Precise Light-Driven\nPolarity of Stationary Phase\nfor Regulating Gradient Separation of Liquid Chromatography

Generally, the traditional stationary phase for liquid\nchromatography\nis the key part, but with an in situ immutable property, leading to\nmany separation limitations. Based on the former exploration of photosensitive\ngas chromatography, we successfully prepared a photosensitive monolithic\ncapillary silica column with high light transmission, taking advantage\nof the reversible cis–trans isomerism of azobenzene. And the\ncis–trans isomerism has launched an effective, reversible,\nand precise control on the liquid chromatographic retention behavior\njust by photoinduction according to the theoretical basis of a good\ncorrelation between photoinduction time, <i>trans</i>-azobenzene\nratio, and chromatographic retention factor (<i>k</i>) (<i>R</i><sup>2</sup> > 0.9586). In this system, the plausible\ncontrol\nmechanism of stationary phase’s polarity is the synergistic\neffect of the self-molecular polarity difference and the occupation\n(or release) of polar groups on the stationary phase surface by cis–trans\nisomerism. Furthermore, a “light control–time ratio\ncycle” strategy was first reported to maintain an arbitrary\nratio of <i>trans</i>-azobenzene corresponding to the polarity\ngradient of the stationary phase during the whole chromatographic\nseparation process, which is verified by a close correlation of the\nretention time of the targets and the ratio of light-controlled time\n(<i>R</i><sup>2</sup> > 0.9977). The “gradient\nelution”\nof the targets was also surprisingly realized due to the gradient\nregulation of stationary phase’s polarity by the programmatic\ncontrol of alternate light induction and time ratio. This work further\nadvances the practical significance of photosensitive chromatography\nin “one column for multiple columns”, even “one\ncolumn for multidimensional chromatography”, and truly provides\nresearch foundation for its development in the separation and analysis\nof complex biological samples.