Feasibility of biogas production from anaerobic co-digestion of herbal-extraction residues with swine manure
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k wacteso94 molidCH4
respectively. These results suggested that biogas production could be enhanced efciently by the anaer-
is little information in the literature about biogas digested fromHER or co-digested from the mixture of HER with other biomasses.
Anaerobic co-digestion has been dened as the treatment of amixture of at least two different biomasses with the aim of improv-ing process efciencies (lvarez et al., 2010). Nowadays, there is anincreasing interest in using anaerobic co-digestion process treating
digestion of agro-residues with swine manures would also aid inovercoming ammonia inhibition related to the digestion of pureswine manure. However, it is unclear whether some agro-residues,e.g., HER, might have adverse effects when added into a stabledigester for swine manure.
The objective of the present work was to investigate the feasi-bility of developing anaerobic co-digestion of HER with swinemanure for efcient biogas production. Batch experiments werecarried out based on a Central Composite Design. The inuenceof the percentage of HER in the substrate and the substrate/inocula
Corresponding authors.E-mail addresses: firstname.lastname@example.org (J.-H. Zhu), email@example.com (W.-B.
Bioresource Technology 102 (2011) 64586463
Contents lists availab
elsZhou).extraction residues (HER) is a specic kind of agricultural wastedue to the major importance of the Chinese herbal medicine indus-try in China (Wang et al., 2010). With the rapid development of theChinese herbal medicine industry, about 1.5 million tons of solidwastes, according to statistics, were produced annually after theextraction of medical active components from natural plants(Cheng and Liu, 2010). Because of its abundance in cellulose, hemi-celluloses, lignin as well as proteins, HER could be employed as arenewable energy source. Its environmentally friendly exploitationbecomes an important topic due to the increase in environmentalpollution and the shortage of energy resource. In addition, there
by co-digestion with other agro-residues (Chae et al., 2008;Lansing et al., 2010; Riao et al., 2011). Moreover, the anaerobicdigestion of agro-residues is often associated with poor bufferingcapacity during anaerobic treatment and its inherent high C/N ra-tio. Anaerobic co-digestion of themwith swine manure could over-come these problems by maintaining a stable pH within themethanogenesis range due to the inherent high buffering capacityof swine manure (Banks and Humphreys, 1998). Additionally,swine manure could present high ammonia content and a widevariety of nutrients needed by the methanogens during the anaer-obic process (Cheng, 2009). From another point of view, the co-Biogas
Biomass is an abundant and rencould be derived from all organic mand natural activities, including induwastes, forestry wastes as well aset al., 2003; Wang and Keshwani,0960-8524/$ - see front matter 2011 Elsevier Ltd. Adoi:10.1016/j.biortech.2011.03.093obic co-digestion of herbal-extraction residues with swine manure. 2011 Elsevier Ltd. All rights reserved.
