1. SECOND GENERATION ETHANOL PRODUCTION FROM AGRICULTURAL RESIDUES
Prof. Rintu Banerjee, FNAAS, FBRS, FBRIAT
Professor, Agricultural & Food Engineering Department
Indian Institute of Technology Kharagpur
India

2. Sector wise global CO2 emission
Carbon dioxide equivalent emissions (CO2 eq) from transportation sector in India
Source: IPCC, 2014)

3. World market for Biofuel production: 2013- 2023

5. Indian scenario in biofuel production
Bioethanol demand with blending targets (%) in India
Demand for ethanol for blending and share in blending

6. Indian scenario in biofuel production
Government of India, is planning to set up twelve (12) 2G Ethanol Bio-refineries across 11 States viz. Punjab, Haryana, U.P., M.P, Bihar, Assam, Odisha, Gujarat, Maharashtra, Karnataka and A.P.
The estimated investment for the 12 Bio-refineries is Rs 10,000 crores. These Bio-refineries shall produce around crore litres of Ethanol annually, thus contributing significantly towards the Ethanol Blended Petrol Programme.
Recently GoI has proposed a new 2G biorefinery in Paradip (Odisha)

7. India to commission world’s largest green field refinery by 2022
Signed a Joint Venture (JV) agreement, June -2017

8. Second Generation (2G) ethanol production

9. Potential source of biomass for 2G Ethanol
Agricultural residues (wheat straw, corn stover)
Energy crops (switchgrass, poplar)
Wood and paper waste
Municipal solid waste, organic
Low cost renewable organic resources

10. Global biofuels market forecast (2017-2014)

11. Gross residue availability from crop production in India
The above scenario illustrates the potential of some of the crop residues which can be utilized for biofuel and biochemical generation

12. Biomass Selection Lignocellulosics Grazing Non- Grazing Rice straw
Maize straw
Ricinus communis
Lantana camara
Wheat straw
Rice husk
Jatropha curcas
Pineapple leaf wastes
Jute waste
Oil seed waste
Bambusa bambos
Datura stramonium
Grazing
Non- Grazing

13. 2G-Ethanol production from Non-conventional and Renewable sources
Lignocellulosic biomass
Bambusa bambos
Lantana camara
Kans Grass
Ricinus communis
Cotton Stalk
Sugarcane Tops
2G-Ethanol production from Non-conventional and Renewable sources
CELLULOSE
HEMICELLULOSE
LIGNIN
Enzymatic Pretreatment
-Promising Green Technology
Rice straw

14. Biochemical Characterization of lignocellulosics
composition
Cellulose
(%, w/w)
Hemicellulose
Lignin
Lantana camara
47.25
16.4
17.26
Ricinus communis
42.00
18.02
19.88
Bambusa bambos
45.00
17.00
19.20
Sugarcane Tops
33.00
22.76
13.45
Kans Grass
38.70
29.00
17.46
Pineapple leaf waste
25.00
13.00
Rice straw 33 16 14
Mixture
43.02 24
14.57

15. Biochemistry of Lignocellulosics

16. Steps involved in 2G ethanol generation: soil to soil technology
Biomass pretreatment biomass pretreatment
Cellulose and Hemicellulose hydrolysis
Hexose and Pentose fermentation
Ethanol recovery
Enzymatic degradation of recalcitrant lignin layer
Hydrolysis of cellulose & hemicellulose to hexose and pentose sugar
Utilization of hexose and pentose sugar to ethanol
Distillation to recover maximum ethanol
Residual solid biomass
Biomethane
Anaerobic
Digestion
Biomanure
Cyanobacterial
treatment

17. Different Methods of Pretreatment
Acid Hydrolysis
Alkali Hydrolysis
Physical Treatment
Physico-chemical Treatment
Biological/ Enzymatic Treatment
Costly, Hazardous, energy and labor intensive, loss of target biomolecules

18. Major Bottlenecks in Lignocellulosic Bioethanol Production
Maximum lignin degradation with minimum loss of cellulose and hemicellulose
Efficient depolymerization of lignin without the production of furfurals and hydroxymethyl furfurals
Simultaneous utilization of Pentose and Hexose sugars
Less tolerance of yeast towards higher concentration of ethanol

19. Water requirement during biomass pretreatment
Biological pretreatment requires litre of water for per litre of ethanol production compared to more than 1 or 5 litre of water requirement in physical, chemical and physico-chemical pretreatment methods

