FERMENTATIVE HYDROGEN PRODUCTION FROM FOOD-INDUSTRY WASTES
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1 Proceedings of the 13 th International Conference on Environmental Science and Technology Athens, Greece, 5-7 September 213 FERMENTATIVE HYDROGEN PRODUCTION FROM FOOD-INDUSTRY WASTES M. ALEXANDROPOULOU 1, 2, G. ANTONOPOULOU 2 and G. LYBERATOS 1, 2 1 School of Chemical Engineering, National Technical University of Athens, GR 1578 Athens, Greece 2 Institute of Chemical Engineering Sciences, P.O. BOX 1414, 2654, Patras, Greece geogant@chemeng.upatras.gr ABSTRACT Τhis study focuses on the exploitation of food-industry wastes as a source for hydrogen production. The wastes which were used were out of date solid baby foods at seven different flavors (solid wastes) and a glucose-based syrup (liquid waste). Preliminary batch hydrogen experiments at mesophilic conditions and initial carbohydrates concentration of 5 g/l were performed to assess the hydrogen potential for both types of wastes. The experiments showed that all substrates were promising for bio-hydrogen production, due to their high content of carbohydrates. Experiments at the same conditions were also performed by using mixtures of solid wastes. The results indicated that all solid wastes could be considered as a sole stream of solid wastes. In the sequel, the effect of initial carbohydrates concentration (2.5-2g/L) and initial ph value (5-7) on hydrogen production from the mixture of solid wastes was assessed. The initial carbohydrates concentration of 15g/L exhibited the highest hydrogen yield of 259.7±3.6 LH 2/kg waste which corresponded to the production of 2.77±.3 mol H 2/mol glucose consumed. The initial ph value of 5.5 also led to maximum hydrogen yield when the initial concentration of carbohydrates in the mixture of solid wastes was 15g/L. The effect of initial carbohydrates concentration in the range of 5 25 g/l on hydrogen production from the liquid waste stream was also assessed. The initial carbohydrates concentration of 15g/L exhibited the highest hydrogen yield of 46.1±.6 LH 2/kg waste which is correlated to the production of 2.61±.2 mol H 2/mol glucose consumed. In the sequel, based on the results from the batch experiments, mesophilic fermentative hydrogen production was investigated in a continuous reactor operating at a hydraulic retention time (HRT) of 24 h, using the mixture of solid wastes at a feed carbohydrates concentration of 15g/L, as substrate. The hydrogen production rate was.39 ±.6 L / L reactor /d, while the yield of hydrogen produced was approximately.22 ±.2 mol H 2 / mol glucose consumed or ± 2.89 L of H 2/kg waste. Keywords: fermentative hydrogen production, food industry wastes, carbohydrates concentration effect, effect of ph value 1. INTRODUCTION Renewable energy resources and alternative fuels have received considerable attention due to growing energy demands in combination with the decreasing reverses of fossil fuels. Hydrogen is a promising energy alternative due to its clean, efficient, renewable and non-polluting characteristics. Anaerobic fermentation is one of the most promising biotechnologies that can be used for hydrogen production, since it is carried out in the dark, at moderate temperatures using different types of wastes as feedstocks (Ntaikou et al., 21). The food industry constitutes the largest industrial sector in Greece and its waste streams CEST213_85
2 could support the development of sustainable technologies for the production of alternative fuels such as biofuels. In this way, food production could be achieved in conjunction with production of energy. Food industry wastes are very rich in carbohydrates, which have been shown to be the main source of hydrogen in fermentative processes. During fermentation, degradation of glucose (or its isomer hexoses or its polymers, starch, glycogen and cellulose) by mixed microbial cultures, is accompanied by the production of hydrogen and various metabolic products, mainly volatile fatty acids (VFAs; acetic, propionic, and butyric acid), lactic acid, and alcohols (butanol and ethanol), in relative proportions that depend on the microbial species present and the prevailing conditions (Antonopoulou et al., 28). Specifically, the production of