ZA200105351B

ZA200105351B - Enzymatic composition for refining wood.

Info

Publication number: ZA200105351B
Application number: ZA200105351A
Authority: ZA
Inventors: Olivier Henry
Original assignee: Robert Stiernon S A N V Ets
Priority date: 1998-12-31
Filing date: 2001-06-28
Publication date: 2002-06-28
Also published as: EP1020264A1

Description

Co 20015351 . The present invention relates to an enzymatic composi- tion and an aqueous bath for seasoning or maturing green wood, and also to a method using this enzymatic composition or this bath.

INTRODUCTION

The use of wood as a material in construction, carpentry or cabinet-making presumes prior drying in order to dimensionally stabilize this wood. For cooperage, there are also organoleptic requirements.

For most industrial applications, this drying takes place in a drying facility at controlled temperature and humidity.

For other applications, in particular when the wood will subsequently be in contact with foods, the drying takes place naturally, outside, over quite long periods of time, with the aim of eliminating certain undesirable compounds. The duration of this natural drying, or ageing, can be varied depending on the final use. This drying can take up to 2 years in order to obtain good organoleptic qualities.

In the case of natural drying, the wood is colonized by a fungal flora which can transform certain compounds of the wood. These transformations of the wood cannot, however, be controlled. Naturally dried wood exhibits great variability of quality.

On the other hand, with artificial drying 1n a drying facility, these transformations do not occur and the : wood thus dried is not suitable for food applications since this drying does not make it possible to eliminate certain undesirable molecules (such as toxic or bitter molecules). -

STATE OF THE ART

EP 0 001 540 A relates to the use of microorganisms Or of enzymes (pectinase or cellulase) with the aim of modifying the structure of wood intended for decoration, by controlling the temperature,. the water content of the wood, and the 0; ‘and CO, content in the wood’s environment. In this method, the wood 1s pretreated before inoculation, post treated in order to terminate the microbiological process, and then dried.

This complex method does not use immersion of the wood in an enzymatic preparation. :

DD 292 864 A relates to improving the impregnability of the sapwood of resinous wood using an enzymatic . preparation (pectinase, cellulase and hemicellulase), at the same time as an aqueous mineral preparation intended to superficially protect this wood.

This method does not relate to the duramen of the wood, or to species other than "conifers, or to a food application.

FR 2 421 039 A relates to the conditioning of wood in a solid medium, using microorganisms, with a view to facilitating the fragmentation and then the agglomera- tion of the elements of the wood.

CE 20015351

This method does not use immersion of the wood or an enzymatic preparation, and does not relate to applica- tion in the food industry.

FR 2 705 607 A relates to the use of microorganisms “having o-amylase ‘and PB-glucosidase activities, with a view to improving the organoleptic qualities of oak : intended for cooperage. The microbiological process is carried out in the solid state using a fungal leaven.

The process, which is supposed to reproduce the effects of natural drying/seasoning, specifies neither the duration of bringing the microorganisms into contact with the wood, nor the temperature, nor the micro- organism content. It does not describe a method of "15 immersion of the wood, nor an enzymatic preparation, and it applies only to species of oak.

DESCRIPTION OF THE INVENTION

The aim of the present invention is to provide a method for seasoning wood which allows, once it has been used, : notably shortened drying of the wood. This method is : suitable for food applications and conserves the : . dimensions of treated. wood. : oC :

A subject of the invention is an enzymatic composition for drying wood, comprising mainly enzymes of the cellulase class, and also an aqueous bath for seasoning wood, comprising an endocellulase activity of between 1 x 10° and 100 x 10° ECU (preferably between 3 x 10° and 30 x 10° ECU) per m®> of wood to be treated, xylanase activity of between 0.6 x 10° and 60 x 10° BXU (preferably between 2 x 10° and 20 x 10° BXU) per m’ of - wood to be treated, a P-mannanase activity of between 0.4 x 10° and 40 x 10° MNU (preferably between 1 x 10°

RM 20015351 and 10 x 10° MNU) per m® of wood to be treated and an a- amylase activity of between 1 x 10° and 100 x 10° TU (preferably between 3 x 10° and 30 x 10° ,TU) per m' of wood to be treated.

A subject of the invention is ‘also a method for seasoning wood, prior to drying, comprising a step of bringing the wood into contact with an aqueous bath } comprising the composition according to the invention.

Particular embodiments of the invention are described in the dependent claims.

