JP2010095450A - Method for producing monocarboxylic acid - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for producing a monocarboxylic acid, which includes a process for removing impurities contained in a solution containing a monocarboxylic acid such as formic acid, acetic acid, etc. <P>SOLUTION: The method for producing a monocarboxylic acid includes a process A for filtering a solution containing a monocarboxylic acid through a nanofiltration membrane and recovering the monocarboxylic acid from a filtrate side and a process B for filtering a nanofiltration membrane filtrate through a reverse osmosis membrane after the process A and recovering the monocarboxylic acid from the filtrate side. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a method for producing a monocarboxylic acid, which includes a step of purifying the monocarboxylic acid. In detail, it is related with the manufacturing method of monocarboxylic acid including the process of obtaining a highly purified monocarboxylic acid containing solution from a permeation | transmission side by letting a monocarboxylic acid containing solution pass to a nanofiltration membrane.

Conventionally, when a monocarboxylic acid (for example, acetic acid) is produced by an organic synthesis method or a fermentation method, the resulting monocarboxylic acid-containing solution contains various impurities in addition to the target monocarboxylic acid. These impurities include, for example, metal catalysts when produced by an organic synthesis method, sugars such as glucose, molasses and starch, and inorganic salts such as ammonium sulfate when produced by a fermentation method, and by-products in the fermentation process. And organic acids such as lactic acid and pyruvic acid. As means for removing these impurities, adsorption separation by ion exchange resin and separation method by extraction operation are generally used. In adsorption separation by ion exchange resin, depending on the target monocarboxylic acid, the adsorption selectivity with other organic acids is poor, and a large amount of regenerated solution is used to maintain the function of the ion exchange resin. There is a problem that the waste liquid treatment is expensive. In the separation method by extraction operation, since the water solubility is high when the monocarboxylic acid is low, distribution to the organic layer is difficult, so it is necessary to use a special organic solvent to improve the recovery rate. There is a problem that it is necessary to repeatedly perform the extraction operation. Further, after the extraction operation, the organic solvent and the aqueous solution containing the organic solvent are discharged as a large amount of waste liquid, which causes a problem of an increase in waste liquid treatment cost and an increase in environmental load (Patent Document 1).
JP-A-9-151158

  The present invention solves the above-described problem, that is, the problem of effectively removing impurities without performing ion exchange and extraction operations when purifying a monocarboxylic acid, and efficiently purifies the monocarboxylic acid. It aims to provide a way to do.

  As a result of intensive studies to solve the above problems, the present inventors have found that high purity monocarboxylic acid can be obtained by filtering a monocarboxylic acid-containing solution through a nanofiltration membrane. It came to complete.

That is, this invention is comprised from following (1)-(7).
(1) A method for producing a monocarboxylic acid, comprising the step A of filtering a monocarboxylic acid-containing solution through a nanofiltration membrane and recovering the monocarboxylic acid from the permeation side.
(2) The production of the monocarboxylic acid according to (1), which comprises the step B of filtering the nanofiltration membrane permeate through the reverse osmosis membrane and recovering the monocarboxylic acid from the permeation side after the step A. Method.
(3) The method for producing a monocarboxylic acid according to (1) or (2), wherein the monocarboxylic acid is one or a mixture of two or more selected from formic acid, acetic acid, and propionic acid.
(4) The method for producing a monocarboxylic acid according to any one of (1) to (3), wherein the functional layer of the nanofiltration membrane is polyamide.
(5) The salt removal rate of the reverse osmosis membrane is 98% or more at an operating pressure of 5.5 MPa, a raw water temperature of 25 ° C., and a raw water sodium chloride concentration of 3.5%, and any one of (2) to (4) A process for producing a monocarboxylic acid as described in 1. above.
(6) The method for producing a monocarboxylic acid according to any one of (2) to (5), wherein the functional layer of the reverse osmosis membrane is polyamide.
(7) The monocarboxylic acid-containing solution recovered from the permeate in Step A or Step B is further subjected to Step C in which distillation is performed at 25 ° C. or more and 200 ° C. or less under a pressure of 1 Pa or more and atmospheric pressure. The method for producing a monocarboxylic acid according to any one of 1) to (6).

  By the method for producing a monocarboxylic acid according to the present invention, impurities other than the monocarboxylic acid contained in the monocarboxylic acid-containing solution can be effectively and economically removed by a simple operation. A monocarboxylic acid can be obtained.

  Hereinafter, the present invention will be described in more detail.

  The present invention is a method for producing a monocarboxylic acid comprising a step of purifying the monocarboxylic acid, and filters the monocarboxylic acid-containing solution through a nanofiltration membrane to remove impurities other than the monocarboxylic acid on the non-permeate side. It includes a step A of removing and recovering the monocarboxylic acid-containing solution from the permeate side.

  The monocarboxylic acid in the present invention is a compound having one carboxyl group (COOH group) in the molecule and no other functional group. Specific examples include formic acid, acetic acid, propionic acid, butanoic acid, pentane. Straight chain aliphatic saturated monocarboxylic acids such as acid, hexanoic acid, branched aliphatic saturated monocarboxylic acids such as isobutanoic acid and 2-methylbutanoic acid, straight chain aliphatic unsaturated monocarboxylic acids such as acrylic acid and crotonic acid, Examples include branched aliphatic unsaturated monocarboxylic acids such as methacrylic acid, aromatic monocarboxylic acids such as benzoic acid and phenylacetic acid. In the present invention, monocarboxylic acids having 1 to 6 carbon atoms are preferred, and formic acid is particularly preferred. One or a mixture of two or more selected from acetic acid and propionic acid is preferably purified.

