In-storage psychrophilic anaerobic digestion: acclimated microbial kinetics

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<ul><li><p>This article was downloaded by: [University of Guelph]On: 14 November 2014, At: 02:24Publisher: Taylor &amp; FrancisInforma Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House,37-41 Mortimer Street, London W1T 3JH, UK</p><p>Environmental TechnologyPublication details, including instructions for authors and subscription information:http://www.tandfonline.com/loi/tent20</p><p>In-storage psychrophilic anaerobic digestion:acclimated microbial kineticsSusan King a , Pierre Courvoisier a , Serge Guiot b &amp; Suzelle Barrington aa Department of Bioresource Engineering , Macdonald Campus of McGill University , 21 111Lakeshore, Ste Anne de Bellevue , QC , H9X 3V9 , Canadab Department of Environmental Bioengineering , Biotechnology Research Institute, NationalResearch Council of Canada , 6100 Royalmount, Montreal , QC , H4P 2R2 , CanadaAccepted author version posted online: 09 Dec 2011.Published online: 24 Jan 2012.</p><p>To cite this article: Susan King , Pierre Courvoisier , Serge Guiot &amp; Suzelle Barrington (2012) In-storagepsychrophilic anaerobic digestion: acclimated microbial kinetics, Environmental Technology, 33:15, 1763-1772, DOI:10.1080/09593330.2011.644867</p><p>To link to this article: http://dx.doi.org/10.1080/09593330.2011.644867</p><p>PLEASE SCROLL DOWN FOR ARTICLE</p><p>Taylor &amp; Francis makes every effort to ensure the accuracy of all the information (the Content) containedin the publications on our platform. However, Taylor &amp; Francis, our agents, and our licensors make norepresentations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of theContent. Any opinions and views expressed in this publication are the opinions and views of the authors, andare not the views of or endorsed by Taylor &amp; Francis. The accuracy of the Content should not be relied upon andshould be independently verified with primary sources of information. Taylor and Francis shall not be liable forany losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoeveror howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use ofthe Content.</p><p>This article may be used for research, teaching, and private study purposes. Any substantial or systematicreproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in anyform to anyone is expressly forbidden. Terms &amp; Conditions of access and use can be found at http://www.tandfonline.com/page/terms-and-conditions</p><p>http://www.tandfonline.com/loi/tent20http://www.tandfonline.com/action/showCitFormats?doi=10.1080/09593330.2011.644867http://dx.doi.org/10.1080/09593330.2011.644867http://www.tandfonline.com/page/terms-and-conditionshttp://www.tandfonline.com/page/terms-and-conditions</p></li><li><p>Environmental TechnologyVol. 33, No. 15, August 2012, 17631772</p><p>In-storage psychrophilic anaerobic digestion: acclimated microbial kinetics</p><p>Susan Kinga, Pierre Courvoisiera, Serge Guiotb and Suzelle Barringtona</p><p>aDepartment of Bioresource Engineering, Macdonald Campus of McGill University, 21 111 Lakeshore, Ste Anne de Bellevue QC H9X3V9 Canada; bDepartment of Environmental Bioengineering, Biotechnology Research Institute, National Research Council of Canada,</p><p>6100 Royalmount, Montreal QC H4P 2R2 Canada</p><p>(Received 20 June 2011; final version received 23 November 2011 )</p><p>In-storage psychrophilic anaerobic digestion develops by microbial acclimation in covered swine-manure storage tanks,producingCH4 and stabilizing organicmatter. To optimize the systems performance, the process kineticsmust be understood.The objective of this study was to evaluate kinetic parameters describing the major stages in the digestion process, and toinvestigate the effect of temperature acclimation on these parameters. Specific activity tests were performed using manureinocula and five substrates at three incubation temperatures. Extant substrate activities were determined analytically foreach case, and intrinsic kinetic parameters for glucose uptake were estimated by grid search fitting to the Monod model.The results demonstrate that this acclimated microbial community exhibits different kinetic parameters to those of themesophilic communities currently modelled in the literature, with increased activity at low temperatures, varying withsubstrate and temperature. For glucose, the higher uptake is accompanied by lower microbial yield and half-saturationconstant. Decomposing these values suggests that active psychrophilic and mesophilic microbial populations co-exist withinthe community. This work also confirms that a new method of assessing microbial substrate kinetics must be developed formanure microbial communities, separating microbial mass from other suspended organics.