Aminopropyl Silica Gel Research Articles

Separation of enantiomers on a chiral stationary phase based on ovoglycoprotein: II. Comparison of chiral recognition properties with crude ovomucoid

Chiral stationary phases based on ovoglycoprotein from chicken egg whites (OGCHI) and crude ovomucoid from chicken egg whites (OMCHI) were compared with regard to the bound amounts of OGCHI and chiral recognition abilities. Crude OMCHI included 11% OGCHI, by weight. Since pure OMCHI had no appreciable chiral recognition ability, the chiral recognition ability of crude OMCHI originated from OGCHI, which was present in crude OMCHI preparations as an impurity. However, a chiral stationary phase based on crude OMCHI showed good chiral recognition ability, despite the 11% OGCHI content in crude OMCHI preparations. When crude OMCHI was reacted with N,N′-disuccinimidylcarbonate (DSC)-activated aminopropyl-silica gels, the ratio of bound OGCHI to that of totally bound protein was 0.23. These results reveal that the good chiral recognition ability of a stationary phase based on crude OMCHI is due to OGCHI being preferentially bound to DSC-activated aminopropyl-silica gels rather than the OMCHI. In addition, OMCHI did not contribute to the enantioselectivity of the solute at all and made little contribution to the retention characteristics.

Retention of benzene derivatives on the aminopropyl stationary phase in high-performance liquid chromatography

The retention of benzene derivatives in the normal-phase mode of high-performance liquid chromatography (HPLC) using a binary mobile phase (n-hexane-THF) on aminopropyl silica gel was considered. The possibility of describing the retention of the sorbates by known models was evaluated. It was established that molecular interactions between a sorbate and components of the mobile phase in solution can significantly affect the retention of the sorbates and the character of the dependence of the retention on the concentration of a modifier in the mobile phase. A quantitative description of this dependence was proposed.

Enantioselectivity of bovine serum albumin-bonded columns produced with isolated protein fragments: II. Characterization of protein fragments and chiral binding sites

Enantioselectivity of bovine serum albumin (BSA)-bonded columns produced with isolated protein fragments has been investigated. The BSA fragment, BSA-FG75, was isolated by size exclusion chromatography following peptic digest of BSA. The isolated BSA-FG75 was further fractionated to two fractions, BSA-F1 and BSA-F2, by anion-exchange chromatography. BSA-F1 and BSA-F2 had molecular mass of about 35 000 daltons, estimated by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry. BSA-F1 has amino acid residues 1–307 estimated by electrospray ionization (ESI) mass spectrometry, while BSA-F2 is an N-terminal half BSA fragment. The BSA, BSA-FG75, BSA-F1 and BSA-F2 proteins were bound to aminopropyl-silica gels activated by N, N′-disuccinimidyl carbonate. The bound amounts of the BSA fragments were 2.2–2.7 times more than that of the intact BSA. Chiral recognition of 2-arylpropionic acid derivatives, benzodiazepines, warfarin and benzoin was obtained with the BSA fragment-bonded columns. The non-enantioselective interactions of benzoin and benzodiazepines except for clorazepate with BSA fragments were increased with protein surface coverages, while those of 2-arylpropionic acid, clorazepate and warfarin were decreased. The BSA fragment columns gave higher enantioselectivity for lorazepam and benzoin, and lower enantioselectivity for other compounds tested, compared with the BSA column. These results might be due to changes in the globular structure of the BSA fragment and/or changes in the local environment around the binding sites.

Analysis of sterols in vegetable oils using off-line SFC/capillary GC-MS

The analysis of sterols in vegetable oils by off-line SFC followed by capillary GC-MS is described. The fractionation of the sterols from the complex oil matrix is achieved by SFC on aminopropyl silicagel in less than 8 minutes. Injection and collection of the sterol fraction is fully automated and time controlled. The sterols are analysed without derivatisation by capillary GC-MS. Identification is performed by full scan electron ionisation and quantitation is carried out by extracted ion chromatography at m/e 107, with cholesterol as internal standard. The analyses of the sterols from the sunflower oil and two olive oils illustrate the possibilities of the method.

Computational chemical analysis of chiral recognition in liquid chromatography, selectivity of N-( R)-1-( α-naphthyl)ethylamino carbonyl-( R or S)-valine and N-( S)-1-( α-naphthyl)ethylamino carbonyl-( R or S)-valine bonded aminopropyl silica gels

The chiral recognition of N-( R)-1-( α-naphthyl)ethylamino carbonyl-( R or S)-valine and N-( S)-1-( α-naphthyl)ethylamino carbonyl-( R or S)-valine bonded aminopropyl silica gels in liquid chromatography was examined using computational chemical analysis. The chiral recognition centers of the model chiral molecules were analyzed using the Extended Hückel CAChe TM program, and then the structures of hydrogen bond complexes with analytes were optimized by molecular mechanics calculations. The differences in final energy values indicated the elution order and enantiomer separation. The optimized density of chiral phases indicated that larger sized analytes may not slip into chiral recognition brush and form a one-to-one complex with the chiral recognition molecule used for analysis.

