Skip to content
Licensed Unlicensed Requires Authentication Published by De Gruyter August 15, 2013

Enzymatic strategies to improve removal of hexenuronic acids and lignin from cellulosic fibers

  • Cristina Valls EMAIL logo , Teresa Vidal and M. Blanca Roncero
From the journal Holzforschung


Different enzymatic strategies were applied to improve lignin and hexenuronic acid (HexA) removal. Three laccases (L) with different redox potentials were applied in combination with 1-hydroxybenzotriazole or methyl syringate to softwood sulfite and hardwood kraft fibers. The enzymes with a high-redox potential from Pycnoporus cinnabarinus and Trametes villosa were found to be the most efficient. The bleaching efficiency was not significantly influenced by the presence or absence of HexA in the different types of lignocellulosic fibers. Subsequently, the lignin or HexA removal in the presence of different mediators was evaluated in eucalyptus fibers with the T. villosa laccase (TvL). Natural mediators removed only lignin, whereas the combinations of TvL with synthetic mediators removed both HexA and lignin. The mediator violuric acid (VA) was the most efficient as judged by the properties of fiber and effluent. A xylanase pretreatment stage was found to boost the access of the LVA system to HexA without affecting lignin in the pulp.

Corresponding author: Cristina Valls, Textile and Paper Engineering Department, Universitat Politècnica de Catalunya (UPC), BarcelonaTech, Colom 11, E-08222 Terrassa, Spain, Phone: +34 937398210, Fax: +34 937398101, e-mail:

This work was funded by the Spanish MICINN BIOSURFACEL- CTQ2012-34109 and BIOFIBRECELL-CTQ2010-20238-CO3-01 projects, by the European Union BIORENEW-NMP2-CT-2006-026456 project, and by the AGAUR 2009SGR 00327. Torraspapel S.A. (Spain) and Domsjö Fabriker AB (Sweden) are gratefully acknowledged for supplying the pulps used. Novozymes (Bagsvaerd, Denmark) and INRA (Marseille, France) are gratefully acknowledged for supplying the enzymes.


Andreu, G., Vidal, T. (2011) Effects of laccase-natural mediator systems on kenaf pulp. Bioresour. Technol. 102:5932–5937.Search in Google Scholar

Andreu, G., Barneto, A.G., Vidal, T. (2013) A new biobleaching sequence for kenaf pulp: influence of the chemical nature of the mediator and thermogravimetric analysis of the pulp. Bioresour. Technol. 130:431–438.10.1016/j.biortech.2012.12.014Search in Google Scholar

Aracri, E., Vidal, T. (2011) Xylanase- and laccase-aided hexenuronic acids and lignin removal from specialty sisal fibers. Carbohydr. Polym. 83:1355–1362.Search in Google Scholar

Aracri, E., Colom, J.F., Vidal, T. (2009) Application of laccase-natural mediator systems to sisal pulp: An effective approach to biobleaching or functionalizing pulp fibers? Bioresour. Technol. 100:5911–5916.10.1016/j.biortech.2009.06.016Search in Google Scholar

Aracri, E., Fillat, A., Colom, J.F., Gutiérrez, A., del Río, J.C., Martínez, A.T., Vidal, T. (2010) Enzymatic grafting of simple phenols on flax and sisal pulp fibers using laccases. Bioresour. Technol. 101:8211–8216.Search in Google Scholar

Aracri, E., Tzanov, T., Vidal, T. (2013) Use of cyclic voltammetry as an effective tool for selecting efficient enhancers for oxidative bioprocesses: importance of pH. Ind. Eng. Chem. Res. 52:1455–1463.10.1021/ie3027586Search in Google Scholar

Babot, E.D., Rico, A., Rencoret, J., Kalum, L., Lund, H., Romero, J., del Río, J.C., Martínez, A.T., Gutiérrez, A. (2011) Towards industrially-feasible delignification and pitch removal by treating paper pulp with Myceliophthora thermophila laccase and a phenolic mediator. Bioresour. Technol. 102:6717–6722.Search in Google Scholar

Borges, M.T., Silva, C.M., Colodette, J.L., De Oliveira, R.C., Tesser, F. (2013) Enzymatic bleaching of eucalyptus kraft pulp: effects on the prebleaching filtrate, pulp quality and paper properties. Holzforschung 67:147–153.10.1515/hf-2011-0130Search in Google Scholar

Buchert, J., Tenkanen, M., Kantelinen, A., Viikari, L. (1994) Application of xylanase in the pulp and paper industry. Bioresour. Technol. 50:65–72.Search in Google Scholar

Buchert, J., Teleman, A., Harjunpaa, V., Tenkanen, M., Viikari, L., Vuorinen, T. (1995) Effect of cooking and bleaching on the structure of xylan conventional pine kraft pulp. Tappi J. 78:125–130.Search in Google Scholar

Cadena, E.M., Vidal, T., Torres, A.L. (2010) Influence of the hexenuronic acid content on refining and ageing in eucalyptus TCF pulp. Bioresour. Technol. 101:3554–3560.Search in Google Scholar