The enzymatic composition used may comprise enzymes of the exocellular or endocellular type. The enzymes may be of bacterial or fungal origin, or of any other possible origin.

DETAILED DESCRIPTION OF THE INVENTION

:

The wood to be treated, the water content of which has ideally been brought tc between 25 and 45% by being rapidly passed through a drying facility, is immersed in a bath containing water or any other suitable solvent, and varying amounts of enzymes mainly of the hydrolase class (International Union of Biochemistry (IUB) class 3), chosen from enzymes of the cellulase type

Co (also known under the name glucosidases) (IUB classification subclass 3.2). In addition, enzymes of the lipase {subclass 3.1), peroxidase (subclass 1, 11.1), etc. type, and mixtures of these enzymes, may be added. Among cellulases which can be used for the present invention, mention may. in particular be made of : cellobiohydrolases, endoglucanases, B-glucosidases, hemicellulases, a-amylases, and xylanases, endo-

SR. 200153 . cellulases, p-mannanases, etc., without losing sight of the fact that the enzymes often have secondary activities besides their main activity.

Depending on whether neutral or acid cellulases are used, the pH of the bath is adjusted to between 3 and 8, preferably between 4 and 7. The temperature of the bath is kept constant at between 20 and 70°C, preferably between 30 and 50°C. After this treatment by immersion, which can last from 30 minutes to 2 weeks, preferably from 1 hour to 1 week, depending on the strength of the effect desired, on the mixture of enzymes used, on the : volume to be treated, on the volume of the bath, on the nature of the wood treated and on the temperature and pH of the bath, the wood is then placed in the drying: facility at controlled temperature and humidity in order : to carry out conventional drying which makes it possible to attain a water content of 15 to 20% for use in cooperage, starting from a wood having a water content of between 25 and 45%, preferably between 30 and 40%, before the bringing into contact with the enzymatic preparation. - The method according to the invention has proved to enable hydrolysis of soluble ellagitannins naturally . present in wood, firstly ellagitannins linked to parietal polysaccharides, and also diverse aromatic molecules (such as digallic acid or heteroside coumarins) which are transformed, by the method according to the invention, into neutrally tasting molecules.

The method according to the invention therefore makes it : possible to improve the impact on taste of the wood extracts by eliminating a ‘significant portion of the undesirable compounds (such as certain phenol compounds) and hydrolysing forms judged to be bitter or astringent.

This is particularly important for wood subsequently coming into centact with foods (typically, wood for cooperage) .

This method superficially modifies the ultrastructure of the wood. By electron microscopy, a decrease in the cell wall is noted over 1 mm. Over 1 to 3 mm, the pores of the cells are open. The effects of the process are felt : | up to a depth of approximately 5 mm.

The present invention also has the advantage of providing a rapid method for ageing wood, while at the same time respecting all of the physicochemical reactions linked to the natural drying of wood.

In addition, the invention has the advantage of providing a reproducible method for seasoning wood and of allowing better homogeneity of the quality of treated wood between different species of wood or species of : wood of different origin, or between the various : portions of the same tree. More constant physicochemical (in particular mechanical and aromatic) properties of the wood treated according to the invention are therefore obtained, with respect to wood dried naturally, this being independent of meteorological factors. oo .

The method according to the invention also makes it possible, through a suitable choice of enzymes having more specific activities, to confer on the wood specific properties such as the expression of specific odours or : flavours.

MATERIALS AND METHODS . 1. Origin of the wood samples

The oak samples consist of the heart-wood transformed into duramen, of trees from 110 to 150 years old, derived from homogeneous and suitably maintained areas of forest. Only the lower quarter of the trunk, generally constituting the wood for cooperage, is used for the study. The samples treated with the enzymatic composition according to the invention are simply dried in a drying facility (1 month, 40°C, with ventilation).

The various randomly-taken samples are planed down to sawdust by crushing in liquid nitrogen, before being sieved so as to keep only the particles less than 250 pm in size. The samples are conserved after lyophilization, to be analysed within a period of 2 months. The species studied is Quercus petraea (Allier, France). 2. Preparation of extracts and control of their composition 1 g of sawdust (250 pm) is extracted with 100 ml of solvent (7:3 by volume acetone/water) for 12 h at room temperature on .a shaking platform; the resulting extract is then filtered through a membrane, lyophilized and weighed. he 100 pg of lyophilized extract are used to identify the major ellagitannins, with an LSIMS mass spectrometer. 1 mg of the extract is taken up with methanol/water (6:4) in order to be analysed by HPLC coupled to a UV- detector and to an LSMIS mass spectrometer, according to the device developed by Vivas et al. (1995). The HPLC separation method is described in the following paragraph. 3. Assaying of phenol compounds 3.1. Coumarins

The coumarins are quantitatively extracted, by extrac- tion with diethyl ether, from 20 ml of a wood extract.

