Process optimization and mass balance studies of pilot scale steam explosion pretreatment of rice straw for higher sugar release

Biomass and Bioenergy 130 (2019) 105390

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Research paper

Process optimization and mass balance studies of pilot scale steam explosion pretreatment of rice straw for higher sugar release Surbhi Semwal a, Tirath Raj a, Rahul Kumar a, Jayaraj Christopher b, Ravi P. Gupta a, Suresh K. Puri a, Ravindra Kumar a, *, S.S.V. Ramakumar b a b

DBT-IOC Centre for Advanced Bioenergy Research, Research & Development Centre, Indian Oil Corporation Limited, Sector-13, Faridabad, 121007, India Research & Development Centre, Indian Oil Corporation Limited, Sector-13, Faridabad, 121007, India

A R T I C L E I N F O

A B S T R A C T

Keywords: Rice straw Steam explosion Glucan conversion Enzymatic hydrolysis Mass balance

The present study deals to visualise the impact of various process parameters, i.e. particle size and impregnation media over sugar released during steam explosion pretreatment. For this, 5, 10 and 20 mm rice straw sizes were impregnated in water and dilute acid media (1%) followed by steam explosion at 180 and 200 � C. Pretreated slurry was further hydrolyzed by 5 and 10 FPU g 1 of residue varying 15 and 20% solid loading. The result showed that 10 mm particle size gave highest glucan conversion (88.7%) in acid impregnated steam explosion at 180 � C using 5 FPU g 1 of residue with 15% solid loading. Comparatively, water impregnated pretreated biomass results significantly lower glucan conversion (61.1%), which was further intensified to 77.7% at 10 FPU g 1 of residue with increased temperature. Furthermore, mass balance, compositional and structural transformation studies support our finding. Overall 30.6–81.1% sugar recovery was achieved with/or without acid pretreatment respectively.

1. Introduction

followed by India at 156.6 million tonnes year 1 [8]. In India, 23% of rice straw is either left in the field for natural decay or burnt for prep­ aration of land for next crop. Biomass burning results in emission including fine particulate matter (PM 2.5), carbon dioxide, carbon mono oxide, sulphur and nitrogen oxides, nitric oxide, and methane, which may directly or indirectly add to the climate change. Moreover, it is also not suitable to use as an animal fodder due to the lower protein and high silica content. Thus, the utilization of surplus rice straw for the ethanol production could be beneficial for sustainable energy production and will improve rural economies and environment [9]. Rice straw is reported to contain >50% of fermentable sugars [2]. However, recalcitrant nature of plant cell wall makes it rigid for any chemical or biochemical deconstruction to fermentable sugars. There­ fore, pretreatment is prerequisite step to alter the structural recalci­ trance of plant cell wall resulting to improve the carbohydrates hydrolysis and increase fermentable sugars yields [10,11]. In general, the pretreatment method includes physical (milling and grinding), physicochemical (steam explosion, hydro-thermolysis, wet oxidation, etc.), chemical (alkali, dilute acid, ionic liquids and organic solvents and biological) [12–16]. Among these, steam explosion (SE) is one of the most effective and favoured methods due to the minimum use of

With an increased population and economic growth, the primary energy consumption was 13,511 million tonnes oil equivalent (Mtoe) in 2017 and is expected to increase to 17,866 Mtoe by 2040 respectively [1]. Thus, it is necessary to explore alternative energy sources, which can meet these energy demands and also mitigate greenhouse gas (GHG) emissions [2]. Various countries across the globe have drawn their plan to reduce GHG emissions by switching over to ethanol, which is considered as an alternative to transportation fuel. Ethanol can be pro­ duced from sugars, starch and lignocellulosic biomass (LCB) [3–5]. The production of ethanol using the sugarcane or grain starch results in food vs. fuel debate. Therefore, lignocellulosic biomass (LCB) such as wheat straw, rice straw, cotton stalk, corn stover, sugarcane bagasse, Acacia mangium and corn cob etc. are the potential feedstock for ethanol pro­ duction [6,7]. Among the crop residues, rice straw is a by-product of rice (Oryza sativa) and is one of the most abundant and promising agricultural waste available across the globe. Based on the Food and Agricultural Organi­ zation (FAO), the annual rice production in the world is 740.3 million tonnes, wherein, China alone produces 209.8 million tonnes year 1

* Corresponding author. E-mail address: [email protected] (R. Kumar). https://doi.org/10.1016/j.biombioe.2019.105390 Received 23 May 2019; Received in revised form 16 September 2019; Accepted 23 September 2019 Available online 25 September 2019 0961-9534/© 2019 Elsevier Ltd. All rights reserved.

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chemicals. SE depolymerise hemicellulose, partly melts lignin, and dis­ rupts complex carbohydrate matrix facilitating the enzymatic hydrolysis of cell wall polysaccharides [17]. SE increases the accessible surface area of biomass due to the removal of hemicellulose and relocation of lignin, which make cellulose fibre accessible to enzymes with improved hydrolytic performance. The limitation of SE is the formation of inhibitors including xylo-oligomers, which are detrimental to the enzyme and fermenting yeast [18]. The condensation and precipitation of soluble lignin components on pre­ treated solid surfaces inhibit enzymatic hydrolysis either as a physical barrier and/or by non-productive protein binding [7]. The impregnation of dilute acid prior to pretreatment can increase the hemicellulose sol­ ubilisation. Kataria et al. [19] studied the acid mediated SE of elephant grass biomass resulted 7-fold increase in saccharification yield as compared to untreated biomass. Fan et al. [20] observed that the con­ version of cellulose in corncob by acid impregnated SE was 85.3%, which was 1.6 times higher than that of without acid. Liu et al. [21] studied steam explosion of corn stover with a different particle size (2.5–0.5 cm) and observed that enzymatic hydrolysis increased with crystallinity index. Harun et al. [22] studied the AFEX with two different particle sizes and observed that a larger particle of rice straw signifi­ cantly gives higher sugar conversion (85.9%) as compared to smaller particle size using high severity. Hence, an optimum particle size of biomass is very important for commercialization. For large scale biorefinery, reduction of particle size is an essential but energy intense process [23]. Hence, the present study deals with the process optimization of steam explosion pretreatment by varying the pretreatment conditions, i.e. biomass particle size, reaction tempera­ ture, impregnation media for high sugar conversion using different solid loading and enzyme doses with minimum by product (inhibitors) for­ mation. In addition, compositional analysis of native and pretreated biomass, FT-IR characterisation and mass balance studies were found to support our findings.

