Octadecylsilyl Silica Gel Research Articles

Trehangelins A, B and C, novel photo-oxidative hemolysis inhibitors produced by an endophytic actinomycete, Polymorphospora rubra K07-0510

Three new natural products, designated trehangelins A, B and C, were isolated by solvent extraction, silica gel and octadecylsilyl silica gel column chromatographies and subsequent preparative HPLC from the cultured broth of an endophytic actinomycete strain, Polymorphospora rubra K07-0510. The trehangelins consisted of a trehalose moiety and two angelic acid moieties. Trehangelins A (IC50 value, 0.1 mg ml(-1)) and C (IC50 value, 0.4 mg ml(-1)), with symmetric structures, showed potent inhibitory activity against hemolysis of red blood cells induced by light-activated pheophorbide a. However, trehangelin B, with an asymmetric structure, displayed only a slight inhibition (IC50 value, 1.0 mg ml(-1)).

Simultaneous Determination of Artesunate and Amodiaquine in Fixed-Dose Combination by a RP-HPLC Method with Double UV Detection: Implementation in Interlaboratory Study Involving Seven African National Quality Control Laboratories

The fixed-dose combination artesunate (AS)–amodiaquine (AQ) is one of the most widely used treatments for uncomplicated falciparum malaria. It is currently proposed to the inclusion in the model list of essential medicines of World Health Organization and has been recently prequalified. Until now, no satisfactory method for the simultaneous determination of the two active ingredients had been available. Thus, a reversed phase high performance liquid chromatography for the quantitative determination of AQ and AS was developed and validated. Chromatography was performed using an end-capped octadecylsilyl silica gel column (100 × 4.6 mm, 3 μm) with a binary gradient using aqueous phase containing potassium dihydrogen phosphate (10 mM) and acetonitrile. Taking into consideration the physico-chemical characteristics of the two compounds related to their ionization, the use of a counter ion was necessary to ensure the retention of AQ in a reversed phase system simultaneously to AS. Thus, aqueous mobile phase was adjusted to pH 3.0 and the chosen counter ion was sodium 1-octanesulfonate (100 mM). In these conditions, the retention times were about 4 min for AQ and 10 min for AS with UV detection at 300 and 210 nm, respectively. Method was then validated according to ICH guideline (specificity/linearity/accuracy/precision) and potential interferences with excipients and degradation products were checked. It has also been used for an interlaboratory study involving seven African National Quality Control Laboratories and Afssaps (Agence francaise de securite sanitaire des produits de sante) laboratory. The results demonstrate that this rapid and simple method can be easily used by official laboratories for routine control, market survey and for the detection of potential substandard medicines which are very frequent in African countries.

Panowamycins A and B, new antitrypanosomal isochromans produced by Streptomyces sp. K07-0010

Two new isochromans, panowamycins A and B, were purified by solvent extraction, silica gel and octadecylsilyl silica gel (ODS) column chromatography followed by preparative HPLC, from a culture broth of Streptomyces sp. K07-0010, together with the known compounds NFAT-133, conglobatin, piericidin C series and dinactin. Structures of panowamycins were elucidated as new analogs of NFAT-133 by spectroscopic studies including various NMR experiments. Panowamycins A and B showed moderate antitrypanosomal activity, with IC(50) values of 0.40 and 3.30 μg ml(-1), respectively.

Mass Transfer of Water-insoluble Organic Compound from Octadecylsilyl-silica Gel into Water in the Presence of a Nonionic Surfactant

The release of perylene from octadecylsilyl (ODS)-silica gel into water using a nonionic surfactant was kinetically studied by single microparticle injection and absorption microspectroscopy techniques. The release of perylene from the porous microparticles significantly depended on the surfactant concentration. The release rate constant was inversely proportional to the microparticle radius; the rate-determining step was the process at the spherical microparticle surface. The mechanism is discussed in terms of the solubilization of perylene at the microparticle surface by the micelle.

