Anaerobic digestion of simulated distillery waste using flocculated and fixed cell reactors

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  • This article was downloaded by: [McGill University Library]On: 17 December 2014, At: 10:18Publisher: Taylor & FrancisInforma Ltd Registered in England and Wales Registered Number:1072954 Registered office: Mortimer House, 37-41 Mortimer Street,London W1T 3JH, UK

    Journal of EnvironmentalScience and Health .Part A: EnvironmentalScience and Engineeringand Toxicology: Toxic/Hazardous Substances andEnvironmental EngineeringPublication details, including instructions forauthors and subscription information:http://www.tandfonline.com/loi/lesa19

    Anaerobic digestion ofsimulated distillery wasteusing flocculated and fixedcell reactorsH.M. Kitsos a c , W.J. Jones b d , R.S. Roberts a

    & T.G. Tornabene ba School of Chemical Engineering , GeorgiaInstitute of Technology , Atlanta, GA, 30332b School of Applied Biology , Georgia Instituteof Technology , Atlanta, GA, 30332c National Starch and Chemical Company ,Plainsfield, NJ, 07063d USEPA , Athens, GA, 30613Published online: 15 Dec 2008.

    To cite this article: H.M. Kitsos , W.J. Jones , R.S. Roberts & T.G. Tornabene(1993) Anaerobic digestion of simulated distillery waste using flocculatedand fixed cell reactors, Journal of Environmental Science and Health .Part A: Environmental Science and Engineering and Toxicology: Toxic/Hazardous Substances and Environmental Engineering, 28:5, 1099-1121, DOI:10.1080/10934529309375931

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    http://dx.doi.org/10.1080/10934529309375931http://www.tandfonline.com/page/terms-and-conditionshttp://www.tandfonline.com/page/terms-and-conditions

  • J. ENVIRON. SCI. HEALTH, A28(5), 1099-1121 (1993)

    Anaerobic Digestion of Simulated DistilleryWaste Using Flocculated and Fixed Cell

    Reactors

    H.M. Kitsos*School of Chemical Engineering

    Georgia Institute of Technology, Atlanta, GA 30332

    W.J. JonestSchool of Applied Biology

    Georgia Institute of Technology, Atlanta, GA 30332

    R.S. RobertsSchool of Chemical Engineering

    Georgia Institute of Technology, Atlanta, GA 30332

    T.G. Tornabene*School of Applied Biology

    Georgia Institute of Technology, Atlanta, GA 30332

    Abstract

    One- and two-stage reactors were used to investigate the reductionin chemical oxygen demand (COD) of simulated stillage waste from anethanol distillery. In both configurations, cells both flocculated and fixed

    * Current address: National Starch and Chemical Company, Plainsfield, NJ 07063. Current address: US-EPA, Athens, GA 30613. To whom correspondence should be sent.

    1099

    Copyright 1993 by Marcel Dekker, Inc.

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  • 1100 KITSOSETAL.

    on a support matrix were utilized. A COD removal above 90% andmethane production of about 4 L/L day were obtained in both the one-and two-staged reactors. Electron microscopy revealed differences in themicrobial structure and bacterial populations comprising the flocculentbiomass and the biofilm attached to the support matrix. Early detectionof hydrogen and propionic acid increases were an indicator of pendingreactor instability.

    Introduction

    Recent advances in understanding the microbiology of diverse anaerobic bac-teria [1-10], biofilm formation [9-11], and development of new reactor con-figurations [12-14] are providing fundamental information that is requiredfor the development of anaerobic methanogenic fermenters for the effec-tive treatment of liquid waste. Different anaerobic reactor systems haveevolved over the last decade varying from laboratory experimental to full-scale size [12-15]. Despite this progress, additional data on the growth kinet-ics, substrate utilization, mass transfer limitations, inoculation procedures,operational parameters, and control of bacterial flocculation/biofilm devel-opment are needed for rigorous design of efficient and reliable systems [15-30].

    A series of experiments with one- and two-stage bioreactors was con-ducted in this laboratory to acquire the additional data required for the designof full-scale systems [31]. Different biofilm matrices were also tested in a spe-cially designed apparatus to measure biofilm development The experimentswere conducted to identify major operational parameters which enhance theattachment of anaerobic bacteria to matrices, reduce the start-up times ofanaerobic biofilm reactors, and achieve high productivity and operational sta-bility. The results reported in this manuscript are the operational parametersof a two-stage and one-stage reactor that culminated from this study. Theresults from the study on the biofilm development have been previously re-ported [32].