  The monocarboxylic acid may be a product produced by organic synthesis or a product produced by fermentation. When the monocarboxylic acid is a product produced by fermentation, the monocarboxylic acid-containing solution may be a fermentation broth.

  The monocarboxylic acid may be dissolved as a monocarboxylate. Examples of monocarboxylic acid salts include inorganic monocarboxylic acid salts. Examples of inorganic salts here include monocarboxylic acid lithium salt, monocarboxylic acid sodium salt, monocarboxylic acid potassium salt, monocarboxylic acid magnesium salt, monocarboxylic acid calcium salt, and monocarboxylic acid ammonium salt. There may be.

  The nanofiltration membrane used in the present invention is also called a nanofiltration membrane or NF membrane, and is a membrane generally defined as “a membrane that transmits monovalent ions and blocks divalent ions”. . It is a membrane that is considered to have a minute gap of about several nanometers, and is mainly used to block minute particles, molecules, ions, salts, and the like in water.

  Also, “through the nanofiltration membrane” means that the monocarboxylic acid-containing solution is filtered through the nanofiltration membrane to remove impurities other than the monocarboxylic acid to the non-permeate side, and the monocarboxylic acid from the permeate side. This means that the contained solution is recovered.

  As a material for the nanofiltration membrane used in the present invention, a polymer material such as cellulose acetate polymer, polyamide, polyester, polyimide, vinyl polymer can be used. It is not limited to this, and a film including a plurality of film materials may be used. In addition, the membrane structure has a dense layer on at least one side of the membrane, and on the asymmetric membrane having fine pores gradually increasing from the dense layer to the inside of the membrane or the other side, or on the dense layer of the asymmetric membrane. Either a composite film having a very thin functional layer formed of another material may be used. As the composite membrane, for example, a composite membrane described in JP-A-62-201606 in which a nanofiltration membrane composed of a functional layer of polyamide is formed on a support membrane made of polysulfone as a membrane material can be used.

  In the present invention, among them, a composite film having a high pressure resistance, high water permeability, and high solute removal performance and having an excellent potential and using polyamide as a functional layer is preferable. Further, in order to maintain durability against operating pressure, high water permeability, and blocking performance, a structure in which polyamide is used as a functional layer and is held by a support made of a porous film or a nonwoven fabric is preferable. In the nanofiltration membrane having a polyamide as a functional layer, preferable carboxylic acid components of monomers constituting the polyamide include, for example, trimesic acid, benzophenonetetracarboxylic acid, trimellitic acid, pyrometic acid, isophthalic acid, terephthalic acid, naphthalene Aromatic carboxylic acids such as dicarboxylic acid, diphenyl carboxylic acid, pyridine carboxylic acid and the like can be mentioned, but considering solubility in a film forming solvent, trimesic acid, isophthalic acid, terephthalic acid, or a mixture thereof is more preferable.

  Preferred amine components of the monomers constituting the polyamide include m-phenylenediamine, p-phenylenediamine, benzidine, methylenebisdianiline, 4,4′-diaminobiphenyl ether, dianisidine, 3,3 ′, 4- Triaminobiphenyl ether, 3,3 ′, 4,4′-tetraaminobiphenyl ether, 3,3′-dioxybenzidine, 1,8-naphthalenediamine, m (p) -monomethylphenylenediamine, 3,3′- Monomethylamino-4,4′-diaminobiphenyl ether, 4, N, N ′-(4-aminobenzoyl) -p (m) -phenylenediamine-2,2′-bis (4-aminophenylbenzimidazole), 2 , 2′-bis (4-aminophenylbenzoxazole), 2,2′-bis (4-aminophenyl) Secondary diamines such as primary diamines having an aromatic ring such as nilbenzothiazole), piperazine, piperidine or derivatives thereof, among which nanofiltration membranes having a functional layer of a crosslinked polyamide containing piperazine or piperidine as a monomer are It is preferably used because it has heat resistance and chemical resistance in addition to pressure resistance and durability. More preferably, it is a nanofiltration membrane mainly composed of the crosslinked piperazine polyamide or the crosslinked piperidine polyamide. Examples of nanofiltration membranes having a functional layer of polyamide containing piperazine polyamide include those described in JP-A No. 62-201606, and specific examples thereof include a crosslinked piperazine polyamide-based half made by Toray Industries, Inc. Examples include permeable membrane UTC60.

  The nanofiltration membrane is generally used as a spiral membrane element, but the nanofiltration membrane used in the present invention can also be preferably used as a spiral membrane element. Specific examples of preferred nanofiltration membranes include, for example, UTC60 manufactured by Toray Industries, Inc., which has a crosslinked piperazine polyamide as a main component and a polyamide containing a component represented by the chemical formula (2) as a functional layer. Nano filter modules SU-210, SU-220, SU-600, and SU-610 can also be used. In addition, NF-45, NF-90, NF-200, NF-400, which is a nanofiltration membrane manufactured by Filmtec Co., which uses a crosslinked piperazine polyamide as a functional layer, or NF99, which is a nanofiltration membrane manufactured by Alfa Laval, which uses polyamide as a functional layer. GE97, a nanofiltration membrane manufactured by GE Osmonics, which is a cellulose acetate-based nanofiltration membrane, and the like.

  In the present invention, the filtration of the monocarboxylic acid-containing solution with the nanofiltration membrane may be performed under pressure. If the filtration pressure is lower than 0.1 MPa, the membrane permeation rate decreases, and if it is higher than 8 MPa, the membrane is damaged. Therefore, the filtration pressure is preferably used in the range of 0.1 MPa to 8 MPa, but 0.5 MPa to 7 MPa. When used below, the membrane permeation flux is high, so that the monocarboxylic acid can be efficiently permeated and is less likely to affect the membrane damage. Is particularly preferred.

