Separation of 15 impurities in semi-synthetic paclitaxel in a single run using supercritical fluid chromatography and greenness assessment by AMGS, GAPI, and AGREE tools.

The semi-synthetic approach for paclitaxel production has emerged as the predominant industrial method. However, the complex structure of paclitaxel leads to by-product formation during synthesis, and it is unstable in acidic, alkaline, and oxidative conditions. To ensure the safety, efficacy, and quality control of paclitaxel, this study successfully developed a rapid, straightforward, sensitive, and environmentally sustainable supercritical fluid chromatography (SFC) method for the quantification of 15 impurities in semi-synthetic paclitaxel raw materials. A comprehensive evaluation was conducted on the effects of stationary phase, mobile phase, column temperature, and back pressure on the retention and separation of analytes. Following method optimization, utilizing CO and methanol as mobile phases, satisfactory results were achieved on the DEA stationary phase within 28 minutes, reducing the analysis time to half compared to the RPLC methods specified in the Chinese Pharmacopoeia (ChP), United States Pharmacopeia (USP), and European Pharmacopoeia (EP). The validation results of the proposed method demonstrate its high sensitivity, with a minimum detection limit for impurities of approximately 0.25 μg·ml. The precision, as indicated by the relative standard deviation (RSD), was consistently below 2.0 %, and the recovery rates ranged from 98.03 % to 104.2 %. Furthermore, the SFC method effectively encompasses 15 known impurities present in the semi-synthetic paclitaxel as listed in the pharmacopoeia. It offers significant advantages in terms of throughput, cost-effectiveness, and operational simplicity. The green assessment results indicate that, compared to reversed-phase liquid chromatography (RPLC), this method is sustainable and environmentally friendly, aligning with the principles of green chemistry. This study provides a valuable reference for further investigations into the impurities of semi-synthetic paclitaxel and supports the application of SFC in the rapid separation of structurally similar compounds.