The organic phase 1s evaporated to dryness and the residue is taken up with methanol. The HPLC analysis is carried out using -a Varian 5060 coupled with a spectrofluorometric detector (Kontron SFM23/B) and an

Ultrasphere ODS column. The pure coumarins were supplied by Extrasyntheése (esculin, esculetin, scopoletin, ombelliferone, methylombelliferone) and Sigma (sporalen).

The coumarins are observed by fluorescence (excitation 425 nm and emission 325 nm). 3.2. Ellagitannins : 3.2.1. Reference products

Vescalagin and castalagin are isolated and purified from the duramen of Q. robur, under the conditions described by Vivas et al. (1995). The various roburins (A-E) and grandinin originate from Scalbert (INA/INRA Thierval-

Grignon) . 3.2.2. Separation and assaying of: ellagitannins by HPLC

The chromatographic technique for separating and assaying ellagitannins is in accordance with the method developed by Scalbert et al. (1990). The wood extracts

- 9g - are analysed by HPLC on an Ultrasphere ODS column. The detection is carried out at A = 280 nm. 3.2.3. Assaying of total ellagitannins a) Estimation of the level of total phenol compounds

The richness of the wood extracts in total phenol compounds is estimated either by the method using the

Folin-Ciocalteu reagent, or by measuring the absorbance © at 280 nm of the extracts diluted 100-fold (Vivas et al., 1993). These two methods' give results which are comparable but not specific for ellagitannins.

Db) Reaction for oxidation with nitrous acid

In this method proposed by Bate-Smith (1972), esters of hexahydroxyphenic acid and of glucose are oxidized with nitrous acid under a nitrogen stream. The reaction produces a blue coloration which is measured at 600 nm. " The results are estimated in mg/g or mg/l of castalagin equivalent (ecsoomm : 983 g™'). : c) Acid degradation

The method proposed is adapted from that developed by

Peng et al. (1991). It is based on the acid hydrolysis of ellagitannins, followed by assaying, by HPLC, of the released ellagic acid. The results are expressed in mg/g of castalagin equivalent, taking one mole of castalagin to give, under these. conditions, one mole of ellagic acid (Peng et al., 1991).

4. Extraction and assaying of polysaccharides 4.1. study of the polysaccharide fraction of the control and treated samples . 5 4.1.1. Extraction of polysaccharides 139 g of dry sawdust are left to soak for 72 h on a shaking platform at room temperature. The solution is 100 then filtered and centrifuged. The extract is then concentrated down to 500 ml. 4.1.2 Isolation of polysaccharides

The extracts undergo 3 precipitations (one at 1:9 of water/ethanol at 95% vol., two at 1:5 of the same mixture). The precipitation is carried out at 3°C for 12 h. Next, the fractions are lyophilized. 4.1.3. Partial characterization

The precipitate has a frothy appearance, very pale in colour, slightly grey, which is probably a complex with the ellagitannins from which it is difficult to isolate the polysaccharide fraction. 4.1.4. Assaying using a chemical method a) Assaying of neutral polysaccharides (nP)

The neutral polysaccharides are assayed using the sulphuric phenol method. The optical density is read at 490 nm and the results are expressed in mg/l of glucose equivalent. :

b) Assaying of acid polysaccharides (aP)

The acid polysaccharides are assayed using the meta- phenylphenol method. The optical density is measured at 520 nm and the results are expressed in mg/l of galacturonic acid equivalent. 4.2. Extraction and assaying of polysaccharides from wood 1 g of wood shavings derived from oak trees from various regions, the wood of which has a characteristic grain, is left to soak for 24 h in water at 20°C on a shaking platform. The solution is collected by filtration and ‘centrifuged. The sawdust recovered is then dried and left to soak once again, in a 5% sodium hydroxide solution, under the same conditions. The alkaline solution is also collected by filtration and centrifuged. Then, the two categories of extracts (water and sodium hydroxide) are supplemented with 95% ethanol in a proportion of 1:5 by volume. The polysaccharides are then collected by centrifugation and taken up with distilled water at 60°C. The nP are assayed with sulphuric phenol and the aP with meta-phenylphenol. The reference solutions are, respectively, an aqueous solution at 100 mg/l of glucose and 50 mg/l of galacturonic acid for nP and aP.

