Chromatographic Properties Research Articles

Can the selectivity of phosphonium based ionic liquids be exploited as stationary phase for routine gas chromatography? A case study: The use of trihexyl(tetradecyl) phosphonium chloride in the flavor, fragrance and natural product fields

Room temperature ionic liquids (ILs) are well established stationary phases (SPs) for gas chromatography (GC) in several fields of applications because of their unique and tunable selectivity, low vapor pressure and volatility, high thermal stability (over 300 °C), and good chromatographic properties. This study is focused on an IL based on a phosphonium derivative (trihexyl(tetradecyl)phosphonium chloride, [P66614+] [Cl−]), previously shown to be suitable as a gas chromatographic SP because of its unique selectivity. In particular, it aims to establish the operative conditions to apply [P66614+][Cl−] to routine analysis of samples containing medium to high volatility analytes with different polarity, organic functional groups and chemical structure. In the first part, the study critically evaluates long term [P66614+][Cl−] column stability and maximum allowable operating temperatures (MAOT). The relatively low MAOT (210 °C) requires the adoption of a dedicated approach for analytes eluting above this temperature based on a suitable combination of efficiency and selectivity, and column characteristics (length, inner diameter and film thickness) and operative conditions. The performance of [P66614+][Cl−] as a GC SP have been validated through the Grob test, a model mixture of 41 compounds of different polarity, structure, and with different organic functional groups in the flavor and fragrance field, a standard mixture of 37 fatty acid methyl esters, some essential oils containing pairs or groups of compounds of different volatility critical to separate in particular peppermint, thyme, oregano, sandalwood and frankincense. The above approach has produced highly satisfactory separations with all of the samples investigated.

Facile preparation of polymer-brush reverse-phase/hydrophilic interaction/ion-exchange tri-mode chromatographic stationary phases by controlled polymerization of three functional monomers

Stationary phases with multiple-mode mechanism are beneficial to meet the needs of complex samples, but there are few multiple-mode stationary phases which can adjust the relative strength among multiple-forces imposed on the solutes. This work presents a facile preparation method of reverse-phase/hydrophilic interaction/ion-exchange tri-mode stationary phase, in which three functional monomers, lauryl methacrylate (LMA), hydroxyethyl methylacrylate (HEMA) and dimethylaminoethyl methacrylate (DMAEMA) as co-monomers underwent surface initiated-atom transfer radical polymerization (SI-ATRP) on the surface of silica. The structure of stationary phases was characterized and their chromatographic properties were investigated using various solutes. By comparison with classical single-mode columns, it is found that the newly designed columns can offer multiple interactions including hydrophobic, hydrophilic and electrostatic interactions and show good separation abilities to the tested solutes. Besides, changing the ratios of LMA, HEMA and DMAEMA in SI-ATRP system can easily adjust the relative strength of three interactions imposed on the solutes, inducing adjustable separation selectivity of the columns. The improved separation of multivitamin sample and the successful use of the columns in Per aqueous liquid chromatography indicate the potential of the tri-mode stationary phases.

Determination of mononitrophenols in water by gas-chromatography

Nitrophenols are highly toxic hydrophilic organic compounds. Nitrophenols prevalence in the biosphere is associated with both human industrial activity and with natural processes in the atmosphere and hydrosphere. Nitrophenols determination using gas chromatography requires chemical modification in order to reduce the hydrophilicity and polarity of the analytes. The derivatization must be carried out before the extraction concentration stage, but in most methods of the nitrophenols determination in water this approach is not implemented. The method of determination of mononitrophenols (2-, 3-, and 4-nitrophenols) in water was developed. It involved the nitrophenols bromination directly in the water, bromoderivatives liquid extraction with toluene, silylation by N-tert-Butyldimethylsilyl-N-methyltrifluoroacetamide (MTBSTFA) extract and gas chromatographic determination with the electron-capture detector ( GC-ECD ). The nitrophenols bromination conditions in aqueous solutions were optimized (pH value, bromination duration, bromide-ions concentration). In order to reduce the oxidizing activity of bromine, the nitrophenols bromination was recommended in the presence of bromide anions. The extraction (degree of extraction) and gas chromatographic (retention indices, ECD relative molar responses) properties and nitrophenols bromoderivatives properties were studied. It was shown that the degree of extraction of brominated nitrophenols in the toluene/water system exceeded 80% and the subsequent silylation of the bromoderivatives significantly improved their chromatographic properties. The analytical range of nitrophenols in water was 0.02-10 µg/dm 3 with an error of 10-35%, the detection limits were 0.05-0.07 μg/dm 3 , the sample volume for analysis was 50 cm 3 , the analysis duration was 90 min. Key words : mononitrophenols, chemical modification, gas-chromatography, electron-capture detector DOI: http://dx.doi.org/10.15826/analitika.2020.24.2.006 I.V. Gruzdev, B.M. Kondratenok, E.I. Lyu-Lyan-Min Institute of Biology of Komi Scientific Centre of the Ural Branch of the Russian Academy of Sciences (IB FRC Komi SC UB RAS), Kommunisticheskaya st., 28, Syktyvkar, 167982, Russian Federation