Photosensing by a fluorescing probe covalently attached to the silica

Starting from aminopropyl silicagel, an anthracene fluorophore has been covalently grafted on the surface of silica, the secondary amino group being left in the linking arm. When one fifth of the initial amino functions are grafted, the silica behaves as a pH probe, the fluorescence of which switches on when the amino groups are protonated. On the contrary, if the silica loading is almost complete, the pH probing is no longer possible. In this case, an excimeric versus monomeric emission can be used in sensing the solvent water content.

Analytical chiral separation of a new quinolone compound in biological fluids by high-performance liquid chromatography

Two methods for the separation of a new racemic quinolone compound, temafloxacin (TMFX), in biological fluids by high-performance liquid chromatography (HPLC) were studied. The first method was coupling of TMFX to S-(−)-N-1-(2-naphthyl sulfonyl)-2-pyrrolidine carbonylchloride (L-NSPC). The diastereomeric derivatives were separated on a silica gel column. The second method was separation on a chiral stationary phase with an ovomucoid conjugated to aminopropyl silica gel. Two enantiospecific methods gave a satisfactory result concerning both accuracy and precision, and the second method was superior to the first one for chromatographic separation. Furthermore, the pharmacokinetics of the enantiomers after oral administration of racemic TMFX to healthy volunteers was investigated by the second method.

Retentive and enantioselective properties of ovomucoid-bonded silica columns. Influence of protein purity and isolation method

The influence of protein purity and isolation method on the chiral resolution of racemates on ovomucoid-bonded materials has been investigated. Chicken ovomucoid (OMCHI) and turkey ovomucoid (OMTKY) were obtained from commercial sources, and were isolated by precipitation with acetone from their corresponding egg whites. Further, crude OMCHI protein was fractionated by cation-exchange chromatography into two fractions, pure OMCHI and an unknown protein fraction. The OMCHI, fractionated OMCHI and OMTKY proteins were characterized by N-terminal sequencing and trypsin-inhibitory activity. Their purity was checked by reversed-phase chromatography. The results reveal that the commercial OMCHI, and the isolated OMCHI and OMTKY are crude preparations and that an unknown protein fraction is present in each crude ovomucoid protein. The OMCHI, fractionated OMCHI and OMTKY proteins were bound to aminopropyl-silica gels activated by N, N′-disuccinimidyl carbonate to test the retentive and enantioselective properties of those materials. The result suggests that the unknown protein fraction could be responsible for chiral recognition ability of the OMCHI and OMTKY columns.

Detection of sugar addition to citrus juices by capillary GC

AbstractA method is described for the detection of citrus juices adulterated by addition of industrial sugar products. The oligosaccharides maltotriose (DP3) and maltotetraose (DP4), both of which are absent from the natural sugar profile of citrus fruits, serve as markers. The sample preparation method includes clean‐up on a solid phase ODS cartridge, fractionation of DP3 and DP4 carbohydrates by LC on aminopropyl silica gel, and derivatization of the collected sugars to form the oxime‐trimethylsilyl derivatives. Capillary GC analysis is performed on a 30 m × 0.32 mm i.d. polydimethylsiloxane fused silica column applying cool on‐column injection and FID detection. The detection limit for maltotriose and maltotetraose is about 1 mg l−1. Several orange and grapefruit juices were evaluated on their authenticity. Adulteration with crystalline sugar products cannot be detected with the described procedure.

Enantioselectivity of bovine serum albumin-bonded columns produced with isolated protein fragments

The enantioselectivity of bovine serum albumin (BSA)-bonded columns produced with isolated protein fragments was investigated. The BSA fragment BSA-FG75 was isolated by size-exclusion chromatography followed by peptic digestion of BSA. The isolated BSA-FG75 was a mixture of three peptides, and was mainly an N-terminal half peptide(s) with an average molecular mass of about 35 000. The BSA and BSA-FG75 proteins were bound to aminopropylsilica gels activated by N,N′-disuccinimidyl carbonate. The amounts of the proteins bound were about 2 and 5.5 μmol/g for the BSA and BSA-FG75, respectively. Chiral recognition of 2-arylpropionic acid derivatives, benzodiazepines, warfarin and benzoin was obtained with the BSA-FG75-bonded columns, but no chiral recognition of tryptophan or kynurenine was obtained. The intact BSA column gave a higher enantioselectivity than the BSA-FG75 column for most of the compounds tested, whereas the BSA-FG75 column gave a higher enantioselectivity than the intact BSA column for lorazepam and benzoin, and had a higher capacity for benzoin. These results are due to a higher density of chiral recognition site(s) on the BSA-FG75 column. Also, the BSA-FG75 column was as stable as the intact BSA column for a continuous flow of eluent.

