WO2010067542A1 - Poly-3-hydroxyalkanoate and process for production thereof - Google Patents

Abstract

Provided is a poly-3-hydroxyalkanoate which, when heat-melted, exhibits reduced discoloration even when other components of biological origin have not completely been removed therefrom.  Also provided is a process for production of same. The discoloration of a poly-3-hydroxyalkanate during heat -melting can be prominently reduced by drying the poly-3 -hydroxyalkanoate under acidic conditions of pH of 3.2 or above.  The poly-3-hydroxyalkanoate thus obtained exhibits a YI of 25 or below as determined by subjecting 2g of the poly-3 -hydroxyalkanoate to melting at 170°C for 40 minutes and then curing at 60°C for 60 minutes.

Description

Poly-3-hydroxyalkanoic acid and process for producing the same

The present invention relates to a poly-3-hydroxyalkanoic acid with reduced coloring during heating and melting and a method for producing the same.

Poly-3-hydroxyalkanoic acid (hereinafter abbreviated as PHA) is a thermoplastic polyester that is produced and stored as an energy storage substance in cells of many microbial species and has biodegradability. Currently, non-petroleum-derived plastics are attracting attention due to increased environmental awareness. Among them, biodegradable plastics such as PHA which are taken into the natural material circulation and do not harm the decomposition products are attracting attention, and their practical application is eagerly desired. In particular, PHA produced and accumulated by microorganisms in the microbial cells is expected to have little adverse effect on the ecosystem because it is incorporated into the natural carbon cycle process.

However, PHA extracted from microorganisms is easily colored when heated and melted when processed into pellets or sheets, and the appearance may be extremely deteriorated, which is a major obstacle to commercialization. Therefore, as a means for solving the problem, a method for decomposing and / or removing a biological component considered as a cause of coloring has been proposed.

As a method of decomposing and / or removing biological components other than PHA, a method of solubilizing and removing biological components other than PHA by physical treatment, chemical treatment or biological treatment has been proposed. For example, a method of combining a treatment for crushing PHA-containing microbial cells and a surfactant treatment (Patent Document 1), a method of performing a crushing treatment after adding an alkali and performing a heat treatment (Patent Document 2), and the like can be mentioned. In addition, a method has been proposed (Patent Document 3) in which an aqueous suspension of microbial cells is treated with sodium hypochlorite or an enzyme to solubilize biological components other than PHA to obtain PHA.

As for the degree of purification of PHA that can be achieved by these treatments, analytical values based on the purity of PHA are exemplified. For example, in a method using an enzyme homogenizer and an oxidizing agent, PHA purity is shown to be 98% to 99% (Patent Document 4). Further, the purity of the method of crushing, fractionation, and oxidizing agent treatment is 97.8% (Patent Document 5). These are calculated | required by the area ratio of the PHA peak obtained when a resin is dissolved in chloroform and analyzed by gas chromatography and the weight ratio before and after extraction.

However, even if these treatments are applied, or even in combination, it is extremely difficult to completely remove biological components other than PHA. In addition, these processes are very difficult to industrialize from the viewpoint of cost. Accordingly, when PHA produced by an existing industrial production method is heated and melted when processed into pellets or sheets, coloring of PHA is inevitable.

Japanese Translation of National Publication No. 08-502415 International Publication No. 2004/065608 JP 2005-348640 A JP 7-177894 A JP 2002-306190 A

In this way, in the production of PHA using microorganisms, it is difficult to completely remove biological components other than PHA, and coloring that occurs at the time of heating and melting when performing sheet production, pellet production, and press molding from PHA. It has been very difficult to reduce. So a solution from another approach was eagerly desired.

An object of the present invention is to provide a PHA in which coloring during heating and melting is reduced and a method for producing the same without completely removing biological components other than PHA.

In the conventional production method, in order to decompose and remove biological components other than the remaining PHA, the pH is generally set to the alkali side during the production method.

