Biofuel Production from Carbon monoxide

1. Biofuel Production from Carbon monoxide

2. Homepage 1.Background 2.Executive Summary3.Introduction4.Summary 4.1 Article 14.2 Article 2 5. Comparative Analysis6. References • (Background picture obtained from http://projectbiofuel.blogspot.com.au/) Next Page

3. 1. Background • Homepage Hello everyone! My name is Geetika Kalleechurn and I am currently doing a major in Biotechnology and Molecular Biology. My topic is biofuel production from acetogenic bacteria that use carbon monoxide as a carbon source and synthesise ethanol.I chose this topic as I found the idea of carbon monoxide utilization in biofuel production quite fascinating and I hope you do too! Previous Next 1.Background 2.Executive Summary3.Introduction4.Summary 4.1 Article 4.2 Article 5. Comparative Analysis6. References

4. 2. Executive Summary • Homepage This work involves a comparative analysis of two experiments based on the parameters affecting the growth of acetogenic Clostridium species that synthesise ethanol via syngas fermentation. The aim is to understand the effects of variation in parameters on the growth of the Clostridium species in order to optimise ethanol production from syngas fermentation. (Image obtained from blogs.cleanfuelsdc.org) Previous Next 1.Background 2.Executive Summary3.Introduction4.Summary 4.1 Article 4.2 Article 5. Comparative Analysis6. References

5. 3.Introduction • Homepage With a worldwide increase in the consumption and gradual diminishing of fossil fuels, the need for renewable energy sources has also increased. Bio-fuels such as ethanol have attractive prospects in the industry. Ethanol can be produced using a series of pathways. Figure 1 from Wei et al, 2008 shows the different pathways that can be used for ethanol production from feedstock.For the purpose of this study the main focus will be on the gasification biosynthesis pathway used by the bacterium Clostridium ljungdahlii Next Page 1.Background 2.Executive Summary3.Introduction4.Summary 4.1 Article 4.2 Article 5. Comparative Analysis6. References

6. 3. Introduction • Homepage 1.Background 2.Executive Summary3.Introduction4.Summary 4.1 Article 14.2 Article 2 5. Comparative Analysis6. References Carbon monoxide is highly toxic to living creatures with the exception of microorganisms such as acetogenic bacteria. These can use carbon monoxide as a carbon source and synthesise ethanol and acetate together with some other products (Kopeke et al.,2011). The metabolic pathway used by these microorganisms is known as the Wood-Ljungdahl pathway (shown in figure 1). The fermentation process has a number of advantages though syngas fermentation is yet to be used on a commercial scale. The advantages include the utilization of the whole biomass for product formation; no pre-treatment steps required; high specificity of biological catalysts (Munasinghe and Khanal,2010). Nevertheless, there are also some challenges involved in the process one of which is the poor solubility of carbon monoxide gas in aqueous solutions (Abubackar et al., 2012) For the purpose of this study, the activity of two Clostridium species grown under different conditions for the optimisation of ethanol production were compared. Previous Next

7. 3. Introduction The Wood-Ljungdahl Pathway-Conversion of carbon monoxide to ethanol Figure 1 adapted from Kopke et al, 2011

8. 4.1 Summary Ethanol and acetate production by Clostridium ljungdahlii and Clostridium autoethanogenum using resting cells Jacqueline L. Cotter , Mari S. Chinn and Amy M.Grunden (*note for the purpose of this study the main focus will be on Clostridium ljungdahlii) OverviewC.ljungdahlii is an acetogenic bacteria, which, when grown on carbon monoxide produces ethanol and acetate as the main end-products. Optimisation of ethanol production requires a shift in metabolism from acetogenesis (Acetic acid formation) to solventogenesis (ethanol production). Resting cells have to found capable of inducing this shift in metabolism and also increase cell stability. In this study, the ability of nitrogen-limited media to initiate a non-growing in C.ljungdahlii and the effects of different growth states and medium pH on ethanol and acetate production were examined. Materials and Methods Table 1 (Cotter et al, 2009) shows the types of media used for C.ljungdahlii.Three types of non-growth media were prepared anaerobically. The cultures were initially grown until a mid-late log phase and then were subjected to a series of centrifugationand resuspension. The suspensionswere then transferred to the appropriate non-growth media. (material used from Cotter et al.,2009) Non-growth media

9. 4.1 Summary Results and Discussion • Homepage Effect of non- growth mediaStudies showed that the addition of vitamins, saltsand trace elements enabled the cultures to maintain highcell densities in nitrogen deficient media. The results from figure 1 showed that relatively few cultures from each typeof non-growth media remained viable. The ethanol production levels were affected by the media type used. Effect of pH on resting cell performanceIt was found that lowering pH values did not affect thestability of the culture but had an adverse effect on theviability. DiscussionThe results obtained in this study are quite differentfrom studies carried out in the past whereby, ethanolis a secondary metabolite in syngas fermentation. A shiftin metabolism as observed with other studies was not successfully established in this experiment. (Materials used from Cotter et al., 2009) 1.Background 2.Execuve Summary3.Introduction4.Summary 4.1 Article 14.2Article 2 5. Comparative Analysis6. References Figure 1 shows the metabolism of C.ljungdahlii – ethanol production over time in initial non-growth media Figure 2 shows the metabolism of C.ljungdahlii on NG.RCM.NA.SVE medium-ethanol production over time at different pH

