Asian Pacific Journal of Tropical Medicine

: 2018  |  Volume : 11  |  Issue : 8  |  Page : 480--485

Comparative phytochemical analysis of Coffea benghalensis Roxb Ex Schult, Coffea arabica L. and Coffea liberica Hiern

Éva Brigitta Patay1, Ágnes Alberti2, Orsolya Csernák2, Szilvia Stranczinger3, Nóra Papp1,  
1 Department of Pharmacognosy, Faculty of Pharmacy, University of Pécs, Rókus 2, 7624 Pécs, Hungary
2 Department of Pharmacognosy, Faculty of Pharmacy, Semmelweis University, Üllői 26, 1085 Budapest, Hungary
3 Department of Plant Biology, Faculty of Sciences, University of Pécs, Ifjúság 6, 7624 Pécs, Hungary

Correspondence Address:
Nóra Papp
Department of Pharmacognosy, Faculty of Pharmacy, University of Pécs, Rókus 2, 7624 Pécs


Objective: To make phytochemical studies of the leaf, pericarp and seed of Coffea benghalensis (C. Benghalensis) compared with those of the widely known Coffea arabica and Coffea liberica. Methods: The sample extracts were prepared by Soxhlet-extraction. Polyphenol content was analyzed by HPLC-ESI-MS/MS, the identification was carried out based on the retention time, UV and mass spectra of standards and literature data of the detected compounds. Results: Phenolic acids like caffeoylquinic acids, dicaffeoylquinic acids, feruloylquinic acids and coumaroylquinic acid, as well as mangiferin were detected as main constituents in all extracts. Procyanidin trimers were present exclusively in the leaves. In C. benghalensis, main constituents were 5-caffeoylquinic acid and 4-caffeoylquinic acid. Flavan-3-ols were described in all immature and mature pericarp and leaf extracts. Even though 4-feruloylquinic acid was described in both immature and mature seed, dicaffeoylquinic acids were identified only in the mature seed extracts. Mangiferin was present in the leaf, mature pericarp and seed. Conclusions: These analyses provide new chemotaxonomical data for the selected coffees, especially for C. benghalensis. Due to its high polyphenol content, our results indicate its significance of providing new data as a possible source for industry.

How to cite this article:
Patay &B, Alberti &, Csernák O, Stranczinger S, Papp N. Comparative phytochemical analysis of Coffea benghalensis Roxb Ex Schult, Coffea arabica L. and Coffea liberica Hiern.Asian Pac J Trop Med 2018;11:480-485

How to cite this URL:
Patay &B, Alberti &, Csernák O, Stranczinger S, Papp N. Comparative phytochemical analysis of Coffea benghalensis Roxb Ex Schult, Coffea arabica L. and Coffea liberica Hiern. Asian Pac J Trop Med [serial online] 2018 [cited 2021 Mar 5 ];11:480-485
Available from:

Full Text

 1. Introduction

Coffea species are well-known and wide-spread all over the world in tropical and subtropical areas, especially in the Equatorial region. The most important coffee producing continents are South America (Brazil, Colombia, Venezuela, Bolivia, Peru, Ecuador), Central America (Mexico, El Salvador, Cuba, Haiti, Dominica, Nicaragua, Guatemala), Africa (Angola, Liberia, Ethiopia, Congo, Kenya, Tanzania, Uganda, Nigeria, Malawi), and Asia (India, Sri Lanka, Malaysia, Indonesia, Java, Sumatra, New Guinea)[1],[2] .

They also have an important role in science because of their pharmacological role. In addition, they are one of the most sought products after petrol and they also provide an income for more than 20 million families in more than 50 countries every year[3],[4].

Because of the great impact on the quality of coffee beverages, many biochemical studies were reported about the chemical composition of the green and roasted coffee beans. By contrast, only few phytochemical and microbiological analyses are available about the leaf and pericarp of wild and cultivated coffee species.

Chlorogenic acids as characteristic components of coffee beans and commercial coffee products compose a group of esters formed between quinic acid and trans-hydroxycinnamic acids (e.g. caffeic, ferulic and p-coumaric acid), and sometimes from dimethoxycinnamic, trimethoxycinnamic and sinapic acid[5],[6]. Among chlorogenic acids, caffeoylquinic, p-coumaroylquinic, feruloylquinic, dicaffeoylquinic and caffeoylferuloyl quinic acids have been reported in coffees[7]. Chlorogenic acids usually form a compound with caffeine which occurs in 1:1 ratio in plants[8].

