Conversion of paper sludge to ethanol in a semicontinuous solids-fed reactor
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- Fan, Z., South, C., Lyford, K. et al. Bioprocess Biosyst Eng (2003) 26: 93. doi:10.1007/s00449-003-0337-x
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Conversion of paper sludge to ethanol was investigated with the objective of operating under conditions approaching those expected of an industrial process. Major components of the bleached Kraft sludge studied were glucan (62 wt.%, dry basis), xylan (11.5%), and minerals (17%). Complete recovery of glucose during compositional analysis required two acid hydrolysis treatments rather than one. To avoid the difficulty of mixing unreacted paper sludge, a semicontinuous solids-fed laboratory bioreactor system was developed. The system featured feeding at 12-h intervals, a residence time of 4 days, and cellulase loading of 15 to 20 FPU/g cellulose. Sludge was converted to ethanol using simultaneous saccharification and fermentation (SSF) featuring a β-glucosidase-supplemented commercial cellulase preparation and glucose fermentation by Saccharomyces cerevisiea. SSF was carried out for a period of 4 months in a first-generation system, resulting in an average ethanol concentration of 35 g/L. However, steady state was not achieved and operational difficulties were encountered. These difficulties were avoided in a retrofitted design that was operated for two 1-month runs, achieving steady state with good material balance closure. Run 1 with the retrofitted reactor produced 50 g/L ethanol at a cellulose conversion of 74%. Run 2 produced 42 g/L ethanol at a conversion of 92%. For run 2, the ethanol yield was 0.466 g ethanol/g glucose equivalent fermented and >94% of the xylan fed to the reactor was solubilized to a mixture of xylan oligomers and xylose.
KeywordsPaper sludgeEthanolSemicontinuous bioreactor
Paper sludges are solid residues arising from wood pulping and papermaking operations. Sludge disposal is viewed as a substantial disposal problem for the paper industry [1, 2] and represents a significant cost at many mills. However, many paper sludges are potentially attractive raw materials for production of fermentation products such as ethanol [3, 4, 5, 6, 7, 8]. In contrast to essentially all native cellulosic materials , a significant fraction of sludges do not require pretreatment in order for near-theoretical yields to be achieved via enzymatic hydrolysis . This feature combined with the availability of utilities and other infrastructural elements at many mills offers significant potential for simplified processing for a facility located at a paper mill as compared to dedicated facilities processing other cellulosic materials. Finally, enzymatic hydrolysis may facilitate recovery and reuse of minerals (e.g., titanium dioxide) from paper sludge. The combined value of ethanol production, avoided disposal, and mineral recovery can potentially exceed $200/dry ton paper sludge. Because of these features, paper sludge processing represents a potential point-of-entry and proving ground for emergent industrial processes featuring enzymatic hydrolysis .
In our previous report of detailed compositional analysis for 15 paper sludges , carbohydrate accounted for an average of 42% on a dry weight basis, with most of the balance consisting of ash (23.2 wt.% on average) and acid-insoluble volatile matter (17.4 wt.% on average). Individual sludges deviated substantially from these averages in some cases. The nature of this unexpectedly large fraction of presumably organic material was not elucidated but is of importance in the context of process design and economics.
Most prior studies of paper sludge processing via enzymatic hydrolysis and fermentation were undertaken primarily for the purpose of determining product yields and, in some cases, rates. Consistent with this objective, rather low substrate concentrations (<20 g/L cellulose in most cases) and batch vessels were employed in these studies, and little or no attention was given to producing ethanol at concentrations that are potentially recoverable in an economic operation. Fed-batch hydrolysis and fermentation of paper sludge was mentioned by Katzen et al. , however, no experimental data were presented. Lark et al.  reported production of 32 and 35 g/L ethanol from 180 and 190 g/L paper sludge, respectively, in 250-ml flasks operated in batch mode. Recovery of ethanol via distillation becomes more costly relative to that for process designs involving cellulosic substrates other than paper sludge at ethanol concentrations less than about 4 wt.% .
In this study we sought to produce ethanol from paper sludge under conditions more nearly representative of those expected in an industrial process and, in particular, at ethanol concentrations in excess of 4 wt.%. Pursuant to this, we developed and subsequently modified a semicontinuous, solids-fed reactor system. In order to close material balances for this system, it was necessary to refine our prior carbohydrate analysis techniques.