20. Leading Pretreatment technologies
Partial hydrolysis of hemicellulose
Ammonia recovery during pilot scale operation is difficult
Lignin is deposited on the surfaces of the material possibly causing blockage of cellulases to cellulose
Expensive
Needs improved method for ILs recovery and reuse

21. Enzymes/microbes involved in second generation bioethanol production
Lignolytic enzymes
Lignin peroxidase
Manganese peroxidase
Laccase
Versatile peroxidase
Cellulolytic enzymes
Endo β-glucanase
β-glucosidase
Cellobiohydrolase
Oxidative cellulases
Cellulose phosphorylase
Xylanase
Swollenin
Expansin
Yeast
Saccharomyces cerevisae
Saccharomyces pastorianus
Pichia pastoris
Saccharomyces fermentati
Saccharomyces paradoxus

22. IIT Kharagpur contribution in biofuel production

23. Enzymatic Delignification of Lignocellulosics
Microbial Biotechnology and DSP Laboratory, Department of AgFE, IIT Kharagpur
Blue laccase from a hyperactive strain of Pleurotus djamor
Yellow laccase from Lentinus squarrosulus

24. Result of Delignification
Biomass
Condition
Solid loading
(% dry weight/vol.)
% Delignification
Ricinus communis
Initial mixing followed by static 30
75-80
Lantana camara
Kans grass
Bamubusa bambos
Cotton stalk 25
77-81
Sugarcane top
Banana waste 20
70-72
Pineapple leaf waste
Rice straw
78-82
Mixture

25. SEM analysis Control Lantana camara Pretreated Control
Ricinus communis
Pretreated
Control
Bambusa Bambos
Pretreated
Control
Pineapple leaf
Pretreated

26. SEM analysis cont… Control Kans grass Pretreated Control Sugarcane top
Mixture
Pretreated

27. Porosity Analysis by BET/BGH Analyzer
Biomass
Pore size
(Before (b); After (a)) Angstrom
Pore volume (Before (b); After (a)) cm3/g
% Delignification
Ricinus communis
(b);
(a)
13.01 x 10-3 (b);
20.50 x 10-3 (a)
75-80
Lantana camara
86.74 (b);
(a)
5.968 x 10-3 (b);
7.496 x 10-3 (a)
Kans grass
61.9 (b);
128.5 (a)
4.259 x 10-3 (b);
5.365 x 10-3 (a)
Pineapple leaf waste
120 (b);
134 (a)
4.26 x 10-3 (b);
6.57 x 10-3 (a)

28. C–H methyl and methylene groups
FTIR analysis
Ricinus communis
Lantana camara
Bambusa bambos
Kans grass
Pineapple leaf
Sugarcane top
Mixture
Wavenumber (cm-1 )
Particular
OH Vibration
C–H methyl and methylene groups
O–H phenolic
lignin

29. XRD Analysis Control Lantana camara Pretreated Control Sugarcane top
Kans grass
Pretreated

30. XRD Analysis cont… Pineapple leaf Mixture Lignocellulosic biomass
Increase in crystallinity (%)
Ricinus communis
6.82
Lantana camara
7.46
Kans grass
8.54
Bambusa bambos
4.62
Pineapple leaf waste
6.25
Sugarcane top
4.30
Mixture
3.66

31. HPLC chromatogram of Lignin degradation Products
With time peak area of the individual peaks increased
Lignin degraded products released over time

32. GC-MS Analysis of lignin degraded compounds
Identified compounds
0 h
1st h
2nd h
3rd h
4th h
5th h
6th h
7th h
Retention time
Propanoic acid - +
4.748
Butanoic acid
5.731
Acetic acid
5.974
Chromone
7.167
Methanone
7.544
Pentacene
8.007
Butanedioic acid
8.194
2-Butenedioic acid
8.747
2(3H)-Furanone
9.186
3,4-Methylenedioxyphenylacetic acid
10.876
3,5-di-tert-Butyl-4-hydroxyphenyl propionic acid
11.282
3-Bromo-5-ethoxy-4-hydroxybenzaldehyde
12.269
2',6'-Dihydroxyacetophenone
15.882
11H-Dibenzo[b,e][1,4]diazepin-11-one
15.863
5(2-Dimethylamino-1-phenyl)-vinyl-thiadiazol
16.264
Trimethylsilyl-3-methoxy-4-(trimethylsilyloxy)cinnamate
17.023
Ferulic acid
17.013
Trimethylsilyl-3,4-bis(trimethylsiloxy)cinnamate
17.452
Ethyl-2-ethoxyquinoline-4-carboxylate
21.628
Salbutamol
23.876
2-(p(Dimethylamino)phenyl) benzimidazole
25.070
3,4-Dimethylbenzamide
25.375
Benzoic acid
26.664
1,2-Benzenedicarboxylic acid
26.425

