Effects of Curcuma Comosa Extracts on Hepatic Cytochrome P450 Activities in Rats
Total Page：16
File Type：pdf, Size：1020Kb
Original Article 1 EFFECTS OF CURCUMA COMOSA EXTRACTS ON HEPATIC CYTOCHROME P450 ACTIVITIES IN RATS Chonticha Kittichanun1, Laddawal Phivthong-ngam2, Nuansri Niwattisaiwong1, Pawinee Piyachaturawat3, Apichart Suksamran4, Khemchart Apipalakul5 and Somsong Lawanprasert1,* 1Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok, Thailand 2Faculty of Medicine, Srinakharinwirot University, Bangkok, Thailand 3Faculty of Science, Mahidol University, Bangkok, Thailand 4Faculty of Science, Ramkamhang University, Bangkok, Thailand 5Faculty of Dentistry, Chulalongkorn University, Bangkok, Thailand ABSTRACT: Curcuma comosa Roxb. (Zingiberaceae) is an indigenous plant of Thailand. The rhizome of this plant has been widely used in Thai traditional medicine for treatment of abnormal uterine symptoms. The purpose of this study was to investigate effects of C. comosa hexane extract and ethanolic extract on hepatic cytochrome P450 (CYP) in rats. Fifty male Wistar rats were randomly divided into 5 groups (10 rats/group). Rats were given orally with C. comosa hexane extract or ethanolic extract (250 and 500 mg/kg/day) or corn oil in a control group for 30 consecutive days. At the end of the treatment, rats were sacrificed and liver microsomes were prepared. Hepatic microsomal total CYP content and CYP activity were determined. The results showed that both C. comosa hexane and ethanolic extracts at 250 mg/kg/day caused a significant increase of total CYP contents and the activity of CYP1A1. Lower dose of both extracts (250 mg/kg/day) caused a more increase of CYP1A1 activity than the higher dose (500 mg/kg/day). Also, both C. comosa hexane and ethanolic extracts caused a dose-dependent increase of CYP2B1/2B2 activities and the increase was higher in the hexane extract group than the ethanolic extract group. Hexane and ethanolic extracts of C. comosa did not affect CYP1A2, CYP2E1 and CYP3A activities. These results indicated the possibilities of C. comosa hexane and ethanolic extracts regarding herb-drug interactions and the increase risk of toxicity, mutagenesis and/or carcinogenesis from drugs or compounds that are metabolized or bioactivated via CYP1A1 and CYP2B1/2B2 Keywords: Curcuma comosa, cytochrome P450, drug interaction INTRODUCTION: Curcuma comosa Roxb. is a estrogenic-like effect4,5), anti-inflammatory effect6). plant in family Zingiberaceae. Rhizomes of C. Other pharmacological effects of C. comosa have comosa have been used extensively in Thai also been documented such as choleretic effect7) traditional medicine as an anti-inflammatory which is the effect of phloracetophenone glucoside2), agent particularly for the treatment of female cholesterol lowering effect7-9) and nematocidal effect1). postpartum uterine inflammation. Rhizomes of C. Cytochrome P450 (CYP), the phase I enzyme comosa comprise various chemical compounds system in the liver, normally plays a key role in include diarrylheptanoids such as trans-1,7- metabolism of most currently used medicines. diphenyl-5-hydroxy-1-heptene; trans-1,7-diphenyl Inhibition and /or induction of CYP by xenobiotics -6-hepten-3-one-5-ol; trans-1,7-diphenyl-3-acetoxy- are characterized as one of the most important 10) 6-heptene; trans-1,7-diphenyl-6-heptene-3-one; etiology of drug-drug interactions . Therefore, on trans,trans-1,7-diphenyl-1,3-heptadien-5-ol; trans, the developmental process of C. comosa for trans-1,7-diphenyl-4,6-heptadien-3-one1), 1,7- medicinal purpose, studying an effect of C. diphenyl-5-hydroxy-(1E)-1-heptene; 5-hydroxy-7-(4- comosa on hepatic CYP is required so as to hydroxyphenyl)-1-phenyl-(1E)-1-heptene; 7-(3,4- investigate the possibilities of herb-drug interactions hydroxyphenyl-5-hydroxy-1-phenyl(1E)-1-heptene2) if C. comosa is administered concomitantly with and phloracetophenone glucoside such as 4,6- other medicines. 2) dihydroxy-2-O-(β-D-glucopyranosyl) acetophe-none . It has been known that CYP plays a role in Many studies have revealed pharmacological carcinogenesis. Metabolic activation of procarcinogens effects of C. comosa, such as uterotrophic effect3), is often catalyzed by CYP including CYP1A1, 1A2, ∗To whom correspondence should be addressed. E-mail: [email protected] Tel./Fax. +66 2218 8324 J Health Res 2010, 24(1): 1-6 2 Original Article 2B6, 2E1 and 3A411). For example, CYP1A1 was approved by the Ethic Committee of the activates polycyclic aromatic hydrocarbons (PAHs) Faculty of Pharmaceutical Sciences, Chulalongkorn to biological reactive metabolites that interact with University (Approval no. 97/2007). DNA resulting in carcinogenesis. Induction of C. comosa some CYPs is a risk factor in several cancers since The rhizomes of C. comosa were collected from these enzymes can convert procarcinogens to Nakhon Pathom, Thailand. The dried rhizome carcinogens. Thus, effect of C. comosa on CYP powder (30 kg) was extracted with n-hexane in a isoforms involving carcinogenic bioactivation is Soxhlet extractor to give a pale browish viscous oil needed to be investigated. The induction effect of (1.01 kg), giving the percent yield of 3.37%. The C. comosa on CYP isoforms that bioactivate pro- marc was subsequently extracted with 95% carcinogens will indicate an increase risk of ethanol, the ethanolic fraction was dried under carcinogens by C. comosa. On the other hand, C. vacuum in rotary evaporator and dried again with comosa may be classified as an anti-carcinogenic high vacuum pump to give a dark reddish-brown agent if it inhibits CYP isoforms which bioactivate viscous oil (1.30 kg), giving the percent yield of carcinogens. 