DESC:FIELD OF THE INVENTION

The present invention relates to a bio-assisted process for transforming carbon dioxide to fuels and chemicals. More particularly, the present invention relates to a process for producing fuels and chemicals using a novel microbial consortium in an electro-biochemical system.

BACKGROUND OF THE INVENTION
CO2 is a greenhouse gas and it is widely accepted that rising atmospheric CO2 levels are responsible for increasing average global temperatures. Burning of fossil fuels is one of the largest overall contributors to CO2 emissions, Thus, utilizing the CO2 generated by manmade sources is critical in the battle against global warming. The CO2 is spent form of energy stored in carbon and hence if CO2 is used as feedstock for producing fuels and chemicals,it can serve as sustainable mitigation options against adverse effect of CO2 accumulation in the atmosphere.
Certain microorganisms called electrogen or electroactive microbes have ability to interact electrochemically with electrodes which can either donate electrons to an anode or accept electrons from a cathode accept or donate electron to facilitate their metabolic activity [Lovley, D. R. (2008) Curr. Opinion Biotechnol. 19:564-571]. These microbes can be utilized for the production of energy from organic material present in wastewater streams [Bond, D. R., et al.(2002), Science 295:483-485; Bond, D. R., and D. R. Lovley(2003), Appl. Environ. Microbiol. 69:1548-1555;Rozendalet al., (2008) Environ. Sci. Technol., , 42, 629-634; Defeng Xing et al. Environ. Sci. Technol., 2009, 43 (10), pp 3953–3958; Clauwaertet al., (2008) Water Sci. Technol., Vol.57, 575-579) and reduction of various contaminants. Few microorganisms form biofilm on cathode under electric current and derive electron from an electrode. These electrons can be used by the microorganisms to reduce carbon dioxide to yield industrially relevant products. The electric current can be derived from any a renewable source of power.
D. R. Lovley (Environmental Microbiology Reports (2011) 3(1), 27–35 illustrates current knowledge on how electrons can be directly transferred from electrodes to microorganisms and the potential practical applications of this novel form of microbial respiration in bioremediation, bioenergy, and chemical production.
Nevin, K. P.et al. (2010. mBio 1(2):e00103-10) describes the demonstration of microbial production of chemicals (including chemicals having more than one carbon atom therein) from carbon dioxide and water using power provided by an imposed electrical current. This system operates in the reverse manner as compared to MFCs. This system is useful to produce chemicals, for example commodity chemicals such as acetic acid, that are useful in their own right, and also serve as chemical precursors for the production of many other commodity and fine chemicals which are well known and have recognized uses.
Kelly P. Nevin,et al.(2011)Appl Environ Microbiol. Vol. 77(9): 2882–2886) provides a process using Sporomusa species, Clostridium ljungdahlii, Clostridium aceticum, and Moorella thermoacetica, using aqueous medium and CO2 in H-Cell where anode and cathode was separated by specific membrane. In this study, in general acetate was the primary product, but 2-oxobutyrate and formate were also formed.

Li et al(2012, Science Volume 335, March 2012, pg 1596) reported conversion of CO2 into formic acid and then formic acid to isobutanol and 3 methyl 1- butanol using a genetically modified bacteria in a integrated electrochemical system where cathode and anode was separated. In this system platinum was used as anode.

Yanming Gong, et al (2013), Environ. Sci. Technol. 2013, 47, 568-573) evaluated the use of sulfide as an electron source for microbial electrosynthesis. Abiotically oxidation of sulfide on the anode yields two electrons. The oxidation product, elemental sulfur, can be further oxidized to sulfate by Desulfobulbus propionicus, generating six additional electrons in the process. The eight electrons generated from the combined abiotic and biotic steps were used to reduce carbon dioxide to acetate on a graphite cathode by Sporomusa ovata at a rate of 24.8 mmol/day•m2. Using a strain of Desulfuromonas as biocatalyst on the anode resulted in an acetate production rate of 49.9 mmol/day•m2, with a Coulombic efficiency of over 90%. These results demonstrate that sulfide can serve effectively as an alternative electron donor for microbial electrosynthesis.

EP2373832 describes a process for producing one or more chemical compounds comprising the steps of providing a bioelectrochemical system having an anode and a cathode separated by a membrane, the anode and the cathode being electrically connected to each other, causing oxidation to occur at the anode and causing reduction to occur at the cathode to thereby produce reducing equivalents at the cathode, providing the reducing equivalents to a culture of microorganisms, and providing carbon dioxide to the culture of microorganisms, whereby the microorganisms produce the one or more chemical compounds, and recovering the one or chemical compounds.

US20120288898 discloses a microbial production of multi-carbon chemicals and fuels from water and carbon dioxide using electric current provides systems and methods for generating organic compounds using carbon dioxide as a source of carbon and electrical current as an energy source. In one embodiment, a reaction cell is provided having a cathode electrode and an anode electrode that are connected to a source of electrical power, and which are separated by a permeable membrane. A biological film is provided on the cathode. The biological film comprises a bacterium that can accept electrons and that can convert carbon dioxide to a carbon-bearing compound and water in a cathode half-reaction. At the anode, water is decomposed to free molecular oxygen and solvated protons in an anode half-reaction. The half-reactions are driven by the application of electrical current from an external source. Compounds that have been produced include acetate, butanol, 2-oxobutyrate, proponal, ethanol, and formate.

US20130130334 discloses a fuel cell comprising an anode electrode, a cathode electrode and a reference electrode electronically connected to each other; a first biocatalyst comprising a consolidated bioprocessing organism (e.g., a cellulomonad or clostridium or related strains, such as Cellulomonasuda (C. uda), C. lentocellum, A. cellolulyticus, C. cellobioparum, alcohol-tolerant C. cellobioparum, alcohol-tolerant C. uda, Clostridium cellobioparum (C. cellobioparum) and combinations thereof) capable of fermenting biomass (e.g., cellulosic biomass or glycerin-containing biomass) to produce a fermentation by product; and a second biocatalyst (e.g., Geobacter sulfurreducens) capable of transferring substantially all the electrons in the fermentation by product (e.g., hydrogen, one or more organic acids, or a combination thereof) to the anode electrode to produce electricity is disclosed. Systems and methods related thereto are also disclosing a consolidated bioprocessing organism.

US20100252443 discloses bioelectrochemical treatment of gaseous byproducts,
WO2012054629 discloses bio-electrochemical systems, US8277984 discloses substrate-enhanced microbial fuel cells, WO2003096467 discloses membrane less and mediator less microbial fuel cell, EP2025033 discloses materials and configurations for microbial fuel cells, WO200907002 discloses production of a product in a microbial fuel cell and Hongqiang Hu, et al. (2008) discloses hydrogen production using single-chamber membrane-free microbial electrolysis cells. Volume 42, Issue 15, September 2008, Pages 4172–4178
From the above, it is evident that:

• Only few microbes have yet known to transform CO2 to organic compounds in electrobiochemical system e.g. Sporomusa ovata, Closridium lijungdahlii, Clostridium aceticum, Moorella thermoacetica, Sporomusa silvacetica, Sporomusa sphaeroides, Geobacter metallireducens, Anaeromyxobacter sp..

The organic product that has been documented to be produced from CO2 reduction include acetic acid, 2-oxobutyric acid, ethanol, proponal, butanol, formate and other as yet unidentified organic acids.
• It is well known fact that solubility of CO2 in aqueous system is very low and microbial cells only take hydrated CO2 for biotransformation. Hence, the productivity is expected to be low mainly because of low CO2 solubility. In the prior art no provision has been disclosed for improving the solubility/availability of the CO2 in the bioelectrochemical system.