  In the present invention, the filtration of the monocarboxylic acid-containing solution with the nanofiltration membrane can improve the recovery rate of the monocarboxylic acid by returning the non-permeate to the raw water again and repeatedly filtering. The recovery rate of monocarboxylic acid can be calculated by Equation 1 by measuring the total amount of monocarboxylic acid before nanofiltration and the total amount of monocarboxylic acid permeating through the nanofiltration membrane.

  Monocarboxylic acid recovery rate (%) = (total amount of monocarboxylic acid permeating nanofiltration membrane / total amount of monocarboxylic acid before nanofiltration) × 100 (Formula 1).

  The membrane separation performance of the nanofiltration membrane used in the present invention is 45% or more when the sodium chloride aqueous solution (500 mg / L) adjusted to a temperature of 25 ° C. and pH 6.5 is evaluated at a filtration pressure of 0.75 MPa. Are preferably used. The salt removal rate here can be calculated by Equation 2 by measuring the permeate salt concentration of the aqueous sodium chloride solution.

  Salt removal rate = 100 × {1− (salt concentration in permeate / salt concentration in feed water)} (Formula 2).

The permeation performance of the nanofiltration membrane is that the membrane permeation flux (m ³ / (m ² · day)) of an aqueous sodium chloride solution (500 mg / L) is 0.3 or more at a filtration pressure of 0.3 MPa. Is preferably used. The membrane permeation flux can be calculated by Equation 3 by measuring the permeate amount and the time when the permeate amount was collected and the membrane area.

Membrane permeation flux (m ³ / (m ² · day)) = permeate amount / (membrane area × water sampling time) (Equation 3).

  In the present invention, it is preferable to adjust the pH to 1 or more and less than 7 by adding an acidic substance to the monocarboxylic acid-containing solution that leads to the nanofiltration membrane. In nanofiltration, a non-ionized (non-dissociated) substance in a solution is easier to permeate than an ionized (dissociated) substance, so that the pH of an aqueous solution containing a monocarboxylic acid is less than 7. By doing so, the proportion of the monocarboxylic acid that is not ionized in the aqueous solution containing the monocarboxylic acid is larger than the monocarboxylic acid that is ionized (non-dissociated monocarboxylic acid / dissociated monocarboxylic acid> 1), The monocarboxylic acid solution can be efficiently recovered from the permeate side. If the pH of the aqueous solution containing the monocarboxylic acid is less than 1, the durability of the nanofiltration membrane may be adversely affected. A more preferable pH range is 1 or more and 6 or less. Examples of the acidic substance added as a pH adjuster include sulfuric acid, hydrochloric acid, carbonic acid, phosphoric acid, and nitric acid.

  In the present invention, impurities separated from the monocarboxylic acid-containing solution to the non-permeate side by the nanofiltration membrane include inorganic substances such as calcium, sodium, sulfuric acid, nitric acid, phosphoric acid, glucose, fructose, xylose, sucrose, galactose, Examples include saccharides such as starch, proteins, and the like, and even a mixture thereof is preferably separated.

  The nanofiltration membrane permeability of the solution containing the monocarboxylic acid in the present invention can be evaluated by calculating the monocarboxylic acid permeability. The monocarboxylic acid permeability is determined by analyzing the concentration of monocarboxylic acid (raw water monocarboxylic acid concentration) in the raw water (solution containing the monocarboxylic acid) and the permeated liquid based on analysis represented by high performance liquid chromatography and gas chromatography. By calculating the monocarboxylic acid concentration (permeate monocarboxylic acid concentration) contained in the (monocarboxylic acid solution), it can be calculated by Equation 4.

  Monocarboxylic acid permeability (%) = (permeate monocarboxylic acid concentration / raw water monocarboxylic acid concentration) × 100 (Equation 4).

  In the present invention, the permeate obtained in the step A is further passed through a reverse osmosis membrane to be subjected to a step B in which the monocarboxylic acid-containing solution is recovered from the permeate side, so that the impurities partially permeated through the nanofiltration membrane are substantially eliminated. It can be completely removed.

  In the present invention, “permeating the nanofiltration membrane through the reverse osmosis membrane” means that the monocarboxylic acid solution that has passed through the nanofiltration membrane is filtered through the reverse osmosis membrane, and the permeate side contains monocarboxylic acid. This means that the solution is recovered by filtration and impurities other than the monocarboxylic acid are removed on the non-permeate side.

  Here, as the membrane separation performance of the reverse osmosis membrane used in the present invention, sodium chloride adjusted to a temperature of 25 ° C. and pH 6.5 (raw water sodium chloride concentration of 3.5%) was evaluated at a filtration pressure of 5.5 MPa. Those having a sodium chloride removal rate of 98% or more are preferably used. The sodium chloride removal rate can be calculated by Equation 5 by measuring the permeate sodium chloride concentration in the seawater.

  Sodium chloride removal rate = 100 × {1− (sodium chloride concentration in permeate / sodium chloride concentration in raw water)} (Formula 5).

The permeation performance of the reverse osmosis membrane is such that sodium chloride (3.5%) has a membrane permeation flux (m ³ / (m ² · day)) of 0.2 or more at a filtration pressure of 5.5 MPa. If it is, it can be preferably used because it can speed up the separation of the permeate side monocarboxylic acid and the non-permeate side impurities.

  As the membrane material of the reverse osmosis membrane used in the present invention, generally available polymer materials such as cellulose acetate polymer, polyamide, polyester, polyimide, vinyl polymer can be used. It is not limited to the film | membrane comprised by these, The film | membrane containing a some film | membrane raw material may be sufficient. In addition, the membrane structure has a dense layer on at least one side of the membrane, and on the asymmetric membrane having fine pores gradually increasing from the dense layer to the inside of the membrane or the other side, or on the dense layer of the asymmetric membrane. Either a composite film having a very thin functional layer formed of another material may be used.

