SE439931B - PROCEDURE FOR GETTING GLUCOSE FROM CELLULOSALLY MATERIALS CONTAINING HEMICELLULOSA AND LIGNIN - Google Patents

Description

Of acids or enzymes, but such attempts have not been completely successful, at least not in developing an economically attractive method which gives a satisfactorily high yield of glucose from cellulosic materials.

The present invention provides a process for obtaining glucose from cellulosic material containing hemicellulose and lignin, wherein hemicellulose is removed from the cellulosic material, the cellulose contained in the cellulosic material is dissolved in a solvent at room conditions, the solvent being a mixture of ethylene and ethylene water, whereby the dissolved cellulose is reprecipitated and the reprecipitated cellulose is hydrolyzed by means of a cellulase enzyme at a pH of about 5 to give glucose. The process may also involve hydrolyzing cellulosic materials, dissolved in a solvent, to produce glucose, after which lignin is separated from the glucose.

As stated above, cellulose molecules form highly ordered crystalline structures. In addition, it forms lignin present in the interlayers in cellulosic materials, a physical seal that surrounds the cellulosic fibers in such materials.

In one embodiment of the process of the present invention, selective solvent extraction is used to fractionate hemicellulose, cellulose and lignin, (the three main components of all cellulosic material). This fractionation is necessary when it has been found that after the cellulose has dissolved, it is no longer protected by the crystalline structure or by the lignin seals, and the cellulose can then be easily precipitated. The recovered cellulose can then be used in a number of ways, such as in the form of "synthetic cotton" or "new paper", and has been found to be particularly useful when the cellulose is hydrolyzed to give a high yield of glucose.

An example of the performance of our process for extracting cellulose from cellulosic materials, as well as the use thereof, including utilization by hydrolysis to produce glucose from cellulose follows below. In this embodiment of our process, the cellulosic material is first extracted, after being physically reduced to a suitable size, if necessary or desired, with a dilute acid or alkali to remove the hemicellulose. The residue after removal of the hemicellulose contains cellulose and lignin. The residue is then dissolved in a suitable solvent. There are a number of solvents reported in the literature that are capable of dissolving cellulose.

After examining a number of these, a solvent known as Cadox was found to give the best results. The cadox is first stated by Jayme and Neuschaffer (see Jayme, G. and _ Neuschaffer, K., 1957, Die Naturwissenschafter, 44 (3), and Jayme, G. and Neuschaffer, K., 1957, Die Makromolekulare Chemie, 28:71 -83). The cadox consists of -30% ethylenediamine and 70-75% water and in addition about 4.5-7% cadmium (added in the form of oxide or hydroxide). Cadox has good dissolution properties, is a clear, colorless, almost odorless liquid, is stable for a virtually unlimited time, causes little degradation of the cellulose and is relatively easy to prepare.

The fact that water is part of the Cadox is of great importance. Most cellulosic residues contain water vapor. The cadox does not require pre-drying of the residues, during dissolution of cellulose, whereby an otherwise very expensive operation can be avoided.

Approximately 10 g of cellulose is dissolved in 100 g of Cadoxen, in order to give an example of the effect thereof. By filtration or centrifugation, lignin, which does not dissolve, can separate; away.

The remaining cellulose-Cadoxen solution is further processed to recover cellulose, e.g. by any of the following treatments: a) The cellulose-Cadoxen solution can be spun through a nozzle into a suitable liquid to recover the cellulose in the form of long threads. This can be a source of "synthetic cotton". b) The cellulose-Cadoxen solution can be dispersed in a suitable liquid to recover the cellulose in the form of sheets. Such sheets should have many uses. Recovered cellulose recrystallizes, which reduces transparency. After adding filler and cover material, a new type of paper can be obtained. c) Cellulose ~ The cadoxene solution can be spun through a nozzle into a suitable liquid, to recover the cellulose in the form of long fibers. This can be the basis for a "boiling" process, which is essential for the removal of lignin and hemicellulose from wood-containing materials. The regenerated cellulose fiber can be made into paper as well as the original pulp.

Cellulose ~ The cadoxene solution is physically stable but this stability, it seems, requires a careful balance between cellulose, water, ethylenediamine and cadmium.

