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Home , Scutellarein, Scutellaria, Scutellarin

Radix Scutellariae – Huangqin

Pharmacopoeia(1): Pharmacopoeia of the People’s Republic of China, English Edition Vol. I, 2005

Of¿ cial drug(1): Baical Skullcap Root is the dried root of baicalensis Georgi (Fam. ). The drug is collected in spring or autumn, removed from rootlet and soil, dashed to peel the rugged outer bark after being sun-dried, and then dried thoroughly.

Origin(6,7): Originated from the provinces Hebei and Shanxi; Inner Mongolia, Japan, Korea and Russia.

Description of the drug(1): Conical, twisted, 8–15 cm long, 1–3 cm in diameter. Externally brownish-yellow or dark yellow, bearing sparse warty traces of rootlets, the upper part rough with twisted longitudinal wrinkles or irregular reticula, the lower part with longitudinal striations and ¿ ne wrinkles. Texture hard and fragile, easily broken, fracture yellow, reddish-brown in the centre; the central part of an old root dark brown or brownish-black, withered or hollowed. Odour, slight; taste, bitter. Cultivar roots slender, mostly branched. Externally yellowish- brown, outer bark closely adhering to wood, with relatively thin longitudinal wrinkles. Fracture yellow or yellowish, slightly horny. Taste bitter.

Substitute drugs: Several other species of Scutellaria from other regions of China have been described as substitutes for . These include S. amoena, S. hypericifolia, S. likiangensis, S. rehderiana, S. tenax and S. viscidula. In Western Herbal Medicine, especially in USA and Canada the aerial parts (herb, ) of Scutellaria lateriÀ ora are commonly used.

Pre-treatment The drug is removed from rootlet and soil, dashed to peel the of the raw drug(1): rugged outer bark after being sun-dried, and then dried thoroughly. a) Processed with water: The dried root is boiled in water for 10 min. (or steamed for 30 min.) to soften thoroughly, then cut into thin slices and dried, protecting from strong sunlight. Occurring in sub rounded or irregular thin slices; externally yellowish-brown, cut surface yellowish-brown or yellowish-green, striated radially.

H. Wagner et al. (eds.), Chromatographic Fingerprint Analysis of Herbal Medicines 755 © Springer-Verlag/Wien 2011 Radix Scutellariae – Huangqin

b) Processed with wine: Radix Scutellariae is described under the method for stir-frying with wine to dryness. Occurring in sub rounded or irregular thin slices; externally brown; cut surface yellowish-brown, striated radially, showing less burnt patches, sometimes appearing brown in the centre.

Medicinal use(7): In TCM to treat fever, nausea and vomiting, acute dysentery, jaundice, coughs, carbuncles and sores, and threatened abortion. In folk medicine to treat allergies, atherosclerosis, diarrhea, dermatitis and hypertension.

Effects and indications of Radix Scutellariae according to Traditional Chinese Medicine(1, 2, 3, 4)

Taste: bitter

Temperature: cold

Channels entered: Orbis cardialis et intestini tenuis, Orbis pulmonalis et intestini crassi, Orbis felleus, Orbis stomachi.

Effects (functions): To remove damp-heat, quench ¿ re and counteract toxicity, arrest bleeding, and prevent abortion.

Symptoms and indications: Discomfort in the chest, nausea and vomiting in epidemic febrile diseases caused by damp-heat or summer-heat; feeling of stuf¿ ness in the abdomen, acute dysentery or jaundice caused by damp-heat; cough due to heat in the lung; high fever with dire thirst; spitting of blood and epistaxis due to heat in blood; carbuncles and sores; threatened abortion.

Main constituents(5, 6, 7, 8, 9,10):

Main À avonoids: • (-7-O-glucuronide) • Baicalein (5,6,7-trihydroxyÀ avone) • Wogonoside (-7-O-glucuronide) • Wogonin (5,7-dihydroxy-8-methoxyÀ avone) • (-7-O-glucuronide) • Scutellarein (5,6,7-4’-tetrahydroxyÀ avone) • Oroxyloside (Oroxylin-A-7-O-glucuronide) • (5,7-dihydroxy-6-methoxyÀ avone)

756 Radix Scutellariae – Huangqin

Minor À avonoids and other constituents: • Viscidulin III-2’-O-glucoside • Viscidulin III • -6-C-arabinosyl-8-C-glucoside • Chrysin • SkullcapÀ avone I and II (I = 5,2’- OH, 7,8-OCH3- À avone; II = 5,2’- OH, 6,7, 8,5’- OCH3-À avone) • trans-Verbascoside • trans-Martynoside • 5,7,2’,6’-TetrahydroxyÀ avone • Dihydro-Oroxylin A

Note:

In the literature(8, 9) at least more than 20 other À avonoids and phenylpropan-derivates (trans- Martynoside, trans-Verbascoside) are described for the various Scutellaria species. Most of them described under “Minor À avonoids” occur in Scutellaria species which are listed as “substitute drugs” of Scutellaria baicalensis.

