NZ615225B2 - Methods of producing biofuels, chlorophylls and carotenoids - Google Patents
Methods of producing biofuels, chlorophylls and carotenoids Download PDFInfo
- Publication number
- NZ615225B2 NZ615225B2 NZ615225A NZ61522512A NZ615225B2 NZ 615225 B2 NZ615225 B2 NZ 615225B2 NZ 615225 A NZ615225 A NZ 615225A NZ 61522512 A NZ61522512 A NZ 61522512A NZ 615225 B2 NZ615225 B2 NZ 615225B2
- Authority
- NZ
- New Zealand
- Prior art keywords
- fraction
- extraction
- algal
- biomass
- ethanol
- Prior art date
Links
- 150000001747 carotenoids Chemical class 0.000 title claims abstract description 49
- 235000019529 tetraterpenoid Nutrition 0.000 title claims abstract description 49
- 235000019804 chlorophyll Nutrition 0.000 title claims abstract description 38
- 229930002875 chlorophylls Natural products 0.000 title claims abstract description 38
- 239000001752 chlorophylls and chlorophyllins Substances 0.000 title claims abstract description 21
- 239000002551 biofuel Substances 0.000 title description 10
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 302
- 150000002632 lipids Chemical class 0.000 claims abstract description 263
- 239000002028 Biomass Substances 0.000 claims abstract description 198
- 239000002904 solvent Substances 0.000 claims abstract description 143
- 239000000727 fraction Substances 0.000 claims abstract description 98
- 230000001264 neutralization Effects 0.000 claims abstract description 96
- 239000007787 solid Substances 0.000 claims abstract description 89
- 241000195493 Cryptophyta Species 0.000 claims abstract description 82
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 76
- 102000004169 proteins and genes Human genes 0.000 claims abstract description 76
- 239000003921 oil Substances 0.000 claims abstract description 59
- 239000007788 liquid Substances 0.000 claims abstract description 37
- 235000020660 omega-3 fatty acid Nutrition 0.000 claims abstract description 23
- ATNHDLDRLWWWCB-AENOIHSZSA-M chlorophyll a Chemical compound C1([C@@H](C(=O)OC)C(=O)C2=C3C)=C2N2C3=CC(C(CC)=C3C)=[N+]4C3=CC3=C(C=C)C(C)=C5N3[Mg-2]42[N+]2=C1[C@@H](CCC(=O)OC\C=C(/C)CCC[C@H](C)CCC[C@H](C)CCCC(C)C)[C@H](C)C2=C5 ATNHDLDRLWWWCB-AENOIHSZSA-M 0.000 claims abstract description 17
- 229940012843 Omega-3 Fatty Acids Drugs 0.000 claims abstract description 16
- 239000006014 omega-3 oil Substances 0.000 claims abstract description 16
- 235000014633 carbohydrates Nutrition 0.000 claims abstract description 9
- 150000001720 carbohydrates Chemical class 0.000 claims abstract description 9
- 239000002031 ethanolic fraction Substances 0.000 claims abstract description 7
- 239000000203 mixture Substances 0.000 claims description 106
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 99
- 239000000446 fuel Substances 0.000 claims description 57
- 239000000463 material Substances 0.000 claims description 37
- 150000002148 esters Chemical class 0.000 claims description 36
- 239000004927 clay Substances 0.000 claims description 11
- 229910052570 clay Inorganic materials 0.000 claims description 11
- PEDCQBHIVMGVHV-UHFFFAOYSA-N glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 11
- 235000011187 glycerol Nutrition 0.000 claims description 10
- 239000003054 catalyst Substances 0.000 claims description 6
- 230000003635 deoxygenating Effects 0.000 claims description 4
- 238000000605 extraction Methods 0.000 description 227
- 238000000034 method Methods 0.000 description 97
- 235000018102 proteins Nutrition 0.000 description 73
- OKKJLVBELUTLKV-UHFFFAOYSA-N methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 64
- 239000007791 liquid phase Substances 0.000 description 58
- 235000019198 oils Nutrition 0.000 description 57
- 239000000047 product Substances 0.000 description 56
- 239000007790 solid phase Substances 0.000 description 48
- 238000000926 separation method Methods 0.000 description 36
- 238000011084 recovery Methods 0.000 description 32
- 241000195649 Chlorella <Chlorellales> Species 0.000 description 29
- 210000004027 cells Anatomy 0.000 description 28
- RTZKZFJDLAIYFH-UHFFFAOYSA-N diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 26
- VLKZOEOYAKHREP-UHFFFAOYSA-N hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 26
- 239000012528 membrane Substances 0.000 description 26
- 239000002253 acid Substances 0.000 description 24
- 239000000284 extract Substances 0.000 description 21
- 239000002002 slurry Substances 0.000 description 21
- 238000009835 boiling Methods 0.000 description 20
- XEKOWRVHYACXOJ-UHFFFAOYSA-N acetic acid ethyl ester Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 18
- CSCPPACGZOOCGX-UHFFFAOYSA-N acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 18
- 238000005194 fractionation Methods 0.000 description 18
- 235000014113 dietary fatty acids Nutrition 0.000 description 17
- 239000000194 fatty acid Substances 0.000 description 17
- 150000004665 fatty acids Chemical class 0.000 description 16
- 150000002500 ions Chemical class 0.000 description 15
- 239000003463 adsorbent Substances 0.000 description 14
- KFZMGEQAYNKOFK-UHFFFAOYSA-N iso-propanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 description 14
- 239000003208 petroleum Substances 0.000 description 14
- 229940096118 Ella Drugs 0.000 description 13
- 239000008079 hexane Substances 0.000 description 13
- 229910052751 metal Inorganic materials 0.000 description 13
- 239000002184 metal Substances 0.000 description 13
- 229960000200 ulipristal Drugs 0.000 description 13
- 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 13
- WEVYAHXRMPXWCK-UHFFFAOYSA-N acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 12
- 239000000243 solution Substances 0.000 description 12
- 239000000126 substance Substances 0.000 description 12
- 238000004821 distillation Methods 0.000 description 11
- 230000002708 enhancing Effects 0.000 description 11
- 229940067631 Phospholipids Drugs 0.000 description 10
- 238000010438 heat treatment Methods 0.000 description 10
- 230000000670 limiting Effects 0.000 description 10
- 150000003904 phospholipids Chemical class 0.000 description 10
- 150000002739 metals Chemical class 0.000 description 9
- 239000011780 sodium chloride Substances 0.000 description 9
- 238000000638 solvent extraction Methods 0.000 description 9
- 239000011877 solvent mixture Substances 0.000 description 9
- 241000894007 species Species 0.000 description 9
- 150000001335 aliphatic alkanes Chemical class 0.000 description 8
- 229940075894 denatured ethanol Drugs 0.000 description 8
- 238000011026 diafiltration Methods 0.000 description 8
- 238000005886 esterification reaction Methods 0.000 description 8
- 238000001914 filtration Methods 0.000 description 8
- 230000002209 hydrophobic Effects 0.000 description 8
- 238000002156 mixing Methods 0.000 description 8
- 239000012454 non-polar solvent Substances 0.000 description 8
- 238000000751 protein extraction Methods 0.000 description 8
- 150000003839 salts Chemical class 0.000 description 8
- 238000004062 sedimentation Methods 0.000 description 8
- 150000001875 compounds Chemical class 0.000 description 7
- 230000000694 effects Effects 0.000 description 7
- 239000012530 fluid Substances 0.000 description 7
- 238000004519 manufacturing process Methods 0.000 description 7
- 235000015112 vegetable and seed oil Nutrition 0.000 description 7
- 239000008158 vegetable oil Substances 0.000 description 7
- -1 coatings Substances 0.000 description 6
- 102100001249 ALB Human genes 0.000 description 6
- 101710027066 ALB Proteins 0.000 description 6
- 101700077116 AVE3 Proteins 0.000 description 6
- 101700006934 AVEA Proteins 0.000 description 6
- 101700044096 AVEE Proteins 0.000 description 6
- 101700047979 AVEF Proteins 0.000 description 6
- 101700064290 GDA0 Proteins 0.000 description 6
- 101700058641 GDA2 Proteins 0.000 description 6
- 101700033989 GDA3 Proteins 0.000 description 6
- 101700012011 GDA4 Proteins 0.000 description 6
- 101700031401 GDA5 Proteins 0.000 description 6
- 101700036788 GDA6 Proteins 0.000 description 6
- 101700014913 GDA7 Proteins 0.000 description 6
- 101700083010 GDA8 Proteins 0.000 description 6
- 101700061165 GDA9 Proteins 0.000 description 6
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 6
- 101700085810 gda1 Proteins 0.000 description 6
- 239000012466 permeate Substances 0.000 description 6
- 239000011347 resin Substances 0.000 description 6
- 229920005989 resin Polymers 0.000 description 6
- 239000012465 retentate Substances 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 150000003626 triacylglycerols Chemical class 0.000 description 6
- 108010001949 Algal Proteins Proteins 0.000 description 5
- 239000003225 biodiesel Substances 0.000 description 5
- 238000004061 bleaching Methods 0.000 description 5
- OKTJSMMVPCPJKN-UHFFFAOYSA-N carbon Chemical group [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 5
- 229910052799 carbon Inorganic materials 0.000 description 5
- 238000005119 centrifugation Methods 0.000 description 5
- 238000010586 diagram Methods 0.000 description 5
- 235000013305 food Nutrition 0.000 description 5
- 230000001965 increased Effects 0.000 description 5
- 238000011068 load Methods 0.000 description 5
- 238000000199 molecular distillation Methods 0.000 description 5
- 235000021122 unsaturated fatty acids Nutrition 0.000 description 5
- 150000004670 unsaturated fatty acids Chemical class 0.000 description 5
- 241000611184 Amphora Species 0.000 description 4
- 240000009108 Chlorella vulgaris Species 0.000 description 4
- 235000007089 Chlorella vulgaris Nutrition 0.000 description 4
- JAZBEHYOTPTENJ-JLNKQSITSA-N Eicosapentaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCCC(O)=O JAZBEHYOTPTENJ-JLNKQSITSA-N 0.000 description 4
- 241000502321 Navicula Species 0.000 description 4
- 241000180701 Nitzschia <flatworm> Species 0.000 description 4
- 239000002585 base Substances 0.000 description 4
- 235000012970 cakes Nutrition 0.000 description 4
- 238000004945 emulsification Methods 0.000 description 4
- 238000001704 evaporation Methods 0.000 description 4
- 238000005188 flotation Methods 0.000 description 4
- 238000006460 hydrolysis reaction Methods 0.000 description 4
- 238000005374 membrane filtration Methods 0.000 description 4