• In the prior art process described for reduction of CO2, has two chambered bioelectrochemical cell with membranes to separate anode and cathode chamber which complicates cell architecture, adds to cost and poses difficulties to upscale.
• The above set up also leads to additional energy burden in deriving the process due to potential losses across the membrane.

In the prior art documents the enzyme carbonic anhydrase is usually used as an externally added substituent to enhance the solubility of CO2 in aqueous media by CO2 hydration. US8480796, US8329498, US8329460, US20130224842, US20120107899CA2813640 and WO2008095057 disclosed improving of the solubility of CO2 by using externally added carbonic anhydrase enzyme from various sources and converting CO2 to inorganic carbonates. However, these disclosures has inherent limitation that enzyme carbonic anhydase need to be added/replenish to keep the process going on. This further adds to cost and limitation towards upscale the process at industrial level.

In order to overcome above deficiencies, we disclose a novel process in which microbes produce carbonic ahydrase enzyme in situ during growth at cathode under electric current and simultaneously bio-transform hydrated CO2 to organic molecules. Prior art is devoid of any such disclosure.

SUMMARY OF THE INVENTION

Accordingly, the main embodiment of the present invention provides a biofilm of electro-active microbe or their synergistic consortium wherein at least one microbe has an ability to solubilise carbon dioxide and convert it to fuels and chemicals under electric current.
Another embodiment of the present invention provides a biofilm as herein described wherein the microbes are selected from the group consisting of Citrobacter sp.IOC-EA-66, Enterobacter aerogenes IOC-EZ-6, Serratia sp. IOC-EA-9 and Clostridium sporogenes IOC-EA-99.
Another embodiment of the present invention provides a method of preparing biofilm of electro-active microbes on cathode said method comprising the steps of:
(a) providing a single chambered electrochemical cell comprising of atleast one cathode and one anode conductive electrode and a growth medium;
(b) adding a carbon dioxide source;
(c) inoculating the cell with consortia of microbes under electric current;
(d) adding plant/microbial extract (s) which improves the biofilm formation;
(e) growing the microbe consortia;
(f) replacing the medium with fresh medium;
( g)obtaining a stable biofilm of electro-active microbes.
One more embodiment of the present invention provides a method as herein described wherein the microbes form biofilm on conductive electrode which is cathode.
One more embodiment of the present invention provides a method as herein described, wherein the microbes are selected from the group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EA-6, Serratia sp. IOC-EA-9 and Clostridium sporogenes IOC-EA-99.
Another embodiment of the present invention provides a single chambered electro-biochemical cell comprising least one cathode, at least one anode, a reaction medium and biofilm of electro-active microbes wherein electro-active microbes having an ability to hydrate carbon dioxide and convert it to fuels and chemicals under electric current.
Another embodiment of the present invention provides an electro-biochemical process for reducing carbon dioxide and water said process comprising the steps of:
(a) providing a single chambered electrochemical system containing at least one anode and one cathode and a medium containing nitrogen, phosphorus and a micronutrient source;
(b) connecting anode and cathode through a source of electrical energy;
(c) operating said source of electrical energy to extract electrons from anode and deliver electrons to cathode;
(d) inoculating the cell with a microbe or consortia of microbes wherein at least one microbe has an ability to produce extracellular carbonic anhydrase enzyme during growth under electric current;
(e) adding the carbon dioxide source;
(f) adding plant/microbial extract (s) which improves the biofilm formation;
(g) allowing the bacteria to form biofilm on cathode;
(h) replacing the medium of step (a) with a fresh medium;
(i) incubating the electrochemical system with microorganisms to produce one or more chemical compounds in a vicinity of said cathode; and
(j) recovering the one or chemical compounds
Another embodiment of the present invention provides an electro-biochemical process as described herein wherein the microbes are selected from the group consisting of Citrobacter intermedius IOC-EA-66, Enterobacter aerogenes IOC-EA-6, Serratia sp. IOC -EA-9, Clostridium sporogenes IOC-EA-99.
Another embodiment of the present invention provides an electro-biochemical process for reducing carbon dioxide and water said process comprising the steps of:
(a) providing a single chambered electrochemical system containing at least one anode and one cathode and a medium containing nitrogen, phosphorus and a micronutrient source, wherein the said cathode (s) contains biofilm of electro-active microbe (s) having ability hydrate CO2 and converting it into fuels and chemicals;
(c) connecting anode and cathode through a source of electrical energy;
(d) operating said source of electrical energy to extract electrons from anode and deliver electrons to cathode;
(e) adding the carbon dioxide source;
(f) incubating the electrochemical system with microorganisms to produce one or more chemical compounds in a vicinity of said cathode; and
(g) recovering the one or chemical compounds

Another embodiment of the present invention provides an electro-active microbe and their synergistic consortia adopted to form a biofilm under electric current, wherein electro-active microbe is capable of solubilizing/ carbon dioxide and converting it into organic molecules under electric current.
Another embodiment of the present invention provides an electro-active microbe described herein wherein electro-active microbe are selected from the group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EA-6, Serratia sp. IOC -EA-9, Clostridium sporogenes IOC-EA-99 alone or in combination.
DETAILED DISCRIPION OF INVENTION
While the invention is susceptible to various modifications and/or alternative processes and/or compositions, specific embodiment thereof has been shown by way of example in the drawings and tables and will be described in detail below. It should be understood, however that it is not intended to limit the invention to the particular processes and/or compositions disclosed, but on the contrary, the invention is to cover all modifications, equivalents, and alternative falling within the spirit and the scope of the invention as defined by the appended claims.

The graphs, tables, formulas, protocols have been represented where appropriate by conventional representations in the drawings, showing only those specific details that are pertinent to understanding the embodiments of the present invention so as not to obscure the disclosure with details that will be readily apparent to those of ordinary skill in the art having benefit of the description herein.
The following description is of exemplary embodiments only and is not intended to limit the scope, applicability or configuration of the invention in any way. Rather, the following description provides a convenient illustration for implementing exemplary embodiments of the invention. Various changes to the described embodiments may be made in the function and arrangement of the elements described without departing from the scope of the invention.
The terms "comprises", "comprising", or any other variations thereof, are intended to cover a non-exclusive inclusion, such that one or more processes or composition/s or systems or methods proceeded by "comprises... a" does not, without more constraints, preclude the existence of other processes, sub-processes, composition, sub-compositions, minor or major compositions or other elements or other structures or additional processes or compositions or additional elements or additional features or additional characteristics or additional attributes.
Definition:
For the purposes of this invention, the following terms will have the meaning as specified therein:
As used herein, the terms “Biofilm/s or Electro-active Biofilm/s or Electrogenic Biofilm/s”, when used in the context of the present invention refers to densely packed communities of microbes herein described in the present invention which grow on conductive materials and can achieve a direct electrochemical connection with the electrode surface using it as electron exchanger, without the aid of mediators.
As used herein, the terms “Electro-active Microbes or Microbes”, when used in the context of the present invention refers to microbes described herein in the present invention which have innate capability to receive electrons from solid electrodes and can use in their cell metabolism .
As used herein, the terms “Consortia of Microbes or Consortium of Microbes”, when used in the context of the present invention refers to a synergistic mixture of microbes described herein the present invention.
As used herein, the term “Conductive Electrode”, when used in the context of the present invention refers to a conductive electrode which may function as anode or cathode.
As used herein, the term “cathode”, when used in the context of the present invention refers to electrode of an electrochemical system that donates electrons to an electrochemical reaction.
As used herein, the term “anode”, when used in the context of the present invention refers to electrode of an electrochemical system that accepts electrons to an electrochemical reaction.
As used herein, the terms “Electro-biochemical system or Electro-biochemical cell”, when used in the context of the present invention refers to an electrochemical sytem in which electroactive microorganisms catalyse the anode and/or the cathode reaction.
The present invention provides a bio-assisted process of transforming carbon dioxide to fuels and chemical using an electro-biochemical process. The present invention relates to providing an efficient and cost-effective means of transforming carbon dioxide to fuels and chemicals using electro-biochemical process.

