Feasibility study of the anaerobic digestion of dewatered pig slurry by means of polyacrylamide

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    fraction, prior to land application, can be achieved by aer-obic composting and/or anaerobic digestion. The secondoption provides a better energy balance and can be comple-

    ment with polyacrylamide (PAM) polymers, prior tomechanical removal or gravity settling, has the potentialto enhance solidliquid separation, thus concentratingnitrogen, phosphorous and organic carbon (Vanotti andHunt, 1999). Since most suspended particles in wastewatersand aqueous solutions, such as livestock and poultry

    * Corresponding author. Fax: +34 935796785.E-mail address: xavier.otats@giroct.irta.es (X. Flotats).

    Available online at www.sciencedirect.com

    Bioresource Technology 991. Introduction

    Modern pig production, which has a very intensive andconcentrated character, generates a large pig slurry surplusthat often cannot be used as an agricultural fertiliser in thesame geographical area, thus making its transport a limit-ing factor. One management strategy consists of separatingthe solid and liquid fractions, then treating the liquid frac-tion prior to using it for irrigation on nearby land, whiletreating the solid fraction in order to stabilise it and toreduce volume before transporting it to areas with nutrientand/or organic matter demand. Stabilisation of the solid

    mented by further aerobic composting in order to producea higher quality end product. The eciency of anaerobicdigestion of this solid fraction can be negatively aectedby high total solids concentration (Itodo and Awulu,1999; Bujoczek et al., 2000).

    The main fraction of organic matter found in pig slurrytakes the form of small suspended particles, mainly in col-loidal form, which are not easily separated by applying asimple mechanical system (Hill and Tollner, 1980). The e-ciency of suspended solids separation using lters andpresses is limited, and for colloids agglutination a chemicalcoagulation process is required (Sievers et al., 1994). Treat-Abstract

    Liquid livestock waste can be managed by separating liquid and solid fractions then treating each separately by applying best avail-able technology, such as anaerobic digestion for the solid fraction. There is an increasing use of polyacrylamide (PAM) as a occulantagent to improve solidliquid separation. In the present work, the anaerobic toxicity of PAM residues and the optimal range of totalsolids concentration for maximum methane production were studied as a function of PAM dosage. Results showed that dry matterand its volatile solids content increased signicantly with increasing PAM dosage. Batch anaerobic tests showed that methane yielddecreased linearly with increasing total solids, while the methane production per unit of raw substrate reached a maximum at 16.4% totalsolids. No PAM toxicity was measured for PAM concentrations below 415 g/kg total solids, but some indirect inhibitory phenomenawere observed, such as a limited hydrolysis rate due to particle aggregation, and inhibition of methanogenesis by high ammoniaconcentration. 2007 Elsevier Ltd. All rights reserved.

    Keywords: Anaerobic digestion; Pig slurry; PAM; Polyacrylamide; Solidliquid phase separationFeasibility study of the anaerobicby means of p

    E. Campos a, M. Almirall a, J. Mtna Laboratory of Environmental Engineering, Centre

    b SELCO MC SL, Pza. Tetuc GIRO Technological Centre, Rambla Pompeu F

    Received 15 January 2004; received in revised foAvailable onlin0960-8524/$ - see front matter 2007 Elsevier Ltd. All rights reserved.doi:10.1016/j.biortech.2006.12.008igestion of dewatered pig slurrylyacrylamide

    Almela b, J. Palatsi c, X. Flotats c,*

    L-IRTA, Rovira Roure 191, E-25198 Lleida, Spain

    6, E-12001 Castellon, Spain

    1, E-08100 Mollet del Valle`s, Barcelona, Spain

    20 September 2006; accepted 6 December 2006February 2007

    (2008) 387395

  • Temanure, have a negative charge at pH values higher than 4,the addition of cationic coagulants to these wastewaterswould be more eective than the addition of anionic ones(Sievers et al., 1994).

    Polyacrylamide (PAM) is widely used in sewage sludgetreatment to enhance dewatering. The polyelectrolyte con-centration in mechanically dewatered cakes is relativelyhigh and typically in the range 2.55.0 g/kg dry matter(TS), and it can be degraded by abiotic processes in culti-vated soils without toxicity problems (ICON, 2001). There-fore, it has been suggested that it presents low toxicity, withLD50 value greater than 5 g/kg TS, but this potential toxic-ity has not been studied yet for anaerobic digestion.