- 238000000874 microwave-assisted extraction Methods 0.000 description 4
- IJGRMHOSHXDMSA-UHFFFAOYSA-N nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 4
- 239000003960 organic solvent Substances 0.000 description 4
- 230000003647 oxidation Effects 0.000 description 4
- 238000007254 oxidation reaction Methods 0.000 description 4
- 239000012071 phase Substances 0.000 description 4
- NBIIXXVUZAFLBC-UHFFFAOYSA-N phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 4
- 229920003053 polystyrene-divinylbenzene Polymers 0.000 description 4
- 238000007670 refining Methods 0.000 description 4
- YXFVVABEGXRONW-UHFFFAOYSA-N toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 4
- ZWEHNKRNPOVVGH-UHFFFAOYSA-N 2-butanone Chemical compound CCC(C)=O ZWEHNKRNPOVVGH-UHFFFAOYSA-N 0.000 description 3
- 210000002421 Cell Wall Anatomy 0.000 description 3
- 241000227752 Chaetoceros Species 0.000 description 3
- 241000195634 Dunaliella Species 0.000 description 3
- 229960005135 Eicosapentaenoic Acid Drugs 0.000 description 3
- 241000196324 Embryophyta Species 0.000 description 3
- KEMQGTRYUADPNZ-UHFFFAOYSA-N Heptadecanoic acid Chemical compound CCCCCCCCCCCCCCCCC(O)=O KEMQGTRYUADPNZ-UHFFFAOYSA-N 0.000 description 3
- HWKQNAWCHQMZHK-UHFFFAOYSA-N Trolnitrate Chemical compound [O-][N+](=O)OCCN(CCO[N+]([O-])=O)CCO[N+]([O-])=O HWKQNAWCHQMZHK-UHFFFAOYSA-N 0.000 description 3
- 238000007792 addition Methods 0.000 description 3
- 239000003513 alkali Substances 0.000 description 3
- 150000001350 alkyl halides Chemical class 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 239000000440 bentonite Substances 0.000 description 3
- 229910000278 bentonite Inorganic materials 0.000 description 3
- 239000010779 crude oil Substances 0.000 description 3
- 230000001186 cumulative Effects 0.000 description 3
- 230000003247 decreasing Effects 0.000 description 3
- 238000001035 drying Methods 0.000 description 3
- 235000020673 eicosapentaenoic acid Nutrition 0.000 description 3
- 239000000839 emulsion Substances 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 239000003502 gasoline Substances 0.000 description 3
- 238000002386 leaching Methods 0.000 description 3
- 150000004668 long chain fatty acids Chemical class 0.000 description 3
- IMNFDUFMRHMDMM-UHFFFAOYSA-N n-heptane Chemical compound CCCCCCC IMNFDUFMRHMDMM-UHFFFAOYSA-N 0.000 description 3
- 239000002417 nutraceutical Substances 0.000 description 3
- 235000016709 nutrition Nutrition 0.000 description 3
- OFBQJSOFQDEBGM-UHFFFAOYSA-N pentane Chemical compound CCCCC OFBQJSOFQDEBGM-UHFFFAOYSA-N 0.000 description 3
- 238000001556 precipitation Methods 0.000 description 3
- 238000000746 purification Methods 0.000 description 3
- 230000002829 reduced Effects 0.000 description 3
- 101700043375 sing Proteins 0.000 description 3
- 238000003860 storage Methods 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 238000005809 transesterification reaction Methods 0.000 description 3
- 239000002351 wastewater Substances 0.000 description 3
- OYHQOLUKZRVURQ-HZJYTTRNSA-N 60-33-3 Chemical compound CCCCC\C=C/C\C=C/CCCCCCCC(O)=O OYHQOLUKZRVURQ-HZJYTTRNSA-N 0.000 description 2
- DKPFZGUDAPQIHT-UHFFFAOYSA-N Butyl acetate Natural products CCCCOC(C)=O DKPFZGUDAPQIHT-UHFFFAOYSA-N 0.000 description 2
- 241000206751 Chrysophyceae Species 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N HCl Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 101700033533 HP20 Proteins 0.000 description 2
- NTIZESTWPVYFNL-UHFFFAOYSA-N Methyl isobutyl ketone Chemical compound CC(C)CC(C)=O NTIZESTWPVYFNL-UHFFFAOYSA-N 0.000 description 2
- 241000199478 Ochromonas Species 0.000 description 2
- 241000514008 Oocystis Species 0.000 description 2
- 241000196250 Prototheca Species 0.000 description 2
- 241000195648 Pseudochlorella pringsheimii Species 0.000 description 2
- QIQXTHQIDYTFRH-UHFFFAOYSA-N Stearic acid Chemical compound CCCCCCCCCCCCCCCCCC(O)=O QIQXTHQIDYTFRH-UHFFFAOYSA-N 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- 101710029677 TIM22-2 Proteins 0.000 description 2
- 229940035295 Ting Drugs 0.000 description 2
- 230000036462 Unbound Effects 0.000 description 2
- 239000000654 additive Substances 0.000 description 2
- 239000005456 alcohol based solvent Substances 0.000 description 2
- 150000001298 alcohols Chemical class 0.000 description 2
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 2
- 238000005815 base catalysis Methods 0.000 description 2
- UHOVQNZJYSORNB-UHFFFAOYSA-N benzene Chemical compound C1=CC=CC=C1 UHOVQNZJYSORNB-UHFFFAOYSA-N 0.000 description 2
- 239000006227 byproduct Substances 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 125000004432 carbon atoms Chemical group C* 0.000 description 2
- 230000015556 catabolic process Effects 0.000 description 2
- HEDRZPFGACZZDS-UHFFFAOYSA-N chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 2
- 238000007796 conventional method Methods 0.000 description 2
- XDTMQSROBMDMFD-UHFFFAOYSA-N cyclohexane Chemical compound C1CCCCC1 XDTMQSROBMDMFD-UHFFFAOYSA-N 0.000 description 2
- 238000010908 decantation Methods 0.000 description 2
- 230000004059 degradation Effects 0.000 description 2
- 238000006731 degradation reaction Methods 0.000 description 2
- 238000011143 downstream manufacturing Methods 0.000 description 2
- 239000003344 environmental pollutant Substances 0.000 description 2
- 230000002255 enzymatic Effects 0.000 description 2
- 238000005189 flocculation Methods 0.000 description 2
- 230000016615 flocculation Effects 0.000 description 2
- 235000013373 food additive Nutrition 0.000 description 2
- 239000002778 food additive Substances 0.000 description 2
- LYCAIKOWRPUZTN-UHFFFAOYSA-N glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 2
- 238000003306 harvesting Methods 0.000 description 2
- 235000013402 health food Nutrition 0.000 description 2
- 239000001257 hydrogen Substances 0.000 description 2
- 229910052739 hydrogen Inorganic materials 0.000 description 2
- UFHFLCQGNIYNRP-UHFFFAOYSA-N hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 2
- 239000012535 impurity Substances 0.000 description 2
- 239000010842 industrial wastewater Substances 0.000 description 2
- 230000002401 inhibitory effect Effects 0.000 description 2
- 150000002576 ketones Chemical class 0.000 description 2
- 229960004232 linoleic acid Drugs 0.000 description 2
- 239000000314 lubricant Substances 0.000 description 2
- 238000003808 methanol extraction Methods 0.000 description 2
- 235000021281 monounsaturated fatty acids Nutrition 0.000 description 2
- 239000010841 municipal wastewater Substances 0.000 description 2
- LRHPLDYGYMQRHN-UHFFFAOYSA-N n-butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 description 2
- 238000001728 nano-filtration Methods 0.000 description 2
- 150000002823 nitrates Chemical class 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- 235000015097 nutrients Nutrition 0.000 description 2
- 235000021317 phosphate Nutrition 0.000 description 2
- 150000003013 phosphoric acid derivatives Chemical class 0.000 description 2
- 231100000719 pollutant Toxicity 0.000 description 2
- 235000020777 polyunsaturated fatty acids Nutrition 0.000 description 2
- 230000000135 prohibitive Effects 0.000 description 2
- ATUOYWHBWRKTHZ-UHFFFAOYSA-N propane Chemical compound CCC ATUOYWHBWRKTHZ-UHFFFAOYSA-N 0.000 description 2
- BDERNNFJNOPAEC-UHFFFAOYSA-N propanol Chemical compound CCCO BDERNNFJNOPAEC-UHFFFAOYSA-N 0.000 description 2
- 235000003441 saturated fatty acids Nutrition 0.000 description 2
- 150000004671 saturated fatty acids Chemical class 0.000 description 2
- 238000001577 simple distillation Methods 0.000 description 2
- 239000011343 solid material Substances 0.000 description 2
- 235000003702 sterols Nutrition 0.000 description 2
- 150000003432 sterols Chemical class 0.000 description 2
- 238000003756 stirring Methods 0.000 description 2
- WYURNTSHIVDZCO-UHFFFAOYSA-N tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 description 2
- 229910052721 tungsten Inorganic materials 0.000 description 2
- XLYOFNOQVPJJNP-ZSJDYOACSA-N water-d2 Chemical compound [2H]O[2H] XLYOFNOQVPJJNP-ZSJDYOACSA-N 0.000 description 2
- 239000001993 wax Substances 0.000 description 2
- 235000008210 xanthophylls Nutrition 0.000 description 2
- 150000003735 xanthophylls Chemical class 0.000 description 2
- XSXIVVZCUAHUJO-HZJYTTRNSA-N (11Z,14Z)-eicosadienoic acid Chemical compound CCCCC\C=C/C\C=C/CCCCCCCCCC(O)=O XSXIVVZCUAHUJO-HZJYTTRNSA-N 0.000 description 1
- KBPHJBAIARWVSC-RGZFRNHPSA-N (1R)-4-[(1E,3E,5E,7E,9E,11E,13E,15E,17E)-18-[(1R,4R)-4-hydroxy-2,6,6-trimethylcyclohex-2-en-1-yl]-3,7,12,16-tetramethyloctadeca-1,3,5,7,9,11,13,15,17-nonaenyl]-3,5,5-trimethylcyclohex-3-en-1-ol Chemical compound C([C@H](O)CC=1C)C(C)(C)C=1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\[C@H]1C(C)=C[C@H](O)CC1(C)C KBPHJBAIARWVSC-RGZFRNHPSA-N 0.000 description 1
- JKQXZKUSFCKOGQ-QAYBQHTQSA-N (1R)-4-[(1E,3E,5E,7E,9E,11E,13E,15E,17E)-18-[(4R)-4-hydroxy-2,6,6-trimethylcyclohex-1-en-1-yl]-3,7,12,16-tetramethyloctadeca-1,3,5,7,9,11,13,15,17-nonaen-1-yl]-3,5,5-trimethylcyclohex-3-en-1-ol Chemical compound C([C@H](O)CC=1C)C(C)(C)C=1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)C[C@@H](O)CC1(C)C JKQXZKUSFCKOGQ-QAYBQHTQSA-N 0.000 description 1
- ZCDMRPMKAQLWAO-PZLFCYFRSA-N (5Z,8Z,11Z,14Z)-icosa-5,8,11,14-tetraenoic acid;(9Z,12Z)-octadeca-9,12-dienoic acid Chemical compound CCCCC\C=C/C\C=C/CCCCCCCC(O)=O.CCCCC\C=C/C\C=C/C\C=C/C\C=C/CCCC(O)=O ZCDMRPMKAQLWAO-PZLFCYFRSA-N 0.000 description 1
- MQZIGYBFDRPAKN-QISQUURKSA-N 6-hydroxy-3-[(1E,3E,5E,7E,9E,11E,13E,15E,17E)-18-(4-hydroxy-2,6,6-trimethyl-3-oxocyclohexen-1-yl)-3,7,12,16-tetramethyloctadeca-1,3,5,7,9,11,13,15,17-nonaenyl]-2,4,4-trimethylcyclohex-2-en-1-one Chemical compound CC=1C(=O)C(O)CC(C)(C)C=1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)C(=O)C(O)CC1(C)C MQZIGYBFDRPAKN-QISQUURKSA-N 0.000 description 1
- 241001133760 Acoelorraphe Species 0.000 description 1
- 102000009027 Albumins Human genes 0.000 description 1
- 108010088751 Albumins Proteins 0.000 description 1
- 241000192542 Anabaena Species 0.000 description 1
- 235000017060 Arachis glabrata Nutrition 0.000 description 1
- 240000005781 Arachis hypogaea Species 0.000 description 1
