Journal of Applied Pharmaceutical Science Vol. 3 (07), pp. 147-155, July, 2013 Available online at http://www.japsonline.com DOI: 10.7324/JAPS.2013.3728 ISSN 2231-3354 Protective and therapeutic activities of Mayodendron igneum Kurz against paracetamol induced liver toxicity in rats and its bioactive constituents Manal H. Shabana1,Fatma A.M. Hashem2, Abdel-Naser Singab3, Sally Khaled2 and Abdel-Razik Farrag4 1Phytochemistry and plant systematic depart. National Research Centre, Cairo, Egypt. 2Pharmacognosy depart. National Research Centre, Cairo, Egypt. 3Pharmacognosy depart. Ain Shams University, Cairo, Egypt. 4Pathology depart. National Research Centre, Cairo, Egypt. ABSTRACT ARTICLE INFO Article history: The total ethanol extract of Mayodendron igneum leaves and its successive extracts ( pet. Ether, chloroform and Received on: 04/06/2013 ethyl acetate combined with ethanol )were examined for their protective and therapeutic effects against Revised on: 29/06/2013 paracetamol induced liver damage in rats. The evaluation was done through measuring the liver function Accepted on: 15/07/2013 enzymes; aspartate and alanine aminotransferases (AST and ALT) as well as cholestatic marker; alkaline Available online: 30/07/2013 phosphatase ( ALP). Liver histopathological analysis were also evaluated. Treatment with the selected extacts significantly decreased AST, ALT and ALP.The biochemical results were in accordance with histopatholical Key words: results. From the successive ethyl acetate extact, three flavonoids were isolated, purified and identified as 6- Mayodendron igneum leaves, methoxyapigenin; fisetin-3-O-β-glucopyranoside and 3-methoxyquercetin-7-O-β-glucopyranoside(6→8”)3”- phenolic constituents, liver methoxykaempferol-7”-O-β-glucopyranoside. Five phytoconstituents were isolated from successive ethanol damage, paracetamol, enzymes, extract and were identified as 4-methoxybenzoic acid; 6,7-dihydroxycoumarin; apigenin-7-O-β- hepatoprotection, histology. neohesperidoside; 6 –methoxyluteolin-7-O-β-neohesperidoside and luteolin-7-O-β-neohesperidoside. Mayodendron leaves extracts resulted in an attractive candidate for the protection and the treatment of liver damage induced by paracetamol.The flavonoids and coumarin present in the plant may be responsible for the pharmacological activity. INTRODUCTION Jacarandic acid, ursolic acid, 2 α-hydroxyursolic acid and 2 α,3 α- dihydroxyurs-12-en-28-oic acid, (Ogura et al., 1977). Tanaecium Mayodendron igneum Kurz, also known as jaroba, a Bolivian plant used in traditional medicine, was reported Radermachera ignea, is a member of family Bignoniaceae which to contain 2 α-hydroxy-12 β-hydroxy-isopimara-8(14),15-diene includes about 750 species of plants in 120 genera within 8 tribes. (diterpene), triterpenes, sterols and fatty acids, (Mitaine-Offer et al., Species within the Bignoniaceae have yielded a number of 2002). phytochemicals especially terpenoids and flavonoids. Aliphatic Work by Jin et al., (2004) led to the isolation hydroxy-ketones compounds have been isolated from the of pentacyclic triterpenoids from leaves of Campsis grandiflora petroleum ether extract of Adenocalymma alliaceum leaves, (Misra that have been characterized by spectroscopic methods as et al., 1989). A Study of Newbouldia laevis, Markhamia oleanolic acid, hederagenin, ursolic acid, tormentic acuminata, Spathodea campanulata and Kigelia africana revealed acid and myrianthic acid. Harborne, (1967) reported that the principal constituents were a homologous series of n- that most species of bignonioids contained flavones rather than alkanes (C -C ), n-alcohols (C -C ) and related carboxylic acids 23 33 18 30 flavonols. 5,6,7,3`,4`-pentahydroxyflavone (6-hydroxy-luteolin) (C -C ), (Gormann et al., 2004). The aqueous ethanol extract 16 36 was identified in leaves of Catalpa bignonioides, C. of Jacaranda caucana yielded β-sitosterol and triterpene acids: . speciosa, C. bungei and Tecoma australis. Subramanian * Corresponding Author and Nair, (1972) isolated from the leaves of Oroxylum Shabana M.H., National Research Centre Phytochemistry & Plant indicum: baicalein, scutellarein and baicalein - 6-glucuronide. Systematic Department, El Behooth St., Dokki Cairo, Egypt. Tel: +202-01001961517 148 Shabana et al. / Journal of Applied Pharmaceutical Science 3 (07); 2013: 147-155 The same authors isolated quercetin, kaempferol, prophylactic and therapeutic activities against paracetamol- kaempferol-3-sophorosides and dihydrokaempferol-7-glucoside induced liver damage in rats. from leaves of Pajanelia longifolia. Tillequin et al., (1977) isolated 5-hydroxy-6,7,4`-trimethoxy flavone from Phylarthrone EXPERIMENTAL madagascariense leaves. Takemura et al., (1995) isolated a Materials and methods flavone from leaves of