The exact balance is not known in detail, but if e.g. If excess water is used, the cellulose is reprecipitated from the solution. The cadox has a high pH value and when an acid is added, the cellulose also precipitates out of the solution. The reprecipitated cellulose, like other linear polymers, can recrystallize spontaneously. The reprecipitated cellulose can then be easily hydrolyzed to give a high yield of glucose, by either an acid or cellulose enzyme. For example, by adding an acid to a cellulose-Cadoxen solution to reduce this value to about 5 (whereby the cellulose precipitates again), followed by the addition of Tricodermavvijdefcellulase solution, the reprecipitated cellulose is hydrolyzed to glucose. Apparently the enzyme is not inactivated by ethylenediamine and the cadmium salt in the solution.

To improve the economy of the process, Cadoxen must be recycled and reused.

Through various recycling techniques, the process can be further divided according to a number of alternatives, which are described below. a) Addition of water for precipitation of the cellulose from a cellulose-Cadoxen solution.

Separation of the cellulose by filtration or centrifugation. The cellulose is then hydrolyzed to glucose using acids or enzymes, or both. To the solution can be added CO 2 to precipitate cadmium carbonate, which in turn can be converted to cadmium oxide for reuse. The amine remaining in the aqueous solution can be recovered using standard methods such as distillation or extraction. b) Addition of CO 2, directly to the cellulose-Cadoxen solution, whereby both cellulose and cadmium carbonate precipitate. After separation from the liquid, which contains ethylenediamine, which can be recovered according to a), the mixture of the two solid precipitates can be treated with cellulase to hydrolyze and dissolve the cellulose, to produce glucose. The remaining cadmium carbonate precipitate can be converted to cadmium oxide for reuse. c) Addition of a solvent, immiscible with water, to extract ethylenediamine for recycling, whereby the cellulose precipitates. Thus, in one step, we can separate the mixture into three fractions: amine in an organic layer, cadmium in an aqueous layer and cellulose as a precipitate. The three fractions are processed separately. d) The reprecipitated cellulose can be hydrolyzed to glucose by either an acid-enzyme two-step process or an all-enzyme process. An acid or cellulase enzyme (probably an endo-1,4-beta-glucanase, but the cellulase mechanisms are still the subject of considerable controversy) can "dissolve" the cellulose by forming cellodextrins. To complete the hydrolysis to glucose, either free or immobilized cellulase was then used. e) Cellulose re-precipitated from Cadoxen has been shown to spontaneously form droplets if left in solution for a number of days. Such drops may possibly support enzyme immobilization.

In a further embodiment of the process according to our invention, the cellulose is dissolved and reprecipitated "in place" and is therefore never removed, purely physically, from the lignin. The process may be preferred where separation of the cellulose is not desirable.

In this process, the cellulose is hydrolyzed to glucose and the remaining lignin, filtered, is filtered off from the resulting glucose solution. This method has been found to be easier than separating the lignin from the cellulose and then hydrolyzing the cellulose. Enzyme can be used to hydrolyze the cellulose and it is believed that acid hydrolysis can also be used.

The following steps are used for this process: 1. Treatment of cellulosic material to remove hemicellulose (this treatment may be the same as that described above with reference to the first discussed embodiment), 2. Dissolution of the cellulosic material in a solvent (again, may this step be identical to that described above with reference to the first discussed embodiment), 3. Addition of water, acid or any other liquid such as e.g. methanol to reprecipitate cellulose; 4. Hydrolysis using enzyme or acid to produce glucose, with water-insoluble lignin as the only residue, and. Separation of lignin from glucose syrup by filtration.

Below are a number of non-limiting examples of our process.

EXAMPLE 1 Pre-treatment and hydrolysis of cellulose. a) Preparation of the solvent (Cadoxen): Ethylenediamine, 25 to 30% in water, was saturated by adding an excess of Cadmium oxide, which had previously been oven dried. Cadmium oxide, which did not dissolve, formed a hydroxide in the form of a white precipitate. or centrifuged off. The procedure of adding cadmium oxide and separating off the hydroxide was performed twice to ensure that a solution saturated with cadmium oxide was obtained.

In this way, an aqueous solution with 25 to 30% of ethylenediamine and 4.5 to T cadmium was obtained. Due to its cadmium and ethylenediamine content, the solution is commonly referred to as Cadoxen. b) Dissolution of OL cellulose l) 0.1 g of crystalline O 6 cellulose (Avicell) was added to 10 ml of the solvent prepared according to a) and mixed with it. The dissolution of the cellulose was complete within 10 minutes at room conditions. 2) The dissolution experiments were repeated with 0.1 g, 0.2 g, 0.3 g, 0.4 g and 0.5 g of crystalline CL-cellulose (Avicell), each in 10 ml of solvent, via a procedure in which the cellulose was mixed with the solvent at room conditions, after which the mixture was cooled by placing it on ice. The times required for complete dissolution are set out in Table 1.

