Objective: This study aimed to establish a hollow fiber ligand fishing method for screening γ- Aminobutyrate transaminase (GABA-T) inhibitory component from the volatile oil of Acorus tatarinowii. Methods: Based on optimizing the enzyme assisted extraction process of volatile oil from A. tatarinowii using response surface methodology, GABA-T solution was injected into the hollow fiber lumen and used as a "bait" to fish out the potential active components in A. tatarinowii volatile oil, and its GABA-T inhibitory activity was verified. Results: The results showed that the optimum extraction conditions of volatile oil from A. tatarinowii were as follows: cellulase content 3 165 U/g, enzymolysis time 1.8 h, enzymolysis temperature 45 ℃, and the yield of volatile oil from A. tatarinowii was 2.56%. Finally, 51 compounds were identified, mainly including olefins, alcohols, ethers, ketones, phenols, acids, and other compounds. 3 mg/mL of GABA-T solution was injected into the cavity of the U-shaped hollow fiber tube, incubated for 160 min, sealed, and placed in the volatile oil of A. tatarinowii for ultrasonic (60 kHz) fishing for 20 min, and then analyzed with LCMS-IT-TOF system. Successfully filtered out α-Asarone has a concentration-dependent inhibitory effect on GABA-T activity, with an IC₅₀ of 57.9 μg/mL. Conclusion: The established hollow fiber ligand fishing method is stable, feasible, and easy to operate.