acetic and butyric acids favours hydrogen production, as glucose fermentation to acetic acid leads to the highest theoretical yield (4 mol H 2/mol glucose consumed) and the conversion to butyric leads to 2 mol H 2 / mol glucose consumed. Production of propionic acid consumes hydrogen, while ethanol and lactic acid are not accompanied by simultaneous production or consumption of hydrogen (Ntaikou et al., 21). The aim of this study is to assess a possible valorisation of the waste streams from a Greek food industry, for the production of biohydrogen. The wastes used were seven different out of date solid baby foods, and one glucose-based syrup. Batch and continuous experiments were conducted to evaluate the fermentative hydrogen production from the solid wastes at 35 C using mixed cultures, while batch hydrogen production tests were conducted to assess the hydrogen production for the syrup waste stream. 2. MATERIALS AND METHODS 2.1 Batch hydrogen production Batch hydrogen experiments were carried out in duplicate at 35ºC in serum bottles of 16 ml with a working volume of 5 ml. 1 mls of mixed anaerobic culture, preheated at 1 o C for 15min (Chen and Lin, 21) were inoculated with 4mLs of a nutrient solution (NaH 2PO 4*2H 2O 8.98g/L, Na 2HPO 4*2H 2O 5.2g/L, yeast extract.625g/l) and fed with the individual solid wastes, mixtures of them and with the liquid waste stream, depending on the experiment.the content of the vials was gassed with a gas mixture of N 2/CO 2 (8/2) and sealed in order to secure anaerobic conditions. Biogas production was measured via syringes. For the individual solid and liquid wastes, hydrogen production experiments were carried out for an initial carbohydrates concentration of 5 g/l. At the same conditions, experiments were also performed by using a mixture of three solid wastes. The effect of initial carbohydrates concentration for the mixture of the seven solid wastes in the range of g/l was investigated. Moreover, the effect of the initial ph value in the range of 5-7 for the mixture of the seven solid wastes was assessed for an initial carbohydrates concentration was 15 g/l. For ph adjustment 6N H 2SO 4 and 6N NaOH were used. Finally, the effect of initial carbohydrates concentration in the range of 5 25 g/l on hydrogen production of the syrup waste stream was also investigated 2.2 Continuous hydrogen production In order to study hydrogen production in a continuous process, a 4 ml active volume mesophilic (35 C) CSTR-type digester (H 2-CSTR) was operated using heat-treated (15 min at 1 o C) anaerobic sludge as inoculum and a mixture of the seven solid wastes at an initial total carbohydrates concentration of 15g/L as substrate. The reactor was operated anaerobically at an HRT of 24h. It was fed intermittently, every 3 h with the mixture of the solid wastes kept at 4 o C. Feeding was programmed always with the stirring on and was supplemented with 1g/L NaHCO 3 in order to maintain the culture ph at CEST213_85
3 suitable levels for biohydrogen production. Biogas production was measured by the method of displacement of acidified water. 2.3 Analytical methods Determinations of total and volatile solids (TS and VS) as well as total and volatile suspended solids (TSS and VSS) were carried out according to Standard Methods (APHA, 1995). The measurement of carbohydrates was carried out according to Joseffson (1983). VFAs were analyzed with a gas chromatograph (VARIAN CP-3), equipped with a flame ionization detector, while gas composition in hydrogen was quantified with a gas chromatograph (SRI 861c MG#1), equipped with a thermal conductivity detector. 3. RESULTS AND DISCUSSION 3.1 Characterization of the substrates The main characteristics of the seven solid wastes used were % TS, % VS, g total carbohydrates/ g substrate and.6.5 g dissolved carbohydrates / g substrate. The main characteristics of the syrup waste stream were g TSS/L, g VSS/L and.15 g dissolved carbohydrates/ g substrate. It is noticeable that these waste streams consisted mainly of carbohydrates, which are an ideal substrate for fermentative hydrogen production. 