    Methods and Materials

    Reactor Systems

    Two reactor systems were used in this study. The first was an 80-L, two-stagereactor consisting of two vertical columns with rectangular cross section di-

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  • SIMULATED DISTILLERY WASTE 1101

    mensions of 18.5 cm x 18.8 cm and a volume ratio of stage 2/stage 1 of approx-imately 2.2 (approximate lengths of 79 cm and 158 cm, respectively, Figure 1).The ratio of 2.2 was estimated on the basis of the longer bioconversion timerequired for some fermentation products (propionate, butyrate) to CH4. Thereactor columns were constructed from plexiglas (1.27 cm thick). The bottomof each column consisted of an inverted pyramid with a 50 side inclinationfrom the horizontal. At the top of each column, removable covers were in-stalled with rubber gaskets and fastened with hexagonal bolts.

    Continuous pH monitoring was accomplished with a pH probe coupled toa pH microprocessor. The pH was adjusted by alkali addition to the first stage.The immobilization matrix was a ceramic material (Johns Manville Corpora-tion, PPR-8623) in tubular configuration with cross-sectional dimensions of23.5 mm (o.d.) and 16 mm (i.d.). Thirty-six matrix tubes per cross section areawere placed in stainless steel cartridges and loaded into the reactor chambers.The immobilization cartridge did not extend into the bottom 10% of eitherstage nor the top 10% and 5% of stages 1 and 2, respectively. The overall spe-cific surface area of the support matrix was 110 m2/m3 of reactor. The totalempty liquid volume of the system was 74 L while the active liquid volume ofthe reactor containing the matrix was 62.5 L. The loading rates given in thetext were calculated on the basis of the internal reactor volume.

    The two-stage reactor was operated in series in an upflow mode with anadditional, separate feed line to the second stage resulting in a parallel-plus-series feeding pattern. The reactor system was operated in a temperaturecontrolled room at 37 C. The feed was delivered with a variable speed Mas-terflex peristaltic pump. A conical-shaped insert was installed at the reactorinlets to enhance feed distribution. A pressure gauge was connected at thefeed inlet of the first stage to monitor the head pressure of the reactor. Atthe exit of the second stage, a T-shaped glass tube was installed for mount-ing a liquid level controller for biogas removal and collection. The controlleractivated a pump for biogas removal when the liquid level fell below the setpoint due to the pressure buildup from the gas accumulation. A two-psi checkvalve was installed in the final effluent line to keep the system under positivepressure. The biogas was collected and its volume measured by water dis-placement.

    The second test system was a single-stage reactor that closely resembledthe first stage of the two-stage reactor (Figure 1) with the exception that itwas made of glass and was water-jacketed for temperature control in a non-incubated room. The dimension of the column was 7.0 cm i.d. x 94 cm length

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  • 1102 KTTSOS ET AL.

    Figure 1: Description of the two-stage, methanogenic reactor for the treat-ment of alcohol stillage. Total volume: 74 L; AS, alkaline solution; LP, liquidpump; GP, gas pump; LS, liquid sampling port; GS, gas sampling port; LC,level controller; SP, stirring plate; GM, gas meter; PV, pressure valve; Gy gasvent; PHC, pH controller; F, feedstock; G, gas; W, water; R, recirculation.

    and contained the same tubular-configured Manville support matrix that wasused in the two-stage reactor. This one-stage system differed from the firststage of the two-stage system by the insertion of a stainless steel screen nearthe midpoint of the reactor preventing the matrix from extending into thelower 40% of the reactor volume. The lower 40% portion of this reactor wascontinually recycled by pumping liquid from just below the matrix materialto the reactor inlet at a ratio of 6:1 of the inlet feed rate. An alkali additionport was installed in the recirculation line near the connection where bothsubstrate feed and recirculation liquid entered the reactor. A pH probe wasinserted at the top of the reactor. Feeding and gas collection were conductedas described above.

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  • SIMULATED DISTILLERY WASTE 1103

    Table 1: Compositions of synthetic feedstocks for enrichment culture ex-periments and bioreactor operations.