    PAM can degrade to acrylamide monomer, which ishighly toxic (IPCS, 1985), followed by a rapid degradationto ammonia and acrylic acid, which is not toxic and in turndegrades to CO2 and water (ICON, 2001). El-Mamouniet al. (2002) demonstrated that PAM is highly recalcitrantto aerobic or anaerobic microbial degradation, suggestingthat this recalcitrance is linked to the high molecularweight, thus making it inaccessible to microbial attack.These authors found that PAM is very susceptible to UVphotolysis, enhancing a further microbial degradation pro-cess, but without intermediate production of acrylamide.Studies on PAM degradation in cultivated soils (Kay-Shoe-make et al., 1998) demonstrated that indigenous bacteriacould use PAM as a source of nitrogen, biotransformingthe polymer to long chain polyacrylate, which may be fur-ther degraded by biological processes without toxicityproblems. In an extended review, Cauleld et al. (2002)concluded that there is no evidence to suggest that PAMcan undergo biodegradation to form free acrylamidemonomer units. Cauleld et al. (2002) also concluded thatPAM can act as a carbon source for microbial growth onlywhen some other physical or chemical process lowers themolecular weight of the polymer beforehand. These resultssuggest that toxicity by acrylamide is not probable duringanaerobic digestion processes without previous physicalor chemical pre-treatment to enhance PAM degradation.

    Chu et al. (2003) studied the anaerobic digestion ofPAM occulated activated sludge, comparing the eect ofcationic, non-ionic and anionic polyacrylamide. Theyfound that anionic and non-ionic PAM had no eect onmethane yield at doses below 15 g/kg TS. For cationicPAM, methane yield decreased with increasing dosage,showing a small variation in relation to the control assayfor 1 and 5 g/kg TS, and a signicant decrease for 15 and40 g/kg TS. Since dosed polymers had no apparent toxicityto the inoculum, although an anaerobic toxicity test wasnot performed, Chu et al. (2003) suggested that the muchgreater oc size obtained with cationic PAM increasedthe mass transfer resistance. No methane production fromPAM degradation was measured in these experiments.Chang et al. (2001) found that the methane productionobtained from anaerobic batch digestion of a commercial

    388 E. Campos et al. / BioresourcePAM, consisting of a copolymer of acrylamide and acry-loyloxyethyltrimethylammonium chloride, was consistentwith a complete degradation of the second monomer, butnot with the degradation of PAM or acrylamide. No inhi-bition phenomena were reported in this study.

    The optimum cationic PAM dose varies with the type ofmanure and the amount of total suspended solids (TSS) inthe liquid manure, the dose increasing with TSS concentra-tion (Vanotti and Hunt, 1999; Chastain et al., 2001). In astudy of the separation of dierent types of pig slurry,Walker and Kelley (2003) found that optimal PAM dosagewas in the range 0.91.8 g/kg TS for ecient separation ofSS, TSS and COD, and in the range 4.210.9 g/kg TS forecient separation of nitrogen (N) and phosphorous (P).In general, the TSS removal eciencies achieved rangefrom 76% to 99%. These values contrast with the ecien-cies of the screening process alone (without using PAM),which range from 5% to 14%.

    The objectives of the present work were to study theanaerobic digestion of the solid fraction of pig slurry sepa-rated using PAM at dierent dosages, to characterize thissolid fraction and to study the anaerobic biodegradabilityand toxicity of PAM, in order to determine whether thepolymer or its possible degradation products can aectanaerobic microorganisms during the pig slurry digestionprocess.

    2. Methods

    2.1. Materials

    The pig slurry and its associated solid fraction camefrom a treatment plant in Modena, Italy. This plant usedthe SELCO-EcopurinTM solid/liquid separation system(Martnez-Almela and Barrera, 2005), using cationicPAM as the coagulant agent. The four dierent materialsidentied in Table 1 were used: raw pig slurry (PS), solidfraction of pig slurry using a PAM dose of 120 mg/l(SFPS), which is the usual dose in the plant, solid fractionof pig slurry using a PAM dose of 140 mg/l (SFPS-1) andsolid fraction of pig slurry without using PAM (SFPS-0).Anaerobically digested sewage sludge from a mesophilicdigester was used as inoculum for batch tests.

    2.2. Analytical methods

    Analytical methods for the determination of total andvolatile solids (TS and VS), total and volatile suspendedsolids (TSS and VSS), total and soluble chemical oxygendemand (CODt and CODs), total Kjeldahl nitrogen(NTK), ammonia nitrogen NH4 N and pH were adaptedfrom Standard Methods for the Examination of Water andWastewater (APHA, 1995). Total and partial alkalinity(TA, PA) were analysed according to the method proposedby Hill and Jenkins (1989).

    Methane and carbon dioxide concentration in the biogaswere measured with a GC 8000 Top Series gas chromato-

    chnology 99 (2008) 387395graph (CE Instruments, Italy), tted with PORAPAK-N(80/100 mesh) packed column (2 m 2 mm) and a Thermal

  • ferent PAM concentrations between 0 and 126 mg/l and




    e Teport (J70), column and TCD were 130, 30 and 120 Crespectively.

