BIOCEN Summer School Valladolid 2011 Work group : Krzysztof Piotrowski Student group :

1. Technological strategies for biohydrogen production BIOCEN SummerSchool Valladolid 2011 Workgroup: KrzysztofPiotrowski Studentgroup: Natalia Alonso-Movilla ChristophKrüger Lina Glittmann Inka Mella AgnieszkaKorus EwaBorowska

2. Contents • Introduction • Fermentativeprocess • Feedstock • Bioreactors • Kinetics • Adaptation of existingplants

3. Hydrogen – General information • Energy carrier • Clean fuel: No CO2 emissions • Used in fuel cells • High energy yield: 122 kJ/g • More orless 3 times greaterthanhydrocarbonfuels • Producedfrom a wide variety of primary energy sources • 5 % fromrenewablessources

4. European hydrogen infrastructure and production • European distribution of hydrogen demonstration projects Source: www.roads2hy.com

5. Role of hydrogen in the future • Key technology for future sustainable energy supply. Source: Int. J. Hydrogen Energ 2010; 74:16-26

6. Hydrogen from biomass • Pathways from biomass to hydrogen: • Fermentative technologies can contribute to a future sustainable hydrogen economy Source: Int. J. Hydrogen Energ 2010; 74:16-26

7. Raw materials for biohydrogen production Categorized as: Lignocellulosicbiomass (i.e. grass, wood, straw), Starchy biomass (i.e. potato, cereals, food, starch-based wastewater), Sucrose containing biomass (i.e. sugar beet, sugar cane, sweet sorghum), Waste oil (POME),

8. Characteristic data of selected kinds of biomass. Source: Journal of cleaner production, Raw materials for fermantative production.

9. In Castile and Leon… • Agriculture-basedeconomy. • Main sourse of organicwaste: • swine manure, poultry manure, cow manure, • fruit wastes, industry (winery, wood). • The crops cultivated: • cereals (Wheat), • legume, • sunflower.

10. Estimated production yields of anaerobic reactors treating agricultural waste. Source: Hydrogen production from agrocultural waste by dark fermentation: review.

11. Factors influencingfermentative biohydrogen production

12. Culture pH • Influence on • hydrogenase activity • metabolism pathway

13. Hydraulic retention time • Depends on different metabolisms • Inhibits or terminates methanogenesis • Ranges from a few minutes to several hours

14. Hydrogen partial pressure • Hydrogen partial pressure ↗ Hydrogen yield ↘ • Excess of the hydrogen must be removed from the system to maintain hydrogen production • Solution: lowering dissolved H2 with N2

15. Nutrients • Necessary supplements: nitrogen, phosphate and other inorganic species • Norg better than Ninorg • Excess of phosphate may favor VFAs and hydrogen production over solvents production • Inorganic ions: Mg2+, Na+, Zn2+, Fe2+

16. Temperature • Mesophilic range (around 37 0C) and thermophilic range (around 55 0C) • T↗improvement of hydrogen production, T↗ ↗ (out of the range) decrease in hydrogen production

17. Seed culture • Clostridium and Enterobacter are themost widely used as inoculum • Problem of pureculturesormixedcultures • Mixedcultures aremore practical Clostridium bifermentans (spores) Source: http://bacterioweb.univ-fcomte.fr/

18. Kinetic model • Mathematical models based on experiments • Description of reaction processes conditions influence of inhibitors /activators • generalized  Application in similar reactions

19. Gompertz model Type of mathematical model for time-dependend functions Here: Kinetic model for Batch fermentative H2-production processes Source: Int. J. (2009) 33:13-23 • Describes the cumulative H2-Production (H) over the cultivation time t • the lag time (λ) stands for the period until the Production starts • The slope (Rm) describes the Production rate • The maximum potential production (P) is described by the upper Asymptote lag time (λ) slope (Rm) Max. potential production (P)

20. Gompertz model Advantages • Accurate • Easy to adopt • Universal • Constants have biological meaning  better understanding of a process • Widely used H(t)=H2-production in ml over cultivation time (t) in h P=Potential (ml) Rm=Rate (ml/h) λ =Lag time (h) e=Euler‘s Number (e = 2.71828...) Rm e P H(t) - t λ P Source: Int. J. (2009) 33:13-23

21. H2fromglucosebatchfermentationParameter • Temperature : Best 41°C // cost-effective 35°C • Cultivation: pH 7 // enhancement: pH 3 • Withoutsludgepreacidification • Pretreatmenttostopmethanogens: BSR: “bacterial stress respond-mechanism" • Best treatment methods: chemical acidification Text: Appl M. Biot. (2002) 58:224-228 Text: Appl M. Biot. (2006) 72:635-643 Abb.: Int. J. (2006) 78:0-5

