Experimental studies of pressure/temperature dependence of protein adsorption equilibrium in reversed-phase high-performance liquid chromatography

Szabelski, Paweł; Cavazzini, Alberto; Kaczmarski, Krzysztof; Liu, X; Horn, Jennifer Van; Guiochon, Georges

doi:10.1016/s0021-9673(01)01614-4

Cited by 77 publications

(52 citation statements)

References 47 publications

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“…Since the changes in retention with pressure represent a volume change in the pressure dependent equilibrium (see below), larger changes can be observed with analytes of a higher molecular weight. The changes with pressure observed in reference [10] were independent of temperature. For the RP retention mechanism of this study, increases in retention were observed.…”

Section: Introductionmentioning

confidence: 71%

“…With a C18 stationary phase and a b-cyclodextrin mobile phase additive, both positive and negative changes in retention with pressure were observed. The RP studies by Guiochon and coworkers [9,10] involved insulin variants, larger molecules than used in the other studies. Since the changes in retention with pressure represent a volume change in the pressure dependent equilibrium (see below), larger changes can be observed with analytes of a higher molecular weight.…”

Section: Introductionmentioning

confidence: 99%

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Influence of pressure on the retention of sugars in hydrophilic interaction chromatography

Neue

Hudalla

Iraneta

2010

J of Separation Science

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Section: Introductionmentioning

confidence: 71%

Section: Introductionmentioning

confidence: 99%

Influence of pressure on the retention of sugars in hydrophilic interaction chromatography

Neue

Hudalla

Iraneta

2010

J of Separation Science

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“…A number of studies about dependence of RPLC retention times on pressure at constant temperature have been performed previously [54,55]. Szabelski et al [53] found that the retention of insulin greatly increases when the average pressure increases from 47 to 147 bar.…”

Section: Resultsmentioning

confidence: 99%

Reverse-High Performance Liquid Chromatography Mechanism Explained by Polarization of Stationary Phase

Andrade-Eiroa¹

2011

CheM

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Original results and conclusions on Reverse Phase High Performance Liquid Chromatography (RPLC) mechanism are here presented. So far, none of the theoretical approaches applied to the RPLC mechanism can explain the retention and elution mechanisms of most of the analytes by RPLC, especially neutral organic compounds. Our experiences allowed us to state that RPLC retention mechanism most likely occurs through polarization of stationary phase (usually dielectric surfaces) submerged into solvents with huge dielectric constant and high dipole moment (i.e. water and/or acetonitrile) at high pressures as those applied in RPLC systems. The surface polarized interacts with polarizable target analytes (i.e. naphthalene, pyrene or benzo(ghi)perylene) in such a way that the retention depends on the medium polarizability of the target compound. The higher is the medium polarizability of the compound retained the higher is its retention time.By using Polycyclic Aromatic Hydro-carbons (PAHs) and 6-nitrochrysene as probes, we found that although the dependence of retention times on pressure is not evident in most cases, the interaction between pressure and the mobile phase dipole moment is evidenced at the light of the constructed mathematical models. Two very different cases can be distinguished:1. On one hand, when low dipole moment mobile phases are used, high pressure means big work developed by the system in such a way that high pressures inside the system reduce retention times compared to low pressures.2. On the another one, when high dipole moment solvents are used as mobile phase, high pressure entails an outstanding increase of the retention times compared to low dipole moment mobile phases. This is most likely due to high pressures that entail closer alignment of mobile phase dipoles and lesser tilt of them, originating an outstanding electric field inside the analytical column.In addition to this, reduced retention times for dipole solutes (6-nitrochrysene, dipole molecule) compared to non-polar solutes (PAHs) were observed when pressure drops. This conclusions would allow to extend the model to basic/acid molecules at different pH. In this case, electrostatic repulsions among analyte molecules should be considered.

