The invention provides a preparation method of a 7-keto-6[alpha]-alkyl cholanic acid derivative. According to the preparation method provided by the invention, a 7-keto-6[beta]-alkyl cholanic acid derivative, as shown by a formula II, is used as a raw material, and the 7-keto-6[alpha]-alkyl cholanic acid derivative is prepared by converting a 6[beta] configuration into a 6[alpha] configuration under an acid or alkali condition. The invention also provides a 7-keto-6[beta]-alkyl cholanic acid derivative and an application thereof in preparation of 3[alpha],7[alpha]-dihydroxy-6[alpha]-alkyl-5[beta]-cholanic acid. The preparation method provided by the invention is simple and convenient, and is high in configuration conversion rate, and the product, the 7-keto-6[alpha]-alkyl cholanic acid derivative, is easy to purify, so that the purification difficulty for preparing the 3[alpha],7[alpha]-dihydroxy-6[alpha]-alkyl-5[beta]-cholanic acid is reduced.