Cadena, E.M., Du, X., Gellerstedt, G., Li, J., Fillat, A., García-Ubasart, J., Vidal, T., Colom, J.F. (2011) On hexenuronic acid (HexA) removal and mediator coupling to pulp fiber in the laccase/mediator treatment. Bioresour. Technol. 102:3911–3917.Search in Google Scholar

Call, H.P., Muecke, I. (1997) History, overview and applications of mediated lignolytic systems, especially laccase-mediator systems (Lignozym-process). J. Biotechnol. 53:163–202.10.1016/S0168-1656(97)01683-0Search in Google Scholar

Chakar, F.S., Ragauskas, A.J. (2000) The Kismet of residual lignins during LMS delignification of high-kappa kraft pulps. Holzforschung 54:647–653.10.1515/HF.2000.109Search in Google Scholar

Chandra, R.P., Ragauskas, A.J. (2005) Modification of high-lignin kraft pulps with laccase. Part 2. Xylanase-enhanced strength benefits. Biotechnol. Prog. 21:1302–1306.Search in Google Scholar

Cusola, O., Valls, C., Vidal, T., Roncero, M.B. (2013) Application of surface enzyme treatments using laccase and a hydrophobic compound to paper-based media. Bioresour. Technol. 131:521–526.Search in Google Scholar

Eugenio, M.E., Santos, S.M., Carbajo, J.M., Martín, J.A., Martín-Sampedro, R., González, A.E., Villar, J.C. (2010) Kraft pulp biobleaching using an extracellular enzymatic fluid produced by Pycnoporus sanguineus. Bioresour. Technol. 101:1866–1870.Search in Google Scholar

Fillat, U., Roncero, M.B. (2010) Bioblanqueo de lino mediante el sistema lacasa mediador a presión atmosférica. Afinidad 67:254–261.Search in Google Scholar

Fillat, U., Roncero, M.B., Bassa, A., Sacón, V.M. (2010a) An approach to industrial application: influence of black liquor and pH on xylanase efficiency in bleaching of eucalyptus kraft pulp. Ind. Eng. Chem. Res. 49:11200–11205.10.1021/ie1014469Search in Google Scholar

Fillat, A., Colom, J.F., Vidal, T. (2010b) A new approach to the biobleaching of flax pulp with laccase using natural mediators. Bioresour. Technol. 101:4104–4110.10.1016/j.biortech.2010.01.057Search in Google Scholar PubMed

Fillat, A., Roncero, M.B., Vidal, T. (2011) Assessing the use of xylanase and laccases in biobleaching stages of a TCF sequence for flax pulp. J. Chem. Technol. Biotechnol. 86:1501–1507.Search in Google Scholar

Fillat, U., Prieto, A., Camarero, S., Martínez, A.T., Martínez, M.J. (2012) Biodeinking of flexographic inks by fungal laccases using synthetic and natural mediators. Biochem. Eng. J. 67:97–103.Search in Google Scholar

Garcia, O., Camarero, S., Colom, J.F., Martínez, A.T., Martínez, M.J., Monje, R., Vidal, T. (2003) Optimization of a laccase-mediator stage for TCF bleaching of flax pulp. Holzforschung 57:513–519.10.1515/HF.2003.076Search in Google Scholar

Garcia-Ubasart, J., Esteban, A., Vila, C., Roncero, M.B., Colom, J.F., Vidal, T. (2011) Enzymatic treatments of pulp using laccase and hydrophobic compounds. Bioresour. Technol. 102:2799–2803.Search in Google Scholar

González Arzola, K., Arévalo, M.C., Falcón, M.A. (2009) Catalytic efficiency of natural and synthetic compounds used as laccase-mediators in oxidising veratryl alcohol and a kraft lignin, estimated by electrochemical analysis. Electrochim. Acta 54:2621–2629.10.1016/j.electacta.2008.10.059Search in Google Scholar

Ibarra, D., Romero, J., Martinez, M.J., Martinez, A.T., Camarero, S. (2006) Exploring the enzymatic parameters for optimal delignification of eucalypt pulp by laccase-mediator. Enzyme Microb. Technol. 39:1319–1327.Search in Google Scholar

Ibarra, D., Köpcke, V., Ek, M. (2010) Behavior of different monocomponent endoglucanases on the accessibility and reactivity of dissolving-grade pulps for viscose process. Enzyme Microb. Technol. 47:355–362.Search in Google Scholar

Magaton, A.S., Silva, T.C.F., Colodette, J.L., Piló-Veloso, D., Milagres, F.R., Resende, J.O. (2013) Behavior of xylans from the Eucalyptus species. Part 2. Characterization of 4-O-methylglucuronoxylans isolated from black liquors of kraft pulping of Eucalyptus grandis and E. urophylla. Holzforschung 67:123–128.10.1515/hf-2012-0035Search in Google Scholar

Martínez, Á.T., Ruiz-Dueñas, F.J., Martínez, M.J., del Río, J.C., Gutiérrez, A. (2009) Enzymatic delignification of plant cell wall: from nature to mill. Curr. Opin. Biotechnol. 20:348–357.Search in Google Scholar