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“…A variação do fator de retenção de um soluto em função da temperatura é descrita pela equação de Van Szabelski et al 13 estudaram a influência da temperatura sobre uma série de proteínas empregando uma coluna Symmetry C8 (150 x 3.9 mm, dp 5 μm) e observaram que o fator de retenção destas proteínas variou em até 10 vezes na faixa de temperatura de …”

Section: -12unclassified

Possibilidades e limitações no uso da temperatura em cromatografia líquida de fase reversa

Borges¹,

Bottoli²

2010

Quím. Nova

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Recebido em 16/9/09; aceito em 30/11/09; publicado na web em 8/4/10 POSSIBILITIES AND LIMITATIONS OF USING TEMPERATURE IN REVERSED PHASE LIQUID CHROMATOGRAPHY. High-temperature liquid chromatography (HTLC) is a technique that presents a series of advantages in liquid phase separations, such as: reduced analysis time, reduced pressure drop, reduced asymmetry factors, modified retentions, controlled selectivities, better efficiencies and improved detectivities, as well as permitting green chromatography. The practical limitations that relate to instrumentation and to stationary phase instability are being resolved and this technique is now ready to be applied for routine determinations.Keywords: high temperature liquid chromatography; stationary phase stability; green chromatography. INTRODUÇÃOO termo alta temperatura em cromatografia líquida (High Temperature Liquid Chromatography, HTLC) se refere ao uso de temperaturas entre 40 a 200 o C. O uso da temperatura como variável em cromatografia líquida é recente, embora Antia e Horvath 1 tenham descrito seu uso em 1988. A principal vantagem desta técnica é a redução no tempo de análise, característica que é de grande interesse prático para os usuários de cromatografia. Mas até o momento, esta técnica reside quase exclusivamente no meio acadêmico por causa de algumas limitações, como a falta de instrumentação específica para tal fim e a instabilidade das fases estacionárias. Desta forma, esta revisão se dedica à discussão das vantagens e limitações desta técnica, bem como os avanços recentes quanto à instrumentação e fases estacionárias, os quais devem impulsionar a migração desta técnica para o meio industrial. ASPECTOS TEÓRICOS Influência da temperatura sobre a pressãoA pressão gerada pela passagem de fase móvel (FM) por uma coluna pode ser descrita pela equação de Kozeny-Carman:2,3 ΔP = (ηFL)/(K 0 πr 2 dp 2 ), onde K 0 é a permeabilidade específica, η é a viscosidade da FM, F é a vazão da FM, r é o raio interno da coluna e dp é o diâmetro médio das partículas da fase estacionária (FE) que recheiam a coluna. A viscosidade decresce exponencialmente em função da temperatura, 2,3 assim, o aumento da temperatura permite: a utilização de maiores vazões; o emprego de partículas com menores diâmetros; a utilização de colunas com maior comprimento ou conectadas em série.Um exemplo de como o aumento da temperatura permite o uso de maiores vazões é o trabalho de Yang et al.2 onde uma coluna ZirChrom-PS (50 x 4,6 mm, dp 2,5 μm) utilizada a 25 o C e com FM acetonitrila:água, 25:75 v/v, a uma vazão de 2 mL/min gera uma pressão de 400 bar, alcançando o limite máximo de pressão de um sistema convencional de cromatografia líquida de alta eficiência (High Performance Liquid Chromatography, HPLC). Quando a temperatura é elevada a 150 o C, a pressão é reduzida a 60 bar, permitindo o uso de uma vazão de 14 mL/min sem exceder o limite de pressão da bomba.As micropartículas (dp < 2 μm) têm pequena resistência à transferência de massa a altas vazões. Por isso podem ser utilizadas com pouc...

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Experimental studies of pressure/temperature dependence of protein adsorption equilibrium in reversed-phase high-performance liquid chromatography

Cited by 77 publications

References 47 publications

Influence of pressure on the retention of sugars in hydrophilic interaction chromatography

Influence of pressure on the retention of sugars in hydrophilic interaction chromatography

Reverse-High Performance Liquid Chromatography Mechanism Explained by Polarization of Stationary Phase

Possibilidades e limitações no uso da temperatura em cromatografia líquida de fase reversa

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