Bio-assisted process for conversion of mixed volatile fatty acids to selective drop-in fuels

Abstract

The present invention relates to a two-stage process for production of drop-in fuels/alcohols (methanol, ethanol or butanol) from volatile fatty acids produced either synthetically from fossil resources or as metabolic intermediates in acidification step of anaerobic digestion process from waste biomass and organic materials.

Claims

1. A bio-assisted process for production of drop-in fuels from CO.sub.2 and a volatile fatty acid rich source, said process comprising: (a) providing a first electrochemical system comprising at least one working electrode, at least one counter electrode, and a medium inoculated with selectively enriched electro-active bacteria selected from the group consisting of Geobacter anodireducens, Clostridium ljungdahlii, Acetobacterium woodii, Sporomusa acidovorans, Propionibacterium acidifaciens, Acetobacterium carbinolicum, Pseudomonas stutzeri MTCC 25027, Pseudomonas fragi MTCC 25025, Pseudomonas aeruginosa MTCC 5389, Psuedomonas alcaligenes, Schewanella putrefaciens, Clostridium aciditolerans, Enterobacter aerogenes MTCC 25016, Shewanella sp. MTCC 25020, and combinations thereof; (b) providing feedstock to the first electrochemical system of step (a), wherein the feedstock comprises CO.sub.2 with the volatile fatty acid rich source; (c) intensifying carboxylic acids in the feedstock of step (b) to obtain an effluent comprising intensified carboxylic acids; (d) providing a second electrochemical system comprising at least one working electrode, at least one counter electrode, and a medium inoculated with selectively enriched electro-active bacteria selected from the group consisting of Sporomusa ovate, Clostridium acetobutylicum, Clostridium butyricum, Clostridium acidurici, Pseudomonas aeruginosa MTCC 5388, Pseudomonas putida MTCC 5387, Clostridium carboxidivorans, Clostridium beijerinckii, Schewanella oneidensis, Geobacter sdfurreducens, Clostridium cellutolyticum, Clostridium cellulosi, Clostridium cellulovorans, Alicaligens sp. MTCC 25022, Serratia sp. MTCC 25017, Lysinibacillus sp. MTCC 5666, and combinations thereof; and (e) reducing the intensified carboxylic acids of step (c) in the second electrochemical system to obtain the drop-in fuels; wherein the working electrode of the first electrochemical system is composed of a material selected from the group consisting of graphite plate, carbon brush, carbon paper, graphite felt, activated carbon cloth, and combinations thereof; wherein the counter electrode of the first electrochemical system is modified with a material selected from the group consisting of carbon nanotube (CNT), graphene, charcoal, activated carbon, stainless steel (SS) mesh, nickel oxide, zinc oxide, and iron oxide; wherein the volatile fatty acid rich source comprises at least 0.5% formic acid, provided that if the formic acid is not already present in the feedstock the feedstock is supplemented with at least 0.5% formic acid; wherein the working electrode of the second electrochemical system is composed of a material selected from the group consisting of graphite plate, graphite rod, carbon brush, carbon paper, carbon plate, graphite felt, activated carbon cloth, and combinations thereof; and wherein the counter electrode of the second electrochemical system is modified with a material selected from the group consisting of CNT, Co.sub.9S.sub.8, graphene, FTO/NiO, Ni/Fe layered double hydroxide, Si/TiO.sub.2 nanowires, charcoal, activated carbon, nickel oxide, zinc oxide, and iron oxide; and wherein the drop-in fuels are selected form the group consisting of methanol, ethanol, butanol, acetic acid, propanoic acid, butanoic acid, valeric acid, and caproic acid.

2. The bio-assisted process as claimed in claim 1, wherein the first electrochemical system is operated at a temperature ranging from 20° C. to 50° C.

3. The bio-assisted process as claimed in claim 1, wherein the first electrochemical system is operated at a pH ranging from 6.5 to 9.0.

4. The bio-assisted process as claimed in claim 1, wherein the first electrochemical system is operated at an applied potential ranging from 1V to 4V.

5. The bio-assisted process as claimed in claim 1, wherein the working electrode of the second electrochemical system is modified with a material selected from the group consisting of CNT, graphene, activated charcoal, noble metal, composite of Zr or Ti or Cd or Fe, metal-free alloy, metal carbide, phosphide, and nickel oxide, zinc oxide, iron oxide, metal-phosphorous alloy, metal-sulfur alloy, metal organic framework, and combinations thereof.

6. The bio-assisted process as claimed in claim 1, wherein the carboxylic acid in step (c) has a length ranging from C2 to C6.

7. The bio-assisted process as claimed in claim 1, wherein the second electrochemical system is operated at a temperature ranging from 20° C. to 50° C.

8. The bio-assisted process as claimed in claim 1, wherein the second electrochemical system is operated at a pH ranging from 4.0 to 6.0.

9. The bio-assisted process as claimed in claim 1, wherein the second electrochemical system is operated at an applied potential ranging from 2V to 6V.

10. The bio-assisted process as claimed in claim 1, wherein the volatile fatty acid rich source is selected from the group consisting of kitchen waste, biomass waste, agricultural waste, biodegradable municipal waste, lignocellulosic waste, and de-oiled algal cake.

11. The bio-assisted process as claimed in claim 1, wherein the reduction of carboxylic acids in step (e) is carried out in presence of a redox shuttler selected from the group consisting of neutral red, methylene blue, phenazine derivative, iron based metal complex, nickel based metal complex, zinc based metal complex, anthraquinone-2,6-disulfonate (AQDS), and combinations thereof.

Description

BRIEF DESCRIPTION OF ACCOMPANYING DRAWINGS

(1) FIG. 1 illustrates a schematic representation of drop-in fuel production.

DETAILED DESCRIPTION OF THE INVENTION

(2) The present invention discloses a two-stage process for production of drop-in fuels/alcohols from volatile fatty acids produced either synthetically from fossil resources or as metabolic intermediates in acidification step of anaerobic digestion process from waste biomass and organic materials.

(3) The present invention also discloses a sustainable two stage process wherein any VFA source, having diverse combination of carboxylic acids, can be used as feedstock/starting material. In the first stage, intensification of specific carboxylic acids having chain length between C2 and C6 is carried out in presence of CO.sub.2 in an electro-assisted fermentation process. In stage-2, the intensified carboxylic acids obtained at the end of Stage 1 are reduced to specific alcohols/drop-in fuels through bio-electrochemical approaches using the in situ generated energy/hydrogen.

(4) Both the reactors in stage 1 and 2 are equipped with two electrodes and inoculated with selectively enriched electro-active bacteria. The reactor for stage-1 is single chambered and can be operated in batch or continuous or semi-continuous mode in continuous stirred tank reactor (CSTR) or up-flow anaerobic sludge blanket reactor (UASB) or plug-flow reactor or sequential batch reactor (SBR). The structure of the reactor for stage-1, specifically its single chambered arrangement presents numerous advantages. For instance, in a single chambered reactor, both the electrodes are kept in the same chamber, which eliminates the requirement of an expensive membrane. This structure allows the present reaction to be more cost-efficient over other processes which have been available in the art.

(5) One of the electrodes in the reactors of stage-1 is for oxidation reaction. In an embodiment, the said electrode can be made of graphite plate, carbon brush, carbon paper, graphite felt, and activated carbon cloth (ACC), and combinations thereof. The other electrode is meant for reduction which is a modified electrode with higher redox potentials. The modification can be done using any electrically conductive and bio-compatible support materials, including but not limited to, CNT, graphene, charcoal, activated carbon, metal oxides of nickel or zinc or iron, etc.

