The present invention relates to a buffer composition for promoting production of calcitriol or calcifediol, and a method for producing calcitriol or calcifediol using the same. More particularly, the present invention relates to a buffer composition for promoting production of calcitriol or calcifediol comprising a metallic compound, an organic solvent, cyclodextrin, tris(hydroxymethyl)aminomethane, sodium succinate, sodium chloride, magnesium chloride, and water, and a method for producing calcitriol or calcifediol using the same. In the method for producing calcitiriol or calcifediol, the production yield of calcitriol or calcifediol is high, and the bioconversion is carried out in an enzyme reaction system instead of in a microorganism culture system. Thus, it is not required to maintain a sterile state. Also, the separation/purification following the completion of a biocatalytic reaction can be carried out in a cleaner state than the microorganism culture method. Accordingly, there is an advantage in that a cost required for separation is low and the quality is improved. Furthermore, the buffer composition for promoting production of calcitriol or calcifediol can provide a high productivity of calcitriol or calcifediol.