- 235000010777 Arachis hypogaea Nutrition 0.000 description 1
- 235000018262 Arachis monticola Nutrition 0.000 description 1
- 235000016425 Arthrospira platensis Nutrition 0.000 description 1
- 240000002900 Arthrospira platensis Species 0.000 description 1
- 244000075850 Avena orientalis Species 0.000 description 1
- 235000007319 Avena orientalis Nutrition 0.000 description 1
- 235000007558 Avena sp Nutrition 0.000 description 1
- 241000271566 Aves Species 0.000 description 1
- UKMSUNONTOPOIO-UHFFFAOYSA-N Behenic acid Chemical compound CCCCCCCCCCCCCCCCCCCCCC(O)=O UKMSUNONTOPOIO-UHFFFAOYSA-N 0.000 description 1
- 229960002747 Betacarotene Drugs 0.000 description 1
- 241000219430 Betula pendula Species 0.000 description 1
- 241001536324 Botryococcus Species 0.000 description 1
- 235000006008 Brassica napus var napus Nutrition 0.000 description 1
- 240000000385 Brassica napus var. napus Species 0.000 description 1
- 235000006618 Brassica rapa subsp oleifera Nutrition 0.000 description 1
- 235000004977 Brassica sinapistrum Nutrition 0.000 description 1
- 235000007575 Calluna vulgaris Nutrition 0.000 description 1
- 240000002804 Calluna vulgaris Species 0.000 description 1
- 235000016401 Camelina Nutrition 0.000 description 1
- 240000008923 Camelina sativa Species 0.000 description 1
- 241000222120 Candida <Saccharomycetales> Species 0.000 description 1
- 241001249699 Capitata Species 0.000 description 1
- 235000003255 Carthamus tinctorius Nutrition 0.000 description 1
- 240000005801 Carthamus tinctorius Species 0.000 description 1
- 241000227757 Chaetoceros sp. Species 0.000 description 1
- 229940112822 Chewing Gum Drugs 0.000 description 1
- 241000704925 Chlorella miniata Species 0.000 description 1
- 241000832151 Chlorella regularis Species 0.000 description 1
- 241001287915 Chlorella sp. 'anitrata' Species 0.000 description 1
- 235000018708 Chlorella vulgaris var vulgaris Nutrition 0.000 description 1
- 240000009282 Chlorella vulgaris var. vulgaris Species 0.000 description 1
- 241000180279 Chlorococcum Species 0.000 description 1
- 241000144274 Chlorococcum infusionum Species 0.000 description 1
- 241000195658 Chloroidium saccharophilum Species 0.000 description 1
- 241000195628 Chlorophyta Species 0.000 description 1
- 241000195492 Chroomonas Species 0.000 description 1
- 241000317914 Chrysis Species 0.000 description 1
- 241000722206 Chrysotila carterae Species 0.000 description 1
- 235000008733 Citrus aurantifolia Nutrition 0.000 description 1
- 235000013162 Cocos nucifera Nutrition 0.000 description 1
- 240000007170 Cocos nucifera Species 0.000 description 1
- 241001301781 Coelastrella vacuolata Species 0.000 description 1
- 241001245609 Cricosphaera Species 0.000 description 1
- 241000195618 Cryptomonas Species 0.000 description 1
- 241000192700 Cyanobacteria Species 0.000 description 1
- 241001147476 Cyclotella Species 0.000 description 1
- 241001147477 Cyclotella cryptica Species 0.000 description 1
- 241000720038 Diplosphaera sphaerica Species 0.000 description 1
- 241000611421 Elia Species 0.000 description 1
- 241000464908 Elliptica Species 0.000 description 1
- 241000354295 Eremosphaera Species 0.000 description 1
- 241000195620 Euglena Species 0.000 description 1
- 241000692361 Fistulifera saprophila Species 0.000 description 1
- 229940098330 GAMMA LINOLEIC ACID Drugs 0.000 description 1
- 235000010469 Glycine max Nutrition 0.000 description 1
- 241000168525 Haematococcus Species 0.000 description 1
- 240000006669 Helianthus annuus Species 0.000 description 1
- 235000003222 Helianthus annuus Nutrition 0.000 description 1
- 241001037825 Hymenomonas Species 0.000 description 1
- 241000221089 Jatropha Species 0.000 description 1
- 241000936931 Lepocinclis Species 0.000 description 1
- 241000283986 Lepus Species 0.000 description 1
- OYHQOLUKZRVURQ-IXWMQOLASA-N Linoleic acid Natural products CCCCC\C=C/C\C=C\CCCCCCCC(O)=O OYHQOLUKZRVURQ-IXWMQOLASA-N 0.000 description 1
- 240000006240 Linum usitatissimum Species 0.000 description 1
- 235000004431 Linum usitatissimum Nutrition 0.000 description 1
- 229960005375 Lutein Drugs 0.000 description 1
- KBPHJBAIARWVSC-NRHWGSPPSA-N Lutein Natural products O[C@H]1C=C(C)[C@H](/C=C/C(=C\C=C\C(=C/C=C/C=C(\C=C\C=C(/C=C/C=2C(C)(C)C[C@H](O)CC=2C)\C)/C)\C)/C)C(C)(C)C1 KBPHJBAIARWVSC-NRHWGSPPSA-N 0.000 description 1
- 241000520876 Merismopedia Species 0.000 description 1
- 241000404165 Microcephala Species 0.000 description 1
- DDLIGBOFAVUZHB-UHFFFAOYSA-N Midazolam Chemical compound C12=CC(Cl)=CC=C2N2C(C)=NC=C2CN=C1C1=CC=CC=C1F DDLIGBOFAVUZHB-UHFFFAOYSA-N 0.000 description 1
- 241000108056 Monas Species 0.000 description 1
- 241001478792 Monoraphidium Species 0.000 description 1
- 241000196305 Nannochloris Species 0.000 description 1
- 241000224476 Nannochloropsis salina Species 0.000 description 1
- 241001442227 Nephroselmis Species 0.000 description 1
- 229910003294 NiMo Inorganic materials 0.000 description 1
- 241000405774 Nitzschia pusilla Species 0.000 description 1
- TVMXDCGIABBOFY-UHFFFAOYSA-N Octane Chemical compound CCCCCCCC TVMXDCGIABBOFY-UHFFFAOYSA-N 0.000 description 1
- 240000007817 Olea europaea Species 0.000 description 1
- 229940033080 Omega-6 Fatty Acids Drugs 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 241000192497 Oscillatoria Species 0.000 description 1
- 241000682093 Oscillatoria subbrevis Species 0.000 description 1
- 241001036353 Parachlorella Species 0.000 description 1
- 241000195646 Parachlorella kessleri Species 0.000 description 1
- 241000196317 Platymonas Species 0.000 description 1
- 231100000614 Poison Toxicity 0.000 description 1
- 241001597169 Prototheca stagnorum Species 0.000 description 1
- 102100000241 RASD2 Human genes 0.000 description 1
- 101710004135 RASD2 Proteins 0.000 description 1
- 241001524101 Rhodococcus opacus Species 0.000 description 1
- 241000905510 Scena Species 0.000 description 1
- 241000195663 Scenedesmus Species 0.000 description 1
- 241000997737 Scenedesmus armatus Species 0.000 description 1
- 241000233671 Schizochytrium Species 0.000 description 1
- 241001535061 Selenastrum Species 0.000 description 1
- 241000196294 Spirogyra Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 235000021355 Stearic acid Nutrition 0.000 description 1
- DCXXMTOCNZCJGO-UHFFFAOYSA-N Stearin Chemical compound CCCCCCCCCCCCCCCCCC(=O)OCC(OC(=O)CCCCCCCCCCCCCCCCC)COC(=O)CCCCCCCCCCCCCCCCC DCXXMTOCNZCJGO-UHFFFAOYSA-N 0.000 description 1
- 229910000831 Steel Inorganic materials 0.000 description 1
- 241000018896 Stigmatophora Species 0.000 description 1
- 241000192560 Synechococcus sp. Species 0.000 description 1
- 240000000785 Tagetes erecta Species 0.000 description 1
- 235000012311 Tagetes erecta Nutrition 0.000 description 1
- 241000891463 Tetraedron Species 0.000 description 1
- 241000196321 Tetraselmis Species 0.000 description 1
- 235000015450 Tilia cordata Nutrition 0.000 description 1
- 235000011941 Tilia x europaea Nutrition 0.000 description 1
- 229940029983 VITAMINS Drugs 0.000 description 1
- 229940021016 Vitamin IV solution additives Drugs 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- JKQXZKUSFCKOGQ-PQJNXSRMSA-N Zeaxanthin Natural products C([C@H](O)CC=1C)C(C)(C)C=1\C=C\C(\C)=C/C=C/C(/C)=C\C=C\C=C(\C)/C=C/C=C(/C)\C=C\C1=C(C)C[C@@H](O)CC1(C)C JKQXZKUSFCKOGQ-PQJNXSRMSA-N 0.000 description 1
- 229940093612 Zein Drugs 0.000 description 1
- 229920002494 Zein Polymers 0.000 description 1
- 241000195647 [Chlorella] fusca Species 0.000 description 1
- 241000857102 [Chlorella] gloriosa Species 0.000 description 1
- 239000002250 absorbent Substances 0.000 description 1
- 230000002745 absorbent Effects 0.000 description 1
- 238000007171 acid catalysis Methods 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 239000000853 adhesive Substances 0.000 description 1
- 230000001070 adhesive Effects 0.000 description 1
- 229940050528 albumin Drugs 0.000 description 1
- 125000003118 aryl group Chemical group 0.000 description 1
- 235000013793 astaxanthin Nutrition 0.000 description 1
- 239000001168 astaxanthin Substances 0.000 description 1
- 229940022405 astaxanthin Drugs 0.000 description 1
- 239000012298 atmosphere Substances 0.000 description 1
- 238000011021 bench scale process Methods 0.000 description 1
- OENHQHLEOONYIE-VYAWBVGESA-N beta-Carotene Natural products CC=1CCCC(C)(C)C=1\C=C\C(\C)=C/C=C/C(/C)=C\C=C\C=C(\C)/C=C/C=C(/C)\C=C\C1=C(C)CCCC1(C)C OENHQHLEOONYIE-VYAWBVGESA-N 0.000 description 1
- 235000013734 beta-carotene Nutrition 0.000 description 1
- 239000011648 beta-carotene Substances 0.000 description 1
- 229920000704 biodegradable plastic Polymers 0.000 description 1
- 235000008984 brauner Senf Nutrition 0.000 description 1
- 125000000484 butyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 1
- 150000001746 carotenes Chemical class 0.000 description 1
- 235000005473 carotenes Nutrition 0.000 description 1
- 239000000919 ceramic Substances 0.000 description 1
- 235000013339 cereals Nutrition 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 235000015218 chewing gum Nutrition 0.000 description 1
- KZBUYRJDOAKODT-UHFFFAOYSA-N chlorine Chemical compound ClCl KZBUYRJDOAKODT-UHFFFAOYSA-N 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 239000012141 concentrate Substances 0.000 description 1
- 239000000356 contaminant Substances 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 235000005824 corn Nutrition 0.000 description 1
- 230000000875 corresponding Effects 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 235000012343 cottonseed oil Nutrition 0.000 description 1
- 238000005336 cracking Methods 0.000 description 1
- 244000038559 crop plants Species 0.000 description 1
- 238000002425 crystallisation Methods 0.000 description 1
- 230000005712 crystallization Effects 0.000 description 1
- 230000001419 dependent Effects 0.000 description 1
- 238000001212 derivatisation Methods 0.000 description 1
- 239000003599 detergent Substances 0.000 description 1
- 239000006280 diesel fuel additive Substances 0.000 description 1
- 235000015872 dietary supplement Nutrition 0.000 description 1
- 238000010981 drying operation Methods 0.000 description 1
- 238000004836 empirical method Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 235000020774 essential nutrients Nutrition 0.000 description 1