Moldes, D., Díaz, M., Tzanov, T., Vidal, T. (2008) Comparative study of the efficiency of synthetic and natural mediators in laccase-assisted bleaching of eucalyptus kraft pulp. Bioresour. Technol. 99:7959–7965.Search in Google Scholar

Monje, P.G., González-García, S., Moldes, D., Vidal, T., Romero, J., Moreira, M.T., Feijoo, G. (2010) Biodegradability of kraft mill TCF biobleaching effluents: application of enzymatic laccase-mediator system. Water Res. 44:2211–2220.10.1016/j.watres.2009.12.047Search in Google Scholar PubMed

Nousiainen, P., Maijala, P., Hatakka, A., Martínez, A.T., Sipilä, J. (2009) Syringyl-type simple plant phenolics as mediating oxidants in laccase catalyzed degradation of lignocellulosic materials: model compound studies. Holzforschung 63:699–704.Search in Google Scholar

Oudia, A., Queiroz, J., Simões, R. (2008) The influence of operating parameters on the biodelignification of Eucalyptus globulus kraft pulps in a laccase-violuric acid system. Appl. Biochem. Biotechnol. 149:23–32.Search in Google Scholar

Roncero, M.B., Colom, J.F., Vidal, T. (2003) Influence of the xylanase enzymatic treatments on the carbohydrate composition of pulp for paper manufacture. Afinidad 60:8–15.Search in Google Scholar

Shatalov, A.A., Pereira, H. (2007) Xylanase pre-treatment of giant reed organosolv pulps: direct bleaching effect and bleach boosting. Ind. Crops Prod. 25:248–256.10.1016/j.indcrop.2006.12.002Search in Google Scholar

Sigoillot, C., Camarero, S., Vidal, T., Record, E., Asther, M., Perez-Boada, M., Martínez, M.J., Sigoillot, J.C., Asther, M., Colom, J.F., Martínez, A.T. (2005) Comparison of different fungal enzymes for bleaching high-quality paper pulps. J. Biotechnol. 115:333–343.10.1016/j.jbiotec.2004.09.006Search in Google Scholar PubMed

Teleman, A., Harjunpaa, V., Tenkanen, M., Buchert, J., Hausalo, T., Drakenberg, T., Vuorinen, T. (1995) Characterization of 4-deoxy-beta-L-threo-hex-4-enopyranosyluronic acid attached to xylan in pine kraft pulp and pulping liquor by H-1 and C-13 NMR-spectroscopy. Carbohydr. Res. 272:55–71.Search in Google Scholar

Valls, C., Roncero, M.B. (2013) Antioxidant property of TCF pulp with a high hexenuronic acid (HexA) content. Holzforschung 67:257–263.10.1515/hf-2012-0114Search in Google Scholar

Valls, C., Colom, J.F., Baffert, C., Gimbert, I., Roncero, M.B., Sigoillot, J.C. (2010a) Comparing the efficiency of the laccase-NHA and laccase-HBT systems in eucalyptus pulp bleaching. Biochem. Eng. J. 49:401–407.10.1016/j.bej.2010.02.002Search in Google Scholar

Valls, C., Vidal, T., Roncero, M.B. (2010b) Boosting the effect of a laccase-mediator system by using a xylanase stage in pulp bleaching. J. Hazard. Mater. 177:586–592.10.1016/j.jhazmat.2009.12.073Search in Google Scholar PubMed

Valls, C., Vidal, T., Roncero, M.B. (2010c) The role of xylanases and laccases on hexenuronic acid and lignin removal. Proc. Biochem. 45:425–430.10.1016/j.procbio.2009.10.015Search in Google Scholar

Valls, C., Quintana, E., Roncero, M.B. (2012) Assessing the environmental impact of biobleaching: Effects of the operational conditions. Bioresour. Technol. 104:557–564.10.1016/j.biortech.2011.10.044Search in Google Scholar PubMed

Valls, C., Cadena, E.M., Roncero, M.B. (2013) Obtaining biobleached eucalyptus cellulose fibers by using various enzyme combinations. Carbohydr. Polym. 92:276–282.Search in Google Scholar

Viikari, L., Ranua, M., Kantelinen, A., Linko, M., Sundquist, J. Bleaching with enzymes. ICBPPI. Proc. 3rd Int. Conf., 1986:67–69.Search in Google Scholar

Xu, F., Kulys, J.J., Duke, K., Li, K., Krikstopaitis, K., Deussen, H.J.W., Abbate, E., Galinyte, V., Schneider, P. (2000) Redox chemistry in laccase-catalysed oxidation of N-hydroxy compounds. Appl. Environ. Microbiol. 66:2052–2056.Search in Google Scholar

Received: 2013-3-4
Accepted: 2013-7-16
Published Online: 2013-08-15
Published in Print: 2014-02-01

©2014 by Walter de Gruyter Berlin Boston

Downloaded on 10.12.2023 from
Scroll to top button