Doctoral Degree Proposal - Sample
Content
Doctoral Degree Research Proposal for Fellowship Programme
Topic: A Simultaneous Processing of Natural Products and Bioethanol as Zero-Waste Management of Cocoa Lignocellulosic Biomass
By Mr. V. Kumaran
1.0 Introduction Fulfilling the basic needs of food, water and shelter had always been one of the prime concerns of mankind. These three elements have played a crucial role in human civilization and in the pursuit of these elements man had negatively impacted the whole eco-balance (Samir, 2009). Agriculture has been the most important turn of event for human civilization ever since first human colony learnt to adapt to the surroundings and started living a non-nomadic life. The use of arable land for cultivation and food production has brought about socio-economic and environmental externalities (Jeong and Foster, 2003). In that aspect modern agriculture practices had increased food production yield tremendously. Parallel to this is the amount of waste generated from this large scale cultivation. It could be inferred from Samir (2009), Jeong and Foster (2003) that the increasing need for food production due to increasing growth of human population had led to the rampant use of fertilizers, pesticides, indiscriminate wastes discharges, contamination of water sources and other serious repercussions such as increased greenhouse gas release. In order to mitigate such damaging events, sustainable agriculture has been introduced over the last decades. rd In Malaysia, the 3 National Agricultural Policy (NAP3 1998-2010) focuses the need of such practices (Faridah, 2001). Nevertheless, the management of agro-waste is still one of the major concerns to the agro-1 -1 industries. Malaysia is the second largest agro-waste producer in Asia, with a production of 0.122 kgcap day while 1.2 million tonnes of agro-waste are land filled annually (Agamuthu, 2009). Globally, part of these waste materials have been used for animal feed and fertilizer (Mamma et al., 2008), however, a large portion of these materials is still deposited annually. This situation is not desirable due to both socio-economic and environmental issues such as waste-disposal health hazard, high transportation costs, insufficient disposal sites, and the land-filling material of high organic content (Tripodo et al., 2004). As a corollary, the extensive study for the use of wastes material as an input for renewable resources and green energy has made its way to the mainstream research community and industry (Dias et al., 2009; Kaparaju et al., 2009; Najafi et al., 2009; Delzeit and Holm-Muller 2009; Sassner, Galbe and Zacchi, 2008). The agro-waste, which is referred as biomass has been used for production of bioethanol and in derivation of functional phytochemicals (Dias et al., 2009; Sassner, Galbe and Zacchi, 2008). In this proposed research, cocoa pod will be used as the targeted agro-waste. The concept of zero waste management will be applied whereby cocoa pod waste volume will be reduced by recovering all resources, and not burning or burying them. Malaysia is known as the third largest producer of cocoa which produces about 200,000 tonnes of cocoa beans per annum (Wong, 2007; Sharifuddin & Zaharah, 2010). The pod, which weighs approximately 0.5 kg, is ovoid in shape with 15-30 cm in length and 8-10 cm in width. The pod
contains about 20-60 cocoa beans which are normally used for production of cocoa butter and chocolate. The pod, being the bulk waste (approximately 2 million tonnes per annum on 1:10 bean to pod weight ratio) however is either been discharged or used as fertilizer after the processing. Even at present, this agriculture residue has never been fully explored in terms of energy production in Malaysia. Extraction of functional components from these materials has also been substantially neglected. The confluence of these preceding points leads to the notion that the cocoa pod agro-waste has very high potential of being new renewable energy source as well as source of natural products in Malaysia, and thus closes the loop for zero-waste management. This will be the prime interest of this doctoral research.
