Treatment of Heterogeneous MixedWastes: Enzyme Degradation ofCellulosic Materials Contaminatedwith Hazardous Organics and Toxicand Radioactive MetalsL A U R A A . V A N D E R B E R G , *T R U D I M . F O R E M A N ,M O S E S A T T R E P , J R . ,J A M E S R . B R A I N A R D , A N DN A N C Y N . S A U E R

Chemical Sciences and Technology Division,Los Alamos National Laboratory, Mailstop E-529,Los Alamos, New Mexico 87545

The redirection and downsizing of the U.S. Department ofEnergy’s nuclear weapons complex requires that manyfacilities be decontaminated and decommissioned (D&D).At Los Alamos National Laboratory, much of the low-levelradioactive, mixed, and hazardous/chemical wastevolume handled by waste management operations wasproduced by D&D and environmental restoration activities.A combination of technologiessair stripping and biodeg-radation of volatile organics, enzymatic digestion of cellulosics,and metal ion extractionswas effective in treating aradiologically contaminated heterogeneous paint-strippingwaste. Treatment of VOCs using a modified bioreactoravoided radioactive contamination of byproduct biomassand inhibition of biodegradation by toxic metal ions in thewaste. Cellulase digestion of bulk cellulose minimizedthe final solid waste volume by 80%. Moreover, the residuepassed TCLP for RCRA metals. Hazardous metals andradioactivity in byproduct sugar solutions were removedusing polymer filtration, which employs a combination ofwater-soluble chelating polymers and ultrafiltration to separateand concentrate metal contaminants. Polymer filtrationwas used to concentrate RCRA metals and radioactivity into<5% of the original wastewater volume. Permeatesolutions had no detectable radioactivity and were belowRCRA-allowable discharge limits for Pb and Cr.

IntroductionThe redirection and downsizing of the U.S. Department ofEnergy’s nuclear weapons complex requires that manyfacilities be decontaminated and decommissioned (D&D).D&D activities are expected to generate waste volumes thatthreaten to overwhelm existing waste management facilities.At Los Alamos National Laboratory (LANL), 68% of the low-level radioactive, mixed, and hazardous/chemical wastevolume handled by waste management operations (11 642m3) was produced by D&D and environmental restorationactivities for the period from Jan 1996 through Oct 1997 (1).Treatment and disposal of this volume of waste is a daunting

challenge; generally, this waste is heterogeneous with respectto physical form and chemical and radiological constituents.Few technologies exist for treating D&D waste, and commonlyit is either stored on-site or buried in landfills. Because thetoxic and radiological constituents are only a small fractionof the total waste, it is highly desirable to develop treatmenttechnologies that reduce the burden on storage and landfillfacilities.

One class of low-level mixed waste (LLMW) generatedduring decontamination of facilities is cellulosic materials(cloth, paper, and other trash) contaminated with toxicmetals, radioactive metals, and organic constituents. Treat-ment of this type of waste for proper disposal requires (1)destruction of the hazardous organic compounds to belowregulatory levels, (2) separation and stabilization of hazardousmetals to meet land disposal restrictions, and (3) volumereduction. Because the bulk of cellulosic waste is readilycombustible, treatment by incineration in a permitted low-level waste incinerator and stabilization of the residual ashis a frequently proposed option. However, incinerationmethods suffer from very low public acceptance and sometechnical problems are associated with handling volatilemetals such as mercury and radioactive metals such asplutonium (2, 3). Biodegradation alone or in combinationwith chemical or physical techniques may offer significanttreatment and permitting advantages. The growing interestin the use of biotechnology for treatment of hazardous wastesstems from the ability of microorganisms or enzymes totransform a broad range of organic substrates in a wide rangeof matrixes (4, 5). In addition, biodegradation is wellunderstood, is accepted by the public, and is usually verycost-effective (2, 6). As a consequence of these attributes,there is a growing interest in using biological systems incombination with other technologies to treat a wider arrayof complex wastes containing both metals and organics.