    Volatile fatty acids (VFA) acetate (Ac), propionate(Pro), iso-butyrate (Iso-But), n-butyrate (n-But), iso-valer-ate (Iso-Val) and n-valerate (n-Val) were determined fromsamples after centrifugation (2790g for 20 min), ltration(0.45 lm) and acidication/extraction (with HCl anddiethyl-ether 1/1) with a Trace 2000 gas chromatograph(Thermo Instruments, Italy), tted with a FFAP capillarycolumn (30 m 0.250 mm 0.25 lm), with ame ioniza-tion detector (FID) and equipped with auto sampler (Auto-sampler AS2000, Italy). The FID was supplied with H2 andsynthetic air, while He was used as make-up gas with a owrate of 30 ml/min. Samples of 1 ll were injected in split-splitless mode, with a constant carrier gas ow rate of1 ml/min, a split ratio of 20/1 and a septum purge ratioof 5/1. The initial oven temperature was 90 C for 4 min,after which it was increased to 155 C at 6 C/min thento 255 C at 12.5 C/min, with a nal isotherm of 2 min.The injector and detector temperatures were set constantat 240 C.

    NH3N concentration was calculated by using Eq. (1),

    NH3N NH4 Nt1

    10pKpH 1

    ; 1

    with a pK value of 8.938 at 35 C (Bonmat and Flotats,2003).

    2.3. Polyacrylamide toxicity and biodegradability testConductivity Detector (TCD). Helium (He) was used as acarrier gas (20 ml/min), and temperatures of the injector

    Table 1Identication and basic characterisation, average of three replicates, of ino

    PAMa dose (mg/l) TS (g/kg) VS (g

    mg/l g/kg TS

    SFPS-1 140 14.27 313.60 41.41 233.58SFPS 120 12.23 136.12 6.06 100.36SFPS-0 0 0 13.57 0.24 7.38PS 9.81 0.36 5.53Inoculum 39.45 0.16 22.65

    a PAM dose applied to raw pig slurry to obtain the corresponding solid

    E. Campos et al. / BioresourcThe PAM toxicity test was carried out according to Sotoet al. (1993). The culture medium consisted of 223 g/l ofdigested sewage sludge as inoculum, macro and micro-nutrient solutions, a mixture of volatile fatty acids (2.95 gacetate/l, 0.59 g propionate/l and 0.25 g butyrate/l) as sub-strate (Soto et al., 1993) and the corresponding PAM con-centration (from 0 control to 2775 mg/kg sludge TS).Batch reactors were 120 ml glass vials lled with 50 ml ofculture medium. After displacement of air from the head-space with N2/CO2 gas (80/20 v/v) for 3 min, the vials weretightly closed with rubber stoppers. Finally, a reducingsolution (0.1 ml of 50 g Na2S/l) was injected into every vialto achieve a reduced medium. The vials were incubated atone additional treatment with a much higher concentra-tion, about 2775 mg/l. These concentrations correspondto 018.7 g PAM/kg TS of sludge, and the sixth concen-trated treatment corresponds to 415 g/kg TS, which is anunusual and extremely high value.

    An anaerobic biodegradability test, following Soto et al.(1993), was also carried out in order to study the biode-gradability of PAM in an anaerobic environment. The120-ml vials were lled with 50 ml of medium, containingmacronutrient and micronutrient solutions, alkalinity solu-tion and anaerobically digested sewage sludge (222 g/l),giving a solids concentration of 6.7 g TS/l and 5.1 g VS/l.The initial concentration of PAM was 259.8 4.4 mg/l inthe culture liquid, corresponding to 38.5 0.6 g/kg TS.pH was adjusted to neutrality. After displacement of airfrom the headspace with N2/CO2 gas (80/20 v/v), the vialswere tightly closed with rubber stoppers. Finally, a reduc-ing solution (0.1 ml of 50 g Na2S/l) was injected into everyvial to achieve a reduced medium. The vials were incubatedat 35 C in a closed and dark incubator for 33 days. Theaccumulated methane production was determined by peri-odic headspace analysis.

    2.4. Batch anaerobic tests

    Four substrate mixtures (Table 1) were prepared andmixed in various ratios to produce ve dierent combina-35 C in a closed and dark incubator for 21 days and theamount of gas accumulated in the headspace was measuredtwice a week.

    Six treatments were carried out (Table 2), with ve dif-

    um and substrates used (g/kg of substrate or inoculum)

    COD (g/kg) NTK (g/kg) NNH4 (g/kg)

    29.98 271.99 36.17 18.81 0.78 3.06 0.003.79 96.30 24.37 7.79 3.45 1.89 0.430.51 17.07 6.15 2.03 0.20 1.51 0.070.70 6.17 3.78 0.90 0.58 0.83 0.000.93 28.22 4.24

    ction used as substrate.

    chnology 99 (2008) 387395 389tions of TS and PAM concentrations, corresponding to vedierent treatments for batch anaerobic tests (Table 3). Anadditional blank treatment water plus inoculum wasalso prepared to evaluate the methane production frominoculum. The methodology of batch tests was adaptedfrom Campos et al. (2000): 120 ml glass vials were lledwith 30 g of mixture (90% substrate and 10% inoculum,digested sewage sludge). After displacement of air fromthe headspace with N2/CO2 gas (80/20 v/v) for 3 min thevials were tightly closed with rubber stoppers. Finally, areducing solution (0.1 ml of 50 g Na2S/l) was injected intoevery vial to achieve a reduced medium. The vials wereincubated at 35 C in a closed and dark incubator, and

  • A0.





