Bioresource Technology 104 (2012) 587–592

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Bioresource Technology

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Effect of steam explosion and microbial fermentation on cellulose and lignindegradation of corn stover

Juan Chang a, Wei Cheng b, Qingqiang Yin a,⇑, Ruiyu Zuo a, Andong Song c, Qiuhong Zheng a,Ping Wang d, Xiao Wang d, Junxi Liu d

a College of Animal Science and Veterinary Medicine, Henan Agricultural University, 95 Wenhua Road, Zhengzhou 450002, Chinab Zhengzhou College of Animal Husbandry Engineering, Zhengzhou 450008, Chinac Key Laboratory of Renewable Energy Ministry of Agriculture, Henan Agricultural University, 95 Wenhua Road, Zhengzhou 450002, Chinad Henan Engineering and Technology Research Center of Feed Microbes, Zhoukou 466000, China

a r t i c l e i n f o a b s t r a c t

Article history:Received 11 September 2011Received in revised form 19 October 2011Accepted 20 October 2011Available online 3 November 2011

Keywords:Corn stoverSteam explosionAspergillus oryzae fermentationCellulose and lignin degradationNutrient metabolic rates

0960-8524/$ - see front matter � 2011 Elsevier Ltd. Adoi:10.1016/j.biortech.2011.10.070

⇑ Corresponding author. Tel.: +86 371 63558180; faE-mail address: QQZ22@yahoo.com.cn (Q.Q. Yin).

In order to increase nutrient values of corn stover, effects of steam explosion (2.5 MPa, 200 s) and Aspergillusoryzae (A. oryzae) fermentation on cellulose and lignin degradation were studied. The results showed thecontents of cellulose, hemicellulose and lignin in the exploded corn stover were 8.47%, 50.45% and36.65% lower than that in the untreated one, respectively (P < 0.05). The contents of cellulose and hemicel-lulose in the exploded and fermented corn stover (EFCS) were decreased by 24.36% and 69.90%, comparedwith the untreated one (P < 0.05); decreased by 17.35% and 38.59%, compared with the exploded one(P < 0.05). The scanning electron microscope observations demonstrated that the combined steam explo-sion and fermentation destructed corn stover. The activities of enzymes in EFCS were increased. The met-abolic experiment showed that about 8% EFCS could be used to replace corn meal in broiler diets, whichmade EFCS become animal feedstuff possible.

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1. Introduction

The crop straw production in the world is about 7 billions tonsannually (Sánchez and Cardona, 2008). The actual application ofstraws for feedstuffs, industrial use and so on is around 30% (Zenget al., 2007), and lots of them have been chopped and plowedunderground in the field as incomplete organic fertilizer, orburned in the open field by human being, which will cause eco-logical and environmental problems. In addition, grain is the mainenergy feedstuffs for animal production. With the rising price andshortage of grain in the world, it is imminent and economical toconvert straws into one kind of feedstuff to replace grain for ani-mal production.

The straws are generally composed of cellulose, hemicelluloseand lignin, and have great potential to be turned to moreedible feedstuffs for animals (Graminha et al., 2008; Eun et al.,2006). However, lignin present in straw is interspersed with hemi-cellulose to form a matrix surrounding the orderly cellulosemicrofibrils to reduce their digestibility (Wan and Li, 2010), sothe degradation of lignin plays an important role in celluloseutilization.