3.2 Batch hydrogen production experiments Preliminary batch hydrogen production experiments were conducted in order to assess the hydrogen potential of each solid and liquid substrate stream. The cumulative hydrogen production of each waste, for initial carbohydrates concentration of 5 g/l, is shown in Figure solid waste 1 solid waste 2 solid waste 3 solid waste 4 solid waste 5 solid waste 6 solid waste liquid waste a Figure 1. Cumulative hydrogen production a) for solid wastes and b) for syrup waste streams, versus time. Hydrogen production experiments showed that all waste streams were promising substrates for bio-hydrogen production, due to their high content of carbohydrates. Moreover, all types of solid wastes led to similar production of hydrogen and biogas. Specifically, the hydrogen potential of solids wastes varied from 173 to 25 L H 2/kg waste, which were correlated with the production of mol H 2/ mol glucose consumed, while the syrup waste yielded L H 2/kg waste, which corresponds to 2.21 mol H 2/ mol glucose consumed. b CEST213_85
4 Experiments at the same conditions (initial carbohydrates concentration of 5 g/l) were also performed by using a mixture of three solid wastes and the hydrogen potential obtained was compared to the respective of an individual solid waste stream (Figure 2). In this case, the mixture of the three solid wastes yielded 17.4 ± 2.5 L H 2/kg, which is a value similar to that of the individual solid wastes. These results verified that the seven different solid wastes could be handled as a single solid waste and mixture of solid wastes and individual solid waste Figure 2. Cumulative hydrogen production from the mixture of three solid wastes and the individual solid waste stream, versus time. In the sequel, the effect of initial carbohydrates concentration in the range of 2.5-2g/L (2.5, 5, 1, 15 and 2 g/l) on hydrogen production from the mixture of the seven solid waste streams was assessed. The respective evolution of hydrogen as well as the production of VFAs are presented in Figure and 5g/L and 7.5g/L and 1g/L and 15g/L and 2g/L a VFAs concentration (mg/l) acetic propionic iso-butyric butyric iso-valeric valeric 5 1 Initial carbohydrates concentration (g/l) 15 b 2 Figure 3. a) Cumulative hydrogen production and b) VFAs concentration for the mixture of the seven solid wastes, at different initial carbohydrates concentration. The experimental results obtained, showed that the increase of initial carbohydrates concentration led to an increase of hydrogen production, and subsequently, to a stoichiometric hydrogen yield increase (table 1). Specifically, the initial carbohydrates concentration of 15 g/l exhibited the highest hydrogen yield of 259.7±3.6 LH 2/kg waste, which corresponded to the production of 2.77±.3 mol H 2/mol glucose consumed. Further increase of the initial concentration does not seem to enhance the hydrogen production. The stoichiometric yields obtained in the present study are remarkably high. Similar results were obtained by Chang et al. (211) who studied the effect of initial substrate concentration in the range of 5 6 g/l on fermentative hydrogen production from glucose. The optimal stoichiometric yield obtained from that study was 1.78 mol H 2/mol glucose when the substrate concentration was 15 g/l. CEST213_85
5 Table 1. Hydrogen yields of the mixture of the solid substrates in different initial carbohydrates concentrations Carbohydrates concentration (g/l) Yield Η 2 (L H 2/kg) Yield Η 2 (mol H 2/mol glucose) ± ± ± ± ± ± ± ± ± ±.31 From Figure 3b it is obvious that the main metabolic products were acetic and butyric acids, while propionic acid was detected at low concentrations. Acetic acid was the dominant metabolic product for low carbohydrates concentrations (2.5 5 g/l), while butyric acid was produced in higher concentration than acetic acid for higher carbohydrates concentrations (1 2 g/l). Similar results were also reported by Giordano et al. (211) who found out that acetic and butyric acids as were the dominant metabolic products from mixed fermentation of food waste. After determining the optimum initial carbohydrates concentration, the effect of the initial ph value (at a range of 5-7) on fermentative hydrogen production from the mixture of the solid wastes stream, was also investigated. The initial carbohydrates concentration was 15 g/l and the results are given in table 2. Table 2. Hydrogen yields