2.0 Literature Review Based on the above introduction section, agro-waste in Malaysia has the potential of bio-energy derivation for zero-waste management. However an in-depth research would be required to ascertain the magnitude and potential value of the agro-waste conversion. Major part of this research focuses on using the lignocellulosic structure of the biomass as the main component for bio-energy conversion. However, in using lignocellulosic agro-waste as potential feedstock for bio-energy production, some of the following issues need to be addressed: (1) The appropriateness of agro-waste for bio-energy production due to its diversity; (2) The availability of value-added components such as functional fibres and polyphenols within crop residues, and (3) the possibility of extracting these value-added components prior to biofuel conversion. Answers to these issues must be based on eco-sustainable approach which is important to address such a way of producing bio-energy without wasting any treasures embedded within the agro-waste. As such, the preliminary literature review of this research focuses on identifying the general solutions currently available as to the above issues and compares the potential processes that could be utilised to carry out the research. The final literature review will encompass wider circle of analysis to produce the specific solution for the targeted agro-waste. Generally, the various reasons that would support the use of cocoa pod biomass for natural product recovery and bioethanol conversion is very much evident as stated by Figueira, Janick, BeMiller (1993). The use of cocoa pod waste as mulching media is detrimental to cocoa plantation especially when the pods become source of disease inoculums. The use of the pod as livestock feed is also restricted by presence of theobromine, which becomes toxic to the livestock at higher feed rate (LD50 of 1254 mgkg-1). These two factors simultaneously limit the disposability options of cocoa pod agro-waste using presently available methods. Studies conducted by Donkoh et al. (2004), Okai et al. (1984) and Adomako (1972) provide chemical composition of cocoa pods cultivated in Ghana. The composition of neutral detergent fibres (comprise of cellulose, hemicelluloses and lignin) account to approximately 522-644 gkg-1 of dry matter of the ripe pod, while pectin makes up 8-11% of ripe pod. A study by Adomafio et al. (2004) however indicates only 26.6% of fibre in the cocoa pod waste, while it lists substantial presence of starch, protein, reduced sugar and lipid. In the analysis by Adomako (1972), fibre content is 45.6% (w/w) for dried ripe pod. The varying results could be partly due to different method of assaying as well as different selection of cocoa pod (Adomako, 1972). These preliminary figures suggest the weighty presence of lignocellulosic components in the cocoa pod for bioethanol conversion. In order to make any bioethanol production cost effective, the cost of feedstock has to be as low as possible. It is interesting to note that the cost of bioethanol production is contributed more than one third by feedstock cost (Dien, Cotta and Jeffries, 2003; Batchelor et al., 1994). Since the feedstock intended for this research is a waste biomass, the cost of bioethanol production could be considerably lower. On the contrary
the cost of bioethanol production is relatively high for lignocellulosic conversion due to lower yield and higher cost of hydrolysis (Sun and Cheng, 2002). This could be compensated by the initial pre-treatment which is expected to extract valuable bioactive compounds that could serve as protein, pectin and antioxidants with monetary values. Pre-treatment of the biomass to solubilise cellulose and hemicellulose is the most significant determinant for successful lignocellulosic bioethanol conversion. This is due to the fact that pre-treatment determines the extent to and the cost at which cellulose and hemicellulose is converted to bioethanol (Balat, Balat and Oz, 2008; Alvira et al., 2010). The use of dilute-acid hydrolysis is the most inexpensive method, but only effective commercially if feedstock is considerably cheap (Alvira et al., 2010; Jeffries and Jin, 2000; Wyman, 1994) however the carbohydrates yield is low (Balat, Balat and Oz, 2008). The yield of sugar for bioconversion can be increased significantly by complementing dilute acid hydrolysis with (supercritical) organosolv process (Aravamuthan et al., 1989), which is more expensive and complex (Wyman, 1996). An additional step of the pre-treatment is the use of alcohol precipitation and lyophilisation to recover pectic-like polysaccharides and the potential antioxidants (Gan, Normaliza and Aishah, 2010), the reason having been mentioned in the previous paragraph. After the bioactive components recovery, the residual lignocellulosic will be treated either via simultaneous saccharification and fermentation (SSF) or separate hydrolysis and fermentation (SHF) process for bioethanol conversion. The SSF process has been used conventionally, but SHF provides an alternative mean with several other benefits (Soccol et al., 2010); however the use and selection of enzymes will depend on the actual analysis of pre-treated residue, since presence of 5-C sugar and 6-C sugar may need specific microbial species and reactor conditions (Okuda et al. 2008,) for high yield fermentation. Since cocoa pod fermentation has not been widely studied, there is little information for any particular microbe selection based on pre-treated sugar composition and its fermentation. Finally the extraction of bioethanol from the fermentation will be carried out and this will also be developed further based on final literature review. These would be one of the primary works in the research and therefore would constitute one of the elements of research hypotheses.