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Biological treatments of straws are safe, environment friendlyand less energy intensive, compared to other pretreatment meth-ods (Liang et al., 2010; Dinis et al., 2009). However, it is limitedby low-rate hydrolysis reaction and long-time incubation, so agreat improvement is needed for its commercial application(Sun and Cheng, 2002). To improve the accessibility of polysac-charides in lignocellulosic complex for enzymatic hydrolysis, pre-treatments such as softening and disruption of lignocellulosestructure are necessary. Steam explosion is one of the most effec-tive pretreatment methods for corn stover because it has themore potential for energy efficiency, lower environmental impactand more soluble carbohydrate production than other pretreat-ment technologies (Alvira et al., 2010). It was reported thatsteam explosion of lignocellulosic biomass could increase thedigestibility of cereal straw for animals (Viola et al., 2008; Liuet al., 1999; De Castro et al., 1995). So far, there have been fewstudies on the combined steam explosion pretreatment and bio-logical treatment for straws. In this study, effects of the com-bined steam explosion and microbial fermentation withAspergillus oryzae (A. oryzae) on corn stover were evaluatedaccording to the changes of compositions, enzyme activities,morphology in the fermented products, and its application inbroiler production as feedstuff. The aims are to improve strawavailability and convert the waste straw to feed materials forsolving grain shortage in animal husbandry.

588 J. Chang et al. / Bioresource Technology 104 (2012) 587–592

2. Methods

2.1. Corn stover preparation

The corn stover was supplied by Key Laboratory of RenewableEnergy of Ministry of Agriculture, Zhengzhou, China. The strawwas ground in a mill, sieved to obtain pieces with the sizes of2–10 mm (accounting for 90%), and stored at room temperature.

2.2. Steam explosion treatment

The steam explosion of corn stover was carried out by steamexplosion equipment (QBS-80), produced in Hebi ZhengdaoMachine Factory, Hebi, China. About 500 g corn stover (dry matter)was put into the steam chamber. The steam was adjusted to2.5 MPa pressure and kept for 200 s, and then suddenly releasedat the end of treatment to give explosion effect. The exploded sam-ples were collected and dried at 65 �C before analysis.

2.3. Microorganism and medium

A. oryzae used in this study was kept in our laboratory, isolatedfrom a cow rumen, and identified by 26S rDNA. It was incubated inthe plates with potato dextrose agar (PDA) at 30 �C for 3 d. Thecompositions of PDA medium were (g/L): soluble amylum 6,glucose 20, yeast extract 2, peptone 5, KH2PO4 2, MgSO4 0.3, agar20. The medium was autoclaved at 121 �C for 20 min. Spore sus-pension was obtained by scraping off the spores from pure culturesand suspending them in sterile 0.9% physiological saline with 0.1%Tween 80. Spore counts were determined to be approximately1 � 108 cfu/mL.

2.4. Solid-state fermentation of corn stover and experimental design

The solid-state fermentation was carried out in the 500 mLErlenmeyer flasks with cotton plug. Twenty grams straws weremixed with 30 mL mineral solution. The compositions of the min-eral solution were as follows (g/L): (NH4)2SO4 1.4, KH2PO4 2.0,MgSO4 0.3, CaCl2 0.3, NaCl 0.5, FeSO4 0.005, MnSO4 0.0016, ZnCl2

0.0017, CoCl2 0.002. The solid-state medium was autoclaved at121 �C for 20 min.

The microbial solid-state fermentation designs were as follows:

Group 1: The untreated corn stover without A. oryzaefermentation.Group 2: The steam exploded corn stover without A. oryzaefermentation.Group 3: The untreated corn stover with A. oryzae fermentation.Group 4: The steam exploded corn stover with A. oryzaefermentation.

The experiments were performed in quadruplicates at 30 �C for7 d, pH was adjusted to 7.0 with Ca(OH)2, and the liquid seed cul-ture was used at 4% (v/w). The fermented products were dried at50 �C to 90% dry matter, and then ground.

2.5. Experimental methods

The neutral detergent fiber (NDF), acid detergent fiber (ADF),cellulose, hemicellulose, and lignin fractions of corn stover weredetermined according to the method of Van Soest et al. (1991).The enzyme activity, soluble carbohydrates such as cellobiose,glucose, xylose, arabinose, xylitol, glycerol, and microbial growthinhibitors such as formic acid, acetic acid, furfural, 5-hydroxymeth-ylfurfural (5-HMF) in corn stover were determined by putting 2 gsample in 40 mL deionizer water, stirred for 2 h at room

temperature. After filtering through filter paper, the supernatantswere kept for the analysis.

