Isolation and identification of impurities in chlorin e6

The tetrapyrrolic compound chlorin e6 is currently used as a pharmaceutical substance for Photolon formulation, which is utilized in photodynamic therapy of various diseases. It was found that chlorin e6 could contain both process- and degradation-related impurities. In order to understand their origin, the most abundant impurities were prepared by liquid extraction, preparative chromatography or chemical synthesis. By means of HPLC-PDA-MS, 1D and 2D NMR spectroscopy, these impurities were identified as chlorin e6 17(4)-ethyl ester, chlorin e4, 15-hydroxyphyllochlorin, rhodochlorin, 15(1)-hydroxymethylrhodochlorin delta-lactone, rhodochlorin-15-oxymethyl delta-lactone, rhodochlorin-15-oxymethyl delta-lactone 17(4)-ethyl ester, 15(1)-hydroxymethylrhodoporphyrin delta-lactone, rhodoporphyrin-15-oxymethyl delta-lactone and purpurin 18. The possible routes of formation of the chlorin derivatives upon the production and storage of chlorin e6 are discussed.