JP5713333B2 - Method for producing dihydroxyacetone - Google Patents

Description

  The present invention relates to a method of using a microorganism in which at least one gene involved in glycerol metabolism is disrupted in a method for producing dihydroxyacetone by a microbial reaction process.

  In recent years, biorefinery has attracted attention as a raw material conversion policy that does not depend only on soaring petroleum resources, or as a technical concept that addresses global warming issues such as carbon dioxide reduction. Biomass is carbon neutral among renewable energy, and the introduction of biofuels such as bioethanol and biodiesel is progressing on a global scale. Biodiesel fuel (hereinafter referred to as “BDF”) is obtained by converting triglycerides, which are the main components of fats and oils, into fatty acid methyl esters by transesterification, and is a method for effectively using glycerin produced as a by-product of this reaction. Development is the key to process development. Considering the rapid increase in BDF usage in recent years, the solution of the glycerin problem is urgent. Also in the oleochemical industry, the effective use of glycerin is also a big problem because processes using vegetable oils and fats, which are alternative and reproducible resources for petroleum, have been introduced.

  Many attempts have been made so far to produce useful substances from glycerin (for example, Non-Patent Documents 1 and 2), and glycerin derivatives such as epichlorohydrin and propylene glycol are being produced at an industrial level. However, for example, when the glycerin price soars with the rise in oil and fat prices as seen from the end of 2007 to 2008, the production process of these low-priced general-purpose chemicals is greatly damaged in cost. Therefore, it is expected to produce a glycerin derivative having higher functionality and higher added value.

  Dihydroxyacetone, which is one of glycerin-inducing substances, is useful in various applications, and is widely used as a high value-added substance such as a raw material for cosmetics and pharmaceuticals.

As a method for producing dihydroxyacetone, a bioprocess (fermentation method) using Gluconobacter oxydans has been industrialized, but in the case of fermentation method, a by-product is present in the reaction solution at the end of fermentation. Since non-target substances are mixed, the separation / purification process of dihydroxyacetone from the fermentation liquor is very complicated, requiring a lot of labor and cost (for example, Patent Document 1). Further, when the glycerin (raw material) concentration in the reaction solution is higher than 10% by weight, there is a problem that the initial glycerin concentration cannot be increased because it adversely affects the biological activity of acetic acid bacteria (for example, Non-Patent Document 3). Therefore, in recent years, research on oxidation reaction of glycerin using a chemical catalyst has also been actively conducted. For example, oxidation reaction using a metal catalyst such as Bi-Pt or Au (for example, Patent Documents 2 and 3, Non-Patent Document 4) Dihydroxyacetone has also been produced by an oxidation reaction using a non-metallic catalyst in subcritical water (Patent Document 4), and both are at the stage of basic research. At present, metal catalysts are not industrialized due to problems such as special and expensive reactions and high temperature / high pressure reactions.

Japanese Patent Publication No.54-28894 Japanese Patent Publication No. 4-356436 Japanese Patent Publication No. 5-245373 Japanese Patent Publication No. 2009-137789

M. Pagliaro, R. Ciriminna, H. Kimura, M. Rossi, C. Della Pina: Angew. Chem. Int. Ed., 46, 2-20 (2007). C.-H. Zhou, J. N. Beltramini, Y.-X. Fan, G. Q. Lu: Chem. Soc. Rev., 37, 527-549 (2008). R. Mishra, S. R. Jain, A. Kumar: Biotechnol. Adv., 26, 293-303 (2008) S. Carrettin, P. McMorn, P. Johnston, K. Griffin, G. J. Hutchings: Chem. Commun., 696-697 (2002).

  An object of the present invention is to solve the above-mentioned problems of the prior art, and specifically, to construct a new technology for efficiently producing dihydroxyacetone by a bioprocess. And a new method for producing dihydroxyacetone that reduces the amount of non-target substances in the fermentation broth.

  In order to achieve the above-mentioned object, the present inventors have intensively studied, and as a result, in the process of producing dihydroxyacetone by a fermentation method, a microorganism that disrupts a gene involved in glycerol metabolism, specifically an alcohol dehydrogenase gene In the present invention, it was found that dihydroxyacetone can be efficiently produced from high-concentration glycerin and the amount of non-target substances such as glyceric acid can be reduced compared to the parent strain when using microorganisms that have disrupted It came to complete.

  That is, the present invention has been completed by obtaining the above knowledge, and is specifically as follows.