An accurate quantitative LC/ESI–MS/MS method for sirolimus in human whole blood

Sirolimus is a widely used immunosuppressant that requires therapeutic drug monitoring (TDM). We optimized a preanalytical procedure that allows for the accurate quantiation of sirolimus in whole blood by LC/ESI–MS/MS with minimal matrix effects. Sirolimus is highly lipophilic, and solvents containing greater than 50% methanol were required to maintain sirolimus recovery. The final pretreatment procedure developed consists of a zinc sulfate protein precipitation, an extraction using octadecyl silyl-silica gel for eliminating water-soluble and hydrophilic compounds, and HybridSPE cartridge treatment to eliminate phospholipids. Using this procedure prior to LC/ESI–MS/MS led to the accurate and reproducible quantitation of sirolimus in human whole blood. The linear range of detection was 0.5–50 ng/mL, a range appropriate for TDM, and the method demonstrated good repeatability and intermediate precision within this quantitative range. In order to investigate the quantitative performance of this method, we compared it to two commercially available sirolimus immunoassays and our previously reported LC/ESI–MS/MS method. The immunoassays gave consistently greater values for the sirolimus concentration, and this may be related to antibody cross-reactivity with sirolimus metabolites and/or other matrix effects. Although our procedure is too long to support real-time TDM for outpatients, it can serve as reference method to assess the performance of other analytical methods that are currently available or may be developed in the future.

Stability Indicating Analytical Method Development and Validation of Efavirenz Quantification by High Performance Liquid Chromatographic Technique

A stability‐indicating high performance liquid chromatography (HPLC) method is developed for the quantification of efavirenz drug substance. Selected mobile phase is a combination of 50 volume of acetonitrile and 50 volume of 0.86% w/v solution of ammonium dihydrogen phosphate, pH adjusted to 3.0 with orthophosphoric acid. Optimized column is a stainless steel column packed with base deactivated octa decylsilyl silica gel and at 252 nm wavelength. Linearity of the method is found 40 μg/mL to 160 μg/mL with regression coefficient of 0.9995. The method is validated according to ICH guidelines.

Selective isolation of bacteria from soil with hydrophobic materials

Bacterial strains having a hydrophobic cell surface have often been considered as degraders of hydrophobic organic pollutants in soil. In this study, bacterial strains were isolated using hydrophobic materials from 12 soil samples, and their cell surface hydrophobicity was determined by evaluating their adherence to n-hexane. Bacterial strains isolated using polytetrafluoroethylene (PTFE) membrane were more hydrophobic on an average than those isolated with styrene–divinylbenzene (DVB) particles or octadecylsilyl silica gel (ODS) particles. Strains closely related to Burkholderia cepacia could be selectively isolated using the PTFE membrane; those closely related to Ralstonia pickettii, using ODS and DVB particles; and those closely related to B. fungorum, using DVB. These results indicate that bacterial strains having a hydrophobic cell surface or within certain phyla can be selectively isolated from soils using hydrophobic materials, and that this isolation method would be useful for collecting candidates for bioremediation of hydrophobic pollutants.

Identification of Eukaryotic and Prokaryotic Methylthiotransferase for Biosynthesis of 2-Methylthio-N6-threonylcarbamoyladenosine in tRNA

Bacterial and eukaryotic transfer RNAs have been shown to contain hypermodified adenosine, 2-methylthio-N(6)-threonylcarbamoyladenosine, at position 37 (A(37)) adjacent to the 3'-end of the anticodon, which is essential for efficient and highly accurate protein translation by the ribosome. Using a combination of bioinformatic sequence analysis and in vivo assay coupled to HPLC/MS technique, we have identified, from distinct sequence signatures, two methylthiotransferase (MTTase) subfamilies, designated as MtaB in bacterial cells and e-MtaB in eukaryotic and archaeal cells. Both subfamilies are responsible for the transformation of N(6)-threonylcarbamoyladenosine into 2-methylthio-N(6)-threonylcarbamoyladenosine. Recently, a variant within the human CDKAL1 gene belonging to the e-MtaB subfamily was shown to predispose for type 2 diabetes. CDKAL1 is thus the first eukaryotic MTTase identified so far. Using purified preparations of Bacillus subtilis MtaB (YqeV), a CDKAL1 bacterial homolog, we demonstrate that YqeV/CDKAL1 enzymes, as the previously studied MTTases MiaB and RimO, contain two [4Fe-4S] clusters. This work lays the foundation for elucidating the function of CDKAL1.

