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A theory for the chromatography of rigid macromolecules on hydroxyapatite columns has been developed under the assumptions of instantaneous thermodynamic equilibrium between adsorbed phase and solution and of negligible longitudinal diffusion of macro-molecules. In the first or static part of this theory the adsorption isotherm of macro-molecules was studied as a function of phosphate concentration in the solvent.

The chromatography of native acid-soluble tropocollagen from calf skin on hydroxyapatite columns has been investigated. Resolution of a number of chromatographic peaks has been obtained by using shallow slopes of the eluting phosphate gradient. The results obtained suggest a heterogeneity of tropocollagen molecules which might be due to different distributions of absorbing sites on the molecules. The results obtained have also been used to study the mechanism of chromatography of proteins on hydroxyapatite and the resolving power of the columns.