The document describes a study on enhancing biohydrogen production from carbohydrate-rich industrial wastewater under anaerobic conditions. A pilot-scale Monitoring Based Agitable Upflow Anaerobic Sludge Blanket (MAUASB) reactor was constructed and operated for 5 months using sugar industry wastewater as the substrate. The maximum COD removal efficiency was 81% at pH 5.0, and hydrogen production peaked at 272.4 ml on the 8th day at pH 5.1 before decreasing due to methanogenesis. The study demonstrated the feasibility of using the MAUASB reactor to treat sugar wastewater and produce biohydrogen as a renewable energy source.

1. Enhanced Biohydrogen
Production Utilizing
Carbohydrate Rich Industrial
Wastewater Under
Anaerobic Microenvironment
Atun RoyChoudhury
M.E. Scholar (Env Engg) & ResearchAsst
Annamalai University (Ramky)

2. Over view
Introduction
Area of Application
What are the Problem Addressed
Limitations/Drawbacks of Current Available Products
Elaborative Description of the Innovation
Area of Immediate and Future Application
Novelty and Usefulness
Stage of the Innovation
Future Research
Market and competitor
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3. Introduction
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4. Waste to Energy Cycle
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5. Wastes suitable for energy production
Solid wastes with high carbon content and heating value
Waste water with high BOD and COD value
Waste gases with high heating value (However, in practice this
option is not normally used).
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6. Scenario of Bio-hydrogen
Synthesis of bio-hydrogen oil from the organic substrate comprises
exclusive advantages like-
• High rate of bacterial growth requires low energy input.
• No oxygen limitation problems.
• Economic feasibility proclaimed by using dark fermentation process
etc.
• Both pure culture such as Clostridium sp. and mixed cultures of
anaerobic bacteria, can used.
• Wide range of carbonaceous waste materials can be used such as
sugar wastewater, starch wastewater, and dairy waste water etc.
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7. Area of Application
Waste to Energy Conversion
Clean and Renewable Energy
Synthesis
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8. What are the Problem Addressed
In the recent era, utmost attention is paid towards exploring the
alternative source of energies, being more specific “Green Energies”.
The conventional energy sources are non-renewable and subjected to
the greater depletion.
Apart from this, due to the rapid urbanization and industrial growth
quantity of waste generated also got increased.
A Sustainable Waste Management technology becomes one of the
prime requirement of the present time.
The present technology is capable of enlightening two prime aspects
of the present waste management scenario namely,
• “Waste Treatment and Minimization”
• “Waste to Energy Conversion”
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9. Limitations/Drawbacks of Current Available Products
Nowadays, universal energy necessities are mostly dependent on fossil fuel.
As per the conventional practice, after extracting juice from the sugarcane the
MOLASSESAND BAGASSE are disposed unscientifically by open dumping.
wastewater from the sugar industries is a “Misplaced Resource”
The ultimate goal is “Sustainable Development”
Drawbacks
• “Leachate formation”
• “Ground Water Contamination”
• “Change in Soil pH”
• “Conversion of primary pollutant to secondary pollutant”
• “Air Pollution Due to the Burning of Fossil Fuel”
• “Generation of Green House Gases”
• “Global warming”
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10. Elaborative Description of the Innovation
“Anaerobic Dark Fermentation Process”
Pilot scale MAUASB reactor was fabricated and operated for the
period of 5 months.
The reactor start-up period was minimized using the Seed Sludge.
Quantifying the overall bio-hydrogen production, a potential
growth was significant between 2nd and 8th day of reactor start up.
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11. Inoculum pre-treatment
Screening &
Identification
16s rRNA
identification
Adaptation of
strain
improvement
strategies
Characterization of
starch and sugar
Industry wastewater
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12. Portrayal of the bioreactor system
Monitoring Based Agitable Up flow Anaerobic Sludge Blanket Reactor
Feed tank has the supply volume of 10 L
Total experimental volume of the reactor was 21 L
5L volume meant for gas collection
16L working volume
Diameter of 212 mm and height of 460 mm
4 different segments-
• Seed sludge introduction
• Substrate configuration
• Biofilm
• Gas collection chamber
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13. Batch Reactor with Different Substrates
With Glucose With Sucrose
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14. An overview of the Pilot Scale MAUASB reactor
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15. Computer Compatibility of the Pilot Scale Reactor
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16. Characteristics of raw starch wastewater effluent
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S.No. Parameter* Tapioca Starch Effluent
1 pH 4-5
2 Total Suspended Solids 400-800
3 Total Dissolved Solids 1000-1400
4 Chlorides 200-500
5 Sulphates 50-200
6 Oil And Grease 3-12
7 BOD 2200-4000
8 COD 4000-6000
9 Phosphates 30-120
10 Ammonical Nitrogen 4-5