[1] A method for producing dihydroxyacetone, comprising culturing a microorganism having an ability to produce dihydroxyacetone from glycerin in which at least one of genes involved in glycerol metabolism is disrupted in a glycerin-containing medium, And the above method comprising the step of isolating and purifying dihydroxyacetone from the medium.
[2] The method according to [1], wherein the gene involved in glycerol metabolism is an alcohol dehydrogenase gene.
[3] The method according to [1] or [2], wherein the microorganism having the ability to produce dihydroxyacetone from glycerin is a microorganism selected from the group of bacteria belonging to the genus Gluconobacter .
[4] The method according to any one of [1] to [3], wherein the microorganism having the ability to produce dihydroxyacetone from glycerin is Gluconobacter oxydans .
[5] The method according to any one of [1] to [4], wherein the glycerin-containing medium contains 10 to 25 wt% glycerin.

  The present invention provides a new technology for efficiently producing dihydroxyacetone by a bioprocess, in particular, a method for producing dihydroxyacetone that uses high-concentration glycerin as a raw material and reduces the amount of non-target substances in the fermentation broth. provide.

FIG. 1 shows the amounts of glycerol, glyceric acid, dihydroxyacetone, and dry cell weight in Example 2 over time. Black circle: Glycerol concentration (g / l); White circle: Glyceric acid (GA) concentration (g / l); White square: Dihydroxyacetone (DHA) concentration (g / l); Black square: Dry cell weight ( g / l) FIG. 2 shows each amount of glycerol, glyceric acid, dihydroxyacetone, and dry cell weight in Example 3 over time. Black circle: Glycerol concentration (g / l); White circle: Glyceric acid (GA) concentration (g / l); White square: Dihydroxyacetone (DHA) concentration (g / l); Black square: Dry cell weight ( g / l) FIG. 3 shows each amount of glycerol, glyceric acid, dihydroxyacetone, and dry cell weight in Example 4 over time. Black circle: Glycerol concentration (g / l); White circle: Glyceric acid (GA) concentration (g / l); White square: Dihydroxyacetone (DHA) concentration (g / l); Black square: Dry cell weight ( g / l) FIG. 4 shows the amounts of glycerol, glyceric acid, and dihydroxyacetone in Example 5 over time. Black circle: Glycerol concentration (g / l); White circle: Glyceric acid (GA) concentration (g / l); White square: Dihydroxyacetone (DHA) concentration (g / l); Black square: Dry cell weight ( g / l) FIG. 5 shows the amounts of glycerol, glyceric acid, and dihydroxyacetone in Example 6 over time. Black circle: Glycerol concentration (g / l); White circle: Glyceric acid (GA) concentration (g / l); White square: Dihydroxyacetone (DHA) concentration (g / l); Black square: Dry cell weight ( g / l) FIG. 6 shows each amount of glycerol, glyceric acid, dihydroxyacetone, and dry cell weight in Comparative Example 1 over time. Black circle: Glycerol concentration (g / l); White circle: Glyceric acid (GA) concentration (g / l); White square: Dihydroxyacetone (DHA) concentration (g / l); Black square: Dry cell weight ( g / l) FIG. 7 shows the amounts of glycerol, glyceric acid, dihydroxyacetone, and dry cell weight in Comparative Example 2 over time. Black circle: Glycerol concentration (g / l); White circle: Glyceric acid (GA) concentration (g / l); White square: Dihydroxyacetone (DHA) concentration (g / l); Black square: Dry cell weight ( g / l)

  The microorganism that can be used in the present invention has the following characteristics.

The microorganism that can be used in the present invention is first characterized in that it can produce dihydroxyacetone from glycerin. Examples of such microorganisms include, but are not limited to, acetic acid bacteria having the ability to convert glycerin to glyceric acid. In particular, Gluconobacter bacteria capable of producing dihydroxyacetone higher than glycerin are desirable. Specific examples include Gluconobacter oxydans , Gluconobacter albidus , and Gluconobacter. · Serinasu (Gluconobacter cerinus), glucoside frateurii (Gluconobacter frateurii), Gluconobacter-Kondoni (Gluconobacter kondonii), Gluconobacter, Thailand Randy dregs (Gluconobacter thailandicus), other Gluconobacter bacteria belonging to the genus (Gluconobacter sp.) NBRC3259 strain etc. are mentioned. Gluconobacter oxydans is preferred. Mutants of these microorganisms can also be used as long as they retain the ability to convert glycerin to dihydroxyacetone. The mutant strain may be derived from a spontaneous mutation, or may be obtained by performing some physical or chemical treatment such as ultraviolet irradiation or chemical mutagen treatment. Furthermore, the case where these microorganisms are used as hosts for genetic recombinants is also included.