2 Materials and methods
2.1 Sources of sludge, yeast, and cellulase
Paper sludge was provided by Pulp and Paper of America from the Cascade mill (bleached Kraft process) in Gorham, NH. Saccharomyces cerevisiae D5A (provided by NREL) was used for simultaneous saccharification and fermentation (SSF). Cellulase (DP151) was from Iogen (Ottawa, Ontario) and β-glucosidase (Novozyme 188) was from Novo Nordisk (Wilton, CT). Sludge was stored at 4°C for 1 to 4 months before use.
2.2 Growth medium and solutions added to the reactor
Solid, moist paper sludge (moisture content about 70%) was packed into a 4-in. feed tube (described subsequently), autoclaved for 12 h at 121°C, and fed to the reactor as described subsequently. In addition, three solutions were fed to the reactor. The first solution was distilled water, which was sterilized for 60 min at 121°C. The second was a nutrient solution for either yeast extract–peptone “rich” medium or corn steep liquor–magnesium sulfate “lean” medium. The rich medium contained yeast extract and peptone as a fivefold concentrated solution to give a final concentration of peptone 20 g/L and yeast extract 10 g/L after combining with paper sludge and the other solutions fed to the reactor. The lean medium contained MgSO4 and corn steep liquor (CPC International, Summit-Argo, IL) as a fivefold concentrated solution to give final concentrations of 5 mM MgSO4 and 0.3% (volume/volume) corn steep liquor. The composition of the growth medium was based on that described by Kadam et al. . The concentrated nutrient solution was prepared by adding MgSO4 to 9 L of distilled water in a 10 L glass carboy, adjusting to pH=4.5 with sulfuric acid, autoclaving for 1 h, and adding filter sterilized corn steep liquor after cooling. The third solution was prepared by adding β-glucosidase at a ratio of 3 units/filter paper unit (FPU) to undiluted Iogen cellulase and filter sterilizing. The Iogen cellulase was centrifuged prior to filter sterilization; the activity of the supernatant was measured to be 100 FPU/ml.
2.3 Analysis of feed and effluent samples
Acid soluble lignin (wt.%) was determined by measuring the absorbance of the first quantitative saccharification hydrolyzate at 205 nM . The acid-insoluble mineral content and acid-insoluble lignin content were determined by heating the dried solid residue from the second quantitative saccharification in a muffle furnace at 500°C.
2.4 Soluble carbohydrate and ethanol
2.5 Power required for mixing
Power consumption for mixing was determined using a variable-speed motor adjusted to rotate at 100 rpm. The motor shaft was connected to a flat rectangular impeller blade of diameter 4 cm and height 1 cm, which was placed in a 600-ml vessel containing 350 ml of sludge. The current drawn by the motor was recorded when the motor was unloaded, turning in air, and when the motor was loaded, stirring sludge at varying concentrations and conversions. The difference in current drawn while loaded versus unloaded was used to determine the power requirement (amps×volts).
2.6 Reactor systems
In the original design (Fig. 1), a 9-in. long conical section of process pipe with diameter starting at 4 in. and tapering to 2 in. was attached at one end to the bottom of the chopper and at the other end to a 2-in. isolation valve. The 2-in. isolation valve was in turn connected to a 2-in. diameter flanged opening in the feedplate to a 4-in. diameter, 800-ml working volume fermentor with a custom two-blade spiral agitator (blade width 16 mm, blade diameter 90 mm, and blade length 115 mm) connected to a shaft that penetrated the bottom of the reactor through a friction seal (Applikon, Foster City, CA). A steam fitting was placed at the top of the chopper assembly, and a drain tube for condensate was placed just above the ball valve. To replace a near-empty feed tube with a new tube filled with paper sludge, the isolation valve was closed to isolate the fermentor from the chopper assembly. The near-empty feed tube was then removed and replaced with a previously autoclaved full feed tube. The feed tube–chopper junction was steamed in place at 121°C for 30 min. Thereafter the 2-in. ball valve was opened and normal operation resumed.
