Retention versatility of silica-supported comb-shaped crystalline and non-crystalline phases in high-performance liquid chromatography

Silica-supported comb-shaped polymer (Sil-ODA18) provided a specific selectivity towards aromatic and non-aromatic solutes. Selectivity of Sil-ODA18 and ODS (polymeric and monomeric) columns towards polycyclic aromatic hydrocarbons and geometric isomers have been discussed in respect of molecular shapes and sizes. The retention versatility of this phase arises from the orientation change on transition from the ordered crystalline to the less ordered non-crystalline state. Selectivity in the crystalline and non-crystalline regions of Sil-ODA18 is similar to that of polymeric and monomeric ODS phases, respectively. Sil-ODA18 column also exhibited the characteristic temperature dependency of liquid-crystalline phases, i.e., an increase in retention with increasing temperature on transition from crystalline to isotropic state.     

Retention of C60 and C70 fullerenes on reversed-phase high-performance liquid chromatographic stationary phases.

The separation of C60 and C70 fullerenes on four different polysiloxane stationary phases was examined. It was determined that polar solvents can be used as mobile phases effectively for the separation of fullerene molecules. Unlike previously published work, a polymeric octadecyl siloxane (ODS) stationary phase provided higher separation factors for C70/C60 than did monomeric ODS stationary phases or phenyl substituted stationary phases. For example, for a methanol-diethyl ether (50:50, v/v) mobile phase and C60, k' approximately 5.0 separation factors, alpha = 3.3, were achieved with polymeric ODS compared to alpha = 2.2, with a monomeric ODS stationary phase. A linear solvation energy relationship (LSER) was used to model the importance of solvent interactions and stationary phase interaction to solute retention.     

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.     

Temperature optimization for the separation of PAHs on micropacked LC ODS columns

The effect of column temperature on the separation of the sixteen polycyclic aromatic hydrocarbons (PAHs) of mixture SRM 1647a of the US Environmental Protection Agency has been studied on different micropacked ODS columns. Isothermal temperature optimization was successfully used for complete separation of the PAHs on a polymeric ODS stationary phase, whereas temperature programmed conditions were selected for separation on a monomeric ODS stationary phase.     

HPLC separation of triacylglycerol positional isomers on a polymeric ODS column

A polymeric ODS column was applied to the resolution of triacylglycerol positional isomers (TAG-PI), i.e. 1,3-dioleoyl-2-palmitoyl-glycerol (OPO) and 1,2-dioleoyl-3-palmitoyl-rac-glycerol (OOP), with a recycle HPLC system. To investigate the ODS column species and the column temperatures for the resolution of a TAG-PI pair, a mixture of OPO and OOP was subjected to an HPLC system equipped with a non-endcapped polymeric, endcapped monomeric, endcapped intermediate, or non-endcapped monomeric ODS column at three different column temperatures (40, 25, or 10 degrees C). Only the non-endcapped polymeric ODS column achieved the separation of OPO and OOP, and the lowest column temperature (10 degrees C) showed the best resolution for them. The other pair of TAG-PI, a mixture of 1,3-dipalmitoyl-2-oleoyl-glycerol (POP) and 1,2-dipalmitoyl-3-oleoyl-rac-glycerol (PPO) was also subjected to the system equipped with a non-endcapped polymeric or monomeric ODS column at five different column temperatures (40, 32, 25, 17, and 10 degrees C). Thus, POP and PPO were also separated on only the non-endcapped polymeric ODS column at 25 degrees C. However, no clear peak appeared at 10 degrees C. These results would indicate that the polymeric ODS stationary phase has an ability to recognize the structural differences between TAG-PI pairs. Also, the column temperature is a very important factor for separating the TAG-PI pair, and the optimal temperature would relate to the solubility of TAG-PI in the mobile phase. Furthermore, the recycle HPLC system provided measurements for the separation and analysis of TAG-PI pairs.     

Separation of carotenoids by high-performance liquid chromatography with polymeric and monomeric octadecylsilica stationary phases

Summary In addition to their value in the nutritional context, the carotenoids have other important functions, including some epidemiological significance in disease prevention. With increasing interest in the carotenoids methods for their characterization and quantification in various matrices are essential particularly with regard to epidemiologic studies dealing with diet and health. Since high-performance liquid chromatography (LC) is the most promising analytical method for this purpose, having high reliability, high selectivity and quantification ability, this work presents the evaluation of reversed-phase LC methods using polymeric and monomeric octadecylsilica (ODS) stationary phases for the highly selective separation of carotenoids. It has been found that the polymeric ODS has a better selectivity for carotenoids, taking account of their molecular shape and size recognition, than the monomeric ODS phase and that the former is more suitable than the latter for separations of the carotenoids.     