- 238000002481 ethanol extraction Methods 0.000 description 1
- 239000000469 ethanolic extract Substances 0.000 description 1
- 150000002170 ethers Chemical class 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
- 238000004880 explosion Methods 0.000 description 1
- 238000001125 extrusion Methods 0.000 description 1
- 239000003925 fat Substances 0.000 description 1
- 235000019197 fats Nutrition 0.000 description 1
- 125000005313 fatty acid group Chemical group 0.000 description 1
- 235000019387 fatty acid methyl ester Nutrition 0.000 description 1
- 239000000835 fiber Substances 0.000 description 1
- 235000004426 flaxseed Nutrition 0.000 description 1
- 230000004907 flux Effects 0.000 description 1
- 239000006260 foam Substances 0.000 description 1
- 235000012041 food component Nutrition 0.000 description 1
- 239000005417 food ingredient Substances 0.000 description 1
- 239000002816 fuel additive Substances 0.000 description 1
- 239000007789 gas Substances 0.000 description 1
- 238000002290 gas chromatography-mass spectrometry Methods 0.000 description 1
- 239000008187 granular material Substances 0.000 description 1
- 229910001385 heavy metal Inorganic materials 0.000 description 1
- 150000002430 hydrocarbons Chemical class 0.000 description 1
- 239000011261 inert gas Substances 0.000 description 1
- 239000000976 ink Substances 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 230000003834 intracellular Effects 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 239000004571 lime Substances 0.000 description 1
- 235000020778 linoleic acid Nutrition 0.000 description 1
- 238000000622 liquid--liquid extraction Methods 0.000 description 1
- 235000012680 lutein Nutrition 0.000 description 1
- 239000001656 lutein Substances 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 238000010907 mechanical stirring Methods 0.000 description 1
- 125000004492 methyl ester group Chemical group 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 235000010755 mineral Nutrition 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000006011 modification reaction Methods 0.000 description 1
- 238000000329 molecular dynamics simulation Methods 0.000 description 1
- 239000012299 nitrogen atmosphere Substances 0.000 description 1
- 235000021315 omega 9 monounsaturated fatty acids Nutrition 0.000 description 1
- 235000020665 omega-6 fatty acid Nutrition 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- MYMOFIZGZYHOMD-UHFFFAOYSA-N oxygen Chemical compound O=O MYMOFIZGZYHOMD-UHFFFAOYSA-N 0.000 description 1
- 238000010979 pH adjustment Methods 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 235000020232 peanut Nutrition 0.000 description 1
- 230000035515 penetration Effects 0.000 description 1
- 238000005191 phase separation Methods 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N phosphorus Chemical class [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 230000000704 physical effect Effects 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920003023 plastic Polymers 0.000 description 1
- 239000002574 poison Substances 0.000 description 1
- 239000002798 polar solvent Substances 0.000 description 1
- 229920001296 polysiloxane 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
- 238000003825 pressing Methods 0.000 description 1
- 108060006613 prolamin family Proteins 0.000 description 1
- XXRYFVCIMARHRS-UHFFFAOYSA-N propan-2-yl N-dimethoxyphosphorylcarbamate Chemical compound COP(=O)(OC)NC(=O)OC(C)C XXRYFVCIMARHRS-UHFFFAOYSA-N 0.000 description 1
- 239000001294 propane Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 238000009877 rendering Methods 0.000 description 1
- 230000000717 retained Effects 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 239000012266 salt solution Substances 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 235000021391 short chain fatty acids Nutrition 0.000 description 1
- 150000004666 short chain fatty acids Chemical class 0.000 description 1
- 239000000741 silica gel Substances 0.000 description 1
- 229910002027 silica gel Inorganic materials 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- 238000002791 soaking Methods 0.000 description 1
- 239000000344 soap Substances 0.000 description 1
- KEAYESYHFKHZAL-UHFFFAOYSA-N sodium Chemical compound [Na] KEAYESYHFKHZAL-UHFFFAOYSA-N 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- 239000008347 soybean phospholipid Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
- 239000008117 stearic acid Substances 0.000 description 1
- 239000010959 steel Substances 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- DKGAVHZHDRPRBM-UHFFFAOYSA-N t-BuOH Chemical compound CC(C)(C)O DKGAVHZHDRPRBM-UHFFFAOYSA-N 0.000 description 1
- 239000004753 textile Substances 0.000 description 1
- 229910052718 tin Inorganic materials 0.000 description 1
- ATJFFYVFTNAWJD-UHFFFAOYSA-N tin hydride Chemical compound [Sn] ATJFFYVFTNAWJD-UHFFFAOYSA-N 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 238000004642 transportation engineering Methods 0.000 description 1
- 229960002415 trichloroethylene Drugs 0.000 description 1
- XSTXAVWGXDQKEL-UHFFFAOYSA-N triclene Chemical group ClC=C(Cl)Cl XSTXAVWGXDQKEL-UHFFFAOYSA-N 0.000 description 1
- 238000000108 ultra-filtration Methods 0.000 description 1
- 239000010913 used oil Substances 0.000 description 1
- 238000005292 vacuum distillation Methods 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 229930003231 vitamins Natural products 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 239000003021 water soluble solvent Substances 0.000 description 1
- 239000010497 wheat germ oil Substances 0.000 description 1
- 235000010930 zeaxanthin Nutrition 0.000 description 1
- 239000001775 zeaxanthin Substances 0.000 description 1
- 229940043269 zeaxanthin Drugs 0.000 description 1
- 239000005019 zein Substances 0.000 description 1
- OENHQHLEOONYIE-JLTXGRSLSA-N β-Carotene Chemical compound CC=1CCCC(C)(C)C=1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C OENHQHLEOONYIE-JLTXGRSLSA-N 0.000 description 1
- VZCCETWTMQHEPK-QNEBEIHSSA-N γ-Linolenic acid Chemical compound CCCCC\C=C/C\C=C/C\C=C/CCCCC(O)=O VZCCETWTMQHEPK-QNEBEIHSSA-N 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C10—PETROLEUM, GAS OR COKE INDUSTRIES; TECHNICAL GASES CONTAINING CARBON MONOXIDE; FUELS; LUBRICANTS; PEAT
- C10L—FUELS NOT OTHERWISE PROVIDED FOR; NATURAL GAS; SYNTHETIC NATURAL GAS OBTAINED BY PROCESSES NOT COVERED BY SUBCLASSES C10G, C10K; LIQUEFIED PETROLEUM GAS; ADDING MATERIALS TO FUELS OR FIRES TO REDUCE SMOKE OR UNDESIRABLE DEPOSITS OR TO FACILITATE SOOT REMOVAL; FIRELIGHTERS
- C10L1/00—Liquid carbonaceous fuels
- C10L1/02—Liquid carbonaceous fuels essentially based on components consisting of carbon, hydrogen, and oxygen only
- C10L1/026—Liquid carbonaceous fuels essentially based on components consisting of carbon, hydrogen, and oxygen only for compression ignition
-
- C—CHEMISTRY; METALLURGY
- C10—PETROLEUM, GAS OR COKE INDUSTRIES; TECHNICAL GASES CONTAINING CARBON MONOXIDE; FUELS; LUBRICANTS; PEAT
- C10L—FUELS NOT OTHERWISE PROVIDED FOR; NATURAL GAS; SYNTHETIC NATURAL GAS OBTAINED BY PROCESSES NOT COVERED BY SUBCLASSES C10G, C10K; LIQUEFIED PETROLEUM GAS; ADDING MATERIALS TO FUELS OR FIRES TO REDUCE SMOKE OR UNDESIRABLE DEPOSITS OR TO FACILITATE SOOT REMOVAL; FIRELIGHTERS
- C10L1/00—Liquid carbonaceous fuels
- C10L1/04—Liquid carbonaceous fuels essentially based on blends of hydrocarbons
-
- C—CHEMISTRY; METALLURGY
- C10—PETROLEUM, GAS OR COKE INDUSTRIES; TECHNICAL GASES CONTAINING CARBON MONOXIDE; FUELS; LUBRICANTS; PEAT
- C10L—FUELS NOT OTHERWISE PROVIDED FOR; NATURAL GAS; SYNTHETIC NATURAL GAS OBTAINED BY PROCESSES NOT COVERED BY SUBCLASSES C10G, C10K; LIQUEFIED PETROLEUM GAS; ADDING MATERIALS TO FUELS OR FIRES TO REDUCE SMOKE OR UNDESIRABLE DEPOSITS OR TO FACILITATE SOOT REMOVAL; FIRELIGHTERS
- C10L1/00—Liquid carbonaceous fuels
- C10L1/04—Liquid carbonaceous fuels essentially based on blends of hydrocarbons
- C10L1/08—Liquid carbonaceous fuels essentially based on blends of hydrocarbons for compression ignition
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B1/00—Production of fats or fatty oils from raw materials
- C11B1/02—Pretreatment
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B1/00—Production of fats or fatty oils from raw materials
- C11B1/10—Production of fats or fatty oils from raw materials by extracting
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B3/00—Refining fats or fatty oils
- C11B3/008—Refining fats or fatty oils by filtration, e.g. including ultra filtration, dialysis
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B3/00—Refining fats or fatty oils
- C11B3/10—Refining fats or fatty oils by adsorption
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11C—FATTY ACIDS FROM FATS, OILS OR WAXES; CANDLES; FATS, OILS OR FATTY ACIDS BY CHEMICAL MODIFICATION OF FATS, OILS, OR FATTY ACIDS OBTAINED THEREFROM
- C11C3/00—Fats, oils, or fatty acids by chemical modification of fats, oils, or fatty acids obtained therefrom
- C11C3/003—Fats, oils, or fatty acids by chemical modification of fats, oils, or fatty acids obtained therefrom by esterification of fatty acids with alcohols
-
- 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
- C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
- C12P7/6436—Fatty acid esters
- C12P7/6445—Glycerides
- C12P7/6472—Glycerides containing polyunsaturated fatty acid [PUFA] residues, i.e. having two or more double bonds in their backbone
-
- 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
Abstract