(6) The energy required for carrying out the reaction can be supplied from any renewable source like solar or wind or geo-thermal or grid etc

(7) The feedstock for reactor in stage 1 can be CO.sub.2 alone or VFA rich leachate along with CO.sub.2 as C-source. The VFA can be obtained from different sources, viz., kitchen waste, biomass waste, agricultural waste, municipal waste (biodegradable), lignocellulosic waste, de-oiled algal cake, etc., with only one clause that it should contain 0.5% formic acid or else to be amended. The CO.sub.2 can be purified or also can be obtained from off-gases of power plants, industries, etc.

(8) The reactor in stage-1 should be inoculated with selectively enriched bacteria of different groups viz., electro-active bacteria (EAB), chemoautotrophic bacteria, homoacetogenic bacteria and acid producing bacteria (APB), which can work alone or in combination with each other. Bacteria that can be used in the present inventions include but not limited to Geobacter anodireducens, Clostridium ljungdahlii, Acetobacterium woodii, Sporomusa acidovorans, Propionibacterium acidifaciens, Acetobacterium carbinolicum, Pseudomonas stutzeri MTCC 25027 deposited on Apr. 20, 2015 at Microbial Type Culture Collection (MTCC), Institute of Microbial Technology, Shanti Path, 39A, Sector 39, Chandigarh, 160036, India, Pseudomonas fragi MTCC 25025 deposited on Apr. 10, 2015, Pseudomonas aeruginosa MTCC 5389 deposited on Dec. 17, 2007, Schewanella putrefaciens, Clostridium aciditolerans, Enterobacter aerogenes MTCC 25016 deposited on Apr. 9, 2015, Shewanella sp. MTCC 25020 deposited on Apr. 9, 2015.

(9) Other bacteria which can work alone or in combination with each other includes but not limited to: Geobacter anodireducens, G. argillaceus, G. bemidjiensis, G. bremensis, G. chapellei, G. daltonii. G. grbiciae, G. hydrogenophilus, G. lovleyi, G. luticola, G. metallireducens, G. pelophilus, G. pickeringit, G. psychrophilus, G. soh, G. sulirreducens, G. thiogenes, G. toluenoxydans. G. uranireducens, Schewanella abyssi, S. aestuarii, S. algae, S. algidipiscicola, S. amazonensis, S. aquimarina, S. arctica, S. atlantica, S. baltica, S. basaltis, S. benthica, S. Canadensis, S. chilikensis, S. colwelliana, S. corallii, S. decolorationis, S. denitrificans, S. dokdonensis, S. donghaensis, S. fidelis, S. fodinae, S. frigidimarina S. gaetbuli, S. gelidimarina, S. glacialipiscicola, S. hafniensis, S. halfaxensis, S. halitosis, S. hanedai, S. indica, S. irciniae, S. japonica, S. kaireitica, S. htorisediminis, S. livingstonensis, S. loihica. S. mangrove, S. marina, S. marinintestina, S. marisflavi, S. morhuae, S. olleyana, S. oneidensis, S. piezotolerans, S. pacifica, S. pealeana, S. piezotolerans, S. pneumatophori, S. profinda, S. psychrophila, S. putrefaciens, S. sairae, S. schegeliana, S. sediminis, S. seohaensis, S. spongiae, S. surugensis, S. upenei, S. vesiculosa, S. violacea, S. waksmanii, S. woodyi, S. xiamenensis, Pseudomonas aeruginosa, P. alcaligenes, P. anguilliseptica, P. argentntensis, P. borbori, P. citronellolis, P. flavescens, P. mendocina, P. nitroreducens, P. oleovorans, P. pseudoalcaligenes, P. resinovorans, P. straminea, P. balearica, P. luteola, P. stutzeri, Sporomusa ovate, Clostridium ljungdahlii, Acetobacterium bakii, Acetobacterium carbinolicum, Acetobacterium fimetarium, Acetobacterium malicum, Acetobacterium paludpsum, Acetobacterium tundra, Acetobacterium wieringae, Acetobacterium woodii, Sporomusa acidovorans, cyanobacterium Synechocystis, Pelotomaculum thermopropionicum, Megasphaera hexanoica, Megasphaera hominis, Megasphaera cerevisiae, Megasphaera elsdenii, Megasphaera micronuciformis, Megasphaera paucivorans, Megasphaera sueciensis, M. cerevisiae, Anaeroglobus geminates, Bacillus subtilis, Propionibacterium acidifaciens, Propionibacterium acidipropionici, Propionibacterium acnes Propionibacterium australense, Propionibacterium avidum, Propionibacterium cyclohexanicum, Propionibacterium damnosum, Propionibacterium freudenreichii, Propionibacterium granulosum, Propionibacterium jensenii, Propionibacterium lymphophilum, Propionibacterium microaerophilu, Propionibacterium namnetense, Propionibacterium olivae, Propionibacterium propionicus, Propionibacterium thoenii, etc.

(10) In an embodiment, the reactor in stage-1 can be operated at temperatures ranging from 20-50° C. In another embodiment, the reactor in stage-1 can be operated at pH ranging between 6.5 to 9.0.

(11) In yet another embodiment, the reactor in stage-1 can be operated under the applied potential in the range of 1-4 V or applied current anywhere in the range of 30-150 A/m.sup.2 vs Ag/AgCl reference electrode. In accordance with the present invention, reactor in stage-1 can be operated in batch mode with a hydraulic retention time (HRT) of 24-72 h or in continuous mode with 18-48 h HRT.

(12) Subsequently, the effluent from reactor in stage-1 enriched with specific type of VFAs is supplied to the reactor in stage-2, where it is converted to alcohols. The chain length of the alcohol depends on the availability of H.sub.2/H.sup.+ in the reactor. In presence of the in situ generated H.sub.2/H.sup.+, the chain length of dominant alcohol of the total product remains same as the VFA obtained in stage-1 but it shortens by 1 carbon when the source of H.sub.2/H.sup.− is absent in the reactor.

(13) The reactor in stage-2 can be single chambered or double chambered operating in batch or continuous or semi-continuous mode in continuous stirred tank reactor (CSTR) or up-flow anaerobic sludge blanket reactor (UASB) or plug-flow reactor or sequential batch reactor (SBR). In double chambered configuration, the membrane used to separate the 2 chambers can be proton exchange membrane (PEM) or cation exchange membrane (CEM) or porcelain disk with micro pores.

(14) Similar to stage-1, one of the electrodes in reactor of stage-2 also meant for oxidation. It should be modified for getting higher oxidation potentials in case of generating H.sub.2/H.sup.+ in situ in the reactor itself or else, the modification is not required. The electrode can be made of different materials viz., graphite plate, carbon brush, carbon paper, graphite felt, activated carbon cloth (ACC), etc., and the modification can be done using metal oxides of zirconium, cadmium nickel, zinc, iron, cobalt, titanium, silicone and their alloy, their phosphorous and sulfur composites etc. The other electrode is meant for reduction which is a carbon graphite electrode modified for obtaining higher redox potentials. The modification can be done using highly conductive materials like CNT, graphene, charcoal, activated carbon, etc.

