An efficient method used to separate five bioactive compounds from Gelidium amansii was optimized by the HCI software. The optimum composition of mobile phase for high-performance liquid chromatographic (HPLC) separation was obtained. The elution profiles were calculated by the polynomial theory based on the retention factor ln k = A + BF + CF2 (F was the volume percentage of acetonitrile with 1.0% acetic acid); then, the theory was applied to calculate the elution profile in both isocratic and gradient modes by modifying different mobile phase conditions with HCI program. The calculated results of mobile phase condition suggested that acetonitrile-water (containing 1.0% acetic acid) with a linear gradient elution of 0∼30 min from 15:85 to 50:50 (v/v) was the optimized component. In the experimental conditions, the agreement between the experimental elution profiles and the calculated values of eluted concentration was relatively good.
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