American Journal of Organic Chemistry

p-ISSN: 2163-1271    e-ISSN: 2163-1301

2016;  6(3): 81-85



Isolation of (-) - Epicatechin from Trichilia emetica Whole Seeds

Abdullahi Usman1, 2, Vera Thoss1, Mohammed Nur-e-Alam1

1School of Chemistry, Bangor University, United Kingdom

2Department of Chemistry, Nasarawa State University Keffi, Nigeria

Correspondence to: Abdullahi Usman, School of Chemistry, Bangor University, United Kingdom.


Copyright © 2016 Scientific & Academic Publishing. All Rights Reserved.

This work is licensed under the Creative Commons Attribution International License (CC BY).


Trichilia emetica whole seeds were refluxed in water for twenty minutes and the extract subjected to column chromatography and preparative-TLC. The mass spectrum of the isolated compound in positive ion mode showed a parent molecular ion peak at m/z 291 which corresponds to the molecular formula C15H15O6. The structure was established on the basis of nuclear magnetic resonance (NMR), and Infrared (IR) spectroscopy, ultraviolet-visible (UV-vis) spectrophotometry as well as by the comparison of the data reported in the literature. It was concluded that the compound isolated is a biologically important polyphenol, (–)- epicatechin.

Keywords: Trichilia emetica, Meliaceae, Epicatechin, Spectroscopy, Chromatography

Cite this paper: Abdullahi Usman, Vera Thoss, Mohammed Nur-e-Alam, Isolation of (-) - Epicatechin from Trichilia emetica Whole Seeds, American Journal of Organic Chemistry, Vol. 6 No. 3, 2016, pp. 81-85. doi: 10.5923/j.ajoc.20160603.01.

1. Introduction

Trichilia emetica (Vahl), also known as Natal or Woodland Mahogany, belongs to the Meliaceae family. It is an evergreen tree reaching 35 metres in height and is widely distributed in the tropical and sub-tropical regions of Africa. They are propagated by cuttings and regenerate naturally by root suckers, and seeds [1, 2]. In Africa, different traditions use different parts of this plant in treating several diseases and disorders. In South Africa, the decoction of the root and stem bark is used as an emetic and also as a remedy for cold, pneumonia, and intestinal disorder [3]. In Mali, the powdered root mixed with milk is used as a purgative and a poison antidote [4, 5]. In Senegal, the macerated root bark is used for epilepsy and leprosy while the leaves are taken against blennorrhoea. Finally, in Zimbabwe, the bark is used to induce abortion, and in Nigeria the leaves are used for treating syphilis [1, 2, 3].
The extensive traditional uses of this plant have prompted researchers to screen the solvent extracts for a wide range of biological activities, such as anti-plasmodia [6], antimicrobial [7], anti-oxidant [8], anti-inflammatory [9], anti-schistosomal [10], anti-trypanosomal [11], anticonvulsant [12] and anticancer activity [13]. The aim of this study was to isolate bioactive compounds responsible for these activities. This work reports isolation of the subject compound for the first time from the seeds of T. emetica.

2. Materials and Methods

Optical rotations were measured in methanol solution on a ADP 440+ polarimeter. The melting point was determined by a Stuart instrument and is uncorrected. The 1H and 13C NMR spectra were recorded on a Bruker Avance (400 MHz) spectrometer with internal references of δH 3.31 and δC 49.0 ppm for CD3OD using TMS (Tetramethylsilane) as an internal standard. A Thermo Instruments HPLC system mass spectrometer with an electrospray ionization (ESI) source was used for recording of the mass and UV spectra. Column chromatography (CC) was performed on Fluorochem silica gel (60Å). Thin layer chromatography (TLC) and Preparative thin layer chromatography (PTLC) were conducted on precoated E. Merck TLC silica gel 60 F254 glass plates, and visualization of the compound was done using UV lamp UVL-14 EL hand held 220V 50Hz 4W 254nm white light by UVP.