experiments, steam explosion digester was flushed 2 to 3 times with steam of 1–1.5 MPa to quickly attain the desired operating temperature in actual experiments. For instance, 450 g (dry basis) of pre-soaked and pressed rice straw (moisture ~60%) was introduced in the digester and temperature was increased by injecting high-pressure steam (1.5 MPa) rapidly. The reactor was maintained at an operating temperature of 180 or 200 � C for 10 min residence time. After incubating for 10 min at desired time and pressure, the outlet ball valve was instantaneously released into reception chamber, this rapid explosive decompression rupture/breaks the intrinsic complex matrix of biomass. Steam exploded pretreated straw was then cooled to room temperature and was recov­ ered from cyclone separator. Slurry received from cyclone separator was fractionated into two fractions: xylose-rich pretreatment hydrolysate and cellulose-rich solid residue, which was further used for enzymatic hydrolysis. For each set of experiments, consecutive 5 explosions were conducted and pretreated rice was collected from the cyclone separator to minimize the error. The overall research methodology is summarized in Fig. 1. 2.3. Analysis of pretreated residue Compositional analysis of cellulose-rich residue was performed by two-stage acid hydrolysis following the standard protocol developed by National Renewable Energy Laboratory (NREL, USA) [24]. Monomeric sugars and inhibitors concentration in xylose-rich hydrolysate received after pretreatment were quantified by HPLC (M/s Waters Gesellschaft Gmbg, Austria) equipped with an Aminex HPX-87P column (300 mm � 7.8 mm, Bio-Rad. Laboratories Inc.) and a refractive index detector as demonstrated by NREL [24,25]. The mobile phase used was Milli-Q water at a flow rate of 0.6 mL min̶ 1 at a temperature of 80 � C. Inhibitors present in liquid hy­ drolyzate were analysed using Bio-Rad Aminex HPX-87H column at 50 � C, 0.005 M H2SO4 at a flow rate of 0.6 mL min̶ 1 as mobile phase equipped with UV detector. Both the columns were equipped with suitable guard columns. Oligomeric sugars in hydrolysate were analysed following NREL LAP using 4% sulfuric acid [24]. Three replications

2. Materials and methods Rice (Oryza sativa) straw was collected from Mathura, Uttar Pradesh (27.28� N 77.41� E) at the time of harvesting. It was air dried and milled to the particle size of 5, 10 and 20 mm by a knife mill and stored in sealed bags at 25 � C until further use. All experiments were conducted using a single lot of rice straw. Cellulase (Cellic CTec3) was obtained from M/s Novozymes, Ban­ galore (India). Cellobiose, glucose, xylose, arabinose, acetic acid, formic acid, hydroxymethylfurfural (HMF), furfural, calcium carbonate and sulfuric acid (purity >98%) were obtained from M/s S.D. Fine Chemicals (India). All the chemicals were of analytical grade and used without any further purification. 2.1. Impregnation of biomass Rice straw (3.0 kg) was initially impregnated in either water or 1% (w/w) dilute sulfuric acid (DA) in a 100 L stainless steel (SS) container at room temperature for 1 h. The ratio of liquid to solid was maintained as 19:1 in each set of experiments. The soaked rice straw was dewatered using a pneumatic hydraulic press at 10 MPa for 15 min. The pressed rice straw was properly mixed to determine moisture content using an IRbased moisture analyzer (MA 150 Sartorius, Germany) and found to contain about ~60% moisture. 2.2. Steam explosion experiments Steam explosion pilot plant was designed in-house, mainly comprising of high pressure and high temperature reactor made of stainless steel of 10 L capacity equipped with feeding hopper, cyclone separator, quick opening pneumatic butterfly valve, steam boiler and a noise absorber was used for experimentation. Before starting the

Fig. 1. Schematic representation of the overall research methodology. 2

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were measured for each biomass samples.

3. Results and discussion

2.4. Enzymatic hydrolysis

3.1. Optimization of straw size

Cellulases cocktail (Cellic CTec3) used in the present study was found to contain β-glucosidase: (5700 IU mL 1, filter paper unit (155 FPU mL 1), endo-glucanase (6555 IU mL 1). β-glucosidase activity was determined as described earlier [26]. Filter paper units (FPU) and endoglucanase (CMCase) activities were analysed according to the method described previously [27]. Cellulose-rich residue obtained after pretreatment was washed with water and enzymatic hydrolysis was conducted at 15 and 20% WIS (water-insoluble solids). To attain the desired solid loading of 15 and 20%, ~7.5 and 10.0 g of pretreated biomass (oven dry weight, ODW) were taken in 500 mL Erlenmeyer flasks and suspended in 0.05 M so­ dium citrate buffer (pH 5.0), making final volume up to 50 mL. The mixture was pre-incubated at 50 � C for 30 min followed by adding cel­ lulases of 5 and 10 FPU g 1 WIS. The mixture was incubated in an orbital shaker at 50 � C up to 72 h at 200 rpm. The samples were withdrawn at various time intervals, centrifuged and heated at 80 � C for 5 min to denature the enzyme followed by centrifugation (8000 g for 10 min). Samples were filtered using a 0.45 μm filter and analysed for sugars by HPLC as described above. The conversion of enzymatic hydrolysis was calculated by using the equation below as referenced by Zhu et al. [28]. For instance, glucan conversion % (Yg) can be calcualted using Equation (1). � � � � � � Cg VVh0h Cg0 * fts0*1xis0 1 � �*100 Yg ð%Þ ¼ (1) ϕG *ρh0 *xG0 þ ϕgos *Cgos0 * fts0*1xis0 1

Pretreatment efficiency was measured by varying the straw sizes from 5 to 20 mm at the different severity and enzymatic hydrolysis of the pretreated slurry was conducted at 20% solid loading for sugar recovery. After SE pretreatment, the pretreated residue was washed and subjected to enzymatic hydrolysis. The detailed process parameters and the glucan conversion of the respective experiments are summarized in Table 1. The glucan conversion after enzymatic hydrolysis of 5, 10 and 20 mm straw size were 53.6, 61.1, 27.9% (W-5-180, W-10-180, W-20-180) and 75.7, 74.8, 74.5% (W-5-200, W-10-200, W-20-200) using water as a pretreatment media at 180 and 200 � C respectively. Whereas, while using dilute acid as an impregnating media, the glucan conversion were 87.7, 88.7, 83.4% (A-5-180, A-10-180, A-20-180) and 88.3, 89.6, 89.3% (A-5-200, A-10-200, A-20-200) respectively. These results suggested that SE at 200 � C while using water and 180 � C and 200 � C while using acid media are the better performers in terms of glucan conversion using 5 FPU g 1 of pretreated residue in an individual set of experiments. The glucan conversion using 5 mm (W-5-180) of straw upon SE at 180 � C in water media results in 53.6%, which is significantly lower than 10 mm (61.1% in W-10-180). However, SE at 200 � C after enzymatic hydrolysis results in almost similar glucan conversion for all sizes, i.e. W-5-200�W10-200�W-20-200. Similarly, SE using the acidic media at 180 and 200 � C after enzymatic hydrolysis results in a very narrow range of glucan conversion (83.4–89.6%) with no clear-cut indication of the su­ periority of the performance. Reduction of biomass size is energy intensive, and higher particle size is always preferred for economic viability [23]. After initial screening based upon glucose release, the study was further focused on two particle sizes, i.e. 10 and 20 mm, only as particle size below 10 mm didn’t significantly enhance the glucose yield.

where, Cg is concentration of glucose (g L 1); Vh is volume of hydrolyzate liquid (L); Vh0 initial volume of liquid (L); Cg0 initial concentration of glucose (g L 1); fts0 initial mass fraction of total solids (soluble solids þ insoluble solids) in slurry; xis0 initial mass fraction of insoluble solids in total solids; ϕG molecular weight ratio of glucose to glucan monomer; (ϕG ¼ 180/162 ¼ 1.11); ρh0 initial density (g L 1) of liquid at 25 � C; xG0 initial mass fraction of glucan in insoluble solids; ϕgos ratio of glucose molecular weight to average monomer weight of glucose olig­ omers (ϕgos ¼ 180/166.5 ¼ 1.08); Cgos0 initial concentration of glucose oligomers (g L 1). Similarly, xylan conversion was also calculated based upon xylose concentration using same equation.

