Fullerenes separation with monomeric type C30 stationary phase in high-performance liquid chromatography

The temperature effect on the separation of fullerenes in LC was examined using monomeric type C30, C18 and C8 alkyl bonded stationary phases. It appears that the C30 phase exhibits superior separation ability for fullerenes. It is observed that the maximum retention temperature of fullerenes on the C30 phase is around 20 degrees C. A strong correlation between the changes in NMR spectra and the retention behavior of the solutes was found. The interpretation of the retention behavior of fullerenes on the alkyl bonded stationary phases, including the behavior in subambient temperature, is discussed using the information obtained by CP-MAS solid-state NMR spectroscopy and LC.     

Selectivity of calixarene‐bonded silica phases in HPLC: Description of special characteristics with a multiple term linear equation at different methanol concentrations

Retention and selectivity characteristics of different calixarene‐, resorcinarene‐ and alkyl‐bonded stationary phases are examined by analyzing a set of test solutes covering the main interactions (hydrophobic, steric, ionic, polar) that apply in HPLC. Therefore Dolan and Snyder's multiple term linear equation has been adapted to fit the properties of calixarene‐bonded columns. The obtained parameters are used to describe retention and selectivity of the novel Caltrex^® phases and to elucidate underlying mechanisms of retention. Here, differences of stationary phase characteristics at different methanol concentrations in the mobile phases are examined. Both selectivity and retention were found to depend on the methanol content. Differences of these dependencies were found for different stationary phases and interactions. The differences between common alkyl‐bonded and novel calixarene‐bonded phases increase with increasing methanol content.     

Influence of stationary phase solvation on shape selectivity and retention in reversed-phase liquid chromatography

In the past few decades, shape selectivity has drawn a great deal of attention from chromatographers. The chemistry and characteristics of bonded stationary phases such as phase type, length of bonded phase, surface coverage, and silica surface material have an effect on the shape selectivity of the columns. Although the effects of bonded phase shape selectivity are relatively well understood, one remaining question is the effect of intercalated solvent on shape selectivity. The intercalation of organic modifier and water molecules into the stationary phase is believed to introduce more rigidity into bonded alkyl chains in RPLC. The use of gas chromatography (GC) opens a new dimension to approach this question. C18 columns 4 cm in length were prepared in our laboratory and used in both LC and GC experiments. Shape selectivity and thermodynamic constants for the transfer of a solute from the mobile phase to the stationary phase have been determined as a function of monomeric octadecyl stationary phase bonding densities over the range of 1.44-3.43 micromol/m2 and a polymeric phase (nominal surface coverage 4.77 micromol/m2). Comparing LC and GC experiments, we observed: (a) similar relationships between shape and phenyl selectivities with monomerically bonded C18 phase densities; (b) different correlation of thermodynamic quantities (DeltaH degrees , DeltaS degrees , and DeltaG degrees ) versus bonded phase densities. The effects of high temperature and residual silanol groups are sources of difficulty in elucidation of the intercalated mobile phase role in selectivity and retention for GC measurements.     

Characterization of calixarene‐bonded stationary phases

Calixarene‐bonded stationary phases received growing interest in HPLC as stationary phases with special retention characteristics and selectivity. The commercially available unsubstituted and p‐tert‐butyl‐substituted Caltrex^® columns have been intensively studied and characterized in our workgroup. They can be used as reversed phases, yet they support additional interactions. Especially, their steric, polar and ionic properties differ from conventional alkyl‐bonded phases. However, also the hydrophobic interaction shows differences since adsorption and partition interactions on or in a bonded layer of calixarenes are not similar to those of alkyl‐bonded layers. The relative strength of the hydrophobic properties of the stationary phases has been found depending on the methanol concentration of the mobile phase. Generally, the dependencies of their interaction strengths on mobile‐phase conditions, e.g. the change of the intensity of the hydrogen‐bonding abilities with decreasing methanol content, are not similar from phase to phase either. This probably gives calixarene‐bonded stationary phases enhanced suitability for analyses at extreme compositions of the mobile phase. An overview about the synthesis, retention and selectivity properties of Caltrex^® columns is given here.     

