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Available online at www.sciencedirect.com

Enzyme and Microbial Technology 43 (2008) 103–108

Cellulase deinking of fresh and aged recyclednewsprint/magazines (ONP/OMG)

Xiao Zhang, Sylvie Renaud, Michael Paice ∗Pulp and Paper Research Institute of Canada (PAPRICAN), 570 boul. St-Jean, Pointe-Claire, Quebec, H9R 3J9 Canada

Received 9 July 2007; received in revised form 7 November 2007; accepted 12 November 2007

bstract

Three commercial cellulases were evaluated for their effects on deinking of both fresh and artificially aged old newsprint (ONP) mixed withresh old magazines (OMG) at a ratio of 7:3. The enzymes were added at the beginning of the repulping stage followed by incubation for up toh. Although these enzymes were able to detach a significant amount of ink from the ONP/OMG, cellulase deinking was less efficient than either

ulphite or alkaline deinking chemistry. None of the three cellulase enzymes tested were able to deink aged ONP/OMG, and a poor deinkabilityas also observed by using either sulphite or alkaline chemistry. However, combining enzymes with sulphite chemistry significantly enhanced

ulphite deinking and provided a potential strategy to achieve effective deinking of aged newsprint at neutral pH.2007 Elsevier Inc. All rights reserved.

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eywords: Cellulase; Deinking; Repulping; Recycling; Old newspapers

. Introduction

A mixture of old newsprint (ONP) and old magazine (OMG)s the primary furnish used in most Canadian recycling mills.onventional ONP/OMG deinking chemistry employs sodiumydroxide in combination with a range of other chemicalsncluding sodium silicate, hydrogen peroxide and surfactants.his high pH and chemical-intensive process causes severalajor problems to the process and recycled paper quality, includ-

ng a high degree of fragmentation of large sticky contaminants,arkening of pulp fibre, increasing COD in process waters andhe need for substantial amounts of acid for neutralization [1].hese adverse effects have hindered the further developmentf alkaline deinking processes. Deinking ONP under neutralr weakly acidic conditions has shown promise to alleviate theegative impact of alkaline deinking [1,2].

Recent studies have shown that sulphite deinking chemistryperating at around pH eight yields a comparable deinkingfficiency to alkaline chemistry [1]. The implementation of a

ulphite deinking process in one Canadian recycling paper millas led to a significant economic savings [2,3]. The bright-ess of the sulphite deinked pulp is however slightly lower

∗ Corresponding author.E-mail address: mpaice@paprican.ca (M. Paice).

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141-0229/$ – see front matter © 2007 Elsevier Inc. All rights reserved.oi:10.1016/j.enzmictec.2007.11.005

han that of alkaline-deinked pulp where peroxide is added.herefore, high dosages of bleaching agents may be required

n the subsequent bleaching step to obtain the same brightness,lthough some mills have observed high or equal brightnessulp after switching from alkaline deinking to sulphite deinking3].

During the past 20 years, a number of enzymes, including cel-ulase, xylanase, laccase and lipase, have been evaluated for theirotential to replace hazardous chemicals in deinking recycledaper [4–7]. Cellulases and hemicellulases have been demon-trated to dislodge inks by peeling off fibres or fines on paperurfaces [4,6]. Lipases have shown some direct action on inkarticles either degrading oil carriers or breaking down pig-ents [5]. Lignin-degrading enzymes, such as laccase, also hold

ome potential for deinking ONP, as they may selectively removeurface lignin, and hence facilitate ink removal [8]. To date, cel-ulolytic enzymes have shown the most promising results foreinking of mixed office waste (MOW). Both pilot plant trialsnd mill applications have demonstrated that cellulase deink-ng can reduce chemical cost, enhance ink and stickies removal,mprove drainage and runnability, and decrease COD and BODontent in process waters and effluent [9–11]. Recent develop-

ents in genetic engineering have also broadened the optimum

onditions of the enzymes, enabling them to work under somextreme mill conditions. The comparable pH range betweennzyme and sulphite deinking make it possible to combine the

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wo deinking methods to create a synergy for improving neareutral deinking.

