Analysis of Phenolic Compounds in Nicotiana tabacum Based on Aqueous Two-Phase Extraction Coupled with Liquid Chromatography

Background: Phenolic compounds in Nicotiana tabacum L. have anti-tumor, antibacterial, anti-inflammatory, and other pharmacological effects. Therefore, either to optimize the utilization of plant resources or to reduce the physiological harm caused by cigarette, it is necessary to separate and analyze the phenolic compounds in Nicotiana tabacum. Aim: To develop a protocol for extraction and analysis of phenolic compounds in Nicotiana tabacum. The method based on aqueous two-phase extraction (ATP) should provide a solid experimental basis for future large-scale medical industrial extraction. Results: Five of phenolic compounds in Nicotiana tabacum including catechol, caffeic acid, coumarin, rutin, and quercetin were extracted with the developed ATP method and analyzed by the optimized liquid chromatography method simultaneously. In the concentration range of 10-200 µg/mL, the linear correlation coefficients of calibration curve for five compounds were all greater than 0.999. The developed ATP extraction method employed 70% acetonitrile water solution as the extractant. The ammonium sulphate was selected as the preferred phase separation salt. The extraction efficiencies of five phenolic compounds were all more than 90%. In concrete sample, the concentrations of five compounds were determined based on acetonitrile/salt ATPE system, Conclusion: Analysis of phenolic compounds in Nicotiana tabacum was achieved based on acetonitrile/ammonium sulphate ATP extraction coupled with liquid chromatography. The developed ATP extraction protocol employed 70% acetonitrile water solution as the extractant. The ammonium sulphate was selected as the preferred phase separation salt. The accuracy and precision were satisfied. The protocol developed in this study is expected to be applied in medical industry.