10. 4.2 Summary Sustainable ethanol fermentation from synthesis gas by Clostridium ljungdahlii in a continuous stirred tank bioreactor Mohammadi M., Younesi H., Najafpour G., Mohamed A.R. • Homepage OverviewSecond generation biofuels such as ethanol by syngas fermentation using C.ljungdahlii represents several challenges, one of which is the cultivation of the bacterium as it is strictly anaerobic. The main aims of this study were to investigate and monitor the fermentation conditions such as fresh medium flow rate, culture pH on cell concentration and viability, substrate uptake and product formation during the continuous cultivation of C.ljungdahlii. (*note for the purpose of this study only the results for substrate uptake and product formation will be used) Materials and methods The inoculum was prepared using a defined medium containing, NaHCO3, a reducing agent, vitamins, trace elements. The pH of the medium was maintained at 6.8 using NaOH solution. The bioreactor was operated under batch conditions for 3 daysfollowing which a continuous mode was started with a liquid flow rate of 0.25mL/min.(note pH of bioreactor was not controlled) Cell growth was monitored by determining the cell dry weight of C.ljungdahlii and acetate and ethanol production was monitored using a gas chromatogram.(Material used from Mohammadi et al., 2012) 1.Background 2.Executive Summary3.Introduction4.Summary 4.1 Article 14.2Article 2 5. Comparative Analysis6. References Figure 1 shows the operating conditions of the bioreactor for optimising the liquid flow rate at a constant stirring rate of 500 rpm and a gas flow rate of 14mL/min

11. 4.2 summary Results and Discussion The results for substrate uptake, product formation and varying pH were analysed.Effect of varying pH The effect of changes in pH can be seen in Figure 2,whereby a drastic reduction in cell density was observed due to a drop in pH to 4.18. A re-adjustment in pH to 6.5 allowed to cells to recover within 5 days. Product formation From figure 3, it was observed that over the first 7 days when there was a drop in the pH of the culture, acetate was the main product. Following this,a drastic increase in both ethanol and acetate was observed, with the product levels reaching a peak of 9g/L on the 10th day of the experiment.Discussion A decrease in pH affects the cell stability and product formation. The production of acetate during the first 7 days led to a decrease in external pH and loss of constantinternal pH. Acetate- a weak organic acid, can accumulate inside the cells and in the presence of H+ ions cause a decrease in internal pH. This in turn leads a metabolic shift from acetogenesis to solventogenesis. Hence , ethanol production occurred after a decrease in internal pH. This study was successful in demonstrating a shift in the metabolic pathway of C.ljungdahlii . Further studies involvingvariation of other parameters are expected to help in increasingthe molar ratio of ethanol to acetate. (Materials used from Mohammadiet al., 2012) Figure 2 shows the change in the cell dry weight with changes in pH over time Figure 3 shows the varying levels ethanol and acetate produced over time

12. 5. Comparative analysis • Homepage The works from both Cotter et al. and Mohammadi et al. focused on the production of ethanol from syngas fermentation using the bacterium Clostridium ljungdahlii . The studies performed in Cotter et al. were relatively complex and the goals of the experiments performed for C.ljungdahlii were not successfully met with. The results obtained showed were different to what was expected-ethanol was produced in the growth phase of the bacteria while the results obtained from other researchers showed otherwise. The work performed by Mohammadi et al. aimed at providing a better understanding and correlation of the biochemical pathway used by C.ljungdahlii. The design of the experiment was far much simpler than the work that was undertaken by Cotter et al. . The goals of the experiment were successfully met with through the demonstration of a shift in metabolism of the bacteria under different parameters. The shift in metabolism from the acetogenic pathway to the solventogenic pathway was found to correlate well with the Wood-Ljungdahl Pathway. Furthermore , the metabolic shift was also observed in a study performed by Gaddy et al. Previous Next 1.Background 2.Executive Summary3.Introduction4.Summary 4.1 Article 14.2Article 2 5. Comparative Analysis6. References

13. 6. References • Homepage 1) Abubackar H.N., Veiga M.C., Kennes C., 2012. “Biological conversion of carbon monoxide to ethanol: Effect of pH, gas pressure, reducing agent and yeast extract”, Bioresource Technology 114:518-522 2) Cotter J.L., Chinn M.S., Grunden A.M., 2009. “Ethanol and acetate production by Clostridium ljungdahlii and Clostridium autoethanogenum using resting cells”, Bioprocess and Biosystems Engineering 32(3):369-380 3) Kopke M., Mihalcea C., Bromley J.C., Simpson S.D., 2011. “Fermentative production of ethanol from carbon monoxide” Current Opinion in Biotechnology (22)3:320-325 4)Mohammadi M., Younesi H., Najafpour G., Mohamed A.R.,2012. “Sustainable ethanol fermentation from synthesis gas by Clostridium ljungdahlii in a continuous stirred tank bioreactor” Journal of Chemical Technology and Biotechnology (87)6:837-843 5)Munasinghe P.C., and Khanal S.K.,2010. “Biomass-derived syngas fermentation into biofuels: Opportunities and challenges” Bioresource Technology (101):13 5013-5022 6) Wei L., Pordesimo L.O., Igathinathane C., Batchelor W.D.,2008. “Process engineering evaluation of ethanol production from wood through bioprocessing and chemical catalysis” Biomass and Bioenergy 33:255-266