(15) The energy required for carrying out the reaction can be supplied from any renewable source like solar or wind or geo-thermal or grid etc. The effluent from reactor in stage-1 acts as feedstock for the reactor in stage 2.

(16) The reactor in stage-2 also should be inoculated with selectively enriched biocatalyst and have the combination of different bacterial groups, viz., electro-active bacteria (EAB), sulfate-reducing bacteria, homoacetogenic bacteria and metal reducing bacteria, which can work in synergistic interaction with each other. Bacteria that can be used in the present inventions include but not limited to Sporomusa ovate, Clostridium acetobutylicum, Clostridium butyricum, Clostridium acidurici, Pseudomonas aeruginosa MTCC 5388 deposited on Dec. 17, 2007, Pseudomonas putida MTCC 5387 deposited on Dec. 17, 2007, Clostridium carboxidivorans, Clostridium beijerinckii, Schewanella oneidensis, Geobacter sulfurreducens, Clostridium cellulolyticum, Clostridium cellulosi, Clostridium cellulovorans, Alicaligens sp. MTCC 25022 deposited on Apr. 10, 2015, Serratia sp. MTCC 25017 deposited on Apr. 9, 2015, Lysinibacillus sp. MTCC 5666 deposited on Oct. 12, 2011, and combinations thereof.

(17) Other bacteria which can work alone or in combination with each other includes but not limited to: Geobacter anodireducens, G. argillaceus, G. bemidjiensis, G. bremensis, G. chapellei. G. daltonii, G. grbiciae, G. suifirreducens, G. thiogenes, G. toluenoxydans, G. uraniireducens, Schewanella abyssi, S. aestuarii, S. algae. S. algidipiscicola, S. amazonensis, S. aquimarina, S. arclica, S. litorisediminis, S livingslonensis, S. loihica, S mangrove, S. marina, S. marinintestina, S. marisfavi, S. morhuae, S. olleyana, S. oneidensis. S. piezotolerans, S. pacifica, S. pealeana, S. piezotolerans, S. pneumatophori, S. profunda, S. psychrophila, S. putrefaciens, S. sairae, S. schegeliana, S. sediminis, S. seohaensis, S. spongiae, S. surugensis, S. upenei, S. vesiculosa, S. violacea, S. waksmanii, S. woodyi, S. xiamenensis, Pseudomonas aeruginosa, P. alcaligenes, P. anguilliseptica, P. argentinensis, P. borbori, P. citronellolis, P. flavescens, P. mendocina, P. nitroreducens, P. oleovorans, P. pseudoalcaligenes, P. resinovorans, P. straminea, P. balearica, P. luteola, P. stutzeri, Sporomusa ovate, Clostridium ljungdahlii, Sporomusa acidovorans, cyanobacterium Synechocystis, Pelotomaculum thermopropionicum, Megasphaera hexanoica, Megasphaera hominis, Megasphaera cerevisiae, Megasphaera elsdeni, Megasphaera micronuciformis Megasphaera paucivorans, Megasphaera sueciensis, M. cerevisiae, Anaeroglobus geminates, Clostridium acetobutylicum, Clostridium butyricum, Clostridium diolis, Clostridium beijerinckii, Clostridium acidisoli, Clostridium aciditolerais, Clostridium acidurici, Clostridium aerotolerans, Clostridium cadaveris, Clostridium caenicola, Clostridium caninithermale, Clostridium carboxidivorans, Clostridium carnis, Clostridium cavenidishii, Clostridium celatum, Clostridium celerecrescens, Clostridium cellobioparum, Clostridium cellulofermentans, Clostridium cellulolyticun, Clostridium celhdlosi, Clostridium cellulovorans, D. acrylicus, D. aerotolerans, D. aespoeensis, D. africanus, D. alaskensis, D. alcoholivorans, D. alkalitolerans, D. aminophilus, D. arcticus, D. baarsii, D. baculatus, D. baslinii, D. biadhensis, D. bizertensis, D. burkinensis, D. butratiphilus, D. capillatus, D. carbinolicus, D. carbinoliphilus, D. cuneatus, D. dechloracetivorans, D. desulfuricans, D. ferrireducens, D. frigidus, D. fructosivorans, D. furfurals, D. gabonensis, D. giganteus, D. gigas, D. gracilis, D. halophilus, D. hydrothermalis, D. idahonensis, D. indonesiensis, D. inopinatus, D. intestinalis, D. legalli, D. alitoralis, D. longreachensis, D. longus, D. magneticus, D. marinus, D. marinisediminis, D. marrakechensis, D. mexicamis, D. multispirans, D. oceani, D. oxamicus, D. oxyclinae, D. paquesii, D. piezophilus, D. pigra, D. portus, D. profindus, D. psychrotolerats, D. putealis, D. salixigens, D. sapovorans, D. senezii, D. simplex, D. sulfodismutans, D. termitidis, D. thermophilus, D. tunisiensis, D. vietnamensis, D. vulgaris, D. zosterae, Klebsiella granulomatis, Klebsiella oxytoca, Klebsiella pneumonia, Klebsiella terrigena, Klebsiella varitcola, Bacillus subtilis, Zvmomonas mobilis, Zymomonas anaerobia etc.

(18) An embodiment of the present invention provides a bio-assisted process for production of drop-in fuels from CO.sub.2 and volatile fatty acid rich source, said process comprising: (a) providing a first electrochemical system comprising at least one working electrode, at least one counter electrode, and a medium inoculated with selectively enriched electro-active bacteria selected from the group consisting of Geobacter anodireducens, Clostridium ljungdahlii, Acetobacterium woodii, Sporomusa acidovorans, Propionibacterium acidifaciens, Acetobacterium carbinolicum, Pseudomonas stutzeri (MTCC 25027), Pseudomonas fragi (MTCC 25025), Pseudomonas aeruginosa (MTCC 5389), Psuedomonas alcaligenes, Schewanella putrefaciens, Clostridium aciditolerans, Enterobacter aerogenes (MTCC 25016), Shewanella sp. (MTCC 25020), and combinations thereof; (b) providing feedstock to the first electrochemical system of step (a), wherein the feedstock comprises CO.sub.2 with or without the volatile fatty acid rich source; (c) intensifying carboxylic acids in the feedstock of step (b) to obtain an effluent comprising intensified carboxylic acids; (d) providing a second electrochemical system comprising at least one working electrode, at least one counter electrode, and a medium inoculated with selectively enriched electro-active bacteria selected from the group consisting of Sporomusa ovate, Clostridium acetobulicum, Clostridium butyricum, Clostridium acidurici, Pseudomonas aeruginosa (MTCC 5388), Pseudomonas putida (MTCC 5387), Clostridium carboxidivorans, Clostridium beijerinckii, Schewanella oneidensis, Geobacter sulfurreducens, Clostridium cellulolyticum, Clostridium cellulosi, Clostridium cellulovorans, Alicaligens sp. (MTCC 25022), Serratia sp. (MTCC 25017), Lysinibacillus sp. (MTCC 5666), and combinations thereof; and (e) reducing the intensified carboxylic acids of step (c) in the second electrochemical system to obtain the drop-in fuels; wherein the working electrode of the first electrochemical system is composed of a material selected from the group consisting of graphite plate, carbon brush, carbon paper, graphite felt, activated carbon cloth, and combinations thereof; wherein the counter electrode of the first electrochemical system is modified with a material selected from the group consisting of CNT, graphene, charcoal, activated carbon, SS mesh, nickel oxide, zinc oxide, and iron oxide; wherein the volatile fatty acid rich source comprises at least 0.5% formic acid, provided that if the formic acid is not already present in the feedstock the feedstock is supplemented with at least 0.5% formic acid; wherein the working electrode of the second electrochemical system is composed of a material selected from the group consisting of graphite plate, graphite rod, carbon brush, carbon paper, carbon plate, graphite felt, activated carbon cloth, and combinations thereof; and wherein the counter electrode of the second electrochemical system is modified with a material selected from the group consisting of CNT, Co.sub.9S.sub.8, graphene, FTO/NiO, Ni/Fe layered double hydroxide, Si/TiO.sub.2 nanowires, charcoal, activated carbon, nickel oxide, zinc oxide, and iron oxide.