In one aspect the present invention provides for unique electro-active microorganisms capable of solubilising carbon dioxide in in vitro aqueous medium and transforming it to fuels and chemicals . The electro-active microbes of the present invention are capable of forming a biofilm under electric current supplied externally. This biofilm formed in capable of driving the solubilisation of Carbon dioxide in a bio-electrochemical process.

In another aspect the present invention provides for electro-active microbes selected from group comprising Citrobacter sp. IOC-EA-66 (deposited at Microbial Type Culture Collection and Gene Bank, Chandigarh, India, dated¬¬¬¬¬¬¬________having Accession No.__________), Enterobacter aerogenes IOC-EA-6 (deposited at Microbial Type Culture Collection and Gene Bank, , Chandigarh, India, dated¬¬¬¬¬¬¬________having Accession No.__________), Serratia sp. IOC -EA-9 (deposited at Microbial Type Culture Collection and Gene Bank, , Chandigarh, India, dated¬¬¬¬¬¬-________having Accession No.__________) and Clostridium sporogenes IOC-EA-99 (, Chandigarh, India, deposited at Microbial Type Culture Collection and Gene Bank, dated¬¬¬¬¬¬-________having Accession No.__________). The characteristics of these microbes are given in Table-1. The 16S rRNA sequences of these microbes are given as SEQ ID 1-4 and identify the type of the micro-organism. Accordingly, the microorganisms are well described. These electro-active microbes are capable of producing carbonic anhydrase enzyme which enables solubilisation of CO2 and convert in into fuels and chemicals in an electro-biochemical process.

In another aspect the present invention provides for a single chambered electro-biochemical cell containing aqueous medium and at least one cathode electrode and one anode electrode where the anode and the cathode being electrically connected to each other and electricity is supplied from external sources along with electrogenic biofilm of consortia of selected electro-active microbes as described herein in the present invention.

In another aspect the present invention provides for a single chambered electro-biochemical process for converting CO2 into fuels and chemicals. In particular the said single chambered electro-biochemical process provides for converting carbon dioxide into fuels and chemicals wherein the carbon dioxide source is selected wherein carbon dioxide source is selected from waste-water effluents, effluents from combustion process of coal, petroleum, methane, natural gas, biomass, organic carbon, an industrial process that releases carbon dioxide, carbon dioxide from geothermal sources and/or atmospheric carbon dioxide, any other fossil fuel source, waste source, etc.In general, any source of CO2 can be used. Another well know source of the supply of CO2 is externally wherein carbon dioxide can also be supplied as gas containing carbon dioxide in concentration of 0.028 to 100 vol% in pure form or in the form of mixture with other gases. .

In another aspect the present invention provides for an electro-biochemical process wherein the transformation of the carbon dioxide into fuels and chemicals is carried out by electo-active microbes or consortia thereof. The electro-active or electrogenic microbes as described herein in the present invention form a biofilm under the electric current on the conductive electrode, preferably cathode electrode. The biofilm of electro-active microbes is capable of solubilising the carbon dioxide in the reaction medium and transforming it into fuels and chemicals. This solubilisation effect is effectively carried out by the electro-active microbes because they are capable of producing carbonic anhydrase enzyme.

In another aspect the present invention provides for a source of external electric current wherein such source can be renewal electric source for example electric current generate from solar power,wind turbines, hydroelectric or biomass-fired electrical generators, or other renewable energy sources of electricity.
In another aspect the present invention provides that the electro-active microbes are capable of extracting electrons from cathode and use the electrons in the metabolic reaction and finally transfer them to CO2 which is reduced to various chemical compounds. The unique advantage of carrying out the electro-biochemical process of the present invention is such that there is no introduction of electron donor or electrons externally in the medium thereby enabling the electro-active microbes to use the cathode as electron source.
One aspect of the present invention provides a method of developing biofilm of electro-active microbes on cathode in the electro-biochemical system based device for reduction of CO2 disclosed in this invention or in some other similar purpose. The biofilm of the electro-active microbes can be developed in single chamber cell having medium containing nitrogen, phosphorus and micronutrient source. The medium in which biofilm of electro-active microbes is developed can also supplemented with microbial or plant extract (s) which facilitates the biofilm formation. Microbial extract include toluene extract of common yeast Saccharomyces cerevisiae, hot water extract of methanol extracted microalgae like Scenedesmus sp., Synechococcus, Chlorella pyrenoidosa. The aqueous extract from plant Grewia optiva and Aloe barbadensis. The microalgae extract or the plant extract used in the present invention is not a by-product neither it is a value added product obtained from the microalgae extract or the plant extract. This extract of microbial algae or plant extract is an unorganized, physically inseparable form of mixture (a raw mixture) which is used in the medium in the present invention. By no means is the micro-algae extract or the plant extract being used for the purpose patenting in the present invention. The micro-algae extract or the plant extract is used in absolute raw form wherein its active ingredients are not isolated neither known nor identified.

In another aspect the present invention provides multicarbon compounds or compounds that are produced from reduction of carbon dioxide or reduction of carbon dioxide source include but are not limited to methanol, ethanol, acetic acid, butanol, proponal, propionic acid, formic acid, butanedioicacid in mixture or individually or any other organic acid, alcohol, aldehyde, ketones, fatty acids, hydrocarbons with at least one carbon atom.
In another aspect the present invention provides exemplary conductive electrode materials may include, but are not limited to, carbon paper, carbon cloth, carbon felt, carbon wool, carbon foam, graphite, porous graphite, graphite powder, iron, carbonnano tubes, functionalized carbonnano tubes, metals, stainless steel, and combinations of these. The physical orientation of the electrodes can be any convenient mutual orientation relative to each other. One embodiment has the cathode and anode of equal dimensions and oriented with surfaces parallel to each other.
One aspect of the present invention provides for another unique advantage is that the reduction of carbon dioxide or the carbon dioxide source to multicarbon compounds with electrons donated from an electrode as the electron donor has been reported in handful pure culture till date. In this reduction process carbon and electron flow is directed primarily to the formation of extracellular products like organic acid, alcohol, aldehyde, ketones, fatty acids, hydrocarbons with at least one carbon atom, rather than biomass. Biomass typically requires extensive additional processing for chemical or fuel production.
In another aspect the present invention provides for a single chamber electro-biochemical cell to produce multi-carbon organic compounds from carbon dioxide and water. The system includes an electrical power supply, an anodic electrode capable of extracting electrons from water, a cathodic electrode having biofilm of microbial consortia that can use electrons derived from the cathodic electrode to fix carbon dioxide into organic compounds. In one embodiment, carbon dioxide is fixed to produce organic chemical having at least two carbon atoms.
In another aspect the present invention the biofilm of electro-active microbes on cathode can be stored for long time using various methods including but are not limited to immobilization of cathode along with biofilm on agarose, lyophilization and/or covering with selective permeable membrane. The stored biofilm can be effectively regenerated by growing them in growth media and this can quickly start the generation of compounds from CO2 under appropriate operating conditions.
In another aspect the present invention provides for production of a wide variety of organic compounds that can be produced, which can include but are not limited to alcohols, amino acids, sugars, fatty acids, aromatic compounds, diols, and the like.
In another aspect the present invention provides for the electric sources comprising of both renewal and non-renewal sources of electricity. The renewal source of the energy include but are not limited to solar energy, wind turbines, hydroelectric or biomass-fired electrical generators, or other renewable energy sources of electricity. In other embodiments, the source of electrical current can be any convenient source of electrical current, including a grid-connected power supply, a battery, a fuel-powered generator, an electrochemical cell, a microfuel cells, a bioelectrochemical cell with anodic chamber having bio-oxidation of wastewater.

