Classifications

• • D—TEXTILES; PAPER
  • D21—PAPER-MAKING; PRODUCTION OF CELLULOSE
  • D21B—FIBROUS RAW MATERIALS OR THEIR MECHANICAL TREATMENT
  • D21B1/00—Fibrous raw materials or their mechanical treatment
  • D21B1/02—Pretreatment of the raw materials by chemical or physical means
  • D21B1/021—Pretreatment of the raw materials by chemical or physical means by chemical means

Definitions

• the present invention relates to a process in accordance with the preamble of Claim 1 for preparing mechanical pulp.
• the invention also relates to a process for decreas- ing the energy consumption of mechanical pulping processes based on refining chips, according to the preamble of Claim 17.
• Chemical and mechanical pulps possess different chemical and fibre-technical properties and thus their use in different paper grades can be chosen according to these properties. Many paper grades contain both types of pulps in different proportions according to the desired properties of the final products.
• Mechanical pulp is used, when necessary, to improve or to increase the stiffness, bulkiness or optical properties of the product. In paper manufacture, the wood material must first be defibred.
• Mechanical pulp is mainly manufactured by means of grinding and refining methods, in which the raw wood material is subjected to periodical pressure impulses. Due to friction heat, the structure of the wood is softened and its structure loosened, finally leading to the separation of the fibres from one another (Virkola, 1983). However, only a small part of the energy brought into the system is used for separating the fibres; the major part being converted into heat. Therefore, the total energy economy of the defi- bration is very poor.
• the said treatment had no effect on the energy consumption of the refining treatment but it has been said to have an influence on the strength of paper or board.
• the energy in defibration is mainly absorbed by the amorphous part of the paper furnish only, i.e., hemicellulose and lignin
• an increase in the amorphousness of the raw material improves the energy economy of the defibration.
• the Patent Specifications WO 94/20666 and WO 94/20667 suggest that the amorphousness of the raw material be increased in connection with mechanical pulping by treating the raw material with a suitable enzyme that reacts with the crystalline, insoluble cellulose of the raw material.
• the Patent Specification WO 94/20666 suggests that the raw material be treated with an enzyme preparation, the main cellulase activity of which consists of cellobiohydrolase activity.
• the Patent Specification WO 94/20667 sug- gests that an enzyme preparation be used for the same purpose, containing cellobiohydrolase activity and mannanase activity.
• the examples of the said specifications deal with rough wood, such as the long-fibre fraction of fractioned TMP spruce pulp, once-refined TMP spruce pulps (with freeness values of CSF 450 - 550) or TMP pulps refined to different freeness levels (30 - 300).
• a synergistically acting cellulase enzyme product i.e., cellulase
• cellulase a synergistically acting cellulase enzyme product
• the treatment resulted in the hydrolysis of the insoluble cellulose and, thus, in the weakening of the strength properties of the pulp.
• the Patent Specification US 6,267,841 describes a manufacturing method of thermo-mechanical pulp, wherein the pulp is treated with enzyme between the first and the second refining processes. It also suggests the treatment of chips with enzymes before the first refining.
• the specification cites enzymes, such as pectinase, xylanase, laccase, cellulase or the mixtures thereof. The specification gives no numerical values of any energy savings obtained.
• the application of growing white rot fungi in the manufacture of mechanical pulp has also been studied.
• the enzymatic treatment is not as effective when applied to chips directly, because it is difficult to make the enzyme preparation to be effectively absorbed into the fibres of a raw material that is in the form of chips.
• the surface area of the raw wood material is not sufficient for an effective enzymatic treatment to take place.
• Another reason is that a major part of the capillaries of the wood are too small to receive any enzyme molecules (Grethlein, H.E. Biotechnology, February 1985, pp. 155 to 160).
• the pulping liquor is made to penetrate the chips used in pulp cooking by treating the chips with pressure shocks in the presence of the pulping liquor.
• the chips are treated in the presence of the cooking liquor by varying the pressure from a pressure of 4.5 kp/cm 2 and a treatment time of 10 - 16s to a pressure of 2 kp/cm 2 and a treatment time of 5 - 6s, the treatment being repeated 6 - 8 times at 1 -minute intervals /Rydholm, 1965).
• the Patent Specification WO 95/09267 suggests treating the chips, which are used in pulp cooking, with a chemical solution by subjecting the chips to a vacuum and making the chemical solution penetrate the wood fibres by means of a pressure shock.
