WO2003023133A1 - Blanchiment biologique de pate au moyen de laccase, de mediateur et d'agent de transfert de chaines - Google Patents

Blanchiment biologique de pate au moyen de laccase, de mediateur et d'agent de transfert de chaines Download PDF

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Publication number
WO2003023133A1
WO2003023133A1 PCT/US2002/013049 US0213049W WO03023133A1 WO 2003023133 A1 WO2003023133 A1 WO 2003023133A1 US 0213049 W US0213049 W US 0213049W WO 03023133 A1 WO03023133 A1 WO 03023133A1
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WO
WIPO (PCT)
Prior art keywords
enzyme
laccase
chain transfer
mediator
pulp
Prior art date
Application number
PCT/US2002/013049
Other languages
English (en)
Inventor
Huai N. Cheng
Simon Delagrave
Qu-Ming Gu
Dennis J. Murphy
Original Assignee
Cedix Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Cedix Inc. filed Critical Cedix Inc.
Publication of WO2003023133A1 publication Critical patent/WO2003023133A1/fr

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Classifications

    • D—TEXTILES; PAPER
    • D21—PAPER-MAKING; PRODUCTION OF CELLULOSE
    • D21C—PRODUCTION OF CELLULOSE BY REMOVING NON-CELLULOSE SUBSTANCES FROM CELLULOSE-CONTAINING MATERIALS; REGENERATION OF PULPING LIQUORS; APPARATUS THEREFOR
    • D21C9/00—After-treatment of cellulose pulp, e.g. of wood pulp, or cotton linters ; Treatment of dilute or dewatered pulp or process improvement taking place after obtaining the raw cellulosic material and not provided for elsewhere
    • D21C9/10—Bleaching ; Apparatus therefor
    • D21C9/1026—Other features in bleaching processes
    • D21C9/1036—Use of compounds accelerating or improving the efficiency of the processes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/0004—Oxidoreductases (1.)
    • C12N9/0055—Oxidoreductases (1.) acting on diphenols and related substances as donors (1.10)
    • C12N9/0057—Oxidoreductases (1.) acting on diphenols and related substances as donors (1.10) with oxygen as acceptor (1.10.3)
    • C12N9/0061—Laccase (1.10.3.2)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y110/00—Oxidoreductases acting on diphenols and related substances as donors (1.10)
    • C12Y110/03—Oxidoreductases acting on diphenols and related substances as donors (1.10) with an oxygen as acceptor (1.10.3)
    • C12Y110/03002—Laccase (1.10.3.2)
    • D—TEXTILES; PAPER
    • D21—PAPER-MAKING; PRODUCTION OF CELLULOSE
    • D21C—PRODUCTION OF CELLULOSE BY REMOVING NON-CELLULOSE SUBSTANCES FROM CELLULOSE-CONTAINING MATERIALS; REGENERATION OF PULPING LIQUORS; APPARATUS THEREFOR
    • D21C5/00—Other processes for obtaining cellulose, e.g. cooking cotton linters ; Processes characterised by the choice of cellulose-containing starting materials
    • D21C5/005—Treatment of cellulose-containing material with microorganisms or enzymes