    Temonitored for 82 days. Vials were shaken by hand once aday. A complete analytical characterisation was performedat both the beginning and the end of the experiment: TSand VS, TSS and VSS, CODt and CODs, NTK, NH

    4 N,

    pH, TA, PA and VFA. The accumulated methane produc-

    Table 2Results from the PAM toxicity test

    Treatment PAM dose (mg/l) PAM dose (g/kg TS)

    T1 0.00 0.00T2 33.29 4.96T3 66.89 10.07T4 99.27 14.99T5 125.55 18.67T6 2775.19 414.80

    Letters: results of Duncan test at 5% signicance; dierent letters indicatea ACm, maximum methanogenic activity, observed at seventh day.

    Table 3Characterisation of treatments used in the anaerobic digestion test (three r

    Treatment PAM dose Substrate composition (% w/w)

    g/kg TS mg/l SFPS-1 SFPS SF

    T1 14.27 140 100 0T2 13.25 130 50 50T3 12.23 120 0 100T4 10.40 100 0 85 1T5 0 0 0 0 10T6 (blank) 0 0 0

    390 E. Campos et al. / Bioresourcetion was determined by periodic headspace analysis.

    2.5. Calculations

    2.5.1. Separation eciency

    The separation eciency (Et) is dened as the total massrecovery of nutrients in the solid fraction as a proportion(%) of the total input of solids or nutrients (Mller et al.,2002),

    Et U M cQ Sc 100; 2

    where U (kg) is the quantity of solid fraction, Mc (g/kg) isthe concentration of TS or NTK in the solid fraction; Q (kg)is the amount of manure treated; and Sc (g/kg) is the con-centration of TS or NTK in the manure.

    2.5.2. Methanogenic activity

    The methanogenic activity, ACm (g COD/g VS day), inthe toxicity test was calculated as methane production dur-ing the maximum growth period, by using the followingexpression adapted from Soto et al. (1993):

    ACm Rf V SSV 3

    where R is the methane production rate (ml CH4/day), f isa factor to transform methane volume to grams of COD(350 ml of CH4/g COD for Normal Conditions), and[SSV] is the concentration of SSV in the culture mediumwith a volume V.

    2.6. Statistical methods

    Cm, 7 daya (g COD/g VS day) Final methane yield (ml CH4/vial)

    118 0.025 A 76.30 12.53 a114 0.025 A 75.24 9.06 a116 0.018 A 76.94 3.51 a117 0.007 A 75.59 6.42 a111 0.012 A 78.42 5.87 a107 0.026 A 86.96 6.55 b

    istically signicant dierences.

    ications per treatment)

    Substrate characterisation (g/kg)

    -0 TS VS COD

    313.60 41.41 233.58 29.98 271.99 36.17210.04 17.34 155.85 15.47 200.65 4.97136.12 6.06 100.36 3.79 96.30 24.3798.74 3.03 71.41 1.83 85.00 7.3413.56 0.24 7.52 0.51 14.84 6.15

    chnology 99 (2008) 387395The condence limits of average values of experimentaldata were calculated using Eq. (4),

    x ts= np ; 4where x is the average value, s is the standard deviation, n isthe number of replicates and t is the corresponding t-statis-tical distribution value, depending on the number of sam-ples and on the degree of condence (95% in the presentstudy).

    Mean separation tests were performed using statisticalanalysis software (SAS Institute, 1989) and by applying aDuncan test with a signicance level of 5%. Signicant dif-ferences have been indicated with dierent letters. Regres-sion analyses were done using the LevembergMarquardtalgorithm.

    3. Results and discussion

    3.1. Eect of polyacrylamide on substrate characteristics

    The basic characterisation of the dierent original mate-rials shown in Table 1 was carried out prior to the anaer-obic tests. The solid fraction of pig slurry obtained from theusual dose of PAM (120 mg/kg) SFPS showed a TSconcentration higher than 13% (136.1 g TS/kg). A slightincrease in the PAM dose, from 120 to 140 mg/kg (SFPS-

  • 1), led to a very signicant increase in the total solids con-centration of the solid fraction, which surpassed 30% ofthe total weight (313.6 g TS/kg). The solid fraction of pigslurry obtained when PAM was not used (SFPS-0) showeda very low TS and NTK content and it was slightly higherthan raw pig slurry (PS). This fact shows that the separationprocess was not eective without PAM dosage, which isconsistent with results from Vanotti and Hunt (1999).