2.1. Collection of Plant Materials

T. emetica seeds were collected from Kumasi, Ghana, in February 2013 and identified by botanist Mr Martin A. Arkoh of Kwame Nkruma University of Science and Technology, Kumasi, Ghana. A voucher specimen TBG-2014-1 was deposited at the herbarium of Treborth Botanical Garden Bangor, UK.

2.2. Preparation of the Plant Materials

2.2.1. Extraction
Seeds (100 g) were refluxed in water (500 ml) at 100°C for 20 minutes and filtered. The filtrate was concentrated to dryness to give 14.23 g dark gummy extract, which was suspended in water. The aqueous extract was sequentially extracted with chloroform and ethyl acetate (EtOAc) at different pH values (pH 3, pH 7 and pH 10), and the pH was adjusted by the addition of 2 M HCl and 2 M NaOH. The TLC and 1H NMR profiles of all the fractions were assessed. The chloroform and aqueous fractions showed poor TLC and 1H NMR profiles and were discarded, while EtOAc fractions at different pH values showed the same major spots under UV lamp and were investigated further.
2.2.2. Isolation
Purification of the ethyl acetate extract (1.50 g) was carried out by column chromatography in a polarity gradient manner. Hexane and EtOAc were used as the eluents at gradient mixtures from 100% hexane: 0% EtOAc to 0% hexane: 100% EtOAc. This process was followed by extraction with 100% methanol. Forty three (43) fractions were collected, and based on their TLC profiles, were pooled together. Fractions 24-27 (97 mg), obtained with 40% hexane in EtOAc, were combined and evaporated to dryness under vacuum at 40°C. The combined extract was further purified by PTLC. The plate was developed using Hexane – EtOAc – MeOH (5:4:1, v/v). The main band with light purple colour visualized under UV lamp was scraped from the plate and eluted with the same developing solvents, yielding epicatechin (23 mg).

3. Results and Discussion

The compound was obtained as a milky white amorphous powder, with the melting point between 238-239°C and optical rotation of (MeOH: c=0.33), which were comparable to literature values [14]. The compound’s molecular formula of C15H15O6 was established on the basis of ESI-HRMS at m/z 291.0867[M + H]+ (Calcd for 291.0869) (Figure 1). The IR spectrum showed a broad band at 3293 cm-1 region corresponding to phenolic and alcoholic O-H stretching. Other bands at 2910 and 1607 cm-1 were due to saturated C-H stretching and aromatic C=C stretching, respectively. Additionally, the UV spectrum showed absorption peaks at 230 and 279 nm, consistant with conjugated transitions arising from the aromatic rings. These results were similar to those reported in the literature [15].
Figure 1. Mass spectrum of epicatechin
The 1H NMR spectrum (Figure 2), showed that proton H-3 was split by H-2 and H-4, resulting in a multiplet at δH 4.19. The protons at H-4, were split by H-3, giving a doublet of doublet at δH 2.73 and δH 2.86. The position of the H-2 chemical shift (δH 4.83) suggests that the flavan structure possesses the correct cis-2,3 stereochemistry, this was supported by the small value for the coupling (< 1 Hz) between the H-2 and H-3 protons, which appeared as a broad singlet at H-2 (δH 4.83) [16, 17]. These signals are attributed to ring C.
Figure 2. The spectrum of epicatechin
The distinctive signals of the B-ring aromatic protons are of the ABX-type. The methine proton H-2’ was split by H-6’ in a meta position resulting in a doublet at δH 6.99 (d, 1.7 Hz), H-5’ was split by H-6’ in an ortho position yielding a doublet at δH 6.77 (d, 8.2 Hz) and H-6’ was split by H-2’ and H-5’ given a doublet of doublet at δH 6.81 (dd, 1.7, 8.2 Hz).
The ring A aromatic protons also showed H-6 and H-8 coupling, with each yielding a doublet at δH 5.93 and 5.96 (d, 2.3 Hz, each). The basic structure of this compound was therefore deduced as 3, 3’, 4’, 5, 7-pentahydroxyflavan.
The 13C and DEPT spectra, showed fifteen carbon signals, which consisted of one methylene, seven methine and seven quaternary carbons. These signals were as follows: δ = 79.9 (C-2), 67.5 (C-3), 29.3 (C-4), 157.4 (C-5), 96.4 (C-6), 157.9 (C-7), 95.9 (C-8), 157.7 (C-9), 100.1 (C-10), 132.3 (C-1’), 115.3 (C-2’), 145.9 (C-3’), 145.8 (C-4’), 115.9 (C-5’), 119.4 (C-6’).
The 1H-1H COSY correlations showed coupling between H-3 and H-2, and also between H-3 and H-4. These positions were further confirmed by long-range coupling observed in the HMBC (Figure 3). The NMR data (Table 1) thus showed signals typical of epicatechin, and it is comparable to the 1H and 13C NMR data reported in the literature [18, 19].
Figure 3. HMBC and COSY correlations of epicatechin
Table 1. 1H and 13C NMR data of epicatechin in deuterated methanol
Epicatechin has been isolated from the leaves and wood of Acacia catechu [19], apple peels [20], mango kernels [21] and pear skin [22]. This compound has been reported to showed antioxidant activity [23], good antilisterial activity at IC50 greater than 200 µg/mg and antimicrobial activity at MIC value greater than 500µg/ml [24].