The retrofitted reactor (Fig. 2) featured the following changes relative to the original design: (1) a 4-in. isolation valve was placed between the feed tube and the chopper assembly, the 2-in. isolation valve and conical section between the chopper and the fermentor in the original design were eliminated, and steam a steam fitting and condensate line were added on the feed-tube side of the 4-in. valve; (2) a common drive (Model 6Z822A, Dayton Electric, Niles, IL) mounted on top of the chopper was used to continuously turn the chopper and reactor agitator at 60 rpm, and the seal at the bottom of the fermentor was eliminated; (3) a 2-in. long section of 4-in. process pipe, added between the chopper and the fermentor, was used to anchor a bearing for the chopper/agitator shaft and for introducing nutrient, enzyme, and distilled water. A horizontal rod at the bottom of the chopper was placed just above the bearing supporter to knock off any sludge that landed there. With these changes, the feed tube replacement procedure featured retracting the plunger, closing the 4-in. ball valve, removing the old feed tube, connecting the new feed tube, steaming in place, opening the 4-in. ball valve, and advancing the paper sludge plug through the ball valve until material is added to the fermentor.
2.7 Reactor operation
2.8 Calculation of flow rates, mass recoveries, and conversion
ΔV=cumulative volume of sludge fed during the duration of the steady-state (L),
ρsludge=sludge density (g dry sludge/L),
Δt=duration of the steady state (days).
f i,in =mass fraction of component i in unreacted paper sludge.
M k=the weight of dry solids removed by the k th withdrawal of effluent (g),
f i, k =mass fraction of component i per mass solids in k th withdrawal of effluent.
V k=the volume of the k th withdrawal of effluent (L),
C j, k =concentration of component j in k th liquid sample (g/L).
3.1 Compositional analysis
Paper sludge composition
Second quant. sacch.
3.2 Mixing characteristics
Our results (Fig. 5) indicate that it is exceedingly difficult to mix the unreacted paper sludge tested here at the solids concentration required in order to produce ethanol at the targeted concentration of 40 g/L. However, the mineral-rich residue present after complete enzymatic hydrolysis was no more difficult to mix than water. Other authors have also noted the pronounced liquification accompanying enzymatic hydrolysis of paper sludge . In light of these results, a continuous or semicontinuous operating strategy would appear to be advantageous as compared to batch operation. In particular, it is desirable in order to have reasonable mixing requirements that the solids in the reactor at any particular time have a high mean conversion.
3.3 Semicontinuous SSF in the original bioreactor design
Using the original reactor design, the solids leaving the reactor increased over time following addition of a new feed tube. Accumulation of solids was less pronounced when reactor effluent was withdrawn from the bottom of the reactor rather than the top. The effluent solids concentration decreased markedly each time the feed tube was changed, which we attribute to compression of the paper sludge plug following changing of the feed tube.
In addition to solids accumulation, the reactor of the original design also had some operational difficulties. Several times during the run shown in Fig. 1 as well as other runs, solids would accumulate in the conical section above the 2-in. isolation valve (Fig. 1), resulting in inconsistent sludge delivery and, in some cases plugging, which required the reactor to be shut down. In addition, the seal for the bottom-mounted agitator (Fig. 1) failed on several occasions.
3.4 Development of a retrofitted reactor and consistency of solids feeding
3.5 Semicontinuous SSF using the retrofitted reactor
Semicontinuous SSF was carried out in the retrofitted reactor for two 1-month runs. For both runs, the reactor was operated at a residence time of 4 days with feeding at 12-h intervals, effluent was withdrawn from the bottom of the reactor, and lean medium was used. No glucose or other soluble glucan was detectable at any point during either of the runs.
Material balance for run 2
Flow rate (g/day)
Recovery, conversion or yield*
Soluble glucose oligomer
Glucose in from enzyme
Soluble xylose out
Soluble oligomer xylan out
Xylan in the solid out
A laboratory-scale solids-fed bioreactor capable of metered aseptic feeding of paper sludge was operated for periods of up to four months (Fig. 6). A retrofitted design displayed more consistent feed delivery, substantially better operational reliability, and allowed good material balance closure. Operation of these reactors involved feeding paper sludge at a solids content of about 30 wt.%, with the balance moisture. At this solids content, which is representative of the material produced by dewatering (e.g., via filter presses) prior to disposal in commercial paper mills, paper sludge is a solid that is slightly moist to the touch. When dilution by liquid feed streams (cellulase, nutrients, water) was factored in, the overall solids content fed to the reactor was about 12 wt.%. In comparison to prior work involving aseptic continuous fermentation of cellulosic feedstocks , this study employed much higher solids concentrations and also involved substantially more fibrous feedstock.