</p><p>Keywords: psychrophilic anaerobic digestion; specific substrate uptake;manure treatment;microbial kinetics; swinemanure</p><p>IntroductionIn-storage psychrophilic anaerobic digestion (ISPAD) isa combined manure treatment and storage system devel-oped for Canadian pork producers [1]. The process occursin manure storage tanks with air-tight covers when theanaerobic microbial community in the manure has accli-mated to the ambient operating conditions; manure solidsare reduced by 24%, and 63% of the potential methane isreleased [2]. This performance could be optimized usingkinetic modelling, if the appropriate parameters are eval-uated. Current modelling of manure decomposition, usingMonod kinetics and Arrhenius temperature correction fac-tors, requires substrate uptake parameters for eachmicrobialpopulation [3,4].</p><p>The substrate uptake kinetics of anaerobic microbialcommunities may be assessed using specific activity assays[5]. Substrates represent the major stages of anaerobicdigestion: glucose is the model substrate for acidogen-esis; propionate and butyrate are used for acetogenesis;acetate for aceticlastic methanogenesis; and H2/CO2 forhydrogenotrophic methanogenesis and homoacetogenesis[6]. Microbial communities acclimated to psychrophilicconditions exhibit increased substrate uptake at lower tem-peratures, compared with non-acclimated communities,but the increases are not uniform across substrates and</p><p>Corresponding author. Email: Susan.King@mail.mcgill.ca</p><p>temperatures [7]. For example, after cultivation of granularsludge for 306 days at 10 C, activities on acetate,propionate and butyrate increased by factors of 3.65, 1.45and 4.1, respectively at 10 C, and 2.44, 1.20 and 2.61respectively at 30 C [8]. Similarly, after operating at 15 Cfor 625 days, activities on acetate and butyrate increasedat both 15 and 37 C, whereas propionate activity remainedlow, and H2/CO2 activity increased continually throughoutthe experimental period [9]. Propionate activity appears tobe the most sensitive to temperature change and the slow-est to acclimate [10,11], while the highest acclimated VFAactivity is reported for butyrate [12].</p><p>When a mesophilic anaerobic microbial communityacclimates to psychrophilic conditions, such as those occur-ring in ISPAD, for at least a year, the component popula-tions generally exhibit maximum substrate uptake at 35 C,which is taken to mean that they are still mesophilic [3,9].At the same time, the greater increases in activity at lowtemperatures indicate that these communities are psychro-active [13]. Occasionally a fully psychrophilic microbialpopulation with a temperature optimum near 15 C is foundwithin the community [14,15]. However, psychrophilic andmesophilic populations consuming a single substrate mayalso coexist in a single community [16,17], in which casethe substrate uptake data may exhibit a bi-modal form with</p><p>ISSN 0959-3330 print/ISSN 1479-487X online 2012 Taylor &amp; Francishttp://dx.doi.org/10.1080/09593330.2011.644867http://www.tandfonline.com</p><p>Dow</p><p>nloa</p><p>ded </p><p>by [</p><p>Uni</p><p>vers</p><p>ity o</p><p>f G</p><p>uelp</p><p>h] a</p><p>t 02:</p><p>24 1</p><p>4 N</p><p>ovem</p><p>ber </p><p>2014</p></li><li><p>1764 S. King et al.</p><p>mesophilic optimum, composed of two superposed uptakecurves [18].</p><p>Experimental substrate uptake datamay be fittedmathe-matically to amodel equation to estimate kinetic parameterssuch as the Monod half-saturation constant, Ks [19,20].The goodness of fit is used to compare the suitability ofdifferent relationship forms to the process represented bythe data [21]. Using this approach, a study of aceticlas-tic methanogenesis data found that both the Monod andHaldane models were accurate at 30 C, but the Haldanemodel produced a better fit between 6 and 22 C [22]. Asimilar study concluded that the Haldane model was pre-ferred at 22 C, while at 11 C both the Haldane and anon-competitive model fit the data equally well, proposingthat differences could be attributed to the representation ofinhibition in each model [23]. However, a key difficulty inassessing and comparing kinetic parameters from substrateuptake data is the evaluation of the size of the active micro-bial population involved [8]. In addition, the endogenousproduction of substrate must be accounted for when eval-uating the substrate uptake behaviour of manure microbialcommunities [24].</p><p>The relationship between the maximum substrateuptake kinetic parameter, qmax, and temperature is usuallydescribed using the exponential Arrhenius equation [25].Accordingly, in a study using granular sludge adapted to10 C for 235 days, the temperature dependence of acetateconversion was well described by an Arrhenius model;however, propionate, butyrate and mixed volatile fattyacid (VFA) activities were better described by a square-root formulation [26]. Conversely, when digested sewagesludge was adapted to 20 C for four months, the tempera-ture dependence of the resulting methane production fromacetatewas poorly described by both theArrhenius andHal-dane equation forms; no better-fitting model was proposedin this study [23].