    Component

    SucroseYeast autolysateSodium glutamateAcetic acidPropionic acidButyric acidEthanolMethanolIsobutanolIsoamyl alcoholn-propanoln-butanolCitric acidNH4CINa2HPO4KC1Na2SO4MgCl2-6H2OCaCl2-2H2ONiCl2-6H2OTrace metals6

    pH

    Alcohol stillage"(perL)12.00 g8.00 g0.30 g1.05 g0.10 g0.19 g0.95 g0.55 g0.04 g0.04 g0.04 g0.04 g0.04 g0.25 g0.64 g1.40 g0.70 g1.20 g1.50 g0.02 g2.00 mL5.6

    VFA mixture"(perL)

    _0.10 g

    -17.60 g4.60 g3.20 g

    -----

    2.10 g0.64 g1.40 g0.70 g1.20 g1.50 g0.02 g2.00 mL4.8

    "Corresponds to 30,000 mg COD/L.^Composition according to Batch et al. [33].

    Inoculation and Feeding Strategy

    The microbial inocula for the 80-L reactor system was generated from twoseparate enrichment cultures. The first stage of the 80-L reactor was inocu-lated with a mesophilic enrichment cultivated for 45 days on synthetic alco-hol stillage (Table 1). This inoculum was enriched to a biomass concentra-tion of 10.6 g volatile solids/L with a methanogenic activity of 4.2 g acetateconsumed/L-day. The second stage of the 80-L reactor was inoculated with a

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  • 1104 KITSOS ET AL.

    methanogenic enrichment cultivated for 45 days at 37 C with a volatile fattyacids (VFA) mixture (Table 1) as primary substrate. The enrichments werecontrolled to select predominantly for acetogens and methanogens. The eval-uations of the enrichments were determined by measuring the acetate con-sumption and methane production rates.

    A fed-batch withdrawing procedure was used throughout the cultureenrichment process to enhance the accumulation of settling (fiocculent)biomass and to eliminate filamentous buoyant cell growth. This was achievedby periodically discontinuing the mixing of the enrichment culture for 30 minfollowed by withdrawal of the upper 20% of the liquid from the vessel. In thisway, floating and suspended biomass was withdrawn from the culture whilesettled biomass remained within the systems. The appropriate feedstock wasthen mixed with continuous stirring. At the time of inoculation into the 80-Lreactor, the enrichment culture had a biomass concentration of 6.2 g volatilesolids/L and a microbial (predominantly methanogenic) activity of 0.66 g ac-etate consumed/L-day. Each reactor stage was inoculated by circulating theinoculum throughout the reactor for 24 h prior to continuous addition of feed-stock.

    The one-stage reactor was inoculated with flocculated biomass and mixedliquid from the lower 40% section of the first stage of the 80-L reactor. A 2:1ratio of feedstock/inoculum was used to maintain the cells in their physiolog-ical growth phase. The mixture was recycled internally for 24 h. After theinoculation period, continuous operation was begun with fresh feedstock.

    The feedstock consisted of synthetic alcohol stillage (Table 1) which wasformulated from the mean composition of stillages produced in ethyl alcoholdistilleries [34] and known trace metal requirements of anaerobes [33]. Theessential ingredients were a soluble mixture of sucrose, yeast extract, gluta-mate, citric acid, volatile fatty acids, alcohols, and mineral salts. In thesestudies, the COD of the feed varied from 1 to 30 g/L.

    Analytical Methods

    The hydraulic retention time was determined by using a pulse-input/responsetechnique using Li+ tracing for flow distribution analysis [35]. Li+ was mea-sured with a Perkin-Elmer Model 2380 Atomic Emission Spectrometer.

    Volatile organic acids were determined with a \&rian 3700 gas chromato-graph (GC) equipped with FID, a Varian 402 data system, and a 1.83-m x0.32-cm glass column packed with Porapack-Q, 80-100 mesh. Reactor sam-

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  • SIMULATED DISTILLERY WASTE 1105

    pies were prepared for GC injection by centrifugation and then acidificationwith 0.05 mL of 18M H2SO4 per milliliter of supernatant. Gas compositionwas determined at room temperature using a thermal conductivity detectorand the following columns: a) 1.83-m x 0.32-cm molecular sieve 5A column,80-100 mesh for the analysis of CH4, N2 and O2; b) 1.83-m x 0.16-cm columnpacked with silica gel, 60-70 mesh for the analysis of CO2; and c) 1.83-m x0.32-cm column containing Porapack Q, 80-100 mesh for measuring H2. He-lium was the carrier gas at a flow rate of 30 mL/min but was replaced by N2for analysis of H2.