3.0 Aim and Objectives of the Research 3.1 Aim The aim of the proposed doctoral research is to apply zero-waste management concept by carrying out laboratory processing of cocoa pod biomass for recovery of valuable natural products followed by conversion of lignocellulosic biomass into bioethanol using both conventional and newer technology as per reviewed literatures. 3.2 Objectives The objectives of the research are: i) To evaluate application of inexpensive extraction methods to determine maximum yield of bioactive compounds; ii) To analyze and classify the bioactive compounds that has potential functional values per basis of biomass unit; iii) To perform an in depth literature review on the most suitable process(es) for cocoa pod biomass lignocellulosic maximum simple sugar recovery; iv) To carry out a laboratory simple sugar recovery process that has optimum yield; v) To develop bioethanol extraction method that would be in-line with current bio-refinery practice;
To compare the yields between SSF and SHF while determining the optimal process conditions for highest bioethanol yield; vii) To evaluate if extraction of functional bioactive compound(s) has any impact on overall bioethanol cost per unit biomass, and viii) To analyze the outcome if a zero-waste management has been achieved while meeting the other research aims. 3.3 Hypotheses to be investigated Based on the above objectives, the following are the hypotheses to be proven: i) The extraction of functional bioactive compounds in the biomass pre-treatment step assist in reducing effective bioethanol cost for cocoa pod lignocellulosic bioethanol production; ii) The yield of simple sugar from cocoa pod lignocellulosic mass could be optimized by using either conventional or combined conventional-contemporary method without adversely affecting bioethanol yield in the fermentation process, and iii) The cocoa pod biomass has sufficient bioethanol yield based on industry standards, biorefinery practices and meets the zero-waste management where the disposal of biomass to landfill or burning of residue is eliminated after final processing.
vi)
4.0 Research Methodology The research methodology would begin with literature review to identify majority of the information required to meet the research objectives. This will provide preliminary work basis to develop the system to meet all the analytical research requirements. This research comprises of three major phases: i) Production of bioactive components from cocoa pod using dilute-acid extraction and supercritical organosolv process. The collection of cocoa pod samples, preparation for assay, extraction and testing of the samples will be done based on set of standard test methods for anticipated compounds. This will also involve lyophilisation of sample and alcohol precipitation method for antioxidant and polysaccharides extraction. The use of supercritical organosolv process will be developed in-line with the second phase requirement. The results will be compared against figures available from the reviewed literatures. The bioactive components will be identified using FTIR (Fourier Transform Infra Red) spectra and chromatography. Production of bio-ethanol using cellulosic waste from cocoa pod which was pre-treated during the bioactive component extraction in Phase 1. In this step, the whole saccharification and fermentation will be carried out using SSF and SHF. Depending on the results from Phase 1 for lignocellulosic composition; and saccharification/hydrolysis of Phase 2 for sugar composition, the specific sets of operating condition for fermentation will be determined by laboratory scale set-up. The selection of fermentation microbe will be part of this phase based on literature review performed in the initial part of the research as well as the sugar composition determined in saccharification/hydrolysis step. Development of a simultaneous extraction-conversion processing line as the protocol for bio-refinery industry. This phase is to be developed based on detailed literature review and upon completion of Phase 1 and the trials during Phase 2. It will be based on standards applied in bio-refinery extractionconversion processes for bio-products from Phase 2.
ii)
iii)
Research Proposal Preliminary Gantt Chart
Preliminary References