  Secondly, the microorganism that can be used in the present invention is characterized in that at least one gene involved in glycerol metabolism is disrupted. The gene involved in glycerol metabolism is not particularly limited, and examples thereof include a glycerol dehydrogenase gene, a glycerol kinase gene, a glycerol-3-phosphate dehydrogenase gene, a dihydroxyacetone kinase gene, and an alcohol dehydrogenase gene. Preferably, it is an alcohol dehydrogenase gene.

  Alcohol dehydrogenase is an enzyme that is localized at various sites such as cytoplasm, cell membrane, and periplasm. Cofactors required for enzyme activity include pyrroloquinoline quinone (PQQ) and nicotine adenine dinucleotide (NAD / NADH). However, PQQ-dependent alcohol dehydrogenase localized in cell membrane-bound or periplasm is preferable.

  The alcohol dehydrogenase gene is a gene encoding an enzyme that catalyzes a reaction for converting alcohol to acetaldehyde. The alcohol dehydrogenase gene has already been identified in various microorganisms. For example, if it is a bacterium belonging to the genus Gluconobacter, it is registered in GenBank as Gene ID: NC_006677.

  The “alcohol dehydrogenase gene” of the present invention includes a base sequence of a known alcohol dehydrogenase gene, such as the base sequence of the alcohol dehydrogenase gene disclosed in GenBank, etc. Also included is a DNA encoding a protein having a deletion, substitution, addition or insertion of several bases and having alcohol dehydrogenase activity. The range of “1 to several” is not particularly limited. For example, 1 to 100, preferably 1 to 70, more preferably 1 to 50, still more preferably 1 to 20, and particularly preferably 1 to 1. 10 or 1 to 5. The “alcohol dehydrogenase gene” of the present invention is complementary to the base sequence of a known alcohol dehydrogenase gene, for example, the base sequence of the alcohol dehydrogenase gene disclosed in GenBank, etc. It also includes a DNA that is capable of hybridizing under stringent conditions with a DNA comprising a base sequence and that encodes a protein having alcohol dehydrogenase activity. In the present invention, “stringent conditions” refers to conditions under which so-called specific hybrids are formed and non-specific hybrids are not formed. For example, 2 to 6 × SSC (composition of 1 × SSC: 0.15M NaCl) , 0.015M sodium citrate, pH 7.0) and a solution containing 0.1 to 0.5% SDS at 42 to 55 ° C., and a solution containing 0.1 to 0.2 × SSC and 0.1 to 0.5% SDS The conditions for washing at 65 ° C. Furthermore, the “alcohol dehydrogenase gene” of the present invention includes a base sequence of a known alcohol dehydrogenase gene, such as the base sequence of the alcohol dehydrogenase gene disclosed in GenBank and the like with the above registration number, For example, a base sequence having an identity of 80% or more, more preferably 90% or more, and most preferably 95% or more when calculated using default or default parameters), and alcohol dehydrogenase activity It also includes DNA encoding a protein having

  “At least one” refers to 1, 2, 3, 4, 5, or more.

  “A gene involved in glycerin metabolism has been disrupted” means that the gene or a transcriptional regulatory region of the gene is deleted, substituted, added, or inserted so that the gene does not function completely or significantly. It means having a mutation. As a result, transcription or translation from the gene is inhibited, or a mutation occurs in the amino acid sequence translated from the incompletely transcribed mRNA, and the protein encoded by the gene can perform its original function. Disappear.

