Hydrophobic interaction chromatography of aliphatic alcohols and carboxylic acids on octyl-sepharose CL-4B: mechanism and thermodynamics

Gelsema, W.J.; Brandts, P.M.; Ligny, C.L. De; Theeuwes, A.; Roozen, A.M.P.

doi:10.1016/s0021-9673(01)87594-4

Cited by 23 publications

(7 citation statements)

References 25 publications

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“…The observed increase in protein retention with temperature, for instance, has been ascribed to enhancement of hydrophobic interactions with increasing temperature due to temperature-induced conformational changes of proteins and concomitant increase in hydrophobic contact area upon binding to the chromatographic surface (16). So far the only thermodynamic study on the effect of temperature in HIC was carried out with aliphatic alcohols and carboxylic acids on octyl-agarose stationary phase with neat aqueous phosphate buffers (19) and hydro-organic mobile phases containing methanol or ethylene glycol (20). Since the salt concentration was very low in these studies, the conditions differed from those employed for protein separation in HIC.…”

Section: Introductionmentioning

confidence: 99%

Temperature effects in hydrophobic interaction chromatography.

Haidacher

Vailaya

Horváth

1996

Proc. Natl. Acad. Sci. U.S.A.

147

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The effect of temperature from 5°C to 50°C on the retention of dansyl derivatives of amino acids in hydrophobic interaction chromatography (HIC) was investigated by HPLC on three stationary phases. Plots of the logarithmic retention factor against the reciprocal temperature in a wide range were nonlinear, indicative of a large negative heat capacity change associated with retention. By using Kirchoff's relations, the enthalpy, entropy and heat capacity changes were evaluated from the logarithmic retention factor at various temperatures by fitting the data to a logarithmic equation and a quadratic equation that are based on the invariance and on an inverse square dependence of the heat capacity on temperature, respectively. In the experimental temperature interval, the heat capacity change was found to increase with temperature and could be approximated by the arithmetic average. For HIC retention of a set of dansylamino acids, both enthalpy and entropy changes were positive at low temperatures but negative at high temperatures as described in the literature for other processes based on the hydrophobic effect. The approach presented here shows that chromatographic measurements can be not only a useful adjunct to calorimetry but also an alternative means for the evaluation of thermodynamic parameters.Hydrophobic interaction chromatography (HIC) is widely used for the separation and purification of proteins in their native state (1, 2). The technique employs weakly hydrophobic stationary phases and the retention is modulated by varying the salt concentration in the aqueous mobile phase (3, 4). The effect of salt on protein interactions in aqueous solutions (5-9) as well as on protein adsorption in HIC (5, 10, 11) has been extensively investigated, and the salt effect on HIC retention has been treated within the hermeneutics of the solvophobic theory (12-14) and Wyman's linked functions (15)(16)(17)(18).Less progress has been made in elucidating the effect of temperature on retention in HIC. The observed increase in protein retention with temperature, for instance, has been ascribed to enhancement of hydrophobic interactions with increasing temperature due to temperature-induced conformational changes of proteins and concomitant increase in hydrophobic contact area upon binding to the chromatographic surface (16). So far the only thermodynamic study on the effect of temperature in HIC was carried out with aliphatic alcohols and carboxylic acids on octyl-agarose stationary phase with neat aqueous phosphate buffers (19) and hydro-organic mobile phases containing methanol or ethylene glycol (20). Since the salt concentration was very low in these studies, the conditions differed from those employed for protein separation in HIC. In order to shed light on the HIC retention behavior of proteins, it is necessary to understand first the physicochemical basis of HIC with simple molecules that undergo no significant conformational changes and to useThe publication costs of this article were defrayed in part by page c...

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Section: Introductionmentioning

confidence: 99%

Temperature effects in hydrophobic interaction chromatography.

Haidacher

Vailaya

Horváth

1996

Proc. Natl. Acad. Sci. U.S.A.

