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Toxicity, Cytotoxicity and Biological Activities of Seeds of Carapa Procera (DC), a Native Oil Tree

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Toxicity, Cytotoxicity and Biological Activities of Seeds of Carapa Procera (DC), a Native Oil Tree Available online at http://www.ifgdg.org Int. J. Biol. Chem. Sci. 13(1): 49-62, February 2019 ISSN 1997-342X (Online), ISSN 1991-8631 (Print) Original Paper http://ajol.info/index.php/ijbcs http://indexmedicus.afro.who.int Toxicity, cytotoxicity and biological activities of seeds of Carapa procera (DC), a native oil tree Balé BAYALA1*, Brahima SOW1, Vinsoun MILLOGO2, Youssouf OUATTARA1 et 3 Hamidou Hamadou TAMBOURA 1Laboratoire de Physiologie Animale. UFR/Sciences de la Vie et de la Terre. Université Ouaga I Pr. Joseph KI-ZERBO. 03 BP 7021 Ouagadougou 03 (BURKINA FASO). 2 Laboratoire de Recherche et d’Enseignement en Santé et Biotechnologie Animales/Institut du Développement Rural/Université Polytechnique de Bobo-Dioulasso 01 BP 1091 Bobo-Dioulasso 01 (BURKINA FASO). 3Institut de l’Environnement et de Recherches Agricoles (INERA)/CNRST). Département Productions Animales. UER/Biologie et Santé Animale. 01 BP 476 Ouagadougou 01 (BURKINA FASO). *Corresponding author; E-mail: [email protected], Tel: (+226) 78823838. ACKNOWLEDGMENTS The present work was done with the support from DANIDA, Danish International Development Agency through QUALITREE project. This work was also supported by IAEA through the BKF5011-5014 project. ABSTRACT Carapa procera is a native tree which seeds are intensively used for oil extraction. So this study deals with the identification of secondary metabolites, the toxicity and cytotoxicity of C. procera hydroalcohol extracts and their estrogenic and androgenic activities. The chemical constituents of C. procera extracts were determined by Thin Layer Chromatography (TLC) methods. The toxicity and cytotoxicity were respectively evaluated by the OECD Guideline 425 “Up and Down procedure” (UDP) and the brine shrimp lethality test. Estrogenic and androgenic activities were evaluated by uterotrophic and Hershberger tests and also by in vitro tests, with the yeast assay and MVLN luceferase assay. The MeOH/H2O extracts of C. procera seeds revealed the presence of steroidal glycosides, triterpene, flavonoids and polyphenols. The DCM fraction revealed the presence of triterpene esters & steroid and carotenoids. The MeOH/H2O has, by in vivo tests, slight toxic effects with mice and shows a lethal effect against the crustacean larvae by in vitro test. The extracts of C. procera seeds also show, by in vivo and in vitro tests an estrogenic and anti-androgenic activities. The results show with the presence of flavonoids, polyphenols and glycosides steroids estrogenic and anti-androgenic activities. The seeds of C. procera contain some phytochemicals, which can disrupt or reinforce endocrine function. © 2019 International Formulae Group. All rights reserved. Keywords: Carapa procera, native oil tree, seeds, toxicity, estrogenic, antiandrogenic. INTRODUCTION (Ouédraogo et al., 2013; Tiétiambou et al., In Burkina Faso, many native tree 2016). species are used for their potential composition Carapa procera (C. procera) is an in oil. For rural communities, these native oils oleaginous plant belongs to Meliaceae family increase and diversify their livelihoods. which is very appreciated by traditional healers Women traditionally extract oil from native to treat various diseases. The oil from C. tree from seeds and this oil is an opportunity procera seeds is known to be used as food, for diversification and income generation cosmetics, veterinary medicine, insecticide and repellent properties (Guillemot, 2004). © 2019 International Formulae Group. All rights reserved. 4032-IJBCS DOI: https://dx.doi.org/10.4314/ijbcs.v13i1.5 B. BAYALA et al. / Int. J. Biol. Chem. Sci. 13(1) : 49-62, 2019 Regarding the importance of oil from C. Animal material procera seeds, there is a need to improve The NMRI mice, 27 days old were extraction method which is still unfortunately obtained from the animal house of Ouaga I Pr. traditional. The rural community extraction is Joseph KI-ZERBO University. The room not able to purify the oil and several toxic temperature was maintained at (22 ± 3) °C with substances have been reported, whereas it is the 12 h light/12 h dark cycle and humidity at known that some oleaginous plants contain 50 ± 10%.The animals were fed with industrial hormone-like substances likely to possess an pellets with 29% protein and have free access effect on animal reproduction function. Many to drinking water. All tests included in the phytochemical studies on the seeds of C. current work were performed according to the procera revealed the presence of flavonoids, protocols already approved by the Department phytosterols, triterpenes and tocopherols of Animal Physiology of Ouaga I Pr. Joseph (Tindo et al., 2012; Goubgou, 2013). Some of KI-ZERBO University and met the these hormone-like substances can bind to the international standards of