• the chemical solution can be cooking liquor that contains, for example, catalysts and enzymes.
• the object of the invention is, thus, to be able to decrease the amount of lignin in order to diminish the need of decreasing the residue lignin at the final stage of cooking.
• the application does not describe in detail, whether or not the enzymes penetrate the wood cells successfully and whether or not the enzymes have any effect on the decrease of the amount of lignin.
• the US Patent 5,374,555 suggests the removal of lignin from the lignocellulose material by means of a protease enzyme.
• the patent suggests a mechanical treatment of chips, for example, in a screw clamp.
• the patent specification reminds that cellulase can be used as a pre-treatment enzyme for the chips or the pulp, but it does not suggest carrying out a treatment with cellulase in connection with the mechanical processes.
• the purpose of the patent is not to save energy but to remove lignin, and there are no observations relating to energy economy.
• the application suggests, among others, fungi from the genera Ceriposiophsis, Phanerochaete and Ophiostoma.
• enzymes lipolytic, proteolytic, linginolytic, cellulolytic and hemicellulolytic en- zymes are mentioned.
• the patent specification describes the absorption of the enzyme preparation Clariant Cartazyme HSTM (xylanase) into the compressed chips after releasing the pressure. Liquid was removed after the treatment, and mechanical pulp was prepared from the chips. In that case, the amount of energy consumed was 7.5% less than in the case of chips that were treated with a buffer only.
• the object of the invention is to provide a pre-treatment method of chips to be used before preparing the mechanical pulp.
• chips can be pre-treated with an enzyme preparation that has synergistically acting enzyme activities.
• the enzyme preparation does not need to contain any certain isolated enzyme activity only, but an enzyme preparation containing different enzyme activities can be used directly as the enzyme preparation.
• the treatment according to the method can be applied to the chips directly. As the enzymatic treatment takes place at an early stage of the pulping process, savings in the refining energy are then as high as possible.
• the chips are pre-treated with an enzyme that is capable of degrading the structural parts of the wood, after which mechanical pulp is manufactured from the chips by refining. It is preferable to carry out the enzymatic treatment by compressing the chips and by bringing the compressed chips in a liquid phase into contact with the enzyme composition to absorb the enzyme composition into the chips.
• the enzyme composition preferably contains both cellobiohydrolase and endoglucanase. It is particularly preferable for the composition to contain an effective amount of both cellobiohydrolase and endoglucanase.
• Enzyme preparations containing cellobiohydrolase and endoglucanase in a ratio of 20:1 - 1 :20, indicated as the weight ratio of the proteins, are preferable.
• the amount of endoglucanase compared with that of cellobiohydrolase is higher than what is naturally produced by the industrial production strains of cellulase, such as Tricho- derma, in their growth media.
• the method according to the invention is mainly characterized in that which is presented in the characterizing part of Claim 1.
• the method according to the invention is also characterized in that which is presented in the characterizing part of Claim 17.
• the invention provides several considerable advantages. When using the methods according to the preferable embodiments of the invention, considerably lower amounts of energy are consumed than in the methods according to prior art.
• the energy saving can be as much as 20% compared with a method, wherein the chips are not treated with the enzyme preparation.
• the strength of the pulp was not weakened; on the contrary, it im- proved to some extent.
• the optical properties also remained good. Thus, when treating the chips with the methods according to the preferred embodiments of the invention, it was possible to improve the quality of the pulp.
• the enzyme preparation is produced in a host organism, which excretes the enzyme preparation out of the cell, whereby the enzyme preparation does not need to be isolated from the cell. It is also especially advantageous to use a genetically modified organism as the production host, producing the desired enzyme preparation directly in the growth medium. This provides the considerable advantage that the used enzyme activity does not need to be isolated from the host organism or its growth medium but, for example, the growth medium of the host organism can be directly used.
• the enzymes that participate in the modification and degradation of cellulose are commonly called "cellulases”. These enzymes include endo- ⁇ -glucanases, cellobio- hydrolases and ⁇ -glucosidase. Countless organisms, such as various wood rotting fungi, moulds and anaerobic bacteria are able to produce some or all of these enzymes. Depending on the type of organism and cultivation conditions, these en- zymes are produced extracellularly in various ratios and amounts.
• enzyme preparation refers to any product that contains at least one enzyme or a structural part of the enzyme. Accordingly, the enzyme preparation can be, for example, a growth medium containing the enzyme ⁇ ), an isolated enzyme or a mixture of two or more enzymes. "Cellulase” or “cellulase enzyme preparation”, in turn, refers to an enzyme preparation containing at least one of the above-mentioned cellulase enzymes.