Separation of polycyclic aromatic hydrocarbons with a C60 bonded silica phase in microcolumn liquid chromatography

A chemically bonded C₆₀ silica phase was synthesized as a stationary phase for liquid chromatography (LC) and its retention behavior evaluated for various polycyclic aromatic hydrocarbons (PAHs) using microcolumn LC. The results indicate that the C₆₀ bonded phase offers selectivity different from that of octadecylsilica (ODS) bonded phases in the separation of isomeric PAHs. With the C₆₀ phase, PAH molecules having a partial structure similar to that of the C₆₀ molecule, e.g. triphenylene and perylene, were retained longer than with ordinary ODS stationary phases. The results also show that good correlation exists between the retention data with this C₆₀ bonded phase and with C₆₀ itself as the stationary phase.     

Effect of column temperature on the retention of peropyrene-type polycyclic aromatic hydrocarbons on various chemically bonded stationary phases in reversed-phase liquid chromatography

The effect of column temperature on the reversed-phase retention of polycyclic aromatic hydrocarbons has been investigated using various chemically bonded phases. Four solutes, coronene, tetrabenzo[a,cd,j,lm]perylene, tetrabenzo[a,cd,f,lm]perylene and benzo[lm]phenanthro[4,5,6-abcd]perylene, were used as the test probes. The temperature dependences of the retention are almost linear (the logarithm of capacity factor is proportional to the reciprocal of the column absolute temperature) with monomeric C18, monomeric C18 with endcapping and diphenyl bonded phases, while non-linear behaviour was observed with polymeric C18 phases. These differences in behaviour of the stationary phases are interpreted in terms of their structural differences.     

Temperature effect in separation of fullerene by high-performance liquid chromatography

Summary The effect of column temperature, especially at low temperatures, on the separation of fullerenes on monomeric and polymeric octadecyl silica (ODS) bonded phases has been studied. Decreasing the column temperature induces an increase in selectivity. The best temperature for the separation of fullerenes was determined for both types of ODS phase with n-hexane eluent. The selectivity for higher fullerenes on monomeric phases becomes similar to that on polymeric phases to low temperature. It has been found that as the carbon content of monomeric phases is increased, the selectivity also becomes similar to polymeric phases.     

Preparation of pyrenebutyric acid bonded silica stationary phases for the application to the separation of fullerenes

A novel immobilization method was proposed for the preparation of pyrenebutyric acid-bonded silica (PYB-silica) stationary phases. The pyrene moiety was grafted to silica gel through spacers of aminoalkyl silanes. The HPLC separation of C60, C70 and higher fullerenes on the new pyrenebutyric acid-bonded silica stationary phases was also studied. Based on the temperature effect, the intermolecular interaction between stationary phases and solutes and the retention mechanism were discussed. The results of column loading capacity test demonstrated the potential for the separation of fullerenes in large amounts on the PYB-silica stationary phases.     

A humic acid stationary phase for the high performance liquid chromatography separation of buckminsterfullerenes: theoretical and practical aspects

The influence of the mobile phase composition and column temperature on the chromatographic separation of five buckminsterfullerenes (C60, C70, C76, C78, C84) on a stationary phase based on silica gel with chemically bonded humic acid (Bonded humic acid column (BHAC)) was studied. The retention behavior of the fullerenes was measured under isocratic conditions with different mobile phase compositions, ranging from 0.05-0.70 (v/v) of toluene in cyclohexane. The column temperature was analysed in the range 35-75 °C. The retention factors of the five fullerenes do not depend linearly on the toluene fraction but follow a quadratic relationship. The best chromatographic conditions for baseline separation of the five fullerenes were selected. The retention of the fullerenes on the HA stationary phase was strongly affected by temperature. Positive values of thermodynamic parameters (changes of enthalpy and entropy) were due to the abnormal solubility behaviour of fullerenes in toluene in the temperature range 35-75 °C. The information obtained in this work makes this BHAC very simple to prepare and low cost, useful for fullerene research applications.     