Recently, the “summer effect” has become a major challengeo recycling paper mills primarily using ONP. “Summer effect”efers to a period during the summer months when mills experi-nce a loss in pulp brightness. The exposure of recovered paper toigh temperatures, especially in closed containers during trans-ortation, accelerates the oxidation and polymerization of inkarticles [12]. The oxidized and/or polymerized inks are proneo fragmentation during alkaline deinking and are difficult toemove by the subsequent flotation process. Consequently, aignificant reduction of brightness and cleanliness of the finalulp occurs. So far, neither conventional nor sulphite deink-ng chemistry has been effective in dealing with the “summerffect”. Compared to chemical deinking methods, enzymaticeinking exerts a different ink removal mechanism which mayive enzymes some advantages. For example, it is proposed thatellulase enzymes remove inks from recycled paper by peelingff the fines and small fibrils on fibre surfaces [4,13].

In this study, we determined the effects of cellulases oneinking fresh and aged ONP and compared the results withoth alkaline and sulphite deinking. Subsequently, we exam-ned the potential of combining enzyme with sulphite chemistryo achieve a better natural deinking strategy for deinking agedNP.

. Materials and methods

.1. Recycled paper and enzymes

A mixture of ONP/OMG at a ratio of 70:30 was used as recycled paper fur-ish. Three commercial cellulases, NS613, NS476 and NS342, obtained fromovozymes North America were used in the study and designated as EA, EB

nd EC, respectively. NS342 is a multi-component cellulase, while NS476 andS613 are mono-component endoglucanases. All three cellulases used in the

tudy have broad pH profiles, exhibiting more than 50% of their maximum activ-ty in a pH range from 6 to 9. An enzyme dosage of 300 U/kg of ONP/OMG wassed in this study for the treatment of ONP/OMG. The protein content of thenzyme preparations was measured by the Bradford Assay using bovine serumlbumin (BSA) as the protein standard (Biorad, Hercules, CA). The enzymectivity was measured as reducing sugars produced from carboxymethylcellu-ose (CMC) substrate.

.2. Aging of ONP

An artificial aging procedure was used to convert fresh ONP to aged ONP12,14]. The old newsprint was initially cut into paper strips of 8 cm × 20 cmn size, and then placed in an oven with a temperature maintained at 60 ± 3 ◦Cor 25 h with air circulation. The relative humidity during the aging was about0%.

.3. Repulping

The repulping of ONP/OMG at a ratio of 70:30 was carried out at 10%onsistency and 50 ◦C for 3 min in a helical pulper at a rotation speed of 800 rpm.charge of 625 g of mixed ONP/OMG furnish was added to the solution which

ad been warmed up at 45 ◦C. The recipes for different deinking methods were

s follows: (1) control: 0.06% surfactant, (2) enzyme: 0.06% surfactant withither of the cellulase enzymes, (3) alkaline: 60 ppm CaCl2, 1.0% NaOH, 2.0%ry Na2SiO3, 0.6% sodium oleate, 0.2% DTPA, and 1% H2O2 (all based on.d. paper), (4) sulphite: based on a near-neutral deinking method previouslyescribed [2,3]. Following the repulping, pulp samples were incubated at 50 ◦C

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l Technology 43 (2008) 103–108

or 0 h, 1 h, 2 h, and 3 h, before either subsequent hyperwashing or preparingandsheets for total ink measurements.

.4. Hyperwashing

After repulping, 10 g of pulp was hyperwashed with de-ionized water fol-owing the protocol described previously [15]. Handsheets (1.2 g) were preparedsing hyperwashed pulp before optical measurements.

.5. Optical measurement

The effective residual ink concentration (ERIC) and brightness values wereetermined according to the procedure previously described [15].

. Results and discussion

.1. The efficiency of enzymes to deink ONP/OMG

Pretreatment of recycled paper with these enzymes hashown some promising effects for deinking [16]. It wasecognized that pretreatment of recycled furnish prior toepulping is not a practical approach in existing deinkinglants. Therefore, in this study, we chose to add the enzymest the beginning of the repulping stage with the surfactant andecycled paper without pH adjustment. It was recognized thatxcessive enzyme dosages would reduce pulp strength andause darkening effects while a low enzyme dosage wouldave little effect on deinking [16,17]. The dosage effect of thehree cellulases during deinking was previously studied and anptimum enzyme dosage range between 200 U/kg and 600 U/kgf recycled paper was suggested [17]. Therefore in this study,e chose an enzyme dosage of 300 U/kg of ONP/OMG. The

nitial pH of the stock mixture at the repulper was 7 and the pHt the end of repulping was about 7.5. After repulping, the stockas incubated at 50 ◦C for up to 3 h. In control experiments,nly surfactant was added with the recycled paper in theepulper. The pulp was subject to a hyperwash for removing inkfter the incubation. Measuring ink content on pulp pads beforend after hyperwash has been shown to be a simple and accurateethod to determine the amount of total and bound ink.The total ink content of enzyme treated and control samples

re shown in Fig. 1. Both EA and EB treatments resulted inower ERIC values than the control samples at all four incubationimes, while the EC-treated sample had a similar ERIC value tohe control. The brightness values of the pulp obtained afterncubation were also determined (Fig. 2).

