In counter-current chromatography (CCC), linear scale-up is an ideal amplification strategy. However, when transferring from analytical to predictable preparative processes with high throughput, linear scale-up would be challenging due to limitations imposed by differences in instrument parameters, such as gravitational forces, tubing cross-section area, tubing length, column volume and flow rate. Some effective scale-up strategies have been studied for different instrument parameters, but so far, these scale-up works have only been tested on standard circular (SC) tubing.The previous research of our group found that rectangular horizontal (RH) tubing can double the separation efficiency compared with conventional SC tubing, and has industrial production potential. This paper used the separation of tilianin from Dracocephalum moldavica L. as an example to demonstrate how to scale up the optimized process from analytical SC tubing to preparative RH tubing. After systematic optimization of solvent systems, sample concentration and flow rate on the analytical CCC, the optimized parameters obtained were successfully transferred to the preparative CCC. The results showed that a crude sample of 2.07 g was successfully separated using a solvent system of n-hexane - ethyl acetate - ethanol - water (1:4:1:5, v/v/v/v) in reversed phase mode, and the three consecutive separations produced a total of 380 mg tilianin in 75 min with high purities of 98.3%, as analyzed by HPLC. The total throughput achieved from the analytical to semi-preparative scale was improved by 138 times (from 12 mg/h to 1.66 g/h), while the column volume was increased by only 46.5 times (from 15.5 mL to 720 mL). This is the successful application of CCC for the separation and purification of tilianin. Given that SC tubing is the traditional configuration for CCC columns, this study is a necessary step to prove the applicability of RH tubing columns for routine use and potential large-scale industrial applications.
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