Retention and selectivity tests of silica-based and metal-oxide bonded stationary phases for RP-HPLC

Chromatographic properties of silica-, zirconia- and alumina-based columns with octadecyl-, polyethylene glycol- and pentafluorophenylpropyl-bonded stationary phases were tested. Selectivities of nine columns for LC were characterized using chromatographic methods including Walters, Engelhardt, Tanaka and Galushko hydrophobicity and silanol activity tests, measurements of methylene selectivity in various aqueous-methanol and aqueous-acetonitrile mobile phases and of gradient lipophilic capacity as a measure of the effect of the sample hydrophobicity on gradient-elution separations. A semi-empirical interaction indices model, assuming a predominant role of the solvophobic interactions of test compounds with different polarities, was compared with the linear free energy relationships approach taking into account selective polar interactions. The interaction indices model was applied to both non-polar stationary phases bonded on silica, alumina and zirconia supports, and to the non-modified adsorbents in the normal-phase LC. The retention data of isomeric naphthalene disulfonic acids were used to compare the attractive and repulsive ionic interactions of the columns in purely aqueous mobile phases. The results of the hydrophobicity and polarity tests were consistent, and allowed column characterization and classification. Silanol activity was important with octadecyl silica columns, but was relatively insignificant with bonded polyethylene glycol and pentafluorophenylpropyl phases on silica gel support. Polar interactions with the alumina and zirconia support materials significantly affect the retention.     

Retention properties of novel β-CD bonded stationary phases in reversed-phase HPLC mode

With the given special structures, the CD bonded stationary phases are expected to have complementary retention properties with conventional C18 stationary phase, which will be helpful to enhance the polar selectivity in RP mode separation. In this work, two β-cyclodextrin (β-CD) bonded stationary phases for reversed-phase HPLC, including 1, 12-dodecyldiol linked β-CD stationary phase (CD1) and olio (ethylene glycol) (OEG) linked β-CD stationary phase (CD2), have been synthesized via click chemistry. The resulting materials were characterized with FT-IR and elemental analysis, which proved the successful immobilization of ligands. The similarities and differences in retention characteristics between the CD and C18 stationary phases have been elucidated by using comparative linear solvation energy relationships (LSERs). The force related to solute McGowan volume has no significant difference, while the hydrogen bonding and dipolar interactions between solutes and CD stationary phases are stronger than between solutes and C18, which is attributed to the special structures (CD and triazole groups) of CD stationary phases. Chemical origins are interpreted by comparison between CD1 and CD2. Similar dispersive interactions of CD1 and CD2 are attributed to their similar length of spacer arms. CD2 which contains OEG spacer arm has relative weaker HBD acidity but stronger HBA basicity. CD stationary phases display no serious different methylene selectivity and higher polar selectivity than in the case of C18. Higher acid selectivity and lower basic selectivity are observed on CD2 than on CD1. Distinctive retention properties and good complementary separation selectivity to C18 make the novel CD bonded stationary phases available for more application in RPLC.     

Retention of cyanoalkanes and cyanoalkylbenzenes by bonded stationary phases in supercritical fluid chromatography

Summary The retention of cyanoalkanes and cyano-alkylbenzenes during SFC was investigated on alkyl and cyanoalkyl-bonded stationary phases and compared to alkane retention. The particular behaviour of short chain homologues was attributed to a silanophilic interaction with surface-OH groups on the silica and inhibition of specific interactions between solutes and C₁₀CN bonded groups.     

Effect of the alkyl chain length of the bonded stationary phase on solute retention in reversed-phase high-performance liquid chromatography

Summary A comparison of the chromatographic retention characteristics of nonpolar bonded stationary phases was investigated. The results show that the interactions between the solutes and the stationary phases having C₂, C₈ and C₁₈ alkyl groups are almost similar in the range of the mobile phase investigated. This interaction is considered as the solvophobic effect.     

Comparison of four cholesterol‐based stationary phases for the separation of steroid hormones

The chromatographic behavior of steroid hormones on four cholesterol‐bonded stationary phases with different structures in binary methanol/water mobile phases was studied. Of the stationary phases tested, the commercially available stationary phases Cogent UDC cholesterol? and COSMOSIL cholester? provided better separations of steroid hormones in comparison to homemade aminocholesterol and diaminocholesterol stationary phases. The results show that the temperature has a significant influence on the retention and selectivity for steroid hormones separation. The temperature increase may cause changes in the elution order. From the dependences of the retention (ln k) on temperature (1/T), the standard partial molar enthalpy and standard partial molar entropy were calculated and their enthalpic and entropic contributions to the retention were compared. The enthalpic effects principally control the retention mechanism.     