Disclose a method of isolating chlorophylls and omega-3 rich oil from algae, comprising: a. dewatering substantially intact algal cells to make an algal biomass; b. adding a first ethanol fraction to the algal biomass in a ratio of about 1 part ethanol to about 1 part algal biomass by weight; c. separating a first substantially solid biomass fraction from a first substantially liquid fraction comprising proteins; d. combining the first substantially solid biomass fraction with a second ethanol fraction in a ratio of about 1 part ethanol to about 1 part solids by weight; e. separating a second substantially solid biomass fraction from a second substantially liquid fraction comprising polar lipids; f. combining the second substantially solid biomass fraction with a third ethanol solvent fraction in a ratio of about 1 part ethanol to about 1 part substantially solid biomass by weight; g. separating a third substantially solid biomass fraction from a third substantially liquid fraction comprising neutral lipids, including omega-3 fatty acids, carotenoids, and chlorophyll, wherein the third substantially solid biomass fraction comprises carbohydrates; and h. isolating at least one of carotenoids, chlorophyll, and omega-3 fatty acids from the third substantially liquid fraction. separating a first substantially solid biomass fraction from a first substantially liquid fraction comprising proteins; d. combining the first substantially solid biomass fraction with a second ethanol fraction in a ratio of about 1 part ethanol to about 1 part solids by weight; e. separating a second substantially solid biomass fraction from a second substantially liquid fraction comprising polar lipids; f. combining the second substantially solid biomass fraction with a third ethanol solvent fraction in a ratio of about 1 part ethanol to about 1 part substantially solid biomass by weight; g. separating a third substantially solid biomass fraction from a third substantially liquid fraction comprising neutral lipids, including omega-3 fatty acids, carotenoids, and chlorophyll, wherein the third substantially solid biomass fraction comprises carbohydrates; and h. isolating at least one of carotenoids, chlorophyll, and omega-3 fatty acids from the third substantially liquid fraction.
Description
CROSS REFERENCE TO RELATED ATIONS
This application claims the benefit under 35 U.S.C. §120 of US. Patent Application
No. 13/149,595, filed May 31, 2011, entitled Methods of Producing Biofuels, Chlorophyiis and
Carotenoids, which is a continuation~in—part of and claims the benefit of US. Application No
13/081,221, filed April 6, 2011, entitled s of and s for Isolating Nutraceutical
Products from Algae, which claims ty to US. Provisional Application No. 61/321,290,
filed April 6, 2010, entitled Extraction with Fractionation of Oil and Proteinaceous Material
from Oleaginous Material, and US. Provisional Application No. 61/321,286, filed April 6,
2010, entitled Extraction With Fractionation of Oil and Co-Products from Oleaginous Material,
the entire contents of which are hereby incorporated by reference herein.
FIELD OF THE INVENTION
The invention is concerned with extracting and fractionating algal ts,
ing, but not limited to, oils and ns. More specifically, the systems and methods
described herein utilize step extraction and fractionation with a slightly nonpolar solvent
process wet algal biomass.
BACKGROUND OF THE INVENTION
Petroleum is a natural resource composed primarily of hydrocarbons. Extracting
petroleum oil from the earth is expensive, ous, and often at the expense of the
environment. Furthermore, world wide reservoirs of oil are dwindling y. Costs also
accumulate due to the transportation and processing required to convert petroleum oil into
usable fuels such as gasoline and jet fuel.
Algae have gained a significant importance in recent years given their ability to
produce lipids, which can be used to produce sustainable biofuel. This ability can be exploited
to produce renewable fuels, reduce global climate change, and treat
wastewater. Algae’s
superiority as a biofuel feedstock arises from a variety of factors, including high
per-acre
productivity ed to typical terrestrial oil crop plants, non-food based feedstock
resources,
use of otherwise non-productive, non-arable land, ation of
a wide variety ofwater sources
W0 2012/138438 PCT/U52012/027537
, brackish, saline, and wastewater), production of both biofuels and valuable co—products
such as noids and chlorophyll.
Several thousand species of algae have been screened and studied for lipid
tion worldwide over the past several decades. Of these, about 300 species rich in lipid
production have been identified. The lipid composition and content vary at different stages of
the life cycle and are ed by environmental and culture conditions. The strategies and
approaches for extraction are rather different depending on individual algal species/strains
employed because of the considerable variability in biochemical composition and the physical
properties of the algae cell wall. Conventional physical extraction processes, such as extrusion,
do not work well with algae given the thickness of the cell wall and the small size (about
2 to
about 20nm) of algal cells. Furthermore, the large amounts of polar lipids in algal oil, as
compared to the typical oil recovered from seeds, lead to refining issues.
Upon ting, typical algal concentrations in cultures range from about 01-10
”/0 (w/v). This means that as much as 1000 times the amount of water
per unit weight of algae
must be removed before attempting oil extraction. Currently, existing oil extraction methods
for oleaginous materials strictly require almost completely dry feed to improve the yield
quality of the oil extracted. Due to the amount ofenergy required to heat the algal mass to dry
it sufficiently, the algal feed to l
process is rendered uneconomical. Typically, the feed is
extruded 0r flaked at high temperatures to enhance the extraction. These
steps may not work
with the existing equipment due to the single cell micrometric nature of algae.
Furthermore,
algal oil is very unstable due to the ce of double bonded long chain fatty acids. The high
temperatures used in conventional extraction methods cause degradation ofthe oil, thereby
increasing the costs of such s.
It is known in the art to extract oil from dried algal
mass by using hexane as a
solvent. This process is energy intensive. The use of heat to dry and hexane
to extract
produces product of lower quality as this type of processing causes lipid and protein
ation.
Algal oil extraction can be classified into two types: disruptive or non-disruptive
methods.
Disruptive methods e cell lies by mechanical, thermal, enzymatic or chemical
methods. Most disruptive methods result in emulsions, requiring
an expensive cleanup
s. Algal oils n a large percentage of polar lipids and proteins which enhance the
W0 2012/138438 PCT/U52012/027537
emulsification of the neutral . The emulsification is further stabilized by the nutrient and
salt components left in the solution. The emulsion is a complex mixture, containing neutral
lipids, polar lipids, proteins, and other algal products, which extensive g processes to
isolate the neutral lipids, which are the feed that is converted into biofucl.
Non—disruptive methods provide low yields. Milking is the use of solvents or
chemicals to extract lipids from a growing algal culture. While sometimes used to extract algal
products, milking may not work with some species of algae due to solvent toxicity and cell
wall disruption. This complication makes the development of a generic
process difficult.
rmore, the volumes of solvents required would be astronomical due to the maximum
attainable concentration of the t in the medium.
hase extractions would require extensive distillations, using x solvent
mixtures, and necessitating mechanisms for solvent recovery and recycle. This makes such
tions impractical and uneconomical for use in algal oil technologies.
Accordingly, to me these deficiencies, there is a need in the art for improved
methods and s for extraction and fiactionating algal products, in particular algal oil,
algal proteins, and algal carotenoids.
BRIEF SUMMARY OF THE ION
ments described herein relate generally to systems and methods for
extracting lipids ofvarying polarities from an oleaginous material, including for example, an
algal biomass. In particular, embodiments described herein concern extracting lipids of
varying polarities from an algal biomass using solvents of varying polarity and/or a series of
membrane filters. In some embodiments, the filter is a mierofilter.
In some embodiments of the invention, a single solvent and
water are used to
extract and fractionate components present in an nous material. In other embodiments,
these components include, but are not limited to, ns, polar lipids, and neutral
lipids. In
still other embodiments, more than one solvent is used. In still other embodiments, a mixture
of solvents is used.
In some ments, the methods and systems described herein
are useful for
extracting coproducts of lipids from oleaginous material. Examples of such coproducts
include, without tion, proteinaceous material, chlorophyll, and carotenoids.
W0 38438 PCT/U52012/027537
Embodiments of the t invention allow for the simultaneous extraction and fractionation
of algal products from algal s in a manner that allows for the production of both fiiels
and nutritional products.
In another embodiment of the ion, a method of isolating nutraeeuticals
ts from algae is provided.
In a further embodiment of the invention, a method of ing carotenoids and
omega-3 rich oil from algae includes dewatering substantially intact algal cells to make an algal
biomass and adding a first ethanol fraction to the algal biomass in a ratio of about 1 part
ethanol to about 1 part algal s by weight. The method also includes separating
a first
substantially solid biomass fraction from a first substantially liquid fraction sing
proteins and combining the first substantially solid biomass fraction with a second ethanol
fraction in a ratio of about 1 part ethanol to about 1 part solids by weight. The method further
includes separating a second substantially solid biomass fraction from
a second substantially
liquid fraction comprising polar lipids and combining the second substantially solid biomass
fraction with a third ethanol t fraction in a ratio of about 1 part ethanol to about
1 part
substantially solid biomass by weight. The method also includes separating a third
substantially solid biomass on from a third substantially liquid fraction comprising l
lipids, wherein the third substantially solid biomass fraction comprises ydrates and
separating the neutral lipids into carotenoids and omega-3 rich oil.
