Isolation, identification and purification of caffeine from Coffea arabica L. and Camellia sinensis L.: A combination antibacterial study

Muthanna J. Mohammed, Firas A. Al-Bayati


The present study was conducted to isolate the most important bioactive compound from Coffea arabica (coffee) beans and Camellia sinensis (green tea) leaves. Caffeine (3,7-dihydro-1,3,7-trimethyl-1H-purine-2,6-dione) was isolated from both plants using a liquid–liquid extraction method, detected on thin layer chromatography (TLC) plates in comparison with standard caffeine, which served as a positive control. Moreover, Fourier transform infrared (FTIR) spectrometer and High performance liquid chromatography (HPLC) analyses were used to confirm the purity and characterization of the extracted caffeine. The isolated material(s) from both plants were investigated for their single and combined antibacterial activities against six selected pathogenic bacteria. The Grampositive bacteria were; Staphylococcus aureus, Bacillus cereus and Gram-negative bacteria included; Escherichia coli, Proteus mirabilis,
Klebsiella pneumonia and Pseudomonas aeruginosa. Both compounds at a concentration of 2 mg/ml showed similar antibacterial activities against all tested bacteria, except for P. mirabilis, and the highest inhibitory effect was observed against P. aeruginosa using a modified agar diffusion method. The minimal inhibitory concentration (MIC) of caffeine was determined using a broth microdilution method in 96 multi-well microtitre plates. MIC values ranged from 62.5 to 250.0 μg/ml for the caffeine isolated from coffee and 62.5
to 500.0 μg/ml for green tea caffeine. Combination results showed additive effects against most pathogenic bacteria especially for P. aeruginosa, using both antibacterial assays.
Key words: Antibacterial activity, Coffea arabica, camellia sinensis, Caffeine

Full Text:



Roberts MF, Wink M. Alkaloids: Biochemistry, ecology and

medicinal applications. New York: Plenum Press; 1998.

Yen WJ, Wang BS, Chang LW, Duh PD. Antioxidant properties

of roasted coffee residues. J Agric Food Chem 2005;53:2658-63.

Farah A, de Paulis T, Moreira DP, Trugo LC, Martin PR. Chorogenic

acids and lactones in regular and water-decaffeinated arabica

coffees. J Agric Food Chem 2006;54:374-81.

Daglia M, Cuzzoni MT, Dacarro C. Antibacterial activity of coffee.

J Agric Food Chem 1994;42:2270-2.

Kohlmeier L, Weteringe KG, Steck S, Kok FJ. Tea and cancer

prevention: an evaluation of the epidemiologic literature. Nut

Cancer 1997;27:1-13.

Roberts EA, Wood DJ. A study of the polyphenols in tea leaf paper

chromatography. Biochem J 1951;49:414-9.

Wang Dongfeng W, Wang Chenghong W, Li Jun L, Zhao Guiwen Z.

Components and activity of polysaccharides from coarse tea.

J Agric Food Chem 2001;49:507-10.

Roedig-Penman A, Gordon MH. Antioxidant properties of

catechins and green tea extracts in model food emulsions. J Agric

Food Chem 1997;45:4267-70.

Mbata TI, Debiao L, Saikia A. Antibacterial activity of the

crude extract of Chinese green tea Camellia sinensis on Listeria

monocytogenes. Int J Microbiol 2006;2.

Zhang ZZ, Li YB, Wan XC. Antifungal activities of major tea

leaf volatile constituents toward Colletorichum camelliae Massea.

J Agric Food Chem 2006;54:3936-40.

Garattini S. Caffeine, coffee, and health. New York: Raven Press

Ltd; 1993.

Serafin WE. Methylxanthines. In: Hardman JG, Limbird LE,

editors. The pharmacological basis of therapeutics. New York:

McGraw-Hill; 1995.

RamanaviÄiene A, Mostovojus V, Bachmatova I, RamanaviÄiene A.

Anti-bacterial effect of caffeine on Escherichia coli and Pseudomonas

fluorescens. Acta Medica Lithuanica 2003;10:185-8.