Although there are many reports demonstrating thedegradation or separation of the individual components thatmake up complex wastes, treatment of these wastes willrequire combinations of technologies with different objectives(in this case, organic destruction and metals separation).Effective coupling of individual technologies into a processfor treating complex wastes will require knowledge of theinteractions between individual treatment steps. Theseinteractions have the potential to be detrimental or beneficial.For example, radioactive and toxic metals are known to inhibitmany biological processes, including degradation of xeno-biotics (7, 8) and cellulose (9). In contrast, degradation of anorganic matrix, such as cellulose, in which RCRA (ResourceConservation and Recovery Act) listed and radioactive metalsare entrained, may enhance the solubilization of metal ionsand thus their separation from the waste (10). Because ofthese interactions, we can expect that the outcome of atreatment process will strongly depend on the order andtype of unit processes used. To begin examining the effectsof interactions between multiple treatment processes andmultiple constituents in a complex waste on the overalltreatment effectiveness, we selected a heterogeneous cel-lulose-containing mixed waste from the storage inventory atLANL. By using this waste, we demonstrated that potentialdetrimental interactions between constituents could beavoided by combinations of physical, chemical, and biologicaltreatment techniques. Our goal was to combine technologiesin such a way as to allow treatment of a heterogeneous mixedwaste by destruction of the RCRA organic components,separation of the hazardous and radioactive metals, and

* Corresponding author. Phone: (505) 665-6493; fax: (505) 665-5052; e-mail: lvanderberg@lanl.gov.

Environ. Sci. Technol. 1999, 33, 1256-1262

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reduction of waste volume, all of which are important in themanagement of radioactive wastes.

Experimental MethodsPaint-Stripping Waste. This waste was generated by paintingover radiologically contaminated surfaces and stripping thepaint and radionuclides with organic solvent-based paintstrippers and cheesecloth wipes. The resulting paint-stripping(PSW) waste, a physically heterogeneous mixture of paint,paint stripper, and cheesecloth, was combined with workers’clothing and other rags and trash, placed into polyethylenebags, and sealed into 55-gal drums for storage. The samplesused in our studies were obtained from one of these drums.

Surrogate Waste. In the development of the treatmentprocess, a nonradioactive waste surrogate was used tooptimize the experimental conditions. This surrogate wasgenerated by stripping painted aluminum plates with acommercial paint stripper containing methanol, dichlo-romethane, and 2-propanol. After wiping the resultingmaterial with cheesecloth, it was sealed in polyethylene bagsand stored at room temperature for approximately 1 monthprior to characterization and use in experiments.

Chemicals and Reagents. All chemicals used in thesestudies were reagent grade. Dichloromethane, 2-propanol,methanol, and chloroacetic acid were obtained from Aldrichand used as received. Polyethylenimine (PEI) was obtainedfrom BASF. Polyethyleneimine carboxylate (PEIC) was syn-thesized by reaction of PEI with chloroacetic acid (11).

Physical Pretreatment of Cellulose. All cellulose waspretreated prior to cellulase degradation. Ball milling ofcellulose was performed using a Fisher ball mill, with a 5.7-Lcontainer, and 30 21-mm carburundum balls for 2.5 days.Greater than 98% (by weight) of the cheesecloth wasrecovered following treatment.

Bacterial and Fungal Cultures. Microbial cultures, Hy-phomicrobium sp. DM-2 (ATCC 43129) and Rhodococcusrhodochrous OFS (ATCC 29672), were obtained from theAmerican Type Culture Collection (ATCC, Rockville, MD)and maintained on agar plates as previously described (12).Trichoderma reesei QM9414 was also obtained from ATCC.A naturally occurring variant of T. reesei was used for theproduction of fungal cellulase; this variant had very highlevels of â-D-glucoside glucohydrolase activity during growthusing microcrystalline cellulose.

Cellulase Enzyme. Cellulase was produced by batchfermentation of T. reesei using microcrystalline cellulose asthe carbon and energy source. The medium employed wasthat of Laubdova and Farkas except that the ionic strengthof the citrate phosphate buffer was 100 mM (13). The enzyme-containing supernate was separated by centrifugation, andthe â-D-glucoside glucohydrolase activity was measuredcolorimetrically using 4-(nitrophenyl)-â-D-glucopyranoside(14). In this work, activity is defined as nmol of p-nitrophenolproduced/(min‚mL of enzyme solution).