(19) In another embodiment of the present invention, the first electrochemical system is operated at a temperature ranging from 20° C. to 50° C.

(20) In yet another embodiment of the present invention, the first electrochemical system is operated at a pH ranging from 6.5 to 9.0.

(21) In still another embodiment of the present invention, the first electrochemical system is operated at an applied potential ranging from 1 V to 4 V.

(22) In a further embodiment of the present invention, the working electrode of the second electrochemical system is modified with a material selected from the group consisting of CNT, graphene, activated charcoal, noble metal, composite of Zr or Ti or Cd or Fe, metal-free alloy, metal carbide, phosphide, and nickel oxide, zinc oxide, iron oxide, metal-phosphorous alloy, metal-sulfur alloy, metal organic framework, and combinations thereof.

(23) Yet another embodiment of the present invention provides that the carboxylic acid in step (c) has a length ranging from C2 to C6.

(24) Still another embodiment of the present invention provides that the second electrochemical system is operated at a temperature ranging from 20° C. to 50° C.

(25) A further embodiment of the present invention provides that the second electrochemical system is operated at a pH ranging from 4.0 to 6.0.

(26) Another embodiment of the present invention provides that the second electrochemical system is operated at an applied potential ranging from 2 V to 6 V.

(27) In another embodiment of the present invention, there is provided that the volatile fatty acid rich source is selected from the group consisting of kitchen waste, biomass waste, agricultural waste, biodegradable municipal waste, lignocellulosic waste, and de-oiled algal cake.

(28) In yet another embodiment of the present invention, there is provided that the reduction of carboxylic acids in step (e) is carried out in presence of a redox shuttler selected from the group consisting of neutral red, methylene blue, phenazine derivative, iron based metal complex, nickel based metal complex, zinc based metal complex, AQDS, and combinations thereof.

(29) In a further embodiment of the present invention, the drop-in fuels are selected from the group consisting of methanol, ethanol, butanol, formic acid, acetic acid, propanoic acid, butanoic acid, valeric acid, and caproic acid.

(30) During stage-2, redox shuttlers such as, neutral red (NR), methylene blue (MB), phenazine derivatives, iron/nickel/zinc based metal complexes, etc., can be added individually or in combinations to enhance the electron exchange between the electrode and biocatalyst. In an embodiment, the reactor in stage-2 can be operated at temperatures ranging from 20-50° C. In an embodiment, the reactor in stage-2 can be operated at pH ranging between 4.0 and 6.0. The reactor in stage-1 can be operated under applied potential in the range of 2-6 V or applied current anywhere in the range of 100-250 A/m.sup.2 vs Ag/AgCl reference electrode.

(31) The reactor in stage-2 can be operated in batch mode or in continuous mode with a hydraulic retention time (HRT) of 24-72.

(32) Having described the basic aspects of the present invention, the following non-limiting examples illustrate specific embodiment thereof.

EXAMPLES

Example 1: General Example

(33) Experiments were carried out in 2 stages, where the intensification of specific VFA was done during stage-1 followed by the VFA reduction to alcohol in stage-2. The existing fermentors were modified by inserting a pair of electrodes (total/working volume, 2.8/2.0 L) and used for this study in both the stage-1 and stage-2. For stage-1 operation, the bioreactor was inserted with graphite rod wrapped with ACC as electrode for oxidation and carbon powder coated graphite rod wrapped with SS mesh as electrode for reduction. Both the electrodes were connected to potentiostat through SS wires and an applied cell potential of +2V was applied to the circuit against Ag/AgCl (3M KCL) reference electrode during stage-1 operation. The current consumption during operation was monitored using chronoamperometry (CA). CO.sub.2 alone or along with VFA rich organic waste leachate was used as substrate for stage-1 operation in batch and continuous modes with a retention time of 72 h and 24 h respectively, at pH 6.5±0.1. Trace metal solution was also added to the reactor, when CO.sub.2 alone was used as substrate to support the microbial activity. Constant mixing was provided to the bioreactor at a rate of 500 rpm experiments were carried out at ambient conditions. Samples were collected at regular time intervals and analyzed for VFAs using gas chromatography at regular time intervals.

(34) During stage-2 operation the effluent from stage-1, enriched with specific VFA was used as substrate and converted those VFAs into alcohols. For this, two approaches were followed, where in approach-1 modified electrode for oxidation was used to generate H.sub.2/H.sup.+ in situ in the reactor and in approach-2, non-modified electrode for oxidation was used. In approach-1, the bioreactor was inserted with graphite rod modified with ZrTi—P as electrode for oxidation and carbon felt and activated carbon filled in cylinder made of SS mesh as electrode for reduction. In approach-2, the electrode for oxidation is replaced with graphite rod wrapped with ACC, keeping the rest as same to approach-1. In both the cases, the electrodes were connected to potentiostat through SS wire and +3V of cell potential was applied to the circuit against Ag/AgCl (3M KCL) reference electrode. NR (0.0001%) was added as redox shuttler in stage-2 bioreactor to facilitate electron exchange between the biocatalyst and electrode. The current consumption during operation was monitored using chronoamperometry (CA). The effluent from stage-1 was used as feedstock for both the approaches of stage-2, after adjusting the pH to 5.0±0.1 and the same was maintained throughout operation. The pH was maintained using a combination of Acid (H.sub.3PO.sub.4)/Base (NaOH). Mixing was provided at a rate of 500 rpm and experiments were carried out at ambient conditions with HRT of 72 h and 24 h during batch and continuous mode operations respectively. Samples were collected at regular time intervals and analyzed for alcohols production using gas chromatography at regular time intervals.

Example 2: Using C02 as Sole C-Source

(35) In one of the examples, the experiment was carried out using CO.sub.2 alone as C-source at a rate of 15 ml/min for 72h of HRT and formic acid (0.5%) as proton source in stage-1. For the reaction, basic media containing NH.sub.4Cl of 500 mg/L, MgCl.sub.2.6H.sub.2O of 160 mg/L, Peptone 250 mg/L Yeast Extract of 26 mg/L along with the trace elements solution (per litre, Nitrilotriacetic acid, 1.5 g; MgSO.sub.4×7H.sub.2O, 2.0 g; MnSO.sub.4×H.sub.2O, 1.5 g; NaCl, 1.0 g; FeSO.sub.4×7H.sub.2O, 0.1 g; CoSO.sub.4×7H.sub.2O, 0.134 g; CaCl.sub.2)×2H.sub.2O, 0.1 g; ZnSO.sub.4×7H.sub.2O, 0.18 g; CuSO.sub.4×5H.sub.2O, 0.01 g; KAl(SO.sub.4).sub.2×12H.sub.2O, 0.02 g; H.sub.3BO.sub.3, 0.01 g; Na.sub.2MoO.sub.4×2H.sub.2O, 0.01 g; NiCl.sub.2×6H.sub.2O, 0.03 g; Na.sub.2SeO.sub.3×5H.sub.2O, 0.50 mg) and vitamin solution (per litre, biotin, 2 mg; pantothenic acid, 5 mg; B-12, 0.1 mg; p-aminobenzoic acid, 0.25 mg; thioctic acid (alpha lipoic), 5 mg; nicotinic acid, 5 mg; thiamine, 5 mg, riboflavin, 5 mg; pyridoxine HCl, 10 mg, folic acid, 2 mg) was used.