Automatic determination of hydroperoxides of phosphatidylcholine and phosphatidylethanolamine in human plasma.

An automatic method for the determination of hydroperoxides of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) is reported. Sample plasma was deproteinized with a fourfold volume of methanol. After centrifugation, the supernatant was injected directly into an HPLC system without further treatment. The hydroperoxides of PC and PE were concentrated and washed on an ODS column followed by introduction into two analytical columns, a silica gel and an aminopropylsilica gel column, which were connected in series, by column switching. After the separation, they were detected by postcolumn detection with diphenyl-1-pyrenylphosphine. The compounds were determined at picomole levels within 30 min with good reproducibilities. By using only a silica gel column as an analytical column, PC hydroperoxides were determined within 20 min, and samples could be injected into it at 15-min intervals. Those methods made it possible to inject a sample of up to 2 ml at one time and up to 8 ml by repeated injections and to determine phospholipid hydroperoxides in human plasma at picomole levels.

Synthesis, properties and applications of silica-immobilized 8-quinolinol: Part 1. Characterization of silica-immobilized 8-quinolinol synthesized via a Mannich reaction

A one-step Mannich reaction has been developed for the immobilization of 8-quinolinol on aminopropylsilica gel that results in a product exhibiting a capacity of 466 ± 20 μmol g −1 for copper(II) ions. Capacity measurements were made by copper(II) uptake and elemental analysis. Characterization of the silica bound ligands in this synthetic context including IR spectrometry, 13C NMR spectrometry, acid-base properties and stabilities were presented. The effect of pH on the extraction of copper(II), nickel(II) and cadmium(II) for silica-immobilized 8-quinolinol was also evaluated.

Direct injection assay of drug enantiomers in serum on ovomucoid-bonded silica materials by liquid chromatography

A high-performance liquid chromatographic (HPLC) method for the determination of drug enantiomers in serum was developed. The method involves direct injection of serum samples on to an ovomucoid-bonded column, which is prepared by bonding of ovomucoid proteins to an aminopropyl-silica gel by the N,N′-disuccinimidyl carbonate activation method and separation of drug enantiomers on the column using a mixture of phosphate buffer and an organic solvent. High recoveries of serum proteins were obtained using eluent pH values of 3, 4, 6 and 7 at phosphate buffer concentrations above 50 m M, whereas the recovery was ca. 70% at an eluent pH of 5. The recovery of each enantiomer of basic and acidic drugs from serum was almost 100%.

Preparation and characterization of covalently bound tetraphenylporphyrin-silica gel stationary phases for reversed-phase and anion-exchange chromatography

A tetraphenylporphyrin-based stationary phase for high-performance liquid chromatography is prepared by attaching a p-carboxyphenyl derivative of the porphyrin to aminopropyl silica gel through an amide bond. This stationary-phase support is used for the reversed-phase separation of several polycyclic aromatic hydrocarbons and is shown to exhibit novel shape selectivity toward planar aromatic solutes. Subsequent metalation of the immobilized tetraphenylporphyrin with either tin(IV) or indium(III) results in a stationary phase with anion-exchange characteristics. Both the tin(IV) and indium(III) metalloporphyrin-based stationary phases exhibit useful selectivity for the separation of aromatic sulfonates and aromatic carboxylates.

High-performance liquid chromatographic separation of enantiomeric amino acids on bis[carbamoyl(alkyl)methylamino]-6- chloro- s-triazine-derived chiral stationary phases

The synthesis and chiral recognition ability of a series of four chiral stationary phases (CSPs) containing 2,4-bis [carbamoyl(alkyl)methylamino]-6-chloro- s-triazine (designated phase A) are described. The synthesis of these CSPs is achieved through amide formation by bonding 2,4-bis[carboxy(alkyl)methylamino]-6-chloro- s-triazine onto 3- aminopropyl silica gel. Such phases are quite effective for high-performance liquid chromatographic separation of a racemic mixture of five typical amino acids. The chromatographic behaviour of these CSPs was studied. Comparison of these CSPs with the s-triazine-derived CSP (designated phase B), which bears a tripeptide chiral moiety, is also discussed. The present study clearly indicates that an s-triazine-terminated CSP derived from certain l- amino acids (phase A), instead of tripeptide, is effective enough for the separation of enantiomeric amino acids.

