The invention belongs to the technical field of synthetic drugs, and particularly relates to a preparation method of hydroxycamptothecine. An acid solvent dissolves camptothecin, an oxidizing agent sodium perborate is added, the reaction liquid is subjected to vacuum concentration after a constant temperature reaction is completed, ice water is added till a crystal is completely separated out, filtration, washing and drying are conducted, and 1-deoxycamptothecine is obtained; dioxane, acetonitrile, water and concentrated sulfuric acid are added into 1-deoxycamptothecine to be dissolved and stirred, the mixture is illuminated with an ultraviolet high pressure mercury lamp, and the ice water is condensed and flows back; vacuum concentration is conducted, the ice water is added till the crystal is completely separated out, filtration, washing and drying are conducted, and a crude product is obtained; and the crude product is purified. The preparation method of hydroxycamptothecine is simple to operate, safe and economic, green and environmentally friendly, and high in total yield.