  As a reverse osmosis membrane preferably used in the present invention, a composite membrane using a cellulose acetate-based polymer as a functional layer (hereinafter also referred to as a cellulose acetate-based reverse osmosis membrane) or a composite membrane using a polyamide as a functional layer (hereinafter, Polyamide-based reverse osmosis membrane). Here, as the cellulose acetate-based polymer, organic acid esters of cellulose such as cellulose acetate, cellulose diacetate, cellulose triacetate, cellulose propionate, cellulose butyrate and the like, or a mixture thereof and those using mixed esters can be mentioned. It is done. The polyamide includes a linear polymer or a crosslinked polymer having an aliphatic and / or aromatic diamine as a monomer.

  In the present invention, the membrane material is not particularly limited as long as it is a reverse osmosis membrane having a high monocarboxylic acid permeability. However, if it is a polyamide-based reverse osmosis membrane, the recovery rate of monocarboxylic acid is high and it is separated from impurities. It is more preferable because of its high efficiency.

  As the membrane form, an appropriate form such as a flat membrane type, a spiral type, and a hollow fiber type can be used.

  Specific examples of the reverse osmosis membrane used in the present invention include, for example, Toray Industries, Ltd. polyamide-based reverse osmosis membrane (UTC) SU-710, SU-720, SU-720F, SU-710L, SU-720L, SU-720LF, SU-720R, SU-710P, SU-720P, SU-810, SU-820, SU-820L, SU-820FA, the company's cellulose acetate reverse osmosis membrane SC-L100R, SC-L200R, SC-1100 SC-1200, SC-2100, SC-2200, SC-3100, SC-3200, SC-8100, SC-8200, NTR-759HR, NTR-729HF, NTR-70SWC, ES10-D manufactured by Nitto Denko Corporation , ES20-D, ES20-U, ES15-D, ES15-U, LF10-D, Alfa Laval O98pHt, RO99, HR98PP, CE4040C-30D, GE made GE Sepa, Filmtec made BW30-4040, TW30-4040, XLE-4040, LP-4040, LE-4040, SW30-4040, like SW30HRLE-4040.

  In the present invention, filtration of the nanofiltration membrane permeate with a reverse osmosis membrane is performed by applying pressure, but if the filtration pressure is lower than 1 MPa, the membrane permeation rate decreases, and if it is higher than 8 MPa, the membrane damage is affected. In order to give, it is preferable that it is the range of 1 MPa or more and 8 MPa or less. Also, if the filtration pressure is in the range of 1 MPa or more and 7 MPa or less, the membrane permeation flux is high, so that the monocarboxylic acid solution can be efficiently permeated and there is little possibility of affecting the membrane damage. It is more preferable that it is in the range of 2 MPa or more and 6 MPa or less.

  In the present invention, impurities other than the monocarboxylic acid separated from the monocarboxylic acid-containing solution by the reverse osmosis membrane to the non-permeate side include organic acids such as lactic acid, pyruvic acid, butyric acid, succinic acid, sodium chloride, ammonia, etc. An inorganic salt is mentioned.

  The concentration of the monocarboxylic acid-containing solution to be subjected to separation by the reverse osmosis membrane is not particularly limited, but a high concentration is suitable for cost reduction because the filtration time can be shortened, for example, 10 g / L. The amount is preferably 100 g / L or less.

  In the present invention, the permeate from Step A or Step B may be subjected to another monocarboxylic acid purification step. Preferably, the monocarboxylic acid-containing solution is subjected to a further distillation step, whereby a high-purity monocarboxylic acid is obtained. Carboxylic acid can be obtained. The distillation step is preferably performed under a reduced pressure of 1 Pa or more and atmospheric pressure (normal pressure, about 101 kPa), more preferably 100 Pa or more and 15 kPa or less. When the distillation is performed under reduced pressure, the distillation temperature is preferably 20 ° C. or higher and 200 ° C. or lower, more preferably 50 ° C. or higher and 150 ° C. or lower.

  As a form of the separation membrane device of the nanofiltration membrane or reverse osmosis membrane of the present invention, a raw water tank for storing a monocarboxylic acid-containing solution, a high-pressure pump for providing a driving force for filtration, and a nanofiltration membrane for mounting It is mainly composed of cells.

  FIG. 1 is a schematic view for explaining an example of a nanofiltration membrane separation membrane device that can be used in the present invention. FIG. 2 is a cell cross-sectional schematic diagram for explaining an example in which a nanofiltration membrane of a nanofiltration membrane separation membrane device that can be used in the present invention is mounted. Next, the form of purification of monocarboxylic acid by the separation membrane device of the nanofiltration membrane of FIG. 1 will be described. The nanofiltration membrane 7 is attached to the cell 2 using the support plate 8. Next, the monocarboxylic acid-containing solution is charged into the raw water tank 1 and the monocarboxylic acid-containing solution is fed to the cell by the high-pressure pump 3 to purify the monocarboxylic acid. The filtration pressure by the high pressure pump 3 can be 0.1 MPa or more and 8 MPa or less. Preferably, it is 0.5 MPa or more and 7 MPa or less, and it is particularly preferably used at 1 MPa or more and 6 MPa or less. The aqueous solution containing the monocarboxylic acid is sent to the cell 2 to obtain the permeate 5 containing the monocarboxylic acid. The concentrated liquid 4 concentrated in the cell is returned to the raw water tank 1 again. At this time, it is also possible to continuously purify the monocarboxylic acid by introducing a new amount of monocarboxylic acid equivalent to the permeate into the raw water tank (not shown). Thus, the monocarboxylic acid which is a desired product and impurities are separated from the monocarboxylic acid-containing solution, and the monocarboxylic acid can be easily purified. In addition, filtration with a reverse osmosis membrane can also be performed by a separation membrane device equivalent to a separation device with a nanofiltration membrane.