Definitions

  • the present invention relates to the use of chain transfer agents that, in combination with laccase and a mediator, can be used to bleach pulp.
  • Paper pulp is typically processed from wood through the Kraft (and other) processes.
  • the process produces a pulp with a dark brown color, mostly due to the presence of lignin and lignin derivatives.
  • the lignin has to be removed by a process known in the art as "bleaching.” This is typically done commercially in several stages in pulp mills, wherein lignin is first oxidized and then removed from the pulp.
  • Bio-bleaching is a methodology whereby an enzyme is used to decrease the optical brightness and/or lignin content of the pulp or paper.
  • the standard measure of bleaching efficiency is "Kappa number.”
  • Enzymes that have most commonly been used include laccase, lignin peroxidase, and manganese peroxidase.
  • An enzyme group that has received particular attention is the laccase family of enzymes, which are copper-containing enzymes that are known to be good oxidizing agents in the presence of oxygen. Laccases are found in microbes, fungi, and higher organisms.
  • the oxidizing efficiency of a laccase can be improved through the use of a mediator that enhances the activity of the laccase.
  • Systems that include a laccase and a mediator are known in the art as laccase-mediator systems (LMS).
  • LMS laccase-mediator systems
  • mediators for use in laccase-mediator systems include HBT (1-hydroxybenzotriazole), ABTS [2,2'-azinobis(3-ethylbenzothiazoline-6-sulfmic acid)], NHA (N-hydroxyacetinilide), DKT 10114
  • NHAA N-acetyl-N-phenylhydroxylamine
  • HBTO 3-hydroxy-l,2,3-benzotriazin-4(3H)-one
  • VIO violuric acid
  • the invention provides the use of chain transfer agents in standard laccase-mediator systems (LMS). These agents themselves are not necessarily mediators that enhance the activity of the enzyme. However, the chain transfer agents have been found to enhance the bleaching ) efficiency of certain laccase/mediator systems.
  • LMS laccase-mediator systems
  • the invention provides a process of bleaching a lignin-containing material.
  • the process comprises the step of treating the material with an oxidative enzyme, a mediator that enhances the oxidative activity of the enzyme, and a chain transfer agent.
  • the i invention provides a process of oxidizing a substrate comprising treating the substrate with an oxidizing enzyme, a mediator that enhances the oxidative activity of the enzyme, and a chain transfer agent.
  • the invention also provides a composition comprising an oxidative enzyme, a mediator that enhances the oxidative activity of the enzyme, and a chain transfer agent.
  • the enzyme has laccase activity.
  • the enzyme is a laccase, a catechol oxidase, a monophenol monooxygenase, a bilirubin oxidase, or a mixture thereof.
  • the invention further comprises adding a hydrolase, such as xylanase.
  • the process of the invention can further include the addition of an oxidizing agent, such as air, oxygen, and/or hydrogen peroxide.
  • the present invention discloses the use of chain transfer agents to enhance the bleach efficiency of laccase/mediator systems. Laccase itself can bleach pulp only to a limited extent. Certain mediators can enhance the activity of laccase in pulp bleaching. It has been discovered DKT 10114
  • LMS can oxidize and degrade lignin and thereby achieve bleaching, it can also polymerize lignin under suitable reaction conditions. The oxidation, degradation, and polymerization reactions compete with one another. In this scenario, it is then possible to have an LMS that gives a high activity for the dye assay (predominant reaction being oxidation), but an increase in Kappa number (if polymerization is the major reaction).
  • Vinyl polymerization can be considered to consist of four reactions: (i) initiation; (ii) propagation; (iii) chain transfer; and (iv) termination. In all four reactions, free radicals are involved.
  • One common method to cut down the molecular weight of a polymer during polymer synthesis is to use chain transfer agents. These agents do not destroy the free radicals, but instead terminate a growing polymer chain, and start a new chain.
  • Chain transfer agents have not been used previously in combination with mediators. In fact, the role of chain transfer in bleaching has not been previously appreciated. It is noteworthy that a chain transfer agent does not generate free radicals by itself, nor does it trap free radicals. Chain agents are described, for example, by G. Scott in Chapter 4, Antioxidants: Radical Chain- Breaking Mechanism from “Atmospheric oxidation and antioxidants", Elsevier Publishing Co., New York, NY, p. 115.
  • the mediator of the invention is a hardwood black liquor or a softwood black liquor.
  • the chain transfer agent is anthracene, carbon tetrabromide, or a mixture thereof.
  • mediators and chain transfer agents of this invention are capable of enhancing the activities of laccase and laccase-related enzymes for the purpose of pulp bleaching.
  • these enzymes include laccase enzymes of the enzyme classification EC 1.10.3.2, catechol oxidase enzymes of the enzyme classification EC 1.10.3.1, the monophenol monooxygenase enzymes of the enzyme classification EC 1.14.99.1, and bilirubin oxidase enzymes of the enzyme classification EC 1.3.3.5.
