научная статья по теме RAPID DETERMINATION OF PAEONOL IN TRADITIONAL CHINESE MEDICINAL PREPARATIONS BY MICROWAVE-ASSISTED EXTRACTION FOLLOWED BY HEADSPACE SOLID-PHASE MICROEXTRACTION AND GAS CHROMATOGRAPHY-MASS SPECTROMETRY Химия

Текст научной статьи на тему «RAPID DETERMINATION OF PAEONOL IN TRADITIONAL CHINESE MEDICINAL PREPARATIONS BY MICROWAVE-ASSISTED EXTRACTION FOLLOWED BY HEADSPACE SOLID-PHASE MICROEXTRACTION AND GAS CHROMATOGRAPHY-MASS SPECTROMETRY»

ЖУРНАЛ АНАЛИТИЧЕСКОЙ ХИМИИ, 2011, том 66, № 3, с. 292-296

ОРИГИНАЛЬНЫЕ СТАТЬИ

УДК 543

RAPID DETERMINATION OF PAEONOL IN TRADITIONAL CHINESE MEDICINAL PREPARATIONS BY MICROWAVE-ASSISTED EXTRACTION FOLLOWED BY HEADSPACE SOLID-PHASE MICROEXTRACTION AND GAS CHROMATOGRAPHY-MASS SPECTROMETRY

© 2011 г. Qing Ye, Dagui Zheng, Zongbao Chen

Key Labortatory of Applied Organic Chemistry, Higher Institutions of Jiangxi Province, Shangrao Normal University

Shangrao 334001, China Received 24.08.2009; in final form 10.06.2010

Microwave-assisted extraction (MAE) followed by headspace solid-phase microextraction (HS-SPME) was developed for rapid determination of paeonol in four traditional Chinese medicinal preparations (TCMPs) including Liuwen Dihuang pills, Maiwei Dihuang pills, Guifu Dihuang pills, and Zhibai Dihuang pills. The optimal MAE conditions obtained were: microwave power of 540 W and irradiation time of 4 min, and HS-SPME optimal conditions were: fiber coating of 100 ^m PDMS/DVB, extraction temperature of 70°C, extraction time of 10 min, stirring rate of 1100 rpm, and NaCl concentration 30%. The optimized method provided satisfactory precision (RSD 6.5%), good linearity (R2 > 0.997) and recovery (90%). The results showed that MAE—HS-SPME—GC-MS is a simple, rapid, efficient, and solvent-free technique for the quantitative determination of paeonol in TCMPs.

Keywords: microwave-assisted extraction, headspace solid-phase microextraction, traditional Chinese medicinal preparations, paeonol, gas chromatography-mass spectrometry.

Traditional Chinese medicinal preparations (TCMPs) are usually made from a variety of Chinese herbs. Liuwei Dihuang pills, one of the most important TCMPs, are composed of six kinds ofherbs including Cortex Moutan and are widely used in Eastern Asia. In China, there are hundreds of medicine manufacturers who produce Liuwei Dihuang pills and its varieties, such as Zhibai Dihuang pills, Guifu Dihuang pills, Mingmu Dihuang pills, Qiju Dihuang pills, Maiwei Dihuang pills, etc [1]. Paeonol used as quality control marker is the main active constituent present in several TCMPs [2], and is known to have many pharmacological effects, including anti-inflammatory and analgesic action [3], inhibition of anaphylactic reaction [4], inhibition of monoamine oxidase A and B [5], anxiolytic effect [6] and inhibition ofplatelet aggregation [7]. It is very interesting to quantitatively determine paeonol concentration in TCMPs. Many methods, such as thin-layer chromatography [8, 9], high-performance liquid chromatography (HPLC) [10, 11], and micellar capillary electrophoresis [12] were developed for the determination of paeonol in several TCMPs. At the same time, gas chromatography—mass spectrometry (GC—MS) is a very simple, rapid, and high-resolution tool for the analysis ofvolatile constituents in TCMs and TCMPs [13—17]. Prior to analysis, it was required to isolate and extract paeonol from the TCMPs. Ultrasonic assisted extraction (UAE) [1, 18] and supercritical fluid extraction (SFE) [11] were developed for the determination of paeonol in TCMPs.

Compared with UAE and SFE, MAE is a fast sample preparation technique (less than 10 min), which requires simple equipment. MAE technique has been applied to the extraction of organic compounds from different matrices including plant materials [19—22]. In MAE, water was usually used as the extraction solvent. MAE becomes a green sample extraction technique. Recently, MAE was successfully applied to fast extraction of active components in TCMs [23—25]. When water was used to extract the analyte from the sample by MAE, prior to GC-MS, the solution required further extraction by using organic solvent. So, it is very interesting to develop a solvent-free procedure for the further extraction and preconcentra-tion of MAE extract.

SPME is an extraction and preconcentration method. It is a solvent-free technique, widely applied to food, environmental, biological and TCM analysis [26—36].

In this work, for the first time, MAE was combined with HS-SPME to develop a procedure for the determination of paeonol in four TCMPs by GC-MS. The experimental parameters ofMAE—SPME were optimized, and validation of the method was performed.