  According to the method for producing monocarboxylic acid of the present invention, separation and purification of monocarboxylic acid by membrane separation operation is suitable for energy cost reduction because it does not require a heating step, and organic solvent waste liquid is not discharged at all. It is suitable for reducing the environmental load.

  EXAMPLES Hereinafter, although this invention is demonstrated in detail using an Example, this invention is not limited to a following example.

(Production of yeast strains capable of producing lactic acid)
Reference Example 1 Production of Yeast Strain Having Lactic Acid Production Capability A lactic acid fermentation broth containing acetic acid as a by-product was used as an acetic acid-containing solution for use in the separation and purification experiment of acetic acid, which is a monocarboxylic acid. Yeast strains capable of producing lactic acid were constructed as follows. Specifically, a yeast strain having L-lactic acid production ability was constructed by linking a human-derived L-LDH gene downstream of the PDC1 promoter on the yeast genome. For polymerase chain reaction (PCR), La-Taq (manufactured by Takara Shuzo) or KOD-Plus-polymerase (manufactured by Toyobo Co., Ltd.) was used according to the attached instruction manual.

  After culturing and recovering human breast cancer cell line (MCF-7), total RNA was extracted using TRIZOL Reagent (Invitrogen), and SuperScript Choice System (Invitrogen) was used with the obtained total RNA as a template. CDNA was synthesized by reverse transcription reaction. Details of these operations followed the attached protocol. The obtained cDNA was used as an amplification template for subsequent PCR.

  The L-ldh gene was cloned by PCR using KOD-Plus-polymerase using the cDNA obtained by the above operation as an amplification template and the oligonucleotides represented by SEQ ID NO: 1 and SEQ ID NO: 2 as a primer set. Each PCR amplified fragment was purified, and the end was phosphorylated with T4 Polynucleotide Kinase (manufactured by TAKARA), and then ligated to a pUC118 vector (which was cleaved with the restriction enzyme HincII and the cut surface was dephosphorylated). Ligation was performed using DNA Ligation Kit Ver.2 (manufactured by TAKARA). Escherichia coli DH5α was transformed with the ligation plasmid product, and plasmid DNA was recovered to obtain a plasmid in which various L-LDH genes (accession number; AY009108, SEQ ID NO: 3) were subcloned. The obtained pUC118 plasmid into which the L-LDH gene was inserted was digested with restriction enzymes XhoI and NotI, and the resulting DNA fragments were inserted into the XhoI / NotI cleavage sites of the yeast expression vector pTRS11. Thus, a human-derived L-LDH gene expression plasmid pL-LDH5 (L-LDH gene) was obtained.

  By PCR using the plasmid pL-LDH5 containing the human-derived LDH gene as an amplification template and the oligonucleotides represented by SEQ ID NO: 4 and SEQ ID NO: 5 as primer sets, a 1.3-kb human-derived LDH gene and Saccharomyces cerevisiae derived A DNA fragment containing the terminator sequence of the TDH3 gene was amplified. In addition, a DNA fragment containing the TRP1 gene derived from Saccharomyces cerevisiae of 1.2 kb was amplified by PCR using the plasmid pRS424 as an amplification template and the oligonucleotides represented by SEQ ID NO: 6 and SEQ ID NO: 7 as a primer set. Each DNA fragment was separated by 1.5% agarose gel electrophoresis and purified according to a conventional method. A product obtained by the PCR method using a mixture of the 1.3 kb fragment and the 1.2 kb fragment obtained here as an amplification template and the oligonucleotides represented by SEQ ID NO: 4 and SEQ ID NO: 7 as a primer set is 1 2.5% agarose gel electrophoresis was carried out to prepare a 2.5 kb DNA fragment linked with human-derived LDH gene and TRP1 gene according to a conventional method. Saccharomyces cerevisiae strain NBRC10505 was transformed with this 2.5 kb DNA fragment in a conventional manner so as not to require tryptophan.

  Confirmation that the obtained transformed cells were cells in which the human-derived LDH gene was linked downstream of the PDC1 promoter on the yeast genome was performed as follows. First, genomic DNA of a transformed cell was prepared according to a conventional method, and a 0.7 kb amplified DNA fragment was obtained by PCR using the oligonucleotides represented by SEQ ID NO: 8 and SEQ ID NO: 9 as a primer set. Confirmed by being. In addition, whether or not the transformed cell has lactic acid production ability is shown below that lactic acid is contained in the culture supernatant obtained by culturing the transformed cell in SC medium (METHODS IN YEAS GENETIC 2000 EDITION, CSHL PRESS). It confirmed by measuring the amount of lactic acid by HPLC method on conditions. The concentration of acetic acid was also analyzed by the HPLC method under the following conditions.

  Column: Shim-Pack SPR-H (manufactured by Shimadzu Corporation), mobile phase: 5 mM p-toluenesulfonic acid (flow rate 0.8 mL / min), reaction solution: 5 mM p-toluenesulfonic acid, 20 mM Bistris, 0.1 mM EDTA · 2Na (flow rate 0.8 mL / min), detection method: electrical conductivity, temperature: 45 ° C.

  As a result of HPLC analysis, 4 g / L L-lactic acid and 1 g / L acetic acid as a by-product were detected. The obtained transformed cells were used in subsequent examples as yeast strain SW-1.