  • the EC (Enzyme Commission) number is based upon the Nomenclature Committee of the International Union of Biochemistry
  • the laccase in this invention may be derived from microbial, fungal, or other sources. It may furthermore be produced by recombinant methods, such as cultivating a host cell transformed with a recombinant DNA vector which includes a DNA sequence encoding the I laccase (and DNA sequences encoding functions that permit the expression of laccase DNA sequence) in a culture medium under the conditions that permits the expression of the laccase, and recovering the enzyme from the culture.
  • Another aspect of the invention provides a process for oxidizing a substrate that comprises treating the substrate with a composition comprising an enzyme exhibiting laccase activity and an enzyme enhancing agent.
  • the enzyme enhancing agent can be selected from one or the above described enzyme enhancing agents.
  • the enhancing agent may be present in concentrations of from about 0.01 micromolar to about 1000 micromolar, more preferably from about 0.1 micromolar to about 250 micromolar and most preferably from about 0.5 to about 100 micromolar.
  • the chain transfer agent may be present in concentrations of from about 0.01 micromolar to about 1000 micromolar, more preferably from about 0.1 micromolar to about 250 micromolar and most preferably from about 0.5 to about 100 micromolar.
  • the enzyme is used in amounts of from about 0.1 to 400 units (defined in Examples using ABTS as substrate) for 1 g dry pulp, more preferably from 1 to 200 units and even more preferably from about 10 to 100 units and most preferably from 20 to 50 units.
  • the process of the invention can further include the step of adding an oxidizing agent, such as at least one of air, oxygen, and hydrogen peroxide.
  • an oxidizing agent such as at least one of air, oxygen, and hydrogen peroxide.
  • One embodiment of the invention provides a process for bleaching a lignin-containing material that comprises treating the material with an enzyme exhibiting laccase activity and an enzyme enhancing agent.
  • the enhancing agent may be present in an amount of from about 0.1% to about 15% based on the weight of the dry lignin containing material, more preferably from about 0.1 % to about 10% and even more preferably from about 0.5% to about 5% and most preferably from about 1% to about 4 %.
  • a lignin containing material is wood pulp.
  • the process for bleaching a lignin-containing material can further include the step of adding an oxidizing agent, such as at least one of air, oxygen, and hydrogen peroxide.
  • the specific activity was determined using ABTS (0.5 mM) as substrate.
  • One unit of activity is equal to the umol of the oxidized product from ABTS per min per mg protein at pH 6.0 at 23 °C.
  • the activity of laccase was determined using syringaldazine as substrate.
  • one unit of activity is equal to the change of 0.001 UV absorbance at A530nm per minute per ug protein in 2 ml of 100 mM, pH 5.5 potassium phosphate buffer, and 0.5 ml of 0.25 mM syringaldazine in methanol at 23 °C.
  • the dried pulp was added to 80 ml of 50 mM phosphate, pH 5.5, and disintegrated in a blender. The pulp was then transferred to a 500 ml conical flask and the blender was washed with 20 ml of the same buffer. The washed buffer and the pulp were combined. The mediator was added at 1-4%) (w/w, based on the dry pulp), followed by the addition of the laccase. The pH of the pulp mixture was adjusted to 5.5 if needed. The flask was covered with an aluminum foil with holes punched through and incubated at 50 °C for 16 hrs on a rotary shaker at 200 rpm.
  • the pulp mixture was filtered through a Buchner funnel, and the pulp was washed with water.
  • the pulp from the control experiment was treated at the same pH and temperature as described above.
  • the washed pulp was then treated with an alkaline solution under the following conditions:
  • the filtered pulp was repulped in the alkaline solution and incubated at 70 °C for 3 hrs.
  • the pH of the pulp mixture should be between 11.7-12.00 during the entire treatment.
  • the pulp mixture was filtered through a Buchner funnel, and the pulp was washed with water extensively and then dried in a hood overnight.
  • Example 1 Chicago Blue Assay Using Hardwood Black Liquor And Softwood Black Liquor As Laccase Mediators
  • Each of the compounds i.e., potential mediators
  • a phosphate buffer and a Chicago Blue solution were dissolved in water, or in ethanol if the potential mediator was not water soluble, and then mixed with a phosphate buffer and a Chicago Blue solution.
  • a solution of a laccase was added to make up 1 ml of the final solution, containing 20 uM mediator, 20 mM buffer at pH 5.5 or 7.0, 0.1-1% laccase (v/v) and Chicago Blue solution, with absorbance at A610 mn between 0.6 to 0.8.
  • A610 nm was measured immediately using a UV-VIS spectrophotometer (UV-1201, Shimadzu Scientific Instruments) after the enzyme was added. The decrease in absorbance was recorded at 30-second intervals for 5 minutes and was used to estimate the efficiency of the mediator.
  • the acid-treated black liquors were prepared using the following procedure: (1) adjust pH of 20 ml of the black liquors to 1.0 using cone. HCl; (2) incubate the acidified solution at 80C for 1 hr; (3) cool down to room temperature; (4) adjust the pH to neutral (6.5-7.5) using 5 M NaOH solution.
  • Acid-treated oak black liquor 100 (13%) 108 225