Many studies focused on cultivated species like Coffea arabica (C. arabica) L. (Arabic coffee), Coffea canephora Pierre ex A. Froehner (Robusta coffee) and Coffea liberica (C. liberica) Hiern. (Liberian coffee) nowadays, but only few data are available about wild coffees like Coffea benghalensis (C. benghalensis) Roxb. ex Schult. (Bengal coffee). Bengal coffee was studied for its histological and phytochemical characters; in our previous work, caffeic acid, sinapic acid and rutin were identified in the immature fruits[4]. Furthermore, caffeine and terpenoids (cafesterol, bengalensol) have been also detected in the leaf and the fruit earlier[9],[10],[11]. In addition, Trevisan et al.[12] denoted higher total mangiferin content in the leaf of plants growing under natural full-sun conditions compared to other ones living in management used organic treatment. The immature pericarp of Bengal and Liberian coffees showed higher amount of tannin and polyphenol components compared with Arabic species[13].

The aim of this study was to investigate and compare the phenolic content of the leaf, pericarp and seed of C. benghalensis, C. arabica and C. liberica. A further purpose was to evaluate the phenolic composition of their immature and mature pericarp, which has not been analyzed up to the present. The phytochemical analysis aimed at the qualitative comparison of phenolic compounds to provide new chemotaxonomical data for the selected coffee species.

 2. Materials and methods

2.1. Plant materials

The leaf, immature and mature pericarp and seed of C. benghalensis, C. arabica and C. liberica were collected at the Botanical Garden, University of Pécs in the spring of 2015. The samples were air-dried at room temperature in the shade. Voucher specimens were deposited and labeled with unique codes at the Department of Pharmacognosy, University of Pécs.

2.2. Chemicals

The following chemicals were used for all analyses: petroleum ether (Molar Chemicals Ltd., Budapest, Hungary), methanol of analytical grade (Reanal, Budapest, Hungary), acetic acid and methanol of HPLC supergradient grade (Sigma-Aldrich, Steinheim, Germany). All aqueous eluents for LC-MS were filtered through MF-Millipore membrane filters (0.45 μm, mixed cellulose esters) (Billerica, MA, USA).

2.3. HPLC-ESI-MS/MS analysis

2.3.1.Sample preparation

All plant samples were powdered and extracted using Soxhlet-extraction method (70:30 v/v % methanol: distilled water). After the evaporation of the solvent, the residues were redissolved in 5 mL 70:30 v/v % methanol-water mixture. Apolar compounds were removed by liquid-liquid extraction with petroleum ether if needed. Prior to evaluation, all samples were submitted to SPE purification (500 mg/3 mL Supelco Supelclean LC-18 SPE cartridges, Sigma-Aldrich, Steinheim, Germany), and they were filtered through Sartorius (Goettingen, Germany) Minisart RC15 (0.2 μm) syringe filters.


Chromatographic analyses were performed on an Agilent 1100 HPLC Series system coupled with an Agilent 6410 Triple Quadrupole mass spectrometer using an electrospray ion source in negative ionization mode. A ZORBAX SB-C18 3.0 mm x 150 mm, 3.5 μm column was used for separation. A total of 0.3 v/v % acetic acid in water and methanol was used as mobile phase A and B respectively with a gradient method as follows: 0 min 10 v/v % B, 30 min 100 v/v % B, 35 min 100 v/v % B. The temperature of the column was kept at 25 °C. The flow rate of the mobile phase was 0.3 mL/min and the injection volume was 5 μL. ESI conditions were as follows: temperature: 350 °C, nebulizer pressure: 40 psi (N2), drying gas flow rate: 9 L/min (N2), capillary voltage: 3 500 V, fragmentor voltage: 100 V. Collision energy was changed between 10-50 eV, according to structural differences. High purity nitrogen was used as collision gas. Full mass scan spectra were recorded over the range m/z 70-1 000 (1 scan/sec). The Masshunter B.01.03 software was used for data acquisition and qualitative analysis. For unambiguous identification, retention times, UV and mass spectra were compared with literature data and with those of authentic standards.

