The ownership structure arrangement has an important impact on the audit costs of listed companies. This article uses the 2007 2018 Shanghai and Shenzhen A share listed companies as a sample, discusses the impact of multiple large shareholders on audit fees and its mechanism. The study found that listed companies with multiple large shareholders will be charged higher audit fees than companies with only a single large shareholder. The mechanism test shows that multiple large shareholders have influenced the companys audit fees through increasing audit demand and reducing the quality of internal control. Cross sectional analysis found that the positive impact of multiple large shareholders on audit fees is more significant in state owned enterprises and Big10 audit firms. The conclusion of the study not only enriches the literature on audit fees and multiple major shareholders, but also provides a useful reference for improving the governance of listed companies and the quality of accounting information.