1. Adomafio, N.A., I.K. Afeke, J. Wepeba, E.K. Ali and F.O. Quaye, 2004. Biomedical composition and in vitro digestibility of cocoa (Theobroma cacao) pod husk, cassava (Manihot esculenta) peel and plantain (Musa paradisiaca) peel. Ghana J. Sci., 44:29-38. Adomako, D., 1972. Cocoa pod husk pectin. Phytochemistry, 11:1145-1148. Agamuthu, P., 2009. Challenges and opportunities in agro-waste management: An Asian perspective. Inaugural Meetings of First Regional 3R Forum in Asia, 11-12 Nov 2009, Tokyo, Japan, http://www.uncrd.or.jp/env/spc/docs/1st_3r_forum_presentation/Session2_2e_Agamuthu.pdf Alvira, P., E. Thomas-Pejo, M. Ballestros and M. J. Negro, 2010. Pretreatment technologies for an efficient bioethanol production process based on enzymatic hydrolysis: A review. Bioresour Technol., 101(13):4851-4861. Aravamuthan, R., W. Chen, K. Zargarian and G. April, 1989. Chemicals from wood: Prehydrolysis/organosolv methods. Biomass, 20:263-276 Balat, M., H. Balat and C. Oz, 2008. Progress in bioethanol processing. Prog. in Ener & Comb. Sci., 34(5):551-573. Batchelor, S.E., P. Cook, E.J. Booth and K.C. Walker, 1994. Economics of bioethanol production from wheat in the UK. Ren Energy, 5(Pt.II):807-809. Delzeit, R. and K. Holm-Muller, 2008. Steps to discern sustainability criteria for a certification scheme of bioethanol in Brazil: Approach and difficulties. Energy, 34:662-668. Dias, M.O.S., A.V. Ensinas, S.A. Nebra, M.F. Rubens, C.E.V. Rossell and M.R.W Maciel, 2009. Production of bioethanol and other bio-based materials from sugarcane bagasse: Integration to conventional bioethanol production process. Chem Eng Res & Design, 87(9):1206-1216. Dien, B.S., M.A. Cotta, T.W. Jeffries, 2003. Bacteria engineered for fuel ethanol production: current status. Appl Microbiol Biotechnol., 63:258-256. Donkoh, A., C.C. Atuahene, B.N. Wilson and D. Adomako, 1991. Chemical composition of cocoa pod husk and its effect on growth and food efficiency in broiler chicks. Animal FS and Technol., 35:161-169. Faridah, A., 2001. Sustainable Agricultural System in Malaysia. Paper presented at Regional Workshop on Integrated Plant Nutrition (IPNS), Development in Rural Poverty Alleviation, 18-20 Sept. 2001, UN Conference Complex, Bangkok, Thailand. p.1-10. Figueira, A., J. Janick and J.N. BeMiller, 1994. Partial characterization of cacao pod and stem gums. Carbohydrate Polymers, 24:133-138.
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10. 11. 12.
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14. Gan, C.Y., H.A.M. Normaliza and A.L.Aishah, 2010. Physico-chemical properties of alcohol precipitate pectin-like polysaccharides from Parkia speciosa pod. Food Hydrocol., 24:471-478. 15. Jeffries T.W. and Y.S. Jin, 2000. Ethanol and thermo tolerance in the bioconversion of xylose by yeasts. Adv Appl Microbiol., 47:221-268. 16. Jeong, H. and L. Forster, 2003. Empirical investigation of agricultural externalities: effects of pesticide use and tillage system on surface water. Working paper AEDE-WP-0034-03, AEDE, Ohio State University. 17. Kaparaju, P., M. Serrano, A.B. Thomsen, P. Kongjan and I. Angelidaki, 2009. Bioethanol, biohydrogen and biogas production from wheat straw in a biorefinery concept. Bioresour Technol., 100(9): 2562-2568 18. Mamma, D., E. Kourtolgou and P. Christakolpoulos, 2008. Fungal multienzyme production on industrial by-products of citrus-processing industry. Bioresour Technol., 99:2373-2383 19. Najafi, G., B. Ghobadian, T. Tavakoli and T. Yusaf, 2008. Potential of bioethanol production from agricultural waste in Iran. Renewable & Sustainable Ener Revs., 13(6-7):1418-1427 20. Okai, D.B., R.A. Easter and G.R. Frank, 1984. The nutritive value of some non-conventional Ghanaian feed ingredients: nutrient composition and effects on the performance of weanling rats. World Rev. Anim. Prod., 20(2):11-16 21. Okuda, N., K. Ninomiya, Y. Katakura and S. Shiyo, 2008. Strategies for reducing supplemental medium cost in bioethanol production for waste house wood hydrolysate by ethanologenic Escherichia coli: Inoculum size increase and co-culture with Saccharomyces cerevisiae. Biosc and Bioeng., 105(2): 90-96 st 22. Samir, D., 2009. Understanding the global environment, 1 ed. Dorling Kindersley (India), p.1-38 23. Sassner, P., N. Galber and G. Zacchi, 2008. Techno-economic evaluation of bioethanol production from three different lignocellulosic materials. Biomass and Bioener., 32(5): 422-430 24. Sharifuddin, H.A.H. and A.R. Zaharah, 2008. Utilisation of organic waste and natural systems in Malaysia agriculture, http://www.infrc.or.jp/english/KNF_Data_Base_Web/PDF KNF Conf Data/C1-4-010.pdf 25. Soccol, C.R., L.P.S. Vandenberghe, A.B.P. Medeiros, S.G. Karp, M. Buckeridge, L.P. Ramos, A.P. Pitarelo, V. Ferreira-Leitao, L.M.F. Gottschalk, M.A. Ferrara, E.B. da Silva Bon, L.M.P. de Moraes, J.A. Araujo and F.A.G. Torres, 2010. Bioethanol from lignocelluloses: status and perspectives in Brazil. Bioresour Technol., 101(3):4820-4825 26. Sun, Y., J. Cheng, 2002. Hydrolysis of lignocellulosic materials for ethanol production: a review. Bioresour Technol., 83:1-11 27. Tripodo, M.M., F. Lanuzza, G. Micali, R. Coppolino and F. Nucita, 2004. Citrus waste recovery: a new environmentally friendly procedure to obtain animal feed. Bioresour. Technol., 91:111-115 28. Wong, C.Y.L., 2007. Development of Malaysia s agricultural sector: agriculture as an engine of growth. Paper presented at the ISEAS Conference on the Malaysian Economy: Development and Challenges , 2526 January 2007, ISEAS, Singapore. p.1-22 29. Wyman, C.E., 1994. Ethanol from lignocellulosic biomass: technology, economics and opportunities. Bioresour. Technol., 50:3-16 30. Wyman, C.E., 1996. Handbook of bioethanol: production and utilization, 1st ed. Taylor & Francis, USA. p.186-208