 3. Results

3.1. HPLC analysis

Altogether 25 phenolic components were identified in the extracts of the studied parts of the selected Coffea species [Table 1] and 22 compounds were detected in Bengal coffee [Figure 1]. Among them, 16 phenolic acid derivatives (e.g. caffeoylquinic acids), 2 flavan-3-ols, 2 procyanidin dimers and 2 procyanidin trimers, a xanthonoid, and 2 aliphatic tricarboxylic acids were qualitatively characterized by comparison of their LC-ESI-MS/MS data with the literature and mass spectral data of reference compounds [Table 1].{Table 1}{Figure 1}

Phenolic compositions of the studied coffee species were similar with minor differences. Chlorogenic acids were observed as the main components in each extract. 4-caffeoylquinic acid (4-CQA) and 5-caffeoylquinic acid (5-CQA) were detected in each sample, except that the latter was absent in C. arabica leaf extract. The most complex composition including 17 compounds was detected in the immature pericarp of Arabic coffee, followed by the extract of the mature pericarp of Bengal coffee (16 compounds), and the leaf extract of Liberian coffee (16 compounds).

3.1.1. Leaf extracts

3-caffeoylquinic acid (3-CQA) was present in all leaf samples being the main compound in C. arabica [Table 2]. 5-CQA was the main component in C. benghalensis and C. liberica. 5-coumaroylquinic acid (5-CoQA) and a further isomer of 5-CQA were present in Liberian coffee. 4-CQA, dicaffeoylquinic acids (diCQAs) and ferulic acid, as well as mangiferin were detected in all samples.{Table 2}

3.1.2.Immature pericarp extracts

5-CQA was characterized as the main component of all samples, additionally, 4-CQA and catechin/epicatechin also were abundant in all studied species [Table 3]. Moreover, Arabic and Liberian coffee contained 3-CQA, a further isomer of 5-CQA, 4-feruloylquinic acid (4-FQA), 3,4-diCQA, 3,5-diCQA and 4,5-diCQA, as well.{Table 3}

A. Leaf, B. Immature pericarp, C. Immature seed, D. Mature pericarp, E. Mature seed.

3.1.3.Immature seed extracts

The main compound of all coffee immature seed samples was 5-CQA, followed by 4-FQA, 4-CQA and 3-CQA [Table 4]. 5-CQA, 5-CoQA and 5-FQA were identified in Arabic and Liberian coffees. The latter was characterized by the presence of a procyanidin dimer and 5-CQA methyl ether that could be detected only in both immature and mature seed of Liberian coffee. The diCQA compounds were described for C. benghalensis and C. arabica, while Liberian coffee contained only 3,4-diCQA and 4,5-diCQA.{Table 4}

3.1.4. Mature pericarp extracts

Similar to the immature seed samples, the main compound of the mature pericarp extracts was 5-CQA, while 4-CQA was present in smaller amount in each species. For Bengal and Arabic coffees, 3-CQA, 5-CQA, 5-CoQA, 4-FQA and diCQA compounds were described, as well. Liberian coffee extract contained only 4,5-diCQA. Flavan-3-ols and a procyanidin compound were detected in Bengal coffee [Table 5].{Table 5}

3.1.5. Mature seed extracts

In the mature seed extracts, 5-CQA was identified as the main compound in Bengal and Arabic coffee, while 4-CQA, 5-CQA,4-FQA and the diCQAs were detected as minor components [Table 6]. In addition, Bengal coffee was characterized by the presence of 5-FQA. The 5-CQA and 4-CQA isomers compounds, as well as the components 7 and 13, respectively, were detected with comparably high intensity in Liberian coffee extracts. Minor components of the mature seed samples were 3-CQA, 5-CoQA, 4-FQA and diCQAs. Citric acid was found only in C. liberica.{Table 6}

 4. Discussion

Based on few literature data of phytochemical investigations of C. benghalensis, which could have formed the basis of the analyses, the comprehensive comparison of the plant with two well-known coffee species is essential to express its possible pharmacological significance.