e energy source whichls produced by humanwastes, municipal solidltural wastes (BerndesAmong them, herbal-
industrial and agricultural organic wastes and swine manure forbiogas production (Kaparaju and Rintala, 2005; Riao et al.,2011; Wu et al., 2010). The advantages of anaerobic co-digestionprocess often lie in balancing the carbon/nitrogen (C/N) ratio inthe mixture, macro and micronutrients, pH, inhibitors/toxic com-pounds as well as dry matter (Hartmann et al., 2003). Thus, themanure specic methane production potential could be improvedKeywords:Herbal-extraction residuesSwine manure
pended solids. Furthermore, specic methane yields were 162, 180 and 220 mL CH4 g1 volatile solidsadded d1 for the reactors co-digesting mixtures with 10%, 25% and 50% herbal-extraction residues,Feasibility of biogas production from anaof herbal-extraction residues with swine
Yan Li a,b, Xi-Luan Yan a, Jie-Ping Fan a, Jian-Hang ZhaCentre for Low-Carbon Biotechnology, School of Environmental and Chemical Engineerb State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang
a r t i c l e i n f o
Article history:Received 16 February 2011Received in revised form 27 March 2011Accepted 29 March 2011Available online 2 April 2011
a b s t r a c t
The objective of this worco-digestion of herbal-extrwere carried out under mspecic biogas yield was 2idues and 3.50 g volatile smanure alone was 207 mL
journal homepage: www.ll rights reserved.obic co-digestionanure
,, Wen-Bin Zhou a,Nanchang University, Nanchang 330031, China047, China
as to examine the feasibility of biogas production from the anaerobicion residues with swine manure. Batch and semi-continuous experimentsphilic anaerobic conditions. Batch experiments revealed that the highestL CH4 g1 volatile solids added, obtained at 50% of herbal-extraction res-s g1 mixed liquor suspended solids. Specic methane yield from swineg1 volatile solid added d1 at 3.50 g volatile solids g1 mixed liquor sus-
le at ScienceDirect
evier .com/locate /bior tech
ratio were evaluated in terms of methane yield. Finally, the effectof the feed component ratio of HER to swine manure on processperformance was investigated in a semi-continuously fed stirredtank reactor (CSTR) under mesophilic anaerobic conditions.
2.1. Origin of HER, swine manure and inocula
The HER investigated in this study was obtained from Jiangz-hong herbal medicine group (Jiangxi, China). Swine manure wasobtained from a pig farm (Jiangxi Evergreen Biotechnology Co.,Ltd.) located in Fengcheng (Jiangxi, China). The anaerobic sludgeused as inocula was collected from the anaerobic biogas digesterin Jiangxi Evergreen Biotechnology Co., Ltd., the mixed liquid sus-pended solids (MLSS) and pH value of which were 21 1.3 g L1
and 7.19 0.6, respectively. The substrates and inocula were indi-vidually homogenized and subsequently stored at 4 C for furtheruse. The characterization of each waste employed was shown inTable 1. All samples were collected in triplicate, and the averageddata of the measurements were presented.
2.2. Batch experiments
Batch experiments were carried out at 35 2 C for 30 daysbased on a Central Composite Design, which is a second order fac-torial design used when the number of runs for a full factorial de-sign is too large to be practical (Box andWilson, 1951). This type offactorial design usually consists of a 2k factorial nucleus, ve repli-
terms of grams of volatile solid (VS) (g VS)/g of MLSS (g MLSS) ra-tio, and the percentage of HER in the substrate (% HER), measuredin terms of VS of HER in relation to the organic carbon of the feed.The selected range for factor one (SC) was 1.15.9 g VS g1 MLSS.The selected range for factor two (% HER) was 0100%. Factorial de-sign levels were codied from 1 to +1. The central point was rep-licated ve times in order to estimate experimental error. Axialpoints ensure design rotatability and their distance to the centralpoint (a) was calculated according to Eq. (1).
a 2k=4 1The selected response for analysis was the methane yield
(ml CH4 ml g1ml VS added), calculated from the methane contentin biogas and the volume of biogas produced from per unit of VSadded. The variables, Xi, were coded as xi according to Eq. (2), suchthat X0 corresponded to the central value:
xi Xi Xi =DXi; where i 1;2;3; . . . ; k; 2where xi is the dimensionless coded value of an independent vari-able, Xi is the actual value of an independent variable for the ith test,Xi is the actual value of an independent variable at the centre pointand DXi is the step change (Chong et al., 2009; Riao et al., 2011).All the evaluated levels were combined in 13 different treatments(T1T13). Codied and real values for both factors are presentedin Table 2.
For predicting the optimal point, a second order polynomialfunction (linear regression) was performed Eq. (3):
Y b0 b1X1 b2X2 b11X21 b22X22 b12X1X2; 3
81.2VS (%) 32.2 1.6 23.5
T7 1 1 1.80
Y. Li et al. / Bioresource Technology 102 (2011) 64586463 6459T8 0 0 3.50T9 0 0 3.50T10 1 1 1.80T11 0 0 3.50T12 1.4142 0 1.10T13 0 0 3.50TKN (g kg ) 4.50 0.17C/N ratio* 36.6pH value 5.69 0.14Moisture content (%) 58.30 1.20
* C/N ratio was dened as the ratio of TOC to TKN.
