Talanta 71 (2007) 784–789 Extraction and determination of morphine in compound liquorice using an aqueous two-phase system of poly(ethylene glycol)/K 2 HPO 4 coupled with HPLC Shehong Li, Chiyang He, Fei Gao, Daobing Li, Zheng Chen, Huwei Liu, Kean Li, Feng Liu ∗ The Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Institute of Analytical Chemistry, College of Chemistry, Peking University, Beijing 100871, China Received 24 February 2006; received in revised form 12 May 2006; accepted 15 May 2006 Available online 27 June 2006 Abstract An aqueous two-phase system (ATPS) of poly(ethylene glycol) (PEG)/K 2 HPO 4 coupled with high performance liquid chromatography (HPLC) method was developed for the separation and determination of morphine in compound liquorice. Morphine and its analogs were used as model compounds to investigate influence of various factors on extraction behaviors of ATPS, such as the types and concentrations of salts, PEG molecular mass, temperature and pH. It was observed that the types of salt had much influence on extraction efficiencies of morphine and its analogs. The results indicated that hydrophobic force cooperating with hydrogen bond interaction between analytes and phases played important role in extraction process. In the optimal system of containing 0.5 g PEG2000 and 1.5 g K 2 HPO 4 , the recoveries of the spiked standards for the analytes were all 91.7–100.3% with relative standard deviation of 1.0–3.0%. Morphine in compound liquorice was determined by the proposed method and the results were consistent with those of LC–MS method. Compared with conventional liquid–liquid extraction or solid-phase extraction, this extraction method can be completed in one operation and is low-cost. Since the entire extraction process is organic solvent-free, this new technique is environmental friendly. © 2006 Elsevier B.V. All rights reserved. Keywords: Extraction; Morphine and its analogs; Aqueous two-phase systems; Compound liquorice; HPLC; LC–MS 1. Introduction Compound liquorice is a commonly used Traditional Chi- nese Medicine, which has excellent curative effect for tussis. It mainly contains liquorice immersed ointment, opium and cam- phor. Morphine is one of the important alkaloids in opium and has stronger inhibition to respiratory center. It is report that long- term taking compound liquorice can result in addiction [1] and over dose of compound liquorice may induce toxicosis or death of newborn [2]. Therefore, “Chinese Codex” (the second edi- tion in 1995) rules that determination of morphine in compound liquorice is a guideline for quality control. Some analytical methods have been reported for the determination of morphine in biomaterials [3–6]. A high performance liquid chromatog- ∗ Corresponding author. Tel.: +86 10 62761187; fax: +86 10 62751708. E-mail address: liufeng@pku.edu.cn (F. Liu). raphy (HPLC) method [7] based on liquid–liquid extraction (LLE) was used for determination of morphine in compound liquorice, but it was difficult to apply it to actual operation due to the tedious sample pretreatment procedure, the unsatisfactory reproducibility and the usage of large volume of organic solvent that was harmful for environment and operator. A solid phase extraction (SPE) followed by HPLC method [8] was attempted, but the toxic organic solvents were still used. Sun et al. [9] reported a capillary zone electrophoresis (CZE) method for the determination of morphine and other components in compound liquorice. Aqueous two-phase system (ATPS) as a useful extraction technique is formed by mixing two aqueous solutions of struc- turally different polymers or by mixing a polymer and a salt aqueous solution together when the concentrations of the com- ponents are higher than a critical value. In the past few years, the aqueous two-phase system was proved a mild system and suitable for separation of biological samples [10] as a result 0039-9140/$ – see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.talanta.2006.05.017