of the mixture of the solid wastes at different initial ph values. Yield Η Initial ph value 2 Yield Final ph value (L H 2/kg) (mol H 2/mol glucose) ± ± ± ± ±.8 4. ± ± ± ± ± ± ± ± ± ±.1 All initial ph values led to similar hydrogen yields, with the value of 5.5 leading to the best hydrogen yield of 26.1 ±. LH 2/kg waste, which corresponded to the production of 2.88 ±.8 mol H 2/mol glucose consumed. At the end of the experiments the ph value was considerably low and varied between The experimental results obtained are also confirmed by other studies demonstrating that the final ph value during fermentative hydrogen production lies in the range of , regardless of the initial ph value (Morimoto et al., 24; Antonopoulou et al., 21). In these experiments, the main metabolic product was butyric acid, while acetic acid was produced at lower concentrations. These results are in agreement with the previous experimental results which indicated that for high carbohydrates concentrations, butyric acid was the dominant metabolic product. Finally, the effect of initial carbohydrates concentration in the range of 5-25g/L (5, 7.5,1, 12.5, 15, 2 and 25 g/l) on hydrogen production from the syrup waste stream was investigated. The respective evolution of hydrogen as well as the production of VFAs, are presented in Figure 4. The experimental results showed that the increase of initial carbohydrates concentration led to an increase of hydrogen production, and CEST213_85
6 subsequently, to an increase of the stoichiometric hydrogen yield (table 3). Specifically, the initial carbohydrates concentration of 15 g/l exhibited the highest hydrogen yield of ±.37 LH 2/kg waste, which corresponded to the production of 2.21 ±.2 molh 2/mol glucose consumed. Further increase of the initial concentration does not seem to enhance the hydrogen production potential. The main metabolic products were acetic and butyric acids, while propionic acid was detected at low concentrations and valeric and iso-valeric acids were not detected at all. The metabolic products distribution profile (figure 4b) followed the same behavior with the experimental results from the mixture of solid wastes streams, with acetic acid being the dominant metabolic product for the lowest carbohydrates concentrations (5-7.5 g/l), while at higher concentrations ( g/l) butyric acid production prevailed and 5g/L and 7.5g/L and 1g/L and 12.5g/L and 15g/L and 2g/L and 25g/L a VFAs concentration (mg/l) acetic propionic iso-butyric butyric iso-valeric valeric b Initial carbohydrates concentration (g/l) Figure 4. a) Cumulative hydrogen production and b) VFAs concentration for the syrup waste, at different initial carbohydrates concentration. Table 3. Hydrogen yields from syrup waste at different initial carbohydrates concentrations Carbohydrates concentration (g/l) Yield Η 2 (L H 2/kg) Yield Η 2 (mol H 2/mol glucose) ± ± ± ± ± ± ± ± ± ± ± ± ± ± Continuous hydrogen production Continuous hydrogen production was conducted in a CSTR-type reactor (H 2-CSTR) with a feed carbohydrates concentration of the solid wastes mixture of 15g/L. The reactor was operated anaerobically at an HRT of 24h and was supplemented with 1g/L NaHCO 3 so as the ph of the feed was 8. The evolution of overall biogas (hydrogen and carbon dioxide) and hydrogen production as well as the production of the main VFAs (acetic, propionic and butyric acid) are presented in Figure 4. CEST213_85
7 The parameters monitored in the hydrogen-producing bioreactor reached levels of low variation from the 4 th day and on. The main characteristics of the hydrogenogenic reactor at steady state are shown in table 3. It should be noted that no methane was detected throughout the experimental period. The percentage of hydrogen in the gas phase at the steady state was 17.4 ± 1.4 %, while the hydrogen production rate reached ml/d (or.39 ±.6 L/L reactor/d). The main products measured were acetic, propionic and butyric acids. Ethanol was also detected, but to a lesser extent, while, butanol, as well as isobutyric, valeric and isovaleric acids, were not detected at all. Contrary to the batch experiments, acetic acid was the dominant metabolic product at steady state.. Gas production rate (L/d) a time (d) Figure 4 a) Biogas and hydrogen