Mass transfer of weak acid in nanometer-sized pores of octadecylsilyl-silica gel

The sorption of bromophenol blue or anthracene-9-carboxylic acid as a weak acid into single octadecylsilyl (ODS)-silica gel microparticles in a solution was analyzed by microcapillary injection-manipulation and absorption microspectroscopy. The distribution ratio and the sorption rate were highly dependent on the pH of the solution. These results are discussed in terms of the acid dissociation/association, distribution between the ODS and solution phases, and intraparticle diffusion of the weak acid in the nanometer-sized pores of the ODS-silica gel.

Liquid Chromatographic Determination of Plasma Ropivacaine for Assessing Pharmacokinetics of the Viscous Preparation

We developed assay method for determination of plasma ropivacaine by using reversed-phase high performance liquid chromatography (HPLC) equipped with ordinary octadecylsilyl silica-gel (ODS) column. Plasma samples spiked with internal standard (bupivacaine) were treated by ethylacetate to extract ropivacaine and internal standard. The ropivacaine and internal standard separated on ODS column were detected by an ultra violet (UV) detector set at 215 nm. The mobile phase solvent consisted of acetonitrile, methanol and 0.05 M phosphate buffer adjusted to pH 4.0 (10 : 30 : 60, v/v) was pumped at a flow rate of 0.8 ml/min. The calibration curve of ropivacaine was linear at the concentration of 25-1,000 ng/ml (r=0.9998). The recoveries of ropivacaine from plasma were greater than 87.9% with the coefficient of variations (CVs) less than 6.1%. The CVs for intra- and inter-day assay of ropivacaine were 2.0-12.0% and 1.7-14.8%, respectively. This HPLC method was applied to determining plasma ropivacaine in two healthy subjects after receiving 0.5% ropivacaine viscous preparation, which was prepared in our hospital. Our preliminary pharmacokinetic data showed that ropivacaine viscous could be used safely based on the plasma ropivacaine concentrations (C(max): 89-125 ng/ml) for pain relief in oral mucosa.

Monolithic octadecylsilyl-silica gel column for the high-speed ion chromatographic determination of acidity

A monolithic ODS-silica gel column modified by saturating it with lithium dodecylsulfate (Li-DS) was used to demonstrate the high-speed separation of H + from other mono- and divalent cations, such as Na +, NH 4 +, K +, Mg 2+ and Ca 2+ using ion chromatography (IC). Using a 5 m M EDTA-2K solution containing 0.10 m M Li-DS (pH 4.80) as eluent, H + was eluted with a sharp and symmetrical peak within 1.0 min before other cations at a flow-rate of 1.5 ml min −1. The rapid elution of H + and its conductimetric detection could be attributed to the presence of EDTA (HY 2−), which can convert H + ions as anions, i.e. H ++H 2Y 2−→H 3Y −. The acidity of rainwater and deionized water samples was determined using this IC system with satisfactory results.

Parameter estimation of perillyl alcohol in RP-HPLC by moment analysis

Parameter estimations were made for the reversed-phase adsorption of perillyl alcohol (POH), a potent anti-cancer agent, on octadecylsilyl-silica gel (ODS). The average particle diame- ter of ODS was about 15 µm, and the particles were packed in the column (3.9 × 300 mm). The mobile phase used was a mixture of acetonitrile and water, in which the acetonitrile ranged be- tween 50 and 70 (v/v %). The first absolute moment and the second central moment were deter- mined from the chromatographic elution curves by moment analysis. Experiments were carried out using POH solutions within the linear adsorption range. The fluid-to-particle mass transfer coefficient was estimated using the Wilson-Geankoplis equation. The axial dispersion coefficient and the intraparticle diffusivity were determined from the slope and intercept of a plot of H vs 1/u0, respectively. The contributions of each mass-transfer step were axial dispersion, fluid-to- particle mass transfer, and intraparticle diffusion.

Improved separation of poly (butylene glycol) oligomers on a C18 column by gradient reversed-phase liquid chromatography and fine-tuning of the organic modifier system

Poly(butylene glycol) (PBG) samples of widely differing molecular weight (M), either in the native form or as the corresponding α, ω-bis(1-naphthyl)urethane derivatives, have been subjected to reversed-phase high-performance liquid chromatography (gRPHPLC) on an octadecylsilyl (C18) silica gel stationary phase with mobile phase gradients comprising binary eluent systems prepared from tetrahydrofuran (THF) and water and from acetone and water, and with ternary systems prepared from acetonitrile, THF and water and from methanol, THF and water.