The filter paper cellulase activity (FPA) and endoglucanaseactivity was measured as described by Ghose (1987) with What-man No. 1 filter paper and 1% (w/v) carboxymethyl cellulose(CMC) as substrates, respectively. The amylase activity was deter-mined by using soluble starch (Sigma Chemical, USA) as a substrate(Yasser et al., 2009). The units of FPA, CMC and amylase activitieswere defined as the amount of enzyme releasing 1 lmol glucoseequivalent per min under these conditions. The protease activitywas determined by the method of Sandhya et al. (2005), one unitof protease was defined as the amount of enzyme that liberated1 lmol tyrosine per min.

The cellobiose, glucose, xylose, arabinose, xylitol, glycerol, formicacid, acetic acid, furfural and 5-hydroxymethylfurfural (5-HMF)contents were quantified by high-performance liquid chromatogra-phy (Agilent 1200 HPLC, Agilent Technologies, USA) with an AminexHPX-87H column (Bio-Rad, Richmond, CA, USA) and a refractiveindex detector (Jasco International Co., Tokyo, Japan).

Scanning electron microscope (SEM) analysis was performedwith a FEI Quanta 200 (FEI Company, Eindhoven, The Netherlands)at acceleration voltages of 10 kV. Samples were air-dried andcoated (golden/palladium) with a sputter coater (SC7640, QuorumTechnology, Newhaven, UK).

2.6. Determination of nutrient metabolic rates

In order to determine the availability of the exploded and fer-mented corn stover (EFCS) as animal feedstuff, Twenty 42-day-old male Arbor Acres broilers were obtained and randomly dividedinto 4 treatments, 5 birds for each treatment. The basal diet (seeTable 4) was formulated to meet nutrient requirements accordingto NRC (1994). Water was given to birds ad libitum, and feed wasgiven with restriction. The preliminary period was 5 d, and exper-imental period was 3 d. The metabolic experiment was designed asfollows:

(1) Basal diet (the corn–soybean meal base).(2) Basal diet with 4% (w/w) corn meal removed + 4% (w/w)

EFCS.(3) Basal diet with 8% (w/w) corn meal removed + 8% (w/w)

FECS.(4) Basal diet with 12% (w/w) corn meal removed + 12% (w/w)

FECS.

During 3-d experimental period, the total excreta was collectedwithout contamination from each bird according to the collectionmethod (Wang et al., 2008). The 3-d fecal collections were dried,ground and mixed to determine the concentrations of nutrients.Dry matter and organic matter in diets and feces were determinedusing the former methods (Zhang et al., 2011). Crude protein andcrude fat in samples were determined with Kjeldahl and ether ex-tract, respectively (Yin et al., 2010). The calculation was made asfollows: Nutrient metabolic rate = (ingested nutrient � excretednutrient)/ingested nutrient. Gross energy values of the sampleswere measured using an adiabatic oxygen bomb calorimeter(IKA-C2000, IKA Instrument Company, Staufen, Germany).

2.7. Statistical analysis

Experimental data were expressed as the means and standarderrors. The data were analyzed using the ANOVA procedures of Sta-tistical Analysis Systems Institute (SAS 8.0). Duncan’s multiplerange test was used to compare treatment means. Differences wereconsidered statistical significance at P < 0.05.