Effect of coverage density and structure of chemically bonded silica stationary phases on the separation of compounds with various properties

The chemical character, geometry, and architecture of chemically formed surface layers determine interactions between stationary phase, analyte, and mobile phase, and therefore the retention mechanisms (partitioning, adsorption, ion exchange, steric exclusion) of separated analytes. These interactions also depend on the structure and chemical character of the solutes and the composition of the mobile phase. High-molecular-weight fullerenes (C60 and C70) and water-soluble selenium-containing peptides (833 and 2607 Da) were used for the evaluation of laboratory-prepared octadecyl stationary phases with high and low coverage density before and after end-capping. The aim of this work was to study differences in surface coverage density and homogeneity and conformational changes of chemically bonded moieties and the influence of these parameters on the separation of mixtures of selenopeptides and fullerenes with significantly different molecular masses. A topographical model of the chemically modified stationary surface is presented.     

Effects of stationary-phase polarity on retention in reversed bonded phase HPLC columns

Summary Variations in retention and selectivity have been studied in cyano, phenyl and octyl reversed bonded phase HPLC columns. The retention of toluene, phenol, aniline and nitrobenzene in these columns has been measured using binary mixtures of water and methanol, acetonitrile or tetrahydrofuran mobile phases in order to determine the relative contributions of proton donor-proton acceptor and dipole-dipole interactions in the retention process. Retention and selectivity in these columns was correlated with polar group selectivities of mobile phase organic modifiers and the polarity of the bonded stationary phases. In spite of the prominent role of bonded phase volume and residual silanols in the retention process, each column exhibited some unique selectivities when used with different organic modifiers.     

Separation and identification of higher fullerenes in soot extract by liquid chromatography-mass spectrometry

Summary Two-step liquid chromatographic separation (LC) has been applied to soot extract and the identification of higher fullerenes has been accomplished by LC-MS measurements using an ESI interface. The first separation step is preparative-scale LC using a 50 mm i.d. column packed with monomeric octadecylisilica (ODS) because elution is mainly controlled by relative molecular mass. 39 batches of five fractions each were collected and then as the second separation step each fraction was analysed by analytical-scale LC using a conventional column of a polymeric ODS phase which can elute fullerenes according to shape and structure. This stationary phase can also separate many isomers of higher fullerenes, consequently the existence of several higher fullerenes larger than C₈₆ has been confirmed and their UV-Vis spectra were obtained by the photodiode array detection system coupled to the analytical LC.     

Insights into the retention mechanism of neutral organic compounds on polar chemically bonded stationary phases in reversed-phase liquid chromatography

The solvation parameter model is used to characterize the retention properties of a 3-aminopropylsiloxane-bonded (Alltima amino), three 3-cyanopropylsiloxane-bonded (Ultrasphere CN, Ultremex-CN and Zorbax SB-CN), a spacer bonded propanediol (LiChrospher DIOL) and a multifunctional macrocyclic glycopeptide (Chirobiotic T) silica-based stationary phases with mobile phases containing 10 and 20% (v/v) methanol-water. The low retention on the polar chemically bonded stationary phases compared with alkylsiloxane-bonded silica stationary phases arises from the higher cohesion of the polar chemically bonded phases and an unfavorable phase ratio. The solvated polar chemically bonded stationary phases are considerably more hydrogen-bond acidic and dipolar/polarizable than solvated alkylsiloxane-bonded silica stationary phases. Selectivity differences are not as great among the polar chemically bonded stationary phases as they are between the polar chemically bonded phases and alkylsiloxane-bonded silica stationary phases.     

Chromatographic Behaviour and Lipophilicity of Estradiol Derivatives

The retention behaviour of estradiol derivatives has been studied by HPLC on polar chemically bonded stationary phases: C₃CN, DIOL and C₃NH₂, commercially available columns. The mobile phases used were: methanol-water and acetonitrile-water in various proportions. Reversed-phase chromatography occurred on polar chemically bonded stationary phases. Correlation between the retention constants of estradiol derivatives obtained on polar chemically bonded phases and log P calculated via different methods was examined too.     

The role of pore size and stationary phase composition in preventing aqueous-induced retention time loss in reversed-phase HPLC

The phenomenon of aqueous mobile phase induced retention time loss, which has also been referred to as phase-collapse behavior, is investigated using a variety of stationary phases with an aqueous mobile phase. The loss of retention of several water-soluble B vitamins is measured as a function of silica pore size, bonded-phase chemistry, and bonded stationary phase density. It is found that these variables influence the magnitude of the retention time loss behavior and that controlling/optimizing these variables can result in a stationary phase with sufficient retention of the analyte without the column exhibiting the so-called phase-collapse behavior.