An off-line two dimensional reversed-phase liquid chromatography/supercritical fluid chromatography (2D RPLC/SFC) method was developed for the separation of the seeds of Trichosanthes kirilowii Maxim. (T. kirilowii.). In the experiment, RPLC was used in the first dimension to analyze the samples. And 12 fractions (F1-F12) were obtained according to the chromatographic peak collection mode, the fractions were re-analyzed in both RPLC and SFC modes. The results clearly showed that this 2D method had good orthogonality. The SFC was used as the second dimension. Ethanol-n-hexane (3:7, v/v) was used as a modifier in SFC. The modifier could provide appropriate elution power. When the sample load increased, it could also ensure the solubility of samples. The 2D RPLC/SFC method had obvious advantages for separating the compounds with similar properties, enriching and purifying the trace components. Finally, 150 peaks could be detected. The off-line 2D RPLC/SFC method will be further enlarged to a preparative level for preparation of compounds. The method can provide a practical reference for the purification of chemical constituents and material basis research of the seeds of T. kirilowii. in the future.
Keywords: reversed-phase liquid chromatography (RPLC); separation; supercritical fluid chromatography (SFC); the seeds of Trichosanthes kirilowii Maxim.(T.kirilowii.); two dimensional (2D) chromatography.