Journal of Chromatography A, 1257 (2012) 34–40 Contents lists available at SciVerse ScienceDirect Journal of Chromatography A j our na l ho me p ag e: www.elsevier.com/locate/chroma Quantification of selected furocoumarins by high-performance liquid chromatography and UV-detection: Capabilities and limits  Angus P. Macmaster a , Neil Owen a , Sylvain Brussaux b , Hugues Brevard c , Richard Hiserodt d , Hans Leijs d , Nikola Bast e , Berthold Weber e , Gerd Loesing e , Alan Sherlock f , Christine Schippa g , Matthias Vey h , Eric Frérot i , Emeline Tissot i , Alain Chaintreau i,∗ a Givaudan, Givaudan UK Ltd, Kennington Road, Ashford, Kent – TN24 0LT, UK b Biolandes, F-40420 Le Sen, France c Robertet, 37, avenue Sidi Brahim, BP 52100, F – 06131 Grasse Cedex, France d IFF, Zevenheuvelenweg 60, NL – 5004 EA Tilburg, The Netherlands e Symrise, Mühlenfeldstrasse 1, D – 37601 Holzminden, Germany f RC Treatt & Co Ltd, Northern Way, Bury St. Edmunds, Suffolk, IP33 2DA, UK g V. Mane Fils, 620 route de Grasse, F – 06620 Le Bar sur Loup, France h IFRA, 6 Avenue des Arts, B-1210 Bruxelles, Belgium i Firmenich S.A. 1 route des Jeunes, PO Box 239, CH-1211 Geneve, Switzerland a r t i c l e i n f o Article history: Received 19 March 2012 Received in revised form 9 July 2012 Accepted 14 July 2012 Available online 25 July 2012 Keywords: Furocoumarin Quantification HPLC–UV Validation Accuracy Limit of quantification a b s t r a c t The performance of HPLC–UV as a means of quantifying selected furocoumarins in essential oils has been evaluated, based on a ring test validation approach. Accuracy profiles were generated, to determine bias and statistical confidence associated with determination at different concentrations, along with lower limits of quantification (LOQ). From these findings, it can be concluded that the method described may only be used in simple cases (essential oils), to measure individual furocoumarin compounds at concentrations greater than 10 mg/l; the non compound-specific nature of detection by absorption in the UV range is unable to overcome the effect of interferences arising from chromatographic coelutions, such as those encountered in the analysis of complex commercial fragrance mixtures. The use of an algorithmically calculated ‘spectral similarity’ function, with reference to authentic standards, may be used to improve reliability in assignment and quantification. © 2012 Published by Elsevier B.V. 1. Introduction Furocoumarins (FC) are bioactive constituents occurring natu- rally in various plants; some have been found to exhibit significant phototoxicity [1]. Among commonly used fragrance raw materials, are essential oils produced from such plants, notably those from citrus peels. Many citrus-derived oils are used in large quantities in the manufacture of finished fragrances. While the FC content of steam-distilled oils is typically tiny (<0.02%), owing to the low volatility of FC, their concentration in cold-pressed oils may rise to significant levels (bergamottin: 10 to >20 g/l) [2]. In response to this, the fragrance industry, via its self-regulatory system of IFRA Standards [3] and other regulatory  Analytical Working Group of the International Fragrance Association (IFRA). ∗ Corresponding author. Chairman, IFRA Analytical Working Group. Tel.: +41 22 780 3471; fax: +41 22 780 3334. E-mail address: alain.chaintreau@firmenich.com (A. Chaintreau). schemes (EU Cosmetics Directive [4,5], ASEAN Cosmetics Directive [6] has followed restrictions on FC content in cosmetic products for a number of years. The current limitation of FC in the EU and ASEAN Cosmetic Regulation (entry 358 in Annex II – List of Substances which must not form part of the composition of cosmetic products) states that FC (e.g. xanthotoxin and bergapten) are prohibited, except when they derive directly from the natural essences used (the use of “essence” reflects the terminology used for “essential oil” in that part of the ASEAN directive, which refers to limitations on the con- tent of furocoumarins in Cosmetic Products). In sun-protection and in bronzing products, those FC resulting from natural essences shall be below 1 mg/kg. The Scientific Committee on Consumer Products (SCCP), an advi- sory body of the European Commission, published in 2005, as an outcome of another review of the data, an opinion expressing that a concentration greater than 1 ppm of any one of the FC in any finished cosmetic product would be of concern [1]. The European Commission has not yet taken a decision on whether there is a need to extend legislation and, if so, what is the most suitable way to 0021-9673/$ – see front matter © 2012 Published by Elsevier B.V. http://dx.doi.org/10.1016/j.chroma.2012.07.048