It appeared that EB-treated samples were about one pointrighter than the controls. There was also a slight increase inrightness of EA- and EC-treated samples. The total ink contentndicates the degree of ink fragmentation during the repulp-ng. The lower ERIC and higher brightness values obtainedfter enzymatic treatments suggested that the cellulases mayelp facilitate ink removal with less fragmentation. The ERICalue measured on the pulp pads obtained after hyperwash deter-

ines the concentration of bound ink attached to fibres and/or

rreversibly redeposited on long fibres after repulping.All three enzyme treatments led to a significant decrease in

ound ink content (Fig. 3). The lowest bound ink content was

X. Zhang et al. / Enzyme and Microbial Technology 43 (2008) 103–108 105

Fig. 1. Total ink content of ONP/OMG with and without enzymatic treatmentfollowed by incubation up to 3 h (standard deviations are within ±10 ppm forall the measurements based on eight replicates).

Fig. 2. The brightness of pulp obtained after repulping ONP/OMG with andwithout enzymatic treatment followed by incubation up to 3 h (standard devia-tions are within ±10 ppm for all the measurements based on eight replicates).

Fig. 3. Bound ink content of ONP/OMG with and without enzymatic treatmentfollowed by incubation up to 3 h (standard deviations are within ±10 ppm forall the measurements based on eight replicates).

Fig. 4. Final brightness of hyperwashed pulp obtained from deinkingO3e

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NP/OMG with and without enzymatic treatment followed by incubation up toh (standard deviations are within ±10 ppm for all the measurements based onight replicates).

btained on the pulp samples treated by EB, a monocompo-ent endoglucanase (approximately 25 ppm lower than controlalues). Endoglucanases have previously been found to removenk particles attached to MOW fibre surfaces through a “peelingction” [13]. The brightness values of hyperwashed pulp sam-les were similar with a slight increase from enzyme-treatedamples (Fig. 4). It was found that different incubation timesad little effect on ink removal or brightness increase. It shoulde mentioned that although the zero time represents the sampleshat were directly subjected to hyperwash without any furtherncubation, it usually took approximately 20 min during the labxperiments to transfer the pulp from the repulper and weightut appropriate amounts of samples for hyperwash. In a typicalill operation, there is also a retention time between repulping

nd flotation deinking. Therefore, the addition of enzyme at the

epulper is likely to be a practical approach in a mill process.

The positive effects of cellulase treatments on reducing inkontent and increasing brightness of ONP have been observed innumber of previous studies [5,16,18]. However, the mechanism

ig. 5. The total and bound ink contents of ONP/OMG obtained after deinkingith either EB (NS476) or BSA at the same protein content (standard deviations

re within ±10 ppm for all the measurements).

106 X. Zhang et al. / Enzyme and Microbia

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ig. 6. Comparing enzyme treatment with alkaline and sulphite chemistries fornk removal from fresh ONP/OMG (standard deviations are within ±10 ppm forll the measurements based on eight replicates).

nvolved in the cellulase deinking is not yet fully understood.ne issue remaining to be clarified is whether the “surfactant

ffect” of enzyme proteins plays a role in enzymatic deink-ng. To gain further understanding, we replaced enzymes with a

odel protein compound, bovine serum albumin (BSA), dur-ng deinking and determined its effect on ink removal. TwoSA dosages, 25 mg and 400 mg, were used to correlate to the

ange of enzyme protein content used in the experiments. Aseen in Fig. 5, the addition of BSA led to little improvement innk removal compared to the control experiment. Therefore, thebserved ink removal after enzymatic treatments was due to theatalytic activity of the enzymes.