For example, a method of combining a treatment for crushing a PHA-containing microbial cell and a surfactant treatment (Patent Document 1) or a method of performing a crushing treatment after adding an alkali and performing a heat treatment (Patent Document 2) The decomposition of biological components other than PHA is promoted by using the alkali side. In addition, a method of decomposing and removing biological components other than PHA by chemical decomposition treatment using hydrogen peroxide has also been proposed, but it is recommended that the treatment be performed while keeping the pH at the alkali side (International Publication No. 04). / 0292266). Furthermore, as a method for decomposing biological components other than PHA, a method using an enzyme can be mentioned. However, since the enzyme used is alkaline and active, the treatment liquid must be made alkaline. Generally, in the conventional method, since the treatment is performed on the alkali side, the PHA obtained therefrom is also alkaline.

On the other hand, the mechanism regarding coloring of PHA that occurs during heating and melting is unknown, and no means for avoiding coloring other than decomposing and / or removing biological components other than PHA have been reported.

Unexpectedly, when the present inventors separated PHA from microorganisms and finally dried PHA to obtain dry PHA, the drying may be performed under acidic conditions. It was found to be extremely effective in overcoming the problem of coloring. Furthermore, the present inventors have completed the present invention by finding a pH region where the coloring reaction during heating and melting is most suppressed.

The present invention relates to a dried poly-3-hydroxyalkanoic acid obtained by drying undried poly-3-hydroxyalkanoic acid under acidic conditions of pH 3.2 or higher.

In the present invention, the dry poly-3-hydroxyalkanoic acid is an undried poly-3-hydroxyalkanoic acid having an organic nitrogen amount of 500 ppm or less, 400 ppm or less, or 300 ppm or less per weight of the poly-3-hydroxyalkanoic acid. It is preferably obtained by drying under acidic conditions of pH 3.2 or higher.

In the present invention, the dried poly-3-hydroxyalkanoic acid is prepared by subjecting an aqueous poly-3-hydroxyalkanoic acid suspension produced by at least one of the following (1) to (3) to pH 3.2 or more. It is preferable to be obtained by drying under acidic conditions. At this time, the dried poly-3-hydroxyalkanoic acid is recovered from the aqueous suspension by a dehydration operation or the like, and the dried poly-3-hydroxyalkanoic acid is dried. Alternatively, the aqueous suspension can be directly dried by spray drying or the like without dehydrating (recovering) the undried PHA.

(1) a step of disrupting cells that have accumulated poly-3-hydroxyalkanoic acid,
(2) a step of decomposing and / or removing biological components other than poly-3-hydroxyalkanoic acid,
(3) A step of washing poly-3-hydroxyalkanoic acid with water and / or an organic solvent.

The present invention also relates to a method for producing poly-3-hydroxyalkanoic acid, which comprises drying undried poly-3-hydroxyalkanoic acid under acidic conditions of pH 3.2 or higher.

In the present invention, undried poly-3-hydroxyalkanoic acid having an organic nitrogen amount per weight of poly-3-hydroxyalkanoic acid of 500 ppm or less, 400 ppm or less, or 300 ppm or less in the production method is adjusted to pH 3.2 or more. Drying under acidic conditions is preferred.

Furthermore, the present invention relates to a poly-3-hydroxyalkanoic acid, which is obtained by melting 2 g of the poly-3-hydroxyalkanoic acid at 170 ° C. for 40 minutes and curing at 60 ° C. for 60 minutes. It also relates to a poly-3-hydroxyalkanoic acid, characterized in that the YI measured for hydroxyalkanoic acid is 25 or less.

In the present invention, in the poly-3-hydroxyalkanoic acid, 2 g of poly-3-hydroxyalkanoic acid having an organic nitrogen amount of 500 ppm or less, 400 ppm or less, or 300 ppm or less per weight of poly-3-hydroxyalkanoic acid is added. It is preferable that YI measured for poly-3-hydroxyalkanoic acid obtained by melting at 40 ° C. for 40 minutes and then curing at 60 ° C. for 60 minutes is 25 or less.

According to the present invention, it is not necessary to completely remove biological components other than PHA, and it is possible to provide a PHA in which coloring generated during heating and melting is reduced and a method for producing the same by an extremely simple technique.

