NZ716079B2 - Processing of Biomass Materials - Google Patents
Processing of Biomass Materials Download PDFInfo
- Publication number
- NZ716079B2 NZ716079B2 NZ716079A NZ71607912A NZ716079B2 NZ 716079 B2 NZ716079 B2 NZ 716079B2 NZ 716079 A NZ716079 A NZ 716079A NZ 71607912 A NZ71607912 A NZ 71607912A NZ 716079 B2 NZ716079 B2 NZ 716079B2
- Authority
- NZ
- New Zealand
- Prior art keywords
- lignocellulosic material
- enzymes
- biomass
- mrad
- particular lignocellulosic
- Prior art date
Links
- 239000000463 material Substances 0.000 title claims abstract description 162
- 239000002028 Biomass Substances 0.000 title description 111
- 102000004190 Enzymes Human genes 0.000 claims abstract description 122
- 108090000790 Enzymes Proteins 0.000 claims abstract description 122
- 244000005700 microbiome Species 0.000 claims abstract description 58
- 108010059892 Cellulase Proteins 0.000 claims abstract description 42
- 229920002678 cellulose Polymers 0.000 claims abstract description 39
- 239000001913 cellulose Substances 0.000 claims abstract description 39
- 229940106157 CELLULASE Drugs 0.000 claims abstract description 28
- 230000001461 cytolytic Effects 0.000 claims abstract description 23
- 230000000051 modifying Effects 0.000 claims abstract description 6
- 235000000346 sugar Nutrition 0.000 claims description 43
- 238000000034 method Methods 0.000 claims description 42
- 238000000855 fermentation Methods 0.000 claims description 24
- 230000004151 fermentation Effects 0.000 claims description 24
- 229920005610 lignin Polymers 0.000 claims description 20
- 235000002017 Zea mays subsp mays Nutrition 0.000 claims description 18
- 235000005822 corn Nutrition 0.000 claims description 18
- 235000005824 corn Nutrition 0.000 claims description 18
- 239000010902 straw Substances 0.000 claims description 15
- 229920002488 Hemicellulose Polymers 0.000 claims description 12
- 108010047754 beta-Glucosidase Proteins 0.000 claims description 11
- 102000006995 beta-Glucosidase Human genes 0.000 claims description 11
- 241000233866 Fungi Species 0.000 claims description 10
- 238000010894 electron beam technology Methods 0.000 claims description 10
- 239000000758 substrate Substances 0.000 claims description 10
- 240000005979 Hordeum vulgare Species 0.000 claims description 9
- 235000007340 Hordeum vulgare Nutrition 0.000 claims description 9
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims description 9
- 241000894006 Bacteria Species 0.000 claims description 8
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 claims description 8
- 240000007594 Oryza sativa Species 0.000 claims description 7
- 241000223259 Trichoderma Species 0.000 claims description 7
- 240000008529 Triticum aestivum Species 0.000 claims description 7
- 239000002245 particle Substances 0.000 claims description 7
- 239000007787 solid Substances 0.000 claims description 7
- 239000010907 stover Substances 0.000 claims description 7
- 241000609240 Ambelania acida Species 0.000 claims description 6
- 244000075850 Avena orientalis Species 0.000 claims description 6
- 235000007319 Avena orientalis Nutrition 0.000 claims description 6
- 235000007558 Avena sp Nutrition 0.000 claims description 6
- 235000007164 Oryza sativa Nutrition 0.000 claims description 6
- 241000209504 Poaceae Species 0.000 claims description 6
- 241000499912 Trichoderma reesei Species 0.000 claims description 6
- 239000010905 bagasse Substances 0.000 claims description 6
- 235000009566 rice Nutrition 0.000 claims description 6
- 235000021307 wheat Nutrition 0.000 claims description 6
- 241001474374 Blennius Species 0.000 claims description 5
- 241000195493 Cryptophyta Species 0.000 claims description 5
- 235000002595 Solanum tuberosum Nutrition 0.000 claims description 5
- 240000001016 Solanum tuberosum Species 0.000 claims description 5
- 238000005119 centrifugation Methods 0.000 claims description 5
- 238000004587 chromatography analysis Methods 0.000 claims description 5
- 108010008885 Cellulose 1,4-beta-Cellobiosidase Proteins 0.000 claims description 4
- 229920000742 Cotton Polymers 0.000 claims description 4
- 240000008042 Zea mays Species 0.000 claims description 4
- 239000002154 agricultural waste Substances 0.000 claims description 4
- 238000000605 extraction Methods 0.000 claims description 4
- 239000002904 solvent Substances 0.000 claims description 4
- 240000009030 Agave Species 0.000 claims description 3
- 240000005337 Agave sisalana Species 0.000 claims description 3
- 240000000116 Alocasia Species 0.000 claims description 3
- 235000016068 Berberis vulgaris Nutrition 0.000 claims description 3
- 241000335053 Beta vulgaris Species 0.000 claims description 3
- 235000006008 Brassica napus var napus Nutrition 0.000 claims description 3
- 240000000385 Brassica napus var. napus Species 0.000 claims description 3
- 235000006618 Brassica rapa subsp oleifera Nutrition 0.000 claims description 3
- 235000004977 Brassica sinapistrum Nutrition 0.000 claims description 3
- 240000000218 Cannabis sativa Species 0.000 claims description 3
- 235000006481 Colocasia esculenta Nutrition 0.000 claims description 3
- 240000000491 Corchorus aestuans Species 0.000 claims description 3
- 235000011777 Corchorus aestuans Nutrition 0.000 claims description 3
- 235000010862 Corchorus capsularis Nutrition 0.000 claims description 3
- 235000009419 Fagopyrum esculentum Nutrition 0.000 claims description 3
- 240000008620 Fagopyrum esculentum Species 0.000 claims description 3
- 235000010469 Glycine max Nutrition 0.000 claims description 3
- 240000007842 Glycine max Species 0.000 claims description 3
- 235000002678 Ipomoea batatas Nutrition 0.000 claims description 3
- 240000003613 Ipomoea batatas Species 0.000 claims description 3
- 240000004322 Lens culinaris Species 0.000 claims description 3
- 235000014647 Lens culinaris subsp culinaris Nutrition 0.000 claims description 3
- 235000004431 Linum usitatissimum Nutrition 0.000 claims description 3
- 240000006240 Linum usitatissimum Species 0.000 claims description 3
- 240000003183 Manihot esculenta Species 0.000 claims description 3
- 235000016735 Manihot esculenta subsp esculenta Nutrition 0.000 claims description 3
- 240000000907 Musa textilis Species 0.000 claims description 3
- 235000008469 Oxalis tuberosa Nutrition 0.000 claims description 3
- 240000000645 Oxalis tuberosa Species 0.000 claims description 3
- 235000010627 Phaseolus vulgaris Nutrition 0.000 claims description 3
- 240000005158 Phaseolus vulgaris Species 0.000 claims description 3
- 240000004713 Pisum sativum Species 0.000 claims description 3
- 235000010582 Pisum sativum Nutrition 0.000 claims description 3
- 235000010575 Pueraria lobata Nutrition 0.000 claims description 3
- 244000046146 Pueraria lobata Species 0.000 claims description 3
- 235000008984 brauner Senf Nutrition 0.000 claims description 3
- 235000009120 camo Nutrition 0.000 claims description 3
- 235000013339 cereals Nutrition 0.000 claims description 3
- 235000005607 chanvre indien Nutrition 0.000 claims description 3
- 235000004879 dioscorea Nutrition 0.000 claims description 3
- 238000001704 evaporation Methods 0.000 claims description 3
- 239000011487 hemp Substances 0.000 claims description 3
- 235000012765 hemp Nutrition 0.000 claims description 3
- 235000012766 marijuana Nutrition 0.000 claims description 3
- 239000004460 silage Substances 0.000 claims description 3
- 235000017166 Bambusa arundinacea Nutrition 0.000 claims description 2
- 240000003917 Bambusa tulda Species 0.000 claims description 2
- 235000017491 Bambusa tulda Nutrition 0.000 claims description 2
- 235000013162 Cocos nucifera Nutrition 0.000 claims description 2
- 240000007170 Cocos nucifera Species 0.000 claims description 2
- 240000005561 Musa balbisiana Species 0.000 claims description 2
- 235000018290 Musa x paradisiaca Nutrition 0.000 claims description 2
- 235000015334 Phyllostachys viridis Nutrition 0.000 claims description 2
- 239000011425 bamboo Substances 0.000 claims description 2
- 238000009837 dry grinding Methods 0.000 claims description 2
- 238000001914 filtration Methods 0.000 claims description 2
- 230000000754 repressing Effects 0.000 claims description 2
- 238000001238 wet grinding Methods 0.000 claims description 2
- 241000193403 Clostridium Species 0.000 claims 1
- 210000003666 Nerve Fibers, Myelinated Anatomy 0.000 claims 1
- 238000000502 dialysis Methods 0.000 claims 1
- 239000007921 spray Substances 0.000 claims 1
- 229940088598 Enzyme Drugs 0.000 description 65
- 239000000047 product Substances 0.000 description 54
- 239000000203 mixture Substances 0.000 description 44
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 40
- 238000004519 manufacturing process Methods 0.000 description 39
- 239000002609 media Substances 0.000 description 33
- 239000000123 paper Substances 0.000 description 26
- WQZGKKKJIJFFOK-VFUOTHLCSA-N β-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 25
- WQZGKKKJIJFFOK-GASJEMHNSA-N D-Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 24
- 239000008103 glucose Substances 0.000 description 24
- 150000008163 sugars Chemical class 0.000 description 24
- 241000196324 Embryophyta Species 0.000 description 22
- 239000000411 inducer Substances 0.000 description 21
- 210000004027 cells Anatomy 0.000 description 20
- -1 card stock Substances 0.000 description 18
- 239000000243 solution Substances 0.000 description 16
- 241000209149 Zea Species 0.000 description 14
- 239000000543 intermediate Substances 0.000 description 14
- 239000002029 lignocellulosic biomass Substances 0.000 description 14
- 235000018102 proteins Nutrition 0.000 description 14
- 102000004169 proteins and genes Human genes 0.000 description 14
- 108090000623 proteins and genes Proteins 0.000 description 14
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 14
- 230000000694 effects Effects 0.000 description 13
- 150000001298 alcohols Chemical class 0.000 description 12
- 230000001939 inductive effect Effects 0.000 description 12
- 239000000446 fuel Substances 0.000 description 11
- 230000012010 growth Effects 0.000 description 10
- SRBFZHDQGSBBOR-SQOUGZDYSA-N Xylose Natural products O[C@@H]1CO[C@@H](O)[C@@H](O)[C@@H]1O SRBFZHDQGSBBOR-SQOUGZDYSA-N 0.000 description 9
- 239000007788 liquid Substances 0.000 description 9
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 8
- GUBGYTABKSRVRQ-CUHNMECISA-N D-Cellobiose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-CUHNMECISA-N 0.000 description 8
- 239000002253 acid Substances 0.000 description 8
- 238000007792 addition Methods 0.000 description 8
- LRHPLDYGYMQRHN-UHFFFAOYSA-N n-butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 description 8
- 239000000126 substance Substances 0.000 description 8
- 150000001720 carbohydrates Chemical class 0.000 description 7
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 7
- 235000013305 food Nutrition 0.000 description 7
- 239000011780 sodium chloride Substances 0.000 description 7
- 239000002023 wood Substances 0.000 description 7
- 206010048652 Enzyme induction Diseases 0.000 description 6
- QTBSBXVTEAMEQO-UHFFFAOYSA-N acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 6
- CSCPPACGZOOCGX-UHFFFAOYSA-N acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 6
- 150000007513 acids Chemical class 0.000 description 6
- 239000000560 biocompatible material Substances 0.000 description 6
- 235000014633 carbohydrates Nutrition 0.000 description 6
- PEDCQBHIVMGVHV-UHFFFAOYSA-N glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 6
- 230000001965 increased Effects 0.000 description 6
- 230000000813 microbial Effects 0.000 description 6
- 238000002156 mixing Methods 0.000 description 6
- 239000001301 oxygen Substances 0.000 description 6
- 229910052760 oxygen Inorganic materials 0.000 description 6
- MYMOFIZGZYHOMD-UHFFFAOYSA-N oxygen Chemical compound O=O MYMOFIZGZYHOMD-UHFFFAOYSA-N 0.000 description 6
- 229920000642 polymer Polymers 0.000 description 6
- 230000001603 reducing Effects 0.000 description 6
- 150000003839 salts Chemical class 0.000 description 6
- 239000002699 waste material Substances 0.000 description 6
- 241001019659 Acremonium <Plectosphaerellaceae> Species 0.000 description 5
- 235000007466 Corylus avellana Nutrition 0.000 description 5
- 240000007582 Corylus avellana Species 0.000 description 5
- UNXHWFMMPAWVPI-QWWZWVQMSA-N D-Threitol Natural products OC[C@@H](O)[C@H](O)CO UNXHWFMMPAWVPI-QWWZWVQMSA-N 0.000 description 5
- 229960003487 Xylose Drugs 0.000 description 5
- 238000006243 chemical reaction Methods 0.000 description 5
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 5
- 238000004880 explosion Methods 0.000 description 5
- 238000000227 grinding Methods 0.000 description 5
- 239000001963 growth media Substances 0.000 description 5
- 150000002500 ions Chemical class 0.000 description 5
- 150000007524 organic acids Chemical class 0.000 description 5
- 235000005985 organic acids Nutrition 0.000 description 5
- 230000003647 oxidation Effects 0.000 description 5
- 238000007254 oxidation reaction Methods 0.000 description 5
- FERIUCNNQQJTOY-UHFFFAOYSA-N Butanoic acid Chemical compound CCCC(O)=O FERIUCNNQQJTOY-UHFFFAOYSA-N 0.000 description 4
- 102000005575 Cellulases Human genes 0.000 description 4
- 108010084185 Cellulases Proteins 0.000 description 4
- BJHIKXHVCXFQLS-UYFOZJQFSA-N Fructose Natural products OC[C@@H](O)[C@@H](O)[C@H](O)C(=O)CO BJHIKXHVCXFQLS-UYFOZJQFSA-N 0.000 description 4
- ZXEKIIBDNHEJCQ-UHFFFAOYSA-N Isobutanol Chemical compound CC(C)CO ZXEKIIBDNHEJCQ-UHFFFAOYSA-N 0.000 description 4
- 229920002472 Starch Polymers 0.000 description 4
- 230000003115 biocidal Effects 0.000 description 4
- 229910052799 carbon Inorganic materials 0.000 description 4
- 238000005984 hydrogenation reaction Methods 0.000 description 4
- 239000011261 inert gas Substances 0.000 description 4
- 238000003801 milling Methods 0.000 description 4
- IJGRMHOSHXDMSA-UHFFFAOYSA-N nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 4
- 238000000527 sonication Methods 0.000 description 4
- 239000004094 surface-active agent Substances 0.000 description 4
- GZCGUPFRVQAUEE-KCDKBNATSA-N D-(+)-Galactose Natural products OC[C@@H](O)[C@H](O)[C@H](O)[C@@H](O)C=O GZCGUPFRVQAUEE-KCDKBNATSA-N 0.000 description 3
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 description 3
- 241000219146 Gossypium Species 0.000 description 3
- 241001520808 Panicum virgatum Species 0.000 description 3
- 229920001131 Pulp (paper) Polymers 0.000 description 3
- 239000000654 additive Substances 0.000 description 3
- 239000003242 anti bacterial agent Substances 0.000 description 3
- 150000001479 arabinose derivatives Chemical class 0.000 description 3
- 235000013405 beer Nutrition 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 239000006227 byproduct Substances 0.000 description 3
- 150000001735 carboxylic acids Chemical class 0.000 description 3
- 239000003795 chemical substances by application Substances 0.000 description 3
- 238000004821 distillation Methods 0.000 description 3
- 230000005611 electricity Effects 0.000 description 3
- 239000000835 fiber Substances 0.000 description 3
- 239000012467 final product Substances 0.000 description 3
- BDAGIHXWWSANSR-UHFFFAOYSA-N formic acid Chemical compound OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 3
- 230000002068 genetic Effects 0.000 description 3
- 239000001257 hydrogen Substances 0.000 description 3
- 229910052739 hydrogen Inorganic materials 0.000 description 3
- 238000002955 isolation Methods 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- VNWKTOKETHGBQD-UHFFFAOYSA-N methane Chemical compound C VNWKTOKETHGBQD-UHFFFAOYSA-N 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 238000006011 modification reaction Methods 0.000 description 3
- 235000015097 nutrients Nutrition 0.000 description 3
- MUBZPKHOEPUJKR-UHFFFAOYSA-N oxalic acid Chemical compound OC(=O)C(O)=O MUBZPKHOEPUJKR-UHFFFAOYSA-N 0.000 description 3
- 239000010893 paper waste Substances 0.000 description 3
- XBDQKXXYIPTUBI-UHFFFAOYSA-N propionic acid Chemical compound CCC(O)=O XBDQKXXYIPTUBI-UHFFFAOYSA-N 0.000 description 3
- 238000000197 pyrolysis Methods 0.000 description 3