Biodegradation in a Modified Gas Lift Loop Bioreactor.Biodegradation of the volatile organic constituents of thepaint stripper was performed in a modified gas lift loopbioreactor (GLLB) (12). Representative 100-g samples ofsurrogate or genuine waste were placed in the waste barrelof the bioreactor system and volatile organics were strippedfrom the waste into the bioreactor vessel by flow of ambientair at 5 scfh. Prior to organics stripping, the bioreactor wasinoculated with a consortium of Hyphomicrobium sp. DM-2(120 mg DWC/L) and R. rhodochrous OFS (140 mg DWC/L).The details of the design and operation of the gas loopbioreactor have been described previously (12). Headspaceand solution samples were taken from the bioreactor andanalyzed for volatile organics by gas chromatography-massspectrometry.

Cellulose Degradation by Cellulase. Ball-milled surrogatewaste or PSW was placed in a 2-L round-bottom flaskequipped with a motorized stirrer. Six hundred milliliters ofT. reesei cellulase enzyme solution (6.7 nmol of p-nitrophenol/min‚mL of solution â-D-glucoside glucohydrolase activity),200 mL of 50 mM acetate buffer, pH 4.8, and 2 mL of 100 mMsodium azide were added to the flask and stirred at roomtemperature (25 °C). Five-milliliter aliquots of reactionsolution were removed from the flask at various times andanalyzed for reducing sugars to monitor the course of thehydrolysis. When assays indicated that more than 3 g ofsoluble reduced sugar had been released, the reactionsolution was removed from the flask, the cellulase separatedfrom the sugars, and the enzyme recycled into the reactionvessel. This was accomplished by pipeting as much solutionas possible containing the soluble sugars and enzyme fromthe reaction flask into 50-mL disposable centrifuge tubes.This mixture was centrifuged at 5000 rpm for 15 min, andthe pellet containing undigested cellulose solids was returnedto the reaction flask. The supernatant was subjected toultrafiltration in a 0.3-L Amicon ultrafiltration cell using a10 000 MWCO polysulfone filter; the retentate (containingthe soluble cellulase) was returned to the reaction flask whilethe permeate (containing the harvested sugars) was held forsugar, metals, and radioactivity analyses. Addition of 50 mMacetate buffer was used after sugar harvests to maintainconstant volume in the reaction flask.

Metal Ion Separation. After cellulase treatment of thesurrogate or genuine waste, the permeates from the sugarharvests were subjected to two polymer filtration (PF) stepsin order to separate and concentrate the radioactive andtoxic metals released from the waste during cellulasetreatment. In the first step, the water-soluble polymer, PEIC,was added to the permeate at a concentration of 1.58 g/L(pH 7.0). The second step of PF used 0.745 g/L PEI (pH 7.0).Ultrafiltration steps were carried out in a 0.3-L Amiconultrafiltration cell using a 10 000 MWCO polysulfone filter.

Analytical Section. Volatile organics were determined bygas chromatography-mass spectrometry, GCMS, using aHewlett-Packard HP 5980 GC equipped with a HP 5972 MSdetector and an HP-5 capillary column. Gas-phase sampleswere obtained using a gas precision sampling syringe andwere manually injected into the GC. Liquid samples fromthe bioreactor were analyzed using a Tekmar 7000 headspaceanalyzer. Standard curves for each of the target organics,methanol, dichloromethane, and 2-propanol, were deter-mined with standards prepared using reagent-grade chemi-cals. The detection limit for the volatiles was 10 ppb. Reducedsugar concentrations were measured colorimetrically usingthe method of Dubois and glucose as the standard (15).

RCRA metal ion concentrations were determined by ICP-AES. All nonradioactive samples (surrogate waste) were runon a Varian model Liberty 220 ICP-AES. All suspect and low-level radioactive samples (authentic waste) were run on aLeeman PS series ICP-AES. Chromium and lead in the liquidsamples were stabilized prior to ICP analysis by acidifyingthe liquid sample to < pH 2 with concentrated nitric acid.Metal ion concentrations in the untreated PSW were obtainedby digestion of 10-g samples in concentrated nitric acid ona hot plate. After digestion and evaporation, the residueswere dissolved for ICP analysis. The analysis of the residualsolids after PSW treatment was performed on separatedcellulosic and paint fractions in order to provide a morehomogeneous sample. For the paint fraction, ∼0.2-g sampleswere digested in concentrated HNO3/HF (5:1 v/v), thesolutions were evaporated to dryness, and the residue wastaken up in water for ICP analysis. For analysis of the cellulosefraction, a ∼0.2-g sample was digested in concentrated H2-SO4, the H2SO4 solution evaporated to dryness, and theresidue heated with concentrated HCl. The solution was

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transferred to a volumetric flask and diluted with distilledwater for ICP analysis. Low residual metal ion concentrationswere anticipated for treated PSW so a series of blanks andspiked samples were also prepared and analyzed. Thedetection limits on these samples using the Leeman PS ICPwere 0.1 mg/L Cr and 0.5 mg/L Pb. The toxic characteristicleaching procedure (TCLP, EPA method 1311) was performedon the solid residue after treatment to determine theleachability of the metals in the waste and the suitability ofthe treated solid for land disposal.