4. Conclusions

(-) Epicatechin is a polyphenol with a high antioxidant property and the isolation of this metabolite from T. emetica seems to support some folkloric use of the plant in African traditional medicine. The isolation of the natural product was carried out by using different chromatographic separation and its identity confirmed by polarimetry and spectroscopic techniques.


This research work was sponsored by Nigerian Tertiary Education Trust Fund (TETFUND).


[1]  Komane, B.M., Olivier, E.I., Viljoen, A.M., 2011. Trichilia emetica (Meliaceae) - A review of traditional uses, biological activities and phytochemistry. Phytochem. Lett. 4, 1–9.
[2]  Orwa, C., Mutua, A., Kindt, R., J., R., Simons, A., 2009. Trichilia emetica. Agroforestree database: a tree reference and selection guide version 4.0. Available at [Accessed: March 19, 2015].
[3]  Van der Vossen, H.A., Mkamilo, G.S., 2007. Vegetable oils of tropical Africa, conclusion and recommendations based on PROTA 14: “Vegetable oils”. Available on: PROTA (Plant Resources of Tropical Africa/Resources vegetales de I’Afrique tropicale), Wageningen, Netherlands PROTA.
[4]  Diallo, D., Paulsen, B.S., Liljeback, T.H.A., Michaelsen, T.E., 2003. The malian medicinal plant Trichilia emetica; studies on polysaccharides with complement fixing ability. J. Ethnopharmacol. 84, 279–287.
[5]  Hutchings, A., Scott, A.H., Lewis, G., Cunningham, A.B., 1996. Zulu Medicinal Plants. University of Natal Press, Pietermaritzburg
[6]  El Tahir, A., Satti, G.M.H., Khalid, S.M., 1999. Antiplasmodial activity of selected Sudanese medicinal plants with emphasis on Maytenus senegalensis (Lam.) Exell. J. Ethnopharmacol. 64, 227–233.
[7]  Germanò, M.P., D’Angelo, V., Sanogo, R., Catania, S., Alma, R., Pasquale, R. De, Bisignano, G., 2005. Hepatoprotective and antibacterial effects of extracts from Trichilia emetica Vahl. (Meliaceae). J. Ethnopharmacol. 96, 227–232.
[8]  Germanò, M.P., D’Angelo, V., Biasini, T., Sanogo, R., De Pasquale, R., Catania, S., 2006. Evaluation of the antioxidant properties and bioavailability of free and bound phenolic acids from Trichilia emetica Vahl. J. Ethnopharmacol. 105, 368–373.
[9]  Frum, Y., Viljoen, A.M., 2006. In vitro 5-lipoxygenase and anti-oxidant activities of South African medicinal plants commonly used topically for skin diseases. Pharmacol. Physiol. 19, 329–335.
[10]  Sparg, S.G., van Staden, J., Jager, A.K., 2000. Efficiency of traditionally used South Africa plants against schistosomiasis. J. Ethnopharmacol. 73, 209–214.
[11]  Kamanzi-Atindedou, K., Schmid, C., Brun, R., Koné, M., Traore, D., 2004. Antitrypanosomal and antiplasmodial activity of medicinal plants from Côte d’Ivoire. J. Ethnopharmacol. 90, 221–227.
