Hepatoprotective Flavonol Glycosides from the Aerial Parts of Rodgersia podophylla

• Abstract
• Material and Methods
• References

Abstract

A new acylated flavonoid, quercetin 3-O-α-L-(5″-O-acetyl)-arabinofuranoside (1), along with six known flavonoids (2 - 7) were isolated from the aerial parts of Rodgersia podophylla. The new flavonoid 1 exhibited 50.1 % hepatoprotective activity at a concentration of 100 μM, and the three known compounds 3, 5 and 6 showed hepatoprotective activities at a concentration of 50 μM (45.7, 50.8 and 57.3 %, respectively) by using the primary cultures of rat hepatocytes injured by H₂O₂.

The rhizomes of Rodgersia podophylla A. Gray (Saxifragaceae) have been used to treat enteritis and bacillary dysentery in China and are also known to have antipyretic and analgesic effects [1]. It has been reported that the rhizomes of this plant contained bergenin, β-peltoboykinolic acid, sterols, monoterpenes, fatty acids and neolignans [1]. During our investigation of hepatoprotective materials from Korean plants, the aerial parts of R. podophylla exhibited hepatoprotective activity in the primary cultures of rat hepatocytes injured by H₂O₂. Therefore, we conducted an activity-guided separation of this plant and isolated the flavonol glycosides 1 - 7 (Fig. [1]) as the active constituents.

Compound 1, yellow powder, gave a pseudomolecular ion peak at m/z = 477.1016 [M + H]⁺ corresponding to a molecular formula of C₂₂H₂₁O₁₂ in the HR-FAB-MS. ¹H-NMR and ¹³C-NMR spectra of 1 showed characteristics of a quercetin moiety with a glycoside. The sugar moiety was assumed to be an α-L-arabinofuranoside, which was supported by comparison of the ¹³C-NMR data of 1 with those of quercetin 3-O-α-L-arabinofuranoside (2) (Table [1]) [2]. Also, the methyl protons of an acetyl group were observed as a singlet peak at δ = 1.89 in the ¹H-NMR and the two carbon peaks belonging to an acetyl group at δ = 20.9 and 170.4 in the ¹³C-NMR spectra. The acetyl group was determined to be attached at the C(5″)-O position of arabinofuranoside based on the downfield-shifted C-5″ signal at δ = 63.8 and the upfield-shifted C-4″ signal at δ = 82.3 in the ¹³C-NMR spectrum [3]. The position of the acetyl group was further confirmed by the cross peaks between δ = 3.72 (H-5″a), 4.05 (H-5″b) and δ = 170.4 (OCOCH₃) in the HMBC spectrum. Therefore, 1 was elucidated as quercetin 3-O-α-L-(5″-O-acetyl)-arabinofuranoside which is isolated for the first time from a natural resource.

The hepatoprotective activities of compounds 1 - 7 were assessed by measuring their effects on the release of glutamic pyruvic transaminase (GPT) from the primary cultures of rat hepatocytes injured by H₂O₂. Compounds 1, 3, 5 and 6 exhibited significant hepatoprotective activities (Table [2]). Compounds 5 and 6 having an α-L-rhamnopyranosyl moiety at the 3-position were more effective than compounds 2 and 4 having an α-L-arabinofuranosyl moiety at the 3-position. Among the quercetin glycosides 1, 2 and 3 having an α-L-arabinofuranosyl moiety at the 3-position, the acylated compounds 1 and 3 were more effective than 2.