    The separation eciency for 120 and 140 mg PAM/kgraw slurry was higher than the usual values reported forother mechanical separation methods (Mller et al., 2000,2002), as shown in Table 4. The dierences were especiallyimportant for the separation eciency of total nitrogen,with values obtained exceeding 50%. Similar or higherremoval eciencies have been obtained by other authorsusing PAM as an additive in the separation process (Vano-tti and Hunt, 1999; Walker and Kelley, 2003). As can beobserved in Table 1, the organic fraction (VS) of totalsolids, the COD/NTK ratio and the NTK=NNH

    4 ratio

    increased with PAM dose, indicating an increasing separa-tion eciency for organic materials.

    Evolution of accumulated methane production (ml) pervial is shown in Fig. 1. Treatments T1T5 showed a verysimilar evolution, without statistically signicant dier-ences between treatments. However, treatment T6 (corre-sponding to 415 g PAM/kg TS) showed higher andstatistically signicant accumulated methane productionat the end of the experiment. The dierence in methaneproduction for this treatment was 112.8 38.0 ml CH4/gPAM added, an average 23% of the maximum theoreticalmethane yield predicted based on molecular composition.Chang et al. (2001) found that low methane productionfrom PAM was due to the degradation of acryloyloxyeth-yltrimethylammonium chloride, a second monomer con-tained in the PAM used for their work. However, PAMused in the work described here did not contain this mono-mer. Therefore, ndings from Chang et al. (2001) cannotexplain the measured methane production. Taking intoaccount the high dose used in treatment T6, this low butsignicant production could have been caused by the pres-ence of impurities and additives.

    Maximum methanogenic activity for the six treatmentswas found on the seventh day of digestion, and showed




    E. Campos et al. / Bioresource Technology 99 (2008) 387395 3913.2. Polyacrylamide toxicity and biodegradability study

    The maximum concentration of PAM that could befound in the solid fraction of pig slurry, assuming that allPAM was associated with the separated solids, was14.27 g PAM/kg TS for 140 mg PAM/l dose (Table 1).With treatments T2T5 (Table 2) the toxicity study coveredusual PAM concentrations. The study was contrasted bothwith an extremely high PAM concentration (T6) and with acontrol assay without PAM dosage (T1). Table 2 shows theresults from the toxicity test.

    Table 4Separation eciencies obtained compared with literature values

    Technology Substrate Reference U/Q (%)

    PAM 120 mg/l PS Present study 6.09PAM 140 mg/l PS Present study 2.59Centrifuge PS Mller et al. (2002) 13.10Centrifuge PS Mller et al. (2002) 8.28Centrifuge PS Mller et al. (2002) 5.67Centrifuge PS Mller et al. (2002) 4.69Centrifuge ADPS Mller et al. (2002) 13.72Centrifuge ADPS Mller et al. (2002) 14.11Centrifuge ADPS Mller et al. (2002) 8.82Centrifuge ADPS Mller et al. (2002) 9.91Screw press PS Mller et al. (2002) 5.23Screw press ADPS Mller et al. (2002) 3.85Screw press ADPS Mller et al. (2002) 2.88Tilted plane screen PS Mller et al. (2000) 30.00Pressing screw PS Mller et al. (2000) 5.00Pressing screw PS Mller et al. (2000) 7.30Two-stage separator PS Mller et al. (2000) 24.00Belt press separator PS Mller et al. (2000) 17.50PS, pig slurry; SFPS, separated solid fraction of pig slurry; ADPS, anaerobicaa Concentration ratio Mc/Sc (see Eq. (2)).no statistically signicant dierences when applying theDuncan test (Table 2 and Fig. 2). These results indicatethat the polymer compound used cannot be consideredtoxic for anaerobic microorganisms at the concentrationsstudied. If PAM is degraded in some way, products are alsonon-inhibitors even at the high concentration used in T6.

    No statistically signicant dierences were found in thebiodegradability test (Fig. 3) between the treatment withPAM as substrate and the control treatment. This fact indi-cates that the polymer is not signicantly biodegradable byanaerobic microorganisms. The low and non-statistically

    SFPS Et Mc/Sca

    NTK TS NTK TS (%) NTK (%) TS NTK

    .81 0.90 136.12 7.79 84.6 53.0 13.9 8.7

    .81 0.90 313.60 18.81 82.9 54.4 32.0 21.0

    .20 4.20 245.60 9.4 60.5 29.3 4.6 2.2

    .90 4.40 279.30 9.88 48.3 18.6 5.8 2.2

    .10 2.20 187.30 7.79 62.1 20.1 11.0 3.5

    .50 3.90 178.20 10.91 32.8 13.1 7.0 2.8

    .20 4.20 280.80 7.41 68.6 24.2 5.0 1.8

    .30 5.00 252.70 10.99 54.6 31.0 3.9 2.2

    .50 3.80 299.90 10.89 74.5 25.3 8.4 2.9

    .40 3.30 201.90 7.89 53.5 23.7 5.4 2.4

    .20 4.20 364.70 6.61 35.9 8.2 6.9 1.6

    .20 4.20 268.40 6.31 18.4 5.8 4.8 1.5

    .40 3.30 298.40 6.89 23.0 6.0 8.0 2.1

    .60 4.10 117.00 4.6 62.0 33.7 2.1 1.1

    .60 4.10 317.00 4.8 28.0 5.9 5.6 1.2

    .60 4.10 219.00 4 28.2 7.1 3.9 1.0

    .60 4.10 167.00 5.3 70.8 31.0 3.0 1.3

    .60 4.10 192.00 6.4 59.4 27.3 3.4 1.6lly digested pig slurry.