EXPERIMENTAL

TCMP sample, chemicals, SPME fiber and microwave oven. Liuwen Dihuang pills-1 was manufactured by Shanghai Leiyingshang Medicine Company, Shanghai,

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China. Liuwen Dihuang pills-2 was manufactured by Henan Wanxi Medicine Company, Henan, China. Zhbai Dihuang pills were manufactured by Foci Medicine Company, Lanzhou, China. Guifu Dihuang pills were manufactured by Shanghai Tonghanchuntang Medicine Company. Mingmu Dihuang pills were manufactured by Lanzhou Foci Medicine Company. Paeonol was obtained from the National Institute for the Control of Pharmaceuticals and Biological Products, Beijing, China. Methanol (analytical grade) and sodium chloride were purchased from Chemical Agent Company, Shanghai, China. The SPME fibers: 100 ^m polydimethylsiloxane (PDMS), 65 ^m poly-dimethylsiloxane/divinylbenzene (PDMS/DVB), 65 ^m carbowax/divinylbenzene (CW/DVB), 75 ^m carbox-en/polydimethylsiloxane (CAR/PDMS), and 85 ^m poly-acrylate (PA) were purchased from Supelco, Bellefonte, USA. The microwave oven with a maximum delivered power of 700 W was purchased From Haier Company (Qingdao, China).

Calibration solution preparation. Stock solution of paeonol (2.0 mg/mL) was prepared in methanol and stored at 4°C. Working solutions with the paeonol concentration from 10 to 200 ^g/mL were prepared by dilution of the stock solution with distilled water.

GC-MS. An HP 6890 GC system, coupled with a HP MD5973 quadrupole mass spectrometer was used. The extracted compounds were separated on an HP-5MS capillary column (30 m x 0.25 mm I.D., 0.25 ^m film). The column oven temperature was programmed to rise from an initial temperature of 40°C (3 min) to 300°C at 15°C/min. The injection temperature and ion source temperature were 250 and 230°C, respectively. Helium was used as the carrier gas with a flow rate of1 mL/min. The ionizing energy was 70 eV. All data were obtained by collecting the full-scan mass spectra within the scan range of40—350 amu.

Optimization of MAE-HS-SPME procedure. In the

proposed procedure, paeonol was extracted from TCMPs using MAE. The aqueous extract was further extracted and concentrated by using SPME fibers. In the work, the experimental parameters of MAE (microwave power, irradiation time) and HS-SPME (fiber coating, extraction temperature, extraction time, stirring rate, and salt effect) were studied. For Liuwen Dihuang pills used in the work, water was selected as the MAE solvent. A Liuwen Dihuang pills sample (0.5 g) was introduced into a 25 mL bottle, and 10 mL water was added as the extraction solvent. MAE of the TCMP sample was performed at different microwave power (230, 380, 540 and 700 W) with different irradiation time of 2, 4, 6 and 8 min. The obtained aqueous extract (5.0 mL) was further extracted and concentrated by using a PDMS—DVB fiber at 50°C for 10 min. The analytes extracted from the fiber were analyzed by GC-MS.

HS-SPME parameters were also studied. Liuwen Di-huang pills sample was extracted under the optimized MAE conditions. The obtained aqueous extract in the

volume of 5.0 mL was introduced into a 10 mL head-space vial with 1 cm stirring bar. Headspace extraction was performed at 50°C for 10 min, using five different fibers. Sequentially, extraction temperature (30—90°C), extraction time (5—40 min), stirring rate (500—1100 rpm) and NaCl concentration (0—30%) were studied.

Quantification of paeonol in TCMP samples by MAE-HS-SPME. Four TCMP samples were ground to fine powder, and 0.5 g of each TCMP sample were added in the extraction bottle containing 10 mL of distilled water. MAE was performed at 540 W for 4.0 min. After MAE, 5.0 mL of the aqueous extract were introduced into a 10 mL headspace vial. Paeonol in the extracts was extracted from the headspace and concentrated by using the 65 ^m-PDMS/DVB fiber, with the optimal HS-SPME conditions. Finally, the analyte on the fiber was determined by GC—MS. To quantify paeonol in the TCMPs, 5 mL of calibration solutions (10—200 ^g/mL) were introduced into 10 mL headspace vials by HS-SPME with the optimal SPME conditions and analyzed by GC—MS. To demonstrate the proposed method, steam distillation (SD) method was used for the determination of paeonol in the four TCMPs, according to the Chinese pharmacopoeia [2].

Linearity, precision, recovery, limit of detection (LOD) and limit of quantification (LOQ). The linearity was investigated by three replicate analyses within the interesting concentration range (10—200 ^g/mL ).The method precision was studied by six replicate analyses of paeonol in Liuwei Dihuang pills by MAE-HS-SPME under the optimum conditions. RSD was calculated based on the obtained peak areas. Recovery was also investigated by adding 20 ^L of standard stock solution (2.0 mg/mL) to Liuwei Dihuang pills (0.5 g) containing known amounts of paeonol. Triplicate measurements were performed by MAE-HS-SPME-GC-MS. The LOD and LOQ were estimated by replicate analyses of the calibration solution of known concentration.

RESULTS AND DISCUSSION

Optimization of MAE-HS-SPME parameters. In

the study, the experimental parameters of MAE and HS-SPMe were studied. The Liuwei Dihuang pills (0.5 g) sample was used. The extraction efficiency ofpae-onol was used to determine the optimal MAE and HS-SPME parameters. The extraction efficiency of paeonol S-

was calculated as = — x 100%, where S, and Se are the

S ' e

concentration of paeonol in Liuwei Dihuang pills sample obtained by the proposed method and steam distillation method, respectively.

At first, the MAE parameters ofmicrowave power and irradiation time were studied. MAE of Liuwei Dihuang pills was performed at different microwave power (230, 380, 540 an

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