Stability of vanadium(V)-protein complexes during chromatography

In order to reduce the risk of having artifacts during the separation of different vanadium containing proteins with chromatographic methods, we carried out some stability tests for selecting the most appropriate eluting conditions without breaking the vanadium(V)-protein binding. Therefore we investigated the stability of the vanadium-protein (transferrin and albumin) binding as function of the pH, salt molarity (NaCl, Na-acetate, NaBr, NaI, LiCl, NH+Cl and CaCl2) and hydrophobicity (acetonitrile). This was performed with a 48-vanadium tracer by means of batch experiments using ultrafiltration techniques to achieve a separation between protein bound and 'free' vanadium. We found that there was a significant pH-dependence. Depending on the eluting salt used, the vanadium(V)-protein binding is also disrupted by a high salt concentration (> 0.3 mol/L). An acetonitrile concentration around 2, mol/L has the same disrupting effect.