Graph showing the relationship between the pH of poly-3-hydroxyalkanoic acid during drying and the color tone of PHA after heat-melting

The microorganism used in the present invention is not particularly limited as long as it is a microorganism that generates PHA in cells. A microorganism isolated from nature, a microorganism deposited at a depositary of a strain (for example, IFO, ATCC, etc.), or a mutant or transformant that can be prepared from them can be used. For example, the genus Capriavidus, the genus Alcaligenes, the genus Ralstonia, the genus Pseudomonas, the genus Bacillus, the genus Azotobacter, the genus Nocardia, Examples include bacteria. In particular, strains such as A. lipolytica, A. latus, Aeromonas caviae, A. hydrophila, and C. necator. Is more preferable. In addition, when a microorganism originally does not have the ability to produce PHA, or when the production amount is low, a transformant obtained by introducing a target PHA synthase gene and / or a mutant thereof into the microorganism Can also be used. The PHA synthase gene used for the preparation of such a transformant is not particularly limited, but a PHA synthase gene derived from Aeromonas caviae is preferred. By culturing these microorganisms under appropriate conditions, it is possible to obtain microbial cells that accumulate PHA in the cells. The culture method is not particularly limited. For example, the method described in JP-A No. 05-93049 is used.

In the present invention, PHA is a general term for polymers having 3-hydroxyalkanoic acid as a monomer unit. The 3-hydroxyalkanoic acid constituting the compound is not particularly limited, and specifically, a copolymer of 3-hydroxybutyrate (3HB) and another 3-hydroxyalkanoic acid, or 3-hydroxyhexanoate ( And a copolymer of 3-hydroxyalkanoic acid containing 3HH). Further, two types selected from the group consisting of 3-hydroxypropionate, 3-hydroxybutyrate, 3-hydroxyvalerate, 3-hydroxyhexanoate, 3-hydroxyheptanoate and 3-hydroxyoctanoate Examples also include copolymers having the above 3-hydroxyalkanoic acid as a monomer unit. Among them, a copolymer containing 3HH as a monomer unit, for example, a two-component copolymer (PHBH) of 3HB and 3HH (Macromolecules, 28, 4822-4828 (1995)), or 3HB and 3-hydroxyvalerate ( 3HV) and 3HH ternary copolymer (PHBVH) (Japanese Patent No. 2777757, Japanese Patent Application Laid-Open No. 08-289797) are more preferred from the viewpoint of the physical properties of the resulting polyester. Here, the composition ratio of each monomer unit constituting the two-component copolymer PHBH of 3HB and 3HH is not particularly limited, but when the total of all monomer units is 100 mol%, 3HH units are 1 to A composition ratio of 99 mol%, preferably 1 to 50 mol%, more preferably 1 to 25 mol% is suitable. Further, the composition ratio of each monomer unit constituting the three-component copolymer PHBVH of 3HB, 3HV, and 3HH is not particularly limited, but when the total of all the monomer units is 100 mol%, for example, The composition ratio of 3HB units is preferably 1 to 95 mol%, the composition ratio of 3HV units is 1 to 96 mol%, and the composition ratio of 3HH units is preferably 1 to 30 mol%.

In the present invention, before decomposing and / or removing impurities such as biological components other than PHA, it is preferable to disrupt cells containing PHA in advance by physical treatment, chemical treatment, or biological treatment. Thereby, the subsequent decomposition and / or removal step can be performed efficiently. The crushing method is not particularly limited, but a fluid shearing force, a solid shearing force, or grinding, such as a conventionally known French press, homogenizer, X-press, ball mill, colloid mill, DYNO mill, or ultrasonic homogenizer, is used. The method can be used. Also, methods using drugs such as acids, alkalis, surfactants, organic solvents, cell wall synthesis inhibitors, methods using enzymes such as lysozyme, pectinase, cellulase, thymolyase, methods using supercritical fluids, and osmotic pressure crushing methods , Freezing method, dry pulverization method and the like. In addition, a self-digestion method using the action of protease, esterase and the like contained in the cell itself is also a kind of disruption method. In the crushing method, it is desirable to select a method for suppressing a decrease in molecular weight of PHA due to a series of treatments. Moreover, these crushing methods may be used independently and may combine several methods. Further, batch processing may be performed, or continuous processing may be performed.