3.2. Compositional analysis of native and pretreated residue The compositional analysis of the native RS was found to be 37.8% of cellulose, 21.6% of hemicelluloses, 13.6% of lignin, 1.4% of acetic acid, Table 1 Pretreatment process parameters and enzymatic hydrolysis of the different straw size of rice straw.

2.5. Structural transformation Diffuse reflectance spectra of untreated and pretreated biomass samples were recorded using Shimadzu (Model FT-IR Prestige-21), FTIR spectrometer to visualise the structural transformation. All spectra were recorded in the absorbance mode from an accumulation of 200 scans at 4 cm 1 resolution over the range 4000 400 cm 1. Microcrys­ talline cellulose (Avicel PH 101) was used as the reference materials.

Set No

Sample ID [IMSS-T]a

Process Parameters Impregnation media

Straw Size (mm)

SE Temperature (� C)

Set 1

W-5-180 W-10180 W-20180

Water

5 10

180

W-5-200 W-10200 W-20200

Water

A-5-180 A-10-180 A-20-180

Dilute acid

5 10 20

180

87.7 � 1.00 88.7 � 0.70 83.4 � 0.50

A-5-200 A-10-200 A-20-200

Dilute acid

5 10 20

200

88.3 � 0.90 89.6 � 1.01 89.3 � 0.80

2.6. Mass balance Mass balance a study was conducted based on weight of biomass (dry basis) received after SE pretreatment and after enzymatic saccharifica­ tion for complete sugar recovery.

Set 2

2.7. Statistical analysis The compositional analysis and enzymatic hydrolysis data have been reported as the average of three replicates. The standard deviation of each data point has been calculated by one-way ANOVA using the posthoc Tukey test (trial version) available at statistica.mooo.com with p < 0.05 was set as the level of statistical significance.

20 5 10

Enzymatic hydrolysis, 72 h (%)b 53.6 � 1.02 61.1 � 1.52 27.9 � 0.82

200

20

75.7 � 0.62 74.8 � 0.65 74.5 � 1.22

a IM-SS-T refers to Impregnation media (W stands for water impregnation and A stands for acid impregnation); the straw size of rice straw (5, 10 and 20 mm); and SE temperature (180 and 200 � C) respectively. b Enzymatic hydrolysis was taken at 15% solid loading using 5 FPU g 1 WIS.

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13.2% of ash and 16.1% of extractives (Table 2). After SE, cellulose was enriched in the pretreated solid residue. The cellulose, hemicellulose, lignin and ash content of pretreated rice straw were in the range of 45.6–52.5, 24.3 to 5.7, 16.8 to 29.0 and 11.5–19.4% respectively. It was observed that the increase in temperature results in the increase in cellulose content in the case of water impregnated pretreated rice straw (Table 2, Set 1). This may be attributed to primarily due to the removal of xylan and/or lignin and due to the removal of extractives. Similar observations of enrichment of biomass with cellulose were also reported in the previous study [13]. This was also noted that the hemicelluloses content increased as compared to native, from 21.6 to 24.1% (W-10-180) and 24.3% (W-20-180) respectively. Whereas, at 200 � C, the hemicellulose content significantly reduced from 21.6 to 8.9% in W-10-200 and to 8.6% in W-20-200. This clearly demonstrates that the temperature has a direct impact on hemicelluloses solubilisation, whereas, the impact of straw size on carbohydrate contents was negli­ gible. This argument was further supported by the hemicellulose sol­ ubilisation. The hemicellulose solubilisation was 28.8 and 78.2% in W-10-180 and W-10-200 and 23.0 and 76.3% in W-20-180 and W-20-200 respectively, which were having nearly the same pattern of increment after SE when the temperature was increased from 180 to 200 � C while keeping the same straw size. Therefore, it may be argued that an increase in the pretreatment severity, due to increase in tem­ perature results in higher xylan solubilisation and consequently reduc­ tion in xylan content in the pretreated residue. Similar pattern while change in temperature over xylan removal has also reported [29,30]. In contrast, in an acid media (Table 2, Set 2) both cellulose and hemicellulose content are decreased when the SE temperature is increased, ranges from 51.0 to 45.6% and 7.4 to 5.7% respectively. The cellulose and hemicellulose content were slightly decreased from 51.0 to 50.0% and 7.4 to 5.7% in A-10-180 to A-10-200 and 49.6 to 45.6% and 7.4 to 6.0% in A-20-180 to A-20-200 respectively. A meagre difference of hemicellulose solubilisation is found in between A-10-180 to A-10200 and A-20-180 to A-20-200. This signifies that the DA impregnation prior to SE has solubilised higher hemicelluloses even at a lower tem­ perature (180 � C) and results similar to 7.4% in pretreated residue using 10 and 20 mm size. This implies a higher degree of hydrolysis of hemicelluloses during pretreatment. Almost similar results are obtained using 20 mm of rice straw. The slightly lower cellulose content with higher straw size, i.e. 49.6 and 45.6% in A-20-180 and A-20-200 are observed. Although the decrease in the cellulose content after SE while using 20 mm size remained unexplained, however, an increase in lignin content to 29.0% justify the degradation of cellulose and the formation of pseudo-lignin. Acetyl is the part of the hemicellulose backbone present in the form

of acetate in pentose at C2 or C3 as O-acetyl. Acetyl content is only observed in acid impregnated pretreated residue ranges from 0.3 to 0.5%, however, in water impregnated pretreated residue, it is not detected. This may be attributed to the lower pH, which facilitated the hydrolysis of the ester bond linking between the acetyl and hemicellu­ lose backbone. Across all the experiments, the lignin content increased from 13.6 in native to 20.8% in the water impregnated pretreated res­ idue and to 29.0% in dilute acid impregnated pretreated residue. It is evident from the results (Table 2) that the higher temperature results in higher lignin content and increasing the severity by reducing the pH results in a further increase in the lignin content. The increase in lignin content with severity can be explained due to the formation of pseudolignin. Similar results of an increase in lignin content while increasing the severity of the pretreatment were also reported in literature [21,31]. 3.3. Analysis of pretreatment hydrolysate The monomeric and oligomeric sugars (C6 and C5) and their degra­ dation products such as HMF, furfural, acetic acid and formic acid formed during pretreatment are summarized in Table 3. In the case of water impregnated SE (Table 3, Set 1) sugars are present mainly in oligomeric form, between 8.2 and 32.9% (% of sugar present in the native biomass) produced by the partial breakdown of hemicelluloses [32]. The concentration of sugars (monomer and oligomers) decreases with increased size using SE at 180 � C. Whereas, at 200 � C the glucose and xylose concentration in hydrolysate of 10 and 20 mm straw size was almost similar, i.e. 1.3 and 5.4% in W-10-200 and 1.2 and 5.2% in W-20-200. Moreover, at a higher temperature in both the size the con­ centration of gluco- and xylo-oligomers was significantly decreased from 4.3 to 3.2% and 28.6 to 5.0% respectively. That is mainly due to the conversion of oligomers to monomers, which can be attributed to the higher degree of hydrolysis while employing higher temperature. It is worth mentioning that acid impregnated SE at 200 � C results in the higher acetic acid formation with respect to temperature, i.e. 820 (W-10-200), 827 (W-20-200), 556 (W-10-180) and 355 (W-20-180) mg/ 100 g of dry biomass residue as given in Table 3, Set 1. This data sup­ ports the argument given in section 3.2, wherein content of acetyl in the pretreated residue is higher at 180 � C SE (0.5%) than the 200 � C (0.3%). Moreover, the high amount of monomeric sugar present in pre­ treatment hydrolysate signifies higher solubilisation of the hemi­ celluloses at 200 � C. It is interesting to note that the high amount of xylose in liquid fraction typically also generated a high conc. of in­ hibitors. In contrast, high amount of glucose in liquid fraction produce a lower amount of HMF. This may be attributed to that the xylan is more labile for hydrolysis and degradation in comparison to glucan. Similar