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 241000894007 species Species 0.000 description 3
- 235000019698 starch Nutrition 0.000 description 3
- 239000008107 starch Substances 0.000 description 3
- WQZGKKKJIJFFOK-PHYPRBDBSA-N α-D-galactose Chemical compound OC[C@H]1O[C@H](O)[C@H](O)[C@@H](O)[C@H]1O WQZGKKKJIJFFOK-PHYPRBDBSA-N 0.000 description 3
- UCSJYZPVAKXKNQ-HZYVHMACSA-N 1-[(1S,2R,3R,4S,5R,6R)-3-carbamimidamido-6-{[(2R,3R,4R,5S)-3-{[(2S,3S,4S,5R,6S)-4,5-dihydroxy-6-(hydroxymethyl)-3-(methylamino)oxan-2-yl]oxy}-4-formyl-4-hydroxy-5-methyloxolan-2-yl]oxy}-2,4,5-trihydroxycyclohexyl]guanidine Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- BTANRVKWQNVYAZ-UHFFFAOYSA-N 2-Butanol Chemical compound CCC(C)O BTANRVKWQNVYAZ-UHFFFAOYSA-N 0.000 description 2
- YJISHJVIRFPGGN-UHFFFAOYSA-N 5-[5-[3,4-dihydroxy-6-(hydroxymethyl)-5-methoxyoxan-2-yl]oxy-6-[[3,4-dihydroxy-6-(hydroxymethyl)-5-methoxyoxan-2-yl]oxymethyl]-3,4-dihydroxyoxan-2-yl]oxy-6-(hydroxymethyl)-2-methyloxane-3,4-diol Chemical compound O1C(CO)C(OC)C(O)C(O)C1OCC1C(OC2C(C(O)C(OC)C(CO)O2)O)C(O)C(O)C(OC2C(OC(C)C(O)C2O)CO)O1 YJISHJVIRFPGGN-UHFFFAOYSA-N 0.000 description 2
- 241001438635 Acremonium brachypenium Species 0.000 description 2
- 229940064005 Antibiotic throat preparations Drugs 0.000 description 2
- 229940083879 Antibiotics FOR TREATMENT OF HEMORRHOIDS AND ANAL FISSURES FOR TOPICAL USE Drugs 0.000 description 2
- 229940042052 Antibiotics for systemic use Drugs 0.000 description 2
- 229940042786 Antitubercular Antibiotics Drugs 0.000 description 2
- HEBKCHPVOIAQTA-QWWZWVQMSA-N Arabitol Chemical compound OC[C@@H](O)C(O)[C@H](O)CO HEBKCHPVOIAQTA-QWWZWVQMSA-N 0.000 description 2
- 241000082175 Arracacia xanthorrhiza Species 0.000 description 2
- 241000193830 Bacillus <bacterium> Species 0.000 description 2
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 2
- FBPFZTCFMRRESA-JGWLITMVSA-N D-glucitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-JGWLITMVSA-N 0.000 description 2
- 241000602080 Dracaena fragrans Species 0.000 description 2
- 229940096118 Ella Drugs 0.000 description 2
- 239000004386 Erythritol Substances 0.000 description 2
- 229940009714 Erythritol Drugs 0.000 description 2
- 210000003608 Feces Anatomy 0.000 description 2
- 239000005715 Fructose Substances 0.000 description 2
- 241001019284 Gliomastix roseogrisea Species 0.000 description 2
- JFCQEDHGNNZCLN-UHFFFAOYSA-N Glutaric acid Chemical compound OC(=O)CCCC(O)=O JFCQEDHGNNZCLN-UHFFFAOYSA-N 0.000 description 2
- 229940093922 Gynecological Antibiotics Drugs 0.000 description 2
- FUZZWVXGSFPDMH-UHFFFAOYSA-N Hexanoic acid Chemical compound CCCCCC(O)=O FUZZWVXGSFPDMH-UHFFFAOYSA-N 0.000 description 2
- 241001480714 Humicola insolens Species 0.000 description 2
- GUBGYTABKSRVRQ-UUNJERMWSA-N Lactose Natural products O([C@@H]1[C@H](O)[C@H](O)[C@H](O)O[C@@H]1CO)[C@H]1[C@@H](O)[C@@H](O)[C@H](O)[C@H](CO)O1 GUBGYTABKSRVRQ-UUNJERMWSA-N 0.000 description 2
- OYHQOLUKZRVURQ-IXWMQOLASA-N Linoleic acid Natural products CCCCC\C=C/C\C=C\CCCCCCCC(O)=O OYHQOLUKZRVURQ-IXWMQOLASA-N 0.000 description 2
- VQHSOMBJVWLPSR-WUJBLJFYSA-N Maltitol Chemical compound OC[C@H](O)[C@@H](O)[C@@H]([C@H](O)CO)O[C@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O VQHSOMBJVWLPSR-WUJBLJFYSA-N 0.000 description 2
- 240000004658 Medicago sativa Species 0.000 description 2
- 240000003433 Miscanthus floridulus Species 0.000 description 2
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N Oleic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 description 2
- IPCSVZSSVZVIGE-UHFFFAOYSA-N Palmitic acid Chemical compound CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 description 2
- 240000006169 Phalaris arundinacea Species 0.000 description 2
- 241000235648 Pichia Species 0.000 description 2
- 241000183024 Populus tremula Species 0.000 description 2
- 241000906075 Simplicillium obclavatum Species 0.000 description 2
- HIWPGCMGAMJNRG-ACCAVRKYSA-N Sophorose Natural products O([C@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O)[C@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HIWPGCMGAMJNRG-ACCAVRKYSA-N 0.000 description 2
- 240000003829 Sorghum propinquum Species 0.000 description 2
- 235000011684 Sorghum saccharatum Nutrition 0.000 description 2
- 241000746413 Spartina Species 0.000 description 2
- QIQXTHQIDYTFRH-UHFFFAOYSA-N Stearic acid Chemical compound CCCCCCCCCCCCCCCCCC(O)=O QIQXTHQIDYTFRH-UHFFFAOYSA-N 0.000 description 2
- KDYFGRWQOYBRFD-UHFFFAOYSA-N Succinic acid Chemical compound OC(=O)CCC(O)=O KDYFGRWQOYBRFD-UHFFFAOYSA-N 0.000 description 2
- 229940024982 Topical Antifungal Antibiotics Drugs 0.000 description 2
- NQPDZGIKBAWPEJ-UHFFFAOYSA-N Valeric acid Chemical compound CCCCC(O)=O NQPDZGIKBAWPEJ-UHFFFAOYSA-N 0.000 description 2
- HEBKCHPVOIAQTA-SCDXWVJYSA-N Xylitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)CO HEBKCHPVOIAQTA-SCDXWVJYSA-N 0.000 description 2
- 229960002675 Xylitol Drugs 0.000 description 2
- 241000588901 Zymomonas Species 0.000 description 2
- IKHGUXGNUITLKF-UHFFFAOYSA-N acetaldehyde Chemical compound CC=O IKHGUXGNUITLKF-UHFFFAOYSA-N 0.000 description 2
- 235000011054 acetic acid Nutrition 0.000 description 2
- 235000017585 alfalfa Nutrition 0.000 description 2
- 235000017587 alfalfa Nutrition 0.000 description 2
- 235000001014 amino acid Nutrition 0.000 description 2
- 150000001413 amino acids Chemical class 0.000 description 2
- 230000003698 anagen phase Effects 0.000 description 2
- 230000000845 anti-microbial Effects 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
- XKRFYHLGVUSROY-UHFFFAOYSA-N argon Chemical compound [Ar] XKRFYHLGVUSROY-UHFFFAOYSA-N 0.000 description 2
- 229910052786 argon Inorganic materials 0.000 description 2
- 244000052616 bacterial pathogens Species 0.000 description 2
- 238000004166 bioassay Methods 0.000 description 2
- 239000003225 biodiesel Substances 0.000 description 2
- 238000009835 boiling Methods 0.000 description 2
- 230000001488 breeding Effects 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- 229910002092 carbon dioxide Inorganic materials 0.000 description 2
- 239000011111 cardboard Substances 0.000 description 2
- 239000000969 carrier Substances 0.000 description 2
- 230000015556 catabolic process Effects 0.000 description 2
- 239000003054 catalyst Substances 0.000 description 2
- 150000001768 cations Chemical class 0.000 description 2
- 101710014331 celS Proteins 0.000 description 2
- 239000007795 chemical reaction product Substances 0.000 description 2
- 235000015165 citric acid Nutrition 0.000 description 2
- 238000002485 combustion reaction Methods 0.000 description 2
- 238000005520 cutting process Methods 0.000 description 2
- 230000004059 degradation Effects 0.000 description 2
- 238000006731 degradation reaction Methods 0.000 description 2
- 229920003013 deoxyribonucleic acid Polymers 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 239000000539 dimer Substances 0.000 description 2
- 230000002708 enhancing Effects 0.000 description 2
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 description 2
- 235000019414 erythritol Nutrition 0.000 description 2
- 239000012530 fluid Substances 0.000 description 2
- WSFSSNUMVMOOMR-UHFFFAOYSA-N formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 2
- 238000005755 formation reaction Methods 0.000 description 2
- 150000004676 glycans Polymers 0.000 description 2
- AEMRFAOFKBGASW-UHFFFAOYSA-N glycolic acid Chemical compound OCC(O)=O AEMRFAOFKBGASW-UHFFFAOYSA-N 0.000 description 2
- 229910052734 helium Inorganic materials 0.000 description 2
- 238000011081 inoculation Methods 0.000 description 2
- 229940079866 intestinal antibiotics Drugs 0.000 description 2
- 235000014655 lactic acid Nutrition 0.000 description 2
- GUBGYTABKSRVRQ-XLOQQCSPSA-N lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 2
- 239000008101 lactose Substances 0.000 description 2
- 108010062085 ligninase Proteins 0.000 description 2
- 239000010871 livestock manure Substances 0.000 description 2
- 238000011068 load Methods 0.000 description 2
- OFOBLEOULBTSOW-UHFFFAOYSA-N malonic acid Chemical compound OC(=O)CC(O)=O OFOBLEOULBTSOW-UHFFFAOYSA-N 0.000 description 2
- 239000011159 matrix material Substances 0.000 description 2
- 239000002207 metabolite Substances 0.000 description 2
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- 229920001542 oligosaccharide Polymers 0.000 description 2
- 150000002482 oligosaccharides Polymers 0.000 description 2
- 229940005935 ophthalmologic Antibiotics Drugs 0.000 description 2
- 230000001590 oxidative Effects 0.000 description 2
- 239000011087 paperboard Substances 0.000 description 2
- 239000003208 petroleum Substances 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- 229920001690 polydopamine Polymers 0.000 description 2
- 229920005862 polyol Polymers 0.000 description 2
- 150000003077 polyols Chemical class 0.000 description 2
- 229920001282 polysaccharide Polymers 0.000 description 2
- 239000005017 polysaccharide Substances 0.000 description 2
- 150000004804 polysaccharides Polymers 0.000 description 2
- 229920000136 polysorbate Polymers 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 230000000644 propagated Effects 0.000 description 2
- BDERNNFJNOPAEC-UHFFFAOYSA-N propanol Chemical compound CCCO BDERNNFJNOPAEC-UHFFFAOYSA-N 0.000 description 2
- DNIAPMSPPWPWGF-UHFFFAOYSA-N propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 description 2
- 230000002829 reduced Effects 0.000 description 2
- 239000010865 sewage Substances 0.000 description 2
- 238000010008 shearing Methods 0.000 description 2
- 108010027322 single cell proteins Proteins 0.000 description 2
- 150000003384 small molecules Chemical class 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 239000000600 sorbitol Substances 0.000 description 2
- DKGAVHZHDRPRBM-UHFFFAOYSA-N t-BuOH Chemical compound CC(C)(C)O DKGAVHZHDRPRBM-UHFFFAOYSA-N 0.000 description 2
- 229960000200 ulipristal Drugs 0.000 description 2
- OOLLAFOLCSJHRE-ZHAKMVSLSA-N ulipristal acetate Chemical compound C1=CC(N(C)C)=CC=C1[C@@H]1C2=C3CCC(=O)C=C3CC[C@H]2[C@H](CC[C@]2(OC(C)=O)C(C)=O)[C@]2(C)C1 OOLLAFOLCSJHRE-ZHAKMVSLSA-N 0.000 description 2
- XSQUKJJJFZCRTK-UHFFFAOYSA-N urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 2
- 229920001221 xylan Polymers 0.000 description 2
- 150000004823 xylans Chemical class 0.000 description 2
- 239000000811 xylitol Substances 0.000 description 2
- 235000010447 xylitol Nutrition 0.000 description 2
- XJCCHWKNFMUJFE-CGQAXDJHSA-N (2S,3R,4R,5R)-4-[(2R,3R,4R,5S,6R)-3,4-dihydroxy-6-(hydroxymethyl)-5-[(2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxyoxan-2-yl]oxyhexane-1,2,3,5,6-pentol Chemical compound O[C@@H]1[C@@H](O)[C@@H](O[C@@H]([C@H](O)[C@@H](O)CO)[C@H](O)CO)O[C@H](CO)[C@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 XJCCHWKNFMUJFE-CGQAXDJHSA-N 0.000 description 1
- LUAHEUHBAZYUOI-KVXMBEGHSA-N (2S,3R,4R,5R)-4-[(2R,3R,4R,5S,6R)-5-[(2R,3R,4R,5S,6R)-3,4-dihydroxy-6-(hydroxymethyl)-5-[(2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxyoxan-2-yl]oxy-3,4-dihydroxy-6-(hydroxymethyl)oxan-2-yl]oxyhexane-1,2,3,5,6-pentol Chemical compound O[C@@H]1[C@@H](O)[C@@H](O[C@@H]([C@H](O)[C@@H](O)CO)[C@H](O)CO)O[C@H](CO)[C@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O[C@@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)O)[C@@H](CO)O1 LUAHEUHBAZYUOI-KVXMBEGHSA-N 0.000 description 1
- ALRHLSYJTWAHJZ-UHFFFAOYSA-N 3-Hydroxypropionic acid Chemical compound OCCC(O)=O ALRHLSYJTWAHJZ-UHFFFAOYSA-N 0.000 description 1
- 229940100198 ALKYLATING AGENTS Drugs 0.000 description 1
- 241001114518 Acaulium acremonium Species 0.000 description 1
- 241001438625 Acremonium dichromosporum Species 0.000 description 1
- 241000228209 Acremonium persicinum Species 0.000 description 1
- 241001019292 Acremonium pinkertoniae Species 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 241001466460 Alveolata Species 0.000 description 1
- BFNBIHQBYMNNAN-UHFFFAOYSA-N Ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 1
- AVKUERGKIZMTKX-NJBDSQKTSA-N Ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 1
- 241000228212 Aspergillus Species 0.000 description 1
- APKFDSVGJQXUKY-INPOYWNPSA-N BRL-49594 Chemical compound O[C@H]1[C@@H](N)[C@H](O)[C@@H](C)O[C@H]1O[C@H]1/C=C/C=C/C=C/C=C/C=C/C=C/C=C/[C@H](C)[C@@H](O)[C@@H](C)[C@H](C)OC(=O)C[C@H](O)C[C@H](O)CC[C@@H](O)[C@H](O)C[C@H](O)C[C@](O)(C[C@H](O)[C@H]2C(O)=O)O[C@H]2C1 APKFDSVGJQXUKY-INPOYWNPSA-N 0.000 description 1
- 229940098773 Bovine Serum Albumin Drugs 0.000 description 1
- 108091003117 Bovine Serum Albumin Proteins 0.000 description 1
- 238000009010 Bradford assay Methods 0.000 description 1
- 101700010451 CELB Proteins 0.000 description 1
- 241000222120 Candida <Saccharomycetales> Species 0.000 description 1
- 229940041514 Candida albicans extract Drugs 0.000 description 1
- 235000007436 Cassia auriculata Nutrition 0.000 description 1
- 244000007668 Cassia auriculata Species 0.000 description 1
- 210000002421 Cell Wall Anatomy 0.000 description 1
- 241000135254 Cephalosporium sp. Species 0.000 description 1
- WIIZWVCIJKGZOK-RKDXNWHRSA-N Chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 1
- 229960005091 Chloramphenicol Drugs 0.000 description 1
- 241001147674 Chlorarachniophyceae Species 0.000 description 1
- 241000123346 Chrysosporium Species 0.000 description 1
- 241001674013 Chrysosporium lucknowense Species 0.000 description 1
- 241000223782 Ciliophora Species 0.000 description 1
- MYSWGUAQZAJSOK-UHFFFAOYSA-N Ciprofloxacin Chemical compound C12=CC(N3CCNCC3)=C(F)C=C2C(=O)C(C(=O)O)=CN1C1CC1 MYSWGUAQZAJSOK-UHFFFAOYSA-N 0.000 description 1
- 229920002261 Corn starch Polymers 0.000 description 1
- FBPFZTCFMRRESA-KAZBKCHUSA-N D-Mannitol Natural products OC[C@@H](O)[C@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KAZBKCHUSA-N 0.000 description 1
- 101700061444 DDX25 Proteins 0.000 description 1
- 241000235035 Debaryomyces Species 0.000 description 1
- GRRNUXAQVGOGFE-UHFFFAOYSA-N Destomysin Chemical compound OC1C(NC)CC(N)C(O)C1OC1C2OC3(C(C(O)C(O)C(C(N)CO)O3)O)OC2C(O)C(CO)O1 GRRNUXAQVGOGFE-UHFFFAOYSA-N 0.000 description 1
- 241000237537 Ensis Species 0.000 description 1
- 239000005977 Ethylene Substances 0.000 description 1
- 241000195623 Euglenida Species 0.000 description 1
- HYBBIBNJHNGZAN-UHFFFAOYSA-N Furfural Chemical compound O=CC1=CC=CO1 HYBBIBNJHNGZAN-UHFFFAOYSA-N 0.000 description 1
- 241000223221 Fusarium oxysporum Species 0.000 description 1
- 101700041462 GUX2 Proteins 0.000 description 1
- FBPFZTCFMRRESA-GUCUJZIJSA-N Galactitol Chemical compound OC[C@H](O)[C@@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-GUCUJZIJSA-N 0.000 description 1
- 229920001706 Glucuronoxylan Polymers 0.000 description 1
- 235000004341 Gossypium herbaceum Nutrition 0.000 description 1
- 240000002024 Gossypium herbaceum Species 0.000 description 1
- 241000206759 Haptophyceae Species 0.000 description 1
- 241000223198 Humicola Species 0.000 description 1
- 229940097277 Hygromycin B Drugs 0.000 description 1
- 241000282619 Hylobates lar Species 0.000 description 1
- FBPFZTCFMRRESA-ZXXMMSQZSA-N Iditol Chemical compound OC[C@@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-ZXXMMSQZSA-N 0.000 description 1
- CDAISMWEOUEBRE-GPIVLXJGSA-N Inositol Chemical compound O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@H](O)[C@@H]1O CDAISMWEOUEBRE-GPIVLXJGSA-N 0.000 description 1
- 229960000367 Inositol Drugs 0.000 description 1
- SERLAGPUMNYUCK-DCUALPFSSA-N Isomalt Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO[C@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O SERLAGPUMNYUCK-DCUALPFSSA-N 0.000 description 1
- SBUJHOSQTJFQJX-NOAMYHISSA-N Kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 description 1
- 244000285963 Kluyveromyces fragilis Species 0.000 description 1
- 230000035832 Lag time Effects 0.000 description 1
- 230000035648 Lag-time Effects 0.000 description 1
- 229940061634 Magnesium sulfate heptahydrate Drugs 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- 241000226677 Myceliophthora Species 0.000 description 1
- 239000005642 Oleic acid Substances 0.000 description 1
- 229950010131 PUROMYCIN Drugs 0.000 description 1
- RXWNCPJZOCPEPQ-NVWDDTSBSA-N PUROMYCIN Chemical compound C1=CC(OC)=CC=C1C[C@H](N)C(=O)N[C@H]1[C@@H](O)[C@H](N2C3=NC=NC(=C3N=C2)N(C)C)O[C@@H]1CO RXWNCPJZOCPEPQ-NVWDDTSBSA-N 0.000 description 1