Accordingly, the main embodiment of the present invention provides a biofilm of electro-active microbes comprising consortia of microbes wherein at least one microbe has an ability to solubilise carbon dioxide under electric current.
Another embodiment of the present invention provides a biofilm as herein described wherein the microbes are selected from the group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EZ-6, Serratia sp. IOC-EA-9 and Clostridium sporogenes IOC-EA-99.
One more embodiment of the present invention provides a biofilm as herein described wherein the consortia of microbes is active at a temperature of 10°C to 52°C.
Another embodiment of the present invention provides a biofilm as herein described, wherein the consortia of microbes is active at a pH ranging from 5-12.
Another embodiment of the present invention provides a biofilm as herein described wherein the biofilm can be stored by immobilization on agarose, lyophilisation and/or covering with selective permeable membrane and the like along with cathode.
Another embodiment of the present invention provides a biofilm as herein described wherein the biofilm is effective in bio-assisted transformation of carbon dioxide into fuels and chemicals.
One embodiment of the present invention provides a biofilm as herein described wherein the biofilms enable production of fuels and chemicals in an bio-electrochemical system.
One embodiment of the present invention provides a biofilm as herein described wherein the biofilm comprising microbial consortia is active in salinity range from 0.05-10%.
One embodiment of the present invention provides a biofilm as herein described wherein at least one of electro-active microbes is capable of producing carbonic anhydrase enzyme.
Another embodiment of the present invention provides a method of preparing biofilm of electro-active microbes on a conductive electrode said method comprising the steps of:
(a) providing a single chambered electrochemical cell comprising of atleast one cathode and one anode conductive electrode and a growth medium;
(b) adding a carbon dioxide source;
(c) inoculating the cell with consortia of microbes under electric current;
(d) adding plant/microbial extract (s) which improves the biofilm formation
(e) growing the microbe consortia;
(f) replacing the medium with fresh medium; and
(g) obtaining a stable biofilm of electro-active microbes.

One more embodiment of the present invention provides a method as herein described wherein the biofilm of electroactive microbes formed on conductive electrode that is cathode.
One more embodiment of the present invention provides a method as herein described, wherein the microbes are selected from the group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EZ-6, Serratia sp. IOC-EA-9 and Clostridium sporogenes IOC-EA-99.
One more embodiment of the present invention provides a method as herein described wherein the consortia of microbes is active at a temperature of 10°C to 52°C.
One more embodiment of the present invention provides a method as herein described wherein the consortia of microbes is active at a pH ranging from 5-12.
One more embodiment of the present invention provides a method as herein described wherein carbon dioxide source is selected from group comprising of is selected from group comprising waste-water effluents, effluents from combustion process of coal, petroleum, methane, natural gas, biomass, organic carbon, an industrial process that releases carbon dioxide, carbon dioxide from geothermal sources and/or atmospheric carbon dioxide.
One more embodiment of the present invention provides a method as herein described wherein the biofilm of the electro-active microbes is formed over or on the cathode electrode.
One more embodiment of the present invention provides a method as herein described where the biofilm formed on cathode can be stored by immobilization on agarose, lyophilisation and/or covering with selective permeable membrane and the like.
One more embodiment of the present invention provides a method as herein described wherein the biofilm is effective in bio-assisted transformation of carbon dioxide into fuels and chemicals.
One more embodiment of the present invention provides a method as herein described wherein the biofilms enable production of fuels and chemicals in a bio-electrochemical system.
One more embodiment of the present invention provides a method as herein described wherein the biofilm comprising microbial consortia is active in salinity range from 0.05-10%.
One more embodiment of the present invention provides a method as herein described wherein at least one of electro-active microbes is capable of producing carbonic anhydrase enzyme.
Another embodiment of the present invention provides a single chambered electro-biochemical cell comprising least one cathode, at least one anode, a reaction medium and biofilm of electro-active microbes wherein electro-active microbes having an ability to solubilise carbon dioxide under electric current.
Another embodiment of the present invention provides a single chambered electro-biochemical cell as herein described wherein the electroactive microbes capable of producing multi-carbon organic compounds from carbon dioxide and water.
Another embodiment of the present invention provides a single chambered electro-biochemical cell as herein described wherein the biofilm electroactive microbes are capable of solubilising carbon dioxide under electric current.
Another embodiment of the present invention provides a single chambered electro-biochemical cell as herein described wherein the electro-active microbes are selected from group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EZ-6, Serratia sp. IOC-EA-9 and Clostridium sporogenes IOC-EA-99.
Another embodiment of the present invention provides a single chambered electro-biochemical cell as herein described wherein the electroactive microbes form a biofilm on the conductive cathode of the single chambered bio-electrochemical cell.
Another embodiment of the present invention provides a single chambered electro-biochemical cell as herein described wherein electric current can be sourced from a renewal electric source selecting from solar energy or solar cell.
Another embodiment of the present invention provides an electro-biochemical process for reducing carbon dioxide and water said process comprising the steps of:
(a) providing a single chambered electrochemical system containing at least one anode and one cathode and a medium containing nitrogen, phosphorus and a micronutrient source;
(b) connecting anode and cathode through a source of electrical energy;
(c) operating said source of electrical energy to extract electrons from anode and deliver electrons to cathode;
(d) inoculating the cell with consortia of microbes wherein at least one microbe has an ability to produce extracellular carbonic anhydrase enzyme during growth under electric current;
(e) adding the carbon dioxide source;
(f) adding plant/microbial extract (s) which improves the biofilm formation
(g) allowing the bacteria to form biofilm on cathode
(h) replacing the medium with fresh medium
(i) getting a stable biofilm of electroactive microbe
(i) incubating the electrochemical system to produce one or more chemical compounds in a vicinity of said cathode; and
(j) recovering the one or chemical compounds

Another embodiment of the present invention provides an electro-biochemical process as described herein wherein the microbes are selected from the group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EA-6, Serratia sp. IOC -EA-9, Clostridium sporogenes IOC-EA-99.
Another embodiment of the present invention provides an electro-biochemical process as described herein wherein the microbes are active at a temperature from 10°C to 52°C.
Another embodiment of the present invention provides an electro-biochemical process as described herein wherein the carbon dioxide source is selected from group comprising waste-water effluents, effluents from combustion process of coal, petroleum, methane, natural gas, biomass, organic carbon, an industrial process that releases carbon dioxide, carbon dioxide from geothermal sources and/or atmospheric carbon dioxide.
Another embodiment of the present invention provides an electro-biochemical process as described herein wherein carbon dioxide is delivered in the cell in the form of macrobubbles ( 1-2 mm in size), in the form of micro- bubbles and nanobuubles ( 25 micron or less in size). Delivering CO2 in microbubble and nanobubble further improves the CO2 dissolution, its availability to cells and improves nutrient solubility. The microbubble and nanobubble can ne generated using the microbubble generator or other such system known for this purpose in prior art and maacrobubble can be used by using sponge diffuser.
Another embodiment of the present invention provides an electro-biochemical process for reducing carbon dioxide and water said process comprising the steps of:
(a) providing a single chambered bio-electrochemical system containing at least one anode and one cathode where cathode having biofilm of electroactive microbes, wherein the electro-active microbe (s) on said cathode having ability hydrate CO2 and converting it into fuels and chemicals
(b) including a medium containing nitrogen, phosphorus and a micronutrient source to the said bio-electrochemical system;
(c) connecting anode and cathode through a source of electrical energy;
(d) operating said source of electrical energy to extract electrons from anode and deliver electrons to cathode;
(e) Delivering CO2 in the microbubble size;
(f) incubating the electrochemical system with microorganisms to produce one or more chemical compounds in a vicinity of said cathode; and
(g) recovering the one or chemical compounds