Usually, the PHA aqueous suspension obtained by crushing PHA-containing cells by the above method contains proteins, nucleic acids, lipids, sugar components, other cell components, and culture substrate residues in cells. It is mixed. Prior to the decomposition and / or removal step described below, it is preferable to carry out a dehydration step for separating water containing water-soluble components such as these proteins. Thereby, the amount of impurities contained in the aqueous PHA suspension can be reduced, and the subsequent decomposition and / or removal process can be efficiently performed. The dehydration method is not particularly limited, and examples thereof include filtration, centrifugation, sedimentation separation, and electrophoresis. The concentration of PHA in the aqueous suspension subjected to the decomposition and / or removal step is not particularly limited, but is preferably 50 g / L or more, more preferably 100 g / L or more, still more preferably 200 g / L or more, and even more preferably It is 300 g / L or more. Moreover, the said dehydration process may be implemented for the purpose of adjusting the density | concentration of PHA in aqueous suspension, and water may be newly added.

The method for decomposing and / or removing impurities such as biological components other than PHA is not particularly limited, and examples thereof include a method using an enzyme. Examples of the enzyme used include proteolytic enzymes, lipolytic enzymes, cell wall degrading enzymes, and nucleolytic enzymes. Specific examples of these enzymes include the following. These may be used alone or in combination of two or more.

(1) Proteolytic enzyme Esperase, Alcalase, Pepsin, Trypsin, Papain, Chymotrypsin, Aminopeptidase, Carboxypeptidase, etc. (2) Lipidase Lipase, Phospholipase, Cholinesterase, Phosphatase, etc. (3) Cell wall degrading enzyme Lysozyme, Amylase, Cellulase, Maltase, saccharase, α-glycosidase, β-glycosidase, N-glycosidase, etc. (4) Nucleolytic enzymes Ribonuclease, deoxyribonuclease, etc. Enzymes used for degrading impurities such as biological components other than PHA are limited to the above. However, any enzyme that has an activity of degrading biological components may be used as long as it can be used for industrial products. In addition, commercially available enzyme detergents for washing can be used. Furthermore, for example, it may be an enzyme composition containing an enzyme stabilizer, a recontamination preventive agent and the like and an enzyme, and is not limited to the enzyme alone. Among the proteolytic enzymes that are included in the above examples, protease A, protease P, protease N (above, Amano Enzyme), esperase, alcalase, zabinase, Everase (above, Novozyme) etc. are industrial. It can be preferably used from the viewpoint of decomposition activity. However, it is not limited to these.

The enzyme treatment time is preferably carried out until a desired degree of treatment is achieved, and is usually 0.5 to 2 hours. The amount of the enzyme used depends on the type and activity of the enzyme and is not particularly limited, but is preferably 0.001 to 10 parts by weight with respect to 100 parts by weight of PHA, and more preferably 0.001 to 5 parts by weight from the viewpoint of cost. Part is more preferred.

Other methods for decomposing impurities such as biological components other than PHA include methods using hypochlorous acid and hydrogen peroxide. When using hypochlorous acid, the pH of the system is set to an alkaline region, and PHA with a low residual amount of chlorine can be obtained by carrying out the process under conditions where heat, light, and contact with metal are suppressed. The pH is desirably 8 or more, more desirably 10 or more, and further desirably 12 or more. The treatment temperature is preferably 40 ° C. or lower, more preferably 30 ° C. or lower, further preferably 20 ° C. or lower, and preferably 10 ° C. or lower in order to reliably exhibit the effect.

As described above, in the dehydration step, filtration, centrifugation, or the like can be performed in order to separate PHA from water containing impurities such as other biological components. The filtration method is not particularly limited, but a method using Nutsche or the like, or a method such as suction filtration or pressure filtration is desirable. Industrially, filtration devices with pressing functions such as filter presses, tube presses, plate presses, gauge presses, belt presses, screw presses, disc presses, centrifugal dehydrators, and multi-chamber cylindrical filters can be selected. is there. In order to increase productivity, a continuous type such as a multi-chamber cylindrical filter is desirable. Examples of a method for removing particles from a continuous filter include a string method, a scraper method, and a precoat scraper method. Alternatively, a membrane separation method may be used. As a filtration method including membrane separation, dead-end filtration or cross-flow filtration can be selected. Any of these can be selected based on filterability, the degree of blockage of the filter medium, membrane, and the like. Further, the pressure may be reduced, vacuumed, or pressurized. Further, a method using centrifugal force may be used. Various materials such as paper, woven fabric, nonwoven fabric, screen, sintered plate, unglazed, polymer film, punching metal, and wedge wire can be selected as the filter medium. Either can be selected based on productivity and the degree of blockage. Moreover, a filter aid may or may not be used. When using a filter aid, there are a method of pre-coating the filter medium (pre-coating method) and a method of pre-adding to the filter stock solution (body feed method).