- 241000235652 Pachysolen Species 0.000 description 1
- 235000021314 Palmitic acid Nutrition 0.000 description 1
- 229940049954 Penicillin Drugs 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- 241000228143 Penicillium Species 0.000 description 1
- 241000364057 Peoria Species 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- 229940066779 Peptones Drugs 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 241000589516 Pseudomonas Species 0.000 description 1
- 241000893045 Pseudozyma Species 0.000 description 1
- 229940100486 RICE STARCH Drugs 0.000 description 1
- 239000007868 Raney catalyst Substances 0.000 description 1
- 229910000564 Raney nickel Inorganic materials 0.000 description 1
- 108020004511 Recombinant DNA Proteins 0.000 description 1
- PNNNRSAQSRJVSB-BXKVDMCESA-N Rhamnose Chemical compound C[C@H](O)[C@H](O)[C@@H](O)[C@@H](O)C=O PNNNRSAQSRJVSB-BXKVDMCESA-N 0.000 description 1
- 241000206572 Rhodophyta Species 0.000 description 1
- HEBKCHPVOIAQTA-ZXFHETKHSA-N Ribitol Chemical compound OC[C@H](O)[C@H](O)[C@H](O)CO HEBKCHPVOIAQTA-ZXFHETKHSA-N 0.000 description 1
- 241000193448 Ruminiclostridium thermocellum Species 0.000 description 1
- 241000235070 Saccharomyces Species 0.000 description 1
- 241000190542 Sarocladium kiliense Species 0.000 description 1
- 235000021355 Stearic acid Nutrition 0.000 description 1
- 241001466451 Stramenopiles Species 0.000 description 1
- 241000187747 Streptomyces Species 0.000 description 1
- 229960005322 Streptomycin Drugs 0.000 description 1
- 210000003283 T-Lymphocytes, Helper-Inducer Anatomy 0.000 description 1
- 241001313536 Thermothelomyces thermophila Species 0.000 description 1
- 241001495429 Thielavia terrestris Species 0.000 description 1
- 241000006364 Torula Species 0.000 description 1
- 241000378866 Trichoderma koningii Species 0.000 description 1
- 241001480015 Trigonopsis variabilis Species 0.000 description 1
- 239000007983 Tris buffer Substances 0.000 description 1
- 241000221533 Ustilaginomycetes Species 0.000 description 1
- OXQKEKGBFMQTML-KVTDHHQDSA-N Volemitol Chemical compound OC[C@@H](O)[C@@H](O)C(O)[C@H](O)[C@H](O)CO OXQKEKGBFMQTML-KVTDHHQDSA-N 0.000 description 1
- 229940100445 WHEAT STARCH Drugs 0.000 description 1
- 229920002000 Xyloglucan Polymers 0.000 description 1
- 241000235017 Zygosaccharomyces Species 0.000 description 1
- 241000588902 Zymomonas mobilis Species 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K [O-]P([O-])([O-])=O Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- NIXOWILDQLNWCW-UHFFFAOYSA-N acrylic acid Chemical compound OC(=O)C=C NIXOWILDQLNWCW-UHFFFAOYSA-N 0.000 description 1
- 230000000996 additive Effects 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 229930013930 alkaloids Natural products 0.000 description 1
- 150000001336 alkenes Chemical class 0.000 description 1
- 239000002168 alkylating agent Substances 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- QGZKDVFQNNGYKY-UHFFFAOYSA-N ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 1
- 150000003863 ammonium salts Chemical class 0.000 description 1
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 1
- 235000011130 ammonium sulphate Nutrition 0.000 description 1
- 239000002280 amphoteric surfactant Substances 0.000 description 1
- 229960003942 amphotericin B Drugs 0.000 description 1
- 229960000723 ampicillin Drugs 0.000 description 1
- 239000004599 antimicrobial Substances 0.000 description 1
- 229920000617 arabinoxylan Polymers 0.000 description 1
- 150000004783 arabinoxylans Chemical class 0.000 description 1
- 230000001580 bacterial Effects 0.000 description 1
- 229960000626 benzylpenicillin Drugs 0.000 description 1
- 238000010364 biochemical engineering Methods 0.000 description 1
- 230000003851 biochemical process Effects 0.000 description 1
- 239000002551 biofuel Substances 0.000 description 1
- 230000031018 biological processes and functions Effects 0.000 description 1
- 238000010504 bond cleavage reaction Methods 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L cacl2 Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 235000012970 cakes Nutrition 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 159000000007 calcium salts Chemical class 0.000 description 1
- 239000004202 carbamide Substances 0.000 description 1
- CURLTUGMZLYLDI-UHFFFAOYSA-N carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 1
- 239000001569 carbon dioxide Substances 0.000 description 1
- 150000001732 carboxylic acid derivatives Chemical class 0.000 description 1
- 150000001733 carboxylic acid esters Chemical class 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 101700046922 cex Proteins 0.000 description 1
- 229960003405 ciprofloxacin Drugs 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- GFHNAMRJFCEERV-UHFFFAOYSA-L cobalt chloride hexahydrate Chemical compound O.O.O.O.O.O.[Cl-].[Cl-].[Co+2] GFHNAMRJFCEERV-UHFFFAOYSA-L 0.000 description 1
- PJQDFOMVKDFESH-UHFFFAOYSA-N cobalt(2+);N-(9H-fluoren-2-yl)-N-oxidoacetamide Chemical class [Co+2].C1=CC=C2C3=CC=C(N([O-])C(=O)C)C=C3CC2=C1.C1=CC=C2C3=CC=C(N([O-])C(=O)C)C=C3CC2=C1 PJQDFOMVKDFESH-UHFFFAOYSA-N 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 239000000356 contaminant Substances 0.000 description 1
- 238000010924 continuous production Methods 0.000 description 1
- 239000008120 corn starch Substances 0.000 description 1
- 229940099112 cornstarch Drugs 0.000 description 1
- 238000002425 crystallisation Methods 0.000 description 1
- 230000005712 crystallization Effects 0.000 description 1
- 239000012228 culture supernatant Substances 0.000 description 1
- 230000000593 degrading Effects 0.000 description 1
- 239000008121 dextrose Substances 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- GOMCKELMLXHYHH-UHFFFAOYSA-L dipotassium;phthalate Chemical compound [K+].[K+].[O-]C(=O)C1=CC=CC=C1C([O-])=O GOMCKELMLXHYHH-UHFFFAOYSA-L 0.000 description 1
- 150000002016 disaccharides Chemical class 0.000 description 1
- 238000004520 electroporation Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 150000002118 epoxides Chemical class 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- OTMSDBZUPAUEDD-UHFFFAOYSA-N ethane Chemical compound CC OTMSDBZUPAUEDD-UHFFFAOYSA-N 0.000 description 1
- VGGSQFUCUMXWEO-UHFFFAOYSA-N ethene Chemical compound C=C VGGSQFUCUMXWEO-UHFFFAOYSA-N 0.000 description 1
- 125000004494 ethyl ester group Chemical group 0.000 description 1
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000002360 explosive Substances 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- CWYNVVGOOAEACU-UHFFFAOYSA-N fe2+ Chemical compound [Fe+2] CWYNVVGOOAEACU-UHFFFAOYSA-N 0.000 description 1
- 239000010419 fine particle Substances 0.000 description 1
- 239000006260 foam Substances 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 239000002816 fuel additive Substances 0.000 description 1
- VZCYOOQTPOCHFL-UHFFFAOYSA-N fumaric acid Chemical compound OC(=O)C=CC(O)=O VZCYOOQTPOCHFL-UHFFFAOYSA-N 0.000 description 1
- 239000007789 gas Substances 0.000 description 1
- 239000003502 gasoline Substances 0.000 description 1
- 230000030279 gene silencing Effects 0.000 description 1
- 230000005017 genetic modification Effects 0.000 description 1
- 235000013617 genetically modified food Nutrition 0.000 description 1
- 235000013922 glutamic acid Nutrition 0.000 description 1
- 150000002307 glutamic acids Chemical class 0.000 description 1
- LYCAIKOWRPUZTN-UHFFFAOYSA-N glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 1
- 238000003621 hammer milling Methods 0.000 description 1
- 238000003306 harvesting Methods 0.000 description 1
- 239000001307 helium Substances 0.000 description 1
- SWQJXJOGLNCZEY-UHFFFAOYSA-N helium(0) Chemical compound [He] SWQJXJOGLNCZEY-UHFFFAOYSA-N 0.000 description 1
- 150000004688 heptahydrates Chemical class 0.000 description 1
- 229920000140 heteropolymer Polymers 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 150000002430 hydrocarbons Chemical class 0.000 description 1
- 125000004435 hydrogen atoms Chemical class [H]* 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- 230000003301 hydrolyzing Effects 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 239000002440 industrial waste Substances 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 238000003780 insertion Methods 0.000 description 1
- 238000010884 ion-beam technique Methods 0.000 description 1
- 239000002563 ionic surfactant Substances 0.000 description 1
- 159000000014 iron salts Chemical class 0.000 description 1
- VQTUBCCKSQIDNK-UHFFFAOYSA-N isobutene Chemical compound CC(C)=C VQTUBCCKSQIDNK-UHFFFAOYSA-N 0.000 description 1
- 239000000905 isomalt Substances 0.000 description 1
- 235000010439 isomalt Nutrition 0.000 description 1
- 229960000318 kanamycin Drugs 0.000 description 1
- 150000002576 ketones Chemical class 0.000 description 1
- 239000002655 kraft paper Substances 0.000 description 1
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 239000000832 lactitol Substances 0.000 description 1
- 235000010448 lactitol Nutrition 0.000 description 1
- 229960003451 lactitol Drugs 0.000 description 1
- 230000000670 limiting Effects 0.000 description 1
- 235000020778 linoleic acid Nutrition 0.000 description 1
- 238000001638 lipofection Methods 0.000 description 1
- 101710026800 lyc Proteins 0.000 description 1
- 159000000003 magnesium salts Chemical class 0.000 description 1
- WRUGWIBCXHJTDG-UHFFFAOYSA-L magnesium sulfate heptahydrate Chemical compound O.O.O.O.O.O.O.[Mg+2].[O-]S([O-])(=O)=O WRUGWIBCXHJTDG-UHFFFAOYSA-L 0.000 description 1
- 239000011976 maleic acid Substances 0.000 description 1
- 239000000845 maltitol Substances 0.000 description 1
- 235000010449 maltitol Nutrition 0.000 description 1
- 229940035436 maltitol Drugs 0.000 description 1
- 150000002696 manganese Chemical class 0.000 description 1
- ISPYRSDWRDQNSW-UHFFFAOYSA-L manganese(II) sulfate monohydrate Chemical compound O.[Mn+2].[O-]S([O-])(=O)=O ISPYRSDWRDQNSW-UHFFFAOYSA-L 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- CERQOIWHTDAKMF-UHFFFAOYSA-M methacrylate Chemical compound CC(=C)C([O-])=O CERQOIWHTDAKMF-UHFFFAOYSA-M 0.000 description 1
- 125000004492 methyl ester group Chemical group 0.000 description 1
- 238000000520 microinjection Methods 0.000 description 1
- 235000010755 mineral Nutrition 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 239000000178 monomer Substances 0.000 description 1
- 150000002772 monosaccharides Chemical class 0.000 description 1
- 230000003505 mutagenic Effects 0.000 description 1
- 239000003471 mutagenic agent Substances 0.000 description 1
- 231100000707 mutagenic chemical Toxicity 0.000 description 1
- 229960004927 neomycin Drugs 0.000 description 1
- PXHVJJICTQNCMI-UHFFFAOYSA-N nickel Chemical compound [Ni] PXHVJJICTQNCMI-UHFFFAOYSA-N 0.000 description 1
- 239000002736 nonionic surfactant Substances 0.000 description 1
- 235000006408 oxalic acid Nutrition 0.000 description 1
- CBENFWSGALASAD-UHFFFAOYSA-N ozone Chemical compound [O-][O+]=O CBENFWSGALASAD-UHFFFAOYSA-N 0.000 description 1
- 238000004806 packaging method and process Methods 0.000 description 1
- 230000036961 partial Effects 0.000 description 1
- OFBQJSOFQDEBGM-UHFFFAOYSA-N pentane Chemical compound CCCCC OFBQJSOFQDEBGM-UHFFFAOYSA-N 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 238000011056 performance test Methods 0.000 description 1
- 150000002978 peroxides Chemical class 0.000 description 1
- QROGIFZRVHSFLM-UHFFFAOYSA-N phenylpropene group Chemical group C1(=CC=CC=C1)C=CC QROGIFZRVHSFLM-UHFFFAOYSA-N 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 150000003013 phosphoric acid derivatives Chemical class 0.000 description 1
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 229920000346 polystyrene-polyisoprene block-polystyrene Polymers 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 159000000001 potassium salts Chemical class 0.000 description 1
- 229920001592 potato starch Polymers 0.000 description 1
- 229940116317 potato starch Drugs 0.000 description 1
- 235000012015 potatoes Nutrition 0.000 description 1
- 238000010248 power generation Methods 0.000 description 1
- 230000002335 preservative Effects 0.000 description 1
- 238000003825 pressing Methods 0.000 description 1
- ATUOYWHBWRKTHZ-UHFFFAOYSA-N propane Chemical compound CCC ATUOYWHBWRKTHZ-UHFFFAOYSA-N 0.000 description 1
- 239000001294 propane Substances 0.000 description 1
- 235000019260 propionic acid Nutrition 0.000 description 1
- 238000002731 protein assay Methods 0.000 description 1
- 238000010926 purge Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 230000000717 retained Effects 0.000 description 1
- 238000011012 sanitization Methods 0.000 description 1
- 238000005549 size reduction Methods 0.000 description 1
- 239000010802 sludge Substances 0.000 description 1
- 239000002002 slurry Substances 0.000 description 1
- 210000004215 spores Anatomy 0.000 description 1
- 238000001694 spray drying Methods 0.000 description 1
- 229910001220 stainless steel Inorganic materials 0.000 description 1
- 239000010935 stainless steel Substances 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
- 239000008117 stearic acid Substances 0.000 description 1
- 239000008223 sterile water Substances 0.000 description 1
- 230000001954 sterilising Effects 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 238000004326 stimulated echo acquisition mode for imaging Methods 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 239000004575 stone Substances 0.000 description 1
- 239000001384 succinic acid Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000001502 supplementation Effects 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 230000002194 synthesizing Effects 0.000 description 1
- 238000005979 thermal decomposition reaction Methods 0.000 description 1
- 229920001187 thermosetting polymer Polymers 0.000 description 1
- RTAQQCXQSZGOHL-UHFFFAOYSA-N titanium Chemical compound [Ti] RTAQQCXQSZGOHL-UHFFFAOYSA-N 0.000 description 1
- 229940005605 valeric acid Drugs 0.000 description 1
- 230000003612 virological Effects 0.000 description 1
- 238000004065 wastewater treatment Methods 0.000 description 1
- 125000000969 xylosyl group Chemical group C1([C@H](O)[C@@H](O)[C@H](O)CO1)* 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
- 150000003751 zinc Chemical class 0.000 description 1
- RZLVQBNCHSJZPX-UHFFFAOYSA-L zinc sulfate heptahydrate Chemical compound O.O.O.O.O.O.O.[Zn+2].[O-]S([O-])(=O)=O RZLVQBNCHSJZPX-UHFFFAOYSA-L 0.000 description 1
- SJZRECIVHVDYJC-UHFFFAOYSA-N γ-Hydroxybutyric acid Chemical compound OCCCC(O)=O SJZRECIVHVDYJC-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
- C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
- C12N9/2405—Glucanases
- C12N9/2434—Glucanases acting on beta-1,4-glucosidic bonds
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
- C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
- C12N9/2405—Glucanases
- C12N9/2434—Glucanases acting on beta-1,4-glucosidic bonds
- C12N9/2437—Cellulases (3.2.1.4; 3.2.1.74; 3.2.1.91; 3.2.1.150)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/02—Monosaccharides
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/04—Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/12—Disaccharides
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/14—Preparation of compounds containing saccharide radicals produced by the action of a carbohydrase (EC 3.2.x), e.g. by alpha-amylase, e.g. by cellulase, hemicellulase
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P2201/00—Pretreatment of cellulosic or lignocellulosic material for subsequent enzymatic treatment or hydrolysis
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P5/00—Preparation of hydrocarbons or halogenated hydrocarbons
- C12P5/02—Preparation of hydrocarbons or halogenated hydrocarbons acyclic
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P5/00—Preparation of hydrocarbons or halogenated hydrocarbons
- C12P5/02—Preparation of hydrocarbons or halogenated hydrocarbons acyclic
- C12P5/023—Methane
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
