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The corrinoid cofactor of the tetrachloroethene reductive dehalogenase of Dehalospirillum multivorans was isolated in its Coβ‐cyano form. This cofactor represents the main corrinoid found in D. multivorans cells. Analysis of the isolated cyano‐corrinoid by a combination of HPLC and UV/VIS‐absorbance spectroscopy revealed it to be nonidentical to a variety of known natural B₁₂ derivatives. From high‐resolution mass‐spectrometric analysis, the molecular formula of the corrinoid isolated from D. multivorans could be deduced as C₅₈H₈₁CoN₁₇O₁₄P. The sample of the novel corrinoid from D. multivorans was further analyzed by UV/VIS, CD, and one‐ and two‐dimensional ¹H‐, ¹³C‐, and ¹⁵N‐NMR spectroscopy, which indicated its structure to be closely related to that of pseudovitamin B₁₂ (Coβ‐cyano‐7″‐adeninylcobamide). By the same means, the corrinoid could be shown to differ from pseudovitamin B₁₂ …