J. Chang et al. / Bioresource Technology 104 (2012) 587–592 589

3. Results and discussion

3.1. The effect of different treatments on chemical compositions of cornstover

The chemical compositions of corn stover in the different groupswere shown in Table 1. The contents of ADF, NDF, cellulose, hemi-cellulose and lignin in groups 2, 3 and 4 were significantly lowerthan that in group 1 (P < 0.05). The steam explosion pretreatmentfor corn stover reduced the contents of cellulose, hemicellulosesand lignin by 8.47%, 50.45% and 36.65%, respectively (P < 0.05).The huge hemicellulose degradation in the exploded straw willcause the bond between lignin and hemicellulose loose, so that cel-lulose in corn stover will be utilized easily by microbes (Zhang et al.,2008). Moyson and Verachtert (1991) found that straw digestibilitywas improved with increasing lignin degradation. Based on thisexperimental result, steam explosion was an important pretreat-ment method for degrading hemicellulose and lignin contents incorn stover.

The contents of NDF, ADF, cellulose and hemicellulose in thefermented corn stover in groups 3 and 4 were lower than that ingroups 1 and 2 (P < 0.05), due to microbial fermentation. It is prob-able that the microbe and excreting enzymes degrade cellulose andhemicellulose. In addition, there was no difference observed in thecontents of lignin between the fermented and unfermented groups(P > 0.05), it is maybe due to the lack of ligninase from microbe. Forlarge-scale microbial straw production used as animal feedstuffs, itis necessary to achieve high-level lignin degradation with cellulosedegradation in an acceptable extent (Viola et al., 2008). The processof steam explosion pretreatment followed by A. oryzae fermenta-tion could significantly decrease the contents of cellulose, hemicel-lulose and lignin in corn stover.

3.2. The effects of different treatments on soluble carbohydrate andmicrobial growth inhibitor contents in corn stover

The contents of soluble carbohydrates and microbial growthinhibitors in corn stover with different groups were shown inTable 2. It was found that the steam explosion pretreatment for cornstover increased the contents of soluble cellobiose, arabinose, glyc-erol, formic acid and furfural, and decreased glucose and xylitol con-tents, compared with the untreated corn stover. However, theexploded and fermented corn stover could decrease soluble cellobi-ose, xylose, arabinose, glycerol, formic acid, acetic acid and furfuralcontents, while increase glucose and xylitol contents, comparedwith the exploded corn stover. It is the microbial fermentation andCa(OH)2 to cause the above changes. Generally, glucose was pro-duced from either the cellulose or hemicellulose degradation. Thehigher glucose content in the exploded and fermented straw maybe due to microbial ability to convert cellulose or hemicellulose tomonosaccharide. The converting mechanisms will need furtherstudy.

There are several kinds of inhibitors for microbial fermentationsuch as formic acids, furfural, 5-HMF, and phenolic compounds

Table 1Main chemical compositions in the different groups (air dry basis, %).

Groups NDF ADF

1 79.26 ± 1.31A 59.23 ± 2.00A

2 60.94 ± 0.34C 51.12 ± 1.12B

3 69.78 ± 1.27B 53.19 ± 0.60B

4 52.76 ± 2.36D 47.28 ± 3.75C

Note: Group 1, the untreated corn stover without A. oryzae fermentation; group 2, the steastover with A. oryzae fermentation; group 4, the steam exploded corn stover with A. oryzasame column, significant differences at P 6 0.05 levels are indicated by the different letsignificantly different from each other (P > 0.05).

produced during lignocellulosic hydrolysates by explosion(Palmqvist and Hhn-Hagerdal, 2000). Some methods have beentaken to reduce the inhibition including addition of activated char-coal, use of laccase, over-liming, molecular sieves, drying and waterwashing (Olsson and Hahn-Hagerdal, 1996; Jurado et al., 2009; Liand Chen, 2008). This result indicated that steam explosion treat-ment for corn stover increased the contents of formic acid and furfu-ral to make pH value lower. In order to maintain microbial growth,Ca(OH)2 was used to adjust the pH values of media for reducingthe negative effects of the microbial growth inhibitors such as formicacid and furfural.