Radiological Analysis. Scintillation counting was used todetermine the total R activities in untreated solid samplesthat were ashed and digested with a 1:1 (v/v) HNO3/HFsolution. Since direct R counting offers increased sensitivity,an Eberline Alpha counter was used to determine the totalR activities in treated and acid-digested solid and liquid PSWresidues. A minimum detection limit of 0.3 pCi was obtainedby taking 3 standard deviations of our counting background(26 counts in 50 min) and converting them to pCi.

Knowledge of the decontamination process suggested thatseveral actinides were present in the waste samples. For Rspectroscopic analysis, 1-g samples of the solid waste weredigested in concentrated HNO3 and the resulting solutionspassed over an anion-exchange column. The column waseluted with HCl-H2O2 (9:1 v/v) to remove Pu and the eluantelectroplated on a platinum disk for counting. A CanberraAlpha Environmental System R pulse height analyzer wasused to obtain the R spectrum that provided information asto the specific identity of the R emitters Am and Pu.

Results and DiscussionPSW is the multicomponent waste that was selected for studyfollowing a review of waste descriptions and characterizationdata for the LLMW in storage at LANL. This waste wasgenerated by a decontamination method used at LANL, whichinvolved painting and stripping of contaminated surfacessuch as glovebox interiors, walls, and floors. PSW is aheterogeneous mixture that contains RCRA-regulated metals(from the paint), RCRA-regulated organics (solvents fromthe paint strippers), radioactive metals, and bulk materials(cheesecloth and protective clothing). Treatment of PSWpresents many of the challenges common to D&D wastesand, consequently, is a good model for development oftreatment strategies for complex wastes and investigation ofthe interdependence of unit operation steps.

An optimum treatment scheme for PSW should ac-complish three objectives: destroy RCRA organics in thewaste, separate and concentrate the RCRA and radioactivemetals, and reduce the bulk waste volume. Accordingly, eachof these individual unit operations was built into ourtreatment scheme. Based on the description of the processand the materials used to generate the waste, as well as theRCRA classification of the stored waste, dichloromethane,2-propanol, and methanol were expected to be the RCRAorganic solvents. Cheesecloth and rags (cellulose-basedmaterials) were anticipated to be the largest contributor tothe waste volume. Chromium, lead, plutonium, and ameri-cium were expected to be the primary metal constituents.Dichloromethane, 2-propanol, and methanol are readilybiodegradable (16-18), so an obvious choice for organicsdestruction was biodegradation. Cellulose degradation bycellulase has been studied extensively as a possible route tobiomass conversion (19) and appeared to be an attractivestrategy for bulk volume reduction. Finally, polymer filtration,PF, has demonstrated advantages for dilute metal ionseparation and was selected to remove and concentrate theRCRA and radioactive metals (20-22). This techniqueemploys water-soluble chelating polymers in conjunctionwith ultrafiltration to accomplish metal ion recovery. A wide

variety of soluble chelating polymers with specific metal ionselectivities have been synthesized and reported (20).

The choice and order of the unit treatment operationswere also driven by consideration of potential interactionsbetween the unit operations and the constituents in the waste.Specifically, we desired to minimize any fugitive emissionsof volatile solvents and the potential inhibition of microbialand enzyme activity by toxic or radioactive metals, minimizegeneration of large volumes of radiologically contaminatedsecondary wastes including biomass, and ensure that anysecondary wastes that were generated could be readilymanaged by existing waste treatment processes at LANL.

Description of Unit Operations. A flow diagram illustrat-ing the treatment steps developed for PSW is shown in Figure1. The shaded boxes in this figure indicate the three objectivesfor treating PSW. RCRA organics are destroyed in a bioreactor,shown in A. Cellulase degradation of the bulk material andsolubilization of metals occur in the second unit operation,shown in B. The solubilized RCRA and R emitters areconcentrated and recovered by PF, shown in C.