Chromatographic property classification of narrowly distributed ZnS quantum dots.

Although optimized synthesis methods for nanoparticles (NPs) on small scale can lead to narrow particle size distributions (PSDs) and thus defined optical properties, in particular during scale-up, an additional classification step must be applied to adjust the particle properties according to the needs of the later application. NP chromatography is a promising separation method, which can be potentially transferred to preparative and industrial scale. Herein, we demonstrate that remarkable classification of ZnS quantum dots (QDs) with respect to the fundamental band gap energy is achieved by chromatography although the PSD of the feed material is already very narrow (1.5-3.0 nm). We investigated the interactions of ZnS QDs with stationary and mobile phase materials in order to select a proper material couple so that irreversible NP adhesion, agglomeration, decomposition or dissolution of the ZnS QDs during the chromatographic experiments are avoided and highly reproducible chromatograms are obtained. Using a fraction collector, the already narrowly size distributed feed material was separated into coarse and fine fractions with distinct band gap energies. For characterization of the chromatographic fractionation, quantities known from particle technology, i.e. separation efficiency, cut size and yield, were adapted to the band gap energy distributions accessible from UV/Vis spectroscopy. The optimization of process conditions (flow rate, temperature, switching time of the fraction collector) allows fine-tuning of the property classification and therefore of the optical properties within the narrow distribution of the ZnS QDs. Our study shows the strength and high potential of chromatography for preparative and continuous separation of NPs even in case of narrow size-distributed sub-10 nm semiconductor QDs.

Preparation and Evaluation of a Cholesterol Derivatized β-Cyclodextrin-bonded Phase for Achiral and Chiral HPLC.

A cholesterol mono-derivatized β-cyclodextrin was synthesized and bonded onto silica gel (SBA-15) to obtain a cholesterol mono-derivatized β-cyclodextrin-bonded stationary phase (CHCDP). The chemical structures of mono-derivatized β-cyclodextrin and CHCDP were characterized by infrared spectroscopy, mass spectrometry, elemental analysis and thermogravimetric analysis, correspondingly. Furthermore, the separation ability of CHCDP in terms of achiral compounds was systematically evaluated by separating benzene homologs, polycyclic aromatic hydrocarbons (PAHs) and some positional isomers. As a result, CHCDP completely separated five benzene homologs and nine PAHs within 30 min under the reversed-phase. In addition, the chiral chromatographic property of CHCDP was also evaluated by separating some racemic compounds including flavanones, triazoles, β-blockers, etc. The results showed that the CHCDP exhibited high enantioselectivities towards most of selected analytes. The enantioresolutions were in the range from 1.43 to 2.51 on CHCDP. Especially the resolutions of 2'-hydroxyflavanone, hexaconazole, Dns-serine and atenolol were as high as 1.94, 1.91, 2.15 and 1.57, respectively. Obviously, the CHCDP was a versatile stationary phase with chiral and achiral separation capabilities in multi-mode chromatography, which was related to the introduction of cholesterol to the port of cyclodextrin, enhancing the hydrophobic interaction of cyclodextrin with achiral compounds, while maintaining the inclusion complexation of it with chiral compounds as well.

