrophobicity contributing to its retention is about 90% and 85% of that on ODS silica, respectively. On the other hand, on TMS and phenyl silicas, the polar functional group on the solute molecule brought about a smaller decrease in retention than on ODS silica.     

Characterisation of RPLC columns packed with porous sub-2 microm particles

Eight commercially available sub-2 microm octadecyl silane columns (C18 columns) have been characterised by the Tanaka protocol. The columns can be grouped into two groups that display large differences in selectivity and peak shape due to differences in hydrophobicity, degree of surface coverage and silanol activity. Measurements of particle size distributions were made using automated microscopy and electrical sensing zone measurements. Only a weak correlation could be found between efficiency and particle size. Large differences in column backpressure were observed. These differences are not related to particle size distribution. A more likely explanation is differences in packing density. In order to take full advantage of 100-150 mm columns packed with sub-2 microm particles, it is often necessary to employ not only an elevated pressure but also an elevated temperature. A comparison between columns packed with sub-2, 3 and 5 microm versions of the same packing indicates potential method transferability problems for several of the columns due to selectivity differences. Currently, the best alternative for fast high-resolution LC is the use of sub-2 microm particles in combination with elevated pressure and temperature. However, as shown in this study additional efforts are needed to improve transferability as well as column performance.     

On the chromatography mechanism of reversed-phase liquid chromatography

Summary An equation is derived which can describe how the retention of solutes is influenced by the composition of the mobile phase in reversed-phase liquid chromatography, the retention of solutes in alkyl bonded stationary phase regarded as the complexation between solute molecule and the active sites on the surface of the stationary phase. When the stationary phase is not fully saturated by the organic modifier, the activity of the active sites, the activity coefficient of the adsorbed solute as well as the activity coefficient of the solute in the mobile phase depend on the composition of the mobile phase. However, when the stationary phase is fully saturated, the composition of the mobile phase mainly influences the activity coefficient of the solute in the mobile phase. In addition, the selectivity of retention is discussed in terms of the derived equation.     

Thiophenepropyltrichlorosilane as chemical modifier on silicagel for reversed phase chromatography

Summary Modification of silica gel with thiophene-propyl-trichlorosilane is described as well as the synthesis of the silane. The chromatographic performance and selectivity are demonstrated with various polyaromatic compounds. Thiophene-modified silica gel was found to exhibit a selectivity comparable with both ODS-and amino-phases.     

Determination of pyridate and its main metabolite CL-9673 in plant materials

Summary The application of a simple reversed-phase, HPLC-UV method for the determination of the contact herbicide pyridate and its main metabolite 3-phenyl-4-hydroxy-6-chloropyridazine-known as CL-9673-in green cereals, grains and straw is described. The method is based on a simple extraction, separation of pyridate from its metabolite, hydrolysis of pyridate to CL-9673, high-performance liquid chromatographic separation of CL-9673 in both fractions using only one reversed phase column and UV-detection.The method is sensitive down to 3 ngg^–1 CL-9673 in green plants and grain and 5ngg^–1 in straw.In the range 0.05–1.0mgkg^–1 recoveries are between 73.7% and 109% from green plants, between 77.7% and 102% from grains and between 77.3% and 87.5% from straw.     

High-performance liquid chromatographic resolution of enantiomers on chiral amine-bonded silica gel

Summary A chiral stationary phase with an immobilized, optically active diamine was prepared for the separation of enantiomers. The synthesis of the phase was carried out by bonding (–)trans-1,2-cyclohexanediamine to microparticulate silica gel through the coupling agent 3-glycidoxypropylsilane. The resolution of the racemic compounds catechin, 2,2-dihydroxy-1,1binaphthyl and trans-1,2-cyclohexandiol, is reported.     

Separation of hordein proteins from european barley by high-performance liquid chromatography: Its application to the identification of barley cultivars

Summary High-performance liquid chromatography using either reversed phase or anion-exchange techniques was used for the fractionation of hordein proteins extracted from European barley. A reversed phase method is presented which utilises an Ultrapore column packed with a wide pore (30 nm) C-3 alkylbonded silica support. Using this method up to 20 components may be separated in 54 min. Elution profiles were found to be reproducible. A further method using rapid anion-exchange chromatography indicated that up to 13 components may be separated, a number which is comparable to that found with electrophoresis. The separation of proteins extracted from different barley cultivars indicated that on the basis of elution profiles high-performance liquid chromatography using either reversed phase or anion-exchange offers considerable potential as a method for barley cultivar identification.     

A potential application for large diameter pellicular supports in the reversed phase study of ultra high molecular mass polystyrenes

Summary Ultra high molecular mass (> 7 million dalton) polystyrenes are prone to degradation in size exclusion chromatography. In gradient elution reversed phase HPLC they do not elute visibly on small particle size porous supports. However, large diameter C18 pellicular particles were successfully employed for reversed phase study of ultra high molecular mass (15 million dalton) polystyrenes without polymer degradation during elution. Although retention for the lower molecular mass polystyrenes was lower than on small diameter porous particles, the medium high molecular mass polystyrenes (0.5–1 million dalton) showed similar retention. The addition of small diameter porous particles in small quantities, to the large diameter pellicular particles, increased the amount of retention of the low molecular mass polystyrenes without affecting the higher molecular mass polystyrenes.     