  • Te80


    T1-0 g/kgTS

    Accumulated CH4 (mL)

    392 E. Campos et al. / Bioresourcesignicant dierence in methane production has the sameorder of magnitude as that obtained for the T6 treatmentin the toxicity test, low enough to be disregarded at usualPAM dosages.

    The NTK and NH4N measurements in the biodegrad-ability and toxicity experiments showed an ammonia





    0 2 4 6 8 10 12 14 16 18 20 22 24Time (days)

    T2-5 g/kgTST3-10 g/kgTST4-15 g/kgTST5-19 g/kgTST6- 415 g/kgTS

    Fig. 1. Accumulated methane production in toxicity test. Condenceintervals calculated for 95% condence level.










    0 2 4 6 8 10 12 14 16 18 20PAM concentration (g/kg TS)

    Methanogenic activity 7d(g COD/g VSd)

    Fig. 2. Maximum methanogenic activity index, found at day 7, as afunction of PAM dose expressed as g of PAM/kg of total dried solids.









    0 5 10 15 20 25 30 35Days of incubation

    PAM-250 mg/lControl

    Accumulated CH4 (ml)

    Fig. 3. Accumulated methane generation in the anaerobic biodegradabil-ity test.release ranging from 17% to 69% of PAM organic N (datanot shown), with high deviation within and between treat-ments and without a dened tendency. Since no anaerobicbiodegradability was measured and no polymer chainbreak took place, this ammonia release suggests that deam-ination of PAM occurred to some extent, as described byKay-Shoemake et al. (1998) and Cauleld et al. (2002).

    It can be concluded from the toxicity and biodegradabil-ity tests that the polymer or its degradation products didnot produce toxicity to anaerobic digestion, suggesting thatacrylamide was not produced, in agreement with Kay-Shoemake et al. (1998); Cauleld et al. (2002) and El-Mamouni et al. (2002).

    3.3. Study of the initial total solid concentration eect toanaerobic batch tests

    Methane production results from the batch anaerobictests are shown in Table 5 and Figs. 46. Methane produc-tion related to substrate weight (M), Table 5 and Fig. 4,increased with TS concentration or PAM dose until treat-ment T3, decreasing with higher PAM doses with a mini-mum methane production value for treatment T1 (14.27 gPAM/kg TS). Treatments T2 and T3 showed no statisti-cally signicant dierences, but treatment T2 showed anincreasing methane production during the last days of theincubation period (Fig. 4), suggesting that methane valueswere not the maximum obtainable in spite of the extendedincubation period.

    Dierent behaviour was detected when methane yield(B) was measured. Yield is expressed as ml CH4 producedrelated to initial added VS or related to initial added COD(Table 5). When related to initial VS concentration, accu-mulated methane showed statistically signicant dierencesfor the ve treatments of the test, with methane yieldincreasing with decreasing TS concentration or PAM doseand a maximum value for the control treatment (T5).When related to initial COD concentrations, the globalresponse was very similar but without statistically signi-cant dierences between treatments T3 and T4. TreatmentT2 showed a clear increasing yield value at the end of theexperiment (Fig. 5), suggesting that the nal value couldhave been higher if incubation time were longer. Althoughwith a lower slope, treatment T1 also showed a slightincrease at the end of the experiment (Figs. 4 and 5), sug-gesting that the digestion process rate was decreased signif-icantly by the increase in TS or PAM dose, but notstopped.

    Studying the relationship between methane yield (B) andinitial TS (Fig. 6), a clear linear decrease associated withthe increase in TS content of the substrate can be observed.Other authors have pointed out this tendency; Bujoczeket al. (2000) found that for total solids concentrationsabove 4%, maximum methane production rate decreasedwith total solids content, following a linear tendency; Itodo

    chnology 99 (2008) 387395and Awulu (1999) observed that methane yield from dier-ent types of animal waste tended to decrease when total

  • 4 yield (B) (ml CH4/g VSinitial) CH4 yield (B) (ml CH4/g CODinitial)

    .46 4.83 A 14.14 4.15 A

    .99 10.33 B 101.74 8.02 B

    .58 7.24 C 215.29 7.55 C

    .81 8.87 D 214.89 7.45 C

    .00 18.92 E 272.56 9.59 D

    e Technology 99 (2008) 387395 393Table 5Accumulated production of methane in the anaerobic batch test

    PAM dose (g/kg TS) CH4 (M) (ml/g sub) CH

    T1 14.27 3.85 1.13 A 16T2 13.25 20.41 1.61 C 130T3 12.23 20.73 0.73 C 206T4 10.40 18.27 0.63 B 255T5 0 4.04 0.14 A 538

    E. Campos et al. / Bioresourcsolids content increased. In the case of pig slurry, thisdecrease only took place for TS values above 10%.