Recognition of critical pairs of polycyclic aromatics on crystalline, liquid-crystalline and isotropic regions of silica-supported polymer in HPLC

The silica-supported comb-shaped polymer (Sil-ODA18) with a “crystalline-to-isotropic” phase transition showed the characteristic behaviors of both polymeric and monomeric ODS phases. This was examined using a SRM 869 test mixture. Furthermore, Sil-ODA18 exhibited a specific selectivity towards the critical pairs of polycyclic aromatic hydrocarbons (PAHs), such as acenaphthene and fluorene, or benz(a)anthracene and chrysene, or benzo(ghi)perylene, indeno(1,2,3-cd)pyrene and dibenz(a,h)anthracene. The separation of these solutes was controlled by selecting temperature and solvent systems. The selectivity on Sil-ODA18 was also compared with a conventional ODS (octadecylsilyl silica gel) column. The selectivity on a Sil-ODA18 matrix is attributable to an orientation change of the long-chain alkyl substituents on the surface of the silica base material on transition from crystalline to isotropic state.

Simple and rapid high‐performance liquid chromatography analysis of propentofylline and its main metabolites in serum using a direct injection technique

A rapid, simple and reliable high-performance liquid chromatography (HPLC) column-switching method with UV detection (270 nm) for the simultaneous determination of propentofylline and its metabolites in human and rat sera was developed. The method involves direct injection of serum onto an HPLC column, which contains a shielded hydrophobic stationary phase for the separation of analytes from proteins in serum, and then loading the analytes onto a short octadecylsilylated silica gel (ODS) column using a switching valve. Propentofylline and its three metabolites in serum were separated from the serum components within 30 min after the injection. The detection limits (S/N = 3) of analytes spiked in human and rat sera ranged from 0.08 to 0.57 nmol/mL, and the net volume of serum used was 20 µL. The relative standard deviations for within- and between-day variations using rat serum were less than 4.3 and 5.6%, respectively. The method was used to determine propentofylline and its main metabolites in rat serum after a single intravenous dose of propentofylline (5 mg/kg). Copyright © 1999 John Wiley & Sons, Ltd.

Simple and rapid high-performance liquid chromatography analysis of propentofylline and its main metabolites in serum using a direct injection technique.

A rapid, simple and reliable high-performance liquid chromatography (HPLC) column-switching method with UV detection (270 nm) for the simultaneous determination of propentofylline and its metabolites in human and rat sera was developed. The method involves direct injection of serum onto an HPLC column, which contains a shielded hydrophobic stationary phase for the separation of analytes from proteins in serum, and then loading the analytes onto a short octadecylsilylated silica gel (ODS) column using a switching valve. Propentofylline and its three metabolites in serum were separated from the serum components within 30 min after the injection. The detection limits (S/N = 3) of analytes spiked in human and rat sera ranged from 0.08 to 0.57 nmol/mL, and the net volume of serum used was 20 microL. The relative standard deviations for within- and between-day variations using rat serum were less than 4.3 and 5.6%, respectively. The method was used to determine propentofylline and its main metabolites in rat serum after a single intravenous dose of propentofylline (5 mg/kg).

Analysis of Surface Diffusion Phenomena in Reversed-Phase Liquid Chromatography

Surface diffusion data obtained for a reversed-phase liquid chromatographic system (octadecylsilyl silica gel and a 70/30 (v/v) methanol/water solution) were analyzed in relation to their molecular diffusivity (Dm). The adsorbate−adsorbent interactions between the studied compounds and the stationary phase were taken into account. The surface diffusion coefficient (Ds) depends on the mobile-phase composition, especially on the nature and concentration of the organic modifiers. Differences between the values of Ds measured under various conditions stem probably from differences in Dm. It also seems that Ds tends toward Dm with decreasing retention factor. The surface diffusion mechanism was assumed to be a restricted molecular diffusion in a potential field of adsorption. A restriction energy for this diffusion (Er) was introduced to correlate Ds with Dm. The ratio of Er to the isosteric heat of adsorption (−Qst) was found to be nearly constant, irrespective of the retention factor, with an average value of 0.32 for our phase system. An estimation procedure for Ds using the enthalpy−entropy compensation effect for the adsorption equilibrium is proposed. From the adsorption equilibrium constant at 298 K only, Ds could be estimated at different temperatures with an error less than ∼50%.