EXAMPLES

The invention is illustrated by the following examples, without, nevertheless, being limited thereto.

Example 1 8 piles, each of approximately 1 n®, of oak staves stacked in staggered rows, arranged to leave a space between each stave in order to allow the solution to circulate, are immersed in a container containing 15 000 litres of water. The temperature of the water is adjusted to 40°C .and the pH to 4.5-5.0, the wood being left to soak for 24 hours. :

Next, 8 kg of an enzymatic formulation are added, comprising mainly: e cellulases (IUB No. EC 3.2.1) with in particular - an activity measured in endocellulase units (IUB

No. EC 3.2.1.4) of 10 000 ECU/g of formulation, : - an activity measured in xylanase units (IUB No. EC 3.2.1.8) of 6 000 BXU/g of formulation, and - an activity measured in p-mannanase units (IUB

No. EC 3.2.1.78) of 4 000 MNU/g of formulation; e o-amylases (IUB No. EC 3.2.1.1) with an activity measured in “a-thermo-stable” units of 10 000 oTU/g of formulation.

These concentrations are, of course, given by way of example. It is clear that they may vary considerably depending on the various parameters used for the treat- ment (treatment time, strength desired, nature of the : wood, volume of the bath, volume of wood to be treated, pH and temperature of the bath, etc.).

The enzymatic solution is left to act while maintaining a gentle circulation in order to ensure homogeneous distribution of the enzymes in the bath. : Co

The wood is then treated for 48 hours and then taken out of the bath and transferred to the drying facility for three weeks in order to allow it to attain a water : content of approximately 18%. The staves are then ready for the manufacturing of barrels, after a total treat- ment of 24 days instead of 2 years in the case of natural ageing, for a final organoleptic quality which is equivalent. The drying will, however, have to be adjusted for each batch of wood in order to avoid the . 10 appearance of cracks.

Example 2

Sawdust and blocks of oak for cooperage (Quercus petraea) were treated with an aqueous solution of enzymes according to Example 1 (1 g of enzymatic preparation in 2.5 1 of water per dm’ of wood), tor 24 h at 35-40°C and pH 5, according to the materials and methods above.

After ‘treatment, the main groups of compounds of the wood were fractionated and the varicus fractions were compared with respect to a nontreated control.

In order to carry out this fractionation, the oak extract is evaporated to dryness. The resulting dry extract 1s taken up with a volume of water, with - stirring for 2 hours at 30°C. The resulting insoluble fraction comprises the lignins. The soluble fraction is again evaporated to dryness. The resulting dry extract is then taken up with a volume of an ethanol/water (9:1 by volume) mixture and stored for 12 h at 4°C. The resulting insoluble fraction comprises the poly- ’ saccharides. With regard to the soluble fraction, it comprises the ellagitannins. In the treated wood, the

C ~ ‘dry extract is 4.2% by weight with respect to the wood, against 12.3% by weight for the green-wood control. The percentages by weight of each of the three fractions, with respect to the dry extract, are given in Table I.

Table I :

Treated according 35% 50% 15% to the invention

This test made it possible to demonstrate hydrolysis of the ellagitannins, in parallel with a relative increase in the polysaccharide content, in the dry extract.

These various fractions were then subjected to more precise assays, demonstrating a decrease in extractable solids, in ellagitannins, in proanthocyanidins and in glucosylated coumarins (see Table II).

Table II

Results in mg/g of wood Control Treated according to the invention

Dry extract on: sawdust 123 42 : wood’ 73 26

Ellagitannins on: sawdust 54 21 wood" 32 14 :

Proanthocyanidins on: } sawdust 0.62 0.12 wood’ 0.47 0.19

Ce .

Results in mg/g of wood to the invention

Coumarins glucosylated coumarins (esculin + scopolin) on: sawdust 5.7 1.4 wood’ 3.2 1.1 aglucone coumarins (esculetin + scopoletin) on: . sawdust : 1.3 © 6.8 wood 4.8 7.1 * Size of the samples: 5 cm x 5 cm x 10 cm

A substantial increase in the polysaccharides was also noted in another test in which the assay made it possible to distinguish between the nP and aP poly- saccharides (see Table III).

