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WO 2017/139420 Al 17 August 2017 (17.08.2017) P O P C T (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (10) International Publication Number (43) International Publication Date WO 2017/139420 Al 17 August 2017 (17.08.2017) P O P C T (51) International Patent Classification: AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY, C07C 45/27 (2006.01) C12P 19/24 (2006.01) BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DJ, DK, DM, DO, DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, (21) International Application Number: HN, HR, HU, ID, IL, IN, IR, IS, JP, KE, KG, KH, KN, PCT/US20 17/0 17069 KP, KR, KW, KZ, LA, LC, LK, LR, LS, LU, LY, MA, (22) International Filing Date: MD, ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, 8 February 2017 (08.02.2017) NI, NO, NZ, OM, PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SA, SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, (25) Filing Language: English TH, TJ, TM, TN, TR, TT, TZ, UA, UG, US, UZ, VC, VN, (26) Publication Language: English ZA, ZM, ZW. (30) Priority Data: (84) Designated States (unless otherwise indicated, for every 62/292,924 9 February 2016 (09.02.2016) US kind of regional protection available): ARIPO (BW, GH, GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, ST, SZ, (71) Applicant: KEMBIOTIX LLC [US/US]; 325 Speen TZ, UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, RU, Street #404, Natick, MA 01760 (US). TJ, TM), European (AL, AT, BE, BG, CH, CY, CZ, DE, DK, EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU, (72) Inventors: MILAN, Jay, L.; 325 Speen Street #404, Nat LV, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK, ick, MA 01760 (US). MANNAN, Ramasamy, Mannar; SM, TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ, 64 Arrowhead Road, Weston, MA 02493 (US). GW, KM, ML, MR, NE, SN, TD, TG). (74) Agent: MANNAN, Ramasamy, Mannar; 64 Arrowhead Published: Road, Weston, MA 02493 (US). — with international search report (Art. 21(3)) (81) Designated States (unless otherwise indicated, for every kind of national protection available): AE, AG, AL, AM, (54) Title: BIOLOGICAL FERMENTATION USING DIHYDROXYACETONE AS A SOURCE OF CARBON (57) Abstract: The present invention relates to the use of hydrocarbons derived from natural gas in the fermentative production of biochemicals including biofuels. More specifically, the present invention provides the method for manufacturing dihydroxyacetone ("DHA") from natural gas, biogas, biomass and CO2 released from industrial plants including electricity-generating plants, steel mills and cement factories and the use of DHA as a source of organic carbon in the fermentative production of biochemicals includ - ing biofuels. The present invention comprises three stages. In the first stage of the present invention, syngas and formaldehyde are produced from natural gas, biogas, biomass and CO2 released from industrial plants. In the second stage of the present invention, formaldehyde and syngas are condensed to produce DHA. In the third stage of the present invention, biochemicals including bio - fuels are produced from DHA using fermentation process involving wild type or genetically modified microbial biocatalysts. BIOLOGICAL FERMENTATIONUSING DIHYDROXYACETONEA S A SOURCE OF CARBON CROSS REFERENCE T O RELATED APPLICATION (001) This application claims the priority to the U.S. Provisional Application Serial No. 62/292,924, filed on February 9, 2016. FIELD OF THE INVENTION (002) This invention is in the field of producing a family of biochemicals including biofuels from natural gas, biogas, biomass and C0 2 using microbial biocatalysts. BACKGROUND OF THE INVENTION (003) There has been an impressive growth in manufacturing chemicals using microbial biocatalysts. Besides reducing toxic by-products, bio-based routes to chemical synthesis involving microbial biocatalysts may allow the use of new class of feedstocks. There is a growing expectation that lowered costs, increase in production speed, flexibility of manufacturing plants, and increased production capacity can be achieved using bio-based routes for chemical synthesis. The bio-based routes for chemical biosynthesis involve biological fermentation process. A number industrial fermentation processes for manufacturing a broad range of biochemicals including biofuels have been commercialized. The industrial biomanufacturing is considered to hold a great promise in meeting the evolving demands of chemical production in the current century and beyond (Clomburg et al. Industrial biomanufacturing: The future of chemical production. Science, 2017, 355: 6320 aag0804). (004) During the year 2015, the demand for the chemicals manufactured through fermentation process was 56.98 million tons and this demand is expected to reach 85.66 million tons by 2024 representing