(36) Rest of the operating conditions was maintained as given in experimental conditions. The combination of Enterobacter aerogenes MTCC 25016, Shewanella sp. MTCC 25020, Pseudomonas stutzferi MTCC 25027, Pseudomonas aeruginosa MTCC 5389 was used as biocatalyst during stage-1. The current consumption on CA during stage-1 started increasing with time and in 6h of operation, it reached 4.1±1.6 A/m2 and sustained afterwards at similar value till 72 h of operation. Results showed enrichment of acetic acid (9.2 g/l) from CO.sub.2 along with formic acid (4.2 g/l) during stage-1 (Table 1).

(37) The effluent from stage-1 operation enriched with acetic acid was used as substrate for stage-2 and no other C-source was provided. The combination of Clostridium butyricum, Serratia sp. MTCC 25017, Pseudomonas aeruginosa MTCC 5388, Pseudomonas putida MTCC 5387, was used as biocatalyst during stage-2. During approaches of stage-2, the current consumption started immediately but reached to a maximum of 8.2±2.1 A/m2 in approach-1, where the in situ H2/H+ generation takes place on the modified electrode, while the current consumption restricted to 2.3±0.6 A/m.sup.2 only in approach-2. Transformation of acetic acid to alcohol was observed during both the approaches of stage-2 but approach-1 resulted in the dominant production of ethanol (6.7 g/l), while the approach-2 produced methanol (5.72). The in situ generated H.sub.2/H.sup.+ along with the formic acid supported the reduction of acetic acid to ethanol without reduction in carbon chain, while in the approach-2, formic acid alone along with the electrode potential supported the transformation which resulted in loss of carbon producing methanol. The CCE for stage-1 is about 26% due to issues in CO.sub.2 solubility. During stage-2, about 92% CCE was recorded during approach-1 but the CCE was about 54% only during approach-2 due to the non-availability of H.sub.2/H.sup.+ in the reactor resulting in the loss of carbon as CO.sub.2.

(38) TABLE-US-00001 TABLE 1 Two stage operation of bioreactor in batch mode using CO.sub.2 as C-source Stage-2/ Stage-2/ Effluent approach-1, approach-2, Influent VFA after Product Product VFA stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 0 0 0.81 5.72 Ethanol 0 0 6.7 0.22 Butanol 0 0 0 0 Formic acid 5 4.2 0.021 0.02 Acetic acid 0 9.2 0.87 0.87 Propanoic acid 0 0 0 0.00 Butanoic acid 0 0 0.49 0.00 Total 0 13.4 8.89 6.83

Example 3: Leachate from Kitchen Waste Along with C02 as C-Source

(39) In another example, the experiment was carried out using leachate from kitchen waste (VFA: 28.24 g/l; TOC: 34.61 g/l) along with CO.sub.2 as C-source (15 ml/min) and formic acid was maintained at 0.5% by external addition of required quantity. The combination of Clostridium ljungdahlii, Enterobacter aerogenes MTCC 25016, Pseudomonas fragi MTCC 25025, Pseudomonas stutzferi MTCC 25027 was used as biocatalyst during stage-1. Here also, the current consumption during stage-1 was observed to be increasing with time and in 3 h of operation, it reached 6.1±0.8 A/m.sup.2 and sustained afterwards at similar range till 72 h of operation. Results showed the intensification of acetic acid (13.78 g/l) and butyric acids (11.96 g/l) during stage-1 at the expense of other constituents of leachate and CO.sub.2 (Table 2).

(40) The effluent from stage-1 operation was made as substrate for stage-2 of both the approaches and no other C-source was provided for stage-2 operation. Rest of the operating conditions remained same. The combination of Pseudomonas putida MTCC 5387, Clostridium acetobutylicum, Clostridium beijerinckii, Serratia sp. MTCC 25017 was used as biocatalyst during stage-2. Similar to the example-1, the current consumption on CA started immediately after start-up during both the approaches of stage-2 and showed a maximum value of 10.2±0.55 A/m.sup.2 and 1.6±0.85 A/m.sup.2 respectively for approach-1 and approach-2. Alcohol production was observed during both the approaches of stage-2 but approach-1 resulted in the dominant production of butanol (11.08 g/l) and ethanol (10.68 g/l), while the approach-2 produced methanol (18.7 g/l). In situ generation of H2/H+ is the major reason behind the variation in alcohol composition. The CCE for stage-1 is about 48%. During stage-2, about 93% CCE was recorded during approach-1 but the CCE was only 50% during approach-2 due to the non-availability of H.sub.2/H.sup.+ in the reactor resulting in the loss of carbon as CO.sub.2.

(41) TABLE-US-00002 TABLE 2 Two stage operation of bioreactor in batch mode using kitchen waste leachate along with CO.sub.2 as C-source Stage-2/ Stage-2/ Effluent approach-1, approach-2, Influent VFA after Product Product VFA stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 1.4 0 0.896 18.7 Ethanol 0.18 0 10.68 0.419 Butanol 0 0 11.08 0 Formic acid 2.23* 3.1 0 0 Acetic acid 8.36 13.78 2.16 1.89 Propanoic acid 3.21 0 0 0 Butanoic acid 7.09 11.96 1.12 0.068 Valeric acid 3.41 2.67 0.316 0 Caproic acid 3.94 2.34 0.06 0 Total 29.82 33.85 26.312 21.077 *Required amount of formic acid was amended to make-up 0.5%

(42) Further to that, the experiment was carried out in continuous mode operation of the both the reactors with a HRT of 24 h at a flow rate of 84 ml/h. Current consumption in this case increased to 7.4±1.08 A/m.sup.2 during stage-1. The cumulative product after 3 days showed the intensification of acetic acid (42.6 g/l) and butyric acid (36.7 g/l) at the expense of other constituents of leachate either completely or partially. Compared to batch mode, the CCE for stage-1 is increased and is about 78%. Subsequently, when the effluent of stage-1 subjected to stage-2, the current consumption recorded as 12.6±0.9 A/m.sup.2 and 1.63±0.7 A/m.sup.2 during approach-1 and 2 respectively. Similar to the batch experiments, the intensified acetic and butyric acids were reduced alcohols during stage-2 (Table 3). Butanol (46.93 g/l) production was higher compared to ethanol (21.12 g/l) in continuous mode operation of approach-1, while the methanol was dominant in approach-2 (71.62 g/l). The CCE during stage-2 operation remained more or less similar for approach-1 (91%), while the CCE increased to 61% in approach-2 compared to batch mode.