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

Summary 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.     

Evaluation of microstructural features of a new polymeric organic stationary phase grafted on silica surface: A paradigm of characterization of HPLC-stationary phases by a combination of suspension-state H NMR and solid-state C-CP/MAS-NMR

Silica-supported poly(octadecylacrylate) (Sil-ODA_n), polymeric octadecylsilyl silica (polymeric ODS), and monomeric octadecylsilyl silica (monomeric ODS) were studied by a combination of suspension-state ¹H NMR and solid-state ¹³C CP/MAS-NMR to probe the mechanisms underlying their functions as stationary phases for RP-HPLC. Sil-ODA_n, with a strong temperature dependent separation behaviour showed correspondent temperature dependent manifestations in both suspension-state ¹H NMR and solid-state ¹³C CP/MAS-NMR experiments. With a gradual increase in temperature, intensity of proton signals (¹H NMR) of octadecyl moieties (mainly methylene groups) rose dramatically. This dramatic rise was at the same temperature of an endothermic peak detectable in its DSC thermogram indicating a relatively complete solid to liquid phase transition. In addition temperature dependencies of the ratio of trans to gauche conformed well to temperature dependencies of the separation factor between naphthacene and triphenylene (as a simple indicator of shape selectivity). Therefore NMR spectra of Sil-ODA_n were used as a reference for ascertaining percentage of octadecyl moieties of liquid type mobility in the two other stationary phases. Using this method we determined percentage of liquid phase in polymeric ODS and monomeric ODS at various temperatures. We suggest a combination of suspension-state ¹H NMR and solid-state ¹³C CP/MAS-NMR for structure-dynamic characterization of various kinds of hydrocarbon chains grafted onto the silica particles.     

Retention behavior of large polycyclic aromatic hydrocarbons on cholesteryl 10-undecenoate bonded phase in microcolumn liquid chromatography

Summary The retention behavior of large polycyclic aromatic hydrocarbons (PAH) in reversed-phase microcolumn liquid chromatography has been found to depend primarily on the inherent planarity of the molecules, the degree of orderliness of the bonded phase, and the concentration of solvent. The planarity recognition capability of cholesteryl 10-undecenoate bonded phase for large PAH is comparable with that of monomeric ODS phases. During solvation, however, the phase characteristics seemed to suggest a fairly rigid structure (an ordered phase), i.e. increased selectivity for non-planar molecules which approaches that of polymeric ODS rather than that of other phases with similarly bulky groups, e.g. naphthylethyl and pyrenylethyl bonded phases.     

Molecular Shape Selectivity for Polycyclic Aromatic Compounds on a Core–Shell Octadecylsilica Stationary Phase at Subambient Column Temperatures

Molecular shape selectivity for polycyclic aromatic compounds on a core–shell-type octadecylsilica (ODS) phase at subambient column temperatures was studied in reversed-phase liquid chromatography. The plate height on the core–shell ODS column was relatively stable at subambient column temperatures when compared with that of a conventional ODS column. In order to compare the sample diffusivities in the conventional and core–shell ODS columns, van Deemter plots were prepared. The plate height of the core–shell column was significantly lower than that of conventional column, suggesting an advantageous feature of the core–shell-type stationary phase especially at a high flowrate of the mobile phase. An enhanced molecular shape recognition capability of the core–shell ODS phase was also confirmed at subambient column temperature. The result could be consistent with an improved shape selectivity as normally observed on conventional ODS phases at low temperatures, however, the enhanced molecular shape recognition capability of the core–shell phase enables a good separation between benz[a]anthracene and chrysene at subambient column temperatures. Similar improved shape selectivities for terphenyl isomers were also confirmed on the core–shell phase.

     

Molecular Shape Selectivity for Polycyclic Aromatic Compounds on a Core–Shell Octadecylsilica Stationary Phase at Subambient Column Temperatures

Molecular shape selectivity for polycyclic aromatic compounds on a core–shell-type octadecylsilica (ODS) phase at subambient column temperatures was studied in reversed-phase liquid chromatography. The plate height on the core–shell ODS column was relatively stable at subambient column temperatures when compared with that of a conventional ODS column. In order to compare the sample diffusivities in the conventional and core–shell ODS columns, van Deemter plots were prepared. The plate height of the core–shell column was significantly lower than that of conventional column, suggesting an advantageous feature of the core–shell-type stationary phase especially at a high flowrate of the mobile phase. An enhanced molecular shape recognition capability of the core–shell ODS phase was also confirmed at subambient column temperature. The result could be consistent with an improved shape selectivity as normally observed on conventional ODS phases at low temperatures, however, the enhanced molecular shape recognition capability of the core–shell phase enables a good separation between benz[a]anthracene and chrysene at subambient column temperatures. Similar improved shape selectivities for terphenyl isomers were also confirmed on the core–shell phase.     