Characterization of Surrogate and Authentic Wastes.Work with a nonradioactive surrogate waste allowed us todevelop and optimize individual unit operations prior totreating actual radioactive waste. The general physicalappearance of the surrogate and PSW was very similar withrespect to texture, color, and heterogeneity. The levels ofchromium and lead in the PSW and surrogate wastes weresimilar, suggesting that the surrogate was a good model forstudy of metal properties in the waste. The biggest differencebetween the PSW and the surrogate was that the PSW didnot contain detectable volatile organics. Presumably, theseconstituents either vaporized from the cheesecloth prior topackaging of the waste for storage or vaporized from thedrums during several decades of storage.

Destruction of RCRA Organics. The first unit operationselected for PSW treatment was biodegradation, principallybecause the PSW organics are volatile and immediatetreatment would minimize fugitive emissions. Based ondegradation studies with surrogate wastes, a modified gaslift loop bioreactor, GLLB, was selected in order to (1) mitigatemetal toxicity toward microbes and (2) eliminate the forma-tion of radiologically contaminated biomass. We havepreviously published our research on and development ofa system in which the toxic organics were removed fromsurrogate PSW by air stripping and transferred to a GLLB forbiodegradation by a defined consortium of R. rhodochrousOFS and Hyphomicrobium DM-2 (12, 23). The destructionof the paint-stripper solvents (methanol, dichloromethane,and 2-propanol) contained in the surrogate waste was fitreasonably by a numerical model for mass transfer andMonod kinetics (12). In tests with the surrogate waste, allRCRA organics in the GLLB were destroyed to below detectionlimits for the target organics. Biomass generated using thisGLLB would not be classified as a radioactive waste sincephysical contact between the biomass and the radioactivitywas avoided. Given the EPA classification and generatorknowledge, it was clear that no nonvolatile RCRA organicswere entrained in the PSW. Additionally, we were disap-pointed to find that the specific sample of PSW we obtaineddid not contain any detectable paint-stripper solvents.Despite this, a 100-g sample of PSW was treated in the GLLBidentically to the surrogate waste for consistency in thetreatment of surrogate and PSW. The absence of volatileorganics was confirmed by the lack of microbial growth inthe bioreactor and gas-phase sampling and GCMS analysisof the vapor phase.

Cellulose Degradation by Cellulase. The second unitoperation selected for PSW treatment was bulk materialdigestion with cellulase enzyme. The multicomponent cel-lulase complex from T. reesei contains three glucohydrolase

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activities: exo-1,4-â-glucanase (EC 3.2.1.4), endo-1,4-â-glucanase (EC 3.2.1.91), and â-D-glucoside glucohydrolase

(EC 3.2.1.21). exo-Glucanases hydrolyze the 1,4-â-glycosidicbonds in cellulose from the reducing sugar end of the linear

FIGURE 1. Flow sheet for the three-stage paint-stripper waste treatment process.

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polymer chain to produce glucose and cellobiose (4-â-D-glucosidoglucose). The endo-glucanases hydrolyze glycosidicbonds in the interior of the cellulose polymer chain to giveoligosaccharides that are susceptible to further hydrolysisby the exo-glucanases. â-D-Glucoside glucohydrolase hy-drolyzes cellobiose to glucose. Together, these enzymeactivities act cooperatively to catalyze the hydrolysis ofcellulose into cellobiose and glucose (19).

Cellulose digestion for volume reduction of a waste hasnot been applied to a complex system such as PSW. Ourstudies were designed to provide information on the deg-radation of heterogeneous systems. Four issues related tooperation of the full process train were examined for cellulosedegradation using both surrogates and PSW: (1) celluloseinhibition by accumulation of products, (2) bulk volumereduction of the treated cellulose, (3) metal ion release bythe action of cellulase on the waste, and (4) toxic effects ofmetals on cellulase.