TABLE l t Avicell Dissolution time in the solvent l 2 minutes 2 2 minutes 3 7 minutes 4 9 minutes 15 minutes 3) The dissolution experiments, performed as above, were repeated. But in this case, amorphous cellulose was dissolved instead of crystalline cellulose. Using the same amounts of cellulose and the same procedure as in (2), the dissolution times listed in Table 2 were observed. TABLE 2% cellulose Dissolution time ilömñmaæüet minutes 2 2 minutes 7 minutes 9 minutes UW-P -UOND-1 minutes 4) Solutions containing a larger amount of cellulose were prepared by modifying the composition of the solvent and by changing the dissolution procedure.Sodium hydroxide was added to the solvent in (a) to give a 0.5 M solution with respect to sodium hydroxide. of the modified solvent was added 300 mg of Avicell, followed by the addition of an excess of Cd0, the solution was mixed and an almost clear, very viscous solution was obtained, with a cellulose concentration of about% c) Preparation of enzyme 1) Cellulase enzyme from the Enzyme Development Corporation, New York ", was treated as follows to give enzyme preparation" CS ". 10 g of enzyme was added to ml of water. The enzyme solution was concentrated using an ultrafilter membrane and diluted with an excess of water, after which it was concentrated again to a volume of 25 ml. This was repeated several times until the salt and carbohydrate content of the enzyme preparation was negligible. 2) The enzyme preparation "CN" was prepared as follows. 10 g of enzyme was dissolved in 100 ml of water. Then 57 g of ammonium sulfate were added. Upon mixing, the ammonium sulfate dissolved and a white precipitate formed. The precipitate was separated by centrifugation and redissolved in 30 ml of water. The solution was then desalted on a Sephadex G-25 column (Pharmacia Corporation) and diluted to a final volume of 100 ml. d) Hydrolysis of crystalline OL cellulose. 1) to 1 unit of volume of 2% crystalline ß-cellulose dissolved in the solvent according to (a), 2 units of volume of 30% of HCl were added, causing the dissolved cellulose to reprecipitate. Then 5 units of volume of sodium acetate buffer and 1 unit of volume of enzyme preparation "CS" were added. Upon mixing and incubating in 5006, up to 50% conversion of the cellulose to glucose was obtained within 30 minutes. Obviously, the enzyme is not inactivated by the presence of cadmium and ethylenediamine. 2) One volume unit of 3% Avicell, in solvent (a), was mixed with 2.5 volume units of water, which resulted in the cellulose being reprecipitated. The reprecipitated cellulose was washed with water, after which the cellulose concentration was 0.5%. After adjusting the pH of the solution to 5 with buffer, a sufficient amount of enzyme preparation "CN" was added to give an enzyme volume: cellulose weight ratio of 18: 1. Incubation of this solution at 50 ° C gave 80% conversion to glucose within 5 hours and 90% within 50 hours. In comparison, a control experiment gave cellulose that did not dissolve in solvent and reprecipitated, ie. untreated cellulose was used, giving only l5% conversion after 5 hours and 47¿ after 50 hours.

EXAMPLE 2 Pretreatment and hydrolysis of agricultural waste Preparation of the solvent and enzyme solutions was done exactly as described above in Example 1. a) Dissolution of cellulose in corn cobs. Maize flasks, which were ground to a particle size of 0.4 mm, were mixed with solvent in a weight ratio of the corn cob. After stirring for 2.5 hours, the solid phase and the liquid phase were separated. The solid phase was washed with water, dried and weighed.

The weight losses, based on the weight of the dry solids before and after, were as high as 77%. This indicates that all cellulosic material has dissolved. b) Dissolution of cellulose in maize residues.

Maize residues, ground to a particle size of 0.4 mm, were mixed with solvent in the weight ratio of maize residue: Cadoxen of 1: 10. The stirring, washing and recovery of the solids were performed according to Example 2 (a). Weight losses were estimated at between 44 and 94% (based on dry weight). c) Dissolution and hydrolysis of maize residues on site.

