Glucose Formation During Biodegradation of Kraft-pretreated Sawdust from the Lagos Area, Nigeria

Transcription

1 Article International Journal of Environment and Bioenergy, 2013, 5(3): International Journal of Environment and Bioenergy Journal homepage: ISSN: Florida, USA Glucose Formation During Biodegradation of Kraft-pretreated Sawdust from the Lagos Area, Nigeria N. A. Ndukwe 1, W. O. Okiei 1, B. I. Alo 1, J. P. H. Van Wyk 2, *, T. M. Mamabolo 2 and C. C. Igwe 3 1 Department of Chemistry, University of Lagos, Akoka, Lagos, Nigeria. 2 Department of Pharmacology and Therapeutics, Medunsa Campus, University of Limpopo, South Africa. 3 Chemical, Fibre and Environmental Technology Department, Federal Institute of Industrial Research, Oshodi, Lagos, Nigeria. * Author to whom correspondence should be addressed; vanwykz@yahoo.com Article history: Received 19 February 2013, Received in revised form 17 March 2013, Accepted 25 March 2013, Published 27 March Abstract: The development of an effective and low-cost biomass to ethanol system has gained global recognition as the depletion of non-renewable energy resources and negative environmental issues surrounding fossil fuel combustion is increased. Environmental pollution and increasing health hazards associated with growing volumes of wood waste generated along the Lagos Lagoon in Nigeria has grown to unbearable limits. Sawdust from twenty different wood species along the Lagos Lagoon was subjected to Kraftpretreatment in solubilizing its lignin component thus releasing cellulose fibers susceptible for saccharification by Trichoderma viride cellulase. The amount of sugars released was calculated after 30 min, 1 h, 3 h and 6 h of incubation with the cellulolytic enzyme. Different sugar releasing patterns were obtained from the various sawdust samples and after 6 h of degradation the highest sugar concentration of mg.ml -1 was released from Lophira alata cellulose while the lowest concentration of 2.30 mg.ml -1 resulted during the hydrolyses of Milicia excels cellulose. Keywords: Sawdust, Trichoderma viride cellulase, Kraft-delignification, bioenergy, lignocellulose.

2 Introduction Lignocellulosic biomass is the most abundantly natural renewable resource and ubiquitous in most parts of the globe (Zheng et al., 2009). The release of fermentable sugars from lignocellulosic waste for bioenergy production has received great industrial interest and is widely considered an optimal feedstock for production of biofuels and high value added products (Van Wyk, 2001). Cellulose is the main biopolymer component of lignocelluloses and due its recalcitrance characteristic, environmental pollution caused by the accumulation of cellulosic biomass has become a global phenomenon (Lynd et al., 2005). It is estimated that the global annual production of lignocellulosic biomass is tons, representing J of energy, ten times greater than the J released from annual petroleum production thus making waste lignocellulose the most abundant renewable energy resource (Binder and Raines, 2010). Lignocellulosic biomass from primary sources such as corn stover, wheat straw, the tops, limbs and bark of trees, as well as secondary biomass sources like wood sawdust represents a much underutilized source of bioenergy and chemical feedstocks. With the growing concerns about the rapid dwindling of fossil fuel reserves coupled with unstable oil prices for the foreseeable future, lignocellulosic biomass materials remain a sustainable alternative and renewable energy resource. The production of fermentable sugars from cellulosic waste is a feasible process as described for the cellulolytic action of sole cellulases as well as combinations of different cellulase systems such as Trichoderma viride and Aspergillus niger (van Wyk, 2011). Much of the current energy demand and feedstocks for bioethanol production could be met by the exploitation of lignocellulosic wastes. Sawdust pollution is a major concern in Lagos, Nigeria as many wood mills are located along the lagoon. During chopping of trees by the local population for manufacturing of wood products huge volumes of sawdust are produced and due to a lack in pollution management these biomass waste accumulates on the banks of the lagoon. To a certain extent this solid waste ends up in the water of the lagoon disturbing the natural eco system. The most popular means of eliminating the excess lignocellulose waste is combustion which aggravates air pollution with negative effects on the local populations health. Sawdust samples from 20 different trees along the Lagos Lagoon have been collected, exposed to delignification by means of Kraft pretreatment and finally the delignified celluloses were saccharified by T.viride cellulase. The relative amount of fermentable sugars released from delignified celluloses during degradation has been determined. 2. Materials and Methods