Improved gram-quantity isolation of malto-oligosaccharides by preparative HPLC

Malto-oligosaccharides up to a degree of polymerization of 8 were isolated in gram quantities by preparative HPLC from mild acid hydrolysates of cyclomaltoheptaose and cyclomatooctaose [β-cyclodextrin (βCD) and γ-cyclodextrin (γCD), respectively]. Dynamax-60A NH 2 preparative aminopropyl silica gel columns (21.4 and 41.4 mm i.d.) and acetonitrile-water mobile phases were used. Isolation rates using the 41.4 mm i.d. preparative column approached 1 g of purified oligosaccharide per hour. The purity of isolated malto-oligosaccharides was at least 98% for dp 7 and dp 8 oligosaccharides derived from βCD and γCD, respectively.

Liquid Chromatography with Aminopropylsilica Gel Modified by Heptakis(2,3,6-tri-O-methyl)-β-cyclodextrin Derivative

Abstract New stationary phases modified by heptakis(2,3,6-O-trimethyl)-β-cyclodextrin were prepared and tested as HPLC columns. The results show that these stationary phases exhibit significant chiral separation abilities for various types of aromatic compounds under reversed-phase conditions. Acetylation and propionylation of the residual amino groups on silica gel improve the retention capacity of the stationary phase and results in better chromatographic chiral separation. One of the most possible reasons for this superior chiral separation observed for the acylated stationary phases is considered to be the increased local concentration of the solutes on the hydrophobic surface of the stationary phase, which increases the possibility for the contact of solutes with the outside of cyclodextrin fixed on the silica gel.

Chromatographic effects of residual amino groups on liquid chromatographic stationary phases derived from aminopropyl-silica gel

Low-capacity phenylazo-8-quinolinol-silica gel (QSG) stationary phases were made by two different procedures. The first began with an aminopropyl-silica gel (APSG), which was converted to nitrobenzamide-silica gel (NBSG) by amidization with p-nitrobenzoyl chloride. The second began with the direct preparation of the NBSG using nitrobenzamidesilane. Retention data for several organic analytes were obtained on APSG and on the NBSG and QSG phases made by both procedures. Anomalously high capacity factors were obtained for p-nitrobenzoic acid on the NBSG material obtained from APSG, indicating the presence of unreacted aminopropyl anion-exchange sites. Only after reamidization of the NBSG using more vigorous conditions did the retention data approach that on the NBSG made by the second method. Even then, the opposite order of elution of p-nitrobenzoic and benzoic acids on the two NBSG and QSG phases indicated that vestiges of amino groups remained on the materials made from the APSG. Several chromatographic and chemical characterization methods confirmed the presence of residual aminopropyl groups.

Gas-phase acylation of aminopropyl-silica gel in the synthesis of some chemically bonded silica materials for analytical applications

Gas-phase acylation of aminopropyl-silica gel with aliphatic dicarboxylic (succinic, adipic and sebacic)_and 4-aminobenzoic acids is proposed as a rapid and efficient one-step method for the synthesis of carboxyalkyl- and 4-aminophenylmidopropyl-silica gels, usually used as zwitterion exchangers for liquid chromatography and matrices for multi-step syntheses of silica-bound aromatic azo reagents for the sorption and chromatographic separation of metal ions. Acylation degrees of 59-90% are achieved after 0.5 h. IR spectra of the acylation products and near-UV-visible spectra for bonded aromatic azo compounds, based on 4-aminobenzamidopropyl-silica gel, are presented. Some positive and negative aspects of the gas-phase acylation are discussed.

Pressure effect on the fluorescence decay of erythrosin bonded to an aminopropyl silica gel particle — measurements under a microscope

An experimental technique to examine the effect of pressure on the fluorescence decay of a microscopic sample has been developed and applied to erythrosin (ER) chemically bonded to an aminopropyl silica gel particle (APS). A dye-bonded particle (ER  APS) of 3 – 10 μm in diameter exhibits a double-exponential fluorescence decay in all solvents studied; this reflects the presence of at least two environmentally different states of the ER moiety. A significant effect of pressure observed in ethylene glycol is ascribed to the pressure-assisted penetration of the solvent molecules into small pores in ER  APS. However, in benzene, ethanol and acetonitrile, no effect of pressure is observed on the fluorescence decay of ER  APS up to 50 MPa despite marked differences in the fluorescence decay in these solvents. This indicates that the two states responsible for the dual exponentiality and their interconversion are not influenced by increasing pressure.