Abstract
We investigated the solubilization and recovery of heat-aggregated cytochrome c using alkylammonium nitrates (RAN-ILs), which are a group of protic ionic liquids (ILs), including methylammonium nitrate (MAN), ethylammonium nitrate (EAN), and propylammonium nitrate (PAN), in a wide range of concentrations (X = 0–40 mol%IL for MAN and 0–100 mol%IL for EAN and PAN) using absorption and circular dichroism spectroscopy to obtain basic information on solubilization/recovery solvents for heat-aggregated proteins using ILs. On the whole, the increase of absorbance at the soret band (~405 nm) of cytochrome c indicated that solubilization ability was observed in aqueous solutions with MAN and EAN (X > 10) and PAN (X > 5). These results indicate that the three RAN-ILs studied have solubilization ability for heat-aggregated cytochrome c, and increasing the alkyl-chain length of the cation causes an enhancement of the solubilization ability for heat-aggregated cytochrome c. The activity, secondary structure and heme coordination state (Fe–His18/Met80) of the solubilized cytochrome c in aqueous PAN solutions, which have the highest solubilization ability among the three RAN-ILs, were recovered up to X = 30 after removing PAN by dialysis. The present findings indicate that aqueous solutions with RAN-ILs have a high potential as recovery agents for heat-aggregated proteins.
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