Cellulose hydrolysis by cellulase is known to be inhibitedby accumulation of products, particularly cellobiose andglucose. Previous work has shown that it may be advanta-geous to remove the inhibitory soluble products from thereaction mixture during the course of the reaction (24, 25).We found this to also be true for our systems. During bothsurrogate studies and treatment of PSW, soluble productsand soluble cellulase enzyme were removed from the reactionmixture. Subsequently, the released sugars and enzyme wereseparated by ultrafiltration, and the enzyme was recycledback into the reaction vessel. In studies on surrogate waste,enzyme activity decreased to ∼40% of the original activityduring the first 5 days of cellulose degradation. Followingseparation of the released sugars, return of the enzymesolution to the reaction mixture restored the total activity to56% of the initial activity. A second sugar harvest resulted ina similar restoration of activity. Despite removal of releasedsugars from the reaction mixture, soluble â-D-glucosideglucohydrolase activity was below our detection limits after28 days; however, significant sugar release continued throughday 37 when the cellulase treatment was terminated for thesurrogate waste. This continued sugar release likely resultedfrom the activity of other enzymes in the cellulase complexor from â-D-glucoside glucohydrolase associated with in-soluble components in the solid residue.

Studies on surrogate waste demonstrated that mass andvolume reduction resulted from cellulase action on cheese-cloth; during the 37-day cellulase treatment on surrogatewaste, 12.2 g of reducing sugars was released (12.2% of thetotal waste mass) and 88% volume reduction was achieved.Similar results were obtained in the treatment of PSW.Reducing sugar release during cellulase digestion of PSW isshown in Figure 2A. During the 65-day treatment of PSWwith cellulase, 18 g of sugar was harvested. The volume ofsolid residue was reduced 80% from the volume of the originalball-milled PSW. Although the mass reduction was modest,the volume reduction was significant. Notably, the massbalance between the sugar harvested and the mass reductionin the solid residue was not met; 18 g of sugar was harvested,but the mass of the solids was reduced by only 8 g (Table 2).This observation suggests that significant enzyme or othercomponents in the crude cellulase preparation were absorbedto the PSW residue during digestion. Enzyme absorption tothe solid residue is also suggested by the observation thatsugar release from PSW during later treatment stagescontinued in the absence of detectable soluble enzymeactivity in the supernatant, as discussed previously. Althoughthe sugars removed represent slightly less than 20% of thetotal mass of waste treated, the degradation process gave aresidue that was greatly reduced (80%) in volume from theoriginal.

The extent of biomass conversion by cellulases is typically50-80% by mass (27, 28) on crystalline cellulose, which ishigh compared to the conversions obtained in this work.However, the makeup of both surrogate waste and PSW wasmuch more complex than typical substrates for cellulasedigestion. Since the substrate-enzyme interaction is by farthe dominant parameter in determining the extent ofconversion (29), it is likely that the limited conversions thatwe observed for our surrogate waste and for PSW were dueto the bound paint stripper and paint, which limited celluloseavailability. Longer digestion periods for PSW would haveresulted in additional degradation of the material and furthermass reduction. Presumably, larger volume reductions wouldalso be obtained.

Metals Solubilization. Mobilization of Pb and Cr as wellas R emitters occurred during enzymatic digestion (Figure2B). Notably, the time course for release of sugars and metalsis nearly identical. This suggests that the two processes arerelated, as has been shown for metals in foods (10). Greaterthan 90% of the solubilized chromium (4.5 mg) and abouthalf of the solubilized lead (8.2 mg) were removed from thereactor during harvest of the sugars and collected in the sugarpermeate after ultrafiltration and recovery of the cellulaseenzyme for recycle. The remainder of the solubilized lead(9.0 mg) was associated with the enzyme retentate. Scintil-lation counting showed that 1070 nCi of radioactivity (totalR) was released during PSW digestion; 52 nCi was detected

FIGURE 2. (A) Total harvested glucose during cellulose degradationof paint-stripping waste. (B) Solubilization of lead, chromium, andradioactivity during cellulose digestion of paint-stripper waste.

TABLE 1. Metal Concentrations Remaining in PermeateSolutions after Treatment of Sugar Harvest Solutions Using PF