17. Characterization of sugar industry effluent
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S.No. Parameter* Sugar effluent
1 pH 6.5-8.8
2 Total solids 870-1950
3 Total suspended solids 220-790
4 Total dissolved solids 400-1650
5 Chlorides 18-40
6 Dissolved oxygen 0-2.0
7 BOD 300-2200
8 COD 1360-2000
9 Sulphate 40-70
10 Oil and grease 60-100

18. Inoculum pretreatment
Heat shock treatment (HST)
Acid pretreatment / acid enrichment
Chloroform pretreatment
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19. 16s rRNA identification for bacterial species
Isolation of bacterial genomic DNA.
Column purification of genomic DNA
Preparation of samples for PCR.
PCR set up
PCR product analysis
PCR product purification
Sequencing
Data analysis
GENOMIC DNA EXTRACTION
AGAROSE GEL ELECTROPHORESIS
COLUMN PURIFICATION
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21. Optimization of monitoring parameters
pH between 4.5 and 8.5
Standard concentration between 0.2 and 0.6 OD at 600 nm
Substrate concentration between 2.5 and 25 g L-1
Temperature range between 250 and 400 C
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22. Adaptation of strain improvement strategies
Selection of stable strains
Selection of non- foaming strains
Selection of strains resistant to the
components of the medium
Selection of morphologically
favourable strains
Selection of strains which are
tolerant to low oxygen tension
Selection of strains based on
mutagenic studies namely
auxotrophic and mutant resistant to
analogues
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23. Bio Hydrogen Production atVarious Substrate
Concentrations
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CumulativeHydrogenproduction(ml)
Time in hours
5 g/l
10 g/l
20mg/l
40 g/l

24. Bio Hydrogen Production atVaried pH
Concentrations
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cumulativehydrogenproduction(ml)
Time in hours
Initial pH 5
Initial pH 5.5
Initial pH 6
Initial pH 6.5
Initial pH 7

25. Percentage of COD removal at different mixing
ratio of substrates
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Cumulative%ofCODremoval
Time in days
60-40
50-50
40-60
30-70
20-80

26. Percentage of COD removal at different pH
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%ofCODremoval
Time in days
pH 9
pH 8
pH 7
pH 6
pH 5

27. COD concentration decrease with increase
Hydrogen production
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CODconcentration(mg/l)
Time in days
Hydrogenyield(ml)
COD concentrations mg/l
hydrogen yield ml