In yet another embodiment of the invention, the method also includes isolating
carotenoids, omega—3 rich oil, carbohydrates and polar lipids; isolating the polar lipids from
components that are not polar lipid components; and/or isolating carotenoids from non—
carotenoid components.
In still a further embodiment of the invention, the method also includes
processing
the polar lipids into at least one of lubricants, detergents, and food additives.
[0020} In another embodiment of the invention, at least
one of the first, second, and third
solvent sets comprises an alcohol. Optionally, the alcohol is ethanol.
Embodiments ofthe present in invention include a method of isolating phylls
and omega-3 rich oil from algae, comprising dewatering ntially intact
algal cells
suspended in a fluid medium to generate an algal biomass; extracting a substantially liquid
fraction comprising neutral lipids, carotenoids, and chlorophylls from the algal
biomass, the
W0 2012/138438 PCT/U82012/027537
neutral lipids including omega-3 fatty acids; and separating the carotenoids and chlorophylls
from the neutral lipids. 1n some ments, the separating is carried out by at least one of
adsorption and membrane diafiltration.
In other embodiments, the methods further comprise esterifying the l lipids
with a st in the presence of an alcohol, and separating a water e fraction comprising
in from a water insoluble on comprising fuel esters. In still other embodiments, the
separating is d out by adsorption with an adsorbent al. In still others, the separating
is carried out by adsorption with a clay. In some embodiments, the clay is selected from the
following group consisting of bleaching clay, bentonite, and fuller’s earth.
Further embodiments of the invention include a method of isolating chlorophylls
and omega-3 rich oil from algae, comprising dewatering substantially intact algal cells
to make
an algal biomass; adding a first ethanol fraction to the algal biomass in
a ratio of about 1 part
ethanol to about 1 part algal biomass by weight; separating a first substantially solid biomass
fraction from a first substantially liquid fraction comprising proteins; combining the first
substantially solid biomass on with a second ethanol on in a ratio of about 1 part
ethanol to about 1 part solids by weight; separating a second substantially solid biomass
fraction from a second substantially liquid fraction comprising polar lipids; combining the
second substantially solid biomass fraction with a third ethanol solvent fraction in
a ratio of
about 1 part ethanol to about 1 part substantially solid biomass by weight; separating
a third
substantially solid biomass fraction from a third substantially liquid fraction comprising neutral
lipids, including omega-3 fatty acids, carotenoids, and chlorophyll, wherein the third
substantially solid biomass fraction comprises carbohydrates; and isolating at least one of
carotenoids, chlorophyll, and omega-3 fatty acids from the third substantially liquid fraction.
In some embodiments of the method, at least one of the first, second, and third solvent
sets
comprises an ethanol. In others, at least one of the first, second, and third solvent sets
ses an alcohol.
In yet other ments, the method further comprises esterifying the
neutral
lipids with a st in the presence of an alcohol, and separating a water soluble fraction
comprising glycerin from a water insoluble fraction comprising fuel . In still other
embodiments, the method further comprises ling the fuel esters under vacuum to obtain a
C16 or shorter fuel ester fraction, a C16 or longer fuel ester fraction, and
a e comprising
omega—3 fatty acids. Some embodiments r comprise deoxygenating the C16 or shorter
W0 2012/138438 PCT/U82012/027537
fuel ester fraction to obtain a jet fuel blend stock and/or the C16 or longer fuel fraction
obtain a diesel blend stock. In still other embodiments, the isolating is d out by
adsorption with an ent material. In still other embodiments, the isolating is carried out
by adsorption with a clay. In some ments, the clay is selected from the following
group
consisting of bleaching clay, bentonite, and fuller’s earth.
BRIEF PTION OF THE DRAWINGS
FIG. IA is a flowchart of steps involved in a method according to an ary
embodiment of the present disclosure.
is a schematic diagram of an exemplary embodiment of
a dewatering
s according to the present disclosure.
is a schematic diagram of an exemplary ment of
an extraction system
according to the present disclosure.
is a comparative graph showing Sohxlet extraction of freeze dried algae
s using an array of solvents encompassing the complete polarity
range showing
maximum non-disruptive algae oil extraction efficiency and the effect of polarity
on the polar
and non—polar lipids extraction.
&B are graphic representations showing neutral lipids (A) Purity and (B)
Recovery in the two step solvent extraction process using methanol and petroleum ether at
three different temperatures.
&B are graphs showing neutral lipids (A) Purity and (B) Recovery in
two step solvent tion process using
aqueous methanol and petroleum ether at three
different temperatures.
is a graph showing lipid
recovery in the two step solvent extraction s
using aqueous methanol and petroleum other at three different temperatures.
is a graph showing the effect of solvents to solid biomass ratio
on lipid
recovery.
is a graph showing the efficacy of different
aqueous tion solutions in a
single step extraction recovery of aqueous methanol on dry biomass.
PCT/U82012/027537
is a graph showing the effect of multiple step methanol tions
on the
cumulative total lipid yield and the neutral lipids .
is a graph showing the cumulative
recovery of lipids using wet biomass and
ethanol.
is a graph showing a comparison of the extraction times of the microwave
assisted extraction and conventional tion systems.
A is a flowchart of steps involved in a method according to
an exemplary
embodiment ofthe present sure which incorporates a step of protein extraction. All of
the units in A are in pounds.
B is a flowchart of steps involved in an exemplary extraction
process
according to the present disclosure.
is a rt and mass balance diagram describing
one ofthe
embodiments ofthe t invention wherein 1000 lbs. of algal biomass
was sed
through extraction and fractionation in order to separate neutral lipids, polar lipids, and protein
from the algal biomass.
is a rt describing one of the ments of the
present invention
wherein an algal mass can be processed to form various products.
is a flowchart describing one of the embodiments of the
present invention
wherein algae neutral lipids are processed to form various products.
is a flowchart describing one of the embodiments of the
present invention
wherein algae neutral lipids are processed to form fuel products.
is a flowchart describing one of the embodiments of the
present invention
wherein algae proteins are selectively extracted from a freshwater algal biomass.
is a flowchart describing one of the embodiments of the
present ion
wherein algae proteins are ively extracted from a saltwater algal s.
is a flowchart describing one of the embodiments of the
present invention
wherein a selected algae protein is extracted from a saltwater
or freshwater algal biomass.
is a flowchart describing one of the embodiments of the
present invention
n a selected algae protein is extracted from a saltwater
or freshwater algal biomass.
W0 38438 2012/027537
is a photograph showing Scenedescemus
Sp. cells before and after
extraction using the methods described . The cells are substantially intact both before
and after tion.
A is a flowchart of steps involved in an method of chlorophyll extraction.
B is a schematic diagram of an exemplary embodiment of chlorophyll
extraction of the present disclosure.
DETAILED DESCRIPTION
Definitions
The term “conduit” or any variation thereof, as used herein, includes
any structure
through which a fluid may be conveyed. Non—limiting es of conduit include pipes,
tubing, channels, or other enclosed structures.
The term “reservoir” or any variation thereof, as used herein, includes
any body
structure capable of retaining fluid. Non-limiting examples of reservoirs e ponds, tanks,
lakes, tubs, or other similar structures.
The term “about” or “approximately,” as used herein,
are defined as being close to
as understood by one of ordinary skill in the art, and in
one non-limiting embodiment the terms
are defined to be within 10%, preferably within 5%,
more preferably within 1%, and most
preferably within 0.5%.
The terms “inhibiting” or “reducing” or
any variation of these terms, as used herein
includes any measurable decrease or complete inhibition to achieve
a desired result.
The term tive,” as used herein, means adequate to accomplish
a desired,
expected, or intended result.
The use of the word “a” or “an” when used in conjunction with the
term
“comprising” herein may mean “one,” but it is also consistent with the meaning of “one
“
more, at least one,” and “one or more than one.”
The term “or” as used herein, means “and/or” unless explicitly indicated
to refer to
alternatives only or the alternatives are ly exclusive, although the
disclosure ts a
definition that refers to only alternatives and “and/or.”
W0 2012/138438 PCT/U82012/027537
The use of the term “wet” as used herein, is used to mean containing about 50% to
about 99.9% water content. Water content may be located either intracellularly or
elluarly.
The use of the term “solvent set” as used herein, is used to mean composition
comprising one or more solvents. These solvents can be amphipathic (also known as
hilic or slightly nonpolar), hydrophilic, or hydrophobic. In some embodiment, thcsc
solvents are water le and in others, they are immiscible in water. Non—limiting example
of ts that may be used to practice the methods of the instant invention include methanol,
ethanol, isopropanol, acetone, ethyl acetate, and acetonitrile, alkanes (hexane, pentane, heptane,
octane), esters (ethyl acetate, butyl e), ketones (methyl ethyl ketone (MEK), methyl
isobutyl ketone (MIBK)), aromatics (toluene, benzene, cyclohexane, tetrahydrofuran),
haloalkanes (chloroform, trichloroethylene), ethers yl ether), and mixtures (diesel, jet
fuel, gasoline).
The term “oil” as used herein includes itions containing neutral lipids and
polar lipids. The terms “algae oil” and “algal oil” as used herein are used hangeably.
The term “diffusate” or “permeate" as used herein
may refer to material that has
passed through a separation device, including, but not limited to a filter or membrane.
The term “retentate” as used herein
may refer to material that remains after the
ate has passed h a separation device.
As used herein, the words “comprising” (and
any form of comprising, such as
“comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”),
“including” (and any form of including, such as “includes” and de”), or ining”
(and any form of containing, such as “contains” and “contain”) are inclusive or open—ended and
do not exclude additional, unrecited elements or method
steps.
The term “polar lipids” or any variation thereof, as used herein, includes, but
is not
limited to, phospholipids and glycolipids.
The term “neutral lipids” or any variation thereof, as used herein, includes, but
not limited to, triglycerides, diglycerides, monoglycerides, carotenoids,
waxes, sterols.
The term “solid phase” as used herein refers to a collection of material that is
generally more solid than not, and is not ed to mean that all of the material in the phase is
solid. Thus, a phase having a substantial amount of solids, while retaining some liquids, is
encompassed within the meaning of that term. Meanwhile, the term “liquid phase”, as used
herein, refers to a collection of material that is generally more liquid than not, and such
collection may e solid materials.
The term “biodiesel” as used herein refers to methyl or ethyl esters of fatty acids
derived from algae.
The term eeutical” as used herein refers to a food product that provides health
and/or medical benefits. Non-limiting examples include carotenoids, carotenes, xanthophylls
such as zeaxanthin, astaxanthin, and lutein.