The identification of mangiferin in the leaf extracts was in concordance with earlier works[12]. They showed that the release of mangiferins depends on the temperature which most effective at 100 °C, and it releases less in 50% methanol extraction because of lower temperature[12]. In contrast to Conéjéro et al. 2014 findings, we did not identify 5-caffeoylquinic acid in C. arabica leaves, but procyanidin trimers were described in each leaf sample[14].

DiCQAs were characteristic for most extracts, and 4,5-diCQA (24) was present in all samples, except Bengal coffee's immature pericarp; while 3,4-diCQA (22) was detected in each sample excluding the mature pericarp of Liberian coffee and the immature pericarp of Bengal coffee[5],[7]. In addition, isocitric acid was described in all samples[15].

The results underline the presence of chlorogenic acids detected in our previous LC/MS studies which analysed their quantity in the non-hydrolysed extracts of the leaf and the immature fruit in Bengal coffee[16]. In our previous and actual investigations, ferulic acid was identified in immature pericarp extracts though the quantified ferulic acid concentration was insignificant in the immature pericarp of Bengal coffee[16]. The ferulic acid concentration was the same in the non-hydrolysed extract made of the immature seed and the leaf of Bengal coffee. The results overlap with the findings of an earlier comprehensive work mentioned the presence of chlorogenic acids[17],[18]. Even though these phenolic compounds were described earlier in the immature seed of Arabic coffee[17], we identified in plus isocitric acid, caffeoyl hexoside, and catechin/epicatechin in the plant. However, this review includes 5-CQA, this compound was not identified in the green seed of Arabic coffee using our methods. Babova et al. underlined the importance of the geographical origine of plants which could influence the caffeine and phenolic content of the cultivated C. arabica and C. canephora[18]. The environmental attributes of the growing area, geographical origine and the growing practice can positively or negatively influence the chemical composition of the coffee plants[19]. In addition, chlorogenic acids content of green coffee beans can depend among others on genes, species, climate and nutrient state of soil[20].

Our results overlap with our earlier work, as the quantity of chlorogenic acids were three times higher in the non-hydrolised extract made of the immature seed of Arabic coffee than that of Bengal coffee[16]. Ky et al.[21]. and Campa et al.[22] also identified and quantified chlorogenic acids in the fruit of Liberian coffee, which confirms our detailed studies. Catechin, epicatechin, flavonols, anthocyanidins, flavan-3-ols and hydroxycinnamic acids like caffeoylquinic acid, its derivatives and p-coumaroylquinic acid detected by Ramirez-Coronel et al.[23] support our investigations and underline that the constitutive units of Arabic coffee fruit were mainly epicatechin, representing more than 90% of the proanthocyanidin units.

In conclusion, the cultivated Coffea species have been evaluated comprehensively; however, the wild Bengal coffee is less investigated, thus our results may contribute to the knowledge regarding its phytochemical composition. Our findings provide relevant new information about less investigated wild coffee taxa that may be of similar significance as Arabic and Liberian coffee. We could summarize that our findings completed the scientific literature of Bengal coffee presenting new opportunities and challenges for further phytochemical and medical studies of the species.


This work was supported by the Research Grant of the University of Pécs (PTE ÁOK KA-2017-27). The present scientific contribution is dedicated to the 650th anniversary of the foundation of the University of Pécs, Hungary.

Conflict of interest statement

The authors declare that there was no conflict of interest.


