Preparation and testing of stationary phases and modified capillaries for affinity chromatography and affinity capillary electrophoresis of pepsin.
Three stationary phases have been prepared for affinity liquid chromatography isolation and separation of porcine and human pepsin. The phases contain 3,5-diiodo-L-tyrosine (DIT) bound to the supports HEMA BIO VS, HEMA BIO E and EPOXY TOYOPEARL. These phases have been tested on a model sample of porcine pepsin A and applied to human pepsin. Fractions have been collected and the chymase activity determined in selected analyses. For affinity CE, capillaries have been prepared by modifying the wall with 3-aminopropyltriethoxysilane, followed either by direct binding of DIT, or by binding L-tyrosine that was subsequently iodated. The dissociation constant K(d) has been determined for the pepsin-DIT complex from the changes in the electrophoretic mobilities.
Varilová
Tereza
T
Department of Analytical Chemistry, Charles University, Prague, Hlavova 2030/8, Praha 2, 12843 Czech Republic. tereza.varilova@seznam.cz
Vránková
Alice
A
Pacáková
Vera
V
Tichá
Marie
M
Stulík
Karel
K
D000818:Animals; D002846:Chromatography, Affinity / Q000295:instrumentation* / Q000379:methods; D004105:Diiodotyrosine / Q000737:chemistry; D019075:Electrophoresis, Capillary / Q000295:instrumentation* / Q000379:methods; D006801:Humans; D010434:Pepsin A / Q000302:isolation & purification*; D012822:Silicon Dioxide / Q000737:chemistry; D013552:Swine
D004105:Diiodotyrosine; D012822:Silicon Dioxide; D010434:Pepsin A