• the "enzyme composition” in this application means the same as the enzyme preparation.
• the enzyme preparation or the enzyme composition may also contain, for example, buffers, stabilizers, preservatives or other necessary additives.
• the "cellobiohydrolase activity" in this application refers to an activity that is capa- ble of modifying the crystalline parts of the cellulose.
• the cellobiohydrolase I and II activities refer to the main activities of the cellobiohydrolase produced by Trichoderma or to the corresponding activities produced by some other organism.
• the endoglucanase activity in this application refers to an activity capable of modifying the amorphous parts of the cellulose.
• the endoglucanase I and the endoglucanase II activities refer to the main activities of the endoglucanase produced by Trichoderma or to the corresponding activities produced by another organism.
• an enzyme preparation containing "an effective amount" of cellobiohydrolase and endoglucanase refers to an enzyme preparation, in which the effect of each enzyme on the chips can be measured as a reduction in the energy consumption of the refin- ing.
• the effective amount of cellobiohydrolase and endoglucanase provides a decrease of at least 3%, preferably at least 5%, more preferably at least 8%, most preferably at least 10% in the energy consumption of the refining.
• the methods according to the invention can be combined with treatments carried out with other enzymes, such as hemicellulases (e.g., xylanases, glucuronidases and mannanases) or esterases.
• Cellobiohydrolase and endoglucanase enzyme preparations are produced by grow- ing suitable micro-organism strains, known to produce cellulase.
• the strains are preferably production strains that are used industrially.
• the growth medium used can be, for example, a simple cellulosic substrate (1% Solka floe), which the necessary trace elements have been added to (Mandels and Weber, 1969).
• the production strains can be bacteria, fungi or moulds.
• the micro-organisms belong- ing to the following families can be mentioned:
• Trichoderma e.g. T. reesei
• Aspergillus e.g. A. niger
• Phanerochaete e.g. P. chrysosporium; Covert et al., 1992
• Penicillium e.g. P. janthinellum, P. digi- tatum
• Streptomyces e.g. S. olivochromogenes, S. flavogriseus
• Humicola e.g. H. insolens
• Bacillus e.g. B. subtilis, B. circulans, Ito et al., 1989).
• white rot fungi can also be used, strains belonging to families, such as Phlebia, Ceri- poriopsis and Trametes. It is also possible to produce cellobiohydrolases, endoglucanases or their structural parts by means of strains, which have been genetically improved to produce specifically these proteins, or by other genetically modified production strains, to which genes coding for these proteins have been transferred. When the genes of the de- sired protein have been cloned (Teeri et al., 1983), it is possible to produce the protein or its part in a desired host organism.
• the desired host may be the Trichoderma mould (EP 244 234, Mitsuishi et al., 1990), yeast (Penttila et al., 1988), some other mould, from families such as Aspergillus (van den Hondel et al., 1992), a bacterium or any other micro-organism, whose genetics are sufficiently well-known.
• the desired cellobiohydrolase and endoglucanase are produced by means of the mould strain Trichoderma, preferably the strain T. reesei.
• the said strain is a generally used production organism and its cellulases are fairly well known. T.
• CBH I and CBH II cellobiohydrolases
• EGI and EGII endoglucanases
• ⁇ -glucosidases Chen et al, 1992.
• Endoglucanases are typically active on soluble and amorphous substrates (CMC, HEC, ⁇ -glucan), whereas the cellobiohydrolases are able to hydrolyze crys- talline cellulose.
• the cellobiohydrolases act clearly synergistically on crystalline cellulose, but their hydrolysis mechanisms are supposed to be different from each other.
• the present knowledge of the hydrolysis mechanisms of cellulases is based on results obtained on pure cellulase preparations, and may not be valid in cases, where the substrate also contains other components, such as lignin or hemicellulose.
• the cellulases of T. reesei do not essentially differ from each other with respect to their optimal external conditions, such as pH or temperature. Instead, they differ from each other with respect to their ability to hydrolyze and modify cellulose in the raw wood material.
• the cellobiohydrolases I and II also differ to some extent from each other, and so do the endoglucanases I and II. In the preferred embodiments of this invention, however, it seems that the ratio of the cellobiohydrolases to the endoglucanases is more important than the interrelation between the various cellobiohydrolases or the various endoglucanases.