- C12P7/06—Ethanol, i.e. non-beverage
- C12P7/08—Ethanol, i.e. non-beverage produced as by-product or from waste or cellulosic material substrate
- C12P7/10—Ethanol, i.e. non-beverage produced as by-product or from waste or cellulosic material substrate substrate containing cellulosic material
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
- C12P7/16—Butanols
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/24—Preparation of oxygen-containing organic compounds containing a carbonyl group
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/24—Preparation of oxygen-containing organic compounds containing a carbonyl group
- C12P7/26—Ketones
- C12P7/28—Acetone-containing products
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
- C12P7/42—Hydroxy-carboxylic acids
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
- C12P7/44—Polycarboxylic acids
- C12P7/46—Dicarboxylic acids having four or less carbon atoms, e.g. fumaric acid, maleic acid
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
- C12P7/44—Polycarboxylic acids
- C12P7/48—Tricarboxylic acids, e.g. citric acid
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
- C12P7/52—Propionic acid; Butyric acids
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
- C12P7/56—Lactic acid
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/62—Carboxylic acid esters
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y302/00—Hydrolases acting on glycosyl compounds, i.e. glycosylases (3.2)
- C12Y302/01—Glycosidases, i.e. enzymes hydrolysing O- and S-glycosyl compounds (3.2.1)
- C12Y302/01004—Cellulase (3.2.1.4), i.e. endo-1,4-beta-glucanase
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/30—Fuel from waste, e.g. synthetic alcohol or diesel
Abstract
method comprising combining a cellulolytic microorganism with an inductant comprising a first quantity of a lignocellulosic material, treated with a dose of less than 40 Mrad of bombardment with electrons, and providing conditions that induce the cellulolytic microorganism to produce a cellulase complex tailored to saccharify the particular lignocellulosic material. The relative concentrations of the one or more enzymes is modulated by selection of the particular dose of bombardment with electrons. Subsequently a second quantity of the lignocellulosic material is saccharified with the enzymes, which has been treated with at least 40 Mrad of electron bombardment to reduce its recalcitrance. Wherein the first quantity of the lignocellulosic material has a greater portion of crystalline cellulose than the second quantity and wherein the cellulase complex exhibits enhanced saccharification of the second quantity of the lignocellulosic material attributable, at least in part, to endoglucanase derived from the first quantity of the lignocellulosic material having a greater portion of crystalline cellulose than the second quantity. omplex tailored to saccharify the particular lignocellulosic material. The relative concentrations of the one or more enzymes is modulated by selection of the particular dose of bombardment with electrons. Subsequently a second quantity of the lignocellulosic material is saccharified with the enzymes, which has been treated with at least 40 Mrad of electron bombardment to reduce its recalcitrance. Wherein the first quantity of the lignocellulosic material has a greater portion of crystalline cellulose than the second quantity and wherein the cellulase complex exhibits enhanced saccharification of the second quantity of the lignocellulosic material attributable, at least in part, to endoglucanase derived from the first quantity of the lignocellulosic material having a greater portion of crystalline cellulose than the second quantity.
Description
PROCESSING OF BIOMASS MATERIALS
by Marshall , Thomas Craig Masterman, Aiichiro Yoshida, Jennifer Moon Yee Fung,
James J. Lynch
CROSS REFERENCE TO RELATED APPLICATIONS
This application claims the benefit of US. Provisional Application Nos. 61/579,550
and 61/579,562, both filed on December 22, 2011. The entire disclosures of the above
applications are incorporated herein by reference.
FIELD OF THE INVENTION
The ion pertains to the ation enzymes useful in the processing of biomass
materials. For example, the invention relates to producing cellulase enzymes or other enzyme
types.
BACKGROUND
As demand for petroleum increases, so too does interest in renewable feedstocks for
manufacturing biofuels and biochemicals. The use of lignocellulosic biomass as a ock for
such cturing processes has been studied since the 1970s. Lignocellulosic biomass is
attractive because it is nt, renewable, domestically ed, and does not compete with
food industry uses.
Many potential lignocellulosic feedstocks are available today, including agricultural
residues, woody biomass, municipal waste, oilseeds/cakes and sea weeds, to name a few. At
present these materials are either used as animal feed, post materials, are burned in a
cogeneration facility or are landfilled.
Lignocellulosic biomass is recalcitrant to degradation as the plant cell walls have a
structure that is rigid and compact. The structure comprises crystalline cellulose fibrils
embedded in a hemicellulose matrix, surrounded by lignin. This compact matrix is difficult to
access by enzymes and other chemical, biochemical and biological processes. Cellulosic
biomass materials (e.g., s al from which substantially all the lignin has been
removed) can be more accessible to enzymes and other conversion processes, but even so,
naturally-occurring cellulosic materials often have low yields (relative to theoretical yields)
when contacted with hydrolyzing enzymes. Lignocellulosic biomass is even more recalcitrant to
enzyme attack. rmore, each type of ellulosic biomass has its own specific
composition of cellulose, llulose and lignin.
While a number of methods have been tried to extract structural carbohydrates from
lignocellulosic biomass, they are either are too expensive, produce too low a yield, leave
undesirable chemicals in the resulting product, or simply degrade the sugars.
Saccharides from renewable biomass s could become the basis of chemical and
fuels industries by replacing, supplementing or substituting petroleum and other fossil
feedstocks. However, techniques need to be developed that will make these monosaccharides
available in large quantities and at able es and prices.
SUMMARY OF THE INVENTION
Provided herein are methods of ng the production of one or more enzymes by a
microorganism, through the use of an inductant.
In one aspect, a method is provided that es combining a cellulosic or
lignocellulosic biomass, which has been treated to reduce its recalcitrance, with a
microorganism, to induce the production of one or more enzyme(s) by the microorganism by
maintaining the microorganism-biomass combination under conditions that allow for the
production of the enzyme(s) by the microorganism. In some implementations, the enzyme(s) are
then used to saccharify cellulosic or lignocellulosic s.
Also provided herein is a method for inducing the production of an enzyme by a
microorganism, where the method includes: providing a first cellulosic or lignocellulosic
biomass; ng the first s with a treatment method to reduce its itrance, thereby
producing a first treated biomass; providing a microorganism; providing a liquid medium;
combining the first treated biomass, the microorganism, and the liquid medium, thereby
producing a microorganism-biomass combination; and ining the microorganism-biomass
combination under conditions allowing for the production of an enzyme by the microorganism,
y producing an inductant-enzyme combination; thereby inducing the production of the
enzyme by the microorganism.
Also provided herein is a composition that includes a liquid medium, a
cellulosic or lignocellulosic biomass treated to reduce its recalcitrance, a
microorganism, and one or more enzymes made by the microorganism.
[0011A] Also provided herein is a method comprising:
selectively combining a cellulolytic microorganism with an inductant comprising a
first quantity of a particular lignocellulosic material, the first quantity of the particular
lignocellulosic material having been treated with a particular dose of bombardment
with electrons, the dose being selected from the range of less than 40 Mrad, and
providing conditions effective to induce the cellulolytic rganism to produce a
cellulase complex sing one or more enzymes having ve concentrations
tailored to saccharify the particular lignocellulosic material, the relative
concentrations of the one or more enzymes being modulated by selection of the
particular dose of dment with electrons; and
subsequently saccharifying a second quantity of the ular lignocellulosic al
with the one or more enzymes, wherein the second quantity of the particular
ellulosic material has been treated with at least 40 Mrad of electron
dment to reduce its recalcitrance;
wherein the first quantity of the ular lignocellulosic material has a greater
portion of lline ose than the second quantity, the dose of electron
bombardment applied to the first quantity being less than the dose of electron
bombardment applied to the second quantity; and
wherein the cellulase complex exhibits enhanced saccharification of the second
quantity of the particular lignocellulosic material attributable, at least in part, to
endoglucanase derived from the first quantity of the particular lignocellulosic material
having a greater portion of crystalline cellulose than the second quantity.
In any of the methods or compositions provided herein, the treatment for
ng the itrance of the biomass material(s) can be any of: bombardment
with electrons, tion, oxidation, pyrolysis, steam explosion, chemical treatment,
mechanical treatment, and freeze grinding. Preferably, the treatment method is
bombardment with electrons.
The methods and compositions can also include mechanically treating the
first or the second cellulosic or lignocellulosic biomass to reduce its bulk density
and/or increase its surface area. The biomass material(s) can be comminuted before
being combined with the microorganism and liquid medium. The comminution can
be dry milling or wet miling. The biomass material can have a particle size of about
to 1400 µm.
In any of the methods and compositions described herein, any of the
cellulosic or lignocellulosic biomasses can be: paper, paper ts, paper waste,
paper pulp, ted papers, loaded papers, coated papers, filled papers, magazines,
printed , r paper, polycoated paper, card stock, cardboard, paperboard,
cotton, wood, le board, forestry wastes, sawdust, aspen wood, wood chips,
grasses, switchgrass, miscanthus, cord grass, reed canary grass, grain residues, rice
hulls, oat hulls, wheat chaff, barley hulls, ltural waste, silage, canola straw,
wheat straw, barley straw, oat straw, rice straw, jute, hemp, flax, bamboo, sisal, abaca,
corn cobs, corn stover, soybean stover, corn fiber, alfalfa, hay, coconut hair, sugar
processing residues, bagasse, beet pulp, agave bagasse, algae, seaweed, manure,
sewage, offal, agricultural or industrial waste, arracacha, buckwheat, banana, barley,
cassava, kudzu, oca, sago, sorghum, potato, sweet potato, taro, yams, beans, favas,
lentils, peas, or mixtures of any of these. Alternatively, the cellulosic or
lignocellulosic s can include material that was ing after a prior
cellulosic or lignocellulosic biomass was previously ted to a product by an
enzyme of a microorganism.
In these methods and compositions, the microorganism can be any of a fungus,
a bacterium, or a yeast. The microorganism can actually be a population of different
microorganisms. The microorganism can be a strain that produces high levels of
ase, and/or it can be genetically engineered. The microorganism can be
Trichoderma reesei, or it can
be Clostrz'dz'um thermocellum, for example. The microorganism can be a T. reesez' strain such as
14, PC3—7, QM9414 or RUT-C30.
In any of these methods and compositions, the cellulosic or lignocellulosic biomass
can be combined with the microorganism at a time when the microorganism is in lag phase.
The s and compositions can also include removing all or a portion of the
liquid from the microorganism-inductant-enzyme combination, to produce an enzyme extract.
The methods and compositions can also include concentrating one or more of the enzymes,
and/or isolating one or more of the enzymes.
The methods and compositions can also include allowing saccharification of the
second osic or lignocellulosic biomass to occur, so that one or more sugars are produced.
The one or more sugars can be isolated and/or trated.
It should be understood that this invention is not d to the embodiments
disclosed in this Summary, and it is intended to cover ations that are within the spirit and
scope of the invention, as defined by the claims.
BRIEF DESCRIPTION OF THE DRAWINGS
The foregoing will be apparent from the following more particular description of
example embodiments of the invention, as illustrated in the accompanying drawings in which
like reference characters refer to the same parts throughout the different views. The drawings are
not necessarily to scale, emphasis instead being placed upon illustrating embodiments of the
present invention.
is a m illustrating the enzymatic hydrolysis of cellulose to glucose.
Cellulosic ate (A) is converted by endocellulase (i) to cellulose (B), which is converted by
exocellulase (ii) to iose (C), which is ted to glucose (D) by cellobiase (beta-
glucosidase) (iii).
is a flow diagram illustrating conversion of a biomass feedstock to one or
more products. Feedstock is physically ated (e.g, to reduce its size) (200), optionally
treated to reduce its recalcitrance (210), rified to form a sugar solution (220), the solution
is transported (230) to a manufacturing plant (e.g., by pipeline, railcar) (or if saccharification is
performed en route, the feedstock, enzyme and water is transported), the saccharified feedstock
is bio-processed to produce a desired product (e.g., alcohol) (240), and the product can be
processed further, e.g., by distillation, to produce a final product (250). Treatment for
recalcitrance can be modified by measuring lignin content (201) and setting or adjusting process
parameters (205). Saccharifying the feedstock (220) can be modified by mixing the feedstock
with medium and the enzyme (221).
is a flow m illustrating the treatment of a first biomass (300), on of
a cellulase producing organism (310), addition of a second biomass (320), and processing the
resulting sugars to make products (e.g, alcohol(s), pure sugars) (330). The first treated s
can optionally be split, and a portion added as the second biomass (A).
is a flow diagram illustrating the tion of enzymes. A cellulase-
producing organism is added to growth medium (400), a treated first biomass (405) is added (A)
to make a mixture (410), a second biomass is added (420), and the resulting sugars are processed
to make products (e.g., alcohol(s), pure ) (430). Portions of the first biomass (405) can
also be added (B) to the second biomass (420).
shows results of protein is using SDS PAGE.
DETAILED DESCRIPTION
Provided herein are methods of inducing the production of one or more enzymes by a
microorganism, through the use of an inductant. The inductant is made from biomass (cellulosic
or lignocellulosic) that has been treated to reduce its recalcitrance. The treatment method can
include subjecting the biomass to bombardment with electrons, sonication, oxidation, sis,
steam explosion, chemical treatment, mechanical treatment, or freeze grinding. As disclosed
herein, biomass that has been treated with such a method can be ed with a microorganism
in a medium (such as a liquid medium), to induce the microorganism to produce one or more
In one aspect, the invention features a method that includes contacting an inducer
comprising a lignocellulosic material with a microorganism to produce an enzyme.
Specifically, the processes described herein include saccharifying cellulosic and/or
lignocellulosic materials using enzymes that have been produced by Trichoderma ’ fungi,
as will be discussed in further detail below.