(43) TABLE-US-00003 TABLE 3 Two stage operation of bioreactor in continuous mode using kitchen waste leachate along with CO.sub.2 as C-source (Cumulative of 3 days) Stage-2/ Stage-2/ Effluent approach-1, approach-2, Influent VFA after Product Product VFA stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 4.2 0 2.41 71.62 Ethanol 0.54 0 21.12 4.12 Butanol 0 0 46.93 0 Formic acid 6.69* 9.83 0 0 Acetic acid 25.08 42.6 2.87 2.87 Propanoic acid 9.63 0 0 0 Butanoic acid 21.27 36.7 2.17 0.981 Valeric acid 10.23 8.74 0.86 0 Caproic acid 11.82 7.62 0.18 0 Total 89.46 105.49 76.54 79.57 *Required amount of formic acid was amended to make-up 0.5%

Example 4: Leachate from Biomass Waste Along with CO.SUB.2 .as C-Source

(44) In another example, the source of leachate was changed from kitchen waste to biomass waste (VFA: 30.16 g/l; TOC: 41.27 g/l), keeping the bioreactor design and operation same. CO.sub.2 was also added at a rate of 15 ml/min and formic acid was maintained at 0.5% by external addition of required quantity. The combination of Shewanella sp. MTCC 25020. Pseudomonas aeruginosa MTCC 5389, Geobacter anodireducens, Pseudomonas stutzeri MTCC 25027, was used as biocatalyst during stage-1. Current consumption during stage-1 was observed to be increasing with time and in 5 h of operation, it reached 7.65±0.7 A/m.sup.2 and sustained afterwards at similar range till 72 h of operation. Similar to the kitchen waste leachate, results showed the intensification of acetic (14.2 g/l) and butyric acids (12.9 g/l) during stage-1 at the expense of other constituents of leachate and CO.sub.2 with a CCE of 51% (Table 4).

(45) The effluent from stage-1 operation was used as substrate for both the approaches of stage-2 and no other C-source was provided. Rest of the operating conditions remained same. The combination of Alicaligens sp. MTCC 25022, Serratia sp. MTCC 25017, Lysinibacilhus sp. MTCC 5666, Clostridium cellulolyticum was used as biocatalyst during stage-2. Here also, the current consumption on CA started immediately after start-up during both the approaches of stage-2 and showed a maximum value of 9.68±0.4 A/m.sup.2 and 1.78±0.6 A/m.sup.2 respectively. Alcohol production was evident similar to the kitchen waste leachate experiment during both the approaches of stage-2. Approach-1 resulted in the dominant production of butanol (12.19 g/l) and ethanol (10.64), while the approach-2 produced methanol (21.13). During stage-2, about 92% CCE was recorded during approach-1 but the CCE was only 64% during approach-2 due to the non-availability of H.sub.2/H.sup.+ in the reactor resulting in the loss of carbon as CO.sub.2.

(46) TABLE-US-00004 TABLE 4 Two stage operation of bioreactor in batch mode using kitchen waste leachate along with CO.sub.2 as C-source Stage-2/ Stage-2/ Effluent approach-1, approach-2, Influent VFA after Product Product VFA stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 0.98 0 0.784 21.13 Ethanol 0.23 0 10.64 3.68 Butanol 0 0 12.19 0 Formic acid 1.68* 3.78 0 0 Acetic acid 10.18 14.2 1.96 1.13 Propanoic acid 6.24 0 0 0 Butanoic acid 5.61 12.9 1.16 0.73 Valeric acid 3.68 2.61 0.398 0 Caproic acid 2.77 2.24 0.09 0 Total 31.37 35.73 27.22 26.64 *Required amount of formic acid was amended to make-up 0.5%

(47) Further to that, the experiment was carried out in continuous mode operation of the both the reactors with a HRT of 24 h at a flow rate of 84 ml/h. Current consumption in this case also increased to 8.3±0.96 A/m.sup.2 during stage-1. The cumulative product after 3 days showed the intensification of acetic acid (4.28 g/l) and butyric acid (36.68 g/l) at the expense of other constituents of leachate either completely or partially, resulting in CCE of 72%.

(48) Subsequently, the effluent of stage-1 subjected to stage-2 and about 11.3±1.09 A/m2 and 1.82±0.6 A/m.sup.2 of current consumptions were recorded during approach-1 and 2 respectively. Similar to the batch experiments, the intensified acetic and butyric acids were reduced alcohols during stage-2 (Table 5). Transformation to butanol (45.17 g/l) is higher compared to ethanol (19.86 g/l) during continuous mode operation of approach-1, while the methanol was dominant in approach-2 (69.82 g/l). The CCE during stage-2 operation remained more or less similar for approach-1 (92%), while the CCE increased to 63% in approach-2 compared to batch mode.

(49) TABLE-US-00005 TABLE 5 Two stage operation of bioreactor in continuous mode using biomass waste leachate along with CO.sub.2 as C-source (Cumulative of 3 days) Stage-2/ Stage-2/ Effluent approach-1, approach-2, Influent VTA after Product Product VFA stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 2.94 0 1.96 69.82 Ethanol 0.69 0 19.86 3.76 Butanol 0 0 45.17 0 Formic acid 5.04* 9.12 0 0.076 Acetic acid 30.54 41.28 1.94 1.94 Propanoic acid 18.72 0 0 0 Butanoic acid 16.83 36.68 2.43 0.873 Valeric acid 11.04 7.36 1.66 0 Caproic acid 8.31 5.98 0.24 0 Total 94.11 100.34 73.26 76.45 *Required amount of formic acid was amended to make-up 0.5%

Example 5: Leachate from Agricultural Waste Along with CO.SUB.2 .as C-Source

(50) In another example, the source of leachate was changed from kitchen waste to agriculture residue (horticulture waste) (VFA: 28.12 g/l; TOC: 38.36 g/l), keeping the bioreactor design and operation same. CO.sub.2 was also added at a rate of 15 ml/min and formic acid was maintained at 0.5% by external addition of required quantity. The combination of Pseudomonas stutzeri MTCC 25027, Pseudomonas aeruginosa MTCC 5389, Shewanella sp. MTCC 25020, Psuedomonas alcaligenes was used as biocatalyst during stage-1. Current consumption during stage-1 was observed to be increasing with time and in 5 h of operation, it reached 8.43±0.6 A/m2 and sustained afterwards at similar range till 72 h of operation. Similar to the kitchen waste leachate, results showed the intensification of acetic (13.68 g/l) and butyric acids (11.42 g/l) during stage-1 at the expense of other constituents of leachate and CO.sub.2 with a CCE of 49% (Table 6).

(51) The effluent from stage-1 operation was used as substrate for both the approaches of stage-2 and no other C-source was provided. Rest of the operating conditions remained same. The combination of Pseudomonas putida MTCC 5387, Pseudomonas aeruginosa strain MTCC 5388, Geobacter sufirreducens, Serratia sp. MTCC 25017, Clostridium celulovorans, was used as biocatalyst during stage-2. Here also, the current consumption on CA started immediately after start-up during both the approaches of stage-2 and showed a maximum value of 10.42±0.38 A/m2 and 2.36±0.6 A/m.sup.2 respectively. Alcohol production was evident similar to the agricultural waste leachate experiment during both the approaches of stage-2. Approach-1 resulted in the dominant production of butanol (10.31 g/l) and ethanol (9.42), while the approach-2 produced methanol (17.18). During stage-2, about 94% CCE was recorded during approach-1 but the CCE was only 58% during approach-2 due to the non-availability of H.sub.2/H.sup.+ in the reactor resulting in the loss of carbon as CO.sub.2.