• Trichoderma reesei naturally produces various cellulase components in its growth medium, the amount and the interrelation of them depending on the production strain and the external conditions used.
• the following relative amounts of the main cellulases have been proposed: CBH I 60%, CBH II 20%, EG I 10% and EG II 10% (Stahlberg, 1991). In that case, the ratio of the cellobiohydrolases to the endoglucanases is about 4: 1.
• the preferred enzyme mixtures for the method according to the invention included those containing both cellobiohydrolase enzymes and endoglucanase enzymes. While not wanting to commit our to any theories, it very strongly seems that the method according to the invention needs both cellobiohydrolase enzymes and endoglucanase enzymes, because the endoglucanase is capable of preparing, in the chips, objects that the cellobiohydrolase is able to act on. As neither activity alone is able to provide the desired effect, the cellobiohydrolases and the endoglucanases must work in synergy.
• the ratio of the cellobiohydrolases to the endoglucanases is preferably 20: 1 - 1 :20, more preferably 9: 1 - 1 :9, more preferably 5: 1 - 1 :5, and more preferably 3:1 - 1 :3, most preferably 2: 1 - 1 :2, and even more preferably about 1 :1.
• the most preferable cellulase compositions are those, wherein the weight ratio of the cellobiohydrolases and the endoglucanases is close to 1 :1.
• an energy saving effect can even be provided by a weight ratio deviating from this, if the endoglucanase used has a very strong activity so that even a small amount is sufficient to provide the desired effect.
• the preferred embodiments of the invention also include an enzyme preparation, wherein the portion of endoglucanases in the preparation is 2 - 60% by weight. Even more preferred is a preparation, wherein the portion of endoglucanases in the preparation is 20 - 55% by weight and the most preferred is one, wherein the por- tion of endoglucanases is 45 - 50% by weight.
• an amount of endoglucanases can be reached by increasing the amount of either EGI or EGII, or both, in the preparation. If the amount of EGI is increased exclusively, the amount of EGI in the preparation should reach a level of 15 - 45% by weight. This is also true, if only the amount of EGII is increased.
• 5,874,293 describes an enzyme preparation that is produced by the strain Trichoderma (ALKO 3529) that overproduces EGII.
• the ratio of CBH: EG in the growth medium produced by the strain is estimated to be 1 - 1.4:1. It would be advantageous to use the growth medium produced by such a strain, for example, in the present invention.
• the publication Karhunen et al. (1993) describes a Trichoderma host that is modified to overproduce the EGI enzyme. The growth medium of this host could also be used in the present invention.
• preferable enzyme mixtures include those, wherein the amount of endoglucanase is higher than what the cellulase-producing micro-organisms, such as Trichoderma, especially T. reesei, would naturally produce in their growth media.
• the modified cellulase preparation herein refers to a preparation, wherein the ratio of CBH and EG components has been changed by methods that are well-known to average experts. Such methods include, e.g., the genetic modification of a host organism so that the host organism produces a novel cellulase compound in its growth medium. Other viable methods of manufacturing modified cellulase preparations include the fractioning of a cellulase-containing growth medium or combining dif- ferent cellulase mixtures.
• the host organism can be modified genetically to produce the desired cellobiohydrolases and endoglucanases in a desired proportion.
• the genetic modification of the strains of the family Trichoderma can be carried out by the methods described in the patent EP 244234 or in the publication Suominen et al., 1993.
• Preferable enzyme preparations to be used in the embodiments of this invention include those, wherein the mould T. reesei is modified to overproduce EG I and/or EG II enzymes.
• the overproduction host may also have been modified so as to produce less of some cellobiohydrolase activities, especially the CBH I or CBH II activities, if any, or to produce less endoglucanases, if any, especially the EG I and/or EG II activities.
• cellobiohydrolase activities especially the CBH I or CBH II activities, if any, or to produce less endoglucanases, if any, especially the EG I and/or EG II activities.
• there should be cellobiohydrolase activities therefore, adding endoglucanase activities to the enzyme preparations is more advantageous than decreasing cellobiohydrolase activities or removing the endoglucanases.
• Corresponding enzyme preparations can also be manufactured by purifying suitable cellobiohydrolase and endoglucanase enzymes and combining the same in advantageous proportions, or by adding to an enzyme preparation, which is produced by a non-modified host, the desired enzyme activities, for example, the EGI and EGII activities.