Trans-Martynoside, trans-Verbascoside, Viscidulin III-2’-O-glucoside, and Viscidulin III, primarily described as constituents of the herb of Scutellaria lateriÀ ora, could be detected also in small amounts in the HPLC ¿ ngerprint of Scutellaria baicalensis root.

Pharmacology: – in vitro and in vivo effects: - anti-inÀ ammatory activity(7,12,15,20) - antihepatotoxic activity(7) - antioxidant activity(7,12,15) - antimicrobial activity(7,8,11) - antitumor activity(7,8,19) - antiviral activity(7,17,21,22) - sedative and activity(7,14,16) - inhibitory effect of neurotoxicity(13) - platelet aggregation inhibition(7,8) - anticonvulsant activity(7,9) - inhibition of mast cell histamine release(18)

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Fig. 1: Formulae of the main compounds as detected in TLC or HPLC of Radix Scutellariae(8, 9)

758 Radix Scutellariae – Huangqin

TLC-¿ ngerprint analysis:

1) Extraction(1): 1 g powdered drug is extracted under reÀ ux for 30 minutes with a mixture of 20 ml ethyl acetate and 10 ml . After cooling and ¿ ltration the extract is evaporated to dryness. The residue is dissolved in 5 ml methanol and once more ¿ ltrated to use the solution for TLC.

2) Reference compounds: each 0.5 mg is dissolved in 0.5 ml methanol.

3) Separation parameters:

Plate: HPTLC Silica gel 60 F254, Merck

Applied amounts: Radix Scutellariae extracts: each 25 —l reference compounds: each 25 —l

Solvent system: : ethyl acetate : methanol : formic acid 10 3 1 2

Detection: Iron-III-chloride (1g in 100 ml EtOH) After spraying the plate is heated at 100°C for 10 minutes.

Fig. 2: Thin layer chromatogram of the methanol root extracts of Scutellaria baicalensis sprayed with Iron-III-chloride reagent (VIS)

759 Radix Scutellariae – Huangqin

4) Description: All ¿ ve extract samples show in VIS a very homogenous ¿ ngerprint pattern of green and violet brown zones respectively. In Fig. 2 only the samples 1, 2 and 3 are shown. Baicalin (T1), one main À avonolglycoside, shows its green zones at Rƒ = 0.31. Scutellarin (T2) at Rƒ = 0.28 is present only in a very low concentration and partly overlapped by Baicalin. The aglycone Baicalein together with Scutellarein appear at Rƒ = 0.85. Directly above Baicalein lies Wogonin at Rƒ = 0.94. In the middle Rƒ-range between Rƒ = 0.5 and 0.65, two main brownish zones, can be assigned to Oroxylin-A-7-glucuronide (Oroxyloside) and Wogonin- 7-O-glucuronide (Wogonoside). In the deep Rƒ-range at 0.2 to 0.25 a further red brown band might be identical with Chrysin-di-C-glycoside or Viscidulin III-2’-O-glucoside analogous to the peaks between Rt 4.0 and 10.3 in the HPLC-¿ ngerprint (see Table 1).

Drug samples Origin

1 Radix Scutellariae / Scutellaria baicalensis Sample of commercial drug obtained from public pharmacy, Munich, Germany

2 Radix Scutellariae / Scutellaria baicalensis Sample of commercial drug obtained from HerbaSinica, Germany (origin: Neimenggu, China)

3 Radix Scutellariae / Scutellaria baicalensis Sample of commercial drug obtained from TCM-Hospital, Bad Kötzting, Germany

4 Radix Scutellariae / Scutellaria baicalensis Sample of commercial drug obtained from China Medica, Germany

5 Radix Scutellariae / Scutellaria baicalensis Sample from German cultivation obtained from China Medica, Germany

6 Radix Scutellariae / Scutellaria baicalensis Sample of commercial extract obtained from China Medica, Germany

7 Radix Scutellariae / Scutellaria baicalensis Province Anhui, China

8 Radix Scutellariae / Scutellaria baicalensis Province Hebei, China

Reference compounds of Figure 2 Rƒ

T 1 Baicalin 0.31

T 2 Scutellarin / Baicalein (Scutellarein) 0.28 / 0.85

T 3 Wogonin 0.94

760 Radix Scutellariae – Huangqin

HPLC-¿ ngerprint analysis(22,23,24,25):

1) Sample preparation(1): 1 g powdered drug is extracted under reÀ ux for 30 minutes with a mixture of 20 ml ethyl acetate and 10 ml methanol. The extract is cooled, ¿ ltered and evaporated to dryness. The residue is dissolved in 5.0 ml of methanol, ¿ ltered over Millipore® ¿ ltration unit, type 0.45 —m and injected into the HPLC apparatus.