soln [Cr], mg/mL [Pb], mg/mL total r, pCi/mL

before PF 1.4 4.3 610after PF with PEIC 1.0 <1.0 60after PF with PEI 0.2 <1.0 BB

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in the soluble enzyme retentate. In the final sugar harvest,the total R was at background levels. The solid residueremaining after treatment was analyzed for its metal ioncontent. Complete acid digestion of representative samplesof the solid for analysis showed that 10.0 mg of Cr and 16.3mg of Pb remained in the treated residues. Thus, 33% of thetotal Cr and 51% of the total Pb in the original sample fortreatment were mobilized during enzymatic digestion. Todispose of this solid residue as low-level waste, it is necessaryto show the absence of leachable RCRA metals. Two 25-gsamples of the treated PSW residue were analyzed by TCLP.Less than 1.0 mg/L Pb and 0.1 mg/L Cr were in the TCLPextractant solutions. TCLP limits are 5 mg/L for both Pb andCr. There was no detectable radioactivity in the TCLPsolutions. Although the cellulose digestion process did leavePb and Cr in the PSW residue, the levels were below TCLPregulatory limits. Further studies to evaluate the mechanismof enzymatic metal mobilization are warranted.

Metals solubilized as a result of enzymatic degradationappeared to not have a significant effect on the cellulosedegradation process. In separate experiments the cellulaseactivity was reduced by only 31% in the presence of 500mg/L Cr(VI) and 5% by 4000 mg/L lead. Since the metal ionconcentrations expected during cellulase treatment of PSWwere 100-1000 times lower than these were, it is very unlikelythat metal inhibition of cellulase activity would significantlyimpact the treatment process. Experiments on both surrogatewaste and PSW showed that once released, Pb and Cr couldbe collected with harvested sugars and removed from thesesolutions by PF to achieve RCRA discharge limits.

Metals Recovery. The third operation in PSW treatmentwas metal ion extraction using PF. Analysis of all sugarharvests from PSW digestion showed that Pb and Cr levelsfor the samples were at or below RCRA discharge limits.However, these solutions represent a significant secondarywaste volume that could contain higher concentrations ofmetals from other drums of PSW. Consequently, we wishedto demonstrate removal and concentration of the metalsand radioactivity from this secondary waste. The sugarsolution harvested on day 13 (Figure 2B) had the highestmetal ion concentrations and, consequently, was selected todemonstrate the third step in the process, PF. This methoduses water-soluble polymers to sequester solubilized metaland ultrafiltration to separate the polymer-bound metal ionsfrom the remainder of the treated solution. The polymer andthe metals bound to it are too large to pass through a size-exclusion ultrafiltration membrane and remain in the re-tentate. The solution that passes through the ultrafiltration

membrane, the permeate, contains uncomplexed metals andsmall molecules such as sugars. Two polymers were evaluatedin this study for metal ion removal from the sugar harvestsolutions, a commercially available polyamine, polyethyl-enimine (PEI), and an aminocarboxylate polymer (PEIC) (11).Previous work in our laboratories has shown that PEICextracted Pb and Cr(III) from both solid matrixes and aqueoussolutions and that PEI removed metal oxyanions such aschromate (30). For PF treatment, a 50-mL aliquot of the day13 sugar solution was mixed with PEIC. After ultrafiltration,ICP-AES analysis of the retentate showed that 80% of the Pb,30% of the Cr, and 92% of the R emitters were bound to thepolymer, leaving <1.0 mg/L Pb, 1.0 mg/L Cr, and 60 pCi/mLof radionuclides in the polished solution. Data are shown inTable 1. Our previous studies have shown that PEIC primarilybinds cationic species, which is consistent with retention oflead and the R emitters, since they would be expected to becationic species under these conditions. The lower removallevels for Cr with PEIC suggest that this metal is present asan oxyanion (CrO4

2-) in the sugar solutions. Subsequenttreatment of the permeate from the first stage of PF withcationic PEI removed the residual Cr and R emitters.Chromium concentrations were reduced by 73% to 0.3 µg/mL, and the total R counting of the solution showedbackground levels of radioactivity. During this process, thePb, Cr, and R emitters, Pu and Am, were concentrated into2 mL of retentate solution, less than 5% of the original volumesent for treatment. The final permeate from the processcontained nonhazardous levels of RCRA metals and nodetectable radioactivity. This nonhazardous aqueous-basedwaste was suitable for discharge to the Los Alamos NationalLaboratory Low-Level Wastewater Treatment Facility.

Table 2 contains a summary of the individual processsteps for the treatment of PSW. For each stage of the process,the residual metal ion concentrations and estimated byprod-uct wastes streams are tabulated.