The term “biofuel” as used herein refers to fuel derived from biological
source.
Non-limiting examples include biodiesel, jet fuel, diesel, jet fuel blend stock and diesel blend
stock.
The term “impurities”, when used in tion with polar lipids,
as used herein,
refers to all components other than the products of interest that are eoextracted
or have the
same ties as the product of interest.
The term “lubricants”, when used in connection with polar lipids,
as used herein
refers to hydrotreated algal lipids such as C16-C20 alkanes.
The term gents”, when used in connection with polar lipids,
as used herein
refers to glycolipids, phospholipids and derivatives thereof.
The term “food additives”, when used in connection with polar lipids,
as used
herein refers to soy lecithin substitutes or phospholipids derived from algae.
The term “non—glycerin matter” as used herein refers to
any impurity that separates
with the glycerin fraction. A further clean
up step will remove most of what is present in order
to produce pharmaceutical grade glycerin.
The term urated fatty acids” as used herein refers to fatty acids with
at least
one double carbon bond. Non-limiting examples ofunsaturated fatty acids include palmitoleie
acid, margaric acid, stearic acid, oleie acid, octadeeenoic acid, linoleic acid, gamma-linoleic
acid, alpha linoleic acid, dic acid, noic acid, homogamma linoleic acid, arachidonie
acid, eicosapenenoic acid, behenic, doeosadienoic acid, heneieosapentaenoic, docosatetraenoie
acid. Fatty acids having 20 or more carbon atoms in the backbone are generally referred to as
PCT/U52012/027537
“long chain fatty acids”. The fatty acids having 19 or fewer carbon atoms in the backbone are
generally referred to as “short chain fatty acids”.
Unsaturated long chain fatty acids include, but are not limited to, omega-3 fatty
acids, omega-6 fatty acids, and omega-9 fatty acids. The term “omega—3 fatty acids” as used
herein refers to, but is not limited to the fatty acids listed in Table 1.
"Z'iiléiél'i‘i"iii’i7""éi'é'c'iééifiéfiéié'éaéi"""""" """""3:
The term “jet fuel blend stock” as used herein refers to alkanes with the carbon
chain lengths riate for use as jet fuels.
The term “diesel blend stock” as used herein refers to alkanes with the carbon
chain
lengths appropriate for use as diesel.
The term l feed” as used herein refers to algae-derived substances that
can be
consumed and used to provide nutritional support for an animal.
The term “human food” as used herein refers to algae—derived substances that
be consumed to provide nutritional support for people. Algae-derived human food products
can contain essential nutrients, such as carbohydrates, fats, proteins, vitamins,
or minerals.
The term “bioremediation” as used herein refers to use of algal growth to
remove
pollutants, such as, but not limited to, nitrates, phosphates, and heavy , from industrial
wastewater or municipal wastewater.
The term “wastewater” as used herein refers to industrial wastewater or municipal
wastewater that contain a variety of contaminants or pollutants, including, but not limited to
nitrates, phosphates, and heavy metals.
The term “enriched”, as used herein, shall mean about 50% or greater content.
The term “substantially”, as used herein, shall mean mostly.
The term “globulin proteins” as used herein refers to salt soluble proteins.
The term in proteins” as used herein refers to water soluble proteins.
The term “glutelin proteins” as used herein refers to alkali soluble proteins.
The term “prolamin proteins” as used herein refers to alcohol soluble proteins.
miting examples ofprolamin ns are n, zein, hordein, avenin.
The term “algal culture” as used herein refers to algal cells in e medium.
The term “algal biomass” as used herein refers to an at least partially dewatered
algal culture.
The term “dewatered” as used herein refers to the removal of at least
some water.
The term “algal paste” as used herein refers to a partially dewatered algal culture
having fluid properties that allow it to flow. Generally an algal paste has a water content of
about 90%.
The term “algal cake” as used herein refers to a partially dewatered algal culture
that lacks the fluid properties of an algal paste and tends to clump. Generally an algal cake has
a water content of about 60% or less.
Saltwater algal cells include, but are not d to, marine and brackish algal
species. Saltwater algal cells are found in nature in bodies ofwater such
as, but not limited to,
seas, oceans, and estuaries. Non-limiting es of saltwater algal species include
Nannochloropsz’s sp., Dumzlz’ella Sp.
PCT/U82012/027537
Freshwater algal cells are found in nature in bodies of water such as, but not limited
to, lakes and ponds. Non-limiting examples of freshwater algal species include Scendescemus
5p. , Haemotococcus Sp.
Non—limiting examples ofmicroalgae that can be used with the methods of the
invention are members of one of the following divisions: Chlorophyta, Cyanophyta
(Cyanobaeteria), and Heterokontophyta. In certain ments, the microalgae used with the
s of the invention are members of one of the following s: Bacillarz’ophyceae,
gmatophyceae, and Chrysophyceae. In certain embodiments, the microalgae used with the
s ofthe invention are s of one of the ing
genera: Nannochloropsz’s,
Chlorella, Dunalz‘ella, Scenedesmus, Selenastrum, Oscillatoria, Phormz'a’z‘um, Spirulz'na,
Amphora, and Ochromonas.
Non-limiting examples of microalgae species that can be used with the methods of
the present invention include: rhes orientalz's, Agmenellum
$1717., Amphipmra ne,
Amp/mm coffézj‘brmz‘s, Amp/mm coffézj‘brmis var. Zinea, a cofl‘eiformis var. punctata,
Amphora cofi’eiformis var. i, Amp/20m cofi’ez'formis var. tenuis, Amphora delicatl'ssz'ma,
Amp/10m delicarz‘ssz’ma var. . capitata, Amphora S11, Anabaena, Ankz'stmdesmus,
Ankz‘strodesmusfalcatus, Boekelovz’a hooglandz'i, Borodinella Sp., Botryococcus braum'z',
Bonyococcus sudetz'cus, Bracteococcus minor, Bracteococcus medionucleatus, Carterz'a,
Chaetoceros is, Chaetoceros muellerz‘, Chaetoceros muellerz' var. subsalsum,
Chaetoceros sp., Chlamydomas perigranulata, Chlorella anitrata, lla antarctica,
Chlorella aureovz'ridz's, Chlorella Candida, Chlorella capsulate, Chlorella desiccate, Chlorella
ellipsoidea, Chlorella emersanii, Chlorellafusca, Chlorellafusca var. vacuolata, lla
glucotropha, Chlorella infitsz‘onum, Chlorella infiisz‘onum var. actophz'la, Chlorella infirsz'onum
var. auxenophila, Chlorella kessleri, Chlorella lobaphora, lla luteoviridis, Chlorella
luteovz‘rz'dis var. aureovz‘rz’dz‘s, Chlore/la luteovz'ridz‘s var. lutescens, Chlorella miniata,
Chlorella minutissz'ma, Chlorella mutabilis, lla nocturna, Chlorella ovalis, Chlorella
paiva, Chlorella plwtophz'la, Chlorella pringsheimii, Chlarella protothecoz‘des, Chlorella
protothecoz‘a’es var. acidicola, Chlorella regularz's, Chlorella ris var. minima, Chlorella
regularis var. umbrz’cata, Chlorella reisz'gliz‘, Chlorella saccharophz‘la, Chlorella saccharophz'la
var. ellipsoidea, Chlorella salina, lla simplex, Chlorella sorokz'm‘ana, Chlorella
Chlorella sphaerica, lla stigmatophora, Chlorella vannz‘e/lz‘z', lla vulgarz's,
Chlorella vulgarisfo. terria, Chlorella vulgaris var. autotrophz‘ca, Chlorella vulgaris
var.
W0 2012/138438 PCT/U52012/027537
vz'rz‘dz'S, Chlorella vulgaris var. vulgaris, lla vulgaris var. vulgarisfo. tertia, Chlorella
vulgariS var. vulgarisfo. viridz‘s, Chlorella xanthella, Chlorella zofingierzSiS, Chlorella
trebouxz'oz'des, Chlorella vulgariS, Chlorococcum infusionum, Chlorococcum Sp,
gorzz'um, Chroomonas Sp, CthSOSphaera Sp, Cricosphaera Sp, Crwthecodz’m’um
colmz'z', Cryptomonas Sp, Cyclotella cryptica, ella menegkim'ana, Cyclotella Sp,
Dzmalz‘ella Sp, 'ella bardawil, Dunaliella bz'oculata, Dunaliella granulate, ella
maritime, Dunalz’ella mimlta, Dunalz'elia parva, Dzmalz’ella pez‘rcez‘, Dunalz’ella primolecta,
Dunaliella , Dzmaliella terricola, Dzmalz'ella terliolecta, Dunalz'ella viridis, Dunalz’ella
terliolecta, Eremosphaera viridz's, Eremasphaera Sp, oidon Sp, Euglena Spp, Francez'a
Sp, Fragilarz'a crotonensz’s, Fragilarz’a Sp, Gleocapsa Sp, Gloeothamniorz Sp, Haematococcus
pluvz'alz'S, Hymenomonas Sp, ysz's ajj‘. galbana, [sochrysz's galbana, Lepocinclis,
filicractinium, Alicractiniwn, Alonoraphz’dium minutum, Monoraphidium Sp, Nannochloris Sp,
Nannochloropsis salina, Nannochlampsz‘s Sp, Navicula accepzata, Navicula biskamerae,
Navicula pseudotenelloz‘des, Navz'cula pellz'culosa, Navicula saprophila, Navicula
Nephrochloris Sp, Nephroselmis Sp, Nz'tschia communis, Nitzschz'a alexandrz‘na, Nitzschia
closterz'um, hia communis, Nitzschz‘a dissipata, Nitzschz‘afiustulum, Nitzschia
chz’ana, Nitzschz’a inconspicua, Nitzschz'a intermedia, Nitzschz‘a microcephala, Nitzschz'a
pusilla, Nitzschia pusilla elliptica, Nitzschia pusz‘lla monoenSZS, Nitzschia quadrangular,
Nitzschz‘a Sp, Ochromonas Sp, OocyStz'S parva, Oocystis pusz'lla, Oocystis
Sp, atorz'a
limaetz’ca, Oscz'llatorz'a Sp, Oscillatoria subbrevis, Parachlorella kesslerz', Pascheria
hila, Pavlova Sp, Phaeodacgzlum tricomutum, Phagus, Phormz’dium, Platymonas Sp,
Pleurochrysis carterae, chrysis e, Pleurochljzsz‘s Sp, Protoz‘heca wickel‘hamii,
Prototheca Stagnora, Prototheca portorz'censz's, heca morzformis, Prototheca zap/ii,
Pseudochlorella aquatica, monas Sp, rys, Rhodococcus
opacus, noz’a’
chrysophyte, Scenedesmus armatuS, Schizochytrium, Spirogyra, Spirulina platenSis,
Stz'chococcus Sp, Synechococcus Sp, ocystz’sf, TageteS erecta, T
ageteS patula,
Tetraedron, Tetraselmz‘s Sp, Tetraselmis suecz’ca, Thalassz'osz'ra weissflogz‘z’, and Viridz'ella
icz'ana.