Table 2 Chemical composition of native and pretreated rice straw. Sample ID [IM-SS-T]a Set No

Chemical Composition (%) Cellulose

Hemicellulose

Acetyl

Ligninb

Ash

Extractivec

N/A

Native RS

37.8 � 0.39

21.6 � 0.10

1.4 � 0.02

13.6 � 0.10

13.2 � 0.01

16.1 � 0.80

Set 1

W-10-180 W-20-180

45.9 � 0.40 46.8 � 0.20

24.1 � 0.30 24.3 � 0.10

0.5 � 0.01 0.5 � 0.01

17.6 � 0.10 16.8 � 0.50

12.0 � 0.10 11.5 � 0.20

n/a n/a

W-10-200 W-20-200

52.1 � 0.51 52.5 � 0.62

8.9 � 0.21 8.6 � 0.32

0.3 � 0.01 0.5 � 0.01

21.7 � 0.20 20.8 � 0.32

16.9 � 0.30 17.6 � 0.40

n/a n/a

A-10-180 A-20-180

51.0 � 0.31 49.6 � 0.43

7.4 � 0.50 7.4 � 0.09

0.0 � 0.00 0.0 � 0.00

25.5 � 0.50 24.6 � 0.50

16.1 � 0.20 18.4 � 0.36

n/a n/a

A-10-200 A-20-200

50.0 � 0.90 45.6 � 0.60

5.7 � 0.05 6.0 � 0.02

0.0 � 0.00 0.0 � 0.00

27.8 � 0.32 29.0 � 0.60

16.5 � 0.50 19.4 � 0.10

n/a n/a

Set 2

n/a- Not Any. a IM-SS-T refers to Impregnation media (W stands for water impregnation and A stands for acid impregnation); the straw size of rice straw (5, 10 and 20 mm); and SE temperature (180 and 200 � C) respectively. b Lignin includes both acid soluble and acid insoluble lignin. c Extractives include water and ethanol extractives. 4

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Table 3 Sugar and Inhibitors released in the pretreatment hydrolysate. Sample ID [IM-SS-T]a

b

c

Monomers (%) Set 1

Set 2

Inhibitors concentration (mg/100 g of impregnated biomass)d

Pretreated Hydrolysate Oligomers (%)

Glucose

Xylose

Gluco-

Xylo-

FAe

AAf

HMFg

Furfural

W-10-180 W-20-180

0.8 � 0.02 1.0 � 0.01

2.3 � 0.01 1.3 � 0.02

5.3 � 0.10 2.0 � 0.06

17.9 � 0.52 10.2 � 0.40

105 0

556 355

8 2

15 2

W-10-200 W-20-200

1.3 � 0.03 1.2 � 0.05

5.4 � 0.03 5.2 � 0.10

4.3 � 0.03 3.2 � 0.12

28.6 � 0.70 5.0 � 0.30

483 202

820 827

92 94

241 225

A-10-180 A-20-180

7.1 � 0.50 7.4 � 0.20

52.6 � 0.20 51.5 � 0.60

n/d n/d

n/d n/d

436 127

839 842

325 204

1221 537

A-10-200 A-20-200

11.3 � 0.10 8.2 � 0.20

40.2 � 0.90 33.2 � 0.45

n/d n/d

n/d n/d

1064 297

1193 1004

1139 797

1993 1285

a

IM-SS-T refers to Impregnation media (W stands for water impregnation and A stands for acid impregnation); straw size of rice straw (5, 10 and 20 mm); and SE temperature (180 and 200 � C) respectively. b Monomer (Glucose/Xylose),% ¼ Glucose/xylose released in pretreatment hydrolysate (g)/Glucose/xylose present in native RS (g) � 100. c Oligomers (Gluco- and Xylo-oligomers),% ¼ Gluco- or Xylo-oligomer released in pretreatment hydrolysate (g)/Glucose/xylose present in native RS(g) � 100. d Inhibitors concentration (FA, AA, HMF and Furfural), mg/100 g of impregnated RS ¼ FA or AA or HMF or Furfural concentration in pretreatment hydrolysate in mg/Impregnated RS taken during pretreatment (g) � 100. e Formic acid. f Acetic acid. g Hydroxymethyl furfural; n/d-not detected.

Fig. 2. Time course of glucan conversion at different straw size and impregnation media. 5

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results of degradation product formation pattern are also reported in earlier studies [13]. Contrary to the above, dilute acid impregnation promotes the con­ version of xylo-oligosaccharide to xylose during SE (Table 3, Set 2). The solubilisation of hemicelluloses to xylose results in reduced residual hemicellulose as determined by compositional analysis (Table 2). Further, it is observed that the concentration of monomers elevated in acid impregnated SE (Table 3, Set 2) as compared with water impreg­ nated SE (Table 3, Set 1). However, no oligomers are detected in acid impregnated SE hydrolysate. Kapoor et al. [13] also reported that dilute acid during pretreatment renders higher monomeric sugars. When the SE temperature was increased from 180 to 200 � C, the glucose concentration is increased in hydrolysate from 7.1 to 11.3% (10 mm) and 7.4–8.2% (20 mm) and xylose concentration decreased from 52.6 to 40.2% (10 mm) and 51.5 to 33.2% (20 mm). It is interesting to note that concentration of total monomers at both temperatures (180 and 200 � C) is decreased from 59.7 to 58.9% and 51.5 to 41.4% with increasing the straw size from 10 to 20 mm. At higher temperature, DA promotes the formation of inhibitors, i.e. HMF and furfural and observed from 325 to 1139 and 1221–1993 mg/100 g in 10 mm straw size and 204 to 797 and 537–1285 mg/100 g of dry biomass in 20 mm straw size. It is reported that these degradation products can inhibit the growth of ethanol fermenting yeast S. cerevisiae and E. Coli [33]. However, the furfural and HMF concentrations in the hydrolysate are lower than the reported levels (~2.0 g L 1) for both impregnation media. Similarly, a higher amount of acetic acid and formic acid are also detected in pre­ treatment hydrolysate at higher severity [21,34].