Another embodiment of the present invention provides an electro-biochemical process as described herein wherein the electrode material includes graphite, graphite felt, porous graphite, graphite powder carbon paper, carbon cloth, carbon felt, carbon wool, carbon nano-tubes, carbon foam, stainless steel as such or chemically modified, and combinations thereof.
Another embodiment of the present invention provides an electro-biochemical process as described herein wherein the voltage applied with a power supply between the anode and the cathode is between 0 and -1 V , preferably between -0.1 and - 0.7 V, more preferably between -0.2 and -0.5 V.
Another embodiment of the present invention provides an electro-biochemical process as described herein wherein the microbial/plant extract include extract Saccharomyces cerevisiae, Scenedesmus sp., Synechococcus sp. Chlorella pyrenoidosa and Grewiaoptiva and Aloe barbadensis.
Another embodiment of the present invention provides an electro-biochemical process as described herein wherein the microbial/plant extract include extract can be used individually or in combination.
Another embodiment of the present invention provides an electro-active microbe adopted to form a biofilm under electric current, wherein electro-active microbe is capable of solubilizing carbon dioxide under electric current.
Another embodiment of the present invention provides an electro-active microbe described herein wherein electro-active microbe are selected from the group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EA-6, Serratia sp. IOC -EA-9, Clostridium sporogenes IOC-EA-99 alone or in combination.
Another embodiment of the present invention provides an electro-active microbe described herein wherein the electro-active microbe is active at a pH 5-12.
Another embodiment of the present invention provides an electro-active microbe described herein wherein the electro-active microbe is active at in salinity ranging from 0.5-15%.
Another embodiment of the present invention provides an electro-active microbe described herein wherein the electro-active microbe is active at a temperature from 10°C to 52°C.
Another embodiment of the present invention provides an electro-active microbes described herein wherein the electro-active microbes are capable of forming consortia of electro-active microbes.

In another embodiment the present invention provides an eletrogenic biofilm of consortia of microorganism on cathode is consisting of four microbes.

In another embodiment, the eletrogenic biofilm of consortia of microorganism on cathode that is able to improving solubility CO2 and generate a chemical having at least one carbon atom using carbon dioxide as a source of carbon

In a further embodiment the present invention provides the enzyme produced by electrogenic or electro-active microbes is carbonic anhydrase.

In a further embodiment, the pH of the medium in which the microbes grow, or the electro-biochemcial process is carried out is > 8.

In another embodiment the present invention provides that the carbon dioxide is transformed to alcohol, organic acid, aldehyde, ketone hydrocarbon, fatty acids having at least one carbon atoms.

Another embodiment of the present invention provides that the carbon dioxide is transformed to alcohol, organic acid, aldehyde, ketone hydrocarbon, fatty acids having more than 5 carbon atoms.
In one embodiment, the biofilm contain microbes tolerant to alcohols (30%) and other product.

In another embodiment, the biofilm comprises microorganisms derived from a source selected from the group consisting of waste water, soil, microbiologically corroded metal surface, activated sludge and coal mining site soil.

The invention will now be explained with the help of following examples. However, the scope of the invention should not be limited to these examples as the person skilled in the art can easily vary the proportion of the ingredients and combinations.
EXAMPLES
Example 1
Electro-biochemical Cell
A single chambered cell with ports for anode, cathode, reference electrode, sampling, CO2 injection and pressure control was constructed. Sampling ports sealed with butyl stoppers, and aluminum crimps were added to the sides and top of each chamber, while electrodes were inerted using respective port and sealing ring. The volume of each chamber, with electrode, was a 250 ml of medium with a 150 ml headspace. Electrodes were unpolished graphite rods. New electrodes were soaked in 1 N HCl, which was changed daily until extractable Fe(II) was below detection. After each use, electrodes were washed in 1 N HCl and 1 N NaOH to remove possible metal and biomass contamination. Connections were made with threaded watertight connectors . Whole assembly was autoclaved before use. A reference electrode was sterilized by immersing the electrode and stopper in 5 N HCl for 5 min, rinsing in ethanol, and allowing the electrode to fully dry before placing it in a sampling port.

The cell was flushed with sterile, anaerobic gas (80:20 N2:CO2), filled with the appropriate medium (described herein below) and connected to a potentiostat. The medium in the cell was stirred at 180 r.p.m. with a magnetic stir bar. The chambers were allowed to equilibrate at -500 mV (versus Ag/AgCl) for 24 h before inoculating with cultures. Control chambers were treated identically, but were not connected to a potentiostat, and no current was able to flow to the working electrode. In yet another control to no microbe was inoculated in cathode and all other conditions were kept identical. The cathode electrode and anode is connected by an electrically conductive wire to facilitate the transfer of electrons generated at anode to the cathode.During operation, the cathode was poised at -600 mV vs Ag/AgClunder 80% nitrogen/20% CO 2 headspace.

Media:
The media used consisted (g/l) of 0.5 Na2 CO3, 2.0 NaHCO3, 2.0 KH2PO4 2.0 K2HPO4 , 0.1 MgSO4, 0.5 (NH4)2SO4, 0.5 KNO3, 2.0 ZnSO4, , 2.0 Yeast exract, 1.5 Tryptone, 0.5 NaCl and 2 ml. Trace element. The trace element solution (gram per liter) comprises Nitrilotriacetic acid (0.1), FeSO4.7H2O (0.2), MnCl2.4H2O (0.005), CoCl2.6H2O (0.02), CaCl2.2H20 (0.08), CuCl2.H2O (0.03), H3BO3 (0.02), Na2MoO4 (0.02), Na2SeO3 (0.06), NiSO4 (0.03), SnCl2 (0.03).
Reaction conditions:
The reaction is conducted at pH ranging from 5-12 and temperature from 5-60°C. Chamber contains single or multiple electrodes which may be made up of same material or of different material. Both electrodes are contacted by electro-conductive wire. A potential difference maintained between anode and cathode by potentiostat or some other method or device employing procedure like chronoamperometry, cycylovoltametery etc. Electrons extracted from water at anode were delivered to cathode at potential difference of - 0.200 to -0.800V. Chamber is inoculated with water (7-10%,w/v) or soil sample (5-10% w/w) obtained from various habits like aeration tank of refinery, coal mining sites, microbially corroded metal piece, soil, sea sediments, mangrove sediments,compost site etc. The chamber was continuously with gas mixture containing Co2 or pure CO2. After 10 days the liquid medium from cathode chamber is removed and fresh medium was filled aseptically. This step is repeated for 4 time in the interval of 12 hours or unless until stable current consumption was not observed. This results in the development of a stable biofilm over conductive electrode which can effectively reduce CO2 using electrons in a bioelectrochemical system.

Subsequently, the cathode is taken out and washed with normal saline aseptically. This cathode containing biofilm of electroactive bacteria can be used in CO2 reduction system disclosed in this invention. This is also useful in microbial fuel cell or other bioelectrochemical system.

Example 2

Extraction of plant/microbial extract:

Addition of plant extract ( 0.5-10 microL/ml of media) from one of these plant or their combination lead to stimulation the biofilm formation and time taken for development of stable and active biofilm was sigficicantly low when these plant extract were used. Their exact mechanism of functioning is not known but we hypothesized thattheseplant/microbial extracts contains some compounds like sesquiterpene alcohols, lactones, polysaccharides which served as a signal for biofilm formation transduced via the kinases. In addition, polysaccharides or other compounds present are probably used as a source of sugars for the initial synthesis of the matrix exopolysaccharide of biofilm.