The method of centrifugation in the dehydration step is not particularly limited, but a centrifugal sedimentator, a centrifugal dehydrator, or the like can be used. If it is a centrifugal settling machine, a separator plate type, a cylindrical type, and a decanter type are mentioned. Examples of the separation plate type include a disk type, a self-cleaning type, a nozzle type, a screw decanter type, and a skimming type. There are a batch type and a continuous type depending on the method of discharging sedimentation components. As for the centrifugal dehydrator, there are a batch type and a continuous type. With these devices, it is possible to separate the sediment containing PHA and the culture solution components by the specific gravity difference.

Other methods that can be used in the dehydration step include flotation, electrophoresis, and cyclone treatment. Methods such as filtration, centrifugation, and flotation may be used alone or in combination. Moreover, in order to improve the filtration performance and sedimentation property of PHA, you may implement aggregation operation. Although it does not specifically limit regarding aggregating operation, For example, the method etc. of international publication 2004/033700 are mentioned.

After recovering the PHA by a method such as filtration or centrifugation in the dehydration step, the recovered PHA can be washed with water or the like to obtain a PHA with a further increased degree of purification. For washing, an organic solvent other than water may be used, or water and an organic solvent may be mixed and used. The pH of water may be adjusted. When an organic solvent is used as a washing solvent, preferably a hydrophilic solvent, specifically methanol, ethanol, acetone, acetonitrile, tetrahydrofuran, ketones, amines, or the like is used. A surfactant or the like may be added to water. A mixture of a plurality of these organic solvents and water may be used. In addition, for a short period of time, it is possible to improve the cleaning performance by spraying water or these organic solvents with heating or vapor.

The cleaning method may be either batch or continuous. The washing method is not particularly limited, but the washing can be carried out by suspending PHA in the washing solution by a stirring method or a circulation method using a pump or the like. Moreover, it can also wash | clean by adding a washing | cleaning liquid to wet PHA obtained by filtration etc. In order to enhance the cleaning effect, it is effective to repeat the cleaning. In order to reduce the amount of the cleaning liquid used, the cleaning liquid can be recycled after being subjected to a general regeneration process that can be considered by those skilled in the art, such as distillation or static separation. It is also effective to reduce the amount of cleaning liquid used by bringing the cleaning liquid and the target PHA into countercurrent contact.

As described above, according to the most preferred embodiment of the present invention, a culture process for culturing microorganisms having the ability to produce PHA in cells, a crushing process for crushing the microorganisms that have accumulated PHA, and an aqueous solution containing the crushed microorganisms A dehydration step for separating water from the suspension, a purification step for decomposing and / or removing impurities such as biological components other than PHA, a washing step for washing PHA with water and / or an organic solvent, and the obtained PHA aqueous solution It is possible to efficiently produce dry PHA by sequentially carrying out a step of recovering the dry PHA from the suspension and a step of drying the dry PHA under acidic conditions of pH 3.2 or more to obtain dry PHA. it can. However, the present invention does not necessarily require that all the steps described above be performed.

The solvent contained in the PHA aqueous suspension may include water, an organic solvent compatible with water, or a mixed solvent of water and the organic solvent. The organic solvent may be used alone or in combination of two or more. In addition, the concentration of the organic solvent in the mixed solvent of water and the organic solvent is not particularly limited as long as it is not more than the solubility of the organic solvent to be used in water. Further, the organic solvent compatible with water is not particularly limited, but for example, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, iso-butanol, pentanol, hexanol , Alcohols such as heptanol, ketones such as acetone and methyl ethyl ketone, ethers such as tetrahydrofuran and dioxane, nitriles such as acetonitrile and propionitrile, amides such as dimethylformamide and acetamide, dimethyl sulfoxide, pyridine and piperidine Can be mentioned. Among these, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, iso-butanol, acetone, methyl ethyl ketone, tetrahydrofuran, dioxane, acetonitrile, propionitrile, etc. are easy to remove. Is preferred. Further, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, iso-butanol, acetone and the like are more preferable because they are easily available. More preferred are methanol, ethanol, and acetone. In addition, as long as the essence of the present invention is not impaired, other solvents and components derived from bacterial cells and compounds generated during purification may be included.

