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Uetimane Jr., E., Jebrane, M., Terziev, N., and Daniel, G. (2018). "Comparative wood anatomy and chemical composition of Millettia mossambicensis and Millettia stuhlmannii from Mozambique," BioRes. 13(2), 3335-3345.

Abstract

The wood anatomy and chemistry of a relatively lesser used wood species, known in Mozambique as nsangala (Millettia mossambicensis J. B. Gillett), was compared to overexploited species jambire (Milletia stuhlmannii Taub.) to provide diagnostic features for safe discrimination. The anatomical results showed that both species shared several similarities such as intervessel pitting size range (8 µm to 11 µm), rays composed of only procumbent cells, fiber dimensions (average length up to 1359 µm and wall thickness up to 10 µm), and banded axial parenchyma. The extractives and lignin content were higher in jambire, while the carbohydrates and acetyl contents were higher in nsangala. The main anatomical feature separating the two species was the porosity pattern with semi-ring porous wood of nsangala compared to the diffuse-porous structure of jambire. Jambire had wider vessel lumina (200 µm) and up to 3 vessels/mm2 compared to nsangala vessel lumina of 86 µm and a frequency of 37 vessels/mm2.


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Comparative Wood Anatomy and Chemical Composition of Millettia mossambicensis and Millettia stuhlmannii from Mozambique

Ernesto Uetimane, Jr.,a Mohamed Jebrane,b Nasko Terziev,b,* and Geoffrey Daniel b

The wood anatomy and chemistry of a relatively lesser used wood species, known in Mozambique as nsangala (Millettia mossambicensis J. B. Gillett), was compared to overexploited species jambire (Milletia stuhlmannii Taub.) to provide diagnostic features for safe discrimination. The anatomical results showed that both species shared several similarities such as intervessel pitting size range (8 µm to 11 µm), rays composed of only procumbent cells, fiber dimensions (average length up to 1359 µm and wall thickness up to 10 µm), and banded axial parenchyma. The extractives and lignin content were higher in jambire, while the carbohydrates and acetyl contents were higher in nsangala. The main anatomical feature separating the two species was the porosity pattern with semi-ring porous wood of nsangala compared to the diffuse-porous structure of jambire. Jambire had wider vessel lumina (200 µm) and up to 3 vessels/mm2 compared to nsangala vessel lumina of 86 µm and a frequency of 37 vessels/mm2.

Keywords: Chemical composition; Illegal logging; Millettia mossambicensis; Millettia stuhlmannii; Wood anatomy

Contact information: a: Eduardo Mondlane University, Forestry Department, Box 257, Maputo, Mozambique; b: Department of Forest Products, Swedish University of Agricultural Sciences, Box 7008, 750 07 Uppsala, Sweden; *Corresponding author: nasko.terziev@slu.se

INTRODUCTION

Wood identification based on anatomical features remains a reliable method in the overall control process of legal timber trade across the globe, but often the identification is only possible down to the genus level (Wheeler and Baas 1998; Hermanson and Wiedenhoeft 2011; Johnson and Laestadius 2011)In a country with a range of very similar commercial timbers, the identification process based on general knowledge is not sufficient. Apart from wood anatomy and dendrochronology, several other methods of wood identification are also employed, namely chemical (near infrared spectroscopy, mass spectrometry, detector dogs, stable isotopes, and radiocarbon), genetic (e.g., DNA barcoding, DNA fingerprinting, population genetics), and phylogeography methods (Dormontt et al. 2015; UNODC 2016).

Every method has its own merits and limitations, and some are complimentary; therefore an integration of methods is recommended (Dykstra et al. 2002; Dormontt et al. 2015). Most of the methods previously mentioned above rely on detailed databases from where models and references are used to cross-check unknown wood samples (Beeckman 2003). Examples include large databases of wood collections, and xylariums consulted by experts and law enforcement officers to inspect both the legality and authenticity of traded wood, especially for either endangered or protected timbers under international treaties such as Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES) and World Wide Fund for Nature (WWF) (United Nation Office on Drugs and Crime-UNODC 2016).

Currently, Mozambique’s forest authorities are discussing the possibility of banning the export of logs regardless of their commercial class, which means that all harvested wood species have to be processed prior to exportation. Therefore, the high similarity amongst sawn timber boards of many wood species in Mozambique will pose new challenges to both customs and law enforcement officers. The timber of jambire (Milletia stuhlmannii) together with a few other species are the most sought after timbers under highly selective logging harvesting regimes that have reduced considerably its growing stock in Mozambique. The forest authorities are also reducing the harvesting quota for this group of species to halt depletion of the growing stock. Thus, the demand for timbers is now extending to relatively lesser-known species, including protected timbers to which very little is known or attention has been paid previously (Uetimane Jr. 2010). Comparative wood anatomy studies of similar species are therefore important to ensure differentiation (Gasson 2011; Maiti et al. 2016) as well as allowing customs officials to correctly enforce tax duties and restrict or ban trade of both endangered and protected timbers (Wheeler and Baas 1998).