An important consideration in designing a waste treat-ment process is its estimated cost in comparison to alterna-tives. These laboratory-scale experiments were primarilydesigned to demonstrate the feasibility of coupling multipletechnologies to effectively treat a complex waste and not togenerate data for cost estimates. However, cost estimates forthree of the unit operationssGLLB (31), ball milling (32),cellulose digestion (33-35)shave been reported in theliterature and can be combined to roughly estimate thetreatment costs for PSW. For PF, a LANL technology, costestimates were obtained from pilot-scale fabrication andoperations. For each of the unit operations, these data were

TABLE 2. Summary of Treatment of Paint-Stripper Waste

process stage mass or vol hazardous component characterization waste for disposal

untreated samplea 100 g 17 mg Cr 100 mg Pb 1500 nCi R no disposal option

(A) VOC destruction 100 g no volatile organics detected nonesno biomass produced

(B) bulk vol reduction 100 g total Cr total Pb total rsolid after treatment 92 g 10 mg 16.3 mg 310 nCi 92 g LLRWb

combined sugar harvests 3180 mL 4.5 mg 8.2 mg 1020 nCi solns to stage Cenzyme concentrate 17 mL 0.4 mg 9.0 mg 52 nCi 17 mL LLMWtotal metal recovered 14.9 mg 33.5 mg 1382 nCiTCLP on treated solids <1.0 mg/L <1.0 mg/L BBc

(C) metals separation [Cr] [Pb] total rcombined sugar harvest 3180 mL 1.4 mg/L 2.6 mg/L 1020 nCi 130 mL LLMWd

sugar harvest day 13 50 mL 1.4 mg/L 4.3 mg/L 305 nCisample after treatment 2 mL 0.3 mg/L <1.0 mg/L BB 3050 mL non-RCRA waste

a Analytical was done on a representative 100-g sample from a 55-gal drum of waste. b Low-level radioactive waste. Digestion gave 8% massloss and a 80% volume reduction. Sugar (18 g) was harvested during the digestion, whereas mass reduction in the solid residue was only 8 g.The difference in the mass of sugars harvested and the mass reduction of the solids is attributed to the mass of cellulase enzyme added to thereactor during the digestion process. c Below Background. d Low-level mixed waste. Estimated using concentration factor (25-fold) for day 13waste treated with PF. Regulations require that all suspect wastes at Los Alamos National Laboratory be sent to low-level wastewater treatment.

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used to estimate the capital and operation costs per massunit of PSW treated. Because of the small volume of wasteto be treated (relative to the industrial scales reported in theliterature), the required treatment capacity will be signifi-cantly less and the capital equipment required to treat PSWwill need to be depreciated more rapidly. Cost estimates forthe capital given in the literature were adjusted by depre-ciating the total capital costs over 12 months. The base costestimate for PSW treatment assumed a treatment capacityof 4 drums/day (91 kg of PSW/day) and PSW compositionof 560 ppm chlorinated solvents, 170 ppm Cr, 1000 ppm Pb,and 15 000 nCi/kg of R radioactivity. These assumptions yieldcost estimates (expressed per kg PSW) for the unit processesof $0.11/kg for GLLB, $1.49/kg for ball milling, $1.07/kg forcellulose hydrolysis, and $0.45/kg for PF, giving a totalestimated treatment cost of $3.12/kg PSW or $70/drum. Inaddition, the solid residue remaining after treatment will besent for disposal at a cost of $2067/m3. This disposal cost willadd $3.80/kg to the total cost of managing the waste, bringingthe total cost to $6.92/kg or $156.5/drum. Because of thedifferences in scale and radiological content between theliterature data and the proposed process, it is likely that theseestimates are low. However, even assuming an order ofmagnitude increase, the estimated treatment cost of $1565/drum is well within the range of treatment cost for otherradiologically contaminated waste streams recently treatedat LANL; for example, uranium turnings in oil, $1500/drum;RCRA organic based scintillation cocktail, $800/drum. Pres-ently, PSW is classified as LLMW. Current estimates fordisposal of LLMW at Los Alamos National Laboratory are$20 000/drum (36). HWP treatment of this waste will reducedisposal costs by over an order of magnitude.

AcknowledgmentsWe thank the Department of Energy Environmental Man-agement Program Office at Los Alamos National Laboratory,for financial support, Brandy L. Duran for assistance withanalytical analysis, and Dr. Wayne Smith for useful discus-sions.

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Received for review April 27, 1998. Revised manuscript re-ceived November 30, 1998. Accepted January 18, 1999.

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