22. H2fromglucosebatchfermentationResults • Clostridium butyricum • No methane production • H2 yield below the yields of pure Clostridium cultures • Example cattle manure: • H2 yield: up to 430ml/g VSS • Lag time: around 8h • Rate: 35 ml/h Appl M. Biot. (2006) 72:635-643

23. Sucrose/Food waste/NFDM results • Variation: Substrate concentration • Low pH <4 : inhibition • Productivity depends on substrates concentration • organic acids Int. J. (2006) Wen-Hsing Chen

24. Starch/glycerol results • Variation: different Inoculum • no production from oil • Sludge influences starch • Activated inoculum (better • 1,3 propanediol from glycerol Int. J. (2009) Yohei Akutsu

25. Glycerolresults • Variable: Sludge concentration • By-products: butyric acid, acetic acid and 1.3 propanediol • low concentration 1,16 g VSS/L • No limitation from nitrogen

26. Glycerol results Int. J. (2009) K. Seifert

27. Bioreactors used for biohydrogen production

28. Anaerobic digestion plant a/b/c: Vertical, completely-stirred tank reactor (a/b: mechanical stirring; c: biogas mixing),

29. Anaerobic digestion plant d/e: Horizontal plug-flow reactor (mechanical stirring)

30. Biogas plants for hydrogen production – new possibilities…

31. Hydrogen production in the biogas plant

32. Hydrogen producers • Species for mesophilicfermentation - Clostridium (C.pasteurianum, C.saccharobutylicum, C. butyricum), Enterobacter (E. aerogenes) and Bacillus • Species for thermophilicfermentation – Thermoanaerobacteriumthermosaccharolyticum, Caldicellulosiruptor (C. saccharolyticus, C. thermocellum), Bacillus thermozeamaize

33. Hydrogen yield depends on: • Feedstock type • Process conditions • Reactor construction • Presence of H2 consumers and metabolic competitors

34. H2consumers and thetreatmentmethods

35. Homoacetogenic bacteria • Anaerobic microorganisms which catalyze the formation of acetate from H2 and CO2. • They decrease significantly the hydrogen yield. Hydrogen is consumed by acetogenic bacteria. • Prevention method: • Heating pretreatment –do not remove some Clostridium • Operating parameters e.g. removing CO2

36. H2 consumers and thetreatmentmethods

37. Sulfate-reducing bacteria (SRB) • The most efficient biochemical reaction usinghydrogen involves the sulfate/nitrate-reducing microorganisms. • Under sulfate-rich conditions – hydrogen (also CO2 and VFA)is immediately consumed. • Prevention method: maintain pH lower than 6

38. H2 consumers and thetreatmentmethods

39. Methanogens (MPB) • Main hydrogen consumers • Prevention methods: • Chemical inhibition – Bromoethanesulfonate (BES), acetylene and chloroform. Not environmental friendly and too expensive. • Low pH maintaining – most methanogens can grow only at pH between 6-8. In absence of pH control during a batch process, an acidic initial pH is strongly recommended. • Heat treatment of the inoculum– 100ᵒC for 10 min – methanogens do not sporulateand do not survive such conditions. The most common treatment. • Short hydraulic retention time – methanogens are unable to create biofilm because of a low growh rate so they can be washed out of the reactor. In most cases – retention time less than 6h.

40. Metabolic competitors

41. Lactic acid bacteria (LAB) • Replacement of hydrogen fermentation by lactic acid fermentation. • Prevention method: increaseinthetemperature above 50ᵒC. • Growth of LAB can be limited only in thermophilic fermentation.

42. Hydrogen vs Methane… Heat value per mass unit H2 CH4 141.88MJ/kg 52.21MJ/kg

43. But… Heat value per volume unit H2 CH4 12,84MJ/m3 40,78MJ/m3

44. Yield… swine manure food waste H2 209ml/gVS 196ml/gVS CH4 266ml/gVS 229ml/gVS

45. Efficiency vs ecology • Hydrogen yield and heat value per volume is lower than methane – less energy can be obtain from biomass. BUT… • Hydrogen combustion does not contribute air contamination – the only product of the reaction is WATER.

46. Recomention of biohydrogen plant for Castile and Leon region. • Agriculturaldomination of wheatwithmaincomponentglucose. • Optimalconditions: pH 6, temperature 35 0C, process time up to 60 h. • Type of inocculum: Clostridiumbutyricum • Preteatmentmethod: • Bioreactor: possibility of adaptation of existingbiogas plant for biohydrogenproduction.