Landfill leachate enhances fermentative hydrogen production from glucose and sugarcane processing derivatives

  • Inara Amoroso da Silva
  • Stella Thomaz de Lima
  • Marcos Rechi Siqueira
  • Márcia Andreia Mesquita Silva da Veiga
  • Valeria Reginatto


Fermentation can use renewable raw materials as substrate, which makes it a sustainable method to obtain H2. This study evaluates H2 production by a mixed culture from substrates such as glucose and derivatives from sugarcane processing (sucrose, molasses, and vinasse) combined with landfill leachate. The leachate alone was not a suitable substrate for biohydrogen production. However, leachate blended with glucose, sucrose, molasses, or vinasse increased the H2 production rate by 2.0-, 2.8-, 4.6-, and 0.5-fold, respectively, as compared with the substrates without the leachate. Determination of metals (Cu, Cd, Pb, Hg, Ni, and Fe) at the beginning and at the end of the fermentative assays showed how they were consumed during the fermentation and demonstrated improved H2 production. During fermentation, Cu, Fe, and Cd were the most consumed leachate metals. The best substrate combination to produce H2 was molasses and leachate, which gave high volumetric productivity—469 ml H2/l h. However, addition of the leachate to the substrates stimulated lactic acid formation pathways, which lowered the H2 yield. The use of leachate combined with sugarcane processing derivatives as substrates could add value to the leachate and reduce its polluting power, generating a clean energy source from renewable raw materials.


Leachate Biohydrogen Metals Molasses Sugarcane derivatives 



Central de Tratamento de Resíduos


3,5-Dinitrosalycilyc acid


Environmental Protection Agency


Gas chromatography



G + L

Glucose + leachate


Glucose_equivalente: mmol of sugar as glucose


Cumulative H2 volume in fermentative tests


Maximum potential of H2 production in ml


High performance liquid chromatography



L + I

Leachate + inoculum





M + L

Molasses + leachate


Mixed liquor volatile suspended solids


Maximum H2 production rate


Refraction index detector



S + L

Sucrose + leachate


Thermal conductivity detector


Total organic carbon


Total sugar



V + L

Vinasse + leachate


Volatile solids


Volatile suspended solids


Yield (mmolH2/mmol glu_eq)


Lag phase or the time elapsed before H2 production started (h)



We acknowledge Conselho Nacional de Pesquisa e Desenvolvimento (CNPq) and Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) for financial support.


