Establishment of HPLC-ECD Fingerprinting of Prunella Vulgaris L. and Study on Its Antioxidant Spectrum-Effect Relationship

OBJECTIVE To establish the fingerprint of Prunella vulgaris L. by high performance liquid chromatography with electrochemical detection(HPLC-ECD), and to evaluate the antioxidant activity in vitro and study the spectrum-effect relationship. METHODS The samples were extracted with 80% methanol. The XBridge BEH shield RP₁₈ column(3.0 mm× 150 mm, 2.5 μm) was selected as the stationary phase, acetonitrile-citrate solution(50 mmol·L^-1, pH 2.8) was selected as the mobile phase with gradient elution at the flow rate of 0.6 mL·min^-1. The detection potential of ECD was 600 mV, the column temperature was 40℃. The antioxidant activity of Prunella vulgaris L. was evaluated by DPPH radical scavenging activity, ABTS radical scavenging activity and ferric reducing antioxidant power. The content of total phenolics was measured by the Folin-Ciocalteau method. The quality of 21 batches of Prunella vulgaris L. were analyzed by similarity evaluation. Furthermore, the spectrum-effect relationship of antioxidation was studied by gray relational analysis and partial least square regression analysis. RESULTS A total of 19 common peaks were obtained by HPLC-ECD fingerprinting, among which 14 components were identified. The similarity of the 21 batches of Prunella vulgaris L. were >0.9. At the same time, Prunella vulgaris L. had superior ability of ABTS, DPPH radical scavenging activity and ferric reducing antioxidant power values. HPLC-ECD fingerprint could specifically reflect the information of antioxidant components in the sample, and there was a significant linear correlation among the total peak area of chromatogram, total phenolics content and antioxidant activity. The results of gray relational analysis showed that there was a high correlation between 19 common peaks and antioxidant activity with the relational degree >0.6. The results of partial least square regression analysis showed that the compounds have the greatest contribution to antioxidative activity were rosmarinic acid, caffeic acid and rutin, etc. CONCLUSION The HPLC-ECD fingerprint combined with antioxidant method can be used to evaluate the quality of Prunella vulgaris L. and investigate the material base of antioxidant activity.