1M ndita D. What do we know and what do not we know about coffee. Bucuresti: Editura Tehnic; 2008.
2Tóth L. Medicinal plants: Drugs phytoterapy. 2nd ed. Debrecen: Debreceni Egyetemi Kiadó; 2010.
3Davis AP, Chester M, Maurin O, Fay MF. Searching for the relatives of Coffea (Rubiaceae, Ixoroideae): The circumscription and phylogeny of Coffeeae based on plastid sequence data and morphology. Am J Bot 2007; 94(3): 313-329.
4Patay ÉB, Németh T, Németh TS, Papp N. Biological, phytochemical and medicinal evaluation of Coffea taxa (in Hungarian: Coffea taxonok biológiai, fitokémiai és gyógyászati értékelése). Botanikai Kôzlemények 2014; 101(1-2): 263-280.
5Jaiswal R, Müller H, Müller A, Karar MGE, Kuhnert N. Identification and characterization of chlorogenic acids, chlorogenic acid glycosides and flavonoids from Lonicera henryi L. (Caprifoliaceae) leaves by LC-MSn. Phytochemistry 2014; 108: 252-263.
6Opitz SEW, Goodman BA, Keller M, Smrke S, Wellinger M, Schenkerc S, et al. Understanding the effects of roasting on antioxidant components of coffee brews by coupling on-line ABTS assay to high performance size exclusion chromatography. Phytochem Anal 2017; 28(2): 106-114.
7Clifford MN, Johnston KL, Knight S, Kuhnert N. Hierarchical scheme for LC-MSn identification of chlorogenic acids. J Agric Food Chem 2003; 51(10): 2900-2911.
8Ky CL, Barre P, Noirot M. Genetic investigations on the caffeine and chlorogenic acid relationship in an interspecific cross between Coffea liberica Dewevrei and C. pseudozanguebariae. Tree Genet Genomes 2013; 9(4): 1043-1049.
9Ashihara H, Crozier A. Biosynthesis and catabolism of caffeine in low-caffeine-containing species of Coffea. J Agric Food Chem 1999; 47(8): 3425-3431.
10Bilkis B, Choudhury MH, Mohammad AR. Caffeine from the mature leaves of Coffea benghalensis. Biochem Syst Ecol 2003; 31(10): 12191220.
11Mohammad AK, Mohammad SR, Mohammed ZR, Choudhury MH, Mohammad AR. A review on phytochemicals from some medicinal plants of Bangladesh. J Pharm Nutr Sci 2011; 1: 87-95.
12Trevisan MTS, Almeida RF, Soto G, Filho EDMV, Ulrich CM, Owen RW. Quantitation by HPLC UV of mangiferin and isomangiferinin coffee (Coffea arabica) leaves from Brazil and Costa Rica after solvent extraction and infusion. Food Anal 2016; 9(9): 2649-2655.
13Patay ÉB, Sali N, K szegi T, Csepregi R, Balázs VL, Németh TS, et al. Antioxidant potential, tannin and polyphenol contents of seed and pericarp of three Coffea species. Asian Pac J Trop Med 2016; 9(4): 366371.
14Conéjéro G, Noirot M, Talamond P, Verdeil JL. Spectral analysis combined with advanced linear unmixing allows for histolocalization of phenolics in leaves of coffee trees. Front Plant Sci 2014; 5: 39.
15Bylund D, Norström SH, Essén SA, Lundström US. Analysis of low molecular mass organic acids in natural waters by ion exclusion chromatography tandem mass spectrometry. J Chromatogr A 2007; 1176(1-2): 89-93.
16Patay ÉB, Németh T, Németh TS, Filep R, Vlase L, Papp N. Histological and phytochemical studies of Coffea benghalensis B. Heyne Ex Schult. compared with Coffea arabica L. Farmacia 2016; 64(1): 125-130.
17Farah A, Donangelo CM. Phenolic compounds in coffee. Braz J Plant Physiol 2006; 18(1): 23-36.
18Babova O, Occhipinti A, Maffei ME. Chemical partitioning and antioxidant capacity of green coffee (Coffea arabica and Coffea canephora) of different geographical origin. Phytochemistry 2016; 123: 33-39.
19Komes D, Vojvodi A. Effects of varieties and growing conditions on antioxidant capacity of Coffea. In: Processing and impact on antioxidants in beverages. Cambridge: Academic Press; 2014.
20Narita Y, Inouye K. Chlorogenic acids from coffee: Coffee in health and disease prevention. Cambridge: Academic Press; 2015.
21Ky CL, Noirot M, Hamon S. Comparison of five purification methods for chlorogenic acids in green coffee beans (Coffea sp.). J Agric Food Chem 1997; 45(3): 786-790.
22Campa C, Doulbeau S, Dussert S, Hamon S, Noirot M. Qualitative relationship between caffeine and chlorogenic acid contents among wild Coffea species. Food Chem 2005; 93(1): 135-139.
23Ramirez-Coronel MA, Marnet N, Kolli VS, Roussos S, Guyot S, Augur C. Characterization and estimation of proanthocyanidins and other phenolics in coffee pulp (Coffea arabica) by thiolysis-high-performance liquid chromatography. J Agric Food Chem 2004; 52(5): 1344-1349.