The semicontinuous operating mode used herein, which we found to be a satisfactory solution to the impracticality of mixing unreacted paper sludge at high feed solids concentrations, may be distinguished from fed-batch operation. As usually interpreted in the biochemical engineering literature and textbooks, fed-batch operation involves initiation of a reaction at low concentrations of cells and substrate with subsequent addition of further substrate (and in some cases catalyst) allowing development of high product concentrations in the fermentation broth, which is harvested in a batch-wise fashion. We suspect that higher reactor productivity (e.g., g ethanol/m3 h) can be achieved in semicontinuous mode as compared to fed-batch mode because the time-averaged concentration of cellulose–cellulase complex and cells is higher in the former than in the latter. The feeding schedule used here (1/8 reactor volume every 12 h) was not varied and is a target for future optimization.
In our previous report of compositional analysis of paper sludges , we found much higher acid-insoluble volatile matter content (averaging 17.43% for 11 sludges) than observed here (3%). We believe that the much lower acid insoluble organic matter content (presumably lignin) reported here is due to our inclusion of a second quantitative saccharification step in our analytical protocol as well as the properties of the particular sludge tested. We suspect that the buffering capacity (alkalinity) of the sludge is a contributing factor to the need for a second quantitative saccharification step.
An ethanol concentration of 50 g/L was observed in run 1, for which the cellulase loading was 15 IU/g and a conversion of 74% was achieved. In run 2, which featured a cellulase loading of 20 IU/g and achieved a cellulose conversion of 92%, the average ethanol concentration was 42 g/L. It is at this point unclear what factors are responsible for the lower conversion observed in run 1 as compared to run 2. The most likely explanations in our view are the lower cellulase loading and/or higher ethanol concentrations in run 1. The kinetic model of South et al.  suggests that the ethanol concentrations observed in this study are near an inhibitory threshhold above which conversion falls sharply for simultaneous saccharification and fermentation featuring yeast fermentation operated at 37°C. Although higher ethanol concentrations are attainable at lower temperatures, this has the undesirable effect of lowering the rate of cellulase-mediated cellulose hydrolysis. Ethanol concentrations in the range 40–45 g/L are typical of process designs for pretreated cellulosic feedstocks [15, 16, 17]. At this concentration, ethanol recovery is relatively inexpensive and does not prevent such processes from having a very favorable overall energy balance .
Although xylose was not converted by the yeast used in this study, several xylose-utilizing ethanol-producing organisms have been developed [18, 19, 20]. It would be of interest to try one or more such organisms for semicontinuous conversion of paper sludge as xylan conversion has the potential to increase the ethanol yield by nearly 20% as compared to conversion of glucan only for the sludge we studied. Over 94% of the xylan orginally present in paper sludge was solubilized, although not all to monomeric xylose. Recovery of the xylan present in the feed was high at 96%, which we take to be an indication of the absence of contamination. Over the course of many runs with the semicontinuous system described herein, contamination was occasionally observed and was accompanied by a drop in xylan recovery.
We continue to believe that paper sludge is one of the most promising feedstocks for near-term commercial application of technology for converting cellulosic feedstocks into commodity products. Several results of this study are seen as significant steps toward the realization of this potential. These include development of a system suitable for laboratory-scale process development and production of recoverable ethanol concentrations. Important tasks associated with the bioreactor system remain in order to more fully evaluate commercial viability. Such tasks include demonstration of high conversion at lower cellulase loadings, optimization of the feeding schedule, testing performance with one or more xylose-utilizing microorganisms, and design and testing of a scaleable feeding device.
This study was partially supported by the Consortium for Plant Biotechnology Research (grant no. OR22072-111) and the State of New Hampshire Governor’s Office of Energy and Community Services (grant no. 622188).