487 M. Bento et al.: Fragmentation of Cork Suberin Introduction Suberin is the main structural component of cork from Quercus suber L. (Pereira 1988). It is located in the cell wall in close association with other structural components, e.g. with lignin (Marques et al. 1996, 1998). The macromolecular structure of suberin is still largely unknown as well as its chemical relationship to the aro- matics in the cell wall and this has led to some controver- sy about the distinction between suberin and other struc- tural components (Kolattukudy and Espelie 1985, 1989; Holloway 1972, 1983; Agulló and Seoane 1982; Guille- monat and Strich 1953; Graça and Pereira 1997, 2000; Marques et al. 1996). Most studies on the chemical com- position of suberin were done after depolymerisation of monomeric units and GC-MS analysis. Depolymerisation using a 3 % NaOMe catalysed methanolysis has allowed identification of the suberinic monomers as esters deriva- tives of saturated alkanoic acids, saturated and unsaturated a, v-alkanedioic acids, saturated and unsaturated v- hydroxyalkanoic acids, epoxyoctadecanedioic acid, mono- hydroxy-epoxyoctadecanoic acid, dihydroxyoctadecane- dioic acid and trihydroxyoctadecanoic acid (Holloway 1983; Arno et al. 1981; Graça and Pereira 2000). Glycerol is also an important component of the suberin methanoly- sis products (Graça and Pereira 1997, 2000). A thermally assisted transmethylation applied directly to cork released the same monomers (Bento et al. 1998). Glyceryl-acyl dimers were recently identified in the products released from cork with a very mild depolymeri- sation procedure and a glyceryl-based structure was pro- posed for suberin (Graça and Pereira 1997, 1999). In this paper we used methanolysis under different cata- lyst concentrations in order to depolymerise suberin to varying extents and analysed the composition of the monomers released as well as the molecular weight and size characteristics of the co-solubilised larger fragments. Mass spectrometric analysis of polymers needs a soft ionisation technique to minimize the fragmentation of molecular ions. Matrix-assisted laser desorption and ionisa- tion (MALDI) is a state-of-art ionisation technique which has been used to analyse synthetic polymers (Bahr et al. 1992) and biomolecules (Karas and Chait 1991; Li et al. 1996). Samples for analysis are prepared by diluting them in a suitable low molecular weight matrix that highly absorbs the UV irradiation. The bombardment of the sam- ples with a short pulse of laser light promotes the fraction of the matrix and the desorption and ionisation of the sam- ple molecules. Coupling MALDI with Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR-MS), a bet- ter resolution and mass accuracy are achieved in principle (Marshall and Schweikhard 1992; Heeren and Boon 1996). Holzforschung / Vol. 55 / 2001 / No. 5 © Copyright 2001 Walter de Gruyter · Berlin · New York Holzforschung 55 (2001) 487–493 Fragmentation of Suberin and Composition of Aliphatic Monomers Released by Methanolysis of Cork from Quercus suber L., Analysed by GC-MS, SEC and MALDI-MS By M.F. Bento 1,2 , H. Pereira 3 , M.Á. Cunha 1 , A.M.C. Moutinho 1,4 , K.J. van den Berg 5 , J. J. Boon 5 , O. van den Brink 5 and R.M.A. Heeren 5 . 1 Centro de Física Molecular da UTL, Instituto Superior Técnico, Lisboa, Portugal 2 Instituto Superior de Engenharia de Lisboa, Lisboa, Portugal 3 Centro de Estudos Florestais, Instituto Superior de Agronomia, Lisboa, Portugal 4 CeFiTec, Dep. de Física, Fac. de Ciências e Tecnologia, Universidade Nova de Lisboa, Monte de Caparica, Portugal 5 FOM Institute for Atomic and Molecular Physics, Amsterdam, The Netherlands Summary Suberin from extractive-free cork from Quercus suber L. was depolymerised by methanolysis using dif- ferent sodium methanolate (NaOMe) concentrations. 1 % and 3 % NaOMe completely removed suberin from cork (54 %–56 % of extractive-free cork), but for lower concentrations there was incomplete solu- bilisation; with 0.05 % NaOMe, only approximately 80 % of total suberin was removed. The monomer- ic composition of the extracts differed significantly: for the 0.05 % NaOMe, only alkanoic acids and diacids were found; the yield of v-hydroxy acids increased with reactant concentration, as well as alka- nols and ferulic acid. Results from light scattering, SEC and MALDI-MS showed that soluble oligomeric fragments con- taining suberinic acids were present in the methanolysis mixture. The molecular weight of these oligomeric fragments decreased with higher sodium methanolate concentrations. Keywords Suberin fragmentation Cork Quercus suber L. SEC MALDI-MS Methanolysis Authenticated | umutsen@isa.utl.pt Download Date | 9/12/13 7:31 PM