Table 2Codied, real values and responses for swine manure co-digestion with HER in batch
Treatmentsa Codied values Real valu
SC (g VS g1 MLSS) %HER SC (g VS
T1 0 1.4142 3.50T2 0 0 3.50T3 1 1 5.20T4 1 1 5.20T5 0 1.4142 3.50T6 1.4142 0 5.90cates of the central point and 2k axial points, where k is the num-ber of factors evaluated. More specically, in the present study thetwo factors selected were the solid concentration (SC), measured in
Table 1Composition of the substrates in batch experiments and semi-continuous digestion: h
Parameters Batch experiments
TOC (g kg1) 164.8 5.2a Data are means of three replicates, except T2, T8, T9, T11 and T13 were performedb The unit is ml CH4 g1 VS added.where Y represents the predicted response, b0, b1, b2, b11, b22, andb12, are the regression coefcients. X1 and X2 are the evaluated fac-tors (SC and %HER). The coefcient of determination (R2) was calcu-
l-extraction residue (HER) and swine manure.
9 3.3 156.8 7.2 71.24 4.2 1.2 31.9 1.4 20.7 + 1.7 0.22 4.65 0.23 4.86 0.19
33.7 14.7 0.21 5.90 0.25 7.59 0.191 1.90 59.20 2.10 72.90 1.70
Real responseb Predicted responseb
0.00 207 9 19850.00 294 29414.64 216 5 22985.36 254 11 260
100.00 243 10 24450.00 279 9 28385.36 275 6 27050.00 301 29450.00 297 29414.46 235 12 23750.00 283 29450.00 282 10 27050.00 298 294once, respectively.
lated to achieve the proportion of data variability that is explainedby the model, thus the quality of t to the model. The p-values ofthe parameter estimation were used to validate the model and p-values less than 0.05 indicate the signicant model terms. Multipleregression analysis for the data sets collected was performed usingDesign-Expert 7.1.6 Trial software.
The anaerobic assays were conducted in 29 bottles (500 mL)with an inocula volume of 150 mL and a total liquid volume of
bottles were closed with a septum and the headspace ushed with
which was monitored daily. The results from the analysis of each
(1992). Samples from the beginning and the end of the experimentwere analyzed. All measurements were conducted in triplicate andthe averaged data were presented.
The volume of biogas produced was measured using water dis-placement method. The fraction of CH4 was periodically analyzedby a gas chromatograph (SP6890) provided by Lunan RuihongCompany (Shandong, China) equipped with a thermal conductivitydetector (Ruihong, Shandong, China) and a 2 m stainless steel col-
6460 Y. Li et al. / Bioresource Technology 102 (2011) 64586463mixture at steady state were used for evaluating the effect of co-digestion on biogas production efciency.
Volatile solid (VS), total organic carbon (TOC), total Kjeldahlnitrogen (TKN), and pH (Sartorius basic pH meter PB-10, Germany)were performed in accordance with APHA Standard Methods
Table 3Characteristics of mixed feedstocks at varied different mixture ratio used in CSTR exp
Parameters HER (%)
Total organic carbon (g L1) 30.20 2.10VS (%) 8.9 0.4Total nitrogen (g L1) 2.10 0.14pure N2 to remove the oxygen. The volume of biogas produced wasmeasured using water displacement method.