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29. Area of Immediate and Future Application
Production of electricity, heat and water for various end uses
Industrial applications
Vehicular transportation
Residential applications
Commercial applications, including in telecom towers for providing
backup power
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30. Novelty and Usefulness
INDIGENOUS ORGANISMS were used to perform the study,
which is isolated from waste itself, therefore no need of
maintaining any pure culture.
Alteration of the conventional anaerobic mechanism (digestion
pathway) to yield more bio hydrogen rather than methane, which
is a green house gas.
Improved Pre-treatment was given in order to suppress the
activity of the methanogenic organisms.
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31. Stage of the Innovation
Pilot Scale Production Done
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32. Future Research
Genetically Engineered Microorganisms can be used to improve
the yield of biohydrogen and inhibit methanogenic activity.
Mutation can be done to improve the quantity of bio hydrogen
generation.
Improvement of symbiotic mechanism can also be improvised by
means of introducing other beneficial organisms.
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33. Market and competitor
Hydrogen is high in energy content as it contains 120.7 kilojoules/gram.
This is the highest energy content per unit mass among known fuels.
Hydrogen can be used for power generation and also for transport
applications.
It is possible to use hydrogen in internal combustion (IC) engines, directly or
mixed with diesel and compressed natural gas (CNG).
Hydrogen can also be used directly as a fuel in fuel cells to produce
electricity.
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34. Global scenario
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Sl No. Country Scenario
1. Germany Largest demonstrator and pioneer of
hydrogen based applications and
having several hydrogen fueling
stations
2. Iceland Plans to be world's first hydrogen
economy with an annual spending
around $ 30M Hydrogen Freedom
Fuel
3. USA Initiative announced in January 2003
with the budget of US $ 2.2 billion
and implemented by setting up IPHE
in November 2003
4. Japan Started hydrogen fueling stations and
plans to spend $20 billion by 2020

35. Indian Scenario (H-CNG Dispensing Station)
Faridabad in Haryana Dwarka in New Delhi
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36. Hydrogen fuelled 3-wheeler Hydrogen-Diesel dual fuel vehicle
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37. Hydrogen fuelled energy-generator
Use of Hydrogen in a telecom tower
for providing back up power
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38. Conclusion
The study claims the feasibility of bio-hydrogen synthesis from sugar
industry wastewater using MAUASB reactor.
It minimizes the environmental intervention by the removal of
pollution load to the optimal from sugary wastewater.
The maximum COD removal efficiency was found to be 81% at pH 5.0.
Maximum H2 production (about 272.4ml) of the MAUASB reactor was
found on 8th day maintained at pH value of 5.1.
Successive production faced depletion due to Methanization.
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39. Acknowledgements and Publications
I sincerely thank UGC for financially supporting the project.
I would like to convey my sincere gratitude to Valliammai
Engineering College for giving me an opportunity to present my
research in front of the jury.
Atun et al. (2017) Synthesis of Bio-Hydrogen Renovated with
Carbohydrate Rich Wastewater, Utilizing Monitoring Based
Agitable UASB Reactor. Bioresource technology, 241(4), 73-84.
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40. References
• 1. Sreela-C, ImaiT, Plangklang P, Reungsang A. Optimization of key factors affecting
hydrogen production from food waste by anaerobic mixed cultures. Int J Hydrogen Energy
2011; 36:14120-33.
• 2. Kapdan IK, Kargi F. Bio-hydrogen production from waste materials. Enzyme Microb
Technol 2006; 38:569-82.
• 3.Wei J, Liu Z-T, Zhang X. Bio-hydrogen production from starch wastewater and
application in fuel cell. Int J Hydrogen Energy 2010; 35:2949-52.
• 4.Wang J,WanW. Factors influencing fermentative hydrogen production: a review. Int J
Hydrogen Energy 2009; 3:799-811.
• 5. Ravi kumar parihar and Dr. kanjan upadhay”production of bio-hydrogen gas from dairy
industry wastewater by anaerobic fermentation process”IJAR 2016; 2(3): 512-515
• 6. Chen-Yeon Chu a,b,c,*, Zulaicha Dwi Hastuti d,e, Eniya Listiani Dewi e,WidodoWahyu
Purwanto d, Unggul Priyanto” Enhancing strategy on renewable hydrogen production in a
continuous bioreactor with packed biofilter from sugary wastewater”
• 7.Taguchi F, Chang JD,Taguchi S, Morimoto M. Efficient hydrogen production from starch
by a bacterium isolated from termites. J FermentTechnol 1992;73:244–5.
• 8. UenoY, KawaiT, Sato S, Otsuka S, Morimoto M. Biological production of hydrogen from
cellulose by natural anaerobic microflora. J Ferment Bioeng 1995;79:395–7.
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