3 Lignocellulosic Samples The common Nigerian names and botanical names (in brackets) of sawdust bioconverted during this investigation are as follows (Ndukwe et al., 2009): Erunobo (Erythropleum suaveolens), Okilolo (Symphona globulifera), Erimado (Ricindendron heudelotii), Oporoporo (Pterygota macrocarpa), Iroko (Milicia excels), Odoko (Ipomoea asarifolia), Abura (Hallea ciliate), Itara (Sacoglottis gabonensis), Akomu (Pycnanthus angolensis), Afara (Terminalia superb), Ofun (Avicennia germinans), Obeche (Triplochiton scleroxylon), Akun (Uapaca guineensis), Opepe (Nauclea diderrichii), Masonia (Masonia altissima), Agba (Entada gigas), Some (Ceiba pentadra), Mahogany (Khaya Ivorensis), Eki-Eki (Lophira alata), Itako (Strombosia pustulata). Prior to delignification all wood samples were dehydrated at 105ºC and these dried samples (2 kg, mm particle size) were subjected to Kraft-pulping (350 g NaOH and 140 g Na 2 S dissolved in 8 L water) followed by delignification in a rotary steel digester at 170ºC at a pressure of 200 kpa for 1 h 45min at a cooking liquor to wood ratio of 4:1. After pre-treatment these delignified lignocellulose fibers were washed with deionized water until free of the Kraft reagents. The pulp air dried samples of delignified cellulose were then treated with a domestic blender to regain their original sawdust particle size Cellulase Enzymes and Sugar Determination Kraft pulped sawdust (10 mg) were mixed separately in triplicate with 0.40 ml Tris (hydroxymethyl aminomethan, 0.005M) buffer solution, ph 4.5. The enzymatic hydrolysis was carried out with 100 µl of a Trichoderma viride [EC ] cellulase solution (10.0 mg.ml -1 ) at an incubation temperature of 40ºC for incubation periods of 30 min, 1h, 3h and 6h. The amount of total reducing sugars released during cellulase action on the delignified celluloses was determined by the dinitrosalicyclic acid (DNS) method at 546 nm using glucose as a standard (Miller, 1959). 3. Results and Discussion The chemical interaction between lignin and cellulose in plant materials as well as the structural composition of cellulose are major stumbling blocks during the enzymatic catalyzed degradation of cellulose into fermentable sugars thus complicating the development of lignocellulose as a resource of bioenergy. Delignification of lignocellulose with the Kraft procedure make cellulose more susceptible for cellulase catalyzed degradation into fermentable sugars (Santos et.al., 2011) and the relative susceptibility of these twenty (20) different delignified Nigerian hardwoods for T. viride cellulase catalyzed bioconversion has been reported (Ndukwe et. al., 2012). This paper however

4 159 reveals the different bioconversion profiles in terms of amount of sugar released from the various delignified celluloses after incubation periods of 30 min, 1 h, 3 h and 6 h (Table 1) with T. viride cellulase. Table 1. Formation of reducing sugars (mg.ml -1 ) during the T. viride cellulase catalyzed bioconversion of Kraft pretreated sawdust waste from twenty different trees along the Lagos Lagoon in Nigeria. Sample 30 min 1h 3h 6h Erunobo (Erythropleum suaveolens) Okilolo (Symphona globulifera) Erimado (Ricindendron heudelotii) Oporoporo (Pterygota macrocarpa) Iroko (Milicia excels) Odoko (Ipomoea asarifolia) Abura (Hallea ciliate) Itara (Sacoglottis gabonensis) Akomu (Pycnanthus angolensis) Afara (Terminalia superb) Ofun (Avicennia germinans) Obeche (Triplochiton scleroxylon) Akun (Uapaca guineensis) Opepe (Nauclea diderrichii) Masonia (Masonia altissima) Agba (Entada gigas) Some (Ceiba pentadra) Mahogany (Khaya Ivorensis) Eki-Eki (Lophira alata) Itako (Strombosia pustulata) The highest concentration of sugar released after 30 min of incubation was obtained during saccharification of delignified cellulose from Nauclea diderrichii at a concentration of 4.97 mg.ml -1 followed by Triplochiton scleroxylon cellulose (4.83 mg.ml -1 ) and cellulose from Strombosia pustulata (3.71 mg.ml -1 ). The lowest sugar concentration was obtained with cellulose from Ricindendron heudelotii at a concentration of 1.03 mg.ml -1. This relative low sugar concentration was

