A retrospective cohort study compared the effectiveness of clown therapy (CCT) and virtual reality distraction (VRD) on anxiety in 220 preschool children (3–6 years old) with venous blood sampling at Dongcheng County Hospital from January 2022 to January 2025. The children were divided into a CCT group (n=110) with an interactive clown game and a VRD group (n=110) with VR devices. The CCT group had significantly lower anxiety scores (t=4.128, p<0.001), smaller increases in heart rate (t=5.217, p<0.001), shorter crying duration (t=6.384, p<0.001), and lower cortisol changes compared to VRD. In CCT there was better cooperation (p<0.001), higher first attempt success rate (χ²=5.143, p=0.023), shorter collection time (t=4.926, p<0.001) and higher parental satisfaction (p<0.001). Multivariate analysis confirmed that CCT was independently associated with greater reduction in anxiety (p<0.001). The study concludes that CCT is more effective than VRD in reducing anxiety, improving physiological responses, clinical effectiveness, and parent satisfaction.