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Development and validation of transient isotachophoretic capillary zone electrophoresis for determination of peptides
Author(s) -
Waterval Joop C. M.,
la Porte Charles J. L.,
van 't Hof Ron,
Teeuwsen Jan,
Bult Auke,
Lingeman Henk,
Underberg Willy J. M.
Publication year - 1998
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150191820
Subject(s) - capillary electrophoresis , chemistry , chromatography , isotachophoresis , capillary action , detection limit , reproducibility , linearity , analytical chemistry (journal) , cationic polymerization , electrolyte , electrophoresis , materials science , electrode , physics , organic chemistry , quantum mechanics , composite material
Although capillary zone electrophoresis (CZE) is known for its high resolution power and low mass detection limits, the concentration detection limits are rather poor when ultraviolet absorbance detection is used. To overcome this limitation, several on‐column transient isotachophoresis (tlTP) protocols have been developed and validated for the determination of both cationic and anionic model peptides, separately. Using this preconcentration method, up to 72% of the capillary can be filled with sample solution, without any loss in resolution. Thus, without any modification of the hardware set‐up, the sensitivity is increased about two orders of magnitude. For the model cationic peptides (gonadorelin, angiotensin II) good linearity and reproducibility is observed in the 20 to 100 ng/mL concentration range. For the anionic peptides ( N‐t ‐Boc‐Pentagastrin and two related peptides), a ITP method was developed using a dynamically coated capillary. The coating was prepared by adding Fluorad FC‐135® to the leading electrolyte buffer. In this way a positively charged bilayer was formed on the inside of the capillary, producing an electroosmotic flow towards the outlet using reversed polarity conditions. In this way, acceptable analysis times were achieved. Using the developed tlTP method, up to 72% of the capillary can be filled with sample solution as well. The anionic peptides are separated even better than when using CZE conditions. Linearity and reproducibility in the 20‐100 ng/mL range proved to be excellent.