Clara Grosso 1 Ana Cristina Figueiredo 2 Jesus Burillo 3 Ana M. Mainar 4 JosØ S. Urieta 4 JosØ G. Barroso 2 JosØ A. Coelho 5 António M. F. Palavra 1 1 Departamento de Engenharia Química e Biológica, IST, Lisboa, Portugal 2 Universidade de Lisboa, Faculdade de CiÞncias de Lisboa, DBV, Centro de Biotecnologia Vegetal, Lisboa, Portugal 3 Centro de Investigación y Tecnología Agroalimentaria, Departamento de Ciencia, Tecnología y Universidad, Gobierno de Aragón, Zaragoza, Spain 4 Química orgµnica y química física, Universidad Zaragoza, Zaragoza, Spain 5 CIEQB/DEQ, ISEL, Lisboa, Portugal Original Paper Enrichment of the thymoquinone content in volatile oil from Satureja montana using supercritical fluid extraction Supercritical fluid extraction (SFE) of the volatile oil from Satureja montana L. was performed under different conditions of pressure (90 and 100 bar), temperature (40 and 508C), mean particle sizes (0.4, 0.6 and 0.8 mm) and CO 2 flow rate (0.8, 1.1 and 1.3 kg/h) to understand the influence of these parameters on the composition and yield of this oil. The results were compared with those obtained for the essential oil isolated by hydrodistillation (HD). The volatile and the essential oil were analysed by GC and GC-MS. The main compounds are carvacrol (52.2 – 62.0% for HD vs. 41.7 – 64.5% for SFE), thymol (8.6 – 11.0% for HD vs. 6.0 – 11.3% for SFE), p-cymene (6.9 – 12.8% for HD vs. 6.0 – 17.8% for SFE), c-terpinene (6.4 – 9.4% for HD vs. 2.3 – 6.0% for SFE) and b-bisabolene (2.0 – 2.7% for HD vs. 2.2 – 3.5% for SFE). The major difference between SFE and HD was the relative amount of thymoquinone, an oxygenated monoterpene with important biological activities, which can be ten-fold higher in volatile oil (1.6 – 3.0 for SFE vs. 0.2% for HD). The morphology of the glandular tri- chomes of S. montana and the effect of the grinding process on them was also eval- uated by SEM. Keywords: Hydrodistillation / Satureja montana / Supercritical fluid extraction / Volatiles / Thymo- quinone / Received: September 2, 2008; revised: October 6, 2008; accepted: October 6, 2008 DOI 10.1002/jssc.200800490 1 Introduction Since ancient times, aromatic and medicinal plants have been used due to the properties of their volatile compo- nents. Everyday new discoveries of the potentialities of these components are reported and pharmaceutical and food industries are testing them as new drugs and food preservers, respectively. Hydrodistillation (HD) and steam distillation are the standard methods to isolate plant volatile components, although some technical problems can be mentioned such as thermal degradation and hydrolysis of some components. These limitations can be overcome using supercritical fluid extraction (SFE), since it can operate at moderate temperatures and there are no hydrolysis reac- tions. Moreover, this method allows a continuous con- trol of the solvent power/selectivity by manipulation of parameters such as pressure and temperature of the supercritical fluid [1]. The most used supercritical fluid is carbon dioxide, because it is safe, readily available, has a low cost, allows supercritical operations at relatively low pressures and near-room temperatures and the extract is practically solvent free [2, 3]. Several works reporting the use of SFE to isolate different groups of plant metabolites have been published in the last years [4 – 9]. According to the Portuguese Pharmacopoeia [10], essential oils are isolated from aromatic plants just by distillation processes such as HD. However, the name of volatile oil is used when the SFE is applied. SFE of the volatile components from plant material may co-extract waxes and fatty acids which contaminate the volatile oil. To minimize the effect of these compo- nents, a fractionation step after the extraction process is required to obtain a pure oil fraction [2, 3]. Satureja montana L. (Lamiaceae), commonly called win- ter savory, is a perennial herb located in the southern Europe, the Caucasus and the North of Africa [11, 12]. As the Lamiaceae is a plant family with many medicinal spe- cies several studies have been performed. Taking into account the Satureja genus the volatiles characterization, Correspondence: Professor António M. F. Palavra, Departamen- to de Engenharia Química e Biológica, IST, Av. Rovisco Pais 1, 1049-001 Lisboa, Portugal E-mail: antonio.palavra@ist.utl.pt Fax: +35-121-8464455 Abbreviations: HD, hydrodistillation; SFE, supercritical fluid ex- traction i 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.jss-journal.com 328 C. Grosso et al. J. Sep. Sci. 2009, 32, 328 – 334