.2. Comparing the deinking efficiency between enzymesnd chemicals

Sulphite deinking operates at near neutral pH conditions andas been implemented in at least two recycling mills. Both sul-hite and alkaline deinking chemistries were also evaluated inur study using the same ONP/OMG furnish and compared withhe results obtained from enzyme deinking. The results obtainedrom EB treatment were used for the comparison as it seemedo be the most efficient cellulase among the three. It was appar-nt that alkaline chemistry resulted in the best deinking effects.he total ink content of alkaline-deinked pulp was about 75 ppm

ower than that obtained from either sulphite or enzyme deinkedulp (Fig. 6). The bound ink content of alkaline-deinked pulpas about 11 ppm and 21 ppm lower than respective sulphite-

nd enzyme-deinked pulp. The brightness value of alkaline-einked pulp after hyperwash was significantly higher than thoserom sulphite- and enzyme-deinked pulp under the same con-itions. The differences in ink content and brightness valuesetween the pulps obtained from the two chemical deinkingethods have been observed during a previous mill trial using

ulphite chemistry [19,20]. It was demonstrated during the sul-hite trial that the ERIC of deinked pulp feeding to the flotation

as approximately 100 ppm higher than conventionally deinkedulp. The deinked pulps collected as flotation accepts were onverage 2 ISO brightness lower than those obtained before therial. The high brightness obtained after alkaline deinking was

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l Technology 43 (2008) 103–108

ikely due to the presence of hydrogen peroxide. Therefore, aigh dosage of sodium hydrosulphite was needed in the sub-equent bleaching step to recover the brightness of sulphiteeinking pulp.

Although enzymes had shown positive effects on ink removal,t must be recognized that enzyme deinking was overall inferioro either sulphite or alkaline deinking methods. The attempt toeplacing chemicals with enzymes in deinking has been pur-ued for over 20 years, with promising applications achievedn recycling plants using mixed office waste. However, enzymeeinking of ONP/OMG has not been successfully demonstratedt a commercial level. The lignified nature of the ONP fibre sur-ace is likely a major factor hindering the effects of cellulasenzymes.

.3. The efficiency of enzymes to deink aged ONP/OMG

So far, there has been little investigation of using cellulasenzymes for deinking aged ONP. Unlike with chemical deink-ng, cellulases can dislodge ink particles through peeling offurface fibres or fines [4,13]. Whether this “unique” mechanismonfers on enzymes some advantages for summer ink removalarrants further investigation. An artificial aging procedure wassed to convert ONP to aged ONP [12]. A previous study hasemonstrated that this procedure could mimic natural newsprintging conditions to provide representative samples for studyinghe aging effects. All three enzymes were used and the treatmentsere carried out in the same manner as for fresh ONP/OMG.s a result of newsprint aging, the total and bound ink contentf the control samples increased from 737 ppm and 135 ppm to24 ppm and 193 ppm, respectively (Table 1). This increase isimilar to the level experienced in a typical recycle mill dur-ng summer time. It was disappointing to find that all threenzymes had few or adverse effects on aged ink removal andulp brightness after hyperwash. The high ink contents fromA and EC treatments suggested that a greater ink fragmenta-

ion occurred during the enzyme treatment of aged ink. This waslso confirmed by the low brightness value of the ensuing pulp.

The efficiencies of either alkaline or sulphite deinking ofged ONP/OMG mixture were also examined. Comparingigs. 6 and 7, it can be seen that alkaline chemistry has a poorerapability to deink aged ONP/OMG than fresh ONP/OMG. Theging of ONP resulted in an increase in bound ink by almost0 ppm with a brightness loss of about 1.7 ISO points on theeinked pulp after hyperwash. The poorer deinking ability oflkaline deinking chemistry on aged ONP has been previouslyemonstrated at an industrial scale: a significant increase in totalnk measured on recycled paper processed during summer timeas observed from 27 North American recycling plants [21].his increase resulted in an average 1.4 ISO-point brightness

oss from these mills. The similar brightness loss obtained fromur artificial aged ONP is similar to this average value validatesur artificial aging method. Sulphite chemistry was also ineffec-

ive in removing aged ink on ONP/OMG (Figs. 6 and 7). Aginged to about 110 ppm and 70 ppm increase in respective total andound ink content with a brightness loss close to two points afteryperwash.

X. Zhang et al. / Enzyme and Microbial Technology 43 (2008) 103–108 107

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ig. 7. Comparing enzyme treatment with alkaline and sulphite chemistries fornk removal from aged ONP/OMG (standard deviations are within ±10 ppm forll the measurements based on eight replicates).

