Studies of proteins adsorption in polyethyleneimine coated capillaries in capillary zone electrophoresis

SPANILÁ, Miroslava, Jiří PAZOUREK a Josef HAVEL. Studies of proteins adsorption in polyethyleneimine coated capillaries in capillary zone electrophoresis. In 11th International Symposium on Separation Sciences ISSS 2005. Book of Abstracts. Pardubice: Univerzita Pardubice, 2005, s. 231. ISBN 80-7194-771-7.

Základní údaje

• Originální název: Studies of proteins adsorption in polyethyleneimine coated capillaries in capillary zone electrophoresis
• Název česky: Studium adsorpce proteinů v polyethyleniminem koutovaných kapilárách v kapilární zónové elektroforéze
• Autoři: SPANILÁ, Miroslava (203 Česká republika, garant), Jiří PAZOUREK (203 Česká republika) a Josef HAVEL (203 Česká republika).
• Vydání: Pardubice, 11th International Symposium on Separation Sciences ISSS 2005. Book of Abstracts. s. 231-231, 2005.
• Nakladatel: Univerzita Pardubice

Další údaje

• Originální jazyk: angličtina
• Typ výsledku: Stať ve sborníku
• Obor: 10406 Analytical chemistry
• Stát vydavatele: Česká republika
• Utajení: není předmětem státního či obchodního tajemství
• Kód RIV: RIV/00216224:14310/05:00025070
• Organizační jednotka: Přírodovědecká fakulta
• ISBN: 80-7194-771-7
• Klíčová slova anglicky: capillary coating; polyethyleneimine; electroosmotic flow; protein adsorption
• Štítky: capillary coating, electroosmotic flow, polyethyleneimine, protein adsorption

Anotace

The magnitude and the direction of electroosmotic flow (EOF) has fundamental influence to separation in capillary zone electrophoresis. Adsorption of proteins on the negative charged silica surface of uncoated capillaries can lead to analyte loss and to irreproducibility of analyses. Therefore, in biomolecules analyses, the inner surface modification of fused-silica capillaries is recommended. One of the used coating is polyethyleneimine (PEI): as a polycationic polymer it creates positively charged covalent coating on the capillary inner wall and hence minimizes the adsorption of basic proteins at acidic pH and maintains the electroosmotic flow stable [1]. In this work, the properties of PEI coated capillaries with respect to different background electrolytes are described. The changes of inner surface of the capillary were followed by electroosmotic flow measurement [2]. Several buffers commonly used in electrophoretic separations of proteins (MES, acetate, borate) were tested, but the main interest was taken into carboxylic acids as a buffers (citrate, malate, malonate, tartrate and succinate). Interesting behaviour of EOF in PEI coated capillaries was observed: the direction (and the value) depend on pKa of the background electrolyte and at certain pH it can reversibly change from positive to negative. With this knowledge the influence of ionic strength and pH of background electrolyte on the separation of basic proteins was studied. As a model, the standard protein mixture of cytochrome C, ribonuclease A and lysozyme in concentration of 0.25 mg/ml was chosen. Finally, the experimental conditions for humanin analyses were optimized.

Anotace česky

The magnitude and the direction of electroosmotic flow (EOF) has fundamental influence to separation in capillary zone electrophoresis. Adsorption of proteins on the negative charged silica surface of uncoated capillaries can lead to analyte loss and to irreproducibility of analyses. Therefore, in biomolecules analyses, the inner surface modification of fused-silica capillaries is recommended. One of the used coating is polyethyleneimine (PEI): as a polycationic polymer it creates positively charged covalent coating on the capillary inner wall and hence minimizes the adsorption of basic proteins at acidic pH and maintains the electroosmotic flow stable [1]. In this work, the properties of PEI coated capillaries with respect to different background electrolytes are described. The changes of inner surface of the capillary were followed by electroosmotic flow measurement [2]. Several buffers commonly used in electrophoretic separations of proteins (MES, acetate, borate) were tested, but the main interest was taken into carboxylic acids as a buffers (citrate, malate, malonate, tartrate and succinate). Interesting behaviour of EOF in PEI coated capillaries was observed: the direction (and the value) depend on pKa of the background electrolyte and at certain pH it can reversibly change from positive to negative. With this knowledge the influence of ionic strength and pH of background electrolyte on the separation of basic proteins was studied. As a model, the standard protein mixture of cytochrome C, ribonuclease A and lysozyme in concentration of 0.25 mg/ml was chosen. Finally, the experimental conditions for humanin analyses were optimized.

Návaznosti

• GA305/03/1100, projekt VaV: Název: Syntéza a studium neuroprotektivních peptidů odvozených od humaninu