Electrospray ionization mass spectrometry fingerprinting of whisky: immediate proof of origin and authenticity Jens K. S. Møller, a Rodrigo R. Catharino b and Marcos N. Eberlin* b Received 5th October 2004, Accepted 14th March 2005 First published as an Advance Article on the web 11th April 2005 DOI: 10.1039/b415422c Authentic samples of whisky produced in Scotland and USA and counterfeit whisky samples commercialized in Brazil have been directly submitted to electrospray ionization mass spectrometry (ESI-MS) analysis in both the negative and positive ion modes to assess the potential of this technique for simple and rapid quality control and proof of authenticity of whisky samples. ESI in the negative ion mode yields the most characteristic whisky fingerprinting mass spectra in just a few seconds by direct infusion of the samples, detecting the most polar or acidic components of each sample in their deprotonated anionic forms. No pre-treatment of the sample, such as extraction or derivatization or even dilution, is required. The analysis of the ESI(2)-MS data both by simple visual inspection but more particularly by chemometric data treatment enables separation of the whisky samples into three unequivocally distinct groups: Scotch, American and counterfeit whisky, whereas single malt and blended Scotch whiskies are also distinguished to some extent. As indicated by ESI-MS/MS analysis, the diagnostic anions are simple sugars, disaccharides and phenolic compounds. Direct infusion ESI-MS therefore provides immediate chemical fingerprinting of whisky samples for type, origin and quality control, as demonstrated herein for American, Scottish and counterfeit samples, whereas ESI-MS/MS analysis of diagnostic ions adds a second dimension of fingerprinting characterization when improved selectivity is desired. Introduction Scotch whisky is one of the UK’s major export commodities, having a value of US$4 billion in 2003 according to the Scotch Whisky Association. 1 Other variants are manufactured around the world, such as in Ireland, USA, Canada and Japan. 2,3 In general, whisky is considered a luxury ‘‘food product’’ of high quality, partly owing to the traditional way of distillation and to the prolonged period of production and maturation. Therefore, whisky belongs to a high value-added segment of the market of alcoholic beverages, which is prone to counter- feiting and other manipulations aimed at imitating or copying authentic whiskies. The nature and origins of flavours in various types of whisky are very complex interactions of several steps in the manufacturing process, and sensory tools such as the flavour wheel have been developed to meet the demands of the distillers and the product. 4 Over the years the unique character of whiskies and the changes occurring during maturation have been studied. 5 The complex chemical composition of whisky has also been investigated, e.g. organic acids and esters, 6,7 mineral content, 8 phenolics 9 and lactones 10 and characteristic low molecular weight fusel alcohols and derived compounds. 11,12 Various attempts have also been made to develop methods for analysis and quality control of different types of whisky. 13,14 Proof of origin and authenticity are major concerns for the distillers, dealers and consumers of genuine whiskies around the world. Attempts have been therefore made to develop standards or methods for quality control and proof of origin and authenticity. The use of sensory profiling and other organo- leptic testing has been valuable to test authenticity, but such methods require a high degree of training. 15 Analysis of copper and other trace elements has been suggested as an indicator of authenticity for Scotch whiskies, 16 whereas the distribution of methyl-branched alcohols (fusel oils) has been found to distin- guish Irish whiskies from other variants. 17 The isotopic ratio of 13 C and 12 C, 18 and the ratio of furfural and 5-hydroxymethyl-2- furaldehyde have also been proposed as whisky markers. 19 Various mass spectrometric (MS) techniques have also been used to analyse and characterize volatiles in whisky. 20,21 A MS study applied pyrolysis followed by MS to analyse the effect and quality of oak casks used for maturation of Scotch whisky. 22 ESI-MS with direct infusion of samples has recently been shown to be an efficient, sensitive, and fast ‘‘MS-only’’ technique able to screen the composition of the most polar, acidic or basic components of complex mixtures such as beer, 23 wine, 24 and bee propolis. 25,26 Herein we present results from direct infusion ESI-MS analysis of whisky samples and demonstrate its suitability for quality control and proof of authenticity of whiskies. Experimental Whisky samples A total of fifteen whisky samples were obtained from reliable sources. These include Scotch, American and presumably counterfeit whiskies commercialized in Brazil. Table 1 displays an outline of the type and number of each variant used. *eberlin@iqm.unicamp.br PAPER www.rsc.org/analyst | The Analyst 890 | Analyst, 2005, 130, 890–897 This journal is ß The Royal Society of Chemistry 2005