(52) TABLE-US-00006 TABLE 6 Two stage operation of bioreactor in batch mode using agriculture waste leachate along with CO.sub.2 as C-source Stage-2/ Stage-2/ Effluent approach-1, approach-2, Influent VFA after Product Product VFA stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 0.63 0 0.662 17.18 Ethanol 0.41 0 9.42 2.82 Butanol 0 0 10.31 0 Formic acid 1.34* 3.78 0 0 Acetic acid 8.87 13.68 2.84 4.18 Propanoic acid 4.26 0 0 0 Butanoic acid 5.18 11.42 1.67 1.07 Valeric acid 2.64 1.63 0.43 0 Caproic acid 3.12 0 0 0 Total 26.45 30.51 25.332 25.25 *Required amount of formic acid was amended to make-up 0.5%

(53) Further to that, the experiment was carried out in continuous mode operation of the both the reactors with a HRT of 24 h at a flow rate of 84 ml/h. Current consumption in this case also increased to 7.16±0.6 A/m.sup.2 during stage-1. The cumulative product after 3 days showed the intensification of acetic acid (39.36 g/l) and butyric acid (31.42 g/l) at the expense of other constituents of leachate either completely or partially, resulting in CCE of 68%.

(54) Subsequently, the effluent of stage-1 subjected to stage-2 and about 12.11±0.93 A/m2 and 1.78±0.34 A/m.sup.2 of current consumptions were recorded during approach-1 and 2 respectively. Similar to the batch experiments, the intensified acetic and butyric acids were reduced alcohols during stage-2 (Table 7). Transformation to butanol (38.18 g/l) is higher compared to ethanol (29.32 g/l) during continuous mode operation of approach-1, while the methanol was dominant in approach-2 (73.16 g/l). The CCE during stage-2 operation remained more or less similar for approach-1 (94%), while the CCE increased to 64% in approach-2 compared to batch mode.

(55) TABLE-US-00007 TABLE 7 Two stage operation of bioreactor in continuous mode using agriculture waste leachate along with CO.sub.2 as C-source (Cumulative of 3 days) Stage-2/ Stage-2/ Effluent approach-1, approach-2, Influent VFA after Product Product VFA stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 0.63 0 2.04 73.16 Ethanol 0.41 0 29.32 3.76 Butanol 0 0 38.18 0 Formic acid 1.34* 10.18 0 0 Acetic acid 8.87 39.36 1.08 2.96 Propanoic acid 4.26 0 0 0 Butanoic acid 5.18 31.42 1.26 0 Valeric acid 2.64 6.66 0.98 0 Caproic acid 3.12 6.04 0 0 Total 26.45 93.66 72.86 79.88 *Required amount of formic acid was amended to make-up 0.5%

Example 6-10: Different Electrode Materials, Microbes and Power Supply Mode Combinations Keeping C-Source Constant i.e., Leachate from Kitchen Waste Along with CO.SUB.2

(56) In examples 5 to 9, all the experiments were carried out using leachate from kitchen waste (VFA: 28.24 g/l; TOC: 34.61 g/l) along with CO.sub.2 as C-source (15 ml/min) and formic acid was maintained at 0.5% by external addition of required quantity. However, the working and counter electrodes for both stage 1 and stage 2 were varied as well as the microbial blend used and the power supply mode (potentiostat or galvanostat) was changed (Table 8). All these experiments also showed current consumption pattern in similar lines. However, the product profile varied slightly with the microbe nature. The example 8 and Example 9, where Propinibacterium Sp. was used, propionc acid production was also dominantly observed along with acetic acid which is not the case with other experiments. (Table 9-13).

(57) The effluent from stage-1 operation was made as substrate for stage-2 of both the approaches and no other C-source was provided for stage-2 operation. Rest of the operating conditions was varied between each experimental set-up as described in Table 8. All the experimental combinations (Example 6-10) showed very good current consumption for approach-1 and lower current consumption for approach-2, similar to the example-1 indicating the role of presence of H2/H+. Alcohol production was observed during both the approaches of stage-2 but approach-1 resulted in the dominant production of butanol and ethanol, while the approach-2 produced methanol. In situ generation of H2/H+ is the major reason behind the variation in alcohol composition. The average CCE lies in the range of 44-52% during stage-1, while it is more than 90% during approach-1 of stage-2 irrespective of the experimental combinations. The CCE was only 50% during approach-2 of stage-2 due to the non-availability of HJH in the reactor resulting in the loss of carbon as CO.sub.2.

(58) TABLE-US-00008 TABLE 8 Different experimental combinations studied using kitchen waste leachate along with CO2 as C-source Example 5 Example 6 Example 7 Example 8 Example 9 Stage-1 Working Carbon brush ACC Graphite Felt Graphite Felt Carbon paper electrode Counter Graphite rod Graphite rod Graphite rod Graphite rod Graphite rod electrode modified with modified with modified with modified with modified with CNT Graphene Charcoal CNT NiO Microbes Enterobacter Clostridium Enterobacter Enterobacter Shewanella sp. aerogenes MTCC ljungdahlii, aerogenes MTCC aerogenes MTCC MTCC 25020, 25016, Pseudomonas 25016, 25016, Pseudomonas Pseudomonas stutzeri MTCC Pseudomonas Pseudomonas fragi MTCC fragi MTCC 25027, aeruginosa MTCC aeruginosa MTCC 25025, 25025, Shewanella sp. 5389, Shewanella 5389, Shewanella Pseudomonas Pseudomonas MTCC 25020 sp. MTCC 25020 sp. MTCC 25020 stutzeri MTCC aeruginosa MTCC 25027, 5389, Shewanella Enterobacter sp. MTCC 25020 aerogenes MTCC 25016 Power Potentiostat mode Potentiostat Galvanostat mode Potentiostat mode Galvanostat supply 2 V mode 3 V 75 mA/m2 4 V mode 100 mode mA/m2 Stage-2 Working Graphite rod Carbon plate Carbon plate Graphite rod Graphite rod electrode modified with modified with modified with modified with graphene CNT graphene CNT and activated activated charcoal charcoal Counter Graphite plate Graphite plate Graphite felt ACC modified Carbon brush electrode modified with modified with modified with with Ni/Fe LDH modified with (Approach 1) Co.sub.9S.sub.8 ZnO nano Si/TiO.sub.2 nanowires (layered double FTO/NiO particles hydroxide) Counter Graphite plate Graphite plate Graphite felt ACC Carbon brush electrode (Approach 2) Electron Neutral red Methylene blue Neutral red and Neutral red AQDS shuttler NiO Microbes Pseudomonas Pseudomonas Clostridium Pseudomonas Serratia sp. aeruginosa strain aeruginosa strain carboxidivorans, aeruginosa strain MTCC 25017, MTCC 5388, MTCC 5388, Pseudomonas MTCC 5388, Clostridium Serratia sp. Lysinibacillus sp. putida MTCC Serratia sp. carboxidivorans, MTCC 25017, MTCC 5666, 5387, MTCC 25017, Lysinibacillus sp. Alicaligens sp. Pseudomonas Alicaligens sp. Clostridium MTCC 5666, MTCC 25022 putida MTCC MTCC 25022, beijerinckii, 5387, Clostridium Lysinibacillus sp. Clostridium beijerinckii MTCC 5666 butyricum Power Potentiostat mode Potentiostat Galvanostat mode Galvanostat mode Potentiostat supply 3 V mode 4 V 100 mA/m2 250 mA/m2 mode 6 V mode