Reference Example 2 Batch Fermentation Test for Lactic Acid-Producing Yeast A batch fermentation test was performed using the fermentation medium shown in Table 1. The medium was used after autoclaving (121 ° C., 15 minutes). The yeast SW-1 strain constructed in Reference Example 1 was used as a microorganism, and the concentration of lactic acid as a product was evaluated using the HPLC shown in Reference Example 1, and the glucose concentration was measured using a glucose test Wako. C (Wako Pure Chemical Industries) was used. The operating conditions of Reference Example 2 are shown below.

  Reaction tank capacity (lactic acid fermentation medium amount): 2 (L), temperature adjustment: 30 (° C.), reaction tank aeration rate: 0.2 (L / min), reaction tank stirring speed: 400 (rpm), pH adjustment: Adjust to pH 5 with 1N calcium hydroxide.

  First, the SW-1 strain was cultured with shaking in a 5 ml lactic acid fermentation medium overnight in a test tube (pre-culture). The culture solution was inoculated into 100 ml of fresh lactic acid fermentation medium and cultured with shaking in a 500 ml Sakaguchi flask for 24 hours (pre-culture). Temperature adjustment and pH adjustment were performed, and fermentation culture was performed. The amount of bacterial cell growth at this time was 15 in terms of absorbance at 600 nm. The results of batch fermentation are shown in Table 2.

Reference Example 3 Evaluation of inorganic salt (magnesium sulfate) rejection rate of nanofiltration membrane 10 g of magnesium sulfate (manufactured by Wako Pure Chemical Industries, Ltd.) was added to 10 L of ultrapure water and stirred at 25 ° C. for 1 hour to prepare a 1000 ppm magnesium sulfate aqueous solution. . Next, 10 L of the magnesium sulfate aqueous solution prepared above was injected into the raw water tank 1 of the membrane filtration apparatus shown in FIG. As a 90φ nanofiltration membrane indicated by reference numeral 7 in FIG. 2, a crosslinked piperazine polyamide semipermeable membrane “UTC60” (Nanofiltration membrane 1; manufactured by Toray Industries, Inc.), a crosslinked piperazine polyamide semipermeable membrane “NF-400” (nanofiltration membrane) 2; Film Tech), polyamide semipermeable membrane “NF99” (nanofiltration membrane 3; manufactured by Alfa Laval), cellulose acetate semipermeable membrane “GEsepa” (nanofiltration membrane 4; manufactured by GE Osmonics), respectively, stainless steel (manufactured by SUS316) The raw water temperature was adjusted to 25 ° C., the pressure of the high-pressure pump 3 was adjusted to 0.5 MPa, and the permeate 5 was collected. The concentration of magnesium sulfate contained in the raw water tank 1 and the permeate 5 was analyzed by ion chromatography (manufactured by DIONEX) under the following conditions, and the transmittance of magnesium sulfate was calculated.

  Anion; column (AS4A-SC (manufactured by DIONEX)), eluent (1.8 mM sodium carbonate / 1.7 mM sodium bicarbonate), temperature (35 ° C.).

  Cation; column (CS12A (manufactured by DIONEX)), eluent (20 mM methanesulfonic acid), temperature (35 ° C.).

  The results are shown in Table 3.

  From the results of Table 3, it was shown that UTC60 (nanofiltration membrane 1: manufactured by Toray Industries, Inc.) has the highest inorganic salt rejection.

Reference Example 4 Acetic Acid Permeability Evaluation of Nanofiltration Membrane 100 g of acetic acid (Wako Pure Chemical Industries, Ltd.) was added to 10 L of ultrapure water and stirred for 1 hour at 25 ° C. to prepare a 1000 ppm aqueous acetic acid solution. Subsequently, the permeate of the nanofiltration membranes 1 to 4 was collected under the same conditions as in Reference Example 1. The acetic acid concentration contained in the raw water tank 1 and the permeate 5 was analyzed by high performance liquid chromatography (manufactured by Shimadzu Corporation) under the following conditions, and the acetic acid permeability was calculated.

  Column: Shim-Pack SPR-H (manufactured by Shimadzu Corporation), mobile phase: 5 mM p-toluenesulfonic acid (flow rate 0.8 mL / min), reaction solution: 5 mM p-toluenesulfonic acid, 20 mM Bistris, 0.1 mM EDTA · 2Na (flow rate 0.8 mL / min), detection method: electrical conductivity, temperature: 45 ° C.

  The results are shown in Table 4.

Examples 1-21 Acetic acid separation experiment by nanofiltration membrane (Preparation of culture solution to be filtered through nanofiltration membrane)
The culture solution (2 L) fermented in Reference Examples 1 and 2 has a pH of 1.9 (Example 1) 2.0 (Examples 2 to 5), 2.2 (Examples 6 to 9), 2.6 ( Examples 10-16) After dripping concentrated sulfuric acid (manufactured by Wako Pure Chemical Industries, Ltd.) until 4.0 (Examples 17-20), the mixture was stirred at 25 ° C. for 1 hour, and calcium lactate in the culture solution, Calcium acetate was converted to lactic acid, acetic acid and calcium sulfate. Next, the precipitated calcium sulfate was filtered by suction filtration using qualitative filter paper No. 2 (manufactured by Advantech Co., Ltd.), and 2 L of filtrate was collected. In addition, a separation experiment was performed on a culture solution (2 L) having a pH of 5 to which no concentrated sulfuric acid was added (Example 21).