3.5. Hemicellulose removal vis-a-vis glucan conversion Hemicellulose solubilisation was expected to improve the enzymatic hydrolysis. The higher hemicellulose content present in W-10-180 and W-20-180 residues as determined by compositional analysis also sup­ ports the relatively poor solubilisation of hemicellulose in these samples (Fig. 3a). In contrast, lower hemicellulose content present in acid impregnated SE support the maximum hemicellulose solubilisation (77.3, 76.3, 83.0 and 82.8%) (Fig. 3b) resulted in much higher enzy­ matic hydrolysis (88.7, 83.4, 89.6 and 89.3%) in A-10-180, A-20-180, A10-200 and A-20-200 respectively (Fig. 2c). Hemicellulose solubilisation was enhanced by using DA prior to SE leading to decompose the sur­ rounding xylan, acid soluble lignin and cellulose fibre results to the restructuring of the lignin, consequently, with more exposure of the cellulose to the enzyme. Almost similar glucan conversion is observed while conducting SE in acid media even after variation in temperature and enzyme dosages. Therefore, it may be concluded that a higher particle size can be dealt to get higher sugar recovery by using the acidified media. 3.6. Impact of solid loading in glucan conversion For the economic feasibility of sustainable bioethanol process from biomass, higher sugar concentration in enzymatic hydrolysate is preferred with reduction of total energy utilization/cost for distillation of the ethanol from fermenting broth. Thus, the threshold of >4 wt % ethanol concentration (theoretical yield of ethanol from glucose is 0.51 g g 1) in the fermentation broth is required for distillation (with minimum energy cost), which require the minimum sugar concentration of ~8–9 wt% in the enzymatic hydrolysate. In order to achieve this target, the minimum solid content of polysaccharide-rich biomasses should be ~20 wt % for enzymatic hydrolysis [40,41]. Most of the literature reported that the used of low solid loading, i.e. <10% attrib­ uted to less sugar concentration in the sugar hydrolysate, which require a larger reactor volume and hence not economical for large-scale pro­ duction [7,42]. Therefore, it would be interesting to conduct the hy­ drolysis at the desired solid loading for maximum sugar conversion with reduced energy inputs. We found that, using 15% WIS, a maximum of 61.1% glucan hydrolysis was achieved for W-10-180 treatment; how­ ever, at 20% WIS, the glucan conversion was reduced to 48.5% using 5 FPU g 1 WIS residue of the enzyme (Fig. 4). While conducting the SE at 200 � C glucan conversion increases to 74.8% at 15% loadings and to 71.9% at 20% loading. These results showed that higher solid loading attributes to lower hydrolysis, which might be due to increase in viscosity, therefore prevents efficient mixing and mass transfer. A decreased enzymatic hydrolysis with higher WIS content has also been reported by others [36]. Although, in acidic media, insignificant decreased of glucan conversion was observed at high solid loading. For example, at 15% solid loading, glucan conversion was 88.7% and 89.6% of A-10-180 and A-10-200; whereas at 20% solid loading, it was reduced to 87.4 and 86.9% respectively (Table SI 1). This might be attribute to prominent solubilisation of hemicellulose during pretreatment leading to more exposed cellulose with the better acces­ sibility of cellulases, subsequently; give higher enzymatic hydrolysis. Table SI 1 shows the glucan conversion of 5, 10 and 20 mm straw size with 5 FPU g 1 WIS using 15 and 20% solid loading in 48 and 72 h in­ cubation time. However, the hydrolysis can be improved by increasing the enzyme dosages. Therefore, it may be argued that there is a trade-off between loadings and glucan conversion, hence, need to be carefully optimised for better economy. Kapoor et al. [29] studies the DA pretreated rice straw at 20 and 25% solid loadings yielded 72.0%, 54.0% glucan con­ version and 87.0, 83.3 g L 1 glucose concentration respectively. Kris­ tensen et al. [38] studied the hydrolysis of filter paper at 5–30% solid loading and revealed that the decreasing yield of enzymatic hydrolysis of cellulose was due to inhibition of enzyme adsorption by the hydrolysis

3.4. Enzymatic hydrolysis of pretreated residue The time course of glucan hydrolysis of the pretreated residue with different straw size at 180 and 200 � C using 15% loading is shown in Fig. 2. The enzymatic hydrolysis of water impregnated pretreated res­ idue is very sluggish for 20 mm straw at 180 � C with 14.3% glucan conversion, whereas the 10 mm straw results in much higher, i.e. 51.8% after 24 h of incubation. The trend of the glucan conversion remains the same till 72 h with final glucan conversion of 27.9% and 61.1% respectively (Fig. 2a). A huge difference in the pretreatment efficiency may be attributed to the (i) high viscosity, which prevents efficient mixing and mass transfer; (ii) product inhibition and (iii) concentration of other compounds (lignin, organic acids) inhibit cellulolytic enzymes [35,36]. High vis­ cosity in enzymatic hydrolysis and fermentation steps poses challenges in stirring and mixing leading to higher energy demand. Reduction of straw size before pretreatment has been demonstrated to reduce the viscosity of the slurry [37]. Product inhibition has been responsible to play an important role in enzymatic hydrolysis [38]. Interestingly, it has recently been observed that glucose and cellobiose strongly inhibit cellulases linearly [39]. However, the enzymatic hydrolysis while con­ ducting SE at 200 � C results in almost similar glucan conversion, i.e. 74.8 and 74.5% of the respective straw size. As discussed, the apparent reason for slow hydrolysis using 20 mm at 180 � C SE is the mass transfer and mixing problem, as the residue after pretreatment remains straw itself, whereas at 200 � C, it is broken into the small pieces resulting to better mass transfer. Moreover, improvement of glucan conversion is observed at a higher temperature, which is due to prominent solubilisation of hemicellulose during pretreatment leading to more exposed cellulose with the better accessibility of cellulases, subsequently results in higher enzymatic hydrolysis. This suggests that the increase in pretreatment temperature may favour breakdown of the hydrogen bonds within the crystalline region of cellulose. With high dosages of the enzyme, i.e. 10 FPU g 1 WIS of pretreated residues results in higher glucan conversion as illustrated in Fig. 2b. However, the marginal difference of glucan conversion, i.e. <2–10% was observed for acid treated residue with increase enzyme dosages from 5 to 10 FPU g 1 WIS of pretreated residue (Fig. 2d). 6

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Biomass and Bioenergy 130 (2019) 105390

Fig. 3. Glucan conversion with 10 and 20 mm straw size as a function of solubilisation during a steam explosion at 15% solid loading using 5 FPU g residue in 72 h incubation time: (a) Water impregnated rice straw SE at 180 and 200 � C; (b) Acid impregnated rice straw SE at 180 and 200 � C.