    Since PAM toxicity was not demonstrated in the presentstudy, the explanation for the lower methane production inthe most concentrated treatments could be: (a) inhibitionof enzymatic hydrolysis due to the colloidal aggregation,decreasing eective particle surface and increasing internalmass transfer resistance due to the increase in oc size, assuggested by Chu et al. (2003), or (b) specic inhibitionof another process step.

    Dierent letters indicate statistically signicant dierences among means by columns, with a signicance level of 5%.







    0 10 20 30 40 50 60 70 80 90Days of incubation

    T1-14.3 g/kgTST2-13.3 g/kgTST3-12.2 g/kgTST4-10.4 g/kgTST5-0 g/kgTS

    M (ml CH4/g subs)

    Fig. 4. Accumulated methane production (M) per gram of substrateobtained in the batch anaerobic test, for dierent PAM doses.








    T1-14.3 g/kg TST2-13.3 g/kg TST3-12.2 g/kg TST4-10.4 g/kg TST5-0 g/kg TS

    B (ml CH4/g CODini)

    0 10 20 30 40 50 60 70 80 90Days of incubation

    Fig. 5. Accumulated methane yield (B) per gram of initial COD in thebatch anaerobic test for dierent PAM doses.

    y = -787.23TS + 257.67TS + 0.5842R2 = 0.9989





    M B B (ml CH4/g CODini)M (ml CH4/g subs)The rate of hydrolysis can be measured by the removal

    y = -897TS + 302.23R2 = 0.9599



    0 5 10 15 20 25 30 35 TS (%)0





    0 10.40 12.23 13.25 14.27g PAM/kg TS

    Fig. 6. Methane production (M) and yield (B) as functions of the totalsolids of the substrate, obtained with the indicated PAM dose.of particulate matter during the batch anaerobic diges-tion, expressed as the reduction of organic nitrogenNTKNNH4 , of TSS, of VSS or of particulate COD(CODp = CODt CODs). The corresponding removalsare shown in Fig. 6. It should be noted that the generaltrend was a decreasing removal rate of particulate matteras TS or PAM dose was increased (see Fig. 7).

    Mass transfer limitation due to high solids concentra-tion could produce a local high accumulation of VFA. Itsaccumulation in the biowaste bed over inhibitory levels(4050 g COD VFA/l) can inhibit the hydrolysis process,











    0 5 10 15 20 25 30 35% TS

    % re



    of p



    e m




    Fig. 7. Average reduction of particulate matter, for dierent initial totalsolids concentration values, expressed as organic N, TSS, VSS orparticulate COD.

  • as has been reported previously (Veeken and Hamelers,2000). The volatile fatty acid concentration at the end ofthe experiment for T1 (highest solid concentration treat-ment) was very high, close to 40 g COD VFA/kg, a levelat which hydrolysis is completely inhibited by VFA (Veekenand Hamelers, 2000). Validated models describing VFAinhibition of hydrolysis of particulate matter have beendeveloped with satisfactory results (Angelidaki et al.,1999; Vavilin and Angelidaki, 2005).

    The high concentration of acetate at the end of the pro-cess in T1 (Table 7) together with the ammonia concentra-tion (Table 6), above 6 g NNH4 =l, could also explain aninhibition of acetoclastic methanogenic microorganismsby free ammonia. Other VFA were also accumulated, butat lower levels than acetate. Accumulation of acetate couldalso have caused an accumulation of other longer chainacids, since a high concentration of acetate can inhibitthe acetogenic process (Ahring and Westermann, 1988).

    In most of the treatments, ammonia nitrogen concentra-tion increased towards the end of the process (Table 6). Intreatments T1T4, ammonia nitrogen concentration practi-cally doubled and reached extremely high levels, above 6 gN/kg. This increase in ammonia nitrogen and of pHthroughout the process caused a signicant increase in freeammonia at the end of the process (Table 6). The measuredvalues were higher than the values described as inhibitoryfor a methanogenic population by some authors (Hashim-

    oto, 1986; Gallert et al., 1998), although below the inhibi-tion threshold value for adapted acetate-utilizing bacteria(Angelidaki and Ahring, 1993; Hansen et al., 1998).

