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Abstract
A subzero temperature liquid–liquid extraction (ST-LLE) method was developed for the determination of five anthraquinone derivatives in a sticky traditional Chinese patent medicine, Xiao'er Resuqing Oral Liquid, by high performance liquid chromatography (HPLC). ST-LLE was based on the phenomenon that a clear phase separation took place in the acetonitrile aqueous solution when it was placed at a subzero-temperature freezer for a short period of time. The high viscosity resulted from the highly concentrated sugar in the medicine sample could be decreased by ST-LLE due to the low solubility of sugar in acetonitrile, and at the same time the five anthraquinone derivatives could be extracted to the acetonitrile phase formed at ST. The optimum conditions for ST-LLE were obtained as follows: 3.5 mL of diluted sample, pH 2.5, 2.0 mL acetonitrile, 30 min of extraction time, and −30°C of extraction temperature. The detection limits of aloe emodin, rhein, emodin, chrysophanol, and physcion were within 0.48 and 1.72 ng mL−1. Average recoveries for the anthraquinone derivatives at three spiked levels were in the range of 84.0–108.1%. Relative standard deviations for five replicate determinations for the real sample were below 5.5%. The established method has been successfully applied to the determination of anthraquinone derivatives in real samples.