• the strains, which are capable of ove ⁇ roducing EGI and EGII enzymes, can be constructed, for example, by transferring genes coding for these enzymes (egll Penttila et al. 1986 and egl2 Saloheimo et al.
• Trichoderma host as several copies or to replace some genes of Trichoderma, such as the cbhl and cbhl genes that code for cellobiohydrolases, as described in the publication Suomi- nen et al. (1993).
• the said genes can be expressed under a strong cbhl promoter, as described in the publication Paloheimo et al. (1993).
• the T. reesei strain QM6a for example, can be used as a host, especially the strains QM9414 and Rut C - 30, which are developed from the same for the production of cellulase, or strains developed from them, which produce less protease.
• the enzyme preparation is manufactured by means of a host organism, which is modified to produce cellobiohydrolases and endoglucanases in a desired proportion in its growth medium.
• a host organism which is modified to produce cellobiohydrolases and endoglucanases in a desired proportion in its growth medium.
• endoglucanase I and/or II enzymes are added to a growth medium, which is produced by a non-modified host organism that naturally pro- prises cellulases in its growth medium, the enzymes having either been produced by a micro-organism that is modified to ove ⁇ roduce these enzymes, or isolated and possibly purified from the growth medium.
• the above-mentioned methods can also be combined.
• the cellobiohydrolase and the endoglucanase can be separated from the growth medium of the production host by means of several known methods.
• various purifying techniques are combined, such as precipitation, ion exchange chromatographic and affinity chromatographic as well as gel chromatographic methods.
• the enzyme preparations can be manufactured by means of the mould Trichoderma or some other production host. Genes that code for cellobiohydrolase and endoglucanase can originate in Trichoderma or some other host that produces the preferable cellobiohydrolase and endoglucanase activities; and the said activities in the enzyme preparation can be from the same or a different origin.
• the treatment according to the present invention is applied to chips.
• the raw wood material is chipped in a normal manner so that the chip length is about 15 - 25 mm.
• the chips can be graded by removing oversize and too thick chips and fines.
• the chip material is typically compressed by at least 10%, generally 10 - 30% of its original bulk volume.
• the chips are com- presses in a ratio of 1:2 - 1:10.
• a ratio of compression of at least 1:4 is preferably used.
• the compression treatment is preferably carried out by a method, wherein the chips are compressed without a considerable circular motion, because the object is not to crush the pieces of chips but make microscopic cracks in the raw wood material.
• the compression can be implemented by various means, e.g., in a screw clamp or by a hydraulic press.
• the impregnated chips are treated for a sufficient time in conditions favourable for the activity of the enzyme, after which the chips are processed in a normal manner before refining, including pre-heating with steam before feeding them into the refiner.
• the method according to the invention is not limited to a certain raw wood material but can generally be applied to both softwood and hardwood, such as the species of the Pinaceae order (e.g., the Picea and Pinus families), the species of the Salica- ceae order (e.g., the Populus family) and the species in the Betula family.
• the compressed chips are brought into contact with the enzyme preparation in a liquid phase. This is best carried out so that the chips are compressed in an enzyme solution.
• the proportion of the liquid and the chips is preferably selected so that the liquid is able to effectively act on the chips.
• the proportion of liquid to the chips can be 10: 1 - 2:1 , and it is preferably 6 - 8:1.
• the compression pressure can be 10 - 20 MPa, and it is preferably 12 - 15 MPa.
• the duration of the compres- sion/abso ⁇ tion stage should be at least 1 min; the duration is preferably 5 - 100 min and generally 10 - 30 min. After releasing the compression, the chips are al- lowed to retum to their original volume under the enzyme solution, whereby the enzyme solution is impregnated into the chips.
• the pH and the temperature of the enzyme solution should be suitable for the functioning of the enzyme preparation.
• the pH should preferably be within a range of pH 3 - 10, preferably pH 4 - 8, and the temperature should be 20 - 55°C, preferably 30 - 45°C.
• the chips are treated for a sufficiently long time in the conditions mentioned above.
• the treatment time greatly varies depending on the properties (size, thickness) of the chips, sort of wood, compression treatment, enzyme preparation, operational condi- tions etc., and a suitable treatment time must be specified for each case separately. In terms of costs, as short a time as possible is advantageous but in terms of process technology, there are no obstacles for a treatment of several hours.
• the treatment time can be within a range of 1 - 24h, preferably 1 - 12h.
