Validation of an Analytical Method to Determine the Content of Sucralose in Beverages - Report on a Method Validation by Collaborative Study Determination of Sucralose in Beverages by Thin Layer Chromatography in Combination with Reagent-free Derivatisation and Ultraviolet/Fluorescence Detection

An inter-laboratory comparison was carried out to evaluate the performance characteristics of a method for the determination of sucralose in beverages, which was developed at the JRC-IRMM. The method is based on high-performance thin layer chromatography (HPTLC), and reagent-free derivatisation followed by ultraviolet/fluorescence detection. It was tested for the determination of Sucralose (C12H19Cl3O8; (2R,3R,4R,5S,6R)-2-[(2R,3S,4S,5S)-2,5-bis(chloromethyl)-3,4-dihydroxy-oxolan-2-yl]oxy-5-chloro-6-hydroxymethyl)oxane-3,4-diol; CAS No: 56038-13-2) in carbonated and still alcoholic and non-alcoholic beverages at proposed European regulatory limits according to Directive 2003/115/EC ( ). Precise determination of Sucralose levels in some of the matrices for which European legislative limits apply, required a robust and reliable analytical method. HPTLC employing reagent-free derivatisation offered such a reliable but simple, fast, cost-effective and environment friendly method (very limited quantities of organic solvents methanol and acetonitrile were used). Separation of Sucralose was performed by direct application of samples (diluted, degassed and/or filtered, if necessary) on amino-bonded silica gel HPTLC plates without prior cleanup and development with acetonitrile:water. The sweetener was determined after heating of the developed plate to 190°C and quantified both in ultraviolet absorption and fluorescence measurement mode. Beverages spiked with sucralose as well as beverages taken from the market and labelled to contain sucralose, were sent to 14 laboratories in five different countries following IUPAC guidelines. A sample that did not contain measureable amounts of sucralose was spiked at levels of 30.5 mg/L, 100.7 mg/L and 299 mg/L. Recoveries ranged from 104 � 125 % with an average of 112 % for ultraviolet detection and from 98 � 101 % with an average of 100 % for fluorescent detection. Based on results for spiked samples (blind duplicates at three levels), as well as samples containing Sucralose (blind duplicates at three levels and one split level), the relative standard deviation for repeatability (RSDr) ranged from 10 � 31 % for ultraviolet detection and from 9 � 16 % for fluorescence detection. The relative standard deviation for reproducibility (RSDR) ranged from 14 � 31 % for ultraviolet detection and from 9 � 21 % for fluorescence detection. The limit of quantification on the basis of 10x the baseline noise was 6 mg/L and response was linear in the range between 30 � 150 ng/spot. The method is therefore considered suitable for the determination of Sucralose in beverages at the proposed European legislative limits.

JRC.D.8 - Food safety and quality