Rapid determination of pK (a) values of 20 amino acids by CZE with UV and capacitively coupled contactless conductivity detections.
A rapid and universal capillary zone electrophoresis (CZE) method was developed to determine the dissociation constants (pK (a)) of the 20 standard proteogenic amino acids. Since some amino acids are poorly detected by UV, capacitively coupled contactless conductivity detection (C(4)D) was used as an additional detection mode. The C(4)D coupling proved to be very successful on a conventional CE-UV instrument, neither inducing supplementary analyses nor instrument modification. In order to reduce the analysis time for pK (a) determination, two strategies were applied: (i) a short-end injection to reduce the effective length, and (ii) a dynamic coating procedure to generate a large electroosmotic flow (EOF), even at pH values as low as 1.5. As a result, the analysis time per amino acid was less than 2 h, using 22 optimized buffers covering a pH range from 1.5 to 12.0 at a constant ionic strength of 50 mM. pK (a) values were calculated using an appropriate mathematical model describing the relationship between effective mobility and pH. The obtained pK (a) values were in accordance with the literature.
Henchoz
Yveline
Y
Laboratoire de Chimie Analytique Pharmaceutique, Section des sciences pharmaceutiques, Université de Genève, Université de Lausanne, Bd d'Yvoy 20, 1211 Geneva 4, Switzerland.
Schappler
Julie
J
Geiser
Laurent
L
Prat
Josiane
J
Carrupt
Pierre-Alain
PA
Veuthey
Jean-Luc
JL
D000596:Amino Acids / Q000032:analysis*; D004553:Electric Conductivity; D053841:Electroosmosis; D019075:Electrophoresis, Capillary / Q000295:instrumentation / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D008962:Models, Theoretical; D009994:Osmolar Concentration; D015203:Reproducibility of Results; D013056:Spectrophotometry, Ultraviolet / Q000379:methods; D013696:Temperature
D000596:Amino Acids