Table III

I = {mg/g of wood) wm | woem

Er a to the invention

The treatment therefore makes it possible to eliminate certain substances judged to be undesirable due to their astringent nature (such as castalagin) -or bitter nature (such as esculin), and to transform them into neutrally tasting molecules (such as gallic acid, ellagic acid and, to a lesser degree, esculetin), and also to increase the polysaccharide ccntent (which will tend to make wine matured in the presence of this wood fuller and fatter).

a + ’

A series of taste tests were also carried out (see

Table IV). They confirm the above experiments in that the amount of wood required to dctect the bitter or astringent natures are much higher in the case of the wood treated enzymatically according to the invention, with respect to green wood dried artificially or naturally.

Table IV

Comparison of the taste detection thresholds as a function of the method of seasoning the wood, indicating the taste detection threshold (Tse) in mg/l of wine model solution

CT wwe | semen

Wood seasoned - according to the 420 350 ’ invention - by natural drying : 300 290 - by artificial drying 165 140 (drying facility) : * Tspy: concentration starting from which a substance is detected by 50% of the tasters.

Example 3

The measurement of phenol heterosidase enzymatic activity PeH is based on the release of carboxylic functions which lower the pH of the reaction medium. 9 mg of an aqueous extract of green oak are added, in a test tube, to 9 ml of a 0.1 M NaCl solution, pH 4.5, containing 1% of enzymatic preparation EP. A control tube 1s prepared under the same conditions, without enzymes. The pH is measured in the two tubes at time 0 (pHco for the control and pHeo for the enzyme) and after

FUR

4 hours of incubation at 25°C (pHcs and pHe,, respec- tively) . The optimum conditions were. established beforehand and there is perfect correlation between the release of glucose and the drop in pH.

The phenol heterosidase activity is calculated from the equation:

PeH = [(pHco - pHcs) — (pHeo — pHes)/4] x 100 (PeH being expressed in 107° units of pH/ml/h)

The activity measurements were carried out on various species of oak treated according to Example 2, in order to determine the differences in possible effectiveness of the enzymatic formulation, as a function of the species of oak (see Table V).

Table V stellata | lobata | prinus [Eee wees ao [as

IE IEE EEC NCE CR ICH IES

IEEE IEE EC EE ICN IS RE

) 20 With regard to the French oaks (Vosges, Allier,

Limousin), no significant difference is observed, the composition of the extracts being, itself, often similar. | :

In the species of American oaks (Q. alba, stellats, lobata, prinus) it is noted that the activity is expressed more strongly than in the French oaks. This is doubtless due to the presence of galloyl-containing compounds, which are present in greater amounts. Since the activity is greater, more significant seasoning of the American species may therefore be expected, which, in terms of taste, should bring them closer to the

French species. This tendency is confirmed during tastings.

Table VI compounds : EE aa (vescalagin) tannins at concentration of reference the concen- : solution tration of

CEERI

EA EE pH/ml/h : ’

In general, all the heteroside substrates are used by the enzymatic activities present. Only the condensed tannins from grape show inhibition of the phenol hetero- sidase activity. This effect is demonstrated in

Table VI, by a decrease in Pel with respect to vescalagin. : ~~ Not only does the enzymatic preparation not act on the procyanidins (tannins from grape and from wine), but it is also greatly inhibited by them. It may be concluded from this that a possible (but highly unlikely) residual phenol heterosidase activity in the cask will not modify the tannic qualities of the wine, developed by the grape. :

’ The condensed tannins of the grape in fact have the property of combining proteins, which explains the inactivation of the activities of our enzymatic solution. This inhibition is irreversible since, when the enzymatic solution is returned to a situation in which condensed tannins are absent, it does not recover its initial activity.

Table VII oo ’ phenol heterosidase activity

Er OE RE RE RE

ICN I IER YX

The activity is inhibited on media which are too rich in pure ellagitannins, as indicated in Table VII. In practice, when wood is treated, this does not represent a limitation of the technique since the local concen- tration of free ellagitannins never reaches these values. The ellagitannins of the wood are in fact associated with glucoside structures which attach them to the walls of the fibres. The immobilized form of : these tannins does not, itself, show any inhibition on the phenol heterosidase activity. As these structures are digested by the enzymatic preparation, the ellagitannins are released "into the medium, but are immediately hydrolysed by the enzymes present.

In addition, applying the enzymatic preparation by soaking enables the wood to be washed, depleting the : © superficial layers and thus decreasing, accordingly, the local concentration of ellagitannins. .