animals study hormone receptor and interfere with (Zimmermann, 1983). endogenous hormone of human and animal reproduction function. Several works suggest Chemical products the beneficial effects of phytoestrogen like Testosterone propionate (Purity 97%) protection against breast and prostate cancer. and the 17-béta-Estradiol (Purity 97%) were On the other hand, these compounds can also purchased from the Sigma Chemical Co. (St. act as endocrine disrupters which could affect Louis, MO, USA).The DMSO 1% was served the endocrine system and may cause as dilution liquid for the preparation of development and reproductive disturbances. different doses. All substances were shipped For these reasons, there is a need to and stored in glass containers at room characterize the hormonal potency of natural temperature. All solvents were analytical compounds with the ability to bind to the grade. estrogen (Diel et al., 2002). Despite the highly sought and used of C. Extraction and isolation procera seeds for oil production very few The dried and committed seeds of C. studies have been interesting in the toxicity and procera (100 g) were successively extracted by the biological effects of these seeds extracts on maceration with dichloromethane (8 L) and animal reproductive function. methanol (10 L) at room temperature during 48 Due to the use of C. procera seeds for h. The extracts were concentrated to dryness oil, the present study investigated the under reduced pressure at 40 °C to yield screening of phytochemicals in this seeds, the dichloromethane (DCM) crude (3.5 g) and a cytotoxicity, the acute toxicity and the effect of the seeds extracts on male and female methanol (MeOH) residue (100 g). Both the reproductive tract by in vivo and in vitro tests. extracts were dried and stored at 4 °C until used. DCM crude (1.5 g) was fractionated by MATERIALS AND METHODS column chromatography over silica gel (40-63 Plant material m, Merck), eluted gradient (from 5/5 to 0/10, C. procera seeds were collected in 2015 µ between 6 and 10 am in West part of Burkina v/v), with / ethyl acetate gradient (from 5/5 to Faso. The seeds were identified by the 0/10, v/v) and afforded thirteen fractions (A to Herbarium of Ouaga I Pr. Joseph KI-ZERBO M). Fraction B was purified by column University where a voucher sample was chromatography over silica gel (40-63 µm, preserved for reference under number 6203. Merck), eluting with n- hexane/ethyl acetate gradient (10/0 to 7/3, v/v) and yielded the compound 1 (25 mg) and three 2 50 50 B. BAYALA et al. / Int. J. Biol. Chem. Sci. 13(1) : 49-62, 2019 other fractions. Fraction D was subjected to periodically during 1, 24, 48 and 72h. The solid phase extraction (SPE) over silica gel (40- lethal dose (LD50) was estimated according to 63 µm, Merck) to yield compound 2 (6 mg) by the method described by Litchfield and using n-hexane/ethyl acetate (0/10 to with n- Wilcoxon (1949). hexane/CH2Cl2 CH2Cl27/3, v/v) as gradient. Subchronic toxicity test The MeOH residue (36 g) was Two groups of 10 mice, each containing suspended in water (600 mL) during 20 hours, an equal number of both male and female, and partitioned with CH Cl (3x200 mL) and 2 2 received intraperitonealy during 28 fraction (0.10 g) and an ethyl acetate fraction consecutive days respectively distilled water (1.03 g). Ethyl acetate fraction was fractionated (control groupe I) and 82 mg/kg (Group II). by column chromatography over Sephadex Mice were weighed every day before extracts LH-20 (Pharmacia) as stationary phase and administration and after a follow-up of 2 hours MeOH as eluent and afforded eight fractions takes place to note signs of toxicity caused by (A.1 to A.8). Fraction A.3 was re- the extract. Each mouse was marked with a chromatographied on silica gel (40-63 µm, unique identification number and behavior was Merck) with n-hexane/ethyl acetate (6/4 to observed daily during the trial period. One day 0/10, v/v) as gradient and ethyl acetate (3x200 after the last administration, all the mice were mL) to yield a CH2Cl2 yielded the compound autopsied and the organs (lungs, heart, spleen, 3 (27 mg). On the other hand, the screening of liver and kidney) were removed and weighed the chemical constituents was carried out with to note the necrotic signs. the extracts of the C. procera seeds, using chemical reagents and thin layer Brine shrimp lethality assay chromatography (TLC) methods according to The bioactivity of the extracts was the methodology suggested by Wagner and monitored by the brine shrimp lethality test (Meyer et al., 1982). Samples were dissolved Bladt (2009). in DMSO and diluted with artificial sea salt water so that final concentration of DMSO did Acute toxicity test not exceed 0.05%. Fifty microliters of sea salt An intraperitoneal study for water were placed in all the wells of the 96-well determining LD50 was performed according to microtiter plate. Fifty microliters (1, 10, 100, the OECD Guideline 425 “Up and Down 1000 μg/ml) seeds extracts were made in procedure” (UDP) (Bruce, 1985; OECD, triplicate.
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