Resolution in Countercurrent Chromatography

Abstract

Using as an example the separation of DNP-glutamic acid and DNP-alanine in a nonplanetary countercurrent chromatograph, it was demonstrated that resolution, R_s, of a solute pair can be satisfactorily predicted from knowledge of the column efficiency, N, the fraction of column volume occupied by stationary phase, S_F, and the partition coefficients of the substances in the solvent system employed. Resolution is a function of the phase volume ratio and separation of rapidly eluted compounds is favored by an increase in S_F. The partition coefficients, K, the separation factor, α, and the column efficiency, N, are independent of S_F. Increasing S_F brings about an increase in R_S by increasing the capacity factor k₁. For S_F of 0.4, characteristic of the horizontal flow-through CCC, baseline separation (R_S = 1.5) is obtained for values of K₁, the first eluted substance, of about 0.5 for N = 1000 or K₁ of 10 for N = 100, where α is 2. Increasing S_F to 0.8, characteristic of the multilayer coil, high speed CCC, favors resolution of rapidly eluted solutes, those with low K values. Baseline resolution is then obtained for K₁ of about 0.07 with N = 1000 or for K₁ of about 1.5 for N = 100.