Enzymatic synthesis and characterization of chlorophyllide derivatives as possible internal standards for pigment chromatographic analysis

In this article the chlorophyllase activity of Dunaliella salina has been employed to generate different chlorophyllide a and b esters that could potentially be used as internal standards in pigment analysis. Chlorophyllide a (8’-hydroxyoctyl) ester was selected due its adequate chromatographic and spectral properties and was fully characterized by UV-Vis, ESI-MS and NMR. An easy room temperature procedure for its synthesis is described. Attachment

Fabrication and application of zwitterionic phosphorylcholine functionalized monoliths with different hydrophilic crosslinkers in hydrophilic interaction chromatography

In this study, a series of zwitterionic phosphorylcholine functionalized monolithic columns were fabricated via the thermally initiated co-polymerization of 2-methacryloyloxyethyl phosphorylcholine (MPC) and different hydrophilic crosslinkers, including 1,4-bis(acryloyl)piperazine (PDA), N,N′-methylenebisacrylamide (MBA) and 2-(methacryloyloxy)ethyl-[N-(2-methacryloyloxy)ethyl]phosphorylcholine (MMPC). The physicochemical and chromatographic properties of these MPC functionalized monoliths, including column morphology, pore size distribution, permeability, column efficiency, retention mechanism and ζ-potential analysis, were systematically compared. Furthermore, the influence of the crosslinker on the chromatographic performance of these MPC functionalized monoliths was evaluated. The chromatographic results indicate that the polarity of MPC functionalized monoliths may be related to the polarity of the crosslinker, which further affects the column selectivity and efficiency. A particularly high column efficiency (88,000 plates/m) was obtained on the novel poly(MPC-co-MMPC) monolith at optimum linear velocity using thiourea as test analyte. Compared to the poly(MPC-co-MBA) and poly(MPC-co-PDA) monoliths, the poly(MPC-co-MMPC) monolith exhibited higher separation selectivity for polar analytes, including nucleobases, nucleosides and benzoic acid derivatives. Moreover, 24 N-glycopeptides could be detected after enrichment with the poly(MPC-co-MMPC) versus 19 and 10 N-glycopeptides with the poly(MPC-co-MBA) and poly(MPC-co-PDA) monoliths, and no N-glycopeptide without enrichment. Therefore, MMPC has a great potential as a new and alternative hydrophilic crosslinker for the development of zwitterionic polymeric monoliths.

Base-deactivated and alkaline-resistant chromatographic stationary phase based on functionalized polymethylsilsesquioxane microspheres.

Vinyl, chloropropyl, and mercaptopropyl functionalized particles were prepared by a two-step acidic/alkaline catalyzed co-hydrolysis/condensation of methyltrimethoxysilane with a different silane precursor that carries chemically reactive functional group including vinyl, chloropropyl, and mercaptopropyl, respectively. The morphology, pore structure, and functional groups of the synthesized packings were studied by SEM, nitrogen adsorption-desorption measurements, and solid-state 13 C 29 Si NMR spectroscopy, respectively. The particles show ordered sphere, narrow particle size distribution, and mesoporous structure. The carbon contents of the microspheres are in the range of 17-19%, comparable to those of octadecyl-bonded silica packings. The three-kind of microspheres were directly used as packing materials for high-performance liquid chromatography without size classification. The chromatographic performance of the columns was evaluated and compared with a commercially available C18 phase. The results revealed that these columns possess typical reversed-phase chromatographic properties with increased hydrophobicity than polymethylsilsesquioxane and symmetric peaks for basic compounds. They were applied to the simultaneous separation of combination bendazol hydrochlorothiazide capsules containing polar and basic drugs with peaks identified by tandem with mass spectrometry. In general, a novel method is provided for the synthesis of different methyltrimethoxysilane-derived microspheres for high-performance liquid chromatography, which are advantageous for separating basic compounds.

Evaluation of silica from different vendors as the solid support of anion-exchange chiral stationary phases by means of preferential sorption and liquid chromatography.