For extraction from algae, five hundred grams of freshly collected wet algae was subjected for sonication at maximum speed for 2 min as subsequently liquid extracted with 200 ml of methanol at room temperature using a glass separating was carried out. Remaining solid residuewas washed twice in distilled water for 5 min and then placed in 200 ml of distilled water and autoclaved at 121°C for 25 min. These preparations were filtered through sterile gauzes of 1-mm porosity subsequently; algal extract was centrifuged at 3000 rpm for 6 min to remove cell coarse material if any. This was lyophilized and suspended in 100 ml of autoclaved water and used.

For extraction of plant species, the stems, branches, leaves were used and above procure was used.

For extraction from yeast, 50 g of spent yeast was autovaed in 100 ml water. The resulting slurry was extracted with toluene (200 ml) and solvent layer was evaporated. The dried powder was dissolved in 50 ml water and used for present purpose.

These extracts can be used in combination or separately. The effectiveness of these plant extract to improve biofilm was studied in single chamber cell and inculcating them by bacteria in presence or absence of plant extract. At the different interval, the current consumption was recorded. It is known that when a stable biofilm is formed, the current consumption become stable. Subsequently, cathode was taken out and planktonic cells were removed, the wells were washed 3 times in 0.85% NaCl and the biofilms were stained with 0.1% crystal violet for 10 min. The dye was solubilised with 150 µl 75% ethanol per well and the absorbance was measured on a plate reader at 550 nm. Higher absorbance at 550 nm shows higher growth of biofilm over the cathode. In some of experiment, cathode was subjected for SEM analysis to see the integrity of the biofilm in presence and absence of plant extract. . Result from this experiment showed that the presence of selected plant/microbial extract, faster and higher biofilm growth is observed. The time for stable current reduced from 10 days to 4 days when the plant/microbial extract used.

Example 3
Development of Consortia
Microbes from the biofilm where purified using the prior art used commonly in microbiological laboratory. Individual bacteria was evaluated its ability to form biofilm on cathode under electric current, reduce CO2 and consume current as well as to produce extracellular carbonic anhydrase (CA) enzyme under the experimental conditions. The bacteria which was highest desired activity was coupled together to reap the benefits of their synergy. In the consortia minimum one bacteria is having ability to produce extracellular carbonic anhydrase enzyme and effectively improve the solubility of the carbon dioxide. The carbonic anhydrase activity was measured by p-nitrophenylacetate hydrolysis by CA following the method of Smith and Ferry (Smith KS, Ferry JG.(1999) J.Bacteriol. Vol.181, pages 6247–6253) and Wilbur, KM Anderson, (1948; J Biol Chem.Vol.176:147–54).

The use of consortia improves, the interspecies cell-cell interaction the electron delivery and metabolic reactions. Moreover, higher CO2 delivery due to extracellular carbonic anhdrase enzyme result with higher yield in less time.
Evaluation of consortia to reduce Carbon Dioxide and identification of product:
The biofilm developed over a cathode electrode transferred to single chambered cells containing suitable media and other electrodes. The cell may contain single cathode or multiple cathode having different or similar biofilm. Electrodes were separated to avoid short circuit. Voltage was applied to circuit through the electric source and cathode poised at -200 to -900 mV. CO2 was continuously sparged at 10-20ml/min rate as only carbon source very near to cathode having biofilm. This assembly was kept under stirring at temperature 10- 65°C. Once, the current consumption become stable and proper biofilm is sample was withdrawn and analyzed by gas chromatography for presence of the fuels and hydrocarbons.
Example 4
Isolation of the microbes:

For the isolation of microbes with desired characteristics i.e., having ability to accept from cathode, form biofilm under electric current on cathode and produce extracellular carbonic anhydrase under electric conditions and reduce CO2, a 150 ml single chamber cell containing 100 ml of medium was taken. The media contained (g/l) of 0.5 Na2 CO3, 2.0 NaHCO3, 2.0 KH2PO4 2.0 K2HPO4 , 0.1 MgSO4, 0.5 (NH4)2SO4, 0.5 KNO3, 2.0 ZnSO4, , 2.0 Yeast exract, 1.5 Tryptone, 0.5 NaCl and 2 ml. Trace element. The trace element solution (gram per liter) comprises Nitrilotriacetic acid (0.1), FeSO4.7H2O (0.2), MnCl2.4H2O (- 0.005), CoCl2.6H2O (0.02), CaCl2.2H20 (0.08), CuCl2.H2O (0.03), H3BO3 (0.02), Na2MoO4 (0.02), Na2SeO3 (0.06), NiSO4 (0.03), SnCl2 (0.03). In this media aqueous extract of Aloe barbadensis was added at the concentration of 2 ppm.
Chamber contains one anode and one cathode both of graphite rods and was sterilized by autoclaving. Both electrodes are contacted by electro-conductive wire. Chamber is inoculated with soil sample (5-10% w/w) obtained from coal mining sites. A potential difference maintained between anode and cathode by potentiostat and electrons extracted from water at anode were delivered to cathode at potential difference -0.800V. The chamber was continuously with gas mixture containing CO2. After 10 days the liquid medium from cathode chamber is removed and fresh medium was filled aseptically. This step is repeated for 4 time in the interval of 12 hours and stable current consumption was observed. This result in the development of a stable biofilm over conductive electrode which can effectively reduce CO2 using electrons in an electro-biochemical system.Subsequently, the cathode is taken out and washed with normal saline aseptically.Microbes from the biofilm where purified using the prior art used commonly in microbiological laboratory. Individual bacteria was evaluated its ability to form biofilm on cathode under electric current, reduce CO2 and consume current as well as to produce extracellular carbonic anhydrase enzyme under the experimental conditions. The bacteria which was highest desired activity was coupled together to reap the benefits of their synergy. In the consortia minimum one bacterium is having ability to produce extracellular carbonic anhydrase enzyme and effectively improve the solubility of the carbon dioxide. A such microbial consortia was made consisted of four bacteria develop in which all bacteria having desired characteristics i.e., having ability to accept from cathode, form biofilm under electric current on cathode and produce extracellular carbonic anhydrase under electric conditions and reduce CO2.

The member of consortia were identified by sequencing their 16S rDNA gene and biochemical testing as Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EA-6, Serratia sp. IOC -EA-9, Clostridium sporogenes IOC-EA-99.(Please provide these sequences).
These microbes singly or in combination used for their ability to reduce Co2 under electric current.
Characteristic Features of the Microbial species
Table-1 Characteristics of microbes used in present invention

Characteristics Citrobacter sp IOC-EA-66 Enterobacter aerogenes IOC-EA-6 Serratia sp. IOC -EA-9, Clostridium sporogenes IOC-EA-99
Shape of the cells Coco-bacillus Rod shaped Coco-bacillus Rod shaped, sub terminal spore
Gram’s reaction Negative Negative Negative Positive
Aerobic/anaerobic Facultative anaerobe Facultative anaerobe Facultative anaerobe Anaerobe
Motility Motile Motile Motile Motile
Temperature tolerance (degree C) 10-52 8-56 15-52 10-70
pH tolerance 5-12 5-11 5-11 5-12
Salinity tolerance Up to 10% Up to 10% Up to 11% Up to 10%
Ethanol and other product tolerance >30% >20% >25% >21%
Ability to grow under electric current Yes Yes Yes Yes
Ability to form biofilm on cathode Yes Yes Yes Yes
Ability to produce carbonic anhydrase under electric current Yes Yes Yes No
Ability to reduce CO2 under electric current Yes Yes Yes Yes
Glucose (gas and/or acid) Gas +
Acid - Gas +
Acid+ Gas +
Acid- Gas +
Acid+
Lactose (gas and/or acid) Gas +
Acid- Gas +
Acid+ Gas +
Acid- Gas +
Acid+
Sucrose (gas and/or acid) Acid –
Gas+ Acid +
Gas + Acid +
Gas - Acid –
Gas-
Salicin (acid) + + + -
Inositol (acid) - + - -
Indole - - - -
Methyl red + - - -
Voges-Proskauer + - - +
Citrate + + + -
Gelatin - + + -
Phenylalanine - - - -
Urease - + + +
Hydrogen sulphide + - - +
KCN + + + ND
Gluconate - + + -
Malonate - + + -
Lysine decarboxyase - + + ND
Ornithine decaroxylase
-
+ + ND
Cellobiose (acid and or gas) Acid +
Gas + Acid +
Gas - Acid -
Gas + Acid -
Gas +
Glycerol ( Acid and/or gas) Acid +
Gas + Acid +
Gas + Acid +
Gas + Acid -
Gas +
Oxidase produced - - - -
Catalase produced + - + +
DNAase produced - - + +
Lipase produced - + - -
Nitrate reduction + + + +