(Separation experiment with nanofiltration membrane)
Next, 2 L of the filtrate obtained in the above example was injected into the raw water tank 1 of the membrane filtration apparatus shown in FIG. As the 90φ nanofiltration membrane denoted by reference numeral 7 in FIG. 2, the nanofiltration membranes 1 to 4 are respectively set in stainless steel (SUS316) cells, and the pressure of the high-pressure pump 3 is adjusted to 1 MPa, 3 MPa, 4 MPa, and 5 MPa, respectively. The permeate 5 at each pressure was recovered. The concentration of sulfate ions and calcium ions contained in the raw water tank 1 and the permeate 5 are the same as those in Reference Example 3, and ion chromatography (manufactured by DIONEX), glucose, lactic acid, and acetic acid concentrations are the same as in Reference Example 1. Analysis was performed by high performance liquid chromatography (manufactured by Shimadzu Corporation) under the conditions. The results are shown in Table 5.

  As shown in Table 5, it was found that calcium sulfate, glucose and lactic acid were removed with high efficiency at all pH and filtration pressures. In Examples 1 to 21, calcium sulfate was removed with high efficiency in the filtration and pressurization without replacing the nanofiltration membrane with a new membrane.

Reference Example 5 Evaluation of sodium chloride removal property of reverse osmosis membrane Sodium chloride (manufactured by Wako Pure Chemical Industries, Ltd.) was added to 10 L of ultrapure water and stirred for 1 hour at 25 ° C. to adjust 3.5% sodium chloride. Next, 10 L of 3.5% sodium chloride prepared above was injected into the raw water tank 1 of the membrane filtration apparatus shown in FIG. As a 90φ reverse osmosis membrane indicated by reference numeral 7 in FIG. 2, a polyamide-based reverse osmosis membrane “UTC70” (reverse osmosis membrane 1; manufactured by Toray Industries, Inc.), a polyamide-based reverse osmosis membrane “RO-99” (reverse osmosis membrane 2; Alfa Laval), polyamide reverse osmosis membrane “HR98PP” (reverse osmosis membrane 3; manufactured by Alfa Laval), polyamide reverse osmosis membrane “GEsepa PA-AK” (reverse osmosis membrane 4; manufactured by GE Osmonics), cellulose acetate reverse The osmosis membrane “GEsepa CF-CA” (reverse osmosis membrane 5; manufactured by GE Osmonics) was set in a cell made of stainless steel (SUS316), and the raw water temperature was adjusted to 25 ° C. and the pressure of the high-pressure pump 3 was adjusted to 5.5 MPa. The permeate 5 was collected. The concentration of sodium chloride contained in the raw water tank 1 and the permeate 5 was analyzed by ion chromatography (manufactured by DIONEX) under the following conditions, and the transmittance of sodium chloride was calculated.

  Anion; column (AS4A-SC (manufactured by DIONEX)), eluent (1.8 mM sodium carbonate / 1.7 mM sodium bicarbonate), temperature (35 ° C.).

  Cation; column (CS12A (manufactured by DIONEX)), eluent (20 mM methanesulfonic acid), temperature (35 ° C.).

  The results are shown in Table 6.

Examples 22 to 41 Acetic acid separation experiment using reverse osmosis membrane 2 L of nanofiltration membrane permeate obtained in Examples 5 to 25 was injected into the raw water tank 1 of the membrane filtration apparatus shown in FIG. As the 90φ reverse osmosis membrane of reference numeral 7 in FIG. 2, the reverse osmosis membranes 1 to 5 are respectively set in cells made of stainless steel (made of SUS316), and the pressure of the high pressure pump 3 is adjusted to 1 MPa, 2 MPa, 4 MPa, and 5 MPa, respectively. The permeate 5 at each pressure was recovered. The concentration of lactic acid and acetic acid contained in the raw water tank 1 and permeate 5 was analyzed by high performance liquid chromatography (manufactured by Shimadzu Corporation) under the same conditions as in Reference Example 1, and recovered from the permeation side of the reverse osmosis membrane. The acetic acid recovery rate was calculated by the method of Formula 6, and the lactic acid content in the acetic acid aqueous solution recovered from the permeation side of the reverse osmosis membrane was calculated by the method of Formula 7.

  Acetic acid recovery rate (%) = total acetic acid recovered from the permeate side / total acetic acid injected into the raw water tank (Formula 6).

Lactic acid content (%) in recovered aqueous acetic acid solution = recovered lactic acid content / recovered total acetic acid amount (Equation 7).
The results are shown in Table 7.

  As shown in Table 7, it was found that high-purity acetic acid was recovered on the permeate side with high efficiency at all filtration pressures of 1 MPa, 2 MPa, 4 MPa, and 5 MPa.

Examples 42 to 57 Formic acid separation experiment using nanofiltration membrane Model aqueous solution in which formic acid (10 g / L), lactic acid (10 g / L), glucose (1 g / L), calcium sulfate (1 g / L) was added to ultrapure water ( 2L) was injected into the raw water tank 1 of the membrane filtration apparatus shown in FIG. As the 90φ nanofiltration membrane denoted by reference numeral 7 in FIG. 2, the nanofiltration membranes 1 to 4 are respectively set in stainless steel (SUS316) cells, and the pressure of the high-pressure pump 3 is adjusted to 1 MPa, 3 MPa, 4 MPa, and 5 MPa, respectively. The permeate 5 at each pressure was recovered. Concentration of sulfate ion and calcium ion contained in raw water tank 1 and permeate 5 under the same conditions as in Reference Example 3, ion chromatography (manufactured by DIONEX), glucose, lactic acid and formic acid concentrations as in Reference Example 1 Analysis was performed by high performance liquid chromatography (manufactured by Shimadzu Corporation) under the conditions. The results are shown in Table 8.

