Analysis of volatiles from black pine (Pinus nigra): significance of wounding and egg deposition by a herbivorous sawfly Roland Mumm a , Tassilo Tiemann a , Stefan Schulz b , Monika Hilker a, * a Institute of Biology, Freie Universita ¨ t Berlin, Haderslebener Str. 9, 12163 Berlin, Germany b Institute of Organic Chemistry, Technische Universita ¨ t Braunschweig, Hagenring 30, 38106 Braunschweig, Germany Received 3 July 2004; received in revised form 13 October 2004 Abstract The composition of headspace volatiles of black pine (Pinus nigra) was analysed by coupled gas chromatography–mass spectrom- etry (GC–MS). It has been shown in a previous study that egg deposition of the sawfly Diprion pini on P. sylvestris induces a quan- titative change of the pine volatile blend . Chrysonotomyia ruforum, an egg parasitoid of D. pini, is known to be attracted by volatiles from egg-carrying P. sylvestris, but not by odour from egg-laden P. nigra. Therefore, the present study focused on the question whether also P. nigra as another host plant of this sawfly responds to egg deposition by change of its volatile blend. The headspace of untreated, egg-carrying, and artificially wounded P. nigra twigs were compared. The artificial damage inflicted to the twigs mim- icked the damage by the sawfly female prior to egg deposition. Thirty five mainly terpenoid compounds that were identified in more than 50% of the egg-carrying P. nigra twigs could also be detected in the headspace of untreated and artificially wounded twigs. Quantitative differences of the blends of volatiles from differently treated P. nigra twigs were compared by multivariate data anal- yses. PLS-DA (projection to latent structures-discriminant analysis) revealed that blends of volatiles from differently treated P. nigra differed significantly. When comparing volatiles from egg-carrying and artificially wounded P. nigra with respective P. sylvestris samples, qualitative and quantitative differences were detected. The differences in volatile composition of P. nigra and P. sylvestris are discussed with special respect to the egg parasitoidÕs response to odours of egg-carrying pine twigs. Ó 2004 Elsevier Ltd. All rights reserved. Keywords: Pinus nigra; Pinus sylvestris; Pinaceae; Sawfly; Egg parasitoid; Monoterpenes; Sesquiterpenes; Terpenoids; Insect oviposition 1. Introduction Like all plants, coniferous trees of the genus Pinus are under a continuous threat by numerous herbivo- rous insects and pathogens attacking nearly all parts and tissues of the plant. To defend themselves, pines produce large amounts of oleoresin that is accumulated in a highly developed network of specialized resin ducts, which are distributed in the wood, bark, and nee- dles (Gijzen et al., 1993; Trapp and Croteau, 2001). Compared to other conifer taxa, pines produce and accumulate large amounts of oleoresin constitutively (Gijzen et al., 1993; Lewinsohn et al., 1991; Trapp and Croteau, 2001). In addition, it has been shown that defence reactions in pines can be induced by attacking herbivores or pathogens (Bonello et al., 2001; Krokene et al., 2000; Litvak and Monson, 1998; Popp et al., 1995; Raffa, 1991; Trewhalla et al., 1997). Conifer resin is composed of a complex blend of terpenoids consist- ing of a volatile turpentine fraction [monoterpenes (C 10 ), sesquiterpenes (C 15 )], and a less volatile so-called rosin fraction [diterpene resin acids (C 20 )], and their derivatives (Gershenzon and Croteau, 1991; Trapp and Croteau, 2001). 0031-9422/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.phytochem.2004.10.010 * Corresponding author. Tel.: +49 30 838 55913; fax: +49 30 838 53897. E-mail address: hilker@zedat.fu-berlin.de (M. Hilker). www.elsevier.com/locate/phytochem Phytochemistry 65 (2004) 3221–3230 PHYTOCHEMISTRY