(59) TABLE-US-00009 TABLE 9 Two stage operation of bioreactor in batch mode using kitchen waste leachate along with CO.sub.2 as C-source with experimental combinations of Example 5 Effluent Stage-2/ Stage-2/ Influent VFA/alco- approach-1, approach-2, VFA/ hols after Product Product alcohols stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 1.4 0 0.896 19.2 Ethanol 0.18 0 9.63 0.212 Butanol 0 0 11.48 0 Formic acid 2.23* 3.4 0 0 Acetic acid 8.36 14.62 3.34 2.06 Propanoic acid 3.21 0 0 0 Butanoic acid 7.09 12.34 0.98 0 Valeric acid 3.41 1.16 0.412 0 Caproic acid 3.94 0.98 0.004 0 Total 29.82 32.50 26.742 21.472 *Required amount of formic acid was amended to make-up 0.5%

(60) TABLE-US-00010 TABLE 10 Two stage operation of bioreactor in batch mode using kitchen waste leachate along with CO.sub.2 as C-source with experimental combinations of Example 6 Effluent Stage-2/ Stage-2/ Influent VFA alco- approach-1, approach-2, VFA hols after Product Product alcohols stage-1 formed formed (g/l) (g/l) (g/l) (g/D Methanol 1.4 0 1.023 16.35 Ethanol 0.18 0 11.22 0.868 Butanol 0 0 10.68 0 Formic acid 2.23* 2.21 0 0 Acetic acid 8.36 12.93 2.34 1.91 Propanoic acid 3.21 0 0 0 Butanoic acid 7.09 13.21 1.31 0 Valeric acid 3.41 0.67 0.042 0 Caproic acid 3.94 1.41 0 0 Total 29.82 30.43 26.615 19.128 *Required amount of formic acid was amended to make-up 0.5%

(61) TABLE-US-00011 TABLE 11 Two stage operation of bioreactor in batch mode using kitchen waste leachate along with CO.sub.2 as C-source with experimental combinations of Example 7 Effluent Stage-2/ Stage-2/ Influent VFA alco- approach-1, approach-2, VFA hols after Product Product alcohols stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 1.4 0 0.414 19.86 Ethanol 0.18 0 8.36 0.061 Butanol 0 0 15.41 0.216 Formic acid 2.23* 2.86 0 0 Acetic acid 8.36 8.61 2.34 2.03 Propanoic acid 3.21 6.42 0.621 0.122 Butanoic acid 7.09 14.13 0.941 0 Valeric acid 3.41 2.12 0.068 0 Caproic acid 3.94 0.34 0 0 Total 29.82 34.48 28.154 22.289 *Required amount of formic acid was amended to make-up 0.5%

(62) TABLE-US-00012 TABLE 12 Two stage operation of bioreaetor in batch mode using kitchen waste leaehate along with CO.sub.2 as C-source with experimental combinations of Example 8 Effluent Stage-2/ Stage-2/ Influent VFA alco- approach-1, approach-2, VFA hols after Product Product alcohols stage-1 formed formed (g/l) (g/l) (g/l) (g/l) Methanol 1.4 0 0.623 18.24 Ethanol 0.18 0 7.45 0.047 Butanol 0 0 13.86 0.188 Formic acid 2.23* 2.72 0 0 Acetic acid 8.36 6.96 2.86 1.641 Propanoic acid 3.21 8.16 0.787 0.063 Butanoic acid 7.09 12.88 0.612 0 Valeric acid 3.41 1.97 0 0 Caproic acid 3.94 1.62 0 0 Total 29.82 34.31 26.192 20.179 *Required amount of formic acid was amended to make-up 0.5%

(63) TABLE-US-00013 TABLE 13 Two stage operation of bioreactor in batch mode using kitehen waste leachate along with CO.sub.2 as C-source with experimental combinations of Example 9 Effluent Stage-2/ Stage-2/ Influent VFA alco- approach-1, approach-2, VFA hols after Product Product alcohols stage-1 formed formed (g/l) (g/l) (g/l) (g/L) Methanol 1.4 0 0.163 20.36 Ethanol 0.18 0 12.68 0 Butanol 0 0 9.64 0 Formic acid 2.23* 3.2 0 0 Acetic acid 8.36 10.08 3.12 3.412 Propanoic acid 3.21 0 0 0 Butanoic acid 7.09 15.16 1.08 1.126 Valeric acid 3.41 1.92 0.621 0 Caproic acid 3.94 2.04 0 0 Total 29.82 32.40 27.304 24.898 *Required amount of formic acid was amended to make-up 0.5%

(64) The results obtained from this study will represent a highly efficient, cost-effective process for the conversion of VFA rich organic waste to drop-in fuels. Hybrid process of biocatalyst activities along with electrochemical reactions, in situ H.sub.2/H.sup.+ generation during stage-2 and the syntrophic association of different groups of bacteria such as acid producing bacteria, chemoautotrophic bacteria, homoacetogens, and EAB are the key element of the process.

(65) Whereas the principal inventive concept has been described in this provisional patent application, the invention will be fully and particularly described in the complete patent application pursuant hereto.

(66) Pseudomonas stutzeri MTCC 25027 is a Gram-negative, rod-shaped, non-spore-forming bacterium, positive for both the catalase and oxidase tests and is electroactive in nature. Pseudomonas fragi MTCC 25025 is a Gram-negative, rod-shaped, non-spore-forming electroactive bacterium, oxidase test-Positive, indole negative, methyl red negative, Voges-Proskauer test negative and citrate positive. Pseudomonas aeruginosa MTCC 5389 is a Gram-negative, rod-shaped, non-spore-forming electroactive bacterium, oxidase test-Positive, indole negative, methyl red negative, Voges-Proskauer test Positive and citrate positive. Enterobacter aerogenes MTCC 25016 is a Gram-negative motile, oxidase negative, catalase positive, citrate positive, indole negative, Methyl Red-negative rod-shaped electroactive bacterium. Shewanella sp. MTCC 25020 is a facultatively anaerobic, Gram-negative rod-shaped bacteria, motile by polar flagella, extracellular electron acceptor. Pseudomonas aeruginosa MTCC 5388 is a Gram-negative, rod-shaped, non-spore-forming electroactive bacterium, oxidase test-Positive, indole Positive, methyl red. negative, Voges-Proskauer test: Positive and citrate positive. Pseudomonas putida MTCC 5387 is a Gram-negative, rod-shaped, catalase positive, cytochrome C oxidase positive, lecithinase/alpha: negative; casein hydrolysis: negative, D-trehalose: negative, Poly-&-hydroxybutyric acid: positive, extracellular electron acceptor. Alicaligens sp. MTCC 25022 is a Gram-negative, aerobic, rod-shaped bacteria, extracellular electron acceptor. Serratia sp. MTCC 25017 is a rod-shaped, Gram-negative bacteria, facultative anaerobe, high catalase-producing, electro-active bacteria, citrate positive, flagella positive, gelatin hydrolysis positive, indole negative, Motility Positive, Methyl Red: Negative, Nitrate Reduction: Positive, Oxidase Negative, rod shaped, spore negative, urease positive, Voges Proskauer: positive. Lysinibacillus sp. MTCC 5666 is a Gram-positive, motile with rod-shaped cells that produce endospores of ellipsoidal or spherical shape; positive for oxidase and catalase tests while negative for indole and H.sub.2S production.