2) Injection volume: Radix Scutellariae extracts: each 10.0 —l

3) HPLC parameter:

Apparatus: MERCK HITACHI D-6000 A Interface MERCK HITACHI L-4500 A Diode Array Detector MERCK HITACHI AS-2000 Autosampler MERCK HITACHI L-6200 A Intelligent Pump

Separation column: LiChroCART® 250-4 LiChrospher® 100 RP-18 (5 ȝm), Merck

Precolumn: LiChroCART® 4-4 LiChrospher® 100 RP-18 (5 ȝm), Merck

Solvent System: A: 0.1 % H3PO4 (phosphoric acid, Merck / water, Millipore Ultra Clear UV plus® ¿ ltered) B: Methanol (VWR)

Gradient: 40 % B in 25 minutes 40–60 % B in 20 minutes 60 % B in 15 minutes total runtime: 60 minutes

Flow: 1.0 ml/min.

Detection: 276 nm

761 Radix Scutellariae – Huangqin

Table 1: Retention times of the main peaks

Peak Rt (min.) Compound 1 ~ 4.1 Viscidulin III-2’-O-glc* 2 ~ 6.5 Chrysin-6-C-ara-8-C-glc* 3 ~ 8.6 / ~ 9.6 ? trans-Verbascoside* 4 ~ 10.3 Viscidulin III* 5 ~ 20.6 Baicalin 6 ~ 28.5 /~ 31.5? trans-Martynoside* 7 ~ 33.6 Oroxylin A-7-O-glucuronide* 8 ~ 36.2 Wogonin-7-O-glucuronide (Wogonoside)* 9 ~ 46.4 Baicalein* 10 ~ 54.4 Wogonin 11 ~ 59.3 Oroxylin A* * identi¿ ed analogous to the HPLC-analysis of American Skullcap(9), the herb of Scutellaria lateriÀ ora, which contains Baicalin as dominant À avonoid and small amounts of Oroxylin- and Wogonin-7-glucuronide.

Fig. 3a: HPLC ¿ ngerprint of Scutellaria baicalensis root extract sample 1

Fig. 3b: HPLC ¿ ngerprint of Scutellaria baicalensis root extract sample 2

762 Radix Scutellariae – Huangqin

Fig. 3c: HPLC ¿ ngerprint of Scutellaria baicalensis root extract sample 3

763 Radix Scutellariae – Huangqin

Fig. 4: On line UV-spectra of the main characteristic peaks of the HPLC-¿ ngerprint of the root extracts of Scutellaria baicalensis

4) Description of the HPLC: All three Scutellaria root extract samples recorded at 276 nm show a very identical pattern of 11 main peaks: The detected À avones are Baicalin (peak 5), Wogonin-7-O-glucuronide (peak 8), Baicalein (peak 9) and Wogonin (peak 10). The other peaks 1 – 4, 6, 7 and 11 could be identi¿ ed according to the publication of Zhang et al.(9) who used a very similar HPLC-system for the HPLC-¿ ngerprint-analysis of the American Skullcap, the herb of Scutellaria lateriÀ ora. The on line UV-spectra are in agreement with those of Zhang’s publication.

Note: The Chinese Pharmacopoeia 2005 describes a content not less than 8 % of baicalin calculated with reference to the dried drug as determined by HPLC.