Timber traders and consumers must be assured on the authenticity of the timbers. It is expected that by reducing both intentional and accidental misclassification of valuable timbers, the government will gain increased revenues of correct duties paid by stakeholders of the timber sector (Dykstra et al.2002). This study is therefore aimed at comparing the wood chemical composition and anatomy of jambire (Millettia stuhlmannii) and its closest sister nsangala (Millettia mossambicensis). Unlike jambire, nsangala is an endemic tree species (in the central region of Mozambique) that has not been studied apart from dendrological and botanical descriptions by Hyde et al. (2017). On the other hand, jambire is widespread across southern Africa, and its wood anatomy and properties are well known and described by several authors (Lemmens 2008; Ali et al 2008; Richter and Dallwitz 2000). The same authors reported that the timber of jambire is very dense (720 to 990 kg m˗³ at 12% moisture content), very durable heartwood, fair workability, dimensionally stable, and mostly used for flooring, framing and luxury furniture. This study is part of a large database project to describe wood anatomical features separating similar timbers to assist with the verification system through a reliable method in the context of reducing the large amount of endangered or protected timbers in the market, thereby prompting conservation of the forests across the country.

EXPERIMENTAL

Materials

The heartwood samples (10×60×200 mm) of M. mossambicensis J. B. Gillett were taken from the herbarium Catapu (listed as number 115, identified and catalogued by Meg Coates Palgrave, Caia, Sofala, Mozambique). The natural habitat of M. mossambicensis is well-described by Remane and Therrell (2015). Heartwood sample size of 20×140×220 mm (in tangential, radial and longitudinal direction) along with reference permanent microscope slides of M. stuhlmannii Taub. were obtained from the Eduardo Mondlane University xylarium (Maputo, Mozambique) listed with numbers 17 to 25-(05/68).

Microscopy Methods

The transverse, radial, and tangential sections (ca. 20 to 25-mm-thick) of M. mossambicensis and additional microscope slides of M. stuhlmannii were prepared using a sliding microtome, bleached in 2 to 3% sodium hypochlorite and stained with 1% w/v aq. Astra blue, followed by 1% safranin in 50% ethanol, dehydrated and mounted in synthetic resin. Fiber and vessel morphological analyses were conducted after maceration using Franklin’s method, as modified by Kraus and Arduin (1997). The prepared slides were examined in light microscopy (LM). Scanning electron microscopy (SEM) was used to examine the fibre wall thickness, lumen widths (fibers and vessels), and structure of vestured pits of M. mossambicensis. For SEM, semi-thin sections were air-dried, mounted on stubs, coated with gold using a Emitec 5000 (EMITEC, Lohmar, Germany) sputter coater, and observed using a FEI (Philips) XL30 ESEM (Now Thermo Fisher Scientific, Waltham, USA) at 15 kV (Daniel et al. 2004). Terminology, definitions, and measurements were performed according to recommendations of the IAWA list of microscopic features (1989).

Micrograph analysis and measurements were processed through Image Pro-Plus Premier software (Media Cybernetics, Inc., version 7, Washington D.C., USA). The microscope slides of M. stuhlmanniiwere mainly used to take micrographs by light microscopy (LM) and cross check the anatomical descriptions previously published (Richter and Dallwitz 2000; Ali et al. 2008; Lemmens 2008).

Chemical Analyses

Extractives content

Wood samples were cut from boards and milled to pass a 40-mesh screen. The milled wood was homogenized and 5 g were taken for extraction. Each sample was extracted in a Soxhlet apparatus with a mixture of toluene/ethanol (2/1; v/v) for 6 h, acetone for 6 h, and water for an additional 6 h, and dried at 103 ºC for 24 h.

Lignin, monosaccharides, and acetyl content

Millettia mossambicensis J. B. Gillett and Millettia stuhlmannii Taub. wood samples were analyzed for acid soluble lignin (ASL), acid insoluble lignin (AIL), monosaccharides, and acetyl content (AC) according to the procedure of Sluiter et al. (2008). The ASL was determined using a Hitachi U-2910 spectrophotometer (Hitachi, Tokyo, Japan) with an absorptivity of 110 L g-1 cm-1 at a wavelength of 205 nm. The monomeric carbohydrates were determined using a Chromaster high-performance chromatography (HPLC; Hitachi, Tokyo, Japan) system equipped with an evaporative light scattering detector (ELSD-90; VWR International GmbH, Darmstadt, Germany), and a Metacarb 87P column (300 mm × 6.5 mm; Santa Clara, CA, USA) with a guard column (Metacarb 87P 50 mm × 4.6 mm). The ELSD-90 was operated at 50 ºC, 2.5 bars, and N2 was used as the nubilizing gas.

The sugars were eluted using ultra-pure water as a mobile phase at a constant flow rate of 0.5 mL min-1 and column temperature of 85 ºC. The acetyl content was determined using a diode array detector (DAD; Hitachi, Tokyo, Japan) operated at 210 nm, and a Metacarb 87H column with a guard column (MetaCarb 87H 50 mm × 4.6 mm). The mobile phase was 0.005 mol L-1 H2SO4 solution (pH 2.1), with a flow rate of 0.6 mL min-1 at 30 ºC.

RESULTS AND DISCUSSION

Wood Anatomical Descriptions

Millettia mossambicensis J. B. Gillett

The growth rings’ distinct boundaries are invariably marked by both smaller vessels and banded axial parenchyma. The heartwood is distinctively dark compared with the pale sapwood, and the grain of the wood is slightly wavy (Fig. 1B).

Fig. 1. Macro features of jambire (A-arrows showing yellow stripes of axial parenchyma) and nsangala (B-arrows showing wavy grain)

The vessels were dispersed as diffuse pores, averaging 37 mm-2 vessels in transverse sections (range of 23 mm-2 to 49 mm-2). The solitary vessel tangential lumen diameter averaged 86 µm (range of 43 µm to 255 µm). There was a semi-ring porosity pattern observed. In transverse sections (TS), there are also multiple vessels along the rays (often clusters of ≥ 4 vessels).