Separation methods used for Scutellaria baicalensis active components

Abstract

Scutellaria baicalensis Georgi is one of the most widely used traditional Chinese herbal medicines. Its roots have been used for anti-inflammation, anticancer, antiviral and antibacterial infections of the respiratory and the gastrointestinal tract, cleaning away heat, moistening aridity, purging fire, detoxifying toxicosis, reducing the total cholesterol level and decreasing blood pressures. Baicalin, baicalein, wogonin and oroxylin A are its main active components. This review provides an overview of various separation, detection, and identification techniques employed for the quantitative and qualitative determination of these active components. Applications of high-performance liquid chromatography, high-speed counter-current chromatography, thin layer chromatography, capillary electrophoresis, and micellar electrokinetic capillary chromatography to the separation and determination of these active components are described. Examples of identification of these active components and their metabolites in complex matrices by high-performance liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry are also presented. The advantages and limitations of these separation and identification methods are assessed and discussed.

Fig. 1

Chemical structures of six S. baicalensis active components.

Fig. 2

Chromatogram of the marker substances gentiopicroside, mangiferin, palmatine, berberine, baicalin, glycyrrhizin and wogonin in Sann-Joong-Kuey-Jian-Tang. Column, Cosmosil 5C₁₈-MS, 150 mm × 4.6 mm i.d.; guard column, Cosmosil 5C₁₈-AR, 50 mm × 4.6 mm i.d., mobile phase, 0.03% phosphoric acid–acetonitrile (0 min, 90:19; 10 min, 87:13; 17–27 min, 77:23; 40 min, 62:38; 50 min, 55:45); flow-rate, 1.0 ml min⁻¹. (1) Standard decoction; (2) commercial preparation; (3) standard decoction without Gentianae scabrae radix. Reprinted from with permission from Elsevier.

Fig. 3

High-performance liquid chromatograms of the standard solution (A) and I-Tzu-Tang (B). AE: aloe–emodin, B: baicalein, BG: baicalin, CA: caffeic acid, E: emodin, FA: ferulic acid, GZ: glycyrrhizin, IS: internal standard (tetralin), OG: oroxylin A 7-O-glucuronide, SA: sennoside A, SB: sennoside B, W: wogonin, WG: wogonin 7-O-glucuronide. Reprinted from with permission from Elsevier.

Fig. 4

Chromatogram of the crude baicalin extracted from S. baicalensis Georgi by HSCCC separation. Curve 2: baicalin. Conditions: column, multilayer coil of 1.6 mm i.d. PTFE tube with a total capacity of 342 ml; rotary speed, 900 rpm; solvent system, n-butanol–water (1:1, v/v); mobile phase, the lower phase; flow-rate, 1.0 ml min⁻¹; detection, 277 nm; sample size, 200 mg; injection volume, 5 ml; retention of the stationary phase, 41.8%; (A) the first separation; (B) the second separation. Reprinted from with permission from Elsevier.

Fig. 5

Electropherograms of the standard solution (A) and the scute-coptis herb couple (B) with internal standard (IS). 1, coptisine; 2, berberine; 3, epiberberine; 4, palmatine; IS, umbelliferone; 5, oroxylin A; 6, wogonin; 7, wogonin 7-O-glucuronide; 8, oroxylin A 7-O-glucuronide; 9, baicalin; 10, baicalein. Reprinted from with permission from Elsevier.

Fig. 6

Electropherograms of the standard solution (A) and I-Tzu-Tang (B). IS: internal standard (salicylic acid), other symbols as in Fig. 3. Reprinted from with permission from Elsevier.

Fig. 7

LC–MSⁿ analysis of wogonin and its conjugated metabolites in plasma after an administration of 5 mg kg⁻¹ wogonin to a Wistar rat. (A) Total ion current and selected ion monitoring chromatograms (A2, wogonin glucuronide; A3, wogonin sulfate; A4, wogonin glucoside; A5, free wogonin); (B) full scan MS–MS spectrum of peak at 6.18 min; (C) full scan MS³ spectrum of peak at 6.18 min. Reprinted from with permission from Elsevier.

Fig. 8

Representative SRM chromatograms of: (A) a blank rat plasma sample; (B) a blank plasma sample spiked with 2.0 ng ml⁻¹ wogonin and the internal standard daidzein (IS, 1000 ng ml⁻¹); (C) plasma sample from a Wistar rat 0.5 h after an oral administration of wogonin (5 mg kg⁻¹). Peak I, wogonin; peak II, IS. Reprinted from with permission from Elsevier.