  Mutations in gene disruption may be naturally introduced, or may be artificially introduced using some physical or chemical treatment or gene recombination technique. The method for introducing a mutation using a gene recombination technique is not particularly limited and can be performed using a known method. An example is double crossover by homologous recombination. Briefly, a target gene involved in glycerol metabolism on the chromosome of the microorganism is cloned into a general vector known to those skilled in the art for gene transfer. Thereafter, a foreign gene (for example, drug resistance gene) is inserted into the cloned sequence of the target gene to prepare a DNA construct containing the target gene divided by the foreign gene (for example, drug resistance gene). By introducing this DNA construct into the microorganism, homologous recombination occurs between the fragmented target gene on the DNA construct and the target gene on the microorganism chromosome, thereby destroying the gene of the microorganism. Alternatively, instead of inserting a foreign gene into the target gene and dividing it, prepare a DNA construct containing the target gene in which part or all of the base sequence of the target gene has been deleted and introduce it into the microorganism. Thus, the target gene having a partial or complete deletion on the DNA construct and the target gene on the microorganism chromosome can undergo homologous recombination, and the gene of the microorganism can be destroyed. Examples of the method for introducing a DNA construct into a microorganism include a calcium phosphate method or a calcium chloride / rubidium chloride method, an electroporation method, an electroinjection method, a method using chemical treatment such as PEG, and a method using a gene gun. .

  In the present invention, the microorganism may be immobilized on a solid support (eg, polymer, mineral, metal, gel, polysaccharide, etc.). The immobilization of the microorganism on the solid support can use a covalent bond or a non-covalent bond.

  As described in detail in the following examples, in the present invention, the microorganism is about 2 times or 3 times in a glycerin-containing medium as compared with a microorganism that can produce dihydroxyacetone from glycerin conventionally used. It is possible to produce 4 times, 5 times or more amounts of dihydroxyacetone. Further, in the present invention, the above-mentioned microorganism has a by-product (for example, glyceric acid) production amount of about 1/10 in a glycerin-containing medium as compared with a microorganism that can produce dihydroxyacetone from glycerin conventionally used. , 1/20, 1/30, 1/40 or less. Furthermore, in the present invention, the above microorganism can grow in a medium containing glycerin at a high concentration and can produce dihydroxyacetone from a high concentration of glycerin as compared to a microorganism that can produce dihydroxyacetone from glycerin that has been conventionally used. it can.

  Dihydroxyacetone can be efficiently produced by the fermentation reaction by the microorganism. The fermentation reaction can be performed by a general microbial culture method known to those skilled in the art.

  Culture of microorganisms for production of dihydroxyacetone can be performed using a nutrient medium containing a carbon source, a nitrogen source, an inorganic salt, and the like. Glycerin is included in the medium as a carbon source. As the nitrogen source, for example, substances that can be assimilated by microorganisms, such as sodium nitrate, ammonium sulfate, ammonium chloride, and urea, can be used alone or in combination. Further, as the inorganic salt, potassium monohydrogen phosphate, potassium dihydrogen phosphate, magnesium sulfate, or the like can be used. In addition to these, nutrients such as yeast extract, peptone, polypeptone, meat extract, corn steep liquor and the like can be appropriately added to the medium as necessary, and these nitrogen-containing organic substances can be substituted for the nitrogen source. The medium can be a solid medium solidified by adding agar or gelatin, a semi-fluid medium added with low-concentration agar, or a liquid medium (also referred to as bouillon or broth) containing only medium components. Is preferred.

  The glycerin as a raw material to be added to the medium as a carbon source can be appropriately selected from various origins and qualities according to the purpose. For example, any of chemically synthesized glycerin, glycerin derived from natural products such as vegetable oil and animal oil, and glycerin derived from waste such as waste oil can be used. Also, any of high-purity purified glycerin and crude glycerin produced as a by-product in the production of BDF can be used. In the present invention, the amount of glycerin added to the medium as a raw material is higher than the amount of glycerin conventionally used, and is 5 to 40 wt%, preferably 10 to 25 wt%, more preferably 15 to 25 wt%. %.

  The pH of the medium is in the range of about pH 3 to 10, preferably about pH 4 to 8 at the start of the reaction.

  Although the culture conditions of microorganisms vary depending on the type of microorganism used, the culture temperature is a temperature condition at which the microorganisms can act, for example, 20 to 50 ° C., preferably 30 ° C., and the culture time is 6 hours to 4 days, preferably 1 ~ 3 days.

  Examples of the fermenter that can be used for the culture include an aeration stirring type, a bubble column type, a fluidized bed type, and a packed bed type. Moreover, culture | cultivation of microorganisms may be performed by a continuous type or a batch type.

  After completion of the culture, the medium is collected and dihydroxyacetone produced and accumulated in the medium is isolated and purified. Isolation and purification of dihydroxyacetone can be performed, for example, by the following method.