Effect of temperature on the mechanism of retention of fullerenes in liquid chromatography using various alkyl bonded stationary phases

Summary The temperature-dependency of the separation of fullerenes in liquid chromatography (LC) has been examined using various alkyl bonded stationary phases. It has been found that a maximum retention temperature exists with long alkyl bonded stationary phases, whereas there is no similar effect with the newly synthesized alkyl bonded phases which have two phenyl groups at the base of the bonded phase. The interpretation of the retention behavior of fullerenes in the low temperature region on alkyl bonded stationary phases is discussed using information obtained by CP-MAS solid-state NMR spectroscopy and LC.     

Selectivity enhancement for the separation of tocopherols and steroids by integration of highly ordered weak interaction sites along the polymer main chain

A novel alternating copolymer-based organic phase was synthesized using a new N-substituted maleimide monomer for the development of alternating copolymer-grafted silica for high-performance liquid chromatographic applications. This new monomer (DGMI) was copolymerized with octadecyl acrylate (ODA) from 3-mercaptopropyltrimethoxysilane-grafted silica to produce Sil-poly(ODA-alt-DGMI). The organic phase was characterized by the elemental analysis and the diffuse reflectance infrared Fourier transform spectroscopy. Tocopherol isomers and steroids were used as analytes for the evaluation of the chromatographic selectivity profiles of this novel stationary phase. The selectivity of this column was then compared with a polymeric ODS column and previously developed another alternating copolymer-grafted silica (without the glutamide-derived moiety) column, Sil-poly(ODA-alt-N-octadecylmaleimide). The complete baseline separation of tocopherol isomers in an isocratic mode has been achieved within 25 min with the Sil-poly(ODA-alt-DGMI). The separation of eight kinds of estrogenic steroids and corticoids has also been achieved in an isocratic mode with this column. Significant differences in separation selectivity between Sil-poly(ODA-alt-DGMI) and polymeric ODS columns were observed towards the steroids, and compared with the reference columns, a better separation profile for these analytes was obtained with the Sil-poly(ODA-alt-DGMI). The results of this investigation indicated that the enhancement of selectivity of Sil-poly(ODA-alt-DGMI) towards the test analytes arose from the multiple interaction mechanism such as hydrophobic effect, carbonyl-π and hydrogen-bonding interactions, and such integrated interactions originated from the addition of two amide groups in the N-substituted maleimide monomer.     

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.     

High performance liquid chromatographic separation of polar and non-polar chlorophyll pigments from algae using a wide pore polymeric octadecylsilica column

A method employing a wide pore polymeric reversed phase column has been developed for the separation of most of the chlorophylls and related compounds previously described as occurring in marine microalgae. The high selectivity toward molecular shape of this kind of stationary phase has enabled compounds of very similar structure, such as chlorophylls c₁, c₂ and Mg-divinylpheoporphyrin a₅ monomethyl ester, and chlorophyll a and the phytol-substituted chlorophyll c-like pigments, which commonly coelute on monomeric bonded phases, to be resolved in a single run. Some of these pigments, formerly thought to be a single compound, have, in fact, been demonstrated to be groups of two or more. The method has been successfully applied to both algal cultures and natural sea water samples. When visible light absorbance detection was used, the method proved suitable for separation of various carotenoids.     

Molecular shape recognition capability of liquid-crystal bonded phases in reversed-phase high performance liquid chromatography

Summary The chromatographic retention behaviour of two liquidcrystal bonded phases have been evaluated using polycyclic aromatic hydrocarbons (PAHs) as the probe samples in reversed-phase high performance liquid chromatography (RP-HPLC). The results clearly indicate that these phases have better planarity and shape recognition capabilities than commercially-avaialble polymeric octadecylsilica (ODS) phases whose strong planarity and shape selectivities were found earlier. It can also be concluded from the chromatographic observations that the shape recognition capability of these phases is dependent on both mobile phase composition and column temperature, but that the effect of mobile phase and temperature on the shape selectivity work independently. The retention behaviour can be explained by changes in the phase structure with changes of eluent composition and temperature.     