Topic: A Simultaneous Processing of Natural Products and Bioethanol as Zero-Waste Management of Cocoa Lignocellulosic Biomass
By Mr. V. Kumaran
1.0 Introduction Fulfilling the basic needs of food, water and shelter had always been one of the prime concerns of mankind. These three elements have played a crucial role in human civilization and in the pursuit of these elements man had negatively impacted the whole eco-balance (Samir, 2009). Agriculture has been the most important turn of event for human civilization ever since first human colony learnt to adapt to the surroundings and started living a non-nomadic life. The use of arable land for cultivation and food production has brought about socio-economic and environmental externalities (Jeong and Foster, 2003). In that aspect modern agriculture practices had increased food production yield tremendously. Parallel to this is the amount of waste generated from this large scale cultivation. It could be inferred from Samir (2009), Jeong and Foster (2003) that the increasing need for food production due to increasing growth of human population had led to the rampant use of fertilizers, pesticides, indiscriminate wastes discharges, contamination of water sources and other serious repercussions such as increased greenhouse gas release. In order to mitigate such damaging events, sustainable agriculture has been introduced over the last decades. rd In Malaysia, the 3 National Agricultural Policy (NAP3 1998-2010) focuses the need of such practices (Faridah, 2001). Nevertheless, the management of agro-waste is still one of the major concerns to the agro-1 -1 industries. Malaysia is the second largest agro-waste producer in Asia, with a production of 0.122 kgcap day while 1.2 million tonnes of agro-waste are land filled annually (Agamuthu, 2009). Globally, part of these waste materials have been used for animal feed and fertilizer (Mamma et al., 2008), however, a large portion of these materials is still deposited annually. This situation is not desirable due to both socio-economic and environmental issues such as waste-disposal health hazard, high transportation costs, insufficient disposal sites, and the land-filling material of high organic content (Tripodo et al., 2004). As a corollary, the extensive study for the use of wastes material as an input for renewable resources and green energy has made its way to the mainstream research community and industry (Dias et al., 2009; Kaparaju et al., 2009; Najafi et al., 2009; Delzeit and Holm-Muller 2009; Sassner, Galbe and Zacchi, 2008). The agro-waste, which is referred as biomass has been used for production of bioethanol and in derivation of functional phytochemicals (Dias et al., 2009; Sassner, Galbe and Zacchi, 2008). In this proposed research, cocoa pod will be used as the targeted agro-waste. The concept of zero waste management will be applied whereby cocoa pod waste volume will be reduced by recovering all resources, and not burning or burying them. Malaysia is known as the third largest producer of cocoa which produces about 200,000 tonnes of cocoa beans per annum (Wong, 2007; Sharifuddin & Zaharah, 2010). The pod, which weighs approximately 0.5 kg, is ovoid in shape with 15-30 cm in length and 8-10 cm in width. The pod
contains about 20-60 cocoa beans which are normally used for production of cocoa butter and chocolate. The pod, being the bulk waste (approximately 2 million tonnes per annum on 1:10 bean to pod weight ratio) however is either been discharged or used as fertilizer after the processing. Even at present, this agriculture residue has never been fully explored in terms of energy production in Malaysia. Extraction of functional components from these materials has also been substantially neglected. The confluence of these preceding points leads to the notion that the cocoa pod agro-waste has very high potential of being new renewable energy source as well as source of natural products in Malaysia, and thus closes the loop for zero-waste management. This will be the prime interest of this doctoral research.