- + is positiveIs negative
- ND: Not done
Growth of Individual bacteria:
Individual member of consortia was grown using media contained (g/l) of 0.5 Na2 CO3, 2.0 NaHCO3, 2.0 KH2PO4 2.0 K2HPO4 , 0.1 MgSO4, 0.5 (NH4)2SO4, 0.5 KNO3, 2.0 ZnSO4, , 2.0 Yeast exract, 1.5 Tryptone, 0.5 NaCl and 2 ml. Trace element. The trace element solution (gram per liter) comprises Nitrilotriacetic acid (0.1), FeSO4.7H2O (0.2), MnCl2.4H2O (- 0.005), CoCl2.6H2O (0.02), CaCl2.2H20 (0.08), CuCl2.H2O (0.03), H3BO3 (0.02), Na2MoO4 (0.02), Na2SeO3 (0.06), NiSO4 (0.03), SnCl2 (0.03) at 45 degree c for 48 hrs. Subsequent to this cells were harvested by centrifugation and wash asceptically using normal saline. Cells were suspended in normal saline at the concentration of 6.0 × 108 cells/ml. If individually used, 2 ml of said suspension was used and when mixture was used 1 ml suspension of each bacteria was used.
Example 5
Evaluation of consortia to reduce Carbon dioxide and identification of product:
Single chambered cells with ports for anode, cathode, reference electrode, sampling, CO2 injection and pressure control were used. The volume chamber, with electrode, was a 250 ml of medium with a 100 ml headspace. The media contained (g/l) of 0.5 Na2 CO3, 2.0 NaHCO3, 2.0 KH2PO4 2.0 K2HPO4 , 0.1 MgSO4, 0.5 (NH4)2SO4, 0.5 KNO3, 2.0 ZnSO4, , 0.5 NaCl and 2 ml. Trace element. The trace element solution (gram per liter) comprises Nitrilotriacetic acid (0.1), FeSO4.7H2O (0.2), MnCl2.4H2O (- 0.005), CoCl2.6H2O (0.02), CaCl2.2H20 (0.08), CuCl2.H2O (0.03), H3BO3 (0.02), Na2MoO4 (0.02), Na2SeO3 (0.06), NiSO4 (0.03), SnCl2 (0.03). In this media aqueous extract of Aloe barbadensis was added at the concentration of 2 ppm.
The graphite stick cathodes and anodes, (both in 1 × 3 cm) were suspended in the reactor. In different set of cell, two, three or more cathode was used parallel and hanged used electroconductive wire. Electrodes were separated to avoid short circuit. The system was autoclaved and subsequently, inoculated with different type of bacteria and their combinations subsequent to polarization for 12 hrs in abiotic conditions. In combination, at least a bacteria producing extracellular carbonic anhydrase was also included.Voltage was applied to circuit through the electric source and cathode poised at -700 mV. CO2 was continuously sparged at 20ml/min rate as only carbon source very near to cathode having biofilm. This assembly was kept under stirring at temperature 35°C. Once, the current consumption become stable and proper biofilm is sample was withdrawn and analyzed by gas chromatography for presence of the fuels and hydrocarbons. Table-2 shows the current consumption and carbonic anhydrase enzyme production by different bacteria.
For higher yield of the consortia we hypothesized that one product from one microbes are used by other bacteria and cell-cell interaction improving the electron deliver to them along with higher CO2 delivery due to extracellular carbonic anhydrase enzyme result with higher yield in less time.

Table:2
Current consumption by individual bacteria and consortia when graphite cathode poised at -0.7V
S.No. Bacteria on cathode Coulombic efficiency Carbonic anhydrase activity
1 Citrobacter sp. IOC-EA-66 76 67 U/ml
2 Enterobacteraerogenes IOC-EA-6 86 12 U/ml
3 Serratia sp. IOC -EA-9 69 8 U/ml
4 Clostridium sporogenes IOC-EA-99 89 8 U/ml
5 Mixture of 1&2&3&4 in one cathode 92 190 U/ml
6 Biofilm of 1&2&3&4 each one in one cathode 97 179 U/ml

When biofilm of all microbes in one cathode, the major product which formed under the experimental conditions are butanol and C-6 fatty acid (Hexanoic acid).The cumulative concentration of the major product formed by the mixture of bacteria was 1.6g/l and 4 g/l for butanol and C-6 fatty acid (Hexanoic acid), respectively. When biofilm of different microbes was developed on different cathode, the major product which formed under the experimental conditions methanol (4g/l). No organic products were produced in the absence of microorganisms. These results showed that consortia could accept electrons from electrodes with the reduction of CO2 and that most of the electrons transferred from electrodes to cells were converted towards extracellular product rather than biomass production.
Experiment 3
In this experiment, four cathodes were used and in each cathode biofilm of one member of consortia was developed. The temperature of cell was kept 20 degree C. Rest all experimental conditions were kept same as mentioned above.
Table 3:
S.No. Bacteria on cathode Coulombic efficiency Carbonic anhydrase activity Yield
1. Biofilm of 1&2&3&4 each one in one cathode 99 200 U/ml C6 fatty acid
5.2 g/l
2. Control with bacteria - - Nil
3. Control without current - 15 u/ml Nil
Experiment-4
It has same experimental conditions, as in experiment-3 except it was run in continuous in flow through mode in which fresh media anaerobically introduced at 0.2ml/min dilution rate of 0.08 per hour. The turbidity in media eve not seen in media after 6 month of continuous operation and biofilm to produce butanol and fatty acid at the 1g/l/d and 1.5 g/l/d, respectively with traces of other components. The total coloumic efficiency was more than 98%.
,CLAIMS:We Claim:
1. A biofilm of electro-active microbe or their synergistic consortium wherein at least one microbe has an ability to solubilise carbon dioxide and convert it to fuel and chemicals under electric current.

2. The biofilm as claimed in claim 1, wherein the microbes are selected from the group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EZ-6, Serratia sp. IOC-EA-9 and Clostridium sporogenes IOC-EA-99.

3. The biofilm as claimed in claim 1, wherein the consortia of microbes is active at a temperature of 10°C to 52°C.

4. The biofilm as claimed in claim 1, wherein the consortia of microbes is active at a pH ranging from 5-12.

5. The biofilm as claimed in claim 1, wherein the pre-formed biofilm of microbes on cathode can be used.

6. The biofilm as claimed in claim 5, wherein the biofilm can be stored by immobilization on agarose, lyophilisation and/or covering with selective permeable membrane and the like.

7. The biofilm as claimed in claim 1, wherein the biofilm is effective in bio-assisted transformation of carbon dioxide into fuels and chemicals.

8. The biofilm as claimed in claim 1, wherein the biofilms enable production of fuels and chemicals in an bio-electrochemical system.

9. The biofilm as claimed in claim 1, wherein the biofilm comprising microbial consortia is active in salinity range from 0.05-10%.

