Abstract:

Twenty-five phenolic compounds in tobacco were analyzed using high performance liquid chromatography-ultraviolet/mass spectrometry (HPLC-UV/MS), including absolute quantification of 11 main phenols by HPLC-UV and relative quantification of other 14 phenols by HPLC-MS. The validation results of this method were satisfactory. Eleven phenol standards showed good linearity with correlation coefficients (r²) of 0.9993-0.9999 over the mass concentration range from 0.90 mg/L to 99.00 mg/L (the range of chlorogenic acid and rutin was from 0.95 mg/L to 380.00 mg/L). The recoveries of the 11 phenol standards were 91.0%-112.4% with the RSDs of 0.33%-8.11% at the three spiked levels of 22.5-24.8, 45.0-49.5 and 67.5-74.3 mg/L. The reproducibility of the method was good with the RSDs of 1.48%-13.40%. In addition, the intra-day and inter-day precisions were also satisfactory with the RSDs of 0.35%-15.54%. Mature fresh tobacco leaves from Yunnan, Henan and Guizhou Provinces, China were analyzed using this method. The results showed that the total amount of main phenols was Guizhou>Yunnan>Henan and there was significant difference between Guizhou and Henan. The amount of rutin was Yunnan>Guizhou>Henan, and there were significant differences among them each other. The method is reproducible with a simple pretreatment covering a wide range of phenols, and can be applied in batch analysis of tobacco leaves.

Key words: high performance liquid chromatography (HPLC), mass spectrometry (MS), phenolic compounds, tobacco leaves, ultraviolet detection