In general, the invention relates to improvements in sing s als
(e.g, biomass als or biomass-derived materials) to produce intermediates and products,
such as fuels and/or other products. For example, the processes may be used to produce sugars,
alcohols (such as ethanol, isobutanol, or n-butanol), sugar alcohols (such as itol), or
c acids.
The invention also relates to the preparation of enzymes useful in the processing of
biomass materials. For example the ion relates to ing cellulase enzymes or other
enzyme types.
A typical biomass resource contains ose, hemicellulose, and lignin plus lesser
amounts of ns, extractables and minerals. The complex carbohydrates contained in the
cellulose and hemicellulose fractions can be converted into sugars, e.g., fermentable sugars, by
saccharification, and the sugars can then be used as an end product or an intermediate, or
converted by fiarther sing, e.g., fermentation or hydrogenation, into a variety of products,
such as alcohols or c acids. The product obtained depends upon the method or
microorganism utilized and the conditions under which the bioprocessing occurs.
In one embodiment, for instance, the invention es a method of inducing the
production of an enzyme. A osic or lignocellulosic s is provided, treated to reduce
its recalcitrance, and then ed with a microorganism in a liquid medium. The resulting
microorganism—biomass combination is then maintained under conditions allowing for the
grth ofthe organism and production of enzymes capable of degrading the biomass. The
treated biomass acts as an inductant, causing the rganism to produce enzymes. The
method produces an inductant-enzyme combination.
Without wishing to be bound by any theory, it is believed that the treatment used to
reduce the recalcitrance of the biomass is important in enzyme induction. The inventors have
found that low levels of treatment result in either low levels of enzyme induction, or extremely
long lag times, presumably because it is difficult for the microorganisms to extract sugars from
the treated biomass material. rly, very high levels of treatment also cause the
microorganisms to produce low levels of s, possibly because relatively easy extraction of
sugars from the treated biomass lessens the need for the microorganisms to produce large
amounts of enzymes.
On a related matter, the recalcitrance treatment also serves to sterilize the material.
Biomass material, by its nature, contains contaminating microbes, which are often embedded
deep within the al itself. Because the enzyme inductions as disclosed herein tend to be
long fermentations (up to a week or more), sterilization is important. It would therefore be
advantageous to treat the material as heavily as possible to ize it. However, such high levels
of treatment would likely be counterproductive because high levels of treatment lessen the
enzyme production by the microorganisms.
As sed herein, there is therefore a large benefit to be gained from carefully
balancing the level of treatment to sterilize the material, yet not over-treat the al so that the
microorganisms fail to produce large amounts of enzyme.
The term “inductant,” as used herein, means a osic or lignocellulosic biomass
that encourages an organism to produce enzyme. An example would be biomass that has been
treated to reduce its recalcitrance. The treated biomass is then used as an enzyme inductant, by
being combined with one or more microorganisms in a liquid medium, and then being
maintained under conditions that allow the microorganism to produce one or more enzymes.
The inductant-enzyme combination can then be combined with another biomass, and
used to saccharify it.
Surprisingly, it has been found that treating the biomass before inoculating it with the
microorganism causes an increased amount of s to be produced by the microbes. In
addition, different enzymes are ed on the treated biomass, ve to the use of untreated
biomass.
As described herein, the cellulosic or lignocellulosic biomass can be d from a
wide variety of materials. In one embodiment, the biomass can be lignin hulls. By “lignin
hulls,” as used herein, is meant material that is remaining after a biomass has been saccharified.
In certain ments, the invention relates to processes for saccharifying a
cellulosic or lignocellulosic material using an enzyme that has been produced by a , e. g.,
by strains of the cellulolytic filamentous fungus Trichoderma reesez‘. In some implementations,
high-yielding cellulase mutants of Trichoderma read are used, e.g., RUT-NG14, PC3-7,
QM9414 and/or RUT-C30. Such s are described, for example, in “Selective Screening
Methods for the Isolation of High Yielding Cellulase Mutants of Trichoderma reesei,”
Montenecourt, BS. and igh, D.E. Adv. Chem. Ser. 181, 1 (1979). These mutants
are hyperproducing and are catabolite repression-resistant, allowing high yields of cellulases to
be achieved.
In preferred embodiments, the enzyme production is conducted in the presence of a
portion of the lignocellulosic material to be saccharified. The lignocellulosic material can act in
the enzyme production process as an inducer for cellulase synthesis, producing a cellulase
complex having an activity that is tailored to the particular lignocellulosic material. In some
implementations, the recalcitrance of the lignocellulosic material is reduced prior to using it as
an inducer. It is believed that this makes the cellulose within the ellulosic al more
readily available to the fungus. Reducing the recalcitrance of the lignocellulosic material also
facilitates saccharification.
In some cases, reducing the recalcitrance of the lignocellulosic material includes
treating the lignocellulosic material with a al treatment. The physical treatment can be, for
example, radiation, e.g., electron bombardment, sonication, pyrolysis, oxidation, steam
explosion, chemical treatment, or ations of any of these treatments. The treatments can
also include any one or more of the treatments sed herein, applied alone or in any desired
combination, and applied once or multiple times.
Enzymes and s—destroying organisms that break down biomass, such as the
cellulose and/or the lignin portions of the biomass, contain or manufacture s cellulolytic
enzymes (cellulases), ligninases or various small molecule biomass-destroying metabolites.
These enzymes may be a x of enzymes that act synergistically to degrade crystalline
cellulose or the lignin portions of biomass. Examples of cellulolytic enzymes include:
ucanases, cellobiohydrolases, and cellobiases (beta-glucosidases).
As shown in for example, during saccharification a cellulosic substrate (A) is
initially hydrolyzed by endoglucanases (i) at random locations producing oligomeric
intermediates (e.g., cellulose) (B). These intermediates are then substrates for exo-splitting
ases (ii) such as cellobiohydrolase to produce cellobiose from the ends of the cellulose
polymer. Cellobiose is a soluble 1,4-linked dimer of glucose. y cellobiase (iii)
cleaves cellobiose (C) to yield glucose (D). Therefore, the endoglucanases are particularly
effective in ing the lline portions of cellulose and increasing the effectiveness of
exocellulases to produce cellobiose, which then es the specificity of the cellobiose to
produce glucose. Therefore, it is evident that depending on the nature and structure of the
cellulosic substrate, the amount and type of the three different enzymes may need to be modified.
The s produced and used in the ses described herein can be produced
by a fiingus, e.g, by one or more strains of the fungus Trichoderma reesez‘. In red
implementations, high—yielding cellulase mutants of Trichoderma reesei, e.g, RUT—NG14, PC3-
7, QM9414 and/or RUT-C30, are used.
It is preferred that enzyme production be conducted in the presence of a portion of the
feedstock that will be rified, thereby producing a cellulase complex that is tailored to the
ular feedstock. The feedstock may be treated prior to such use to reduce its recalcitrance,
e.g. one or more of the recalcitrance-reducing ses described herein, so as to make
, using
the cellulose in the feedstock more y available to the fungus.
In a preferred embodiment the enzyme-inducing biomass can be treated by electron
dment. The biomass can be treated, for instance, by electron bombardment with a total
dose of less than about 1 Mrad, less than about 2 Mrad, than about 5, about 10, about
, less 20,
about 50, about 100 or about 150 Mrad. Preferably, the enzyme-inducing biomass is treated with
a total dose of about 0.1 Mrad to about 150 Mrad, about 1 to about 100 Mrad, preferably about 2
to about 50 Mrad, or about 5 to about 40 Mrad.
As will be discussed further below, once the enzyme has been produced, it is used to
saccharify the ing feedstock that has not been used to produce the enzyme. The process
for converting the feedstock to a desired product or intermediate generally includes other steps in
addition to this saccharification step.
For example, referring to a process for manufacturing an alcohol can include,
for example, optionally mechanically treating a ock, e.g. to reduce its size (200), before
and/or after this treatment, optionally treating the feedstock with another physical treatment to
further reduce its recalcitrance (210), then saccharifying the feedstock, using the enzyme
complex, to form a sugar solution (220). ally, the method may also include transporting,
e.g. truck or barge, the solution (or the feedstock, enzyme and water, if
, by pipeline, railcar,
saccharification is performed en route) to a manufacturing plant (230). In some cases the
rified feedstock is r bioprocessed (e.g., fermented) to produce a desired product e.g.,
alcohol (240). This resulting product may in some implementations be processed further, e.g,
by distillation (250), to produce a final product. One method of reducing the recalcitrance of the
feedstock is by electron bombardment of the feedstock. If desired, the steps of measuring lignin
content of the feedstock (201) and setting or adjusting process parameters based on this
2012/071091
measurement (205) can be performed at various stages of the process, as described in US. Pat.
App. Pub. 2010/0203495 A1 by Medoff and Masterman, published August 12, 2010, the
complete disclosure of which is incorporated herein by reference. Saccharifying the feedstock
(220) can also be modified by mixing the feedstock with medium and the enzyme (221).
The manufacture of the enzyme complexes will now be described first, followed by a
description of the method steps discussed above with reference to and the materials used
in the process.
A number of different conditions were tested, and the results are as follows. In one
embodiment, the enzyme induction biomass is corn cob. In this embodiment, the s is
treated by electron bombardment with a 35 Mrad on beam. Preferably, the biomass is
comminuted to a particle size of 0 um, more preferably less than 200 um, most preferably
less than 50 um. The treated biomass (in either wet or dry form) is added in a total amount of
about 25 to about 133 g/L of inoculated medium, more preferably 100 g/L. The inductant
biomass can be added at any point in the growth of the microorganisms up through the third day
after inoculation, but is ably added 1-3 days after inoculation. The total amount of biomass
to be added as an inductant can be added all at once, or in aliquots, for instance, in two parts, or
in five parts. Preferably the corncob biomass is added all at once.
The enzyme induction biomass can be presented to the rganisms as a solid, or
as a slurry. Preferably it is added as a .
ENZYME PRODUCTION
Filamentous fungi, or bacteria that produce cellulase, lly require a carbon
source and an inducer for production of cellulase. Without being bound by any theory, it is
believed that the enzymes of this disclosure are ularly suited for saccharification of the
substrate used for inducing its production.
Lignocellulosic materials comprise ent combinations of ose, hemicellulose
and lignin. Cellulose is a linear polymer of e forming a fairly stiff linear structure without
cant coiling. Due to this structure and the disposition of hydroxyl groups that can
hydrogen bond, cellulose contains crystalline and non-crystalline portions. The crystalline
portions can also be of different types, noted as I(alpha) and I(beta) for example, depending on
the location of hydrogen bonds between strands. The polymer lengths themselves can vary
lending more y to the form of the ose. Hemicellulose is any of several
heteropolymers, such as xylan, glucuronoxylan, arabinoxylans, and xyloglucan. The primary
sugar monomer present is xylose, although other monomers such as mannose, galactose,
rhamnose, arabinose and glucose are present. Typically hemicellulose forms branched structures
with lower molecular weights than cellulose. Hemicellulose is therefore an amorphous material
that is generally susceptible to enzymatic hydrolysis. Lignin is a complex high molecular weight
polymer generally. Although all lignins show variation in their composition, they have
been described as an amorphous dendritic network polymer of phenyl propene units. The
amounts of cellulose, hemicellulose and lignin in a specific biomaterial depends on the source of
the biomaterial. For example wood derived biomaterial can be about 38-49% cellulose, 7—26%
hemicellulose and 23-34% lignin depending on the type. Grasses typically are 33-38% cellulose,
24-32% hemicellulose and 17-22% lignin. Clearly ellulosic biomass constitutes a large
class of substrates.
The diversity of biomass materials may be further increased by pretreatment, for
example, by changing the crystallinity and molecular s of the polymers.
The cellulase producing organism when contacted with a s will tend to
produce enzymes that release molecules advantageous to the organism’s growth, such as glucose.
This is done through the phenomenon of enzyme induction as described above. Since there are a
variety of substrates in a particular biomaterial, there are a y of cellulases, for example, the
endoglucanase, exoglucanase and cellobiase discussed usly. By selecting a particular
lignocellulosic material as the inducer the relative concentrations and/or activities of these
enzymes can be modulated so that the ing enzyme complex will work efficiently on the
lignocellulosic material used as the r or a similar material. For example, a biomaterial
with a higher n of crystalline cellulose may induce a more effective or higher amount of
endoglucanase than a biomaterial with little crystalline cellulose.
Therefore, there may be many methods for optimal formation and use of ases.
Some s of these ses will be described with reference to the .
For example, referring to a first biomass is optionally pre-treated (300), for
example to reduce its recalcitrance, and is then mixed with an aqueous medium and a cellulase
producing organism (310). After an adequate time has passed for the cells to grow to a desired
stage and enough s have been produced, a second biomass is added (320). The action of
the enzyme on the second and any remaining first biomass produces a mix of sugars, which can
be further processed to useful products (e.g., alcohols, pure ) (330). The first and second
biomass can be portions of the same biomass source material. For example, a portion of the
biomass can be combined with the cellulase producing organism and then another portion added
at a later stage (A) once some of the enzymes have been produced. Optionally, the first and
second biomass may both be ated to reduce recalcitrance. The aqueous media will be
discussed below.
ing now to the cellulase producing organism (400) can be grown in a
growth medium for a time to reach a specific growth phase. For example, this growth period
could extend over a period of days or even weeks. Pretreated first biomass (405) can then be
contacted (A) with the enzyme producing cells (410) so that after a time enzymes are produced.
Enzyme production may also take place over an extended period of time. The enzyme
containing solution is then combined with a second biomass (420). The action of the enzyme on
the second and remaining first biomass produces mixed sugars which can be further processed to
useful products (430). The first and second s can be portions of the same biomass or
could be r but not identical (e.g., ated and non-pretreated) material (B).
In addition to the s discussed above in the cellulase producing
organism (400) may optionally be harvested prior to being combined with the first pretreated
biomass (410). Harvesting may include partial or almost complete removal of the solvent and
growth media components. For example the cells may be collected by centrifugation and then
washed with water or another solution.
In another embodiment, after the enzyme(s) is produced (410), it can be concentrated
(e.g, between steps 410 and 420 of . Concentration may be by any useful method
including chromatography, centrifugation, filtration, is, extraction, evaporation of solvents,
spray drying and adsorption onto a solid support. The concentrated enzyme can be stored for a
time and then be used by on of a second biomass (420) and tion of useful products
(430).
The s media used in the above described methods can contain added yeast
extract, corn steep, peptones, amino acids, ammonium salts, phosphate salts, potassium salts,
magnesium salts, calcium salts, iron salts, manganese salts, zinc salts and cobalt salts. In
addition to these components, the growth media typically contains 0 to 10% glucose (e.g., l to
2012/071091
% glucose) as a carbon source. Additionally the inducer media can n, in addition to the
biomass discussed previously, other inducers. For example, some known inducers are lactose,
pure cellulose and sophorose. Various components can be added and removed during the
processing to optimize the desired tion of useful products.
The concentration of the biomass typically used for inducing enzyme production is
r than or equal to 0.1 wt.% and less than or equal to 50 wt.%, greater than or equal to 0.5
wt.% and less than or equal to 25 wt.%, greater than or equal to 1 wt. %, and less than or equal to
wt.%, and greater than or equal to 1 wt.% and less than or equal to 10 wt. %.
Any of the processes described above may be performed as a batch, a fed-batch or a
continuous process. The processes are useful for ally industrial scale tion, 6.g.
having a culture medium of at least 50 liters, preferably at least 100 liters, more preferably at
least 500 liters, even more preferably at least 1,000 liters, in particular at least 5,000 liters
0 liters or 500,000 liters. The process may be carried out aerobically or anaerobically.
Some enzymes are produced by submerged cultivation and some by e cultivation.
In any of the process described above, the enzyme can be manufactured and stored
and then used to saccharify at a later date and/or different location.
Any of the processes described above may be conducted with agitation. In some
cases, agitation may be performed using jet mixing as described in US. Pat. App. Pub.
2010/0297705 A1 by Medoff and Masterman, published November 25, 2010, US. Pat. App.
Pub. 2012/0091035 A1 to Medoff and Masterman, published April 19, 2012, and US. Pat. App.
Pub. 100572 A1 by Medoff and Masterman, published April 26, 2012, the full disclosures
of which are incorporated by reference herein.
Temperatures for the growth of enzyme producing organisms are chosen to enhance
organism growth. For example for Trichoderma reesei the optimal ature is generally
between 20 and 40°C (e.g., 30°C), The temperature for enzyme tion is optimized for that
part of the process. For example for Trichoderma reesez’ the optimal ature for enzyme
production is between 20 and 40°C (e.g., 27°C).
FEEDSTOCK, BIOMASS MATERIALS, AND/OR INDUCERS
The feedstock, which may also be the inducer for enzyme production, is preferably a
lignocellulosic material, although the processes described herein may also be used with
osic materials, e.g., paper, paper products, paper pulp, cotton, and mixtures of any of these,
and other types of biomass. The processes described herein are particularly useful with
lignocellulosic materials, because these processes are particularly effective in reducing the
recalcitrance of lignocellulosic materials and allowing such materials to be processed into
ts and intermediates in an economically viable manner.