764 Radix Scutellariae – Huangqin

References: (1) Pharmacopoeia of the People’s Republic of China (English Edition, Vol. I), People’s Medical Publishing House, Beijing (2005) (2) Porkert, Klinische Chinesische Pharmakologie, Verlag für Medizin Dr. Ewald Fischer, Heidelberg (1978) (3) Junying et al., Materia medica der Chinesischen Arzneimitteltherapie, Band 2, Verlag für Ganzheitliche Medizin, Bad Kötzting (1993) (4) Hempen C-H, Fischer T, Leitfaden Chinesische Phytotherapie 2. AuÀ age, Urban & Fischer, Munich (2007) (5) Stöger, EA, Arzneibuch der chinesischen Medizin, Dtsch. Apotheker-Verlag, Stuttgart (2001) (6) Keys, Chinese Herbs, Charles E. Tuttle Company, Japan (1976) (7) Traditional Medicine Team, Department of Technical Co-operation for Essential Drugs and Traditional Medicine, World Health Organisation: WHO monographs on selected medicinal : Revised Draft monograph on Radix Scutellariae, May 2001 (8) Tang W, Eisenbrand G, Chinese drugs of origin, Springer-Verlag Berlin Heidelberg (1992) (9) Zhang Z, Lian X-Y, Li S, Stringer J, Characterization of chemical ingredients and anticonvulsant activity of American skullcap, Phytomedicine 16 (5):485-493 (2009) (10) Tomimori T, Jin H, Miyaichi Y, Toyofuku S, Namba T, Studies on the constituents of Scutellaria species. VI. On the À avonoid constituents of the root of Scutellaria baicalensis GEORGI (5). Quantitative analysis of À avonoids in Scutellaria roots by high- performance liquid chromatography, Yagugaku Zasshi. 105 (2): 148-55 (1985 Feb) (11) Wu J, Hu D, Wang KX, Study of Scutellaria baicalensis and Baicalin against antimicrobial susceptibility of Helicobacter pylori strains in vitro, Zhong Yao Cai. 31 (5): 707-10 (May 2008). (12) Huang WH, Lee AR, Yang CH, Antioxidative and anti-inÀ ammatory activities of polyhydroxy-À avonoids of Scutellaria baicalensis GEORGI, Biosci Biotechnol Biochem. 70 (10): 2371-80 (Epub 2006 Oct 7) (13) Heo HJ, Kim DO, Choi SJ, Shin DH, Lee CY, Potent Inhibitory effect of À avonoids in Scutellaria baicalensis on amyloid beta protein-induced neurotoxicity, J Agric Food Chem. 52 (13): 4128-32 (2004 Jun 30) (14) Wang H, Hui KM, Chen Y, Xu S, Wong JT, Xue H, Structure-activity relationships of À avonoids, isolated from Scutellaria baicalensis, binding to site of GABA(A) receptor complex, Planta Med. 68 (12): 1059-62 (2002 Dec) (15) Schinella GR, Tournier HA, Prieto JM, Mordujovich de Buschiazzo P, Rios JL, Antioxidant activity of anti-inÀ ammatory plant extracts, Life Sci. 70 (9): 1023-33 (2002 Jan 18) (16) Hui KM, Wang XH, Xue H, Interaction of À avones from the roots of Scutellaria baicalensis with the benzodiazepine site, Planta Med. 66 (1): 91-3 (2000 Feb) (17) Huang R-L, Chen Ch-Ch, Huang H-L, Chang Ch-G, Chen Ch-F, Chang Ch, Hsieh M-T, Anti-Hepatitis B Virus Effects of Wogonin Isolated from Scutellaria baicalensis, Planta Med. 66 (8): 694-8 (2000 Dec) (18) Kubo M, Matsuda H, Kimura Y, Okuda H, Arichi S, Scutellariae Radix. X. Inhibitory Effects of various on Histamine Release from Rat Peritoneal Mast Cells in Vitro, Chem Pharm Bull. 32 (12): 5051-5054 (1984 Apr) (19) Ryu SH, Ahn BZ, Pack MY, The Cytotoxic Principle of Scutellariae Radix against L1210 Cell, Planta Med. 51 (4): 355 (1985) (20) Li BQ, Fu T, Gong W-H, Dunlop N, Kung H-F, Yan Y, Kang J, Wang JM, The À avonoid baicalin exhibits anti-inÀ ammatory activity by binding to chemokines, Immunopharmacol 49: 295-306 (2000) (21) Chen F, Chan KH, Jiang Y, Kao RYT, Lu HT, Fan KW, Cheng VCC, Tsui WHW, Hung IFN, Lee TSW, Guan Y, Peiris JSM, Yuen KY, In vitro susceptibility of 10 clinical isolates of SARS corona-virus to selected antiviral compounds, J Clin Virol 31: 69-75 (2004) (22) Ma S-C, Du J, But PP-H, Deng X-L, Zhang Y-W, Ooi VE-C, Xu H-X, Lee SH-S, Lee SF, Antiviral Chinese medicinal herbs against respiratory syncytial virus, J Ethnopharmacol 79: 205-211 (2002) (23) The Fourth International Advisory Broad Meeting on Hong Kong Chinese Materia Medica Standards, 20 to 24 October 2008, Hong Kong (24) Kim YH, Jeong DW, Paek IB, Ji HY, Kim YC, Sohn DH, Lee HS, Liquid chromatography with tandem mass spectrometry for the simultaneous determination of baicalein, baicalin, oroxylin A and wogonin in rat plasma, J Chromatogr B Analyt Technol Biomed Life Sci. 844 (2006 Dec 5) (25) Horvath CR, Martos PA, Saxena PK, Identi¿ cation and quanti¿ cation of eight À avones in root and shoot tissues of the medical plant huang-qin (Scutellaria baicalensis GEORGI) using high-performance liquid chromatography with diode array and mass spectrometric detection, J Chromatogr A. 1062 (2): 199-207 (2005 Jan 14)

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