High-performance chromatofocusing using linear and concave pH gradients formed with simple buffer mixtures I. Effect of buffer composition on the gradient shape

Numerical calculations together with simplified analytical relations based on local equilibrium theory are used to determine the factors which govern the shape of the gradient formed during chromatofocusing when simple mixtur es of buffering species are employed to produce linear or concave pH gradie nts. The numerical and analytical development is also used to determine the relation between the gradient shape and the buffering capacities of the ad sorbed and liquid phases. Experiments which verify the theoretical methods are described! where internally generated, retained pH gradients of various shapes are formed using high-performance chromatography columns. The resul ting experimental and theoretical basis can be employed as means for the se lection of the buffer composition for use in chromatofocusing. (C) 2000 Els evier Science B.V. All rights reserved.