Analysis of commercial proanthocyanidins. Part 1: The chemical composition of quebracho (Schinopsis lorentzii and Schinopsis balansae) heartwood extract Pieter B. Venter, Mirek Sisa, Marthinus J. van der Merwe, Susan L. Bonnet, Jan H. van der Westhuizen ⇑ Department of Chemistry, University of the Free State, Nelson Mandela Avenue, Bloemfontein 9301, South Africa article info Article history: Received 4 April 2011 Received in revised form 23 June 2011 Available online 5 November 2011 Keywords: Schinopsis lorentzii and Schinopsis balansae Anacardiaceae Quebracho Electrospray mass spectrometry Proanthocyanidins Natural polymer abstract Quebracho (Schinopsis lorentzii and Schinopsis balansae) extract is an important source of natural poly- mers for leather tanning and adhesive manufacturing. We combined established phyto- and synthetic chemistry perspectives with electrospray mass spectrometry experiments to prove that quebracho pro- anthocyanidin polymers consist of an homologous series of flavan-3-ol based oligomers. The starter unit is always catechin which is angularly bonded to fisetinidol extender units. By comparison of the MS 2 frag- mentation spectra of the oligomer with product ion scans of authentic catechin and robinetinidol sam- ples, we proved that quebracho extract contains no robinetinidol, as is often reported. Quebracho proanthocyanidins have acid resistant interflavanyl bonds, due to the absence of 5-OH groups in fisetin- idol, and the aDP cannot be determined via conventional thiolysis and phloroglucinolysis. We used the MS data to estimate a conservative (minimum value) aDP of 3.1. Ó 2011 Elsevier Ltd. All rights reserved. 1. Introduction The wild quebracho forests in the Gran Chaco region of Argen- tina, Bolivia, and Paraguay have been harvested for more than 100 years as an important source of vegetable tannins and timber. The timber is durable and extremely hard and the name quebracho is derived from the Spanish word quiebrahacha which means ‘‘axe- breaker’’. To obtain a warm water soluble quebracho extract, the heartwood is stripped of its bark, chipped, and extracted with boil- ing water. A cold water soluble extract (sulfited extract) is obtained upon treatment of the warm water soluble extract with bisulfite or direct extraction of wood chips with a boiling aqueous bisulfite solution. Higher extraction rates are obtained with boiling aqueous bisulfite solution than with boiling water alone. Quebracho extract is obtained from Schinopsis balansae (red ‘‘chaqueno’’ quebracho, pure tannin content 20–21%) from the Eastern Chaco region and Schinopsis lorentzii (red ‘‘santiagueno’’ quebracho, pure tannin content 15–18%) from the Western Chaco region. These two species were previously referred to as Quebracho colorado chaqueño and Quebracho colorado santiagueño (Schinopsis quebracho-colorado) and belongs to the family Anacardiaceae. A third tree species, Aspidosperma quebracho-blanco of the family Apocynaceae, is commonly referred to as white quebracho. Quebracho extract consists of about 95% proanthocyanidins (PAs) and 5% water soluble sugars on a dry basis. The term pro- anthocyanidin (PA) refers to the characteristic development of a red color upon heating PAs with dilute acid (Roux, 1992). PAs are also referred to as condensed tannins to distinguish them from hydrolysable tannins which do not produce a red color when heated with aqueous acid. Hydrolysable tannin oligomers are esters of gallic acid and D-glucose. Important industrial sources of PAs are mimosa bark extract (Acacia mearnsii) and quebracho heartwood extract, and of hydrolysable tannins, tara pods, chest- nut bark, and oak gall extracts. Progress in defining quebracho PA composition has been slow, mainly due to the complexity of the extracts and the difficulty of isolating pure PAs with silica gel based chromatography materials. Uncertainties include different hydroxylation patterns of the con- stituent flavan-3-ol aromatic rings, different configurations at the C-2, C-3 and C-4 stereogenic centers, the possibility of a second ether interflavanyl bond (A-type PAs), the average chain length (degree of polymerization), and the presence of angular oligomers. Progress is further hampered by the absence of 5-OH groups in the constituent monomers, which imparts stability to the interflav- anyl bond against acid hydrolysis (Roux and Paulus, 1962; Roux et al., 1975). This renders the classical method to analyse PAs via acid hydrolysis of the interflavanyl bond and subsequent trapping of intermediates with toluene-a-thiol or phloroglucinol (thiolysis and phloroglucinolysis) (Thompson et al., 1972; Foo and Porter, 1978; Kennedy and Taylor, 2003; Rigaud et al., 1991) and analysis 0031-9422/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.phytochem.2011.10.006 ⇑ Corresponding author. Tel.: +27 51 4012782; fax: +27 51 4448463. E-mail address: vdwestjh@ufs.ac.za (J.H. van der Westhuizen). Phytochemistry 73 (2012) 95–105 Contents lists available at SciVerse ScienceDirect Phytochemistry journal homepage: www.elsevier.com/locate/phytochem