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Bioactivity-guided chromatographic methods are of great significance for the isolation of the active compounds in complex samples. In this study, four antifungal compounds were located by activity screening and successfully isolated from a microbial fermentation sample by preparative high-performance liquid chromatography. Separation performance of columns including C18, positively charged C18, negatively charged C18 and C8 were firstly investigated. And it showed a better capacity of mixed-mode stationary phases for retention and separation. Therefore, the positively charged C18 column was used to separate the sample into several fractions, among which the active one was identified by the antifungal test. And then the active fraction was enriched and separated again by successively using the negatively charged C18 and C8 columns to obtain four compounds, which were identified as polyoxins A, K, F and H. With activity verification, four polyoxins were found to have good inhibitory effects against the three fungal plant diseases including rice sheath blight, tomato grey mould disease, and apple spot leaf disease.

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Developing methods for the isolation of highly polar compounds from complex samples is of great significance. In this study, three polyoxins were successfully isolated from a complex sample (PN1-1 # ) by preparative high-performance liquid chromatography. Separation was carried out on five polar reversed-phase stationary phases, using pure aqueous as mobile phase, where the C18HC column can provide the best performance for PN1-1 # . Next, the effects of the mobile phase composition were studied. It was found that adding NaClO 4 can enhance the retention and resolution, and adding NaH 2 PO 4 was beneficial to maintain good peak shapes when the sample loading increased. Therefore, the optimized mobile phase consisting of 20 mmol NaH 2 PO 4 and 20 mmol NaClO 4 (adding H 3 PO 4 to adjust pH 2) was used to separate PN1-1 # . This method of using 100% aqueous phase can effectively improve both the retention and the solubility of polar samples. Eventually, through further purification, three compounds, namely, polyoxins B, D, and G, were obtained. This paper provided an effective and eco-friendly strategy for the preparative-scale separation of polar samples.

An online supercritical fluid extraction with supercritical fluid chromatography system could provide sequential extraction and quantitative analysis of lignans in Schisandra chinensis. Supercritical fluid extraction conditions were optimized at 15 MPa, 50 • C, and 4 min with supercritical CO 2 adding 1% methanol; the elution volume and flow rate were set at 6 mL and 2 mL/min to blow extract out of the tank completely. The split-flow rate was confirmed at 2.5%, which determines injection volume and accuracy of quantitative detection. The factors having negative influences on supercritical fluid chromatography retention in the online system, including sample loading forms and backpressure settings, are discussed in the paper. At last, an extraction-quantitative method for lignans in Schisandra chinensis was developed, which could be finished within 19.5 min. The total content percentage of four lignans (Schisandrin, Schisandrin A, Schisandrin B and Schisandrol B) in four batches was respectively measured to be 1.42, 1.54, 1.62, and 1.90%. K E Y W O R D S quantitative analysis, Schisandra chinensis, supercritical fluid chromatography, supercritical fluid extraction 2444