The causes of poor deinkability of aged ONP have beenreviously studied [22]. It is postulated that some of the inkomponents, such as oil vehicles and/or resin, can undergoxido-polymerization reactions at high temperatures to formthree-dimensional polymer network, resulting in increased

onding between the ink and the paper surface. The tighternteractions between ink and paper cause a higher degree ofnk fragmentation during repulping and higher undetachable inkontent. Several strategies have been attempted to combat thesummer effects”. In mill operations, shortening repulping timend increasing repulper temperature were shown to reduce inkragmentation [21]. Addition of antioxidants in offset inks waslso proposed as a preventive means to control the autoxida-ion of oil ink vehicles and enhance the deinkability of ensuingewsprint [23].

As mentioned, sulphite deinking is typically carried out atpH around 8 and temperature around 60 ◦C, which are sim-

lar to the optimal conditions for the cellulase enzymes usedn this study. This provides an opportunity to combine the twoeinking mechanisms and may provide some synergistic effectso enhance sulphite chemistry for summer ink removal. Cellu-ase B (EB) was chosen to supplement the sulphite deinkinghemistry and the effects of this combination on deinking freshnd aged ONP/OMG were studied. The addition of enzyme to

ulphite chemistry during fresh ONP/OMG deinking slightlyecreased the bound ink content from 95.3 to 89.5 with a simi-ar brightness value of 56 compared to 55.8 obtained by sulphiteeinking with enzyme. However, using a combination of sulphite

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able 1nk content and brightness value of aged ONP/OMG after enzymatic deinking (staneplicates)

ample Total ink content ERIC ppm Bound

0 h 2 h 0 h

ontrol 824 812 193ellulase A treated 856 853 203ellulase B treated 808 815 220ellulase C treated 837 838 211

ulphite chemistries for ink removal from aged ONP/OMG (standard deviationsre within ±10 ppm for all the measurements based on eight replicates).

nd enzyme to deink aged ONP/OMG produced a DIP with sig-ificantly lower total and bound ink content than obtained fromither sulphite or alkaline chemistry (Fig. 8). The lower totalnk content indicated that combined enzyme and sulphite deink-ng reduced ink fragmentation during the repulping. The boundnk content of combined enzyme- and sulphite-deinked pulpas approximately 30 ppm and 24 ppm lower than sulphite- and

lkaline-deinked pulps respectively, suggesting that the enzymeddition enhanced the removal of undetached ink. This was alsoonfirmed by the higher brightness value obtained by a combinednzyme sulphite recipe compared with sulphite chemistry alone.owever, this brightness value was still slightly lower than thatbtained by alkaline deinking where hydrogen peroxide wassed.

The residual enzyme activity in the combined recipe afterepulping was also determined to assess the effects of sulphitehemistry on the enzymes. Aliquots from the repulper beforend after pulping were taken for measuring enzyme activity. TheH of ONP/OMG stock after the addition of sulphite chemistryas around 8.6; therefore the enzyme activity was also deter-ined at pH 8.6. It was found that approximately 64% of the

ellulase activity remained after repulping at pH 8.6. This resultuggested that neither the repulping conditions nor the presencef sulphite had a significant negative effect on enzyme activity.

t seems possible that combining enzyme with sulphite deinkingan provide a potential effective strategy for neutral deinking ofged ONP.

dard deviations are within ±10 ppm for all the measurements based on eight

ink content ERIC ppm Brightness after hyperwash (% ISO)

2 h 0 h 2 h

192 53.1 52.3224 52.3 51.5204 52.9 51.7203 52.6 52.1

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. Conclusions

Adding cellulase enzymes at the repulping stage without pHdjustment was tested as a possible approach for enzymaticeinking. All three cellulase treatments showed some reductionn both total and bound ink content of ONP/OMG, when com-ared with the use of surfactant alone. The addition of Novozyme76 (EB) reduced ERIC content of hyperwashed pulp by 30 ppmnd increased final pulp brightness by 1% ISO point. It was veri-ed that the effect of cellulase enzyme on ink removal was likelyue to the catalytic action of the enzyme rather than the surfac-ant effects from enzyme protein. However, cellulase deinkingas less efficient than either alkaline or sulphite chemistry. Cel-

ulase treatment alone was not effective for deinking aged oldewsprint, but the combination of cellulase (Novozyme 476) andulphite deinking chemistry showed promise to improve summernk removal.

cknowledgements

The authors are thankful to Drs. Gilles Dorris, Luc Lapierre,homas Browne and Ms. Natalie Page for their advice and helpuring this work. We also thank Novozymes North America forroviding us with enzymes. The financial support from Papricanember Companies is greatly appreciated.

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