33. Some of the detected compounds

34. Compounds detected during laccase mediated delignification
Low molecular weight compounds like
2(3H)-Furanone (RT 9.186),
3,4-Methylenedioxyphenylacetic acid (RT ),
Ethyl-2-ethoxyquinoline-4-carboxylate (RT )
Aromatic compounds
Trimethylsilyl-3-methoxy-4-(trimethylsilyloxy) cinnamate (RT ),
3-Bromo-5-ethoxy-4-hydroxybenzaldehyde (RT ) and
Ferulic acid (RT )
Low molecular organic acids
Propanoic acid (RT 4.748),
Butanoic acid (RT 5.731),
Acetic acid (RT 5.974),
Butanedioic acid (RT 8.194)

35. Microscopic analysis of raw and pretreated biomass
Raw biomass
Pretreated biomass

36. Established biomass pretreatment technology with different substrates
Blue laccase
Substrates: L. camara, R. communis, S. spontaneum, B. bambos, S. Officinarum tops, A. comosus leaf waste, Lignocellulosics Mixture
Optimum conditions: ºC, 6-8 h, 15-25% Solid loading, 6-7 pH, IU/mL laccase titre, 75-86% delignification %
Ref: Gujjala et al., 2016; Mukhopadhyay et al., 2011; Rajak and Banerjee, 2015; Kulia et al., 2011; Avanthi and Banerjee, 2016.
Yellow laccase
Substrates: L. camara, R. communis, S. spontaneum, B. bambos, S. Officinarum tops, A. comosus leaf waste, Lignocellulosics Mixture
Optimum conditions: ºC, 4-6 h, 15-25% Solid loading, 5-7 pH, IU/mL laccase titre, 65-87% delignification %
Ref: Mukhopadhyay et al., 2011; Rajak and Banerjee, 2016.

37. Enzymatic Saccharification of Pretreated Lignocellulosic Biomass
Delignification
A mixture of carbohydratases produced
from T. reesei RUT C30

38. Correlation between Delignification, Reducing sugar and Ethanol Production from Lignocellulosic Biomass

39. Strategies to improve the ethanol yield from lignocellulosic biomass
Ethanol (%, v/v)
SHF
SSF
CBP
PCBP
SSCF
Ricinus communis
2.78
3.58
6.81
7.12
6.24
Lantana camara
2.51
3.38
6.5
6.9
6.95
Sugarcane top
3.21
6.02
5.91
5.96
7.50
Kans grass
3.50
6.30
4.9
7.80
8.00
Pineapple leaf waste
3.25
6.42
6.78
7.18
Mixture
3.00
5.14
7.52
7.65

40. Integrated approach towards soil to soil technology
Processing
Processed solid biomass
Ethanol, butanol, lactic acid
Residual fermented biomass
By-products (Biogas)
Removal of CO2 by water stripping
Left over residual solid biomass
Enriched biomethane
CO2 as carbon source used in algal cultivation
Enrichment with cyanobacteria
Biomanure
applied back to the soil
Fermentation (MAC)
Fermentation
Microbes / Enzymes
Biodiesel
Agricultural residue

41. Pilot Plant facility for Ethanol Production
(Capacity: 500 kg fresh biomass processing per batch)

42. Lignocellulosic
Biomass
Hot Air Oven
Pulverizing unit

43. Saccharification tank
Pilot plant
Pre-treatment tank
Saccharification tank
Fermentation tank
Tincture press
Gantry system
Control panel

44. Distillation assembly
Boiler assembly
Distillation assembly

45. Gantry system for lifting the immobilization tray
Biomass collection
tank
Gantry system for lifting the immobilization tray

46. Students’s Contribution
Dr. Mainak Mukhopadhyay
Dr. Arindam Kuila
Mr. Rajiv Ch. Rajak
Mr. Sanjeev Kumar
BIOENERGY GROUP AT P.K.SINHA CENTRE FOR BIOENERGY,
IIT KHARAGPUR
Ms. Althuri Avanthi
Ms. Anjani
Devi Chintagunta
Ms. Knawang Ch. Sherpa
Mr. G. Lohit K. Srinivas
Mr. Debajyoti Kundu Ms. Tania Dev Ms. Reddhy Mahle

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