CHROMATOFOCUSING OF BIPOLAR COMPOUNDS WITHIN CATION-EXCHANGE COLUMNS USING THE SIMPLE MOBILE PHASE

A.V.Ivanov¹, M.S.Wakstein²

¹M.V.Lomonosov Moscow State University, Chemistry Department; 119992, Moscow, GSP-2, Lenin Hills; e-mail: sandro@analyt.chem.msu.ru
²Applied Acoustics Research Institute,
High Technologies Center, Dubna, Moscow region

The increasing pH-gradient formation within the columns, packed with carboxylic or sulfonic cation exchangers, using the simple eluents (containing 1 – 2 components) was studied. The optimal mobile phases for carboxylic columns consist of citric acid (2 – 5 mM) and Tris (1,5 – 20 mM), for sulfonic cation-exchange column – NaH₂PO₄ (3 – 10 mM) and Tris (up to 6 mM) as active components. The smooth and more linear pH gradients were formed within carboxylic columns at adjusting the ionic strength (0,05 – 0,5) in one or both mobile phases (i.e., in the start buffer and in the eluent). For sulfonic cation-exchange column it is preferably to adjust the ionic strength up to 0,1 – 0,3 in the mobile phases. The gradients obtained are comparable in a wide pH range with the gradients formed by synthetic polyampholytic "Polybuffer-96". The potentialities of the analytical application of proposed simple eluents in chromatofocusing were demonstrated by the separation of proteins model mixture, formate dehydrogenase isoforms and tripsin hydrolyzate of human serum albumin.

Key words: chromatofocusing, simple eluents, linear increasing pH-gradient, carboxylic and sulfonic cation-exchange columns

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