    Total suspended solids (TS) and volatile suspended solids (VS) were de-termined by standard methods [36] with the modification that 10-mL sam-ples were centrifuged for 15 min at 15,000 rpm. The pellet was rinsed withdistilled water, recentrifuged and then transferred to preweighed aluminumdishes. The dishes were placed in an oven at 105 C for 12 h, weighed, andtransferred to a muffle furnace at 550 C for 30 min. The difference betweendried and ashed weights represented the VS while the difference between thedried and the empty dish weight gave the TS. Alkalinity was determined bystandard procedures [36]. The gas phase concentration of hydrogen sulfidewas determined by Cline's method [37] after trapping biogas in zinc acetatesolution (2N). Carbohydrate and protein were determined spectrophotomet-rically using the phenol-sulfuric acid [38] and Folin-Lowry [39] methods, re-spectively.

    Scanning electron microscopy (SEM) was accomplished with a Cam-bridge SEM using 20 kV accelerating voltage and 10 kA current inten-sity. Preparatory procedures were essentially those described by Dawes [40].Transmission electron micrographs (TEMs) of thin sections of biomass weretaken with a Philips E-200 Electron Microscope. Biomass and biofilm ac-cumulation on matrix materials were estimated by liquid displacement tech-niques described by Characklis et al. [41]. The difference between the dis-placement of water by the original wetted-matrix and the biofilm covered ma-trix was determined as an estimate of biomass/biofilm accumulation duringthe course of the experiment. The bacterial flocs/granular biomass particleswere measured with a diagnostic microscope (x 10) and a Venier micrometerafter the samples were appropriately diluted in an isotonic buffer and spreadon a pyrex plate for single particle isolation. The granules/bacterial floes werealso examined by SEM, TEM, Light diffraction microscopy (LDM) and X-raydiffraction analysis.

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  • 1106 KITSOS ET AL.

    U g

    100

    80

    60

    40

    20

    COD Removal

    COD Load

    CH4-stage1

    CH4-stage2

    0 10 20 30 40 50 60 70

    Time (days)

    Figure 2: Profile of COD load, COD removal, and CH4 composition of thegas phase of the two stage methanogenic reactor.

    Results

    Performance of the TVvo-Stage Reactor

    The two-stage bioreactor was designed to anaerobically treat a range of or-ganic loads. The reactor, containing a combined granular sludge-bed andimmobilization matrix, was operated for a period of 65 days on a syntheticwaste stream (Table 1). Fresh feedstock was supplemented to reactor stagetwo at a ratio of one part fresh feed to two parts of stage one effluent. Thisfeeding method was sufficient to maintain an active biomass in both reactorstages during start-up and periods of low substrate levels.

    During the continuous operation with a hydraulic retention time (HRT) of45 h, the COD level of the feedstock was increased gradually from 1,500 mg/Lto 30,000 mg/L within a period of 53 days (Figure 2, Table 2), correspondingto an organic load of 0.5-14.6 g COD/L-day. The relative concentrations ofVFAs in the effluents of both reactor stages are given in Table 3. The dif-

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  • Table 2: Performance data of the two-stage 80-L reactor."

    Time(days)

    13814162022252628303234384044525364

    Feed C O D(mg/L)

    1500300030003000450045006000600090009000116001200015000150002000025000250003000030000

    C O D load(g/L-day)

    0.51.11.21.01.41.52.12.23.53.43.94.16.66.88.611.112.414.613.4

    COD removal(%)

    48.077.079.081.587.092.093.595.295.095.695.494.594.494.793.394.694.394.191.9

    COD (mg/L)a

    S-l

    90084040029075054077553086589599010552080170023102450337041204810

    S-2

    78069064055559537539029045040053066084080013501345143017802440

    Biogas yield(L/gCOD)

    0.130.140.190.270.250.290.320.310.280.300.270.320.280.300.300.270.300.260.27

    % MethaneS-l

    77.474.088.589.083.590.790.986.076.282683.080.475.173.770.768.068.968.671.3

    S-2

    78.281.074.073.171.973.069.271.066.267.268.173.477.875.573.372.670.367.373.0

    pHS-l

    7.17.07.17.37.17.37.37.37.27.47.47.37.27.37.27.27.27.27.3

    S-2

    7.47.06.97.06.87.17.07.07.07.17.17.27.37.37.37.37.37.37.5

    1OCO

    p

    *

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