production rates and b) VFAs production (acetic, propionic and butyric acid) versus time of the H 2-CSTR. Table 3 The main characteristics of the H 2-CSTR at steady state Characteristic Value ph 5.5 ±.1 Percentage of H 2, (%) 17.4 ± 1.4 Hydrogen production rate, (L/L/d).39 ±.6 Hydrogen yield, (mol H 2 /mol consumed glucose).22 ±.2 Hydrogen yield, (L H 2 / kg waste) ± 2.89 It is obvious from the table, that the yield of hydrogen produced was approximately.22 ±.2 mol H 2 / mol glucose consumed and, consequently, L H 2/ kg waste, which is much lower than the maximum theoretical ones. Furthermore, those yields are much lower than the respective yields obtained by the batch hydrogen production experiment with initial carbohydrates concentration of 15 g/l. An explanation could be that a considerable amount of the produced hydrogen is probably consumed by hydrogen consuming microorganisms such as homoacetogenic bacteria which may be established in the reactor, producing acetic acid (reaction 1): 4H 2 + 2CO 2 CH 3COOH + 2H 2O (1) 4. CONCLUSIONS Biogas Hydrogen VFAs concentration (mg/l) time (d) In this study, it was demonstrated that solid and liquid wastes of the food industry can be exploited for hydrogen production due to their high content in carbohydrates, which are the ideal substrate for fermentative hydrogen production. Batch tests of fermentative hydrogen production proved that all solid wastes could be considered as a single b Acetic acid Propionic acid Butyric acid CEST213_85
8 combined waste stream of all solid wastes. The experimental results showed that the initial carbohydrates concentration of 15g/L either for the mixture of solids waste or for the liquid waste stream exhibited the highest hydrogen yields, while the initial ph value of 5.5 seemed to be the optimum for the solid waste stream. Continuous fermentative hydrogen production was carried out at an HRT of 24h, using the mixture of solids wastes at an initial carbohydrates concentration of 15g/L. At a steady state the hydrogen production rate was.39 ±.6 L / Lreactor /d, while the yield of hydrogen produced was approximately.22 ±.2 mol H 2 / mol glucose consumed or L H 2 / kg waste. This yield is much lower than that obtained through batch operation. ACKNOWLEDGEMENTS The authors would like to acknowledge General Secretariat for Research and Technology (GSRT) for the financial support of this work under Nutrifuel SYNERGASIA_9SYN_32_621 REFERENCES 1. Antonopoulou G., Gavala HN., Skiadas IV and Lyberatos G. (21) Influence of ph in fermentative hydrogen production from sweet sorghum extract. Int. J. Hydrogen Energy, 35, Antonopoulou G., Gavala, H.N., Skiadas I.V., Angelopoulos K. and Lyberatos G. (28). Biofuels generation from sweet sorghum: fermentative hydrogen production and anaerobic digestion of the remaining biomass. Bioresour. Technol., 99, Ntaikou I., Antonopoulou G. and Lyberatos G. (21) Biohydrogen production from biomass and wastes via dark fermentation: a review. Waste Biomass Valor., 1, APHA, AWWA, WPCF. (1995) Standard Methods for the examination of water and wastewater. Franson MA, editors. Washington, DC: American Public Health Association 5. Chang S., Li J.-Z., Liu F. and Wang S.-J. (211) Effect of initial substrate and biomass concentrations on anaerobic fermentative hydrogen production in batch reactors. J. Harbin Institute of Technol., 18(6), Chen C.C. and Lin C.Y. (21) Start up of anaerobic hydrogen producing reactors seeded with sewage sludge. Acta Biotechnol., 21, Chen W-H., Chen S-Y., Khanal S.K. and Syng S. (26) Kinetic study of biological hydrogen production by anaerobic fermentation. Int. J. Hydrogen Energy, 31(15), Giordano A., Cantu C. and Spangi A. (211) Monitoring the biochemical hydrogen and methane potential of the two-stage dark fermentative process Bioresour. Technol., 12, Joseffson B Rapid spectrophotometric determination of total carbohydrates. In: Grasshoff K, Ehrhardt M, Kremling K, editors. Methods of seawater analysis, Verlag Chemie GmbH, Ntaikou I, 1. Morimoto M., Atsuko M., Atif A.A.Y., Ngan M.A, Fakhrul-Razi A and Iyuke S.E. (24). Biological production of hydrogen from glucose by natural anaerobic microflora. Int. J. Hydrogen Energy, 29, CEST213_85
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