</p><p>These results illustrate that acclimated anaerobic com-munities may not always be treated as mesophilic, and thatparameters must be evaluated for each intermediate sub-strate. In addition, the assumptions of Monod kinetics andArrhenius temperature dependence may not apply. As such,the substrate uptake kinetics of an ISPAD community can-not be estimated from known data, and must be defined inthe laboratory. Therefore the objective of this study wasto define the kinetic parameters and temperature depen-dence relationships of the main microbial populations inan ISPADmicrobial community. To accomplish this, swinemanure inocula from a three-year-old ISPAD installationwere evaluatedusing specific substrate activity tests.Assayswere performed at three temperatures, 8, 18 and 35 C,using glucose, acetate, propionate, butyrate and H2/CO2 assubstrates. Inocula from a similar uncovered storage tank,and freshly produced manure were evaluated as controls.Extant substrate uptake activity and optimal temperaturewere defined for each population. Intrinsic activity wasestimated by curve fitting to the Monod model.</p><p>Materials and methodsManure inoculaIn 2004, a full-scale ISPAD system was established in St.Francois Xavier Quebec, Canada. The facility used a con-crete tank, 30m in diameter by 3.66mdeep, covered withan air-tightmembrane (GTI, Fredericton, NB,Canada). Thetank received swinemanure on a regular basis. The contentswere removed for land-spreading twice yearly, except for adepth of 0.30.6m.Manure from this facility (Xf ) was usedto represent ISPAD in this study. The two controls, freshlyproduced manure (Xm) and one-year-old manure containedin an uncovered storage tank (Xo), were obtained from theswine research facility of theMcGill UniversityMacdonaldCampusExperimental Centre, located inMontreal, Quebec,Canada. Manures from the two operations were consideredcomparable in terms of solids and nutrients, as they are pro-duced by hogs fed a standard corn and soybean-based ration[27]. Samples of each manure were collected in June 2007as described previously [28].</p><p>Manure characterizationSub-samples of all three manures (Xf , Xo and Xm) wereanalysed according to standard methods [29] to estab-lish: solids, chemical oxygen demand (total and soluble)and pH. The quantity of active microbial biomass ineach manure sample was estimated using the Luminultrawastewater ATP kit (Luminultra, NB, Canada) and a lumi-nometer (Sirius, model V3.2, Bethold Detection Systems,TN, USA).</p><p>Specific substrate activity testsThree sets of substrate activity tests (SAT) were performed[30]: one using the microbial community contained in theISPAD manure as active biomass (Xf ), one using the com-munity in the uncovered tank manure (Xo) and the thirdusing the community in fresh manure (Xm). Each set com-prised three batches, one each at 8, 18 and 35 C. Each batchincluded five individual substrate assays: glucose, acetate,propionate and butyrate were the liquid substrates, andH2/CO2 was the gaseous substrate used. All combinationswere run in triplicate.</p><p>For each batch, twelve 120mL bottles (for liquid sub-strates) and three 60mL bottles (for gaseous substrate) wereprepared. Manure inoculum was added to each bottle, toprovide 5 gVSS L1 for liquids and 2 gVSS L1 for thegaseous substrate. Phosphate buffer was added, bringingthe volume of liquid in each bottle to 20mL. The bot-tles were sealed, flushed with N2/CO2 gas (80%/20%)and placed in a shaker (New Brunswick Scientific, Edi-son, NJ, USA) operating at 100 rpm (400 rpm for gaseoussubstrates), in a thermostatically controlled environment, inthe dark. The bottles were allowed three or four days underthese conditions for acclimation to the assay conditions.</p><p>Dow</p><p>nloa</p><p>ded </p><p>by [</p><p>Uni</p><p>vers</p><p>ity o</p><p>f G</p><p>uelp</p><p>h] a</p><p>t 02:</p><p>24 1</p><p>4 N</p><p>ovem</p><p>ber </p><p>2014</p></li><li><p>Environmental Technology 1765</p><p>Following the acclimation period, liquid substrate wasinjected through the cap of each 120mL bottle, and the60mL bottles were flushed and pressurized to 140 kPa withH2/CO2. The first sample was immediately taken: 0.5mLof liquid (for liquid substrate bottles) or 100l of headspacegas (for H2/CO2 bottles). Liquid samples were centrifugedto remove solids. Sub-samples of supernatant were anal-ysed for glucose in the bottles with this substrate. For thebottles fed acetate, butyrate or propionate, sub-samples ofsupernatant were diluted fivefold for VFA analysis. Forthe H2/CO2 bottles, the headspace gas samples were anal-ysed immediately by gas chromatography. Sampling wasrepeated at regular intervals during each assay period. Atthe end of each assay, the bottle contents were analysed todetermine solids and pH.</p><p>Analytical methodsGlucose was measured using high-pressure liquid chro-matography (Waters Chromatography Division, Milford,MA, USA) equipped with an injector (model 717+), photo-diode array detector (model 2996), pump (model 600) andrefractive indexdetector (model 2414). The columnused forthe separation was an ICSep IC ION-300 column (Transge-nomics, San Jose, CA, USA) of 300mm 7.8mm i.d. andan ion guard GC-801 column (Transgenomics). The mobilephase consisted of 0.035N H2SO4 at a pH of 4, flowing ata rate of 0.4mLmin1. The measurements were conductedusi...</p></li></ul>