147

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“…Although many studies were carried out on the thermodynamic behavior of small solutes in reversed-phase liquid chromatography (RPLC) [1][2][3][4] and hydrophobic interaction chromatography (HIC) with linear, or non-linear plot by van't Hoff plot, [5][6][7] none have been found for biopolymers. Due to many different characteristics between biopolymers and small solutes, both show varied chromatographic behaviors and different mechanisms in RPLC.…”

Section: Introductionmentioning

confidence: 99%

Studies on the Thermodynamics During the Biopolymer Retention Process in Reversed‐Phase Liquid Chromatography. I. The Free Energy Change and Its Fractions, Adsorption, and Desorption Free Energy Changes

Bai

Geng

2003

Journal of Liquid Chromatography & Related Technologies

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Based on the stoichiometric displacement theory for retention of solute (SDT-R), the changes of free energy of biopolymers during retention process in reversed-phase high performance liquid chromatography (RPLC) can be divided into two independent fractions, the adsorption and desorption free energy changes. The totally net free energy change and the two fractions of seven kinds of proteins in RPLC were accurately measured. thermodynamics, with the comparisons of the adsorption and desorption free energy changes between small solutes and biopolymers in RPLC, the following experimental facts were first explained quantitatively: [Robbat, A., Jr.; Liu, T.Y. J. Chromatagr. 1990, 513, 117] The elution range of organic solvent for small solutes in the mobile phase in RPLC is usually broad, while that for biopolymers is very narrow. [Tan, L.C.; Carr, P.W. J. Chromatogr. 1993, 656, 521] Small solutes in RPLC can be eluted with isocratic elution, but biopolymers only with gradient elution. [Tchapla, A.;Heron, S.;Colin, H.; Guiochon, G. Anal. Chem. 1988, 60, 1443 The resolution of small solutes in RPLC depends on column length, while that of biopolymers is almost independent of it.

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“…In addition, if the small solutes are homologues, establishing whether the parameters obtained obey the homologue rule would help to confirm the reliability of the result. So far temperature effect in HIC have been studied with aliphatic alcohols and carboxylic acids on an octylagarose stationary phase with aqueous phosphate buffers [7] and hydro-organic mobile phases containing methanol or ethylene glycol [8], but the conditions differed from those employed for the usual protein separation with very high salt concentration in mobile phase. Recently Horvfith and co-workers [9,10] investigated the effect of temperature on the retention of densyl amino acids with an (NH4)2SO 4 mobile phase, and Byun et al [11] reported the effects of salt and temperature on peptide retention on a SynChropak-HIC column.…”

Section: Introductionmentioning

confidence: 99%

Influences of the mobile phase composition and temperature on the retention behavior of aromatic alcohol homologues in hydrophobic interaction chromatography

et al. 2002

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SummaryTo eliminate the very complicated effects of chromatographic thermodynamics in hydrophobic interaction chromatography (HIC) with biopolymers as solutes, homologues of neutral aromatic alcohols were selected as solutes for investigating their thermodynamic behavior in HIC. The effects of the mobile phase composition and temperature (O ~ 80 ~ on the retention behavior of the homologues were studied extensively. The retention behavior of the homologue was characterized by the linear parameters in the stoichiometric displacement model for retention (SDM-R). The retention of small molecules is essentially controlled by non-specific interaction in HIC as well as in reversed phase liquid chromatography (RPLC), and the parameters obtained were found to follow the homologue rule. Plots of the logarithm of retention of solutes in four kinds of salt solution versus the reciprocal of the absolute temperature over a wide range were nonlinear, indicating a large heat capacity change associated with retention. The thermodynamic parameters demonstrate the retention of small molecules in HIC to be entropy-driven at low temperature and enthalpy-driven at high temperature.

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Hydrophobic interaction chromatography of aliphatic alcohols and carboxylic acids on octyl-sepharose CL-4B: mechanism and thermodynamics

Cited by 23 publications

References 25 publications

Temperature effects in hydrophobic interaction chromatography.

Temperature effects in hydrophobic interaction chromatography.

Studies on the Thermodynamics During the Biopolymer Retention Process in Reversed‐Phase Liquid Chromatography. I. The Free Energy Change and Its Fractions, Adsorption, and Desorption Free Energy Changes

Influences of the mobile phase composition and temperature on the retention behavior of aromatic alcohol homologues in hydrophobic interaction chromatography

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