[12]  Bah, S., Paulsen, B.S. Diallo, D., Johansen, H.T. 2006. Characterization of cysteine proteases in Malian medicinal plants. J. Ethnopharmacol. 107, 189-198.
[13]  Verschaeve, L., and Van Staden, J., 2008. Mutagenic and antimutagenic properties of extracts from South African traditional medicinal plants. J. Ethnopharmacol. 119, 575-587.
[14]  Seto, R., Nakamura, H., Nanjo, F., Hara, Y., 1997. Preparation of epimers of tea catechins by heat treatment. Biosci. Biotech. Biochem. 9, 1434–1439.
[15]  Sun, J., Jiang, Y., Wei, X., Shi, J., You, Y., Liu, H., Kakuda, Y., Zhao, M., 2006. Identification of (-) - epicatechin as the direct substrare for polyphenol oxidase isolated from litchi pericarp. Food Research Int. 39, 864–870.
[16]  Fan, P., Lou, H., Yu, W., Ren, D., Ma, B., Ji, M., 2004. Novel flavanol derivatives from grape seeds. Tetrahedron Lett. 45, 3163–3166.
[17]  Pizzolatti, M. G., Venson, A. F., Smania, A. J., Smania, E. F., & Braz-Filho, R. 2002. Two Epimeric Flavalignans fromTrichilia catigua (Meliaceae) Antimicrobial Activity. Zeitschrift fu¨r Naturfors-chung, 57, 483–488.
[18]  Oh, M.H., Park, K.H., Kim, M.H., Kim, H.H., Park, K.J., Heo, J.H., Lee, M.W., 2014. Anti-oxidative and anti-inflammatory effects of phenolic compounds from the stems of Quercus acuta Thunberg. Asian J. Chem. 26, 4582–4586.
[19]  Shen, D., Wu, Q., Wang, M., Yang, Y., Lavoie, E.J., Simon, J.E., 2006. Determination of the Predominant Catechins in Acacia catechu by Liquid Chromatography/Electrospray Ionization−Mass Spectrometry. J. Agric. Food Chem. 54, 3219–3224.
[20]  Alonso-Salces, R. M., Ndjoko, K., Queiroz, E. F., Ioset, J. R., Hostettmann, K., Berrueta, L.A., 2004. On-line characterisa-tion of apple polyphenols by liquid chromatography coupled with mass spectrometry and ultraviolet absorbance detection. Journal of Chromatography A, 1046, 89–100.
[21]  Arogba, S. S. (2000). Mango (Mangifera indica) kernel: chromatographic analysis of the tannin, and stability study of the associated polyphenol oxidase activity. Journal of Food Composition and Analysis, 13, 149–156.
[22]  Siegelman, H. W. (1955). Detection and identification of polyphenoloxi-dase substrates in apple and pear skins. Archives of Biochemistry and Biophysics, 56, 97–102.
[23]  Gadow, A.V., Joubert, E., Hansmann, C.F., 1997. Comparison of the antioxidant activity of rooibos tea (Aspalathus linearis) with green, oolong and black tea. Food Chem. 60, 73–77.
[24]  Nyila, M.A., Leonard, C.M., Hussein, A.A., Lall, N., 2012. Activity of South African medicinal plants against Listeria monocytogenes biofilms, and isolation of active compounds from Acacia karroo. South African J. Bot. 78, 220–227.