• the amount of enzyme preparation used in the invention in the treatment of chips is selected so that the amount of free sugars released in the solution is preferably about 0.1 - 1.0% of the original dry matter.
• a suitable dosage, determined as total protein, is 0.1 - 7mg of protein per g of chips, preferably 3 - 6mg of protein per g of chips (as dry matter).
• mechanical pulp is manufactured by refining chips that are treated with an enzyme to obtain a drainability value, which is preferably at least
• the method according to the invention yielded energy savings of 13%, preferably 15% and most preferably as much as 20%. It seems that the enzymatic treatment according to the present invention is advantageous, when combined with the manufacture of mechanical pulp by the refining method, in particular, and when refining the pulp into drainability of 100 CSF or lower.
• the invention provides considerable advantages. Accordingly, it can be used to considerably reduce the specific energy consumption of refining; in accordance with the preferred embodiments of the invention, as much as 20% lower energy consumption can be achieved than with untreated source materials, as the examples below indicate.
• a suitable enzyme preparation the properties of the mass can also be improved. Using the solutions according to the preferred embodiments of the invention, a high yield is obtained in the manufacture of mechanical pulp by refining, the quality of the pulp is good, the strengths are maintained, the optical properties are good, and the method is easy to connect to the present processes.
• thermo-mechanical pulp TMP
• RMP refined mechanical pulp
• Enzymatic treatments with a cellulase mixture were carried out on sorted spmce sapwood chips ( ⁇ 7mm), using an enzyme dosage of 6.3mg of protein per g of chips (as dry matter), [ ⁇ u][KL2]a commercial enzyme preparation produced by the Trichoderma strain, wherein the weight proportions of CBH:EG were defined as 1 :1.
• a compression treatment was exerted on the chips using a PREX hydraulic press. In the hydraulic compression, a chip lot (200g) was compressed in the enzyme solution into a volume that was about 20% smaller than the original, using a compression load of 48t (14 Mpa).
• the ratio of liquid to wood was 11:4 and the duration of the compression/abso ⁇ tion stage was lOmin.
• the chips were allowed to return to their original volume under the enzyme solution, whereby the enzyme solution was impregnated into the chips.
• an otherwise similar treatment was used, but without the enzyme.
• the compression treatment neither visual nor microscopic changes were perceived in the chips.
• the chips (+ the compression solution) were transferred into a rotary air oven for further processing. The treatment was carried out in atmospheric pressure and at a temperature of 45°C.
• the amount of carbohydrates released in the treatment solution, as reducing sugars, was defined after 6 and 22 h.
• the result obtained was compared with a treatment, wherein the compression treatment of chips was omitted. The results are shown in Table 1.
• the effects of the combined compression/abso ⁇ tion and enzymatic treatments on the beatability of the chips were examined by means of a blade refiner.
• the equipment used in the tests contained the actual refiner and an accurate energy measuring system connected thereto.
• the refiner chamber of the blade refiner consisted of a cylinder provided with counter blades (20 in number) and a rotary rotor having four wing-like blades.
• Several refining operations (125g dry matter per refining) were carried out for each specific energy consumption curve (SEC) by varying the refining time (3 - 12min) and, thus, also the energy level of the refining.
• the total energy consumption of the refining was obtained from a watt-hour meter by means of a cumulative pulse counter.
• the energy consumption value obtained per amount of defibred chips was corrected by a zero load.
• the defibration times for the treated spruce sapwood chips were 3 - 12min.
• the treatments were carried out as described in Example 3 (45°C, 22h).
• the compres- sion/abso ⁇ tion treatments were carried out using a treated cellulase mixture, as in Example 1 , CBH I, and an EG-rich enzyme preparation.
• the dosages for the treated mixture were 0.63 and 6.3 of protein per g of chips (as dry matter).
• the dosage for CBH I and the EG-rich enzymes was 5.0mg of protein per g of chips (dry matter).
• Example 3 Effect of the enzymatic treatment on the sheet properties of the pulp
• the chips were impregnated and treated with a cellulase mixture (a dosage containing 0.63mg of protein per g of chips (dry matter), 45°C, 22h), as described in Example 1. After this, the chips were refined by the blade refiner in accordance with Example 2. Laboratory sheets were prepared from the refined pulps and tested in accordance with SCAN methods. The sheet properties are shown in Table 3. Table 3.
• the cellulase treatment which was applied to the chips, improved the strength properties of the pulp; the tensile strength and the z-strength (Scott Bond) in particular. Also the optical properties were well-preserved.

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