Estimation of the Surface-Diffusion Coefficient in Gas-Phase Adsorption Using Octadecylsilyl-Silica Gel

Abstract Surface-diffusion data in a gaseous adsorption system using an octadecylsilyl-silica gel as an adsorbent were analyzed according to a restricted Knudsen diffusion model. A linear correlation was confirmed between the ratio of the restriction energy for Knudsen diffusion to the isosteric heat of adsorption, Qst, and 1/(−Qst). It was proved that a linear correlation could be observed under the conditions that a linear free-energy relation and an enthalpy–entropy compensation in adsorption equilibrium were established. An estimation procedure of the surface-diffusion coefficient (Ds) was proposed based on the results. It was demonstrated that Ds of various adsorbates at given temperatures could be estimated with an error of less than about 50% from only one datum of the adsorption equilibrium constant measured at a different temperature. Similarly, Ds in gas-phase adsorption could be estimated from the evaporative energy of the adsorbates without experimental data.

Investigations on Radiolabeling Substance P Derivatives and Reversed-Phase Liquid Chromatographic Separation of the Products

In investigations of radiolabeling substance P1–11 (SP) with 125I isotope, its [Tyr8]-derivative was subjected to the lactoperoxidase (LPO) technique, whereas the native substance P undecapeptide was coupled with 125I-tyrosin-N-hydroxy-succinimidyl ester to the corresponding 125I-Bolton-Hunter SP derivative. For separation of the desired components from their by-products, labeled [Tyr8]-SP was pre-purified by ion-exchange chromatography (IEC) on carboxymethyl cellulose and subsequently investigated by gradient high performance reversed-phase liquid chromatography (gRP-HPLC) with 0.1 M triethyl-ammonium phosphate in acetonitrile (pH 2.5) on an octadecylsilyl silica gel (RP-18) stationary phase. When the total amount of labeled peptide affording a specific activity of approx. 2 × 106 Ci/M was checked by gRP-HPLC, the LPO technique yields about an equimolar mixture of 125I-[Tyr8]-(Met11)-SP and 125I-[Tyr8]-(Met11O)-SP. However, in contrast, substantial enrichment of 125I-[Tyr8]-(Met11)-SP was observed when more retained IEC fractions were analyzed. The 125I-Bolton-Hunter reagent was coupled in about 54 % yield to the SP undecapeptide at slightly alkaline conditions and separated from by-products only by gRP-HPLC yielding a specific activity of approx. 2 × 106 Ci/M. Both radiolabeled derivatives of SP are further characterized by their binding properties to a specific antibody raised in rabbits against synthetic SP1–11. When individual fractions of either radioiodinated [Tyr8]-SP or the corresponding 125I-Bolton-Hunter derivative were investigated with respect to their binding properties to the antibody, only those fractions coinciding with the maximum amount of radioactivity showed a satisfactory ratio of specific vs. non-specific antibody-binding, especially in the case of the 125I-Bolton-Hunter derivative. Therefore, the observed results are in good accordance with the existence of the desired target components.

Effect of the Type of Organic Modifiers on Mass Transfer in Reversed-Phase Liquid Chromatography

The mass-transfer characteristics were studied in a reversed-phase liquid chromatography consisting of an octadecylsilyl-silica gel and acetonitrile/water mixture (70/30, vol), and were compared with those obtained by using methanol/water (70/30, vol). In the two chromatographic systems, similar tendencies were observed for various adsorption characteristics. However, their absolute values were not entirely identical. Irrespective of the type of organic modifiers, surface diffusion was dominant for intraparticle diffusion. Similar correlations were also demonstrated with regard to the surface diffusion, i.e., the enthalpy-entropy compensation and the linear free-energy relation, suggesing a similarity in the surface migration mechanism in the two chromatographic systems. A few estimation procedures of the surface diffusion coefficient were proposed.