  A medium obtained by culturing for a predetermined time is collected, and cultured cells of microorganisms are separated and collected from the medium. The method for separating / recovering the cultured cells is not particularly limited, and known methods such as centrifugation and membrane separation can be used. The collected cultured cells can be used as a microbial source for the subsequent production of dihydroxyacetone.

  Isolation and purification of dihydroxyacetone produced and accumulated in the medium can be performed by any method, but from the medium after removing the cells by a method such as centrifugation, for example, separation methods by various chromatography, It can be isolated and purified using a separation method by crystallization, a separation method using a membrane, or the like.

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

  In the following examples, glycerin and dihydroxyacetone are quantified using a column that analyzes sugar alcohol (SC1011 manufactured by Shodex), and glyceric acid is quantified using a column that simultaneously analyzes sugar and organic acid (SH1011 manufactured by Shodex). Chromatography (HPLC) was performed.

Example 1
For the gene (adhA) encoding the membrane-bound alcohol dehydrogenase of Gluconobacter oxydans IFO12528, the following primers were designed based on the known sequence information (GenBank NC_006677):
5'-GAAGATCTTACAGCCCGCTCGACCAG-3 '(SEQ ID NO: 1)
5′-ACGCGTCGACACAGGGGTGGGGACGCTT-3 ′ (SEQ ID NO: 2).

Genomic DNA of Gluconobacter oxydans IFO12528 was extracted according to a conventional method, and a DNA fragment containing a gene encoding a membrane-bound alcohol dehydrogenase (adhA) was amplified by PCR using the above primers. After constructing a plasmid by ligating the amplified 3.6 kb DNA fragment to the vector pGEM T-easy, insert a BamHI fragment containing the approximately 1.2 kb kanamycin resistance gene into the BamHI site inside the adhA gene to disrupt the gene. Plasmid pGEM-ΔadhA was prepared.

The prepared plasmid pGEM-ΔadhA was introduced into Gluconobacter oxydans IFO12528 by electroporation, and a disrupted strain Gluconobacter oxydans IFO12528ΔadhA was obtained by homologous recombination. The genomic DNA of Gluconobacter oxydans IFO12528ΔadhA was extracted according to a conventional method, and PCR was performed in the same manner to confirm the destruction of the adhA gene.

(Example 2)
Gluconobacter oxydans IFO12528 △ adhA prepared by the above method was inoculated into 5 tubes of 5 mL of preculture medium having a composition of 3% glucose, 0.5% yeast extract and 0.2% polypeptone, and reciprocated at 30 ° C for 48 hours. Shaking culture was performed. Subsequently, in the main culture medium having the composition shown in Table 1, a total amount (25 mL) of the above preculture was inoculated into a 1-liter miniger containing 500 mL of a medium having a glycerin concentration of 10 wt%. The culture was performed at a temperature of 30 ° C., an aeration rate of 1 vvm, and stirring at 500 rpm.

  The pH in the reaction solution was controlled at 5, and 10M NaOH was used as a pH adjuster. The amount of dihydroxyacetone produced from the start of the main culture by the above-described method until the third day was quantitatively determined as shown in FIG. 1, and the amount of dihydroxyacetone in the reaction for three days was 73.5 g / l. . The production of glyceric acid as a by-product was 1.9 g / l. The produced dihydroxyacetone was 0.693 mol with respect to 1 mol of glycerin consumed during the reaction period.

(Example 3)
Gluconobacter oxydans IFO12528ΔadhA was subjected to jar culture in the same manner as in Example 2. However, in the main culture medium having the composition shown in Table 1, the glycerin concentration was 15 wt% and the aeration rate was 0.5 vvm. The amount of dihydroxyacetone produced was quantified as shown in FIG. 2. The amount of dihydroxyacetone was 108 g / l after 3 days of reaction. The production of glyceric acid as a by-product was 2.2 g / l. The produced glyceric acid was 0.676 mol per 1 mol of glycerin consumed during the reaction period.

Example 4
Gluconobacter oxydans IFO12528ΔadhA was subjected to jar culture in the same manner as in Example 2. However, in the main culture medium having the composition shown in Table 1, the glycerol concentration was set to 20 wt%. The produced dihydroxyacetone was quantified as shown in FIG. 3. The amount of dihydroxyacetone was 125 g / l after 3 days of reaction. The production of glyceric acid as a by-product was 1.7 g / l. The produced glyceric acid was 0.801 mol per 1 mol of glycerin consumed during the reaction period.