Sakurani H, Mitsuhashi N, Tamaki Y, Akimoto T, Murata O,

Kitamoto Y, et al. Interaction between low-rate irradiation, mild

hyperthermia and low-dose caffeine in a human lung cancer cell

line. Int J Rad Biol 1999;75:739-45.

RamanaviÄiene A, AÄaitÄ— J, DringelienÄ— A, MarkeviÄius A,

RamanaviÄiene A. Effect of caffeine on mice immonucometent

cells. Acta Medica Lithuanica 2003;2:86-9.

Almeida AA, Farah A, Silva DA, Nunan EA, Glória MB.

Antibacterial activity of coffee extracts and selected coffee

chemical compounds against Enterobacteria. J Agric Food Chem


Si W, Gong J, Tsao R, Kalab M, Yang R, Yin Y. Bioassay-guided

purification and identification of antimicrobial components in

Chinese green tea extract. J Chromatogr A 2006;1125:204-10.

Belay A, Ture K, Redi M, Asfaw A. Measurement of caffeine in

coffee beans with UV/vis spectrometer. Food Chem 2008;108:310-15.

Wagner H, Bladt S. Plant drug analysis: A thin layer chromatography

atlas. 2nd ed. Berlin: Springer-Verlag; 1996.

Mothana RA, Lindequist U. Antimicrobial activity of some

medicinal plants of the island Soqotra. J Ethnopharmacol


Sarker SD, Nahar L, Kumarasamy Y. Microtitre plate-based

antibacterial assay incorporating resazurin as an indicator of cell

growth, and its application in the in vitro antibacterial screening

of phytochemicals. Methods 2007;42:321-4.

Silverstein RM, Bassler GC, Morrill TC. Spectrometric identification

of organic compounds. 4th ed. New York: John Wiley and Sons,

Inc; 1981.

Sadek PC. The HPLC solvent guide. 2nd ed. New York: John Wiley

and Sons, Inc; 2002.

Wang X, Wan X, Hu S, Pan C. Study on the increase mechanism

of the caffeine content during the fermentation of tea with

microorganisms. Food Chem 2008;107:1086-91.

Nartowicz V, Buchanan R, Segall S. Aflatoxin production in regular

and decaffeinated coffee beans. J Food Sci 1979;44:446-8.

Daglia M, Tarsi R, Papetti A, Grisoli P, Dacarro C, Pruzzo C,

Gazzani G. Antiadhesive effect of green and roasted coffee

on Streptococcus mutans adhesive properties on saliva-coated

hydroxyapatite beads. J Agric Food Chem 2002;50:1225-9.

Ibrahim S, Salameh M, Phetsomphou S, Yang H, Seo C. Application

of caffeine, 1,3,7-trimethylxantine to control Escherichia coli

O157:H7. Food Chem 2006;99:645-50.

Labbe RG, Nolan LL. Inhibition of macromolecular synthesis by

caffeine in Clostridium perfringes. Can J Microbiol 1978;33:589-92.

Kaufmann WK, Heffernan TP, Beaulieu LM, Doherty S, Frank AR,

Zhou Y, et al. Caffeine and human DNA metabolism: The magic

and the mystery. Mut Res 2003;532:85-102.

Tenover FC. Mechanisms of antimicrobial resistance in bacteria.

Am J Med 2006;119:S3-10.

Daglia M, Papetti A, Grisoli P, Aceti C, Spini V, Dacarro C,

Gazzani G. Isolation, identification, and quantification of roasted

coffee antibacterial compounds. J Agric Food Chem 2007;55:


Charles BG, Rawal BD. Synergistic effect of methyl substituted

xanthines and neomycin sulphate on Staph. aureus and

P. aeruginosa in vitro. Lancet 1973;1:971-3.

Hosseinzadeh H, Bazzaz B, Sadati M. In vitro evaluation of

methylxanthines and some antibiotics: Interaction against

Staphylococcus aureus and Pseudomonas aeruginosa. Iranian Biomed

J 2006;10:163-7.



  • There are currently no refbacks.