4.33%. Hexane and ethanolic extracts of C. In this study, effects of C. comosa hexane comosa were characterized by thin layer chromato- extract and ethanolic extract were ex vivo examined graphy. The major constituent of hexane extract on rat hepatic microsomal CYPs especially the was 1,7-diphenyl-5-hydroxy-(1E)-1-heptene. The pivotal isoforms involved in drug metabolism and major constituents of ethanolic extract were 5- activation of mutagens and/or carcinogens. These hydroxy-7-(4-hydroxyphenyl)-1-phenyl-(1E)-1- CYP isoforms include CYP1A1, 1A2, 2B1/2B2, 2E1 heptene and 7-(3,4-dihydroxyphenyl)-5-hydroxy- and 3A. 1-phenyl-(1E)-1-heptene. MATERIALS AND METHODS: Preparation of microsomes and CYP activity Experimental animals assays Fifty adult male Wistar rats of body weight Rat liver microsomes were prepared according between 250-300 g were obtained from the to the method described by Lake BG12). Liver National Laboratory Animal Center, Mahidol microsomal protein concentrations were determined University, Salaya, Nakorn Pathom, Thailand. according to the method of Lowry OH, et al13). Animals were housed, two per cage at the Faculty Total CYP content was determined according to of Medicine, Srinakharinwirot University, Bangkok the method of Omura T and Sato R14). Rate of and acclimatized for at least a week prior to the hepatic microsomal alkoxyresorufin O-dealkylation experiment. All animals were in a controlled was determined according to the method of Burke humidify room at a constant temperature of 25ºC MD and Mayer RT15) and Lubet RA, et al16). and maintained on a 12-hour alternate light-dark Benzyloxyresorufin and pentoxyresorufin were used cycle. They were allowed free access to food (C.P. as selective substrates of CYP2B1/2B2. Ethoxy- company, Thailand) and drinking water. Rats resorufin and methoxyresorufin were used as were randomly divided into 5 groups. Each group selective substrates of CYP1A1 and CYP1A2, comprised 10 rats. The experimental groups were respectively. The catalytic activity of CYP2E1 was received C. comosa hexane extract or ethanolic determined based on the rate of hepatic microsomal extract orally at doses of 250 and 500 mg/kg/day aniline 4-hydroxylation, according to the method whereas rats in the control group were given corn of Schenkman JB, et al17). Aniline hydrochloride oil at 1 ml/kg/day for 30 consecutive days. At the was used as a selective substrate in this reaction. end of the treatment, animals were fasted for 12 Rate of hepatic microsomal erythromycin N-demethyl- hours before anesthesized with diethyl ether and ation was determined using the method of Nash sacrificed by cervical dislocation. The protocol of T18) and Friedli GL19). Erythromycin stearate was animal housing and treatment used in this study used as a selective substrate of CYP3A. J Health Res 2010, 24(1): 1-6 Original Article 3 RESULTS: Total CYP content Effect of C. comosa extracts on rat hepatic microsomal total CYP content and CYP 1 0.9 * activities * 0.8 C. comosa hexane extract caused significant * 0.7 increase of hepatic total CYP content while C. 0.6 comosa ethanolic extract increased total CYP 0.5 0.4 content only at the dose of 250 mg/kg/day 0.3 (Figure 1). (nmol/mg protein) (nmol/mg Total CYP content 0.2 CYP1A1 activity which represented by the rate 0.1 of ethoxyresorufin O-dealkylation (EROD) reaction 0 control C. comosa 1 C. comosa 2 C. comosa 3 C. comosa 4 was significantly increased in hepatic microsomes of rats treated with both doses of C. comosa Figure 1 Effect of C. comosa hexane extract and ethanolic extract on total CYP contents. Rats were administered orally with 1 ml/kg/day hexane extract (250 and 500 mg/kg/day) and C. of corn oil (Control), 250 and 500 mg/kg/day of the C. comosa comosa ethanolic extract only at the dose of 250 hexane extract (C. comosa 1 and C. comosa 2), 250 and 500 mg/kg/day of the C. comosa ethanolic extract (C. comosa 3 and C. mg/kg/day as compared to the control group. The comosa 4) for 30 days. The individual bar graph represented mean ± SEM (n = 9-10). lower dose (250 mg/kg/day) of both hexane and * p < 0.05; C. comosa group vs control group. ethanolic extracts caused a significant higher increase of CYP1A1 activity than the higher dose EROD (500 mg/kg/day) (Figure 2).