2.0 Literature Review Based on the above introduction section, agro-waste in Malaysia has the potential of bio-energy derivation for zero-waste management. However an in-depth research would be required to ascertain the magnitude and potential value of the agro-waste conversion. Major part of this research focuses on using the lignocellulosic structure of the biomass as the main component for bio-energy conversion. However, in using lignocellulosic agro-waste as potential feedstock for bio-energy production, some of the following issues need to be addressed: (1) The appropriateness of agro-waste for bio-energy production due to its diversity; (2) The availability of value-added components such as functional fibres and polyphenols within crop residues, and (3) the possibility of extracting these value-added components prior to biofuel conversion. Answers to these issues must be based on eco-sustainable approach which is important to address such a way of producing bio-energy without wasting any treasures embedded within the agro-waste. As such, the preliminary literature review of this research focuses on identifying the general solutions currently available as to the above issues and compares the potential processes that could be utilised to carry out the research. The final literature review will encompass wider circle of analysis to produce the specific solution for the targeted agro-waste. Generally, the various reasons that would support the use of cocoa pod biomass for natural product recovery and bioethanol conversion is very much evident as stated by Figueira, Janick, BeMiller (1993). The use of cocoa pod waste as mulching media is detrimental to cocoa plantation especially when the pods become source of disease inoculums. The use of the pod as livestock feed is also restricted by presence of theobromine, which becomes toxic to the livestock at higher feed rate (LD50 of 1254 mgkg-1). These two factors simultaneously limit the disposability options of cocoa pod agro-waste using presently available methods. Studies conducted by Donkoh et al. (2004), Okai et al. (1984) and Adomako (1972) provide chemical composition of cocoa pods cultivated in Ghana. The composition of neutral detergent fibres (comprise of cellulose, hemicelluloses and lignin) account to approximately 522-644 gkg-1 of dry matter of the ripe pod, while pectin makes up 8-11% of ripe pod. A study by Adomafio et al. (2004) however indicates only 26.6% of fibre in the cocoa pod waste, while it lists substantial presence of starch, protein, reduced sugar and lipid. In the analysis by Adomako (1972), fibre content is 45.6% (w/w) for dried ripe pod. The varying results could be partly due to different method of assaying as well as different selection of cocoa pod (Adomako, 1972). These preliminary figures suggest the weighty presence of lignocellulosic components in the cocoa pod for bioethanol conversion. In order to make any bioethanol production cost effective, the cost of feedstock has to be as low as possible. It is interesting to note that the cost of bioethanol production is contributed more than one third by feedstock cost (Dien, Cotta and Jeffries, 2003; Batchelor et al., 1994). Since the feedstock intended for this research is a waste biomass, the cost of bioethanol production could be considerably lower. On the contrary
the cost of bioethanol production is relatively high for lignocellulosic conversion due to lower yield and higher cost of hydrolysis (Sun and Cheng, 2002). This could be compensated by the initial pre-treatment which is expected to extract valuable bioactive compounds that could serve as protein, pectin and antioxidants with monetary values. Pre-treatment of the biomass to solubilise cellulose and hemicellulose is the most significant determinant for successful lignocellulosic bioethanol conversion. This is due to the fact that pre-treatment determines the extent to and the cost at which cellulose and hemicellulose is converted to bioethanol (Balat, Balat and Oz, 2008; Alvira et al., 2010). The use of dilute-acid hydrolysis is the most inexpensive method, but only effective commercially if feedstock is considerably cheap (Alvira et al., 2010; Jeffries and Jin, 2000; Wyman, 1994) however the carbohydrates yield is low (Balat, Balat and Oz, 2008). The yield of sugar for bioconversion can be increased significantly by complementing dilute acid hydrolysis with (supercritical) organosolv process (Aravamuthan et al., 1989), which is more expensive and complex (Wyman, 1996). An additional step of the pre-treatment is the use of alcohol precipitation and lyophilisation to recover pectic-like polysaccharides and the potential antioxidants (Gan, Normaliza and Aishah, 2010), the reason having been mentioned in the previous paragraph. After the bioactive components recovery, the residual lignocellulosic will be treated either via simultaneous saccharification and fermentation (SSF) or separate hydrolysis and fermentation (SHF) process for bioethanol conversion. The SSF process has been used conventionally, but SHF provides an alternative mean with several other benefits (Soccol et al., 2010); however the use and selection of enzymes will depend on the actual analysis of pre-treated residue, since presence of 5-C sugar and 6-C sugar may need specific microbial species and reactor conditions (Okuda et al. 2008,) for high yield fermentation. Since cocoa pod fermentation has not been widely studied, there is little information for any particular microbe selection based on pre-treated sugar composition and its fermentation. Finally the extraction of bioethanol from the fermentation will be carried out and this will also be developed further based on final literature review. These would be one of the primary works in the research and therefore would constitute one of the elements of research hypotheses.