Material and Methods

The aerial parts of Rodgersia podophylla were collected from Jinbu, Kangwon province in 2001. A voucher specimen (SNUPC-011) was deposited at the Seoul National University. Silibinin was purchased from Sigma Chemical Co. (St. Louis, USA). Dried materials (2 kg) were extracted with MeOH giving a crude extract (173 g). The MeOH extract was partitioned with n-hexane, CH₂Cl₂ and n-BuOH, successively. The n-BuOH extract (44 g) was subjected to HP-20 gel (Diaion, 400 g) chromatography (10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 100 % acetone, 2 L each) which gave nine fractions and then the fifth fraction (1.8 g) was applied to silica gel (230 - 400 mesh, Merck, 40 g) column chromatography using a CH₂Cl₂-MeOH (7 : 1, 6 : 1, 5 : 1, 4 : 1, 3 : 1, 2 : 1, 1 : 1, 0 : 1, 750 mL each) gradient that afforded eight fractions. The first fraction (320 mg) was subjected to HPLC (J’sphere ODS-H80) with an AcCN-H₂O [17 : 83 (v/v), 2 mL/min] solvent system to provide compounds 1 (5.4 mg, t_R: 35.7 min), 2 (22 mg, t_R: 23.2 min) [2] and 6 (7 mg, t_R: 52.1 min) [4]. From the second fraction (210 mg), compounds 3 (7.8 mg, t_R: 43.1 min) [5], 4 (12.3 mg, t_R: 30.6 min) [6], 5 (13.3 mg, t_R: 35.4 min) and 7 (7.4 mg, t_R: 26.6 min) [4] were isolated by HPLC [YMC-Pack Ph, AcCN-H₂O 17 : 83 (v/v), 2 mL/min].

These compounds were demonstrated to be pure as evidenced by NMR and HPLC analysis (purity > 95 %).

Quercetin 3-O-α-L-(5″-O-acetyl)arabinofuranoside (1): yellow powder; m. p. 186 °C; C₂₂H₂₀O₁₂; HR-FAB-MS: m/z = 477.1016 [M + H]⁺; calcd. for C₂₂H₂₁O₁₂ : 477.1033; [α]_D ²⁰: -96.3^o (c 0.1, MeOH); UV (MeOH): μ_max (log ε) = 257 (3.93), 355 nm (3.82); IR (KBr): λ_max = 3239, 1738, 1653, 1606, 1199, 1008 cm^-1; ¹H-NMR (400 MHz, DMSO-d6): δ = 1.89 (3H, s, OCOCH ₃), 3.62 (2H, brs, H-2″, 3″), 3.72 (1H, dd, J = 10.7, 6.3 Hz, H-5″a), 4.05 (1H, d, J = 10.7 Hz, H-5″b), 4.15 (1H, brs, H-4″), 5.46 (1H, s, H-1″), 6.17 (1H, s, H-6), 6.38 (1H, s, H-8), 6.85 (1H, d, J = 8.4 Hz, H-5′), 7.44 (1H, s, H-2′), 7.46 (1H, d, J = 8.4 Hz, H-6′);¹³C-NMR: Table [1]. Copies of the original spectra are obtainable from the author of correspondence.

Compounds 1, 2 and 6 were hydrolyzed according to the procedure in the literature [7]. The specific optical rotations of arabinose from 1 and 2 are + 226.2^o and + 227.4^o (c 0.1, H₂O), respectively, whereas rhamnose from 6 has + 13.0^o (c 0.1, H₂O).

Details of bioassay on hepatoprotective activity have been described elsewhere [8],[9].

