Original Paper Fiori, Hudaib, Valgimigli, Gabbanini, Cavrini 703 Jessica Fiori a) , Mohammad Hudaib a) , Luca Valgimigli b) , Simone Gabbanini c) , Vanni Cavrini a) a) Dipartimento di Scienze Farmaceutiche, Università di Bologna, Via Belmeloro 6, 40126 Bologna, Italy b) Dipartimento di Chimica Organica “A. Mangini”, Università di Bologna, Via S. Donato 15, 40127 Bologna, Italy c) R & D Department, BeC s.r.l., Via C. Monteverdi 49, 47100 Forli, Italy Determination of trans-anethole in Salvia sclarea essential oil by liquid chromatography and GC-MS Liquid chromatographic (HPLC) and GC-MS methods were developed to confirm the unusual presence of trans-anethole in Salvia sclarea essential oil. The selectivity of the HPLC method with a diode array detector (DAD) was enhanced by a post-column on-line photochemical derivatization. When a photoreactor (equipped with a 254 nm UV lamp) was arranged between the analytical column and the DAD, photoinduced modification of the anethole chromophore resulted in modified spectral properties. Therefore, two UV spectra (photoreactor Off and On) were achieved for the photo- reactive trans-anethole, providing an information rich detection system. Using both HPLC and GC-MS approaches the unambiguous confirmation of the occurrence of trans-anethole in some commercial samples of S. sclarea oil was obtained. Genuine samples of S. sclarea essential oil were also prepared from the “drug” by each of the following techniques: steam-distillation, Soxhlet extraction in n-hexane, cold extrac- tion in diethyl ether. GC-MS analysis of all the above extracts revealed the presence of trans-anethole. Although not previously recognized, trans-anethole is to be consi- dered a “normal” component of S. sclarea essential oil. Key Words: trans-Anethole; Essential oil; Salvia sclarea; Received: December 21, 2001; revised: February 19, 2002; accepted: March 21, 2002 1 Introduction In previous investigations [1], carried out to evaluate the phytochemical impact of viral infections on the composi- tion of the essential oil from Salvia sclarea, evidence of the presence of a significant level of trans-anethole in a commercial oil was observed. trans-Anethole, 1-methoxy- 4-(1E)-1-propenylbenzene, used as an expectorant and carminative in cough preparations [2] and as flavor enhan- cer in the food and drink industry [3], is the main compo- nent of anise and fennel oils [3, 4], but is not reported as a component of S. sclarea oil. The main components of this oil are monoterpene esters (linalyl acetate), alcohol (lina- lol), hydrocarbons (myrcene), and sesquiterpenes hydro- carbons (germacrene D, b-caryophyllene) [5 – 12]; reports about the presence of trans-anethole have not come to our knowledge. This surprising preliminary result led us to extend the investigations over a wider range of commercial S. sclarea oils, using both liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS) techniques, as independent methods able to provide unambiguous confirmatory data. These approaches were chosen according to the physico-chemical properties of anethole (volatile and UV-absorbing compound); GC-MS is a well established technique suitable for the analysis of complex volatile mixtures such as the essential oils, and liquid chromatography with a photodiode array detector (DAD) offers the opportunity to obtain the UV spectrum for each analyte and to compare it with that of a standard. This technique has been recently applied to the determi- nation of trans-anethole in fennel teas [13]. In this application, to improve the intrinsic selectivity of the HPLC-DAD method, the photoreactivity of anethole, known as a photolabile compound [2, 4], was positively exploited by introducing a post-column on-line photoche- mical derivatization. In fact, when a photochemical reactor is arranged on-line between the analytical column and the detector, the column effluent is subjected to UV irradiation at 254 nm and the analyte can undergo light-induced structural modifications resulting in modified UV spectral and electrochemical properties [14, 15]. Using a diode array detector (DAD) two UV spectra can be recorded for each analyte (photoreactor switched on and off). In pre- vious studies this approach was found to be an informa- tion-rich detection system able to enhance the (HPLC) method selectivity and useful for confirming peak identity in the analysis of pharmaceutical [16, 17] and cosmetic [18, 19] preparations of complex composition. Therefore, GC-MS and HPLC, with and without post-col- umn photochemical derivatization, were applied to verify the presence of trans-anethole in commercial S. sclarea oils. The presence of trans-anethole in some products Correspondence: Vanni Cavrini, Dipartimento di Scienze Farmaceutiche, Università di Bologna, Via Belmeloro 6, 40126 Bologna, Italy. E-mail: vcavrini@alma.unibo.it Fax: +39 051-2099734 J. Sep. Sci. 2002, 25, 703–709 i WILEY-VCH Verlag GmbH, 69469 Weinheim 2002 1615-9306/2002/1007–0703$17.50+.50/0