1

WIS pretreated

Fig. 4. Glucan conversion and total sugar of water impregnated pretreated rice straw (10 mm) SE at 180 and 200 � C at 15 and 20% solid loading using 5 FPU g 1 WIS for 72 h incubation time.

product. It is clear from the above discussion that low solid loading re­ sults in higher glucan conversion with reduced overall sugar concen­ tration. Lower sugar concentration results in lower ethanol titre leading higher distillation cost. 3.7. FT-IR spectral analysis The FT-IR spectral analysis of native, water and acid impregnated SE pretreated rice straw are employed to investigate the alteration in the chemical structure transformation (Fig. 5). The absorption peak at – O and C–O bonds of 1720 cm 1 is mainly attributed to ester linkage C– acetyl group in hemicellulose structure. The decrease in the intensity of these bands is attributed to the deacetylation/degradation of hemi­ celluloses [7,42]. Intensity at 1720 cm 1 is pronounced in water impregnated pretreated residue at 180 � C (W-10-180 and W-20-180). Furthermore, the intensity of this band was diminished in W-10-200 and W-20-200 with increased temperature from 180 to 200 � C. This effect was much more pronounced in the case of DA treatment. Since the addition of acid in the pretreatment media catalyze about ~80–85% hemicellulose solubilisation. This positively enhances the enzymatic hydrolysis with ~89% of glucan conversion as shown in Fig. 3. The peaks 1614, 1512 and the 1429 cm 1 are attributed to aromatic ring vibrations of lignin compounds, together with the bands at – O conjugated to aromatic rings) and at 1657 cm 1 (stretching of C– – O unconjugated to aromatic rings). It 1726–1710 cm 1 (stretching of C– might be hypothesized that the increased lignin content mighty be attributed to the grafting process, where soluble phenols generated by SE were incorporated into the biomass fiber. The peaks intensity around

Fig. 5. FT-IR of native and pretreated rice straw with different straw size: (a) Water impregnated RS (SE at 180 and 200 � C); (b) Acid impregnated RS (SE at 180 and 200 � C).

1083, 1135 and 1172 cm 1 were associated with cellulose C–O–H, C–O–C stretching, which increases with increased glucan content, i.e. 52.1 and 52.5% in W-10-200 and W-20-200 respectively as compared to untreated straw (37.8%) (Fig. 5a and Table 2). A similar observation is also visualized in the case of DA-SE treated rice straw (Fig. 5b). Intensification of these peaks after water (SE at 200 � C) and acidic (SE at 180 and 200 � C) pretreated rice straw might be due to increased 7

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Biomass and Bioenergy 130 (2019) 105390

lignin content from 20.8 to 29.0% (Fig. 5 and Table 2). With that, the decreased intensity of above bands in water impregnated at lower SE temperature (W-10-180 and W-20-180) could be correlated with the low amount of lignin in pretreated biomass. Interestingly, the FT-IR spectral data showed that SE disrupts the cell wall structure of rice straw in a similar fashion; however, the results showed that intensity of these peaks was independent to the size of a straw. The absorption band at 898 cm 1 was attributed to β-(1,4)-glycosidic linkages (C–O–C stretch­ ing), which was enhanced due to the transformation of the crystalline structure of cellulose to amorphous after SE pretreatment.

Whereas, 95.0, 89.2, 92.4 and 79.9% (for C6 sugars) and 74.9, 72.9, 56.4 and 49.7% (for C5 sugars) for A-10-180, A-20-180, A-10-200 and A-20-200 respectively. The respective pretreated and washed solid res­ idues were further subjected to enzymatic hydrolysis. The final sugar recovery (C6þC5) in both steps was found in order of W-10-200 (60.9%)>W-10-180 (55.9%)>W-20-200 (49.3%)>W-20-180 (30.6%) for water impregnated SE. In contrary, a higher sugar recovery (C6þC5) was observed for DA assisted SE and was found in order of this A-10-180 (81.1%)>A-10-200 (75.6%)>A-20-180 (74.6%)>A-20-200 (64.6%). 4. Conclusions

3.8. Mass balance

Steam explosion of rice straw has been conducted to achieve high glucan conversion by varying the straw size and process parameters. The study reveals that the rice straw with 10 and 20 mm impregnated with DA followed by SE at 180 � C and hydrolyzed by 5 FPU g 1 of cellulase at 15% solid loading resulted in 88.7 and 83.4% of glucose conversion respectively. Whereas, water impregnated SE pretreated straw gave only 61.1 and 27.9% glucan conversion for 10 and 20 mm size within same reaction condition. Lower straw size, i.e., 5 mm generates a high amount of degradation products leading to the formation of pseudo-lignin as compared to 10 and 20 mm. DA assisted SE at 200 � C with 10 mm effectively solubilize hemicel­ lulose leading to increase glucan conversion to 89.6% as compared to water assisted SE (74.8%). Effect of pretreatment temperature was more profound in case of water impregnated SE as compared to DA. Maximum overall sugar recovery (81.1%), combining the pretreatment and enzy­ matic saccharification steps was observed for DA assisted SE at 180 � C with 15% solid loading using 5 FPU g 1 of biomass. Therefore, higher particle size can be dealt by varying the process parameters by consuming less energy and low production cost.

Table 4 illustrated the mass balance and overall sugar recovery after pretreatment and enzymatic hydrolysis of 10 and 20 mm of rice straw. Initially, 3.0 kg of rice straw was taken on dry basis which was reduced to 96.6 to 94.0% after impregnation in water and or dilute acid with removal of extractives and dirt. The variation in impregnation condition and severity due to temperature significantly affected the sugar recovery after pretreatment. Impact of impregnation/soaking of biomass with water/dilute acid prior to pretreatment resulted in swelling of biomass thus increase hemicellulose solubilisation and reduce the formation of xylo-oligomers, thereby offering more exposed cellulose with higher potential towards enzymatic saccharification [20,43]. The soaked biomass was thus subjected to steam explosion at 180 and or 200 � C for 10 min. Total solid recovery after pretreatment was reduced from 68.3 to 56.7%, which may be due to the removal of left-over extractives and solubilisation of hemicellulose. The sugar recovery in pretreated residue and hydrolysate (both monomers and oligomers) were 83.8, 78.3, 83.7 and 80.3% (for C6 sugars) and 91.6, 80.1, 57.3 and 32.0% (for C5 sugars) for W-10-180, W-20-180, W-10-200 and W-20-200 respectively.

Table 4 Mass balance and overall sugar recovery of pretreatment of RS and enzymatic hydrolysis.