Maize residues containing 38% O 6 cellulose were mixed with Cadoxen in a weight ratio of maize residue: Cadoxen of 1: 4.2. After allowing the mixture to stand for 12 hours, buffer, water and enzyme preparation "Cw" were added to give a 2.5% solution of residues at a pH of 5. Hydrolysis of the mixture at 45 ° C gave 72% conversion of Since the pretreatment with the solvent and the subsequent reprecipitation of the cellulose was done without first separating the solvent and the dissolved cellulose from the solid residue, this technique is referred to as "in situ" solution (and reprecipitation). . d) Dissolution and hydrolysis of bagasse (sugar cane residue) on site.

Bagasse residues containing 33% cellulose were mixed with Cadoxen in a weight ratio bagasse: Cadoxen of 1: 4.2. Under the same conditions as in Example 2 (c), 70% conversion was obtained in 19 hours.

EXAMPLE 3 Hydrolysis with immobilized enzyme. a) Preparation of immobilized enzyme.

A filter consisting of chemically active, porous PVC membrane material (obtained from Amerace Corporation, Butter New Jersey) was immersed in a solution of 10 H-glutaraldehyde buffer with phosphate, to a pH of 7, for 2 hours under room temperature. 2 conditions. After washing with water and sodium acetate buffer (pH 5), the disk was immersed in enzyme preparation "CW" for 12 hours at 4 ° C. The disc was then washed with buffer, placed in a column apparatus and used in the manner described below. b) Hydrolysis of cellodextrins using immobilized enzyme, with complete resolution where 79% was converted to glucose and excess was obtained as cellodextrins. This clear solution was passed through the disk with immobilized enzyme 5 times, giving an increase in glucose conversion from 79% to 81%.

Glucose was formed by hydrolysis of soluble cell oligosaccharides to glucose.

Subsequently, a control experiment was performed using only soluble enzyme. In this experiment, the glucose conversion was constant at 79%, during the same time period.

EXAMPLE 4 Preparation of cellulose drops A solution of 2% Avisell in Cadoxen was mixed with 50% HCl in a volume ratio of solution acid of 1: 1.5. This caused the cellulose to precipitate. The mixture was heated to 50 ° C for 30 minutes and then cooled to room temperature.

During the 12 hours that the solution was then allowed to stand, the flocculating cellulose precipitate spontaneously formed small cellulose droplets.

From the foregoing, the present invention provides a process for treating cellulosic material and producing glucose therefrom, comprising a process for dissolving and reprecipitating cellulose "on site", and another process for recovering cellulose from cellulosic material, which can then be used to a high production of glucose.

Claims (10)

10 20 25 30 Q 7901898-2 PATENT REQUIREMENTS A process for obtaining glucose from cellulosic material containing hemicellulose and lignin, wherein hemicellulose is removed from the cellulosic material, the cellulose contained in the cellulosic material is dissolved in a solvent at room conditions, the solvent being a mixture ethylenediamine, cadmium oxide and water, characterized in that the dissolved cellulose is reprecipitated and that the reprecipitated cellulose is hydrolyzed by means of a cellulase enzyme at a pH of about 5 to give glucose. A method according to claim 1, characterized in that the method comprises physically reducing the cellulosic material to a maximum particle size of 0.4 mm before the hemicellulose is removed therefrom. 3. A process according to claim 1, characterized in that the hemicellulose is removed by extraction with a dilute acid or alkali. A method according to claim 1, characterized in that said lignin is separated from the cellulosic material by filtration or centrifugation after the cellulose has been hydrolyzed to give glucose. A process according to any one of claims 1 to 4, characterized in that said cellulase enzyme which is added is an endo-1,4-beta-glucanase. 6. A method according to claim 5, characterized in that the added cellulase enzyme is a Tricoderma viride cellulase solution. A process according to any one of claims 1 to 6, characterized in that said solvent is recovered for the purpose of reuse. 8. A process according to claim 7, characterized in that said solvent is recovered by the addition of CO 2 to precipitate cadmium carbonate which can be converted to cadmium oxide for the purpose of reuse. Process according to claim 7, characterized in that said solvent is recovered by adding a water-immiscible solvent to extract ethylenediamine for recycling and to separate the resulting mixture into three fractions comprising an organic layer of amine, a aqueous layer of cadmium and a precipitate of cellulose. Process according to any one of claims 1 to 9, characterized in that the hydrolysis is completed by the use of antigen-free or immobilized EH Zy fl 1.