As used herein, the term “biomass materials” includes lignocellulosic, osic,
y, and microbial materials.
Preferably the enzyme—inducing biomass materials are agricultural waste such as corn
cobs, more preferably corn stover. Most preferably, the enzyme-inducing biomass material
comprises grasses.
Lignocellulosic materials include, but are not limited to, wood, particle board,
forestry wastes (e.g., sawdust, aspen wood, wood chips), grasses, (e.g., switchgrass, miscanthus,
cord grass, reed canary grass), grain residues, (e.g, rice hulls, oat hulls, wheat chaff, barley
hulls), agricultural waste (e.g., silage, canola straw, wheat straw, barley straw, oat straw, rice
straw, jute, hemp, flax, , sisal, abaca, corn cobs, corn stover, soybean stover, corn fiber,
alfalfa, hay, t hair), sugar processing residues (e.g., bagasse, beet pulp, agave bagasse),
algae, seaweed, manure, sewage, and es of any of these.
In some cases, the lignocellulosic material includes comcobs. Ground or
hammermilled comcobs can be spread in a layer of relatively uniform thickness for irradiation,
and after irradiation are easy to disperse in the medium for further processing. To facilitate
t and collection, in some cases the entire corn plant is used, ing the corn stalk, corn
kernels, and in some cases even the root system of the plant.
Advantageously, no additional nutrients (other than a nitrogen , e.g, urea or
a) are required during fermentation of comcobs or cellulosic or lignocellulosic materials
containing significant amounts of comcobs.
Corncobs, before and after comminution, are also easier to convey and se, and
have a lesser tendency to form explosive mixtures in air than other cellulosic or lignocellulosic
materials such as hay and grasses.
Cellulosic materials include, for example, paper, paper products, paper waste, paper
pulp, pigmented papers, loaded papers, coated papers, filled papers, magazines, printed matter
(e.g, books, catalogs, manuals, labels, calendars, ng cards, brochures, prospectuses,
newsprint), printer paper, polycoated paper, card stock, cardboard, paperboard, materials having
a high alpha-cellulose content such as cotton, and mixtures of any of these. For example paper
products as described in US. App. No. 13/396,365 (“Magazine ocks” by Medoff et al.,
filed February 14, 2012), the fill disclosure of which is incorporated herein by nce.
Cellulosic materials can also e ellulosic materials which have been de—
lignified.
Starchy materials include starch itself, e.g., corn starch, wheat starch, potato starch or
rice starch, a derivative of starch, or a material that includes starch, such as an edible food
product or a crop. For example, the starchy material can be arracacha, buckwheat, ,
barley, cassava, kudzu, oca, sago, sorghum, regular household potatoes, sweet potato, taro, yams,
or one or more beans, such as favas, lentils or peas. Blends of any two or more starchy materials
are also starchy materials. Mixtures of starchy, cellulosic and or lignocellulosic materials can
also be used. For example, a biomass can be an entire plant, a part of a plant or different parts of
a plant, e. g., a wheat plant, cotton plant, a corn plant, rice plant or a tree. The y materials
can be d by any of the methods described herein.
Microbial materials include, but are not limited to, any lly occurring or
cally modified microorganism or organism that contains or is capable of providing a
source of carbohydrates (e.g, cellulose), for example, protists, e.g., animal protists (e.g,
protozoa such as flagellates, amoeboids, ciliates, and oa) and plant protists (e.g., algae
such alveolates, chlorarachniophytes, cryptomonads, euglenids, phytes, haptophytes, red
algae, stramenopiles, and viridaeplantae). Other examples include seaweed, plankton (e.g.,
macroplankton, mesoplankton, microplankton, nanoplankton, picoplankton, and
femptoplankton), phytoplankton, bacteria (e.g., gram positive ia, gram negative bacteria,
and extremophiles), yeast and/or es of these. In some instances, microbial biomass can be
obtained from natural sources, e.g., the ocean, lakes, bodies of water, e.g., salt water or fresh
water, or on land. Alternatively or in addition, microbial biomass can be obtained from culture
systems, e.g, large scale dry and wet culture and fermentation systems.
The biomass material can also include offal, and similar sources of material.
In other embodiments, the biomass materials, such as cellulosic, starchy and
lignocellulosic feedstock materials, can be obtained from transgenic rganisms and plants
that have been modified with respect to a wild type variety. Such modifications may be, for
example, through the iterative steps of selection and breeding to obtain d traits in a plant.
Furthermore, the plants can have had genetic material removed, modified, silenced and/or added
with respect to the wild type variety. For e, cally d plants can be produced
by recombinant DNA methods, where genetic modifications include introducing or modifying
specific genes from parental varieties, or, for example, by using transgenic breeding wherein a
c gene or genes are introduced to a plant from a ent species of plant and/or bacteria.
Another way to create genetic ion is through on ng wherein new alleles are
artificially created from endogenous genes. The artificial genes can be created by a variety of
ways including treating the plant or seeds with, for example, al mutagens (e.g., using
alkylating agents, epoxides, alkaloids, peroxides, formaldehyde), irradiation (e.g., X-rays,
gamma rays, neutrons, beta particles, alpha particles, protons, deuterons, UV radiation) and
temperature shocking or other al stressing and subsequent selection techniques. Other
methods of providing modified genes is through error prone PCR and DNA shuffling followed
by insertion of the desired modified DNA into the desired plant or seed. Methods of introducing
the desired genetic variation in the seed or plant include, for example, the use of a bacterial
carrier, biolistics, m phosphate precipitation, electroporation, gene splicing, gene silencing,
lipofection, microinjection and viral carriers. Additional genetically modified materials have
been described in US. Application Serial No 13/396,369 filed February 14, 2012 the full
disclosure of which is incorporated herein by reference.
Any of the methods described herein can be practiced with mixtures of any s
materials described herein.
BIOMASS -- MECHANICAL ATION
Mechanical treatments of the feedstock may include, for example, cutting, milling,
e.g., hammermilling, wet milling, grinding, pressing, shearing or chopping. The initial
mechanical ent step may, in some implementations, include reducing the size of the
feedstock. In some cases, loose feedstock (e.g, recycled paper or switchgrass) is initially
prepared by cutting, shearing and/or shredding.
In addition to this size reduction, which can be performed initially and/or later during
processing, mechanical ent can also be advantageous for “opening up, ,3 CEstressing,”
breaking or ring the feedstock materials, making the cellulose of the materials more
tible to chain scission and/or disruption of crystalline structure during the structural
modification treatment.
Methods of mechanically treating the feedstock include, for example, g or
grinding. Milling may be performed using, for example, a hammer mill, ball mill, d mill,
conical or cone mill, disk mill, edge mill, Wiley mill or grist mill. ng may be performed
using, for example, a g/impact type grinder. Specific examples of grinders include stone
grinders, pin grinders, coffee grinders, and burr rs. Grinding or milling may be ed,
for example, by a reciprocating pin or other element, as is the case in a pin mill. Other
mechanical treatment methods include mechanical ripping or tearing, other s that apply
pressure to the fibers, and air attrition milling. Suitable mechanical treatments further include
any other technique that continues the disruption of the internal structure of the material that was
initiated by the previous processing steps.
Mechanical treatments that may be used, and the characteristics of the mechanically
treated feedstocks, are described in further detail in US. Serial No. 13/276,192, filed r 18,
2011, and published on April 26, 2012 as US. Pat. App. Pub. 2012/0100577 Al, the fiill
disclosure of which is hereby incorporated herein by reference.
BIOMASS TREATMENT -- ELECTRON DMENT
In some cases, the feedstock may be treated with electron bombardment to modify its
structure and thereby reduce its recalcitrance. Such treatment may, for example, reduce the
average molecular weight of the feedstock, change the crystalline structure of the feedstock,
and/or increase the surface area and/or porosity of the feedstock.
Electron bombardment Via an electron beam is generally preferred, e it
provides very high throughput and because the use of a relatively low voltage/high power
electron beam device eliminates the need for expensive concrete vault shielding, as such s
are “self-shielded” and e a safe, efficient process. While the “self-shielded” devices do
include ing (e.g., metal plate shielding), they do not require the uction of a concrete
vault, greatly reducing capital expenditure and often allowing an existing manufacturing facility
to be used without expensive modification. Electron beam accelerators are available, for
example, from IBA (Ion Beam Applications, Louvain-la-Neuve, Belgium), Titan Corporation
(San Diego, California, USA), and NHV Corporation (Nippon High Voltage, Japan).
2012/071091
Electron bombardment may be performed using an electron beam device that has a
nominal energy of less than 10 MeV, e.g, less than 7 MeV, less than 5 MeV, or less than 2 MeV,
e.g., from about 0.5 to 1.5 MeV, from about 0.8 to 1.8 MeV, from about 0.7 to 1 MeV, or from
about 1 to about 3 MeV. In some entations the nominal energy is about 500 to 800 keV.
The electron beam may have a relatively high total beam power (the combined beam
power of all accelerating heads, or, if multiple accelerators are used, of all accelerators and all
heads), e.g., at least 25 kW, e.g., at least 30, 40, 50, 60, 65, 70, 80, 100, 125, or 150 kW. In
some cases, the power is even as high as 500 kW, 750 kW, or even 1000 kW or more. In some
cases the electron beam has a beam power of 1200 kW or more.
This high total beam power is usually achieved by utilizing multiple accelerating
heads. For example, the electron beam device may include two, four, or more accelerating
heads. The use of multiple heads, each of which has a relatively low beam power, prevents
excessive temperature rise in the al, thereby preventing burning of the al, and also
increases the mity of the dose through the thickness of the layer of material.
In some entations, it is desirable to cool the material during electron
bombardment. For e, the material can be cooled while it is being conveyed, for example
by a screw extruder or other conveying equipment.
To reduce the energy required by the recalcitrance-reducing process, it is desirable to
treat the material as quickly as possible. In general, it is preferred that treatment be performed at
a dose rate of greater than about 0.25 Mrad per second, e.g, greater than about 0.5, 0.75, 1, 1.5,
2, 5, 7, 10, 12, 15, or even greater than about 20 Mrad per second, e.g., about 0.25 to 2 Mrad per
second. Higher dose rates generally require higher line speeds, to avoid thermal decomposition
of the material. In one implementation, the accelerator is set for 3 MeV, 50 mAmp beam
t, and the line speed is 24 feet/minute, for a sample thickness of about 20 mm (e.g.,
comminuted corn cob material with a bulk density of 0.5 g/cm3).
In some embodiments, electron bombardment is performed until the material receives
a total dose of at least 0.5 Mrad, e.g., at least 5, 10, 20, 30 or at least 40 Mrad. In some
embodiments, the treatment is performed until the material receives a dose of from about 0.5
Mrad to about 150 Mrad, about 1 Mrad to about 100 Mrad, about 2 Mrad to about 75 Mrad, 10
Mrad to about 50 Mrad, e.g, about 5 Mrad to about 50 Mrad, from about 20 Mrad to about 40
Mrad, about 10 Mrad to about 35 Mrad, or from about 25 Mrad to about 30 Mrad. In some
2012/071091
entations, a total dose of 25 to 35 Mrad is preferred, applied ideally over a couple of
seconds, e.g, at 5 Mrad/pass with each pass being applied for about one second. ng a
dose of greater than 7 to 8 Mrad/pass can in some cases cause thermal degradation of the
feedstock material.
Using le heads as discussed above, the material can be treated in multiple
passes, for e, two passes at 10 to 20 Mrad/pass, e.g., 12 to 18 Mrad/pass, separated by a
few seconds of cool-down, or three passes of 7 to 12 Mrad/pass, e.g., 9 to 11 Mrad/pass. As
discussed above, treating the material with several relatively low doses, rather than one high
dose, tends to prevent overheating of the material and also increases dose uniformity through the
thickness of the material. In some implementations, the al is stirred or otherwise mixed
during or after each pass and then ed into a uniform layer again before the next pass, to
r enhance treatment uniformity.
In some embodiments, electrons are accelerated to, for example, a speed of greater
than 75 t of the speed of light, e.g, greater than 85, 90, 95, or 99 percent of the speed of
light.
In some embodiments, any processing described herein occurs on lignocellulosic
material that remains dry as acquired or that has been dried, e.g. heat and/or reduced
, using
pressure. For e, in some embodiments, the cellulosic and/or lignocellulosic material has
less than about five percent by weight retained water, measured at 25°C and at fifty percent
relative humidity.
Electron bombardment can be applied while the cellulosic and/or lignocellulosic
material is exposed to air, oxygen—enriched air, or even oxygen itself, or blanketed by an inert
gas such as en, argon, or helium. When maximum oxidation is desired, an oxidizing
environment is utilized, such as air or oxygen and the distance from the beam source is
optimized to maximize reactive gas formation, e.g., ozone and/or oxides of nitrogen.
BIOMASS TREATMENT -— SONICATION, SIS, OXIDATION, STEAM
EXPLOSION
If desired, one or more sonication, pyrolysis, oxidative, or steam explosion processes
can be used in addition to or instead of electron bombardment to reduce the recalcitrance of the
feedstock. These processes are described in detail in US. Pat. No. 7,932,065 to Medoff, the full
disclosure of which is incorporated herein by reference.
USE OF TREATED S MATERIAL
The biomass material (e.g., plant biomass, animal biomass, paper, and municipal
waste biomass) can be used as ock to produce useful intermediates and products such as
organic acids, salts of organic acids, ides, esters of organic acids and fuels, e.g, fuels for
internal combustion engines or feedstocks for fuel cells. Systems and processes are described
herein that can use as feedstock cellulosic and/or lignocellulosic materials that are readily
available, but often can be difficult to s, e.g, municipal waste streams and waste paper
streams, such as streams that include newspaper, kraft paper, corrugated paper or mixtures of
these.
In order to convert the feedstock to a form that can be readily processed, the glucan-
or xylan-containing cellulose in the feedstock can be hydrolyzed to low molecular weight
carbohydrates, such as , by a saccharifying agent, e.g., an enzyme or acid, a process
referred to as rification. The low molecular weight carbohydrates can then be used, for
example, in an existing manufacturing plant, such as a single cell protein plant, an enzyme
manufacturing plant, or a fuel plant, e.g., an ethanol manufacturing facility.
The feedstock can be yzed using an enzyme, e.g., by combining the materials
and the enzyme in a solvent, e.g, in an aqueous solution. The enzymes can be made/induced
according to the methods described herein.
Specifically, the enzymes can be supplied by organisms that are capable of breaking
down biomass (such as the cellulose and/or the lignin portions of the biomass), or that contain or
manufacture s cellulolytic enzymes (cellulases), ligninases or various small molecule
biomass—degrading metabolites. These enzymes may be a x of enzymes that act
synergistically to degrade crystalline cellulose or the lignin portions of biomass. Examples of
cellulolytic s include: endoglucanases, cellobiohydrolases, and cellobiases (beta-
glucosidases).
During saccharification a osic substrate can be initially hydrolyzed by
endoglucanases at random locations producing eric intermediates. These intermediates
are then substrates for litting glucanases such as iohydrolase to produce cellobiose
from the ends of the cellulose polymer. Cellobiose is a water—soluble 1,4—linked dimer of
glucose. y, iase cleaves cellobiose to yield glucose. The efficiency (e.g., time to
hydrolyze and/or completeness of hydrolysis) of this s depends on the recalcitrance of the
cellulosic material.
INTERMEDIATES AND PRODUCTS
Using the processes described herein, the biomass material can be converted to one or
more products, such as energy, fuels, foods and materials. c examples of products
include, but are not limited to, hydrogen, sugars (e.g, glucose, xylose, arabinose, mannose,
galactose, fructose, disaccharides, oligosaccharides and polysaccharides), alcohols (e.g,
monohydric ls or dihydric alcohols, such as ethanol, n—propanol, isobutanol, sec-butanol,
tert-butanol or n-butanol), ed or hydrous alcohols (e.g., containing greater than 10%, 20%,
% or even greater than 40% water), biodiesel, organic acids, hydrocarbons (e.g.
, methane,
ethane, propane, isobutene, pentane, ne, biodiesel, bio-gasoline and mixtures f), co-
products (e.g., proteins, such as cellulolytic proteins (enzymes) or single cell proteins), and
mixtures of any of these in any ation or relative concentration, and optionally in
ation with any additives (e.g, fuel additives). Other examples include ylic acids,
salts of a carboxylic acid, a mixture of carboxylic acids and salts of carboxylic acids and esters of
carboxylic acids (e.g., methyl, ethyl and n-propyl esters), ketones (e.g., acetone), des (e.g.,
acetaldehyde), alpha and beta unsaturated acids (e.g., acrylic acid) and olefins (e.g., ethylene).
