Efficient Production of Antioxidants from Rape Pollen via a Chromatographic Strategy

Rape pollen has always been considered as a research hotspot in health foods and pharmaceuticals due to its abundance of natural active ingredients. In this work, a compound with antioxidant activity was directly isolated from the methanol extract of rape pollen using a two-step procedure, under the supervision of online HPLC-1,1-diphenyl-2-picrylhydrazyl (HPLC-DPPH) detection. Firstly, online HPLC-DPPH detection was used to identify the active peaks in the methanol extract of rape pollen, and then the methanol extract was pretreated via medium-pressure liquid chromatography (MPLC) to obtain the target fraction 3 (Fr3). Fr3 was further purified using HPLC to finally obtain the target fraction 3-1, which was identified as kaempferol 3,4 '-di-O-beta-D-glucopyranoside through NMR and mass spectrometry. To further explore the free radical scavenging activity of this compound, its DPPH scavenging ability was determined, and two proteins related to the antioxidant pathway were used for molecular docking. The results revealed that the chromatographic strategy used in this study was efficient and reliable in separating high-purity antioxidants from rape pollen. A strategy such as this, meanwhile, also holds promise for qualitatively identifying and specifically isolating active compounds from other natural products.