2.3. Semi-continuous digestion of different mixtures
Semi-continuous co-digestion of HER with swine manure wascarried out in two identical continuously stirred tank reactors(CSTRs) with a total volume of 10 L and a working volume of 7 L,namely R1 and R2. A water bath was used to maintain the temper-ature of the digesters at 35 2 C. Digesters were mounted sepa-rately on a mechanical stirrer, stirring continuously at 100 rpm.The outlets provided on the top of each digester were used forfeeding inuent, withdrawing efuent and for collecting biogas.Biogas was daily measured by displacement of water.
Feeding VS was maintained constant during the whole experi-ment resulting in a VS loading rate of about 2.90 g VS L1 d1 underhydraulic retention time (HRT) of 30 days. Varied feeding VS ratiosof HER and swine manure were evaluated. After inoculating the di-gester with 7 L of digested anaerobic sludge, R1 was used to co-di-gest HER with swine manure in a feeding VS ratio of 75% swinemanure and 25% HER, whereas R2 was performed with swine man-ure alone. After 30 days, R1 was fed with swine manure and HER,in a feeding VS ratio of 50% swine manure and 50% HER, whereasthe feed in R2 was made up 90% swine manure and 10% HER.The digesters were fed once a day every weekday. Prior to eachfeeding, a volume equal to the feeding volume was removed tomaintain a constant digester volume. The characteristics of sub-strates are shown in Table 3.
The semi-continuous digestion experiments on R1 and R2 wereperformed two batches, respectively. And the composition of inu-ent and efuent were determined twice a week except the pH375 mL, respectively. All treatments were performed in three rep-licates, except Treatment 2 (T2), T8, T9, T11 and T13 was per-formed in one bottle, respectively. Two blanks containing 150 mLof inocula and 225 mL of distilled water were also run to determinethe endogenous biogas production of the anaerobic sludge. TheC/N ratio 14.38 1pH value 7.48 0.18 7umn (Ruihong, Shandong, China) packed with 5 molecular sieves(its pore size is about 5 ). High purity nitrogen gas (99.99%) wasused as the carrier gas with a ow rate of 30 mL min1. The tem-peratures of the injection port, oven and detector were 120, 50and 150 C, respectively. Gas analyses were carried out two tothree times a week during the rst 2 weeks of the experiments. A250 ll pressure-locked gas tight syringe (SP-6800A) (Ruihong,Shandong, China) was used for the gas sampling. All measurementswere conducted in triplicate and the averaged data werepresented.
3. Results and discussion
3.1. Chemical characteristics of swine manure and HER
There were signicant differences in the composition of the twobiomass wastes (Table 1). HER had higher C/N ratios in the range of33.736.6, as compared to swine manure, which presented C/N ra-tios in the range of 14.414.7. HER, as shown for pH values, wassignicantly acidic, whereas swine manure had a pH value ofaround 7.6. The characterization of the two wastes indicated thatco-digestion of HER with swine manure could be a good solutionto balance C/N ratio, overcoming the problems of digesting bothsubstrates separately.
3.2. Batch experiments
The experimental design data, real responses and predicted re-sponses were presented in Table 2. Regression analyses wereshown in Table 4 and resulted in the following second order poly-nomial Eq. (4):
YCH4 294:20 4:47 SC 16:11 %HER 8:79 SC2
36:54 %HER2 0:50 SC %HER 4The regression showed that the model was signicant because
the Model F-value of 23.30 was greater than the calculated one(0.0003). The determined R2 coefcient obtained was 0.9433,meaning that the model explained 97.15% of the variability data.Although the Pred R-squared of 0.7061 is in reasonable agree-ment with the Adj R-squared of 0.9028, the Lack of Fit F-valueof 2.51 implies the Lack of Fit is not signicant relative to the pureerror. There is a 19.79% chance that a Lack of Fit F-value this largecould occur due to noise (Table 5).
Moreover, the 0.7061 correlation coefcient indicated that thecombination of both factors of SC and% HER had high importancein the yield of methane. p-values for the entire model terms were
ent. All parameters were measured in triplicate.