Other alcohols and alcohol tives include propanol, propylene glycol, l,4-butanediol, 1,3-
ediol, sugar alcohols and polyols (e.g., glycol, glycerol, erythritol, threitol, arabitol,
xylitol, ribitol, mannitol, sorbitol, galactitol, iditol, inositol, volemitol, isomalt, maltitol, lactitol,
maltotriitol, maltotetraitol, and polyglycitol and other polyols), and methyl or ethyl esters of any
of these alcohols. Other products include methyl te, methacrylate, lactic acid, citric
acid, formic acid, acetic acid, propionic acid, butyric acid, succinic acid, valeric acid, caproic
acid, 3-hydroxypropionic acid, palmitic acid, stearic acid, oxalic acid, malonic acid, glutaric
acid, oleic acid, linoleic acid, glycolic acid, gamma-hydroxybutyric acid, and mixtures thereof,
salts of any of these acids, mixtures of any of the acids and their respective salts.
Any combination of the above products with each other, and/or of the above products
with other products, which other products may be made by the processes described herein or
otherwise, may be packaged together and sold as products. The products may be combined, e.g.,
mixed, blended or co-dissolved, or may simply be ed or sold together.
Any of the products or combinations of products bed herein may be sanitized or
sterilized prior to selling the products, e. g., after purification or isolation or even after packaging,
to neutralize one or more potentially undesirable contaminants that could be t in the
product(s). Such sanitation can be done with electron bombardment, for example, be at a dosage
of less than about 20 Mrad, e.g., from about 0.1 to 15 Mrad, from about 0.5 to 7 Mrad, or from
about 1 to 3 Mrad.
The processes described herein can produce various by-product streams useful for
generating steam and electricity to be used in other parts of the plant (co-generation) or sold on
the open market. For example, steam generated from burning by-product s can be used in
a lation process. As another example, electricity ted from burning duct
s can be used to power electron beam generators used in pretreatment.
The by—products used to generate steam and electricity are derived from a number of
sources throughout the process. For example, anaerobic digestion of ater can produce a
biogas high in methane and a small amount ofwaste biomass (sludge). As another example,
post-saccharification and/or post-distillate solids (e.g., unconverted lignin, cellulose, and
hemicellulose remaining from the pretreatment and primary processes) can be used, e.g., burned,
as a fuel.
Many ofthe products obtained, such as ethanol or n-butanol, can be utilized as a filel
for powering cars, trucks, tractors, ships or trains, e.g., as an internal combustion fuel or as a fuel
cell feedstock. Many of the products obtained can also be utilized to power aircraft, such as
, e. g., having jet engines or helicopters. In addition, the products described herein can be
utilized for electrical power generation, e.g., in a tional steam generating plant or in a fuel
cell plant.
Other intermediates and products, including food and ceutical products, are
described in US. Pat. App. Pub. 2010/0124583 A1, published May 20, 2010, to Medoff, the full
disclosure of which is hereby incorporated by reference herein.
RIFICATION
The reduced-recalcitrance feedstock is treated with the enzymes discussed above,
generally by ing the material and the enzyme in a fluid medium, e.g., an aqueous
solution. In some cases, the ock is boiled, steeped, or cooked in hot water prior to
saccharification, as described in US. Pat. App. Pub. 2012/0100577 A1 by Medoff and
Masterman, published on April 26, 2012, the entire contents of which are incorporated herein.
The saccharification process can be partially or completely med in a tank (e.g.,
a tank having a volume of at least 4000, 40,000, or 500,000 L) in a manufacturing plant, and/or
can be partially or completely performed in transit, e.g, in a rail car, tanker truck, or in a
supertanker or the hold of a ship. The time required for complete saccharification will depend on
the process conditions and the biomass material and enzyme used. If saccharification is
med in a manufacturing plant under controlled conditions, the cellulose may be
substantially entirely converted to sugar, e.g., glucose in about 12-96 hours. If saccharification is
performed lly or completely in transit, saccharification may take longer.
It is generally preferred that the tank contents be mixed during sacchariflcation, e.g.,
using jet mixing as described in International App. No. l , filed May 18,
2010, which was published in English as W0 2010/135380 and designated the United States, the
full disclosure of which is incorporated by reference herein.
The addition of surfactants can e the rate of saccharification. Examples of
surfactants include non-ionic surfactants, such as a Tween® 20 or Tween® 80 polyethylene
glycol surfactants, ionic surfactants, or amphoteric surfactants.
It is generally preferred that the concentration of the sugar solution resulting from
saccharification be relatively high, e.g, greater than 40%, or greater than 50, 60, 70, 80, 90 or
even greater than 95% by weight. Water may be d, e.g., by evaporation, to increase the
concentration of the sugar solution. This reduces the volume to be shipped, and also inhibits
microbial growth in the solution.
Alternatively, sugar solutions of lower concentrations may be used, in which case it
may be desirable to add an crobial ve, e.g. , a broad spectrum antibiotic, in a low
concentration, e.g., 50 to 150 ppm. Other le antibiotics include amphotericin B, ampicillin,
chloramphenicol, ciprofloxacin, icin, hygromycin B, kanamycin, neomycin, penicillin,
puromycin, streptomycin. Antibiotics will inhibit growth ofmicroorganisms during transport
and e, and can be used at appropriate concentrations, e.g., between 15 and 1000 ppm by
weight, e.g., between 25 and 500 ppm, or between 50 and 150 ppm. If d, an antibiotic can
be included even if the sugar concentration is relatively high. Alternatively, other additives with
anti-microbial of preservative properties may be used. Preferably the antimicrobial additive(s)
are food-grade.
A relatively high concentration solution can be obtained by limiting the amount of
water added to the biomass material with the enzyme. The concentration can be controlled, e.g.
by controlling how much rification takes place. For example, concentration can be
increased by adding more biomass material to the solution. In order to keep the sugar that is
being produced in solution, a surfactant can be added, e.g., one of those discussed above.
Solubility can also be increased by increasing the temperature of the on. For example, the
solution can be maintained at a temperature of 40-50°C, 60—80°C, or even .
SACCHARIFYING AGENTS
Suitable cellulolytic enzymes include ases from species in the genera Bacillus,
CaprinuS, Myceliophthora, Cephalosporz'um, Scytalz'dl'um, Penicillium, AspergilluS,
Pseudomonas, Humicola, um, Thielavz'a, Acremonz’um, ChrySOSporz'um and Trichoderma,
especially those produced by a strain selected from the species ASpergz’lluS (see, e.g., EP Pub.
No. 0 458 162), Humicola insolens ssified as Scytalz'dz'um thermophilum, see, e.g., US. Pat.
No. 4,435,307), CaprinuS cinereuS, um oxysporum, Myceliophthora thermophila,
Merz’pz'luS giganteus, Thielavia terrestris, nium Sp. (including, but not limited to, A.
persicinum, A. acremonium, A. brachypenium, A. dichromosporum, A. obclavatum, A.
pinkertoniae, A. roseogriseum, A. incoloratum, and A. furatum). Preferred strains include
Humicola insolens DSM 1800, Fusarium oxySporum DSM 2672, Mycelz'ophthora thermophila
CBS 117.65, osporz'um Sp. RYM-202, Acremonium Sp. CBS 478.94, Acremonium Sp.
CBS , Acremonium perSicz'num CBS 169.65, Acremonium acremom’um AHU 9519,
Cephalosporium Sp. CBS , Acremonium brachypenium CBS 866.73, Acremanium
dichromosporum CBS , Acremonium obclavatum CBS 311.74, Acremom’um pinkertoniae
CBS 157.70, Acremonium roseogriseum CBS 134.56, Acremonium ratum CBS 146.62,
and Acremoniumfuratum CBS 299.70H. Cellulolytic enzymes may also be obtained from
ChrySOSporium, preferably a strain of Chrysosporium lucknowense. Additional strains that can
be used include, but are not limited to, derma (particularly T. , T. reesez', and T.
koningii), alkalophilic Bacillus (see, for example, US. Pat. No. 3,844,890 and EP Pub. No. 0 458
162), and Streptomyces (see, e.g., EP Pub. No. 0 458 162).
Many microorganisms that can be used to saccharify biomass material and produce
sugars can also be used to ferment and convert those sugars to useful products.
SUGARS
In the processes described , for example after saccharification, sugars (e.g.
glucose and xylose) can be isolated. For example sugars can be isolated by precipitation,
crystallization, chromatography (e.g, simulated moving bed chromatography, high pressure
chromatography), fiagation, extraction, any other isolation method known in the art, and
combinations thereof.
HYDROGENATION AND OTHER CHEMICAL ORMATIONS
The processes described herein can include hydrogenation. For example glucose and
xylose can be hydrogenated to sorbitol and xylitol respectively. Hydrogenation can be
accomplished by use of a catalyst (e.g., Pt/gamma-A1203, Ru/C, Raney Nickel, or other catalysts
know in the art) in combination with H2 under high pressure (e.g., 10 to 12000 psi). Other types
of chemical transformation of the products from the ses described herein can be used, for
example production of c sugar derived products such (e.g., furfural and al-derived
products). Chemical transformations of sugar derived products are described in U.S. Prov. App.
No. 61/667,481, filed July 3, 2012, the disclosure of which is incorporated herein by nce in
its entirety.
TATION
The sugars produced by saccharification can be isolated as a final product, or can be
fermented to produce other products, e.g, alcohols, sugar alcohols, such as erythritol, or organic
acids, e.g., lactic, glutamic or citric acids or amino acids.
Yeast and Zymomonas bacteria, for example, can be used for fermentation or
conversion of sugar(s) to alcohol(s). Other microorganisms are discussed below. The optimum
pH for fermentations is about pH 4 to 7. For example, the optimum pH for yeast is from about
pH 4 to 5, while the optimum pH for Zymomonas is from about pH 5 to 6. Typical fermentation
times are about 24 to 168 hours (e.g., 24 to 96 hrs) with temperatures in the range of 20°C to
40°C (e.g., 26°C to 40°C), however thermophilic microorganisms prefer higher temperatures.
In some embodiments, e.g., when anaerobic organisms are used, at least a portion of
the fermentation is conducted in the absence of oxygen, e.g., under a blanket of an inert gas such
as N2, Ar, He, C02 or mixtures thereof. Additionally, the mixture may have a constant purge of
an inert gas flowing through the tank during part of or all of the fermentation. In some cases,
anaerobic condition, can be achieved or maintained by carbon dioxide production during the
fermentation and no additional inert gas is needed.
In some embodiments, all or a portion of the fermentation process can be interrupted
before the low molecular weight sugar is completely converted to a product (e.g., ethanol). The
intermediate fermentation ts include sugar and ydrates in high trations. The
sugars and carbohydrates can be ed via any means known in the art. These intermediate
fermentation products can be used in preparation of food for human or animal ption.
Additionally or alternatively, the intermediate fermentation products can be ground to a fine
particle size in a stainless-steel laboratory mill to produce a flour—like substance.
Jet mixing may be used during fermentation, and in some cases saccharification and
fermentation are performed in the same tank.
Nutrients for the rganisms may be added during saccharification and/or
fermentation, for example the food-based nutrient packages bed in US. Pat. App. Pub.
2012/0052536, filed July 15, 2011, the complete disclosure of which is incorporated herein by
reference.
“Fermentation” includes the methods and products that are disclosed in US. Prov.
App. No. 61/579,559, filed December 22, 2012, and US. Prov. App. No. 61/579,576, filed
December 22, 2012, the contents of both of which are orated by reference herein in their
entirety.
Mobile fermenters can be utilized, as described in International App. No.
PCT/U82007/074028 (which was filed July 20, 2007, was published in h as WO
2008/01 1598 and designated the United States), the ts of which is incorporated herein in
its entirety. Similarly, the saccharification equipment can be mobile. Further, saccharification
and/or tation may be performed in part or entirely during transit.
FERMENTATION AGENTS
The rganism(s) used in fermentation can be naturally-occurring
microorganisms and/or engineered microorganisms. For example, the rganism can be a
bacterium ding, but not limited to, e.g., a olytic bacterium), a fungus, (including, but
not limited to, e.g., a , a plant, a protist, e.g. a protozoa or a fungus-like protest (including,
but not limited to, e.g., a slime mold), or an alga. When the organisms are compatible, mixtures
of organisms can be utilized.
Suitable fermenting microorganisms have the y to convert ydrates, such
as glucose, fructose, xylose, arabinose, mannose, galactose, oligosaccharides or polysaccharides
into fermentation products. Fermenting microorganisms include strains of the genus
Saccharomyces spp. (including, but not d to, S. cerevisiae (baker’s yeast), S. distatz'cas, S.
avaram), the genus Klayveromyces, (including, but not limited to, K. marxianas, K. fragilis), the
genus a (including, but not limited to, C. pseudotropicalz’s, and C. brassz'cae), Pichz'a
stz‘pz‘tz’s (a relative of Candida ae), the genus Clavz'spora (including, but not limited to, C.
lusz'tanz'ae and C. opantz'ae), the genus Pachysolen (including, but not limited to, P. tannophilas),
the genus Bretannomyces (including, but not limited to, e.g., B. clausenz'z' (Philippidis, G. P.,
1996, Cellulose bioconversion technology, in Handbook on Bioethanol: Production and
Utilization, Wyman, C.E., ed., Taylor & Francis, Washington, DC, 179-212)). Other le
microorganisms include, for example, Zymomonas mobilis, Clostrz’dz’um spp. (including, but not
limited to, C. thermocellum (Philippidis, 1996, supra), C. robulylacetonicum, C.
saccharobalylz‘cum, C. Paniceum, C. bez’jernckl‘z', and C. acetobulyll‘cum), Monilz'ella pollinz's,
Monilz'ella megachz'lz'ensis, Lactobacz'llus spp. Yarrowz'a lipolytica, Aureobasidz'um sp.,
sporonoz'des 319., Trigonopsis variabilis, sporon sp., Monilz'ellaacetoabatans Sp.
Typhala variabilis, a magnoliae, Ustilaginomycetes Sp., Pseudozyma tsukabaensis, yeast
species of genera Zygosaccharomyces, Debaryomyces, Hansenula and Pichia, and fungi of the
dematioid genus Torula.
For instance, Clostrz'dz'um spp. can be used to produce ethanol, butanol, butyric acid,
acetic acid, and acetone. Lactobacz'llus spp., can be used to produce lactice acid.
Many such microbial strains are publicly available, either commercially or through
depositories such as the ATCC (American Type Culture Collection, Manassas, Virginia, USA),
the NRRL (Agricultural Research Sevice Culture Collection, Peoria, Illinois, USA), or the
DSMZ che Sammlung von Mikroorganismen und Zellkulturen GmbH, Braunschweig,
Germany), to name a few.
Commercially available yeasts include, for example, Red Star®/Lesaffre Ethanol Red
(available from Red Star/Lesaffre, USA), FALI® (available from Fleischmann’s Yeast, a division
of Burns Philip Food Inc., USA), SUPERSTART® (available from Alltech, now Lalemand),
GERT STRAND® (available from Gert Strand AB, Sweden) and FERMOL® (available from
DSM Specialties).
Many microorganisms that can be used to saccharify biomass material and produce
sugars can also be used to ferment and convert those sugars to useful products.
DISTILLATION
After fermentation, the resulting fluids can be distilled using, for e, a “beer
” to separate ethanol and other ls from the majority of water and residual solids.
The vapor exiting the beer column can be, e.g., 35% by weight ethanol and can be fed to a
rectification column. A mixture of nearly azeotropic (92.5%) ethanol and water from the
rectification column can be purified to pure (99.5%) ethanol using vapor-phase lar sieves.
The beer column bottoms can be sent to the first effect of a three-effect evaporator. The
cation column reflux condenser can e heat for this first effect. After the first effect,
solids can be separated using a centrifuge and dried in a rotary dryer. A portion (25%) of the
centrifuge effluent can be recycled to tation and the rest sent to the second and third
evaporator effects. Most of the evaporator condensate can be returned to the process as fairly
clean condensate with a small portion split off to waste water treatment to prevent build-up of
low-boiling compounds.
Other than in the examples herein, or unless otherwise expressly ed, all of the
numerical ranges, amounts, values and percentages, such as those for amounts of materials,
tal contents, times and temperatures of on, ratios of amounts, and others, in the
following portion of the specification and attached claims may be read as if ed by the word
“about” even though the term “about” may not expressly appear with the value, amount, or
range. Accordingly, unless indicated to the contrary, the numerical parameters set forth in the
following cation and attached claims are approximations that may vary depending upon
the desired properties sought to be obtained by the present invention. At the very least, and not
as an t to limit the application of the doctrine of equivalents to the scope of the claims,
each numerical parameter should at least be construed in light of the number of reported
significant digits and by applying ordinary rounding ques.
Notwithstanding that the numerical ranges and parameters g forth the broad
scope of the invention are approximations, the numerical values set forth in the specific
examples are reported as precisely as possible. Any numerical value, however, inherently
contains error arily resulting from the standard ion found in its underlying respective
testing measurements. Furthermore, when numerical ranges are set forth , these ranges are
inclusive of the recited range end points (i.e., end points may be used). When percentages by
weight are used herein, the numerical values reported are ve to the total weight.