Chromatographic performance of a chiral stationary phase is significantly influenced by the employed solid support. Properties of the most commonly used support, silica particles, such as size and size distribution, and pore size are of utmost importance for both superficially porous particles and fully porous particles. In this work, we have focused on evaluation of fully porous particles from three different vendors as solid supports for a brush-type chiral stationary phase based on 9-O-tert-butylcarbamoyl quinidine. We have prepared corresponding stationary phases under identical experimental conditions and determined the parameters of the modified silica by physisorption measurements and scanning electron microscopy. Enantiorecognition properties of the chiral stationary phases have been studied using preferential sorption experiments. The same material was slurry-packed into chromatographic columns and the chromatographic properties have been evaluated in liquid chromatography. We show that preferential sorption can provide valuable information about the influence of the pore size and total pore volume on the interaction of analytes of different size with the chirally-modified silica surface. The data can be used to understand differences observed in chromatographic evaluation of the chiral stationary phases. The combination of preferential sorption and liquid chromatography separation can provide detailed information on new chiral stationary phases.

Structural Analysis of Polygalaxanthones, C-Glucosyl Xanthones of Polygala tenuifolia Roots.

Polygalaxanthone III, a xanthone glycoside that is a major constituent of "Polygala Root" (Polygala tenuifolia roots, Onji in the Japanese Pharmacopoeia), has been used as a standard in the quality control of crude drugs. However, we previously noted differences in the chromatographic properties of one of three samples of polygalaxanthone III. Therefore, standardization of the standard itself is extremely important. The structures of three standard samples commercially available as polygalaxanthone III were characterized by LC/MS and NMR. LC/MS analysis revealed that two molecular types exist. Both types are chromatographically separable but have an identical mass number with distinguishable MS/MS spectra. One dimensional (1D)-NMR analyses demonstrated that both had the same xanthone moiety and heteronuclear multiple bond correlation (HMBC) analyses revealed that they are structural isomers at the connecting position of glucose to apiose 1-position. Consequently, the isomers were identified as polygalaxanthone III and its regioisomer, polygalaxanthone XI. Based on the findings, we recommend using the LC-MS/MS detection method, which discriminates polygalaxanthone III and XI, to confirm the quality of the standard.

Chromatography of Hedysarum extracts on modified Sepharose CL sorbents

Aim. To study the chromatographic efficiency of modified Sepharose CL-4B sorbents for separation, purification, and preparation of major components from Hedysarum theinum Krasnob. and Hedysarum neglectum Ledeb. root extracts.Materials and Methods. Sorption capacity of the native and modified sepharose was evaluated in both isocratic and gradient elution modes. 4-pyridinecarboxylic acid derivative [(4-hydroxy-3-methoxyphenyl)methylene]hydrazide (ftivazide) was used as a sorbent modifier ligand. We studied chromatographic properties of Sepharose CL-4B, Sepharose CL-6B, and azoepoxyadsorbent Sepharose CL-4B-DEP-p-nitrobenzohydrazide-ftivazide, a chemically modified analogue of Sepharose CL-4B.Results. Sepharose CL-4B and Sepharose CL-4B-DEP-p-nitrobenzohydrazide-ftivazide were found efficient carriers for liquid column chromatography of water-ethanol Hedysarum extracts. Complete elution of bioactive substances has been achieved exclusively by using sodium tetraborate as an eluent. We were able to identify flavonoids and tannins of different condensation in chromatographic fractions.Conclusion. Comparative chromatography of bioactive substances from Hedysarum root extracts is more efficient if employing Sepharose CL-4B instead of Sepharose CL-6B as a sorbent. Sepharose CL-4B-DEP-p-nitrobenzohydrazide-ftivazide demonstrates excellent separation capacity for tannins and flavonoids.

Gas chromatographic properties of methacrylate polymers modified by silver nanoparticles

The gas chromatographic properties of the porous methacrylate polymers, copolymers of 2,3-epoxypropyl methacrylate (2,3-EPMA) and ethylenedimethacrylate (EDMA), modified by silver nanoparticles were studied. The modification of the initial methacrylate copolymer by silver nanoparticles almost does not change the ability of the copolymer to dispersion interaction but results in an increase in polarity and enhances the possibility of specific interactions to occur. The aminated copolymer (copolymer of 2,3-EPMA and EDMA modified by diethylenetriamine using the reaction at the epoxy group) modified by silver nanoparticles demonstrates a higher ability to dispersion interactions and a lower polarity. Possibly, the sorption of silver particles on amino groups of the polymer is a reason for decreasing polarity of the polymer.