Reversible solvent and temperature induced “monomeric-like” —“Polymeric-like” transitions in alkyl bonded silica

Summary The study of behaviour of homologous series (and more precisely, the plots of methylene selectivity versus the carbon number, n, of the alkyl chain of the homologue) on monomeric and polymeric phases in liquid chromatography enabled us to characterize two kinds of retention mechanism. These mechanisms are not independent; changes in the temperature or mobile phase composition allow passage from one to the other. The first, for which the methylene selectivity versus n decreases with a discontinuity, defines a retention mechanism where solutes are inserted between the grafted chains. It occurs for monomeric phases at ambient temperature, for polymeric phases, at high temperature or with a strong solvent like THF, CH₂Cl₂, AcMe or CHCl₃. The second gives a plot of methylene selectivity versus n with a maximum. It is consistent with a partition mechanism with total immersion of the solute in the bed of the stationary phase. It occurs for a polymeric phase at ambient temperature or for a monomeric one at low temperature.     

Comparison of a polymeric pseudostationary phase in EKC with ODS stationary phase in RP‐HPLC

Poly(stearyl methacrylate‐co‐methacrylic acid) (P(SMA‐co‐MAA)) was induced as pseudostationary phase (PSP) in electrokinetic chromatography (EKC). The n‐octadecyl groups in SMA were the same as that in octadecylsilane (ODS) C18 column. Thus, the present work focused on the comparison of selectivity between polymeric PSP and ODS stationary phase (SP), and the effect of organic modifiers on the selectivity of polymeric PSP and ODS SP. 1‐butanol could directly interacted with PSP as a Class I modifier, and improved both of the methylene selectivity and polar group selectivity. When the analysis times were similar, the polymeric PSP exhibited better methylene selectivity and polar group selectivity. Although the hydrophobic groups were similar, the substituted benzenes elution order was different between polymeric PSP and ODS SP. Linear solvation energy relationships (LSER) model analysis found that polymeric PSP and ODS SP exhibited two same key factors in selectivity: hydrophobic interaction and hydrogen bonding acidity. But polymeric PSP exhibited relatively strong n‐ and π‐electrons interaction to the analytes.     

Selectivity comparisons of monolithic silica capillary columns modified with poly(octadecyl methacrylate) and octadecyl moieties for halogenated compounds in reversed-phase liquid chromatography

Stationary phase selectivities for halogenated compounds in reversed-phase HPLC were compared using C18 monolithic silica capillary columns modified with poly(octadecyl methacrylate) (ODM) and octadecyl moieties (ODS). The preferential retention of halogenated benzenes on ODM was observed in methanol/water and acetonitrile/water mobile phases. In selectivity comparison of selected analytes on ODM and ODS, greater selectivities for halogenated compounds were obtained with respect to alkylbenzenes on an ODM column, while similar selectivities were observed with a homologous series of alkylbenzenes on ODM and ODS columns. These data can be explained by greater dispersive interactions by more densely packed octadecyl groups on the ODM polymer coated column together with the contribution of carbonyl groups in ODM side chains. For the positional isomeric separation of dihalogenated benzenes (ortho-, meta-, para-), the ODM column also provided better separation of these isomers for the adjacently eluted isomers that cannot be completely separated on an ODS column in the same mobile phase. These results imply that the ODM column can be used as a better alternative to the ODS column for the separation of other halogenated compounds.     

Contributions to reversed-phase column selectivity. I. Steric interaction

In reversed-phase chromatography (RPC), the restricted retention of "bulky" solutes can occur in one of two ways, giving rise to either "shape selectivity" or "steric interaction." Starting with data for 150 solutes and 167 monomeric type-B alkylsilica columns, the present study examines the steric interaction process further and compares it with shape selectivity. The dependence of column hydrophobicity and steric interaction on column properties (ligand length and concentration, pore diameter, end-capping) was determined and compared. The role of the solute in steric interaction was found to be primarily a function of solute molecular length, with longer solutes giving increased steric interaction. We find that there are several distinct differences in the way shape selectivity and steric interaction are affected by separation conditions and the nature of the sample. Of particular interest, steric interaction exhibits a maximum effect for monomeric C(18) columns, and becomes less important for either a C(1) or C(30) column; shape selectivity appears unimportant for monomeric C(1)-C(18) columns at ambient and higher temperatures, but becomes pronounced for C(30) - as well as polymeric columns with ligands ≥C(8). One hypothesis is that shape selectivity involves the presence or creation of cavities within the stationary phase that can accommodate a retained solute (a primarily enthalpic process), while steric interaction mainly makes greater use of spaces that pre-exist the retention of the solute (a primarily entropic process). The related dependence of hydrophobic interaction on column properties was also examined.