    The high concentration of propionate and the high valueof the propionate/acetate (P/A) ratio (Table 7) at the endof treatment T2 (corresponding to 130 mg/kg PAM dose)showed that the process had been strongly inhibited. Thisalso explains the shape of the accumulated methane curve,with a longer lag phase than in the most diluted treatments(Figs. 3 and 4), probably indicating an overloading of themethanogenic population due to the high ratio of organicmatter/inoculum.

    The other treatments (T3T6) showed much lower levelsof VFA at the end of the experiment (Table 7), and in manycases only acetate was detectable, showing that themethanogenic phase was not inhibited, as can also bededuced from the shape of accumulated methane produc-tion curves.

    Summing up, the decrease in solids removal rate andmethane yield of the solid fraction of pig slurry separatedby PAM as PAM dose increased, with consequentincreased TS concentration, can be explained by a combi-nation of two phenomena: resistance to enzymatic hydroly-sis due to organic matter aggregation and transportlimitations, and inhibition of the methanogenic step dueto high NH4N concentration, which increased in the solidfraction separated as PAM dose increased.





    394 E. Campos et al. / Bioresource Technology 99 (2008) 387395Table 6Nitrogen measurements at the beginning and at the end of the anaerobic b

    PAM dose (mg/kg TS) NTK (g N/kg) NH4 N g N=k


    T1 14.27 15.68 0.67 3.07 0.28T2 13.25 11.73 0.59 2.19 0.12T3 12.23 7.87 0.66 2.46 0.09T4 10.40 6.45 0.25 1.78 0.10T5 0 2.01 0.09 1.37 0.08T6 0.35 0.06 0.25 0.21

    Average of three replicates.

    Table 7Volatile fatty acid concentrations at the beginning and at the end of batch

    PAM (mg/kg TS) Individual volatile fatty acid concentration (m

    Ac Pro Iso-But n-But

    InitialT1 14.27 75.48 5.84 6.99 3.44T2 13.25 59.33 10.51 6.92 5.90T3 12.23 50.23 22.90 2.63 6.27T4 10.40 48.00 19.64 2.30 5.60T5 0 42.34 9.56 1.38 4.42T6 0.52 0.33 0.08 0.01

    FinalT1 14.27 206.23 54.71 27.00 39.53T2 13.25 8.14 77.59 12.55 0.16T3 12.23 1.44 0.10 0.06 0.00T4 10.40 1.03 0.00 0.01 0.00

    T5 0 0.13 0.00 0.00 0.00T6 0.13 0.00 0.00 0.00h tests (mixture of substrate and inoculum)

    pH NH3 (mg N/kg)

    Final Initial Final Initial Final

    6.47 1.10 7.42 7.60 90.52 286.234.96 0.12 7.29 7.87 48.10 387.893.57 0.27 7.99 7.96 248.91 337.703.29 0.27 7.26 7.98 36.23 328.971.62 0.08 7.58 8.19 57.68 246.310.20 0.05 7.00 7.65 2.86 9.93

    periments (average of three replicates)

    ) Total VFA (mM) Total VFA (g COD/kg)

    Iso-Val n-Val

    10.96 0.35 103.05 13.73 9.50 1.1410.19 0.84 93.69 35.72 9.32 3.253.99 1.57 87.60 2.60 8.35 0.373.51 1.37 80.43 4.74 7.55 0.431.88 1.04 60.61 4.55 5.31 0.480.10 0.02 1.06 1.09 0.07 0.07

    39.49 2.11 369.08 31.34 38.61 4.8718.98 0.19 117.61 13.26 15.21 1.210.08 0.00 1.68 1.03 0.13 0.080.33 0.00 1.37 0.98 0.14 0.24

    0.00 0.00 0.13 0.21 0.01 0.010.00 0.00 0.13 0.12 0.01 0.01

  • e Te4. Conclusions

    The solidliquid phase separation process applied to pigslurry is very sensitive to the dose of PAM used as coagu-lant agent. An increase from 12 to 14 g PAM/kg TS iscapable of almost tripling the total solids content of thesolid fraction, reaching a total solids concentration as highas 31% w/w.

    The use of a PAM concentration higher than 12 g/kg TSis not recommended for further anaerobic treatment, sincesymptoms of inhibition of the hydrolysis step, probablydue to the strong colloidal aggregation, and of inhibitionof the methanogenic step by free ammonia nitrogen, wereobserved in the most concentrated treatments.

    Anaerobic toxicity by PAM, or by its degradation prod-ucts, was not observed for concentrations lower than 415 gPAM/kg TS.


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    Feasibility study of the anaerobic digestion of dewatered pig slurry by means of polyacrylamideIntroductionMethodsMaterialsAnalytical methodsPolyacrylamide toxicity and biodegradability testBatch anaerobic testsCalculationsSeparation efficiencyMethanogenic activity

    Statistical methods

    Results and discussionEffect of polyacrylamide on substrate characteristicsPolyacrylamide toxicity and biodegradability studyStudy of the initial total solid concentration effect to anaerobic batch tests



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