3.3. The effect of different treatments on morphology of corn stover

The color of the untreated corn stover was brownish yellow.SEM micrograph (�1000 and �5000) analysis indicated that thesurface of the untreated corn stover was smooth and regular. Withsteam explosion, the color turned to dark brown, and the size ofcorn stover tended to be smaller. The surface of the exploded cornstover was uneven and covered with a thin layer of deposits. Thesedeposits of globular shapes were characteristic of lignin deposits asreported before (Kristensen et al., 2008; Selig et al., 2007). It is wellknown that lignin and hemicellulose encase cellulose, hinderingcellulase from reaching cellulose fibrils. However, the lignin layerof the exploded corn stover is easily removed, due to lignin beingless strongly bound to carbohydrate polymers, compared with itsnative linkages. It is a hypothesis that the migration of lignin tothe outer surface exposes more internal cellulose surfaces to in-crease enzyme accessibility.

For the untreated corn stover with microbial fermentation, thecolor of corn stover turned to bright yellow due to microbial fer-mentation. Most of the surface was smooth and intact becausethe straw was only partially defibrated. For the exploded corn sto-ver with microbial fermentation, its color turned to greenishyellow, and the surface was softer and more fragile than that ofthe unfermented one. The SEM observations demonstrated thatthe combined steam explosion with microbial fermentation de-stroyed corn stover structure, and the globular shapes of depositedlignin were not seen. In addition, the microbes on the surface andtheir penetration into the exploded straw were observed. It can beconcluded that steam explosion followed by microbial fermenta-tion was effective in altering the structure of corn stover.

3.4. The optimum periods for microbial fermentation

Table 3 indicated that the contents of NDF, ADF, cellulose andhemicellulose were generally decreased with microbial fermenta-tion time prolonging. The hemicellulose contents of the fermentedstraw were decreased after the third day (P < 0.05). There was no sig-nificant difference in the compositions of ADF, cellulose, hemicellu-lose in straw between the fifth and seventh day fermentation(P > 0.05).

The important consideration in straw solid-state fermentation isto determine the optimum fermentation time of harvesting by

Cellulose Hemicellulose Lignin

41.30 ± 2.31A 20.03 ± 1.18A 12.58 ± 1.07A

37.80 ± 0.40B 9.82 ± 0.86C 7.97 ± 1.21B

33.77 ± 1.23C 16.59 ± 1.02B 14.58 ± 2.18A

29.78 ± 2.92D 5.48 ± 1.03D 9.54 ± 0.97B

m exploded corn stover without A. oryzae fermentation; group 3, the untreated corne fermentation. Each value represents mean ± SE of 4 replicates per treatment. In theters (A, B, C and D). Data followed by the same letter in the same column are not

Table 4Main compositions and nutrient levels of basal diet for broilers.

Ingredients (%) Nutrient levels

Corn meal 65.03 Metabolic energy (MJ/kg) 12.74Soybean meal 26.10 Crude protein (%) 19.00Fish meal 2.00 NDF (%) 15.73Soybean oil 3.00 ADF (%) 5.62Calcium carbonate 1.30 Calcium (%) 0.90Dicalcium phosphate 1.15 Total phosphorus (%) 0.61

D–L Methionine 0.07 Available phosphorus (%) 0.40

Salt 0.35 Lysine (%) 1.02Premix compounda 1.00 Methionine (%) 0.40Total 100.00

a Compositions of premix compounds (per kilogram of diet): Vitamin A (VA) 12000 IU, VD3 3000 IU, VE 20 IU, VK3 1.0 mg, VB1 2.0 mg, VB2 6.0 mg, VB3 12 mg, VB5

3.6 mg, VB9 1.2 mg, VB12 0.01 mg, Biotin 0.15 mg, Cu (copper sulfate) 8 mg, Fe(ferrous sulfate) 100 mg, Mn (manganese sulfate) 80 mg, Zn (zinc oxide) 60 mg, I(calcium iodate) 0.45 mg, Se (sodium selenite) 0.35 mg.

Table 2The soluble carbohydrate and inhibitor contents of corn stover in different treatments(mg/g).