Examples 58-77 Formic acid separation experiment using reverse osmosis membrane 2 L of nanofiltration membrane permeate obtained in Examples 42-57 was injected into the raw water tank 1 of the membrane filtration apparatus shown in FIG. As the 90φ reverse osmosis membrane of reference numeral 7 in FIG. 2, the reverse osmosis membranes 1 to 5 are respectively set in cells made of stainless steel (made of SUS316), and the pressure of the high pressure pump 3 is adjusted to 1 MPa, 2 MPa, 4 MPa, and 5 MPa, respectively. The permeate 5 at each pressure was recovered. The concentration of lactic acid and formic acid contained in raw water tank 1 and permeate 5 was analyzed by high performance liquid chromatography (manufactured by Shimadzu Corporation) under the same conditions as in Reference Example 1, and recovered from the permeation side of the reverse osmosis membrane. The formic acid recovery rate was calculated by the method of Formula 8, and the lactic acid content in the acetic acid aqueous solution recovered from the permeation side of the reverse osmosis membrane was calculated by the method of Formula 9.

  Formic acid recovery rate (%) = total amount of formic acid recovered from the permeate side / total amount of formic acid injected into the raw water tank (Equation 8).

Lactic acid content (%) in recovered formic acid aqueous solution = recovered lactic acid content / recovered total formic acid amount (Equation 9).
The results are shown in Table 9.

  As shown in Table 9, it was found that high-purity formic acid was recovered on the permeate side with high efficiency at all filtration pressures of 1 MPa, 2 MPa, 4 MPa, and 5 MPa.

Comparative Example 1 Purification of acetic acid without using a nanofiltration membrane In the same manner as in Example 1, concentrated sulfuric acid (Wako Pure Chemical Industries, Ltd.) was used until the pH of the fermentation broth (2 L) cultured in Reference Examples 1 and 2 was 1.9. The product was stirred at 25 ° C. for 1 hour to convert calcium lactate and calcium acetate in the culture solution into lactic acid, acetic acid and calcium sulfate. Next, the precipitated calcium sulfate was filtered by suction filtration using qualitative filter paper No. 2 (manufactured by Advantech Co., Ltd.), and 2 L of filtrate was collected. Subsequently, 2 L of the filtrate was concentrated by evaporating water under reduced pressure (50 hPa) using a rotary evaporator (manufactured by Tokyo Rika). At this time, precipitation of calcium sulfate was observed. The obtained filtrate contained lactic acid 45 g / L and acetic acid 12 g / L.

Comparative Example 2 Purification of acetic acid by extraction operation 2 L of chloroform (manufactured by Wako Pure Chemical Industries, Ltd.) was added to 2 L of the filtrate obtained in Comparative Example 1, and extracted with a separatory funnel. When the acetic acid concentration contained in the extracted organic layer and aqueous layer was analyzed by HPLC, the acetic acid recovery rate to the organic layer was 10%. Chloroform 2L was again added to the aqueous layer after extraction, and extraction was repeated with a separatory funnel, but the recovery rate did not exceed 40%. Moreover, the organic solvent waste liquid of 10 L or more was produced by extraction operation.

  From the results of the above Examples and Comparative Examples, it was clarified that the nanofiltration membrane can remove impurities from the acetic acid-containing solution with high efficiency and recover acetic acid with high yield. That is, according to the present invention, it was clarified that the monocarboxylic acid can be purified in a high yield without performing an extraction operation using an organic solvent by filtering the monocarboxylic acid-containing solution using a nanofiltration membrane. .

It is a schematic diagram showing one embodiment of the nanofiltration membrane and reverse osmosis membrane separation device used in the present invention. It is the schematic which shows one Embodiment of the cell sectional view with which the reverse osmosis membrane of the nanofiltration membrane and reverse osmosis membrane separation apparatus used by this invention was mounted | worn.

Explanation of symbols

1 Raw Water Tank 2 Cell with Nanofiltration Membrane or Reverse Osmosis Membrane 3 High Pressure Pump 4 Flow of Membrane Concentrate 5 Flow of Membrane Permeate 6 Flow of Culture Solution or Nanofiltration Membrane Permeate Flowed by High Pressure Pump 7 Nanofiltration membrane or reverse osmosis membrane 8 Support plate

Claims (7)

  A method for producing a monocarboxylic acid, comprising the step A of filtering a monocarboxylic acid-containing solution through a nanofiltration membrane and recovering the monocarboxylic acid from the permeation side.   The manufacturing method of monocarboxylic acid of Claim 1 including the process B which collect | recovers monocarboxylic acid from the permeation | transmission side by filtering a nanofiltration-membrane permeation | transmission liquid through a reverse osmosis membrane after the said process A.   The method for producing a monocarboxylic acid according to claim 1 or 2, wherein the monocarboxylic acid is one or a mixture of two or more selected from formic acid, acetic acid and propionic acid.   The method for producing a monocarboxylic acid according to any one of claims 1 to 3, wherein the functional layer of the nanofiltration membrane is polyamide.   The monocarboxylic acid according to any one of claims 2 to 4, wherein a salt removal rate of the reverse osmosis membrane is 98% or more at an operating pressure of 5.5 MPa, a raw water temperature of 25 ° C, and a raw water sodium chloride concentration of 3.5%. Acid production method.   The method for producing a monocarboxylic acid according to any one of claims 2 to 5, wherein the functional layer of the reverse osmosis membrane is polyamide.   The monocarboxylic acid-containing solution recovered from the permeate in Step A or Step B is further subjected to Step C in which distillation is performed at 25 ° C. or more and 200 ° C. or less under a pressure of 1 Pa or more and atmospheric pressure or less. 6. A process for producing a monocarboxylic acid according to any one of 6 above.

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