10. The biofilm as claimed in claim 1, wherein at least one of electro-active microbes growing on cathode is capable of producing carbonic anhydrase enzyme.

11. The biofilm as claimed in claim 1, wherein at least one of electro-active microbes is capable of producing carbonic anhydrase enzyme.

12. A method of preparing biofilm of electro-active microbes on a conductive electrode said method comprising the steps of:
(a) providing a single chambered electrochemical cell comprising of at least one cathode and one anode conductive electrode and a growth medium;
(b) adding a carbon dioxide source;
(c) inoculating the cell with consortia of microbes under electric current;
(d) adding plant/microbial extract (s) which improves the biofilm formation;
(e) growing the microbe consortia (f) Replacing the medium with fresh medium; and
( g) obtaining a stable biofilm of electro-active microbes.

13. The method as claimed in claim 12, wherein biofilm of electroactive microbes formed on conductive electrode that is cathode.

14. The method as claimed in claim 12, wherein the microbes are selected from the group consisting of Citrobacter sp.IOC-EA-66, Enterobact eraerogenes IOC-EZ-6, Serratia sp. IOC-EA-9 and Clostridium sporogenes IOC-EA-99 and their combinations

15. The method as claimed in claim 12, wherein the consortia of microbes is active at a temperature of 10°C to 52°C.

16. The method as claimed in claim 12, wherein the consortia of microbes is active at a pH ranging from 5-12.

17. The method as claimed in claim 12, wherein carbon dioxide source is selected from group comprising of is selected from group comprising waste-water effluents, effluents from combustion process of coal, petroleum, methane, natural gas, biomass, organic carbon, an industrial process that releases carbon dioxide, carbon dioxide from geothermal sources and/or atmospheric carbon dioxide.

18. The method as claimed in claim 12, wherein carbon dioxide is delivered in the cell in the form of macrobubbles (1-2 mm in size) and/or in the form of micro- bubbles and nanobuubles ( 25 micron or less in size).

19. The method as claimed in claims 12-143 wherein the biofilm of the electro-active microbes is formed over or on the cathode electrode.

20. The method as claimed in claim 12, where the biofilm formed on the cathode can be stored by immobilization on agarose, lyophilisation and/or covering with selective permeable membrane and the like.

21. The method as claimed in claim 12, wherein the biofilm is effective in bio-assisted transformation of carbon dioxide into fuels and chemicals.

22. The method as claimed in claim 12, wherein the biofilms enable production of fuels and chemicals in a bio-electrochemical system.

23. The method as claimed in claim 12, wherein the biofilm comprising microbial consortia is active in salinity range from 0.05-10%.

24. A single chambered bio-electrochemical cell comprising least one cathode, at least one anode, a reaction medium and biofilm of electro-active microbes wherein electro-active microbes having an ability to solubilise carbon dioxide under electric current.

25. The single chambered bio-electrochemical cell as claimed in claim 24, wherein the electroactive microbes capable of producing multi-carbon organic compounds from carbon dioxide and water.

26. The single chambered bioelectrochemical cell as claimed in claims 24-25, wherein the biofilm electroactive microbes are capable of solubilising carbon dioxide under electric current.

27. The single chambered bio-electrochemical cell as claimed in claim 24, wherein the electro-active microbes are selected from group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EZ-6, Serratia sp. IOC-EA-9 and Clostridium sporogenes IOC-EA-99.

28. The single chambered bio-electrochemical cell as claimed in claim 24, wherein the electroactive microbes form a biofilm on the conductive cathode of the single chambered bio-electrochemical cell.

29. The single chambered bio-electrochemical cell as claimed in claim 24, wherein electric current can be sourced from a renewal electric source selecting from solar energy or solar cell.

30. The bioelectrochemical process for reducing carbon dioxide and water said process comprising the steps of:
(a) providing a single chambered electrochemical system containing at least one anode and one cathode and a medium containing nitrogen, phosphorus and a micronutrient source;
(b) connecting anode and cathode through a source of electrical energy;
(c) operating said source of electrical energy to extract electrons from anode and deliver electrons to cathode;
(d) inoculating the cell with consortia of microbes wherein at least one microbe has an ability to produce extracellular carbonic anhydrase enzyme during growth under electric current;
(e) adding the carbon dioxide source;
(f) including plant/microbial extract (s) which improves the biofilm formation;
(g) allowing the bacteria to form biofilm on cathode
(h) replacing the medium of steps (a-g) with fresh medium
(i) getting a stable biofilm of electroactive microbe
(j) incubating the electrochemical system to produce one or more chemical compounds in a vicinity of said cathode; and
(k) recovering the one or chemical compounds

31. The process as claimed in claim 30, wherein the microbes are selected from the group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EA-6, Serratia sp. IOC -EA-9, Clostridium sporogenes IOC-EA-99.

32. The process as claimed in claims 30-31, wherein the microbes are active at a temperature from 10°C to 52°C.

33. The process as claimed in claim 30, wherein the carbon dioxide source is selected from group comprising waste-water effluents, effluents from combustion process fo coal, petroleum, methane, natural gas, biomass, organic carbon, an industrial process that releases carbon dioxide, carbon dioxide from geothermal sources and/or atmospheric carbon dioxide.

34. The process as claimed in claim 30, wherein the electrode material includes graphite, graphite felt, porous graphite, graphite powder carbon paper, carbon cloth, carbon felt, carbon wool, carbon nano-tubes carbon foam, stainless steel as such or chemically modified, and combinations thereof.

35. The process as claimed in claim 28, wherein the voltage applied with a power supply between the anode and the cathode is between 0 and -1V , preferably between -0.1 and - 0.7 V, more preferably between -0.2 and -0.5 V.

36. The process as claimed in claim 30, wherein the microbial/plant extract include extract Saccharomyces cerevisiae, Scenedesmus sp., Synechococcus sp. Chlorella pyrenoidosaandGrewiaoptiva and Aloe barbadensis.

37. The process as claimed in claim 30, wherein carbon dioxide is delivered in the cell in the form of macrobubbles (1-2 mm in size), in the form of micro- bubbles and nanobuubles (25 micron or less in size).

38. The process as claimed in claim 30, wherein the microbial/plant extract include extract can be used individually or in combination.

39. An electro-active microbe adopted to form a biofilm under electric current, wherein electro-active microbe is capable of solubilizing carbon dioxide under electric current.

40. The electro-active microbe as claimed in claim 39, wherein electro-active microbe are selected from the group consisting of Citrobacter sp. IOC-EA-66, Enterobacter aerogenes IOC-EA-6, Serratia sp. IOC -EA-9, Clostridium sporogenes IOC-EA-99 alone or in combination.

41. The electro-active microbe as claimed in claims 39-40, is active at a pH 5-12.

42. The electro-active microbe as claimed in claims 39-40, is active at in salinity ranging from 0.5-10%.

43. The electro-active microbe as claimed in claims 39-40, is active at a temperature from 10°C to 52°C.

44. The electro-active microbe as claimed in claims 39-40, wherein the said microbes are capable of forming consortia of electro-active microbes.

45. An electro-biochemical process for reducing carbon dioxide and water said process comprising the steps of:
(a) providing a single chambered bio-electrochemical system containing at least one anode and one cathode where cathode having biofilm of electroactive microbes, wherein the electro-active microbe (s) on said cathode having ability hydrate CO2 and converting it into fuels and chemicals;
(b) adding a medium containing nitrogen, phosphorus and a micronutrient source to the said bio-electrochemical system;
(c) connecting anode and cathode through a source of electrical energy;
(d) operating said source of electrical energy to extract electrons from anode and deliver electrons to cathode;
(e) Delivering Co2 in the microbubble size;
(f) incubating the electrochemical system with microorganisms to produce one or more chemical compounds in a vicinity of said cathode; and
(g) recovering the one or chemical compounds