The composition of stains produced by the oxidation of Methylene Blue

The composition of some stains produced by the oxidation of Methylene Blue has been studied by thin-layer chromatography. Various named methods for the production of Polychrome Methylene Blue, Azure A, Azure B, Azure C and Methylene Violet Bernthsen have been found to give complex mixtures of varying proportions of up to eleven dyes. Ten of these, namely Methylene Blue, Azure B, Azure A, sym-Dimethylthionine, Azure C, Thionine, Methylene Violet Bernthsen, Methyl Thionoline, Thionoline and Thionol, have been identified by their visible absorption spectra. The remaining dye could not be identified. When used on a laboratory scale, these methods give stains of constant composition independent of the batch of Methylene Blue. Stain composition as revealed in the present study has been compared with that previously indicated by other, less effective, analytical techniques. Reasons are presented why the latter give equivocal results.