a compound annual growth rate of 4.6% during the period 2016-2024. (005) Representative examples of biochemicals that are suitable industrial scale production using biological fermentation include but not limited to ethanol, acetic acid, propionic acid, lactic acid, 3-hydroxy propionic acid, 1-3, propanediol, butanol, succinic acid, and muconic acid. (006) In the industrial fermentation process for the production of chemicals through bio-based routes, organic carbon in the form of fermentable carbohydrates such as glucose, sucrose and glycerol have been the primary raw material and often account for the largest single input cost. At present, dextrose derived from starch in grains and sucrose from sugarcane are primarily used in the industrial production of chemicals through biological fermentation. There has been some effort to use glycerol, obtained as a by-product from the biodiesel industry and fermentable sugars derived from the hydrolysis of cellulosic materials as a source of carbon in the industrial scale fermentation. Hexose and pentose sugars derived from cellulose are considered to yield cost-effective fermentable sugars. However, the technology to produce fermentable sugars from cellulose is not yet matured enough to support industrial scale fermentation process. (007) The cost of the feedstock used in the fermentation processes generally accounts for over 50%-70% of the product cost. It can be higher than 70% where the fermentation yields are low. When the biochemical products are made using fermentation processes, the cost of feedstocks is an important factor in the overall economy of the process. In most cases, petroleum feedstocks are abundant and cheap, making the products derived from petrochemical processes economically more competitive than the similar products derived from biological fermentation using microbial biocatalysts. For a fermentation process to be competitive against petrochemical processes in manufacturing a desired biochemical, the feedstock used in the fermentation process needs to be cost competitive. The cost of the feedstocks used in the biological fermentation process can be significantly reduced if they can be derived from fossil hydrocarbons. Methane is a fossil hydrocarbon and is abundantly available around the world. Methane can be converted into dihydroxyacetone ("DHA") which can be used as a feedstock in the biological fermentation as described in this invention. The method to convert methane to DHA is scalable and relatively inexpensive process. One of the biggest advantages of using methane and fossil hydrocarbons to produce DHA according to the present invention is that methane is a gas and DHA is a solid which can be easily stored and transported from one place to another. On the other hand, the conventional dextrose feedstock used in the industrial fermentation process is available as 70% solution in water and needs to be kept above ambient temperature during transportation and storage to prevent crystallization. (008) Natural gas primarily composed of methane is reported to be present in abundant quantities in several regions of the world and is fast becoming a cheap feedstock to replace petrochemical feedstock. For example, natural gas is replacing the feedstocks derived from naptha-crackers for the manufacturing many chemical products. (009) There has been growing interest in manufacturing value added commodity chemicals from methane present in natural gas as feedstock. For examples, oxidative coupling of methane (OCM) has been developed to produce methanol, ethylene, propylene and butadiene using natural gas as a feedstock. Methanol to olefins (MTO) process is used to convert methanol into dimethyl ether (DME) and water. DME derived from MTO process is converted to olefins by a pyrolysis reaction. In another industrial application, methane present in the natural gas is converted to Fisher-Tropsch (FT) liquid and subsequently the FT liquid is converted to olefins by means of steam cracking. The olefins derived from natural gas are used as a primary building block to produce various polymers and consumer-focused functional materials. However, most of the current processes suffer from poor selectivity, high-energy cost and large C02 emission. (010) There is a growing interest in using methane and its derivatives such as methanol, syngas, formate or formaldehyde as a source of organic carbon in the biological fermentation. A number of methanogenic microorganisms are considered as potential biocatalysts for the fermentative production of industrial chemicals using methane as a feedstock in the biological fermentation. However, we need to overcome several challenges before we develop a biocatalyst useful in commercial scale fermentative production of industrial chemicals using methane as a feedstock since
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