Low-temperature derivatization followed by vortex-assisted liquid-liquid microextraction for the analysis of polyamines in Nicotiana Tabacum

Polyamines are ubiquitous polycationic molecules that play a key role in many biological processes such as nucleic acid metabolism, protein synthesis, cell growth, and nicotine synthesis precursors. This work describes a rapid, sensitive, convenient, green, and cost-effective method for the determination of polyamines in Nicotiana tabacum by ultra high performance liquid chromatography with photodiode array detection. The analytes were derivatized with 3,5-dinitrobenzoyl chloride at low temperature (about 4 degrees C) and then extracted with vortex-assisted liquid-liquid microextraction. The experimental designs based on quarter-fractional factorial design and Doehlert design were used to screen and optimize the important factors in microextraction process. Under the optimal conditions, the method was linear over 0.05-8.00 mu g/mL with an r(2) >= 0.992 and exhibited good repeatability and reproducibility less than 6.0 and 6.9%, respectively. The limit of detection ranged between 0.013 and 0.029 mu g/g. The newly developed method was successfully employed to analyze different leaf samples of Nicotiana tabacum, among which the polyamines contents were found to be very different. Moreover, tyramine, 1,3-diaminopropane, homospermidine, and canavalmine were tentatively identified with the electrospray ionization quadrupole time-of-flight mass spectrometry. To our knowledge, this is the first report of identification of canavalmine in Nicotiana Tabacum.