In order to recover the undried PHA from the aqueous PHA suspension, the methods such as filtration, centrifugation, flotation method, electrophoresis method, and cyclone treatment described above can be used.

In the drying step, there is no particular limitation on the method of setting the pH condition for drying undried PHA to 3.2 or more. The aqueous PHA adjusted to an acidic region having a pH of 3.2 or higher is drained to recover the dry PHA, whereby the pH of the dry PHA may be 3.2 or higher. After the liquid is removed and undried PHA is recovered, the pH of the undried PHA may be 3.2 or more by adding an aqueous solution whose pH is adjusted to acidic to the undried PHA. The method for adjusting the pH of the aqueous PHA suspension to an acidic region of 3.2 or higher is not particularly limited. For example, a method of adding an acid, a method of adding an organic salt or an inorganic salt, or the like can be used. When the acid addition method is used, the acid is not particularly limited, and may be either an organic acid or an inorganic acid, and may be volatile. Further, for example, any strong acid such as sulfuric acid and hydrochloric acid, and weak acid such as phosphoric acid and acetic acid can be used, and the same color tone improving effect can be obtained. In addition, when adding an aqueous solution whose pH has been adjusted to acidic after dehydrating the aqueous suspension of PHA, the acid species used for adjusting the pH of the aqueous solution is not limited. Can be used. Addition of organic salt or inorganic salt can also be applied, and the same effect as the addition of aqueous pH solution can be obtained as long as it can be adjusted to an acidic region of pH 3.2 or higher. It is also possible to substitute for acid addition using an aqueous solution adjusted to a predetermined pH during dehydration washing.

When drying PHA under acidic conditions, the lower limit of the pH is 3.2 or more, preferably 3.3 or more, more preferably 3.4 or more, and even more preferably 3.5 or more. If it deviates from this pH range, the coloration during heating and melting is promoted. Although details of this mechanism are still unclear, it is considered that the reaction that occurs during heating and melting is suppressed in the pH range. The upper limit of the pH is preferably less than 7 and more preferably 6.5 or less from the viewpoint of suppressing coloring during heating and melting. The drying method is not particularly limited, and a drying method that can be considered by those skilled in the art, such as an air dryer, a vibration dryer, a fluidized bed dryer, a band dryer, or a rotary dryer, can be used. Drying with can also be applied. In the present invention, dry PHA refers to PHA substantially free of volatile matter, whereas undried PHA contains volatile matter, and the volatile matter is reduced by the drying method described above. Refers to PHA that can be removed.

Especially, PHA produced by microorganisms has many impurities such as proteins, polysaccharides and lipid components. The effects of the present invention can be remarkably obtained by decomposing and / or removing at least these components in an existing method, for example, the method described above. By making the amount of organic nitrogen contained in PHA preferably 500 ppm or less, more preferably 400 ppm or less, even more preferably 300 ppm or less, and even more preferably 100 ppm or less per PHA weight, the effects of the present invention can be achieved more remarkably. Since the organic nitrogen component is non-volatile, the amount of organic nitrogen contained in PHA does not change before and after the PHA drying process.

Further, as long as the pH of the undried PHA in the drying step is set to the acidic range, the step of adjusting the pH may be performed at any stage. However, it is preferably performed after decomposing and / or removing the biological component adhering to the PHA, and more preferably, it is performed on an aqueous suspension of PHA immediately before the drying step from which the biological component has been sufficiently removed. . In addition, regarding the decomposition and / or removal of the biological component, the above methods may be performed alone or in combination. These operations may be continuous or batch.

According to the present invention, PHA having a YI value of 25 or less can be produced industrially. Furthermore, PHA having a YI value of 20 or less and 10 or less can be produced industrially. Here, the YI value is a yellow index, which is an index of the color of the finally obtained dry poly-3-hydroxyalkanoic acid. The lower the yellowness, the better. The YI value in the present invention refers to a value measured for PHA cured through heating and melting. Specifically, PHA obtained by melting 2 g of PHA at 170 ° C. for 40 minutes and then curing at 60 ° C. for 60 minutes. The measured value. The lower this value, the lower the degree of coloring by heating and melting.