Influence of photo-initiators in the preparation of methacrylate monoliths into poly(ethylene-co-tetrafluoroethylene) tubing for microbore HPLC

In this study, poly(butyl methacrylate-co-ethyleneglycol dimethacrylate) polymeric monoliths were in situ developed within 0.75 mm i.d. poly(ethylene-co-tetrafluoroethylene) (ETFE) tubing by UV polymerization via three different free-radical initiators (α,α’-azobisisobutyronitrile (AIBN), 2,2-dimethoxy-2-phenylacetophenone (DMPA) and 2-methyl-4′-(methylthio)-2-morpholinopropiophenone (MTMPP). The influence of the nature of each photo-initiator and irradiation time on the morphological features of the polymer was investigated by scanning electron microscopy, and the chromatographic properties of the resulting microbore columns were evaluated using alkyl benzenes as test substances. The beds photo-initiated with MTMPP gave the best performance (minimum plate heights of 38 μm for alkyl benzenes) and exhibited a satisfactory reproducibility in the chromatographic parameters (RSD < 11%). These monolithic columns were also successfully applied to the separation of phenylurea herbicides, proteins and a tryptic digest of β-casein.

Quantitative structure – retention relationships of amino acids on the amino acid- and peptide-silica stationary phases for liquid chromatography

Quantitative structure – retention relationships analysis was applied to investigate the molecular retention mechanism of proteinogenic and non-proteinogenic amino acids on the amino acid- and peptide-silica stationary phases. Twelve stationary phases with chemically bonded amino acids of different types (glycine, alanine, phenylalanine, leucine, methionine, aspartic acid, and N-(9-Fluorenylmethoxycarbonyl)-O-tert-butyl-l-tyrosine) and chains lengths (amino acid, dipeptide, and tripeptide) were tested. In order to compare chromatographic properties of the prepared materials with the conventional columns, the amino-bonded phases (laboratory-prepared and commercial one) were also studied. For each of analyte, the molecular descriptors were calculated using quantum mechanics method. The QSRR models were determined using 13 molecular descriptors mainly related to the surface area, hydrophobicity, polarity, ion-exchange and hydrogen bonding capabilities of the analytes. Finally, the prediction potency of the molecular modeling descriptors-based models was also independently studied for the tested stationary phases using 15 training set and 6 test set of amino acids.

Polarity of chloroform eluent critical to quantification of arbuscular mycorrhizal biomass in soil using the neutral lipid fatty acid biomarker C16:1cis11

Arbuscular mycorrhizal fungi (AMF) form mutualisms with the majority of terrestrial plants. The fatty acid biomarker C16:1cis11, abundant in the neutral lipids, has become the preferred method for quantification of AMF biomass within the soil. The neutral lipids are eluted from silica gel with chloroform; however, commercially available chloroform is stabilized with varying amounts of ethanol (0.5–1.9%) which influences its chromatographic properties. At ethanol contents below 1.0%, neutral lipids are being lost to the glycolipid fraction thereby underestimating AMF biomass in soil. Thus, quantification of AMF biomass in soil using C16:1cis11 found in neutral lipids is highly dependent on the chloroform used. Combining the neutral and glycolipid fractions is proposed as one alternative to address this issue.

Application of reversed-phase thin layer chromatography and QSRR modelling for prediction of protein binding of selected β-blockers

Chromatographic properties of sixteen β-blockers were studied using planar chromatography. The aim of presented study was to investigate influence of different organic solvents (acetonitrile, methanol, dioxin and tetrahydrofurane) on β-blockers’ retention on C18 bonded silica gel stationary phase. Group of sixteen, diverse in terms of structure, beta blockers was used as a model set. The main goal of this study was to compare chromatographically estimated lipophilicity parameters with values obtained with the use of computational methods. Furthermore, in order to understand molecular mechanisms of retention better, quantitative structure-retention relationships (QSRR) analysis was performed. The next step was focused on application of chromatographically obtained lipophilicity parameters for prediction of protein binding (PB), based on quantitative retention–activity relationships (QRAR) approach. The obtained results showed that reversed-phase thin layer chromatography (RP-TLC), especially with tetrafydrofurane used as an organic modifier of mobile phase, is a useful tool for lipophilicity estimation, as well as for prediction of β-blockers’ biological properties. The QRAR model included C0 parameters for tetrahydrofuran-water as a mobile phase, as well as maximal projection area, and can be easily applied for prediction of systematically synthesized β-blockers structures’ PB.