5 160 five (5) times less than sugar produced at the highest concentration. The average sugar concentration produced after 30 min of incubation from all the delignified celluloses was calculated at a concentration of 2.50 mg.ml -1 with 10 of the sawdust samples producing more than this average sugar concentration. The highest sugar concentration after 1 h of cellulase catalyzed biodegradation was again obtained from Nauclea diderrichii cellulose at a concentration of 6.95 mg.ml -1 followed by a lower concentration of 6.68 mg.ml -1 released from delignified Triplochiton scleroxylon cellulose. The third highest sugar concentration was released during the degradation of cellulose from Lophira alata at a concentration of 5.16 mg.ml -1. At a concentration of 1.50 mg.ml -1 cellulose from Milicia excels released the lowest concentration of sugar that was 4.6 times less than the highest sugar concentration released after 1 h of saccharification. The average concentration of sugar released after 1 h of degradation was calculated at 3.47 mg.ml -1 with thirteen sawdust samples releasing sugar at a concentration less than this average value. The average amount of sugar released after 1 h of degradation was 1.4 times higher than the average sugar concentration released after 30 min of incubation with the highest sugar concentration produced after 1 h of incubation 1.4 times higher than the highest sugar concentration released after 30 min of incubation. The lowest sugar concentration released after 1 h of incubation was 1.5 times higher than the lowest amount of sugar released during 30 min of cellulase catalyzed bioconversion of the cellulose. After 3 h of T. viride cellulase catalyzed bioconversion the highest sugar concentration was obtained from Triplochiton scleroxylon cellulose at a concentration of mg.ml -1 followed by a concentration of 9.94 mg.ml -1 released by Nauclea diderrichii cellulose. The third highest degree of saccharification was calculated for Lophira alata cellulose which resulted in a sugar concentration of 8.52 mg.ml -1 while the lowest concentration of 2.08 mg.ml -1 was obtained from Milicia excels that was five (5) times less than the highest sugar concentration. The average sugar concentration produced during three hours of bioconversion was 5.27 mg.ml -1 with thirteen (13) samples producing sugar concentrations less than the average sugar concentration. The highest amount of sugar obtained after 3 h of incubation was 1.5 times more than the highest sugar concentration obtained after 1 h of incubation and double the concentration obtained after 30 min of incubation. At the lowest sugar concentration level the sugar concentration obtained after 3 h of incubation was 1.4 times higher than the concentration calculated during the one hour incubation period and double the lowest sugar concentration produced after 30 minutes of cellulase catalyzed degradation. The average concentration of sugar released after three (3) h of cellulase catalyzed hydrolysis was 1.5 times higher than the average sugar amount obtained after one (1) hour of incubation and 2.1 times the sugar concentration obtained after 30 min of incubation.

6 161 When these different Kraft-pretreated sawdust samples were bioconverted with T. viride cellulase during a six (6) h incubation period the maximum degree of degradation was observed with cellulose from Lophira alata producing sugars at a concentration of mg.ml -1 followed by a concentration of mg.ml -1 obtained from Terminalia superb cellulose. The third highest extent of saccharification was calculated for sugars released by Pycnanthus angolensis cellulose at a concentration of mg.ml -1 while the lowest amount was released from cellulose from Milicia excels at a concentration of 2.30 mg.ml -1. This lowest sugar concentration was almost six (6) times less than the highest sugar concentration. After six (6) hours of incubation an average sugar concentration of 8.10 mg.ml -1 was calculated with ten sawdust celluloses releasing sugars at concentrations higher than the average value. The highest sugar concentration was 1.3 times more than the maximum amount of sugar released after 3 h of degradation, 1.9 times more than the amount obtained after 1 h of incubation and 2.7 times more than the maximum sugar concentration obtained after 30 min of incubation. The lowest sugar concentration after 6 h of degradation resulted in a value that was 1.1 times higher than the amount of sugar released after 3 h, 1.5 times more than the lowest sugar production after 1 h and 2.2 times more than the concentration after 30 min of T. viride cellulase catalyzed degradation. These different sugar releasing patterns from the twenty different Kraft-pretreated lignocellulosic wood waste materials could be explained in terms of the following. (i) Lignin not completely removed from the cellulose as a result of its chemical associations with cellulose. Due to its complex structure lignin is the most recalcitrant component of lignocellulose leading to the inability of pretreatment chemicals to completely remove it from the lignocellulosic substrates and as a result the enzymatic hydrolysis cellulose into glucose is less effective (Baptista et al., 2008). (ii) The chemical structure of pure cellulosic consists of two sections, an amorphous component that is susceptible for cellulase attack and a crystalline section offering more resistance to the hydrolytic action of the cellulase enzymes. The relative amount of these two structural components will also influence the efficiency of cellulose bioconversion into glucose (Habibi et. al., 2010). (iii) The complex composition of the cellulase enzyme is another variable that could influence the efficiency of cellulose degradation. Cellulase is a multi-component enzyme system of which each component performs a specific role during the saccharification procedure. The relative presence of each component in a specific cellulase system will also affect the biodegradation of cellulose into fermentable sugars (Jeoh et al., 2007).

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