REV. CHIM. (Bucharest) ♦ 61♦ Nr. 12 ♦ 2010 http://www.revistadechimie.ro 1173 Simultaneous Analysis of Taurine and Caffeine in Energy Drinks using Hydrophilic Interaction Chromatography with UV and Evaporative Light Scattering Detection on line RALUCA-IOANA CHIRITA 1,2 , CATALINA DASCALU 1,2 , LUCIAN GAVRILA 2 , CLAIRE ELFAKIR 1 * 1 Institut de Chimie Organique et Analytique CNRS FR 2708 UMR 6005, University of Orléans, F-45067, Orléans, France 2 “Vasile Alecsandri” University of Bacau, 157 Calea Marasesti, 600115, Bacau, Romania This paper presents the validation of a hydrophilic interaction liquid chromatographic method for taurine and caffeine dosing. Acceptable levels of specificity, linearity, precision, accuracy and limits of detection were achieved during the validation step. The method was applied successfully to the analysis of three commercial energy drinks containing about 100 g L -1 of carbohydrates, amounts of taurine in the range 4 g L -1 and ten times lower amounts of caffeine. The determination does not require any preliminary treatment of the samples except dilution and only basic LC instrumentation is necessary for this procedure. Keywords: taurine, caffeine, energy Drink, hydrophilic interaction chromatography,e evaporative light scattering detection Over the last decade, stimulant drinks have developed a considerable share of the global soft drinks market. Legislation controlling their sale and marketing and scientific research into their ingredients lags behind the development of these “functional” beverages. The success of energy drink is related to their reputation of increasing physical performances and mental concentration level. These types of products not only quench thirst but also have physiological and functional effects due to the fact that they contain important amounts of carbohydrates, amino acids, methylxanthines and vitamins. It has been proven that there is a high correlation between the production and maintenance of muscle and the content of amino acids, especially that of taurine [1]. Taurine (2- aminoethanesulfonic acid) is a non-essential sulphur- containing amino acid, which functions with glycine and γ -amino butyric acid as a neurotransmitter [2]. Nonetheless, excess of taurine has also been associated with cardiomiopathy, hepatic toxicity [3] while other possible side effects are still poorly known. Another key component of the energy drinks is caffeine (1,3,7-trimethyl- xanthine), which shows physiological stimulatory effects on various body systems, including the central nervous and cardiovascular system [4, 5]. In this context, fast, reliable and low-cost analytical methods have to be developed to monitor simultaneously the caffeine and taurine content for quality control. Each new method is of interest if it uses a single sample aliquot and conventional instrumentation usually present in routine laboratory. The levels of caffeine in different food samples have been determined by numerous techniques including spectroscopic and chromatographic methods like planar chromatography (HPTLC) [6, 7] and liquid chromatography (HPLC) coupled with ultraviolet (UV) detection [8-11]. On the other hand, some papers described direct determination of taurine in pharmaceutical preparation or energy drink by HPLC with evaporative light scattering detection (ELSD) [12,13] or MS detection [14, 15]. Few references are available for the simultaneous determination of caffeine and taurine by HPTLC or HPLC [7, 16]. Nevertheless, in a routine analysis context, most quality * email: claire.elfakir@univ-orleans.fr; Tel.: +33 238 494 587 control laboratories are not equipped with the relatively expensive instruments nor have the highly trained personnel required for performing such determinations. Thus, the choice of common detection systems for the simultaneous analysis of caffeine and taurine is limited. ELSD was preferred for taurine determination, as it is generally considered to be a very convenient LC detector for analytes without UV chromophore [17]. Considering the inconvenience of the already described methods, the aim of our study was to validate a direct, fast, isocratic and simultaneous analysis of taurine and caffeine by hydrophilic interaction chromatography (HILIC). We present here the validation in terms of linearity, mean recovery and repeatability in order to meet the accepted criteria for bioanalytical method validation [18, 19]. The validation process is very important as it permits analytical methods transposition between laboratories [20, 21]. The validated method was then applied to determine the levels of taurine and caffeine in three different energy drink samples. Experimental part The HPLC system consisted of a Merck-Hitachi (Darmstadt, Germany) model L-6200 ternary pump, a Rheodyne (Cotati, CA, USA) model 7725 injection valve fitted with a 10 µL loop, a 785A UV visible HPLC Detector (Applied Biosystems) set at a wavelength of 272 nm (maximal absorbance wavelength of caffeine). The detection of taurine was carried out using an ELSD detector, Sedex 85 from Sedere (Alfortville, France), coupled in-line after the UV detector. Nitrogen was used as the ELSD nebulizer gas (3 bars), at a temperature of 40°C, and the gain was set to 10. The chromatographic data handling was accomplished using EZChrom Server software (Merck, Darmstadt, Germany). Separation was carried out on an Astec apHera TM NH 2 polymer (150 x 2.1 mm I.D., 5μm) (Whippany, NJ, USA). Flow–rate was 0.15 mL min -1 . Column temperature was regulated by a Croco-Cil oven (Cluzeau, France) at 45°C. HPLC-grade acetonitrile (MeCN) and methanol (MeOH) were purchased from J.T. Baker (Noisy le Sec, France).