SR

Table VIII

Influence of pH on the phenol heterosidase activity

HE EN EE AE EE

‘As indicated in Table VIII, the phenol heterosidase activity is optimal at pH 5, which corresponds to the general optimum of activity of the enzymatic preparation, measured by the release of glucose into the medium.

Table IX

Enhancement of the value of chestnut through the use of the enzymatic preparation according to the invention .Digallic Glucosylated ) acid coumarins

Treated according to the invention

Digallic Glucosylated : acid coumarins

Gironde chestnut 0.8 : 3.5

Chestnut extracts (rich in digallic acid) from diverse origins were subjected (see Table IX) to enzymatic

Tl. s hydrolysis with the enzymatic preparation according to

Example 2. Virtually total hydrolysis of the digallic acid to gallic acid emerges from this, and also a large decrease in the glucosylated coumarins (the aglucone coumarins being insipid). :

These results are of great importance since they show the possibility of enhancing the value of a species which has been used very little to date. Chestnut in fact has all the mechanical qualities required for making cooperage wood, but 1s used very little because of the considerable bitterness which it produces.

The enzymatic treatment very greatly decreases this bitterness and makes it possible to envisage chestnut being used more in cooperadge, for specific uses in which oak 1s not suitable. Thus, for maturing colourless spirits, chestnut is absolutely indicated because, since it has no ellagitannins, it will hardly modify the fruit taste of the alcohol and, since it has no pigments, : neither will ‘it colour the alcohol left to mature in these casks. ;

Claims

} - . v' a CLAIMS

1. Use of an enzymatic composition comprising enzymes of the IUB cellulase class for seasoning green wood, : 5 characterized in that this wood is intended to come into contact with food.

2. Use of an enzymatic composition according to claim 1, characterized in that the wood consists of duramen.

3. Use of an enzymatic composition according to any one of the preceding claims, characterized in that the wood is chosen from species of oak or of chestnut.

4. Use of an enzymatic composition according toc any one of the preceding claims, characterized in that the wood 1s intended for cooperage.

5. Use of an enzymatic composition according to any one of the preceding claims, characterized in that this: ‘composition also contains enzymes chosen from peroxidases, lipases and mixtures of these enzymes.

6. Use of an enzymatic composition according to any one of the preceding claims, characterized in that the } enzymes of the cellulase type are chosen from a-amylases

(3.2.1.1), cellobiohydrolases (3.2.1.91), endocellulases

(3.2.1.4), endoglucanases (3.2.1.4), B-glucosidases

(3.2.1.21), hemicellulases (3.2.1.4), xylanases

(3.2.1.8) and P-mannanases (3.2.1.78).

7. Use of an enzymatic composition according to any one of the preceding claims, for modifying the aromatic characteristics of wood.

NE

8. Use of an aqueous bath comprising enzymes of the IUB cellulase class for seasoning green wood intended to come into contact with food.

Coo. Use of an aqueous bath according to claim 8, characterized in that the bath has an endocellulase : activity of between 1 x 10° and 100 x 10° ECU per m’ of wood to be treated. . :

10. Use of an aqueous bath according to either of Claims 8 and 9, characterized in that the bath has a xylanase activity of between 0.6 x 10° and 60 x 10° BXU per m’ of wood to be treated.

11. Use of an aqueous bath according to any one of Claims 8 to 10, characterized in that the bath has a B-mannanase activity of between 0.4 x 10° and 40 x 10° MNU per m® of wood to be treated.

12. Use of an aqueous bath according to any one of Claims 8 to 11, characterized in that the bath has an a-amylase activity of between 1 x 10° and 100 x 10° ,TU per m’ of wood to be treated. 2513.

Method .for seasoning green wood intended to come into contact with food, characterized in that it. comprises a step of bringing the wood into contact with a bath comprising enzymes of the IUB cellulase class.

14. Method according to claim 13, characterized in that it comprises a subsequent step of artificial drying.

15. Method according to either of claims 13 ~and 14, characterized in that the wood is brought into contact

EE with the bath by immersing the wood for a period of time ranging from 30 minutes to 2 weeks.

16. Method according to any one of claims 13 to 15, : characterized in that the pH of the bath is between 3 and 8.

17. Method according to any one of claims 13 to 16, ‘characterized in that the temperature of the bath is between 20 and 70°C.

18. Method according to. any one of claims 13 to 17, characterized in that the water content of the wood to be treated is adjusted to between 25 and 45% before subjecting the wood to the enzymatic preparation.