Since the PHA produced as described above is excellent in color tone, it can be expected to be widely applied to general-purpose products such as films and bottles. Furthermore, since PHA with a reduced amount of organic nitrogen per PHA weight is a low allergen, it can be expected to be used for medical purposes.

Hereinafter, the present invention will be described in more detail with reference to examples, but the present invention is not limited to these examples.

Example 1
The Ralstonia eutropha KNK-005 strain described in International Publication No. 2008/010296 [0049] is cultured by the method described in [0050-0053] to obtain a cell culture solution containing cells containing PHA. It was. Ralstonia and Eutropha are now classified as Capriavidas Necka.

The obtained bacterial cell culture was sterilized at 80 ° C. for 1 hour. Thereto was added 0.2% sodium dodecyl sulfate. Furthermore, after adding sodium hydroxide so that pH might be set to 11.5, it heat-retained at 50 degreeC for 1 hour. Thereafter, high-pressure crushing was performed at a pressure of 450 to 550 kgf / cm ² using a high-pressure crusher (high-pressure homogenizer model PA2K manufactured by Niroso Avi).

After centrifuging the crushed liquid after high-pressure crushing, the supernatant was removed. To the precipitated PHA, add the same amount of water as the excluded supernatant, suspend, and add 0.2% sodium dodecyl sulfate and 1/100 amount of PHA protease (Esperase, manufactured by Novozyme). The mixture was stirred for 2 hours while being kept at 50 ° C. at pH 10. PHA was recovered from the resulting reaction solution by centrifugation and washed thoroughly with water and then with methanol. The obtained aqueous suspension of PHA was adjusted so that the concentration of PHA was 10% by weight.

Thereafter, the pH of the aqueous PHA suspension was adjusted to the value shown in FIG. 1 using an appropriate amount of phosphoric acid, acetic acid, sulfuric acid, or hydrochloric acid, and after stirring for 30 minutes, PHA was recovered. The recovered undried PHA was dried at 45 ° C. for 12 hours.

2 g of dried PHA was put into a screw tube (No. 3 screw tube 21 La45 manufactured by Maruem) and heated with a block heater (EYELA MG-2200) for 40 minutes so that the internal temperature became 170 ° C., and PHA was melted. . The melted PHA was cured for 60 minutes in a thermostatic bath at 60 ° C., and then YI of this cured PHA was measured with a color difference meter SE-2000 (manufactured by Nippon Denshoku Industries Co., Ltd.). In addition, it measured by reflected light using (phi) = 10mm of a color difference meter lens. The measurement was performed in the dark. As a result, the result shown in FIG. 1 was obtained. From this, it was found that a preferred YI value (25 or less) was exhibited by drying PHA under acidic conditions of pH 3.2 or higher.

Moreover, 5M NaOH was added to the dry PHA obtained above, and a hydrolysis reaction was carried out at 95 ° C. This hydrolyzed solution was neutralized with an equal amount of 60% aqueous acetic acid solution, and an acetic acid buffer solution and a ninhydrin solution were added to perform a color reaction at 100 ° C. The absorbance of this color reaction solution was measured with a ratio beam spectrophotometer model U-1800 manufactured by Hitachi, Ltd. The amount of organic nitrogen per PHA weight in PHA before hydrolysis was calculated by comparing this absorbance with a calibration curve prepared using a leucine sample. The amount of organic nitrogen per weight of PHA before hydrolysis was approximately 500 ppm or less.

(Comparative Example 1)
10% PHA having a pH in the alkaline region (pH 8) by adding a 30% sodium hydroxide solution to the PHA purified by the method described in Example 1 using the same cell culture solution as in Example 1. An aqueous suspension of was prepared. Subsequently, PHA was recovered and dried at 45 ° C. for 12 hours. With respect to the dried PHA, YI measured by the method described in Example 1 was 29. The amount of organic nitrogen per PHA weight contained in this PHA was measured by the method described in Example 1 and found to be 381 ppm.

