1370 J. Sep. Sci. 2013, 36, 1370–1378 Johanna Rudb ¨ ack 1 Nurul Islam 1 Ulrika Nilsson 2 Ann-Therese Karlberg 1 1 Department of Chemistry and Molecular Biology, Dermatochemistry and Skin Allergy, University of Gothenburg, Gothenburg, Sweden 2 Department of Analytical Chemistry, Stockholm University, Stockholm, Sweden Received September 11, 2012 Revised January 26, 2013 Accepted January 29, 2013 Research Article A sensitive method for determination of allergenic fragrance terpene hydroperoxides using liquid chromatography coupled with tandem mass spectrometry Different compositions of monoterpenes are utilized for their pleasant scent in cosmetics and perfumes. However, the most commonly used fragrance terpenes easily oxidize upon contact with air, forming strongly skin-sensitizing hydroperoxides. Due to their thermola- bility and low UV absorbance, detection methods for hydroperoxides are scarce. For the first time, a simple and sensitive method using LC/ESI-MS/MS was developed to quantitatively determine hydroperoxides from the common fragrance compounds linalool, linalyl acetate, and limonene. The method was applied to autoxidized petitgrain oil and sweet orange oil. A separation was accomplished using a C 3 column. The method LOD for the investigated hydroperoxides in the essential oils was below 0.3 g/mL, corresponding to 0.3 ppm. For prevention purposes and according to EU regulations, concentrations in cosmetics exceed- ing 100 ppm in “rinse-off” and 10 ppm in “stay-on” products of linalool and limonene must be labeled. However, the products may still contain allergens, such as hydroperoxides, formed by oxidative degradation of their parent terpenes. The sensitivity and selectivity of the presented LC/MS/MS method enables detection of hydroperoxides from the fragrance terpenes linalool, linalyl acetate, and limonene. However, for routine measurements, the method requires further validation. Keywords: Autoxidation / Contact allergy / Fragrance terpenes / Hydroperoxides / Liquid chromatography–tandem mass spectrometry DOI 10.1002/jssc.201200855  Additional supporting information may be found in the online version of this article at the publisher’s web-site 1 Introduction Fragrances are, after nickel, the most common cause of con- tact allergy. It is estimated that the frequency of contact allergy to fragrance ingredients in the general population in Europe is 1–3% [1–3]. Contact allergy is a life-long altered specific re- activity in the immune system that can develop following skin contact with a sufficient amount of allergenic compounds. Allergic contact dermatitis develops upon re-exposure to the specific allergen(s) that, in the case of fragrance allergy, is due to contact with every-day products such as cosmetics and household products. Eczema from fragrance compounds typ- Correspondence: Professor Ann-Therese Karlberg, Department of Chemistry and Molecular Biology, Dermatochemistry and Skin Allergy, University of Gothenburg, SE-412 96 Gothenburg, Sweden E-mail: karlberg@chem.gu.se Fax: +46-31-772 1394 Abbreviations: APCI, atmospheric pressure chemical ion- ization; FID, flame ionization detection; IS, internal stan- dard; LimOOH, limonene-2-hydroperoxide; LinOAcOOH, lina- lyl acetate hydroperoxides; LinOOH, linalool hydroperoxides; MRM, multiple-reaction monitoring ically involves the face, the armpits, and the hands [4, 5]. Pa- tients appear to become sensitized to fragrances primarily from deodorants and perfumes and to a lesser extent from other cosmetic types [6]. In order to reduce the risk of skin sensitization and eczema in the population, the EU legislation lists 26 fragrance compounds for which information should be provided to con- sumers about their presence in cosmetic products. For prod- ucts that are designed to stay on the skin for a prolonged time and not to be rinsed off, the limit for labeling is set to 0.001% or 10 ppm (e.g., lotions, deodorants). For those compounds that are rinsed off directly after application, the limit is 0.01% or 100 ppm (e.g., shampoos). The 26 fragrance compounds were introduced into Annex III of the Cosmetics Directive by the 7th amendment (2003/15/EC) on the basis of the opin- ion of The Scientific Committee on Cosmetic Products and Non-Food Products intended for Consumers (SCCNFP) from 1999 [7]. It is important that analytical methods sensitive enough for quantification of the allergenic fragrance ingredients are available to control their presence in complex mixtures. Such methods are in progress or have been developed following the EU legislation [8, 9]. C  2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.jss-journal.com