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In this study, a simple analytical method for the determination of γ-aminobutyric acid, gabapentin, and baclofen by using high-performance liquid chromatography with fluorescence detection was developed. An amidogen-reactive fluorescence labeling reagent, 4-(carbazole-9-yl)-benzyl chloroformate was first used to sensitively label these analytes. The completed labeling of these analytes can be finished rapidly only within 5 min at the room temperature (25°C) to form 4-(carbazole-9-yl)-benzyl chloroformate labeled fluorescence derivatives. These labeled derivatives expressed strong fluorescence property with the maximum excitation and emission wavelengths of 280 and 380 nm, respectively. The labeled derivatives were analyzed using a reversed-phase Eclipse SB-C18 column within 10 min with satisfactory shapes. Excellent linearity (R² > 0.995) for all analytes was achieved with the limits of detection and the limits of quantitation in the range of 0.25−0.35 and 0.70−1.10 μg/L, respectively. The proposed method was used for the simultaneous determination of γ-aminobutyric acid and its analogs in human serum with satisfactory recoveries in the range of 94.5-97.5%.