Also, it should be understood that any numerical range recited herein is intended to
include all sub-ranges subsumed therein. For example, a range of “1 to 10” is intended to
include all sub-ranges between (and including) the recited minimum value of 1 and the recited
maximum value of 10, that is, having a minimum value equal to or greater than 1 and a
maximum value of equal to or less than 10. The terms “one, a, or “an” as used herein are
intended to include “at least one” or “one or more,” unless ise indicated.
EXAMPLES
Materials & Methods
The following ures and materials were used in the following examples.
Cell banking: The following Trichiderma ’ strains were banked: ATCC 66589,
PC3-7; ATCC 56765, RUT-C30; ATCC 56767, NG-l4; ATCC 26921, QM 9414.
Each cell was rehydrated and propagated in potato dextrose (PD) media at 25°C.
For production of master cell banks, each strain was rehydrated overnight in 0.5ml
sterile water. To propagate cells, 40 ul of rehydrated cells were used to inoculate potato se
agar (PDA) solid medium. Rehydrated cells were also inoculated into 50 ml of PD liquid
medium and incubated at 25°C and 200 rpm. After 2 weeks culture in PDA media, spores were
resuspended in sterile NaCl (9g/L), 20% glycerol solution, and stored in -80°C freezer for use as
a cell bank.
WO 96698
Protein measurement and cellalase assay: Protein concentration was measured by
the Bradford method using bovine serum albumin as a standard.
Filter paper assay (FPU), cellobase activity and CMC activity was carried out using
the IUPAC method (T.K. Ghose, Pure Appl. Chem. 59:257-68, 1987).
The reaction product (glucose) was analyzed on a YSI 7100 Multiparameter
Bioanalytical System (YSI Life es, Yellow s, Ohio, USA) or HPLC.
Media: The media included com steep (2 g/L), ammonium sulfate (1.4 g/L),
potassium ide (0.8 g/L), Phosphoric acid (85%, 4mL/L), phthalic acid dipotassium salt (5
g/L), magnesium sulfate heptahydrate L), calcium chloride (0.3g/L), ferrous e
heptahydrate (5 mg/L), manganese sulfate mono hydrate (1.6 mg/L), zinc sulfate heptahydrate
(5 mg/L) and cobalt chloride hexahydrate (2mg/L). The media is described in Herpoel-Gimbert
et al., Biotechnologyfor Biofaels, 2008, 1:18.
Bio-reactor: The freezer stock from the cell banking was used to make the seed
culture using the media described above, with 2.5% additional glucose. The seed culture was
typically made in a flask using an incubator set at 30°C and 200rpm for 72hrs. Seed e
broth (50mL) was used as an inoculum in the 1L starting medium in a 3L fermenter. In growth
phase, 35 g/L of lactose was added to the medium. The e conditions were as follows:
27°C, pH 4.8 (with 6M ammonia), air flow 0.5 VVM, stirring 500 rpm, and dissolved oxygen
(DO) was maintained above 40 % oxygen saturation. In the ion phase, the desired inducer
(discussed below) was added. During fermentation, Antiform 204 (Sigma) was ed into the
culture when the foam reached the fermentor head.
Shakeflask: In addition to the media described above, for the flask culture, Tris
buffer (12.1g/L), maleic acid (11.06g/L) and sodium hydroxide (2.08g/L) were added. A starter
culture was prepared in the media with added glucose. After 3 days of growth, the cell mass was
harvested by centrifugation. The cell mass was re-suspended in 50 m1 of media with the desired
inducer. The flasks were placed in a shaker incubator set at an agitation speed of 200 rpm and
temperature of 30°C.
Example 1. Cellulase Performance Test on Paper, Treated Corn Cob and Untreated
Corn Cob
Various inducers (treated biomass (TBM), untreated biomass (UBM), paper (P) and
carboxylmethylcellulose (CMC, Aldrich)) were used to produce enzymes. The s (TBM
and UBM) was milled corn cob collected n mesh sizes of 15 and 40. Treatment of the
s (UBM) to produce the TBM involved electron bombardment with an electron beam to a
total dose of 35 Mrad. The paper was ed and screened to have a nominal particle size
smaller than 0.16 inch. The r experiments were conducted using shake flasks and PC3-7
and RUT-C30 strains. After 3 days of the growth culture, the harvested cell mass was added to a
series of shake flasks each containing 50 ml of medium and 1 wt. % of one of the inducers.
The induction experiment was allowed to proceed for 11 days. The culture
supernatant was then harvested by centrifugation at 14,500 rpm for 5 minutes, and stored at 4°C.
Protein concentration ofculture supernatant: Using the cell culture grown in the
shake flasks and derived from PC3-7, protein concentrations after 11 days were 1.39, 1.18, 1.06
and 0.26 mg/mL for TBM, UBM, P and CMC tively. For RUT-C30, the protein
concentrations were 1.26, 1.26, 1.00 and 0.26 mg/mL for TBM, P, UBM and CMC respectively.
Cellulase ty: The cellulase activities were assessed and are listed in the table
below.
Table 1. Cellulase activity for different inducers and cell strains.
Inducer Cell type FPU (U/mL) Cellobiase activity FPU Cellobiase activity
/mL U/m U/m
TBM PC—3—7 0.57 0.47 1.04 0.86
UBM PC—3—7 0.45 0.39 1.08 0.93
P PC-3—7 0.57 0.39 0.96 0.66
CMC PC7 0.06 0.11 0.55 0.99
TBM RUT-C30 1.02 0.53 1.97 1.03
UBM RUT-C30 0.72 0.42 1.76 1.03
P RUT-C30 0.71 0.40 1.31 0.74
CMC RUT-C30 0.24 0.18 1.77 1.31
These results show that treated biomass serves to induce enzyme production at a
greater rate than untreated biomass.
Example 2. Enzyme Production in Different Concentrations ofTBM Inducer
This e was done using bioreactors. Cell strain RUT-C30 was propagated in
the media with 2.5 % glucose. After 3 days of growth the e was centrifuged and the cells
was re-suspended in 50 ml of media with 1, 3, 5, 7 and 9 wt.% TBM. The protein concentrations
and activities after 11 days of incubation at 27°C and 200 rpm are shown in the table below.
Table 2. Amounts of Protein and Enzyme Made With Differing Amounts of Inducer
r amount (wt.%) n (g/L) FPU (U/mL) CMC (U/mL)
1 0.7 1.4 1.3
2 1.4 3.1 1.7
3.4 6.2 2.6
7 2.9 2.5 1.5
9 1.5 0.6 1.0
These results show that higher levels of enzymes were produced when the treated
biomass (TBM) was added at a rate of 5 %.
Saccharification ofbiomass with enzymes: Saccharification of biomass (TBM) using
enzymes produced by addition of 2, 5 and 7 wt.% treated biomass inducer (TBM) versus a
commercial enzyme (DuetTM Accellerase, Genencor) was conducted. The biomass, 10wt. %
TBM, was combined with either 0.25 ml/g of enzyme culture broth or commercial enzyme. The
saccharification was carried out at 50°C and 200 rpm in a shaking incubator. After 24 hours the
amount of generated glucose was measured by YSI. The amount of e produced per L of
solution and mg of protein is shown in the table below.
Table 3. Amount of Glucose Produced From Varying Levels of Inducer
Enzyme produced from Glucose (g/L) Glucose (g/mg)
2% TBM r 4.04 2.31
% TBM inducer 4.06 1.08
7% TBM inducer 3.02 0.83
Commercial enzyme 14.4 0.50
Example 3. SDS—PAGE of Enzyme Produced With Treated Biomass
A bioreactor culture was prepared using the method described above except that the
mixing was done at 50 rpm rather than 500 rpm. The protein assay showed that 3.4 g/L protein
was produced.
The is of the protein using SDS PAGE is shown in Lane 1 and 5 are
molecular weight markers, Lane 2 is a 30 uL load of the protein, Lane 3 is a 40 uL load of the
protein, Lane 4 is DuetTM Accelerase enzyme complex (Genencor).
Example 4. Range of Conditions Tested
Induction Parameters Range Working Best
Tested Tested Range Range
Amount Added 25-133g/L 25-133g/L 100g/L
Timing of on Day 0-3 1-3 Day 1-3
Frequency of on 1, 2, and 5 1, 2, and 5 1
Presentation wet or dry wet or dry wet or dry
Treatment Levels 35 35 35
e Timing of Addition Day 3 Day 3 Day 3
Amount Added 4.7-40g/L/d 4.7-18.7g/L/d 18.7g/L/d
continuous continuous uous
Frequency of Addition feed feed feed
Any patent, publication, or other disclosure material, in whole or in part, that is said
to be incorporated by reference herein is incorporated herein only to the extent that the
incorporated material does not conflict with existing definitions, statements, or other disclosure
material set forth in this disclosure. As such, and to the extent necessary, the disclosure as
explicitly set forth herein supersedes any conflicting material incorporated herein by nce.
Any material, or portion thereof, that is said to be incorporated by reference herein, but which
conflicts with existing definitions, ents, or other disclosure material set forth herein will
only be incorporated to the extent that no conflict arises between that incorporated material and
the existing sure material.
2012/071091
While this invention has been particularly shown and described with references to
preferred embodiments thereof, it will be understood by those skilled in the art that various
changes in form and details may be made n without departing from the scope of the
invention encompassed by the appended claims.
Claims (33)
1. A method comprising: selectively combining a cellulolytic rganism with an inductant comprising a first quantity of a particular lignocellulosic material, the first quantity of the particular lignocellulosic material having been d with a particular dose of bombardment with ons, the dose being selected from the range of less than 40 Mrad, and ing conditions effective to induce the cellulolytic microorganism to produce a cellulase complex comprising one or more enzymes having relative concentrations tailored to saccharify the particular lignocellulosic al, the relative concentrations of the one or more enzymes being modulated by selection of the particular dose of bombardment with electrons; and subsequently saccharifying a second quantity of the particular lignocellulosic material with the one or more enzymes, wherein the second quantity of the particular lignocellulosic material has been d with at least 40 Mrad of electron bombardment to reduce its recalcitrance; wherein the first quantity of the ular lignocellulosic material has a greater portion of crystalline cellulose than the second quantity, the dose of electron bombardment applied to the first quantity being less than the dose of electron bombardment applied to the second quantity; n the cellulase complex exhibits enhanced saccharification of the second quantity of the particular lignocellulosic material utable, at least in part, to endoglucanase derived from the first quantity of the particular lignocellulosic material having a greater portion of crystalline cellulose than the second quantity.
2. The method of claim 1, wherein the total dose of electron bombardment d to the first quantity of the particular lignocellulosic material is from 0.5 to 40 Mrad.
3. The method of claim 1 or 2, wherein the total dose of electron bombardment applied to the first quantity is less than about 5 Mrad.
4. The method of any one of claims 1-3, wherein the total dose of electron bombardment applied to the second quantity is from 40 to 150 Mrad.
5. The method of any one of claims 1-4, n the total dose of on bombardment applied to the first quantity is less than about 5 Mrad, and the total dose of electron bombardment applied to the second quantity is 40 to 150 Mrad.
6. The method of any one of claims 1-5, wherein the dose rate of electron bombardment d to the second quantity of the particular lignocellulosic material is from about 0.25 to about 20 ec.
7. The method of any one of claims 1-6, wherein the electron bombardment of the second quantity of the particular lignocellulosic material is provided by an electron beam device having an energy of about 0.5 to about 10 MeV.
8. The method of any one of claims 1-7, wherein the first quantity of the particular lignocellulosic material and/or the second quantity of the particular lignocellulosic material has been mechanically treated to reduce its bulk density and/or increase its surface area.
9. The method of any one of claims 1-8, wherein the first quantity of the particular lignocellulosic material has been comminuted before being combined with the cellulolytic microorganism.
10. The method of any one of claims 1-9, wherein the second quantity of the particular ellulosic material has been comminuted before subsequently saccharifying the second quantity of the particular lignocellulosic material with the one or more enzymes.
11. The method of claim 9 or 10, wherein the comminution comprises dry milling.
12. The method of any one of claims 9-11, n the ution comprises wet milling.
13. The method of any one of claims 1-12, wherein the first quantity of the particular lignocellulosic material and/or the second quantity of the particular lignocellulosic material has a particle size of about 30 to 1400 µm.
14. The method of any one of claims 1-13, wherein the cellulolytic microorganism is selected from the group consisting of a fungus, a bacterium, and a yeast.
15. The method of any one of claims 1-14, wherein the cellulolytic microorganism is a strain of cellulolytic filamentous fungus.
16. The method of any one of claims 1-15, where the cellulolytic microorganism comprises a cellulase producing fungus.
17. The method of any one of claims 1-16, wherein the cellulolytic microorganism is cally engineered.
18. The method of any one of claims 1-17, wherein the cellulolytic rganism is selected from the group consisting of Trichoderma reesei, and Clostridium cellum.
19. The method of any one of claims 1-18, wherein the cellulolytic microorganism is selected from among the strains of Trichoderma .
20. The method of any one of claims 1-19, wherein the cellulolytic microorganism is a lite repression-resistant mutant of Trichoderma reesei.
21. The method of any one of claims 1-20, wherein the olytic microorganism is selected from the group consisting of: RUT-NG 14, PC3-7, QM9414 and/or RUT-C30.
22. The method of any one of claims 1-21, wherein the one or more enzymes are cellulase enzymes.
23. The method of any one of claims 1-22, wherein the one or more enzymes comprise a ase complex of two or more of the following: endoglucanases, cellobiohydrolases, cellobiases, and lulases.
24. The method of any one of claims 1-23, wherein relative concentrations of the one or more enzymes correlates to the relative proportion of one or more substrates in the first quantity of the particular ellulosic material, the one or more substrates comprising one or more of: cellulose, a hemicellulose, and lignin.
25. The method of any one of claims 1-24, wherein the relative proportions of the one or more enzymes correlates to the crystallinity of one or more ates in the first quantity of the particular lignocellulosic material.
26. The method any one of claims 1-25, n the particular lignocellulosic material comprises agricultural residues.
27. The method of any one of claims 1-26, wherein the particular lignocellulosic material comprises one or more of: cotton, grasses, grain residues, rice hulls, oat hulls, wheat chaff, barley hulls, silage, canola straw, wheat straw, barley straw, oat straw, rice straw, jute, hemp, flax, bamboo, sisal, abaca, corn cobs, com stover, soybean stover, corn fiber, a, hay, coconut hair, sugar processing residues, bagasse, beet pulp, agave bagasse, algae, seaweed, cha, buckwheat, banana, barley, cassava, kudzu, oca, sago, m, potato, sweet potato, taro, yams, beans, favas, lentils, peas, and agricultural waste material from any one or more of these.
28. The method of any one of claims 1-27, wherein the particular lignocellulosic material comprises a residue of a rification or fermentation process.
29. The method of any one of claims 1-28, wherein the second quantity of the particular lignocellulosic material comprises material from which the first quantity was selected.
30. The method of any one of claims 1-28, wherein the first quantity of the particular lignocellulosic material was obtained from a different source than the source of the second quantity of the ular ellulosic material.
31. The method of any one of claims 1-30, wherein the electron bombardment of the second quantity of the particular lignocellulosic material is applied in multiple doses.
32. The method of any one of claims 1-31, further comprising concentrating the cellulase complex and storing the cellulase complex prior to subsequently saccharifying the second quantity of the particular lignocellulosic material.
33. The method of claim 32, where concentrating comprises one or more of: chromatography, centrifugation, filtration, dialysis, extraction, evaporation of solvents, spray , and adsorption onto a solid support.
Applications Claiming Priority (5)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US201161579562P | 2011-12-22 | 2011-12-22 | |
US201161579550P | 2011-12-22 | 2011-12-22 | |
US61/579,550 | 2011-12-22 | ||
US61/579,562 | 2011-12-22 | ||
NZ625162A NZ625162B2 (en) | 2011-12-22 | 2012-12-20 | Processing of biomass materials |
Publications (2)
Publication Number | Publication Date |
---|---|
NZ716079A NZ716079A (en) | 2017-07-28 |
NZ716079B2 true NZ716079B2 (en) | 2017-10-31 |
Family
ID=
Similar Documents
Publication | Publication Date | Title |
---|---|---|
AU2016235003B2 (en) | Processing of Biomass Materials | |
AU2017200438B2 (en) | Production of Sugar and Alcohol from Biomass | |
EP2794902B1 (en) | Processing biomass for use in fuel cells | |
NZ716079B2 (en) | Processing of Biomass Materials | |
NZ625162B2 (en) | Processing of biomass materials | |
OA16925A (en) | Processing of biomass materials. | |
NZ739541B2 (en) | Processing Biomass For Use In Fuel Cells | |
NZ625335B2 (en) | Production of sugar and alcohol from biomass | |
NZ716083B2 (en) | Production of Sugar and Alcohol from Biomass | |
NZ719871B2 (en) | Production Of Sugar And Alcohol From Biomass |