0 25 50
1.91 2.22 33.89 1.81 37.33 2.15.7 0.5 8.8 0.4 8.6 0.6.93 0.09 2.84 0.13 1.56 0.11
6.54 18.33 23.9.59 0.21 7.42 0.13 7.30 0.24
Table 4Regression of a full quadratic model for methane production.
Factor Coefcient estimate Degree of freedom
Intercept 294.20 1AA 4.47 1BB 16.11 1AB 0.50 1A2 8.79 1B2 36.54 1
Standard deviation = 10.00; R-squared = 0.9433; Adj R-squared = 0.9028; C.V.% = 3.75; Pr
Table 5Analysis of variance for the regression model of methane production.
Source Sum of squares Degrees of freedom M
Model 11639.45 5 23AA 159.82 1 1
Y. Li et al. / Bioresource Technololower than 0.0003, except for the quadratic term associated withSC (Table 5), showing %HER and (%HER)2 are more signicant mod-el terms, as compared to SC and SC2. As an overall, the second orderpolynomial model tted the experimental results corresponding tomethane yield quite well. Here, the methane yield could be calcu-lated from the produced biogas volume and methane content inthe biogas.
The averaged methane yield of all treatments (29 bottles) was261.2 mL CH4 g1 VS added. Methane content was above 58% (datanot shown) for all treatments. The highest specic methane yieldwas 294 mL CH4 g1 VS added (the mean of ve treatments), ob-tained at the value of 3.50 g VS g1 MLSS and 50% for factors SC
BB 2077.28 1AB 1.00 1A2 537.18 1B2 9286.88 1Residual 699.32 7Lack of Fit 456.52 3Pure error 242.80 4Cor total 12338.77and% HER, respectively. The specic methane yield for HER andswine manure alone were 243 (T5) and 207 (T1) mL CH4 g1 VSadded, respectively, both of which were lower than those obtainedfrom the mixture when HER content was 50%, showing the advan-tages of anaerobic co-digestion of mixed biomass wastes for biogasproduction.
Fig. 1 illustrated the accumulated methane production through-out the co-digestion time for T1, T3, T4, T5, T6 and Tm (the
Fig. 1. Accumulated methane production for HER and swine manure by anaerobicco-digestion in batch tests. All data were measured in triplicate.accumulated methane production of Tm was the means of T2, T8,T9, T11 and T13), all of which SC were equal to or above3.5 g VS g1 MLSS. The maximal methane production of 6.19 Lcould be obtained on day 12 for T1 with the 100% of swine manureas the digestion substrate, while the maximal methane productionof 7.25 L was delayed to be observed on day 22 for T5 with 100% ofHER as the digestion substrate. The delayed maximal methane pro-duction might be resulted from the higher C/N ratio of the sub-strate as well as the lower biodegradability of HER (Cheng andLiu, 2010), as compared to swine manure. As also shown inFig. 1, two-stage methane production could be observed when var-ied percentages of HER were co-digested with swine manure. At
Standard error 95% CI low 95% CI high VIF
4.47 283.63 304.773.53 12.83 3.89 1.003.53 7.76 24.47 1.005.00 12.32 11.32 1.003.79 17.75 0.17 1.023.79 45.50 27.58 1.02
ed R-squared = 0.7061; Adeq precision = 14.118.
ean value F prob > F p-Value Signicance
27.89 23.30 0.0003 Signicant59.82 1.60 0.246477.28 20.79 0.00261.00 0.01 0.9231
37.18 5.38 0.053586.88 92.96
Chong, M.L., Rahman, N.A., Rahim, R.A., Aziz, S.A., Shirai, Y., Hassan, M.A., 2009.
noloand these values increased up to 1002 mL CH4 d1 and220 mL CH4 g1 VS added d1 with 50% HER addition. The methanecontent increased from 57.3% with SM alone to 63.8% with a 50%HER addition. When the %HER was within the range of 050%,the reactor fed with higher concentrations of HER showed higherbiogas production rates, higher compositions in methane as wellas high specic methane yield. The specic methane yield in-creased 7.3%, 19.2% and 45.7% as compared to that obtained fromthe digestion of swine manure alone when 10%, 25% and 50% ofHER was added, respectively. These results were in accordancewith those found in literature (Amon et al., 2006; Liu et al., 2009;Cavinato et al., 2010; Gelegenis et al., 2007) that indicated thatanaerobic co-digestion could increase CH4 production of manuredigesters, depending on the operating conditions and the co-sub-strates used.