the Brazilian plant Arrabidaea chica f. Plant material cuprea, its structure was established by spectroscopic analysis. Fresh leaves of Mayodendron igneum Kurz plant Hase et al., (1995) isolated from Millingtonia hortensis 5,7- (Bignoniaceae) growing in Giza Zoo, Giza, Egypt, were collected. dihydroxy-6,4`-dimethoxy flavone (Pectolinaringenin), hispidulin, The plant was identified by Dr. M. El-Gibaly, a plant taxonomist, apigenin7-O-glucuronide, hispidulin 7-O-glucoside and hispidulin and Mrs. Trease Labib, Consultant of Plant Taxonomy at the 7-O-glucuronide methyl ester. Chen et al., (2003) isolated four Ministry of Agriculture and Director of Orman Botanical Garden, flavonoids from the seeds of Oroxylum indicum: chrysin, Giza, Egypt. A voucher specimen has been placed in the baicalein, baicalein-7-O-glucoside, baicalein-7-O-diglucoside herbarium at the Orman Botanical Garden (Voucher number (Oroxylin B) by high-speed counter-current chromatography. M.20). From Mayodendron igneum Kurz bark a new diterpenoid, igeumone, was isolated together with the following Plant extracts flavonoids: quercetin, naringenin and naringin, (Guo et al., 2007). total ethanol extract was prepared from 400 g of air- While Hashem et al., (2007a) isolated four apigenin glycosides dried, powdered leaves of M. igneum by repeatedly extraction in a from the ethyl acetate extract of M. igneum leaves, identified as Soxhlet apparatus with 80% ethanol until exhaustion. The extract apigenin 7-O-glucoside, 6-methoxy apigenin 7-O-glucoside, 6- was filtered, concentrated under reduced pressure at 55°C to methoxy apigenin-7-O-rhamnoglucoside and6-hydroxy apigenin- dryness, yielding 95 g of crude extract, divided to 90g for 7-O-rhamnoglucoside. successive extraction with different solvents and the remaining 5g Many plants in the Bignoniaceae family have been total ethanol extract used for biological study (hepatoprotection). reported to possess anti-inflammatory (Alguacil et al., 2000), Successive fractions were made using 90 g of the crude extract (Munoz-Mingarro et al., 2003) and (Silva et al., 2006), suspended in water and partitioned successively with petroleum antioxidant(Kumar et al., 2005) and (Hashem et al., 2007b), ether (60-80oC), chloroform, and ethyl acetate. The solvents were antimicrobial(Pereira et al., 2006) and (Eyong et al., 2006), evaporated to dryness to yield fractions of 25 g of petroleum ether cytotoxic (Alguacil et al., 2000) and (Marzouk et al., 2006), extract, 19 g of chloroform extract, 10 g of ethyl acetate extract, antimalarial (Onegi et al., 2002) and antimutagenic activities and 30 g of remaining residue considered as successive ethanol (Nakahara et al., 2002). Earlier studies have shown the anti- extract. inflammatory, analgesic and antipyretic activities of 80% ethanol extract of M. igneum Kurz leaves, (Hashem et al., 2007a). Isolation and purification of phytoconstituents In a study of the total alcohol and successive extracts of The successive ethyl acetate fraction was examined by M. igneum leaves against human cancer cell lines, (Hashem et al., paper chromatography (Whatmann No 1) using the solvent 2011), the total ethanol extract demonstrated strong cytotoxic systems: n-butanol; acetic acid; water (3:1:1) (S1), 15% acetic acid activity against the cervix cancer cell line (IC50= 0.60 μg/mL), (S2) and n-butanol; acetic acid; water (4:1:5 upper phase) (S3). breast cancer cell line (IC50=0.54 μg/mL) and liver cancer cell Chromatograms were visualized under UV, UV & NH3 and UV & line (IC50=0.87 μg/mL) when compared to doxorubicin AlCl3. The presence of the detected spots, their Rf values and their (anticancer drug). colours were recorded. Isolation of detected compounds were Antioxidant activity of total ethanol extract and carried out by preparative paper chromatography using Whatmann successive fractions of M. igneum was measured by electron spin No.3MM sheets using S3 solvent system. The eluted substances resonance (ESR) and using DPPH as the stable free radical, were further purified by repetitive chromatography on Sephadex (Hashem et al., 2012). The strongest inhibition of DPPH stable LH-20 column and eluted with methanol to yield compounds 1-3. free radicals was exhibited by the successive ethanol fraction The successive ethyl alcohol fraction of M. igneum was (equal to 100% of free radical scavenging activity relative to fractionated over a polyamide 6S column. The elution process ascorbic acid), followed by ethyl acetate fraction which had occurred in a gradient mode using water/methanol mixtures of 64.96% of the antioxidant activity of ascorbic acid. The petroleum decreasing polarities as eluent. Fractions of similar PC profiles in ether and chloroform fractions had 12.46% and 0%, respectively, S1 and S2 were