Claims (18)

Dry poly-3-hydroxyalkanoic acid obtained by drying undried poly-3-hydroxyalkanoic acid under acidic conditions of pH 3.2 or higher. 2. An undried poly-3-hydroxyalkanoic acid having an organic nitrogen content of 500 ppm or less per weight of poly-3-hydroxyalkanoic acid is obtained by drying under acidic conditions of pH 3.2 or more. The dry poly-3-hydroxyalkanoic acid described. 3. An undried poly-3-hydroxyalkanoic acid having an organic nitrogen content of 400 ppm or less per weight of poly-3-hydroxyalkanoic acid is obtained by drying under acidic conditions of pH 3.2 or more. The dry poly-3-hydroxyalkanoic acid described. 4. An undried poly-3-hydroxyalkanoic acid having an organic nitrogen content of 300 ppm or less per weight of poly-3-hydroxyalkanoic acid is obtained by drying under acidic conditions of pH 3.2 or more. The dry poly-3-hydroxyalkanoic acid described. Poly-3-hydroxyalkanoic acid is from 3-hydroxypropionate, 3-hydroxybutyrate, 3-hydroxyvalerate, 3-hydroxyhexanoate, 3-hydroxyheptanoate, and 3-hydroxyoctanoate The dry poly-3-hydroxyalkanoic acid according to any one of claims 1 to 4, which is a copolymer composed of two or more monomers selected from the group consisting of: 6. The dried poly-3-hydroxyalkanoic acid according to claim 5, wherein the poly-3-hydroxyalkanoic acid is a two-component polymer of 3-hydroxyhexanoate and 3-hydroxybutyrate. The dry poly-3-hydroxyalkanoic acid according to any one of claims 1 to 6, wherein the poly-3-hydroxyalkanoic acid is produced by a microorganism. The dried poly-3-hydroxyalkanoic acid according to claim 7, wherein the microorganism is Capriavidas neceta. 8. The dried poly-3-hydroxyalkanoic acid according to claim 7, wherein the microorganism is a transformant into which a poly-3-hydroxyalkanoic acid synthase gene derived from Aeromonas caviae and / or a mutant thereof is introduced. . It is obtained by drying a poly-3-hydroxyalkanoic acid aqueous suspension produced through at least one of steps (1) to (3) below under acidic conditions of pH 3.2 or higher. The dry poly-3-hydroxyalkanoic acid according to any one of claims 1 to 9.
(1) a step of disrupting cells that have accumulated poly-3-hydroxyalkanoic acid,
(2) a step of decomposing and / or removing biological components other than poly-3-hydroxyalkanoic acid,
(3) A step of washing poly-3-hydroxyalkanoic acid with water and / or an organic solvent. A method for producing poly-3-hydroxyalkanoic acid, comprising drying undried poly-3-hydroxyalkanoic acid under acidic conditions of pH 3.2 or higher. 12. The poly-3-hydroxyalkanoic acid having an organic nitrogen content of 500 ppm or less per weight of the poly-3-hydroxyalkanoic acid is dried under an acidic condition of pH 3.2 or more. A process for producing -3-hydroxyalkanoic acid. 13. The poly-3-hydroxyalkanoic acid having an organic nitrogen content of 400 ppm or less per weight of the poly-3-hydroxyalkanoic acid is dried under acidic conditions of pH 3.2 or more. A process for producing -3-hydroxyalkanoic acid. 14. The poly-3-hydroxyalkanoic acid having an organic nitrogen content of 300 ppm or less per weight of the poly-3-hydroxyalkanoic acid is dried under acidic conditions of pH 3.2 or more. A process for producing -3-hydroxyalkanoic acid. Poly-3-hydroxyalkanoic acid,
YI measured for poly-3-hydroxyalkanoic acid obtained by melting 2 g of the poly-3-hydroxyalkanoic acid at 170 ° C. for 40 minutes and then curing at 60 ° C. for 60 minutes is 25 or less. Poly-3-hydroxyalkanoic acid. The poly-3-hydroxyalkanoic acid according to claim 15, wherein the amount of organic nitrogen per weight of poly-3-hydroxyalkanoic acid is 500 ppm or less. The poly-3-hydroxyalkanoic acid according to claim 16, wherein the amount of organic nitrogen per weight of poly-3-hydroxyalkanoic acid is 400 ppm or less. 18. The poly-3-hydroxyalkanoic acid according to claim 17, wherein the amount of organic nitrogen per weight of poly-3-hydroxyalkanoic acid is 300 ppm or less.

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