The invention provides a pharmaceutical composition containing ginkgolide B. The pharmaceutical composition comprises the ginkgolide B and thrombin inhibitor. The invention further provides a preparation method and application of the pharmaceutical composition. The pharmaceutical composition has the advantages that the ginkgolide B and the thrombin inhibitor are combined to achieve a synergic effect, and the pharmaceutical composition is excellent in anticoagulation effect, low in side effect and promising in clinical application prospect; the pharmaceutical composition is novel in prescription, simple in component, clear in action mechanism, evident in curative effect, less prone to tolerance generating and capable of achieving large-scale industrial production.