Clujul Medical - Iuliu Haţieganu

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DETERMINATION OF CAFFEINE FROM INSTANT COFFEE AND CAPPUCCINO BY LIQUID CHROMATOGRAPHY – MASS SPECTROMETRY LORENA FILIP 1 , L. VLASE 2 , IOANA MÎNDRUŢĂU 3 , DOINA MIERE 1 , DANIELA HANGANU 4 Faculty of Pharmacy, University of Medicine and Pharmacy “Iuliu Haţieganu”, Cluj-Napoca 1 Department of Environmental Chemistry 2 Department of Pharmaceutical Technology and Biopharmaceutics 3 Department of Physical Chemistry. 4 Department of Phytotherapy Summary This paper describes the application of a high performance liquid chromatography-mass spectrometry (HPLC-MS) method for rapid analysis of caffeine from different brands of instant coffee and cappuccino. Caffeine is an alkaloid naturally found in coffee beans, green tea and cola nuts. Some soft drinks contain it, including instant coffee and cappuccino. The developed method utilises a Zorbax SB- C18, 100 mm x 3.0 mm i.d., 3.5 µm column, with a mobile phase containing methanol : 0.2% formic acid in water. Detection and quantification of caffeine was carried out by monitoring the protonated molecular abundance (m/z 194.9). The quantification was made using the external standard method. The calibration curves were in range of 0.26- 26 µg/ml for caffeine with a determination coefficient greater than 0.995 for all compounds. Keywords: caffeine, LC/MS, instant coffee, cappuccino. Introduction Coffee is one of the most widely consumed beverages in the world. It is prepared from the roasted seeds, commonly referred to as beans, of the coffee plant. Coffee is considered as the most popular processed beverage, with over one third of the world's population consuming it in some form. Instant coffee and cappuccino are among the most frequently-drunk coffee beverages. Instant coffee is a beverage derived from brewed coffee beans, which are transformed through various manufacturing processes into the form of powder or granules. The advantages of instant coffee are speed of preparation, reduced weight and volume, and long shelf life. Cappuccino is a large Italian beverage prepared with espresso, hot milk and milk foam 1 . Caffeine (1,3,7-trimethylxanthine) (Figure 1) is a naturally occurring substance and the world's primary source of caffeine is the coffee bean (the seed of the coffee plant), from which coffee is brewed. Caffeine represents a pharmacologically active substance and can be, depending on the dose, a central nervous system stimulant, a weak bronchodilator, a diuretic and a cardiac muscle stimulant 2 . Methods for determination of xanthines usually involve the use of HPLC methods 3 . This paper describes the results of caffeine quantification using a simple LC/ESI-MS technique. 190
Clujul Medical 2006 vol. LXXX - nr. 1 Materials and method Fig. 1. Chemical structure of caffeine Chemicals Caffeine was purchased from Sigma-Aldrich Germany. Methanol and formic acid 98% were from Merck KGaA (Darmstadt, Germany). Apparatus and chromatographic conditions The concentration of caffeine was determined using a LC/MS analytical method. An Agilent 1100 Series (Agilent, USA) chromatographic system was used coupled with an 1100 LC/MSD Ion Trap detector (Agilent, SUA). The analytical column was a Zorbax SB-C18, 100 mm x 3.0 mm i.d., 3.5 µm, (Agilent, USA) with an on-line filter (0.2µm). The mobile phase was a mixture 24:76 (v/v) of methanol: 0.2% formic acid in water. The flow rate was 0.6 ml/min and the injection volume of 5 µl. The mass analyser was operated with an ESI source, in positive ion mode. For caffeine, the quantification was made by measuring the protonated molecular ion [M+H] + at m/z 194.9. Standard solutions Stock solution of caffeine (1-2 mg/ml) was obtained by dissolving corresponding compounds in methanol. Standard solutions (7 concentrations) were obtained by diluting appropriate volumes of stock solution with bidistilled water. Sample preparation A set of 7 samples of commercially available instant coffee and 11 samples of different brands of cappuccino were selected for the study. Because of detection specificity due to MS detector, the sample preparation was reduced to a simple extraction of caffeine with boiled water and dilution of this solution. About 150 mg of instant coffee and 300 mg of cappuccino were extracted for 10 min with 15 ml boiled water. The volume was adjusted to 25 ml and then 0.1 ml from this solution were diluted to 5 ml. 5 µl from final solution was injected into LC/MS system. Results and discussions Caffeine was identified in all samples by comparing its retention time with that of caffeine in standard solutions. Under the above mentioned operating conditions the retention time for caffeine was 2.75 min, as shown in a chromatogram of an instant coffee sample (Figure 2). Fig. 2. Chromatogram of an instant coffee sample 191
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Clujul Medical Revistă de Medicin
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Articole de orientare Clujul Medica
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VOICHITA SĂBĂDUŞ, D. BORZEA, DIA
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CAMELIA GEORGETA RĂCĂREANU, GABRI
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