Groups 1 2 3 4

Cellobiose 1.06 2.62 0.80 NDGlucose 5.07 3.30 1.49 6.28Xylose 6.98 6.46 1.23 1.77Arabinose 0.41 2.04 0.32 0.41Xylitol 2.07 1.09 1.16 1.63Glycerol 0.99 2.35 2.02 0.63Formic acid 0.70 7.78 ND 0.53Acetic acid 3.50 3.57 0.17 NDFurfural ND 0.35 ND ND5-HMF ND ND ND ND

Note: Group 1, the untreated corn stover without A. oryzae fermentation; group 2,the steam exploded corn stover without A. oryzae fermentation; group 3, theuntreated corn stover with A. oryzae fermentation; group 4, the steam explodedcorn stover with A. oryzae fermentation.ND: not detected.

590 J. Chang et al. / Bioresource Technology 104 (2012) 587–592

considering acceptable limits and fermentation cost. In the fifth dayof fermentation, the contents of cellulose and hemicellulose in theexploded straws with microbial fermentation were decreased by24.36% and 69.90%, compared with the untreated corn straw(P < 0.05); decreased by 17.35% and 38.59% (P < 0.05), comparedwith the exploded straw. According to corn stover degradation,the optimum harvesting time of the fermented straws was 5 d.

3.5. The enzyme activity of the exploded corn stover during microbialfermentation

The enzyme activities during the different fermentation timewere shown in Fig. 1. The FPA activity on the fifth day fermentation

Table 3Main compositions of the exploded straws during the different microbial fermentation tim

Time (d) NDF ADF

0 60.94 ± 0.34A 51.12 ± 1.12A

1 59.12 ± 1.85A 51.02 ± 1.51A

3 58.59 ± 0.48AB 50.47 ± 0.09AB

5 56.02 ± 0.90B 49.99 ± 1.40AB

7 52.76 ± 2.36C 47.28 ± 3.75B

Note: Each value represents mean ± SE of 4 replicates per treatment. In the same column,and D). Data followed by the same letter in the same column are not significantly differ

0

5

10

15

20

25

30

35

40

0 1 2 3 4

Time (days)

Enz

yme

activ

ity(U

/g)

Fig. 1. Enzyme activity of the exploded s

was relatively lower than that on the sixth day (P < 0.05), and thendecreased significantly on the seventh day (P < 0.05). The CMCactivity was relatively higher on the first, fourth and sixth day fer-mentation than that on the other days (P < 0.05). The CMC activitywas higher than the FPA activity during fermentation, which is cor-respondent with the former reports (Rodrigues et al., 2008; Diniset al., 2009). In addition, the amylase activity was relatively higherfrom the fifth to seventh day fermentation than that on the otherdays (P < 0.05). The protease activity was higher between the thirdand fourth day fermentation than that on the other days. On sixthday fermentation, FPA, CMC, amylase and protease activities in thefermented products reached 5.59, 18.98, 32.57 and 1.11 U/g,respectively. With microbial fermentation, the enzyme activity in

e (air dry basis, %).

Cellulose Hemicellulose Lignin

37.80 ± 0.40A 9.82 ± 0.86A 7.97 ± 1.21A

34.11 ± 0.85B 8.10 ± 0.53B 9.54 ± 0.49A

32.67 ± 0.49BC 8.12 ± 0.50B 9.22 ± 0.06A

31.24 ± 0.92CD 6.03 ± 0.52C 9.74 ± 0.37A

29.78 ± 2.90D 5.48 ± 1.03C 9.54 ± 0.97A

significant differences at P 6 0.05 levels are indicated by the different letters (A, B, Cent from each other (P > 0.05).

5 6 7

FPA

CMCase

Amylase

Protease

traw during microbial fermentation.

Table 5Effect of EFCS on nutrient metabolic rates of broilers (%).

