The invention belongs to the field of traditional Chinese medicine constitution healthcare, and particularly relates to a medicine composition for treatment of yin deficiency. The medicine composition for treatment of yin deficiency is mainly prepared from, by weight, 3-25 parts of the root of straight ladybell, 3-20 parts of radix polygonati officinalis, 3-22 parts of fructus mume, 4-35 parts of the root of rehmannia, 3-15 parts of lily and 2-25 parts of asparagus cochinchinensis. Meanwhile, the invention provides a preparation method and application of the medicine composition. The medicine composition for treatment of yin deficiency and the preparation method and application thereof have the advantages that the medicine composition for treatment of yin deficiency is reasonable in formula, all component raw materials are matched with one another, and a better treatment effect can be achieved when yin deficiency is treated. Meanwhile, the medicine composition obtained through the preparation method has the advantages of being easy and convenient to manufacture, high and even in effective constituent content, low in manufacturing cost and the like.