3.0 Aim and Objectives of the Research 3.1 Aim The aim of the proposed doctoral research is to apply zero-waste management concept by carrying out laboratory processing of cocoa pod biomass for recovery of valuable natural products followed by conversion of lignocellulosic biomass into bioethanol using both conventional and newer technology as per reviewed literatures. 3.2 Objectives The objectives of the research are: i) To evaluate application of inexpensive extraction methods to determine maximum yield of bioactive compounds; ii) To analyze and classify the bioactive compounds that has potential functional values per basis of biomass unit; iii) To perform an in depth literature review on the most suitable process(es) for cocoa pod biomass lignocellulosic maximum simple sugar recovery; iv) To carry out a laboratory simple sugar recovery process that has optimum yield; v) To develop bioethanol extraction method that would be in-line with current bio-refinery practice;
To compare the yields between SSF and SHF while determining the optimal process conditions for highest bioethanol yield; vii) To evaluate if extraction of functional bioactive compound(s) has any impact on overall bioethanol cost per unit biomass, and viii) To analyze the outcome if a zero-waste management has been achieved while meeting the other research aims. 3.3 Hypotheses to be investigated Based on the above objectives, the following are the hypotheses to be proven: i) The extraction of functional bioactive compounds in the biomass pre-treatment step assist in reducing effective bioethanol cost for cocoa pod lignocellulosic bioethanol production; ii) The yield of simple sugar from cocoa pod lignocellulosic mass could be optimized by using either conventional or combined conventional-contemporary method without adversely affecting bioethanol yield in the fermentation process, and iii) The cocoa pod biomass has sufficient bioethanol yield based on industry standards, biorefinery practices and meets the zero-waste management where the disposal of biomass to landfill or burning of residue is eliminated after final processing.
vi)
4.0 Research Methodology The research methodology would begin with literature review to identify majority of the information required to meet the research objectives. This will provide preliminary work basis to develop the system to meet all the analytical research requirements. This research comprises of three major phases: i) Production of bioactive components from cocoa pod using dilute-acid extraction and supercritical organosolv process. The collection of cocoa pod samples, preparation for assay, extraction and testing of the samples will be done based on set of standard test methods for anticipated compounds. This will also involve lyophilisation of sample and alcohol precipitation method for antioxidant and polysaccharides extraction. The use of supercritical organosolv process will be developed in-line with the second phase requirement. The results will be compared against figures available from the reviewed literatures. The bioactive components will be identified using FTIR (Fourier Transform Infra Red) spectra and chromatography. Production of bio-ethanol using cellulosic waste from cocoa pod which was pre-treated during the bioactive component extraction in Phase 1. In this step, the whole saccharification and fermentation will be carried out using SSF and SHF. Depending on the results from Phase 1 for lignocellulosic composition; and saccharification/hydrolysis of Phase 2 for sugar composition, the specific sets of operating condition for fermentation will be determined by laboratory scale set-up. The selection of fermentation microbe will be part of this phase based on literature review performed in the initial part of the research as well as the sugar composition determined in saccharification/hydrolysis step. Development of a simultaneous extraction-conversion processing line as the protocol for bio-refinery industry. This phase is to be developed based on detailed literature review and upon completion of Phase 1 and the trials during Phase 2. It will be based on standards applied in bio-refinery extractionconversion processes for bio-products from Phase 2.
ii)
iii)
Research Proposal Preliminary Gantt Chart
Preliminary References
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