Native RS

Sample ID [IM-SS-T]

W-10180

C6 Sugars (3000*Glucan*1.1/ 100) (g) C5 Sugars {3000* (XL þ AR)*1.1/ 100} (g)

1247.4

c d

Solid recovered post soaking (%) Soaked residue (c*3000/100)(g)

e

a b

Soaking/Extraction Pretreatment (P)

Pretreated residue

f g h

a

Liquid hydrolysate

i j k l

Enzymatic hydrolysis (E)

m n o p q r

Overall sugar recovery (P þ E) (%)

W-20180

W-10200

W-20200

A-10180

A-20180

A-10200

A-20200

95.2 2856

96.4 2892

95.2 2856

96.4 2892

94.0 2820

94.2 2826

94.0 2820

94.2 2826

Solid recovered post pretreatment (%) Pretreated residue (d*e/100) (g) C6 Sugars (f* Glucan*1.1/100) (g) C5 Sugars [f* (XL þ AR) *1.1/ 100](g)

67.2

63.3

59.5

56.7

68.3

66.2

65.2

63.1

1919.2 969.0

1829.5 941.8

1699.3 973.9

1639.8 947.0

1926.1 1080.5

1870.0 1020.3

1838.6 1011.3

1782.4 894.1

508.8

489.0

166.4

155.1

156.8

152.2

115.3

117.6

C6 sugars (C þ G þ Glu-olg) (g) C5 sugars (X þ A þ Xy-olg) (g) C6 recovery w.r.t. native (i/ a*100) (%) C5 recovery w.r.t. native (j/ b*100) (%)

76.1 144.0 83.8

37.4 82.0 78.5

69.9 242.4 83.7

54.9 72.7 80.3

88.6 374.9 95.0

92.3 367.1 89.2

141.0 286.5 92.4

102.3 236.6 79.9

91.6

80.1

57.3

32.0

74.9

72.9

56.4

49.7

Glucan Conversion (%) Xylan Conversion (%) C6 sugars (m*g*1.1/100) (g) C5 sugars (n*h*1.1/100) (g) C6 recovery w.r.t. native (o/ a*100) (%) C5 recovery w.r.t. native (p/ b*100) (%)

61.1 40 651.3 224.4 52.2

27.9 35 289.0 190.4 23.2

74.8 44 801.3 80.5 64.2

74.5 37 776.0 63.3 62.2

88.7 41 1054.3 71.2 84.5

83.4 39 936.0 66.6 75.0

89.6 45 996.7 57.2 79.9

89.3 38 878.3 49.3 70.4

31.5

26.7

11.3

8.9

10.0

9.3

8.0

6.9

[(i þ j þ o þ p)*100/(a þ b)]

55.9

30.6

60.9

49.3

81.1

74.6

75.6

64.6

712.8

The starting amount of rice straw is taken 3.0 kg. Where GL, XL, AR, C, G, X, Glu-olg and Xy-olg represents the glucan, xylan, arabinan, cellobiose, glucose, xylose, gluco-oligomer and xylo-oligomer respectively. Monomers and oligomers sugar are calculated in the total volume of hydrolysate obtained after steam explosion. Sugars in solid residue are reported based on total solid recovery after steam explosion. a The oligomers (glu-and xy-oligomers) are not found in pretreatment hydrolysate of acid treated rice straw. 8

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Biomass and Bioenergy 130 (2019) 105390

Declaration of interest

[19] R. Kataria, A. Mol, E. Schulten, A. Happel, S.I. Mussatto, Bench scale steam explosion pretreatment of acid impregnated elephant grass biomass and its impacts on biomass composition, structure and hydrolysis, Ind. Crops Prod. 106 (2017) 48–58. [20] X. Fan, G. Cheng, H. Zhang, M. Li, S. Wang, Q. Yuan, Effects of acid impregnated steam explosion process on xylose recovery and enzymatic conversion of cellulose in corncob, Carbohydr. Polym. 114 (2014) 21–26. [21] Z.-H. Liu, L. Qin, F. Pang, M.-J. Jin, B.-Z. Li, Y. Kang, B.E. Dale, Y.-J. Yuan, Effects of biomass particle size on steam explosion pretreatment performance for improving the enzyme digestibility of corn stover, Ind. Crops Prod. 44 (2013) 176–184. [22] S. Harun, V. Balan, M.S. Takriff, O. Hassan, J. Jahim, B.E. Dale, Performance of AFEX™ pretreated rice straw as source of fermentable sugars: the influence of particle size, Biotechnol. Biofuels 6 (1) (2013) 40. [23] J.Y. Zhu, X.J. Pan, G.S. Wang, R. Gleisner, Sulfite pretreatment (SPORL) for robust enzymatic saccharification of spruce and red pine, Bioresour. Technol. 100 (8) (2009) 2411–2418. [24] A. Sluiter, B. Hames, R. Ruiz, C. Scarlata, J. Sluiter, D. Templeton, D. Crocker, Determination of Structural Carbohydrates and Lignin in Biomass, National Renewable Energy Laboratory; 2010 Jul, Golden, Colorado, 2011, p. 17. Report N. TP-510-42618. [25] A. Sluiter, B. Hames, R. Ruiz, C. Scarlata, J. Sluiter, D. Templeton, Determination of Sugars, Byproducts, and Degradation Products in Liquid Fraction Process Samples, National Renewable Energy Laboratory Analytical Procedure, Golden, CO, 2006. [26] S. J€ ager, A. Brumbauer, E. Feh�er, K. R� eczey, L. Kiss, Production and Characterization of β-glucosidases from Different Aspergillus Strains, Springer, 2001, pp. 455–461. [27] T. Panda, V.S. Bisaria, T.K. Ghose, Effect of culture phasing and a polysaccharide on production of xylanase by mixed culture of trichoderma reesei D1-6 and aspergillus wentii Pt 2804, Biotechnol. Bioeng. 30 (7) (1987) 868–874. [28] Y. Zhu, M. Malten, M. Torry-Smith, J.D. McMillan, J.J. Stickel, Calculating sugar yields in high solids hydrolysis of biomass, Bioresour. Technol. 102 (3) (2011) 2897–2903. [29] M. Kapoor, S. Soam, R. Agrawal, R.P. Gupta, D.K. Tuli, R. Kumar, Pilot scale dilute acid pretreatment of rice straw and fermentable sugar recovery at high solid loadings, Bioresour. Technol. 224 (2017) 688–693. [30] M.P. Tucker, K.H. Kim, M.M. Newman, Q.A. Nguyen, Effects of Temperature and Moisture on Dilute-Acid Steam Explosion Pretreatment of Corn Stover and Cellulase Enzyme Digestibility, Biotechnology for Fuels and Chemicals, Springer, 2003, pp. 165–177. [31] P. Sannigrahi, D.H. Kim, S. Jung, A. Ragauskas, Pseudo-lignin and pretreatment chemistry, Energy Environ. Sci. 4 (4) (2011) 1306–1310. [32] J.C. L� opez-Linares, I. Ballesteros, J. Tour� an, C. Cara, E. Castro, M. Ballesteros, I. Romero, Optimization of uncatalyzed steam explosion pretreatment of rapeseed straw for biofuel production, Bioresour. Technol. 190 (2015) 97–105. [33] J.P.C. Pereira, P.J.T. Verheijen, A.J.J. Straathof, Growth inhibition of S. cerevisiae, B. subtilis, and E. coli by lignocellulosic and fermentation products, Appl. Microbiol. Biotechnol. 100 (21) (2016) 9069–9080. [34] T.-C. Hsu, G.-L. Guo, W.-H. Chen, W.-S. Hwang, Effect of dilute acid pretreatment of rice straw on structural properties and enzymatic hydrolysis, Bioresour. Technol. 101 (13) (2010) 4907–4913. [35] T.I. Georgieva, X. Hou, T. Hilstrøm, B.K. Ahring, Enzymatic hydrolysis and ethanol fermentation of high dry matter wet-exploded wheat straw at low enzyme loading, Appl. Biochem. Biotechnol. 148 (1–3) (2008) 35–44. [36] K. Olofsson, M. Bertilsson, G. Lid�en, A short review on SSF – an interesting process option for ethanol production from lignocellulosic feedstocks, Biotechnol. Biofuels 1 (2008) 7. [37] R. Koppram, E. Tom� as-Pej� o, C. Xiros, L. Olsson, Lignocellulosic ethanol production at high-gravity: challenges and perspectives, Trends Biotechnol. 32 (1) (2014) 46–53. [38] J.B. Kristensen, C. Felby, H. Jørgensen, Yield-determining factors in high-solids enzymatic hydrolysis of lignocellulose, Biotechnol. Biofuels 2 (1) (2009) 11. [39] R. Kumar, C.E. Wyman, An improved method to directly estimate cellulase adsorption on biomass solids, Enzym. Microb. Technol. 42 (5) (2008) 426–433. [40] H. Jørgensen, J. Vibe-Pedersen, J. Larsen, C. Felby, Liquefaction of lignocellulose at high-solids concentrations, Biotechnol. Bioeng. 96 (5) (2007) 862–870. [41] J. Larsen, M. Østergaard Petersen, L. Thirup, H. Wen Li, F. Krogh Iversen, The IBUS process–lignocellulosic bioethanol close to a commercial reality, Chem. Eng. Technol.: Industrial Chemistry-Plant Equipment-Process EngineeringBiotechnology 31 (5) (2008) 765–772. [42] T.C. Hsu, G.L. Guo, W.H. Chen, W.S. Hwang, Effect of dilute acid pretreatment of rice straw on structural properties and enzymatic hydrolysis, Bioresour. Technol. 101 (2010) 4907–4913. [43] W.-H. Chen, B.-L. Pen, C.-T. Yu, W.-S. Hwang, Pretreatment efficiency and structural characterization of rice straw by an integrated process of dilute-acid and steam explosion for bioethanol production, Bioresour. Technol. 102 (3) (2011) 2916–2924.