Void volume determination and experimental probes in reversed phase chromatography

Summary In reversed-phase liquid chromatography with n-alkyl bonded silica, the dead volume (V₀) of the column is theoretically indeterminate owing to adsorption of organic modifier on n-alkyl chains and of water on silanol groups. With binary mobile phases, retention volumes of the mobile phase components and of their deuterated species are relaeed to the adsorption isotherms and V₀ by equations which can be solved with some assumptions on the adsorbed layer composition. Methanol-water and acetonitrile-water systems are studied. As the experimental excess isotherm shows a linear part in the concentration range 50–80% in organic modifier, the hypothesis of an adsorbed layer of constant composition in this range is possible. When increasing the water content of the mobile phase, adsorption of water occurs up to saturation of silanol groups. Then the assumption of a constant water content for a mobile phase having more than 50% of water is applied. With the hypothesis of a constant adsorbed content of organic modifier when the eluent has more than 80% of organic modifier, V₀ and the absolute isotherms are calculated over the entire range of mobile phase composition. Experimental retention behavior of the mobile phase components are totally explained by these V₀ determinations. The retention times of commonly used V₀ markers are compared with V₀ values. It is shown that, when buffering the eluent, no visible effect on the distribution equilibrium is observed, so that injection of concentrated potassium nitrate is a convenient method to measure V₀. With a few solutes with are UV detectable it is possible to measure V₀ whatever the mobile phase composition in methanol-water and acetonitrile-water systems.     

Adsorption isotherms of organic modifiers and the determination of the dead volume in RPLC

Summary In RPLC the dead volume can be defined as the difference between the maximum column hold-up volume and the volume of the adsorbed phase. The composition of the adsorbed phase depends on the composition of the mobile phase and therefore, the dead volume also varies with it. In this work, the alkyl bonded phase acetonitrile (ACN)-water mobile phase system is investigated. In the system, deuterated water (D₂O) and deuterated acetonitrile (D-ACN) are retained due to the isotopic dilution effect. By means of D₂O and D-ACN, the absolute adsorption isotherm of the organic modifier ACN is measured. Based on the isotherm, the chromatographic behaviour of ACN, D-ACN and D₂O, the variation of the dead volume with the composition of the mobile phase, and the approach to determine the maximum column hold-up volumn are explained. In addition, the various approaches to determine the dead volume are compared and the recommendations are given for the case of common unbuffered binary systems (MeOH/H₂O, THF/H₂O and ACN/H₂O).     

Capillary liquid chromatography as a tool for separation of hydrophobic basic drugs. Relation between tests for column characterization and real analysis

Two capillary columns for reversed phase (RP) capillary liquid chromatography (CLC), viz. Nucleosil 100‐5 C18 and LiChrosorb RP‐select B, were characterized by the Walters test, i.e. the chromatographic test proposed for RP stationary phases. Hydrophobicity indices were determined not only in acetonitrile/water mobile phase, as proposed in the test, but they were also measured in buffered systems. This approach was used to quantify the influence of mobile phase composition on the modification of the surface of the stationary phases. In the next step, small basic compounds differing in their hydrophobicity and basicity were selected and their retention on the stationary phases in mobile phases of the same composition as used for column testing was examined. Furthermore, the retention of newly synthesized drugs, chemotherapeutics derived from thioacridine and pyridoquinoline, differing in their structures, basicity, and hydrophobicity, was also studied. The composition of the mobile phases had to be shifted to higher contents of organic modifiers – acetonitrile or methanol – in order to elute these hydrophobic compounds from the columns. The question we wanted to answer was: How is the method for testing of reversed phases related to retention, separation efficiency, and peak symmetry of various analytes?     

Correlation between retention and the C18 silica surface coverage in reversed-phase liquid chromatography

Summary Members of three homologous series and five non-homologous solutes with various functional groups were chromatographed on a series of well-characterized C₁₈ reversed stationary phases with a range of methanol-water mobile phases. Measured capacity factors of solutes were correlated with the concentration of C₁₈ ligands on the stationary phase. A linear relationship with the slope depending on a solute molecular structure and the volume fraction of methanol in the mobile phase was obtained. A method for the evaluation of phase ratio is also proposed. Presented at the 17th International Symposium on Chromatography, September 25–30, 1988, Vienna, Austria.     

含酯基包覆聚合物液相色谱柱填料

以丙烯酸甲酯或辛酯和二乙烯苯为原料，在溶液中用游离基聚合法制备了一系列含酯基包覆聚合物反相液相色谱柱填料．用傅立叶红外光谱、电子显微镜和元素分析鉴定了聚合物层，并评价了诸如硅羟基、柱压降、柱效和峰对称性等特性．该类填料适合于含氨基和羟基化合物的分离，作为应用实例，对洛伐他汀（Lovastatin）的分析展示其优良的色谱性能。     

Use of molecular connectivity indices to predict LC retention of dansylamides in six different eluent systems

Summary Molecular connectivity indices (x) through the sixth order were calculated and compared with measured reversed-phase liquid chromatographic retention data of sixteen dansyl derivatives of amines (dansylamides). Retention measurements were performed in the capacity factor, k′, range 1–30 with organic solvent-water eluents using acetone, acetonitrile, ethanol, methanol, 1-propanol and 2-propanol. Mainly valence level indices were selected to describe retention. Indices of different order were selected in the different eluents, suggesting that there are differences in the retention mechanism in these eluents. Retention can be predicted with great accuracy by the molecular connectivity indices, as the high correlations between the calculated and observed retention indicate. Presented at the 15th International Symposium on Chromatography, Nürnberg, October 1984