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  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Wood Science & Technology (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Zoology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Genetics & Genomics (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Biomedical Technology (AREA)
  • Biotechnology (AREA)
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  • Molecular Biology (AREA)
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Abstract

L'invention concerne un procédé d'oxydation d'un substrat. Ce procédé consiste à traiter le substrat à l'aide d'une enzyme oxydante, d'un médiateur qui augmente le pouvoir oxydant de l'enzyme, et d'un agent de transfert de chaînes. Dans une réalisation, le substrat est un matériau à base de lignine et le procédé conduit au blanchiment de matériau ligneux. Des exemples d'enzymes utiles comprennent laccase, catéchol oxydase, monophénol monooxygénase, et bilirubine oxydase. L'invention concerne aussi une composition comprenant une enzyme oxydante, un médiateur augmentant le pouvoir oxydant de l'enzyme et un agent de transfert de chaînes.
PCT/US2002/013049 2001-09-10 2002-04-24 Blanchiment biologique de pate au moyen de laccase, de mediateur et d'agent de transfert de chaines WO2003023133A1 (fr)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US31829201P 2001-09-10 2001-09-10
US60/318,292 2001-09-10
US10/027,149 US20030047295A1 (en) 2001-09-10 2001-12-20 Bio-bleaching of pulp using laccase, mediator, and chain transfer agent
US10/027,149 2001-12-20

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WO2003023133A1 true WO2003023133A1 (fr) 2003-03-20

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Families Citing this family (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20040112555A1 (en) * 2002-12-03 2004-06-17 Jeffrey Tolan Bleaching stage using xylanase with hydrogen peroxide, peracids, or a combination thereof
WO2009069143A2 (fr) * 2007-08-12 2009-06-04 Advanced Enzyme Technologies Ltd. Composition synergique et procédé de blanchiment biologique de pâte lignocellulosique
FI20085345L (fi) * 2008-04-22 2009-10-23 Kemira Oyj Menetelmä valon aikaansaaman ligniinipitoisen materiaalin kellertymisen vähentämiseksi
CN112301779A (zh) * 2020-10-28 2021-02-02 天津科技大学 一种漆酶辅助纤维素酶处理提高竹浆纤维柔软度的方法

Citations (2)

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Publication number Priority date Publication date Assignee Title
US6162260A (en) * 1999-05-24 2000-12-19 Novo Nordisk Biochem North America, Inc. Single-bath biopreparation and dyeing of textiles
US6242245B1 (en) * 1997-09-26 2001-06-05 Consortium für elektrochemische Industrie GmbH Multicomponent system for modifying, degrading or bleaching lignin or lignin-containing materials, and processes for its use

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
ES2122299T3 (es) * 1993-06-16 1998-12-16 Lignozym Gmbh Proceso para modificacion, degradacion o blanqueo de lignina, materiales que contienen lignina o carbon.
DK77393D0 (da) * 1993-06-29 1993-06-29 Novo Nordisk As Aktivering af enzymer

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6242245B1 (en) * 1997-09-26 2001-06-05 Consortium für elektrochemische Industrie GmbH Multicomponent system for modifying, degrading or bleaching lignin or lignin-containing materials, and processes for its use
US6162260A (en) * 1999-05-24 2000-12-19 Novo Nordisk Biochem North America, Inc. Single-bath biopreparation and dyeing of textiles

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