References

• 1 Chin Y -W, Kim J. Four novel neolignans from Rodgersia podophylla .  Tetrahedron Lett. 2004;  45 339-41
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• 2 Lu Y, Foo L Y. Identification and quantification of major polyphenols in apple pomace.  Food Chemistry. 1997;  59 187-94
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• 3 Merfort I. Acylated and other flavonoid glycosides from Arnica chamissonis .  Phytochemistry. 1988;  27 3281-4
  CrossrefPubMedSearch in Google Scholar
• 4 Agrawal P K, Bansak M C. Flavonoid Glycosides. In: Agrawal PK, editor Carbon-13 NMR of Flavonoids. Amsterdam; Elsevier 1989: pp 335-8
  Search in Google Scholar
• 5 Takemoto T, Yoshida S, Kobayashi M, Hasegawa M. Studies on the constituents of Boehmeria tricuspis Makino.  Yakugaku Zasshi. 1974;  94 1597-602
  PubMedSearch in Google Scholar
• 6 Murakami T, Satake T, Hirasawa C, Ikeno Y, Saiki Y, Chen C -M. Chemical and chemotoxonomical studies on Filices.  Yakugaku Zasshi. 1984;  104 142-6
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• 7 Plaza A, Cinco M, Tubaro A, Pizza C, Piacente S. New triterpene glycosides from the stem of Anomospermum grandifolium .  J Nat Prod. 2003;  66 1606-10
  CrossrefPubMedSearch in Google Scholar
• 8 Kanno S, Ishikawa M, Takayanagi M, Takayanagi Y, Sasaki K. Exposure to hydrogen peroxide induces cell death via apoptosis in primary cultured mouse hepatocytes. Biol. Pharm.  Bull.. 1999;  22 1296-1300
  PubMedSearch in Google Scholar
• 9 Reitman S, Frankel S A. A colorimetric method for the determination of serum glutamic pyruvic transaminase. Am. J. Clin.  Pathol.. 1957;  28 56-63
  PubMedSearch in Google Scholar

Prof. Jinwoong Kim

College of Pharmacy

Seoul National University

San 56-1

Shinlim-dong

Kwanak-gu

Seoul 151-742

Korea

Phone: +82-2-880-7853

Fax: +82-2-887-8509

Email: jwkim@snu.ac.kr

References

• 1 Chin Y -W, Kim J. Four novel neolignans from Rodgersia podophylla .  Tetrahedron Lett. 2004;  45 339-41
  CrossrefPubMedSearch in Google Scholar
• 2 Lu Y, Foo L Y. Identification and quantification of major polyphenols in apple pomace.  Food Chemistry. 1997;  59 187-94
  CrossrefPubMedSearch in Google Scholar
• 3 Merfort I. Acylated and other flavonoid glycosides from Arnica chamissonis .  Phytochemistry. 1988;  27 3281-4
  CrossrefPubMedSearch in Google Scholar
• 4 Agrawal P K, Bansak M C. Flavonoid Glycosides. In: Agrawal PK, editor Carbon-13 NMR of Flavonoids. Amsterdam; Elsevier 1989: pp 335-8
  Search in Google Scholar
• 5 Takemoto T, Yoshida S, Kobayashi M, Hasegawa M. Studies on the constituents of Boehmeria tricuspis Makino.  Yakugaku Zasshi. 1974;  94 1597-602
  PubMedSearch in Google Scholar
• 6 Murakami T, Satake T, Hirasawa C, Ikeno Y, Saiki Y, Chen C -M. Chemical and chemotoxonomical studies on Filices.  Yakugaku Zasshi. 1984;  104 142-6
  PubMedSearch in Google Scholar
• 7 Plaza A, Cinco M, Tubaro A, Pizza C, Piacente S. New triterpene glycosides from the stem of Anomospermum grandifolium .  J Nat Prod. 2003;  66 1606-10
  CrossrefPubMedSearch in Google Scholar
• 8 Kanno S, Ishikawa M, Takayanagi M, Takayanagi Y, Sasaki K. Exposure to hydrogen peroxide induces cell death via apoptosis in primary cultured mouse hepatocytes. Biol. Pharm.  Bull.. 1999;  22 1296-1300
  PubMedSearch in Google Scholar
• 9 Reitman S, Frankel S A. A colorimetric method for the determination of serum glutamic pyruvic transaminase. Am. J. Clin.  Pathol.. 1957;  28 56-63
  PubMedSearch in Google Scholar

Prof. Jinwoong Kim

College of Pharmacy

Seoul National University

San 56-1

Shinlim-dong

Kwanak-gu

Seoul 151-742

Korea

Phone: +82-2-880-7853

Fax: +82-2-887-8509

Email: jwkim@snu.ac.kr