The higher methane yields achieved in co-digestion of HER withSM, as compared with those achieved with SM alone at the sameloading rate in the present study, were apparently due to the high-er methane potential of HER, as demonstrated by batch experi-ments. This high methane potential achieved by the co-digestionof HER was probably due to the high anaerobic biodegradabilityof the cellulose and sugars, the main components of HER. On theother hand, the main components in typical pig manures are carbo-hydrates, hemi-cellulose and cellulose followed by proteins, fatsand lipids and a small amount of lignin (4.4%) and starch (1.6%)(Iannotti et al., 1979). The biodegradability of pig manure has beenreported to be dependent upon the lignin content, which is notonly considered as refractory to anaerobic degradation, but also re-duces the availability of other components, especially cellulose(Gelegenis et al., 2007; Gonzlez-Fernndez et al., 2008; Liuet al., 2009).
Finally, NH4 -N concentrations in the present study were far toreach reported toxics levels of >4 g L1 which would cause ammo-nia inhibition (De Baere et al., 1984; Cheng, 2009). The high con-tent of ammonia in swine manure makes it possible to degradeHER biologically without the addition of external alkalinity andwithout addition of external nitrogen source (Table 1). On theother hand, as reported by previous study, signicant increasesin volumetric biogas production can be achieved by adding car-bon rich agricultural residues to the co-digestion process withswine manure (Wu et al., 2010). These authors found that the
Table 6Performance data of CSTR treating the mixture of HER with swine manure.
Biogas production rate (mL d1) 762 14Methane content (%) 57.3 0.9Specic methane yield (mL g1 VS added d1) 151 7TOC reduction (%) 73.2 2.2
6462 Y. Li et al. / Bioresource TechC:N ratio of 20:1 was the best in terms of biogas productivityin the anaerobic co-digestion of swine manure with three cropresidues as an external carbon source. HER addition to swinemanure widened TOC:TKN ratio from 14.38 to 23.9 in 0% HERand 50% HER, respectively (Table 6). As mentioned, in this work,the swine buffer capacity contributed to the stability of theprocess.
Co-digestion in the present study should be considered as a pro-cess for the simultaneous treatment of two different biomasswastes and as a solution for the problems of ammonia inhibitiongenerally encountered during anaerobic digestion of pig manure.However, the process stability and efciency have not been inves-tigated. Our following works should be focused on the processOptimization of biohydrogen production by Clostridium butyricum EB6 frompalm oil mill efuent using response surface methodology. Int. J. HydrogenEnergy 34, 74757482.optimization, especially in improving process stability via the elu-cidation and regulation of the microbial consortia.
Co-digestion of HER with swine manure is very promising forbiogas production. The specic methane yield increased 7.3%,19.2% and 45.7%, as compared to that obtained from swine manurealone when 10%, 25% and 50% HER was added, respectively. More-over, the addition of HER to the anaerobic digestion of swine man-ure increased methane content in the biogas. The results of thepresent laboratory study revealed that the use of HER as co-sub-strate in the anaerobic digestion of swine manure has the advan-tage of balancing the C/N ratio.
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Feasibility of biogas production from anaerobic co-digestion of herbal-extraction residues with swine manureIntroductionMethodsOrigin of HER, swine manure and inoculaBatch experimentsSemi-continuous digestion of different mixturesAnalyses
Results and discussionChemical characteristics of swine manure and HERBatch experimentsSemi-continuous single-stage digestion of different mixtures