Differentiation of o-, m-, and p-fluoro-α-pyrrolidinopropiophenones by Triton B-mediated one-pot reaction

Positional isomer differentiation is crucial for the analysis of forensic drugs. Presently, it is difficult to distinguish among ortho, meta, and para positional isomers of ring-fluorinated synthetic cathinones, a major class of new psychoactive substances (NPSs), because they exhibit similar chromatographic properties and mass spectral patterns. We describe herein that the ring-fluorinated synthetic cathinone positional isomers, viz. o-, m-, and p-fluoro-α-pyrrolidinopropiophenones (o-, m-, and p-FPPPs), can be discriminated by their benzyltrimethylammonium hydroxide (Triton B)-mediated one-pot reaction with methanol at ambient temperature, followed by chromatographic and mass spectral analyses of the corresponding products. For p-FPPP, fluorine was nucleophilically substituted by the methoxy group to afford p-methoxy-α-pyrrolidinopropiophenone, while o- and m-FPPPs afforded the corresponding FPPP-enamine–pyrrolidine adducts, which allowed the above positional isomers to be unambiguously differentiated by comparing the reaction product chromatograms and mass spectra. The adopted approach, which does not require excess heating or use of metallic catalysts and features the advantages of simplicity and convenience, is expected to contribute toward practical NPS identification.

Manipulating selectivity of covalently-bonded hyperbranched anion exchangers toward organic acids. Part III: Effect of diamine structure in the external part of the functional layer

The chromatographic properties of three hyperbranched anion exchangers having various diamines in the external part of the functional layer are studied in order to reveal diamine influence on selectivity toward mono- and divalent organic acids. The obtained stationary phases have the same structure of the internal part of the functional layer formed by repeating 4 modification cycles including alkylation with 1,4-butanediol diglycidyl ether (1,4-BDDGE) and amination with methylamine (MA) and differ by the structure of diamine used in the 5th modification cycle. For the first time several diamines (ethylenediamine, (2-aminoethyl)aminoethanol, and N,N’-bis(2-hydroxyethyl)ethylenediamine) are used for completing the last modification cycle in hyperbranching. The performance of three prepared anion exchangers is investigated using KOH and NaHCO3 as eluents and discussed with respect to the differences in hydrophilicity of the external part of the functional layer showing its effect on the separation of organic acids.

Rapid and Simple Detection of Ochratoxin A using Fluorescence Resonance Energy Transfer on Lateral Flow Immunoassay (FRET-LFI).

The detection of mycotoxins is crucial because of their toxicity in plants, animals, and humans. It is very important to determine whether food products are contaminated with mycotoxins such as ochratoxin A (OTA), as mycotoxins can survive heat treatments and hydrolysis. In this study, we designed a fluorescence resonance energy transfer (FRET)-based system that exploits antibody-antigen binding to detect mycotoxins more rapidly and easily than other currently available methods. In addition, we were able to effectively counteract the matrix effect in the sample by using a nitrocellulose membrane that enabled fluorescence measurement in coffee samples. The developed FRET on lateral flow immunoassay (FRET-LFI) system was used to detect OTA at a limit of detection (LOD) of 0.64 ng∙mL−1, and the test can be completed in only 30 min. Moreover, OTA in coffee samples was successfully detected at a LOD of 0.88 ng∙mL−1, overcoming the matrix effect, owing to the chromatographic properties of the capillary force of the membrane. We believe that the developed system can be used as a powerful tool for the sensitive diagnosis of harmful substances such as mycotoxins and pesticides for environmental and food quality control monitoring.

Indole Alkaloids from the Aerial Parts of Kopsia fruticosa

A phytochemical investigation of the ethanol extract of Kopsia fruticosa resulted in the isolation of two new indole alkaloids, 3-oxo-11,12-dimethoxy-N1-decarbomethoxy-14,15-didehydrochanofruticosinate (1) and 11,12-dimethoxydanuphylline (2). Their structures were established on the basis of extensive chromatographic properties and chemical and physicochemical methods.