(Example 5)
Gluconobacter oxydans IFO12528 △ adhA was pre-cultured in the same manner as in Example 2 and then a 1 liter miniger containing 500 mL of a medium having a glycerin concentration of 5 wt% among the main culture media having the composition shown in Table 1. Then, the whole amount of the preculture was inoculated. The culture was performed using 10M NaOH as a pH adjusting agent at a temperature of 30 ° C., an aeration rate of 0.5 vvm, and stirring at 500 rpm for 24 hours. Thereafter, whole cells were collected by centrifugation, suspended in 500 mL of a medium having a glycerin concentration of 15 wt% of the main culture medium having the composition shown in Table 1, and then reacted in a 1 liter minijar. went. The reaction was carried out using 10M NaOH as a pH adjuster at a temperature of 30 ° C., an aeration rate of 0.5 vvm, and stirring at 500 rpm, and 75 g of glycerin was added 2 days after the start of the reaction, and the reaction was continued for 4 days. The amount of dihydroxyacetone produced was quantified, as shown in FIG. 4, and the amount of dihydroxyacetone was 140 g / l after 4 days of reaction. Moreover, almost no production of glyceric acid as a by-product was observed.

(Example 6)
Gluconobacter oxydans IFO12528ΔadhA was collected in the same manner as in Example 5, suspended in 500 mL of a 25 wt% aqueous glycerin solution, and then reacted in a 1 liter miniger. The reaction was performed using 10M NaOH as a pH adjuster at a temperature of 30 ° C., an aeration rate of 0.5 vvm, and stirring at 500 rpm, and the reaction was continued for 4 days. The amount of dihydroxyacetone produced was quantified as shown in FIG. 5, and the amount of dihydroxyacetone was 86.4 g / l after 4 days of reaction. Although residual glycerin was observed, dihydroxyacetone could be produced even at a high concentration of 25 wt%. Moreover, almost no production of glyceric acid as a by-product was observed.

(Comparative Example 1)
As a comparative experiment of Example 2, Gluconobacter oxydans IFO12528 (parent strain) was subjected to jar culture in the same manner as in Example 2. When the produced dihydroxyacetone and glyceric acid were quantified, the amount of dihydroxyacetone produced in the reaction for 3 days was 63.5 g / l and the by-product glyceric acid was 18.5 g / l as compared with FIG. 1 of Example 2. (FIG. 6). The produced dihydroxyacetone was 0.617 mol with respect to 1 mol of glycerin consumed during the reaction period.

(Comparative Example 2)
As a comparative experiment of Example 3, Gluconobacter oxydans IFO12528 was subjected to jar culture in the same manner as in Example 3. When the produced dihydroxyacetone and glyceric acid were quantified, the amount of dihydroxyacetone produced in the reaction for 3 days was 38.1 g / l and the byproduct glyceric acid was 47 g / l as compared with FIG. 2 of Example 3. (FIG. 7). The amount of produced dihydroxyacetone was 0.357 mol with respect to 1 mol of glycerin consumed during the reaction period.

(Comparative Example 3)
As a comparative example of Example 4, Gluconobacter oxydans IFO12528 was subjected to jar culture in the same manner as in Example 4. As a result, the cells hardly grew after 3 days of culture.

  According to the present invention, dihydroxyacetone, which has a high industrial utility value as a raw material for pharmaceuticals and cosmetics, etc., only reduces the amount of by-products by destroying a gene involved in glycerol metabolism of microorganisms that have been used so far. In addition to being able to be manufactured, the production amount of dihydroxyacetone using high-concentration glycerin as a raw material increases dramatically, so that the efficiency of the manufacturing process and cost reduction can be expected.

Claims (4)

A method for producing dihydroxyacetone, comprising a step of culturing a microorganism having an ability to produce dihydroxyacetone from glycerin in which an alcohol dehydrogenase gene is disrupted in a glycerin-containing medium, and dihydroxyacetone from the medium. The said method including the process to isolate | separate and refine | purify. The method according to claim 1, wherein the microorganism capable of producing dihydroxyacetone from glycerin is a microorganism selected from the group of bacteria belonging to the genus Gluconobacter. The method according to claim 1 or 2 , wherein the microorganism having the ability to produce dihydroxyacetone from glycerin is Gluconobacter oxydans. The method according to any one of claims 1 to 3 , wherein the glycerin-containing medium contains 10 to 25 wt% glycerin.

Images