Items Basal diet 4% EFCS 8% EFCS 12% EFCS

Dry matter 76.01 ± 3.45A 73.98 ± 4.34A 73.92 ± 2.48A 70.37 ± 5.96A

Organic matter 79.96 ± 2.75A 78.17 ± 3.82A 77.96 ± 2.09A 74.37 ± 5.29A

Energy 80.93 ± 2.40A 80.61 ± 3.10A 78.46 ± 2.54AB 75.16 ± 4.80B

Crude protein 56.43 ± 3.92A 61.66 ± 3.05A 58.64 ± 2.98A 59.53 ± 1.69A

Crude fat 87.91 ± 1.50A 83.66 ± 5.19A 84.73 ± 1.83A 86.11 ± 4.72A

NDF 46.22 ± 6.97B 63.11 ± 3.20A 62.27 ± 3.32A 60.65 ± 8.81A

ADF 20.82 ± 7.97B 35.99 ± 8.04A 36.12 ± 6.15A 42.67 ± 6.86A

Note: Each value represents mean ± SE of 5 replicates per treatment. In the same row, significant differences at P 6 0.05 levelsare indicated by the different superscript letters (A, B, C and D).

J. Chang et al. / Bioresource Technology 104 (2012) 587–592 591

the exploded corn stover was lower than that of the untreated one,indicating that some products from the explosion actually inhibitenzyme excretion (Chang, 2011).

Solid-state fermentation is a unique process with great poten-tial for recycling some agro-industrial by-products into someuseful materials such as animal feedstuffs, enzymes and so on.Aspergillus species have been found to be able to produce enzymesfor hydrolyzing cellulosic materials, and contribute to the develop-ment of desirable, flavor and aroma products (Gao et al., 2008;Hachmeister and Fung, 1993). In many studies, enzymes as feedadditives played an important role in improving animal productionperformance (Marquardt et al., 1996; Yang et al., 1999; Gado et al.,2009). The fermented corn stover with high activity of multi-enzymes in this study made it more useful for destructing corn sto-ver and compounding animal diets.

3.6. The nutrient metabolic rates of EFCS for broilers

Table 5 showed that the metabolic rates of dry matter, organicmatter, energy, crude protein and crude fat were not affected byreplacing 4–8% corn meal with EFCS in broiler diets (P > 0.05);however, the NDF and ADF metabolic rates were increased by EFCSaddition significantly (P < 0.05). When 12% corn meal in broilerdiet was replaced by EFCS, the energy metabolic rate was de-creased significantly (P < 0.05), compared with the basal diet. Thereduction in energy metabolic rate could be attributed to the highfiber level in the diet with 12% EFCS. The detrimental effect of fiberon animal production performance has been previously reported(Abiola et al., 2002). Because energy is very important for broilergrowth, the optimum amount of EFCS used in broiler diets shouldbe less than 8%. The reasons why EFCS can replace corn meal tokeep the same nutrient metabolic rates are as follows: (1) thesteam explosion and microbial fermentation can degrade the poly-merization of lignin and hemicellulose, and convert cellulose intouseful nutrients such as glucose and other compounds; (2) the en-zymes in the fermented products play an important role in animalperformance and health (Wang and Gu, 2010). There are many re-ports about enzyme positive effect on nutrient utilization (Menget al., 2005; Wu and Ravindran, 2004). In this experiment, the high-er NDF and ADF metabolic rates are probably due to high activitiesof FPA and CMC enzymes in EFCS.

4. Conclusions

Steam explosion is an effective pretreatment method fordestructing corn stover. The cooperation of steam explosion andA. oryzae fermentation for corn stover treatment could further in-crease cellulose, hemicellulose and lignin degradation, destroy itsstructure, and increase its nutritive values. The metabolic experi-ment of broilers showed that EFCS could be used as feedstuff to re-place about 8% grain in bird diets.

Acknowledgements

The project was supported by the National Science Foundationof China (No. 30571346), the authors are grateful to Zhengdaocompany for technical assistances in steam explosion treatmentfor corn stover.

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