Hepatoprotective Phenylpropanoids from Scrophularia buergeriana Roots against CCl₄-Induced Toxicity: Action Mechanism and Structure-Activity Relationship

Abstract

Phenylpropanoids isolated from the roots of Scrophularia buergeriana MIQ. (Scrophulariaceae) protected primary cultures of rat hepatocytes from toxicity induced by carbon tetrachloride (CCl₄). In this report, we show that two of these phenylpropanoids, 4-O-E-p-methoxycinnamoyl-α-L-rhamnopyranoside ester (1) and p-methoxycinnamic acid (3) have significant hepatoprotective activity; another phenylpropanoid used for comparison, isoferulic acid (11), was equally active. To determine the mechanism(s) by which these three phenylpropanoids exerted their hepatoprotective activity, we measured activities of enzymes involved in the glutathione (GSH) redox system and assayed the level of hepatic mitochondrial GSH. The GSH levels in primary cultures of rat hepatocytes were significantly reduced with CCl₄ insult, but were significantly preserved by the treatment with these three phenylpropanoids. The activities of glutathione disulfide reductase and glutathione-S-transferase which normally decrease in CCl₄-injured rat hepatocytes were significantly preserved by the treatment with these three phenylpropanoids. In addition, in CCl₄-injured rat hepatocytes, the increased formation of malondialdehyde, a byproduct of lipid peroxidation, was reduced by the treatment with these phenylpropanoids. We determined the essential structural moiety within these three phenylpropanoids needed to exert hepatoprotective activity. The α,β-unsaturated ester moiety seemed to be essential for exerting hepatoprotective activity.

Abbreviations

CCl₄:carbon tetrachloride

GSH:glutathione

MDA:malondialdehyde

GST:glutathione-S-transferase

GPx:glutathione peroxidase

GR:glutathione disulfide reductase

Scrophularia buergeriana - Scrophulariaceae - phenylpropanoids - carbon tetrachloride - hepatoprotective activity - structure-activity relationship

References

• 1 Zhu Y -P. Chinese Materia Medica: chemistry, pharmacology and applications. Amsterdam; Harwood Academic Publishers 1998: 161-2
• 2 Yamamoto A, Miyase T, Ueno A, Maeda T. Scrophulasaponins II-IV, new saikosaponin homologs from Scrophularia kakudensis FRANCH.  Chemical and Pharmaceutical Bulletin. 1993;  41 1780-3
• 3 Kajimoto T, Hidaka M, Shoyama K, Nohara T. Iridoids from Scrophularia ninpoenesis .  Phytochemistry. 1989;  28 2701-4
• 4 Calis I, Gross G, Sticher O. Phenylpropanoid glycosides isolated from Scrophularia scopolii .  Phytochemisty. 1987;  26 2057-61
• 5 Li Y M, Jiang S H, Gao W Y, Zhu D Y. Iridoid glycosides from Scrophularia ningpoensis .  Phytochemistry. 1999;  50 101-4
• 6 Kim S R, Kim Y C. Neuroprotective Phenylpropanoid esters of rhamnose isolated from roots of Scrophularia buergeriana .  Phytochemistry. 2000;  54 503-9
• 7 Fan W Z, Tezuka Y, Xiong Q B, Hattori M, Namba T, Kadota S. Apocynins A-D - New phenylpropanoid-substituted flavan-3-ols isolated from leaves of Apocynum venetum (Luobuma-Ye).  Chemical and Pharmaceutical Bulletin. 1999;  47 1049-50
• 8 Li J X, Shi Q, Xiong Q B, Prasain J K, Tezuka Y, Hareyama T, Wang Z T, Tanaka K, Namba T, Kadota S. Tribulusamide A and B, new hepatoprotective lignanamides from the fruits of Tribulus terrestris: indications of cytoprotective activity in murine hepatocyte culture.  Planta Medica. 1998;  64 628-31
• 9 Xiong Q, Fan W, Tezuka Y, Adnyana I K, Stampoulis P, Hattori M, Namba T, Kadota S. Hepatoprotective effect of Apocynum venetum and its active constituents.  Planta Medica. 2000;  66 127-33
• 10 Meister A, Anderson M E. Glutathione.  Annual Review of Biochemistry. 1983;  52 711-60
• 11 Berry M N, Friend D S. High yields preparation of isolated rat liver parenchymal cells.  Journal of Cell Biology. 1984;  43 506-20
• 12 Kim S Y, Kim H P, Huh H, Kim Y C. Antihepatotoxic zeaxanthins from the fruits of Lycium chinense .  Archives of Pharmacal Research. 1997;  20 529-32
• 13 Reitman S, Frankel S. A colorimetric method for the determination of glutamic oxaloacetic and glutamic pyruvic transaminase.  American Journal of Clinical Pathology. 1957;  28 56-63
• 14 Gibson G G, Skelf P. Techniques and experiments illustrating drug metabolism. In: Gibson GG, Skelf P, editors Introduction to Drug Metabolism. New York; Chapman and Hall 1988: 239-71
• 15 Tietz F. Enzymatic method for quantitative determination of nanogram amounts of total and oxidized glutathione.  Analytical Biochemistry. 1969;  27 502-22
• 16 Habig W H, Pabst M J, Jakoby W B. Glutathione S-transferases. The first enzymatic step in mercapturic acid formation.  Journal of Biological Chemistry. 1974;  249 7130-5
• 17 Flohe L, Gunzler W A. Assays of glutathione peroxidase.  Method in Enzymology. 1984;  105 114-21
• 18 Carlberg I, Mannervik B. Purification and characterization of the flavoenzyme glutathione reductase from rat liver.  Journal of Biological Chemistry. 1975;  250 5475-80
• 19 Ohkawa H, Ohishi N, Yagi K. Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction.  Analytical Biochemistry. 1979;  95 351-8
• 20 Lowry O, Rosebrough H, Farr A, Randall R. Protein measurement with the Folin Phenol reagent.  Journal of Biological Chemistry. 1951;  193 265-75
• 21 Hikino H, Kiso Y, Wagner H, Fiebig M. Antihepatotoxic actions of flavonolignans from Silybum marianum fruits.  Planta Medica. 1984;  50 248-50
• 22 Slater T F. Free-radical mechanisms in tissue injury.  Biochemical Journal. 1984;  222 1-15
• 23 Ip S P, Poon M KT, Che C T, Ng K H, Kong C Y, Ko K M. Schisandrin B protects against carbon tetrachloride toxicity by enhancing the mitochondrial glutathione redox status in mouse liver.  Free Radical Bioloy and Medicine. 1996;  21 709-12
• 24 Boyer T D, Vessey D A, Holcomb C, Saley N. Studies of the relationship between the catalytic activity and biding of non-substrate ligands by the glutathione-S-transferase.  Biochemical Journal. 1984;  217 179-85
• 25 Mosialou E, Morgenstern R. Activity of rat microsomal glutathione transferase toward products of lipid peroxidation and studies of the effect of inhibitors on glutathione-dependent protection against lipid peroxidation.  Archives of Biochemistry and Biophysics. 1989;  275 289-94
• 26 Mosialou E, Ekstroem G, Adang A EP, Morgenstern R. Evidence that rat liver microsomal glutathione transferase is responsible for gluthathione-dependent protection against lipid peroxidation.  Biochemical Pharmacology. 1993;  45 1645-51