None. Acknowledgements The authors are thankful to the Department of Biotechnology (DBT) India for funding under grant number BT/PB/08/03/2007. We are also thankful to IOC-DBT Centre for Advanced Bioenergy Research, Indian Oil Corporation Limited (R&D Centre), Faridabad for providing the necessary facilities. Appendix A. Supplementary data Supplementary data to this article can be found online at https://doi. org/10.1016/j.biombioe.2019.105390. References [1] B.E. Outlook, 2019 edition, 2019. London, United Kingdom, https://www.bp.com/ en/global/corporate/energy-economics/energy-outlook.html. [2] T. Raj, M. Kapoor, R. Gaur, J. Christopher, B. Lamba, D.K. Tuli, R. Kumar, Physical and chemical characterization of various Indian agriculture residues for biofuels production, Energy Fuels 29 (5) (2015) 3111–3118. [3] P. Kumar, D.M. Barrett, M.J. Delwiche, P. Stroeve, Methods for pretreatment of lignocellulosic biomass for efficient hydrolysis and biofuel production, Ind. Eng. Chem. Res. 48 (8) (2009) 3713–3729. [4] D.M.d. Carvalho, O. Sevastyanova, J.H.d. Queiroz, J.L. Colodette, Cold alkaline extraction as a pretreatment for bioethanol production from eucalyptus, sugarcane bagasse and sugarcane straw, Energy Convers. Manag. 124 (2016) 315–324. [5] M. Balat, Production of bioethanol from lignocellulosic materials via the biochemical pathway: a review, Energy Convers. Manag. 52 (2) (2011) 858–875. [6] R. Agrawal, A. Satlewal, R. Gaur, A. Mathur, R. Kumar, R.P. Gupta, D.K. Tuli, Pilot scale pretreatment of wheat straw and comparative evaluation of commercial enzyme preparations for biomass saccharification and fermentation, Biochem. Eng. J. 102 (2015) 54–61. [7] S. Semwal, R. Gaur, S. Mukherjee, A. Chopra, R.P. Gupta, R. Kumar, D.K. Tuli, Structural features of dilute acid pretreated Acacia mangium and impact of sodium sulfite supplementation on enzymatic hydrolysis, ACS Sustain. Chem. Eng. 4 (9) (2016) 4635–4644. [8] S. Muthayya, J.D. Sugimoto, S. Montgomery, G.F. Maberly, An overview of global rice production, supply, trade, and consumption, Ann. N. Y. Acad. Sci. 1324 (1) (2014) 7–14. [9] S. Soam, M. Kapoor, R. Kumar, P. Borjesson, R.P. Gupta, D.K. Tuli, Global warming potential and energy analysis of second generation ethanol production from rice straw in India, Appl. Energy 184 (2016) 353–364. [10] Z. Zhang, R. Yang, W. Gao, X. Yao, Investigation of [Emim][OAc] as a mild pretreatment solvent for enhancing the sulfonation efficiency of alkali lignin, RSC Adv. 7 (49) (2017) 31009–31017. [11] C.K. Phwan, H.C. Ong, W.-H. Chen, T.C. Ling, E.P. Ng, P.L. Show, Overview: comparison of pretreatment technologies and fermentation processes of bioethanol from microalgae, Energy Convers. Manag. 173 (2018) 81–94. [12] T. Raj, R. Gaur, P. Dixit, R.P. Gupta, V. Kagdiyal, R. Kumar, D.K. Tuli, Ionic liquid pretreatment of biomass for sugars production: driving factors with a plausible mechanism for higher enzymatic digestibility, Carbohydr. Polym. 149 (2016) 369–381. [13] M. Kapoor, T. Raj, M. Vijayaraj, A. Chopra, R.P. Gupta, D.K. Tuli, R. Kumar, Structural features of dilute acid, steam exploded, and alkali pretreated mustard stalk and their impact on enzymatic hydrolysis, Carbohydr. Polym. 124 (0) (2015) 265–273. [14] A.A. Elgharbawy, M.Z. Alam, M. Moniruzzaman, M. Goto, Ionic liquid pretreatment as emerging approaches for enhanced enzymatic hydrolysis of lignocellulosic biomass, Biochem. Eng. J. 109 (2016) 252–267. [15] I. Ballesteros, M. Ballesteros, P. Manzanares, M.J. Negro, J.M. Oliva, F. S� aez, Dilute sulfuric acid pretreatment of cardoon for ethanol production, Biochem. Eng. J. 42 (1) (2008) 84–91. [16] M.D. Ferro, M.C. Fernandes, A.F.C. Paulino, S.O. Prozil, J. Gravitis, D.V. Evtuguin, A.M.R.B. Xavier, Bioethanol production from steam explosion pretreated and alkali extracted Cistus ladanifer (rockrose), Biochem. Eng. J. 104 (2015) 98–105. [17] R. Gaur, S. Semwal, T. Raj, B. Yadav Lamba, E. Ramu, R.P. Gupta, R. Kumar, S. K. Puri, Intensification of steam explosion and structural intricacies impacting sugar recovery, Bioresour. Technol. 241 (2017) 692–700. [18] P. Alvira, E. Tom� as-Pej� o, M. Ballesteros, M.J. Negro, Pretreatment technologies for an efficient bioethanol production process based on enzymatic hydrolysis: a review, Bioresour. Technol. 101 (13) (2010) 4851–4861.

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