  1. 1.
    Wang J, Wan W (2008) Effect of Fe2+ concentration on fermentative hydrogen production by mixed cultures. Int J Hydrog Energy 33:1215–1220CrossRefGoogle Scholar
  2. 2.
    Chaubey R, Sahu S, James OO, Maity S (2013) A review on development of industrial processes and emerging techniques for production of hydrogen from renewable and sustainable sources. Renew Sust Energ Rev 23:443–462CrossRefGoogle Scholar
  3. 3.
    Li C, Fang HP (2007) Inhibition of heavy metals on fermentative hydrogen production by granular sludge. Chemosphere 67:668–673CrossRefGoogle Scholar
  4. 4.
    Lo YC, Chen WM, Hung CH, Chen SD, Chang JS (2008) Dark H2 fermentation from sucrose and xylose using H2-producing indigenous bacteria: feasibility and kinetic studies. Water Res 42:827–842CrossRefGoogle Scholar
  5. 5.
    Ren N, Li J, Li B, Wang Y, Liu S (2006) Biohydrogen production from molasses by anaerobic fermentation with a pilot-scale bioreactor system. Int J Hydrog Energy 31:2147–2157CrossRefGoogle Scholar
  6. 6.
    Guo W-Q, Ren N-Q, Wang X-J, Xiang W-S, Meng Z-H, Ding J, Qu Y-Y, Zhang L-S (2008) Biohydrogen production from ethanol-type fermentation of molasses in an expanded granular sludge bed (EGSB) reactor. Int J Hydrog Energy 33:4981–4988CrossRefGoogle Scholar
  7. 7.
    Lay C, Wub H, Hsiao C, Chang J, Chen C, Lin C (2010) Biohydrogen production from soluble condensed molasses fermentation using anaerobic fermentation. Int J Hydrog Energy 35:13445–13451CrossRefGoogle Scholar
  8. 8.
    Bittencourt E, Larroche SC, Novak AC, Nouaille R, Sarma SJ, Brar SK, Letti LAJ, Soccol VT, Soccol CR (2014) Economic process to produce biohydrogen and volatile fatty acids by a mixed culture using vinasse from sugarcane ethanol industry as nutrient source. Bioresour Technol 159:380–386CrossRefGoogle Scholar
  9. 9.
    Hafez H, Nakhla G, Naggar HE (2010) An integrated system for hydrogen and methane production during landfill leachate treatment. Int J Hydrog Energy 35:5010–5014CrossRefGoogle Scholar
  10. 10.
    Liu Q, Zhang X, Yu L, Zhao A, Tai J, Liu J, Qian G, Xu ZP (2011) Fermentative hydrogen production from fresh leachate in batch and continuous bioreactors. Bioresour Technol 102:5411–5417CrossRefGoogle Scholar
  11. 11.
    Watanabe H, Yoshino H (2011) Biohydrogen using leachate from an industrial waste landfill as inoculums. J Mater Cycles Waste Manag 13:113–117CrossRefGoogle Scholar
  12. 12.
    Ratti R, Botta LS, Sakamoto IK, Varesche MBA (2013) Microbial diversity of hydrogen-producing bacteria in batch reactors fed with cellulose using leachate as inoculum. Int J Hydrog Energy 38:9707–9717CrossRefGoogle Scholar
  13. 13.
    Liu Q, Zhang X, Zhou Y, Zhao A, Chen S, Qian G, Xu ZP (2011) Optimization of fermentative biohydrogen production by response surface methodology using fresh leachate as nutrient supplement. Bioresour Technol 102:8661–8668CrossRefGoogle Scholar
  14. 14.
    Renou S, Givaudana JG, Poulain S, Dirassouyan F, Moulin P (2008) Review: landfill leachate treatment: Review and opportunity. J Hazard Mater 150:468–493CrossRefGoogle Scholar
  15. 15.
    Yu HQ, Fang HHP (2001) Inhibition on acidogenesis of dairy wastewater by zinc and copper. Environ Technol 22:1459–1465CrossRefGoogle Scholar
  16. 16.
    Yu HQ, Fang HHP (2001) Inhibition by chromium and cadmium of anaerobic acidogenesis. Water Sci Technol 43:267–274Google Scholar
  17. 17.
    Buitrón G, Carvajal C (2010) Biohydrogen production from Tequila vinasses in an anaerobic sequencing batch reactor: effect of initial substrate concentration, temperature and hydraulic retention time. Bioresour Technol 101:9071–9077CrossRefGoogle Scholar
  18. 18.
    APHA, AWWA, WEF (1995) Standard methods for the examination of water and wastewater. 19th edn. American Public Health Association. Washington, DCGoogle Scholar
  19. 19.
    Gonzalez-Gil G, Kleerebezem R, Lettinga G (2002) Assessment of metabolic properties and kinetic parameters of methanogenic sludge by on-line methane production rate measurements. Appl Microbial Biotechnol 58:248–254CrossRefGoogle Scholar
  20. 20.
    García-Morales JL, Nebot E, Romero LI, Sales D (2001) Comparison between acidogenic and methanogenic inhibition caused by liner alkylbenzzene-sulfonate (LAS). Chem Biochem Eng Q 15:13–19Google Scholar
  21. 21.
    Wang J, Wan W (2009) Kinetic models for fermentative hydrogen production: a review. Int J Hydrog Energy 34:3313–3323CrossRefGoogle Scholar
  22. 22.
    Zwietering MH, Jongenburger I, Rombouts FM, van´t Riet K (1990) Modelling of the bacterial growth curve. App Environ Microbiol 56:1875–1881Google Scholar
  23. 23.
    United States Environmental Protection Agency—USEPA (2007) Microwave assisted acid digestion of sediments sludge, soils, and oils. EPA SW 846(3051a):30pGoogle Scholar
  24. 24.
    Miller GL (1959) Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal Chem 31:426CrossRefGoogle Scholar
  25. 25.
    Oliveira LA, Amorim TS, Santos DV, Silva J (2007). Composição físico-química de variedades de mandioca de mesa cultivadas no sistema orgânico. Revista Raízes e Amidos Tropicais. UNESP, 3Google Scholar
  26. 26.
    Lutz IA (2005) Métodos físico-químicos para a análise de alimentos. 4° Edição: Ministério da Saúde, ISBN:9788533410381Google Scholar
  27. 27.
    Sá LRV, Oliveira MAL, Cammarota MC, Matos A, Leitão VSF (2010) Simultaneous analysis of carbohydrates and volatile fatty acids by HPLC for monitoring fermentative biohydrogen production. Int J Hydrog Energy 36:15177–15186Google Scholar
  28. 28.
    Han SK, Shin HS (2004) Biohydrogen production by anaerobic fermentation of food waste. Int J Hydrog Energy 29:569–577CrossRefGoogle Scholar
  29. 29.
    Sír M, Podhola M, Patocka T, Honzajková Z, Kocurek P, Kubal M, Kura M (2012) The effect of humic acids on the reverse osmosis treatment of hazardous landfill leachate. J Hazard Mater 207–208:86–90CrossRefGoogle Scholar
  30. 30.
    Abu-Daabes M, Qdais HA, Alsyouri H (2013) Assessment of heavy metals and organics in municipal solid waste leachates from landfills with different ages in Jordan. J Environ Prot 4:344–352CrossRefGoogle Scholar
  31. 31.
    Baun DL, Christensen TH (2004) Speciation of heavy metals in landfill leachate: a review. Waste Manage Res 22:3–23CrossRefGoogle Scholar
  32. 32.
    Zheng XJ, Yu HQ (2004) Biological hydrogen production by enriched anaerobic cultures in the presence of copper and zinc. J Environ Sci Health A 39:89–101CrossRefGoogle Scholar
  33. 33.
    Lin CY, Shei SH (2008) Heavy metal effects on fermentative hydrogen production using natural mixed microflora. Int J Hydrog Energy 33:587–593CrossRefGoogle Scholar
  34. 34.
    Srikanth S, Mohan SV (2012) Regulatory function of divalent cations in controlling the acidogenic biohydrogen production process. RSC Adv 2:6576–6589CrossRefGoogle Scholar
  35. 35.
    Bao MD, Su HJ, Tan TW (2013) Dark fermentative bio-hydrogen production: effects of substrate pre-treatment and addition of metal ions or l-cysteine. Fuel 112:38–44CrossRefGoogle Scholar
  36. 36.
    Krishna RH, Gilbert WB (2014) Toxification and detoxification of heavy metals in anaerobic reactors used in the production of biohydrogen: future fuel. Int J Environ Eng Res 3:1–6Google Scholar
  37. 37.
    Intanoo P, Rangsunvigit P, Namprohm W, Thamprajamchit B, Chavadej J, Chavadej S (2012) Hydrogen production from alcohol wastewater by an anaerobic sequencing batch reactor under thermophilic operation: nitrogen and phosphorous uptakes and transformation. Int J Hydrog Energy 37:11104–11112CrossRefGoogle Scholar
  38. 38.
    Chaganti SR, Kim DH, Lalman JA (2012) Dark fermentative hydrogen production by mixed anaerobic cultures: effect of inoculum treatment methods on hydrogen yield. Renew Energy 48:117–121CrossRefGoogle Scholar

Copyright information

© Springer Japan KK 2017

Authors and Affiliations

  • Inara Amoroso da Silva
    • 1
  • Stella Thomaz de Lima
    • 1
  • Marcos Rechi Siqueira
    • 1
  • Márcia Andreia Mesquita Silva da Veiga
    • 1
  • Valeria Reginatto
    • 1
  1. 1.Departamento de Química, Faculdade de Filosofia, Ciências e Letras de Ribeirão PretoUniversidade de São PauloRibeirão PretoBrazil

Personalised recommendations