In other embodiments, the s can be plant material, including but not limited
to soy, corn, palm, camelina, jatropha, canola, coconut, peanut, safflower, cottonseed, linseed,
sunflower, rice bran, and olive.
Systems and methods for extracting lipids and coproducts (e.g., proteins) of varying
polarity from a wet oleaginous material, including for e, an algal biomass, are disclosed.
In particular, the methods and systems bed herein concern the ability to both t and
fractionate the algae components by doing sequential extractions with a hydrophilic
t/water mixture that becomes progressively less polar (z'.e., water in solvent/water ratio is
progressively reduced as one proceed from one extraction step to the next). In other words, the
interstitial solvent in the algae (75% of its weight) is initially water and is replaced by the
slightly nonpolar solvent gradually to the azeotrope of the organic solvent. This results in the
extraction of ents soluble at the ty developed at each step, thereby leading to
simultaneous fractionation of the extracted components. Extraction ofproteinaceous
byproducts by acid ng and/or alkaline extraction is also disclosed.
In some embodiments of the invention, a single solvent and water are used to
extract and fractionate components present in an oleaginous material. In other embodiments, a
solvent set and water are used to extract and fractionate components present in an oleaginous
al. In some embodiments the oleaginous material is wet. In other embodiments, the
oleaginous material is algae.
Polar lipid ry depends mainly on its ionic , water solubility, and
location (intracellular, extracellular or membrane bound). Examples ofpolar lipids e, but
are not limited to, phospholipids and glycolipids. Strategies that can be used to separate and
purify polar lipids can roughly be divided into batch or continuous modes. Examples ofbatch
modes include precipitation (pH, organic t), solvent extraction and crystallization.
es of continuous modes include centrifuging, adsorption, foam separation and
precipitation, and membrane technologies (tangential flow ion, diafiltration and
precipitation, ultra filtration).
Other objects, features and advantages of the present invention will become
nt from the following detailed description. It should be understood, however, that the
detailed description and the examples, while indicating specific embodiments of the invention,
are given by way of illustration only. Additionally, it is contemplated that s and
modifications within the range and scope of the invention will become apparent to those skilled
in the art from this detailed description.
Surprisingly, the proposed non-disruptive extraction process results in over 90%
ry. The small amount of polar lipids in the remaining biomass enhances its value when
PCT/USZOIZ/027537
the remaining s is used for feed. This is due, at least in part, to the high long chain
unsaturated fatty acid content of the biomass. In addition, ethanol extracts can further be
directly transesterified. Furthermore, unlike the existing conventional methods, the methods
and systems described herein are generic for
any algae, and enable recovery of a significant
portion of the le components, including polar lipids, in the algae by the use of a water
miscible organic solvent gradient.
The neutral lipid on ed by the use of the
present invention possesses a
low metal content, thereby enhancing stability of the lipid fraction, and reducing
subsequent
sing steps. Metals tend to make neutral lipids unstable due to their ability to catalyze
oxidation. Furthermore, metals inhibit hydrotreating catalysts, necessitating their removal
before a neutral lipid mixture can be refined. The systems and methods sed herein
allow
for the extraction of metals in the protein and/or the polar lipid fractions. This is advantageous
e proteins and polar lipids are not highly affected by metal
exposure, and in some cases
are actually stabilized by metals.
The systems and methods disclosed herein can start with wet biomass, reducing
drying and dewaterin g costs. Compared to conventional tion processes, the disclosed
extraction and fractionation ses should have relatively low ing
costs due to the
moderate temperature and pressure conditions, along with the solvent recycle.
Furthermore,
conventional extraction processes are cost prohibitive and cannot meet the demand
ofthe
Another aspect of the systems and methods described herein is the ability
accomplish preliminary refining, which is the separation of polar lipids from l lipids
during the extraction process. The differences between algal oil used in exemplary
embodiments and vegetable oils used in previous embodiments include the
percentage of
dual classes of lipids. An exemplary algal crude oil composition is
compared with
vegetable oil shown in Table 2 below:
Table 2
Algal Crude Oil (w/W) Vegetable Oil (W/w)
Neutral lipids 30-90% 90-98%
WO 38438 PCT/U52012/027537
Phospholipids 10-40%
Glycolipids 10-40%
Free fatty acids l-10%
Pigments
Degumming (physical and/or chemical) of vegetable oil is done in order to remove
polar lipids (e.g., glycolipids and phospholipids). Vegetable oil that has been chemically
degummed retains a cant quantity of neutral lipid. This neutral lipid fraction is further
removed from the degummed al using solvent extraction or supercritical/subcritieal fluid
extraction or membrane technology. In contrast, separation of the neutral lipids from an
oleaginous algal biomass is far more difficult than from a vegetable oil feedstock due to the
presence of large quantities of polar lipids lly found in algal oil (see Table 2). This is
because the larger percentage of polar lipids present in algal oil enhances the emulsification of
the neutral lipids. The emulsification is further stabilized by the nutrient and salt
components
left in the solution. The presence of polar lipids, along with metals, results in processing
difficulties for separation and ation of neutral lipids. However, because polar lipids have
an ng market, their recovery would add significant value to the
use of algal oil to generate
fuels.
Polar lipids are surfactants by nature due to their molecular structure and have
huge existing market. Many of the existing technologies for producing polar lipids are raw
material or cost prohibitive. Alternative feedstocks for glycolipids and phospholipids are
mainly algae oil, oat oil, wheat germ oil and vegetable oil. Algae oil typically contains about
—85 % (w/w) polar lipids depending on the species, physiological status of the cell,
culture
conditions, time of harvest, and the solvent utilized for extraction. r, the glycerol
backbone of each polar lipid has two fatty acid
groups attached instead of three in the neutral
lipid triacylglycerol. Transesterification of polar lipids may yield only irds ofthe end
product, z’.e., esterified fatty acids, as compared to that of neutral lipids, on a per mass basis.
Hence, l and ry of the polar lipids would not only be highly beneficial in
W0 2012/138438 PCT/U52012/027537
producing high y biofuels or triglycerides from algae, but also te value-added co-
products glycolipids and phospholipids, which in turn can offset the cost associated with algae-
based biofuel production. The ability to easily recover and fractionate the various oil and
products produced by algae is advantageous to the economic success of the algae oil s.
A further aspect of the methods and systems bed herein is the ability to
extract
proteins from an oleaginous material, such as algal biomass. The methods disclosed herein of
extraction of proteinaceous material from algal biomass comprise a flexible and highly
customizable process of extraction and fractionation. For e, in some ments,
extraction and fractionation occur in a single step, thereby providing a highly efficient
process.
Proteins sourced from such biomass are useful for animal feeds, food ingredients and industrial
products. For e, such proteins are useful in applications such as fibers, adhesives,
coatings, ceramics, inks, cosmetics, textiles, chewing gum, and biodegradable plastics.
Another aspect of the methods and systems described herein involves g the
ratio of algal biomass to solvent based on the components to be ted. In one embodiment,
an algal biomass is mixed with an equal weight of solvent. In another embodiment, an algal
biomass is mixed with a lesser weight of solvent. In yet r embodiment, an algal biomass
is mixed with a greater weight of solvent. In some embodiments, the amount of solvent mixed
with an algal biomass is calculated based on the solvent to be used and the desired polarity
the algal biomass/solvent mixture. In still other embodiments, the algal mass is extracted in
several steps. In an exemplary embodiment, an algal biomass is sequentially extracted, first
with about 50-60% of its weight with a slightly nonpolar, water miscible solvent. Second, the
ing algal solids are extracted using about 70% of the solids’ weight in t. A third
extraction is then med using about 90% ofthe solid’s weight in solvent. Having been
informed of these aspects of the invention, one of skill in the art would be able to
use different
solvents of different polarities by adjusting the ratios of algal biomass and/or solid residuals
the desired polarity in order to selectively t algal ts.
For example, in preferred embodiment, the solvent used is ethanol. Components
may be selectively isolated by g the ratio of solvent. Proteins can be extracted from an
algal biomass with about 50% ethanol, polar lipids with about 80% ethanol, and neutral lipids
with about 95% or greater ethanol. Ifmethanol were to be used, the solvent concentration to
extract proteins from an algal biomass would be about 70%. Polar lipids would require about
90% methanol, and neutral lipids would require about 100% methanol.
W0 2012/138438 PCT/U82012/027537
ments ofthe s and methods described herein exhibit surprising and
cted results. First of all, the recovery/extraction process can be done on a wet biomass.
This is a major economic advantage as exemplary ments avoid the
use of large amounts
of energy required to dry and disrupt the cells. Extraction of neutral lipids from a dry algal
biomass is far more effective using the systems and methods of the
present invention. The
yields ed from the disclosed processes are significantly higher and purer than those
obtained by conventional extractions. This is because conventional extraction frequently
results in emulsions, ing component separations extremely difficult.
Exemplary embodiments may be applied to any algae or gae oleaginous
al. Exemplary embodiments may use
any water-miscible slightly nonpolar solvent,
including, but not limited to, methanol, ethanol, isopropanol, acetone, ethyl acetate, and
acetonitrile. Specific embodiments may use a green renewable solvent, such as ethanol. The
alcohol solvents tested resulted in higher yield and purity of isolated neutral lipids.
Ethanol is
relatively economical to purchase as compared to other solvents disclosed herein. In some
exemplary embodiments, extraction and fractionation can be performed in one step followed by
membrane-based purification ifneeded. The resulting biomass is almost devoid of
water and
can be completely dried with lesser energy than an aqueous algae slurry.
In some exemplary embodiments, the solvent used to extract is ethanol. Other
ments include, but are not d to, cyclohexane, eum ether,
pentane, hexane,
heptane, diethyl ether, toluene, ethyl acetate, form, oromethane, acetone,
acetonitrile, isopropanol, and methanol. In some embodiments, the same solvent is used in
sequential tion steps. In other embodiments, different solvents are used in each
extraction step. In still other embodiments, two or more solvents
are mixed and used in one or
more extraction steps.
In some embodiments of the methods described herein,
a mixture oftwo or more
ts used in any of the extraction steps includes at least
one hydrophilic solvent and at least
one hydrophobic t. When using such a mixture, the hydrophilic solvent
extracts the
material from the biomass via ion. Meanwhile, a relatively small amount of hydrophobic
solvent is used in combination and is involved in a liquid—liquid separation such that
material of interest is concentrated in the small amount of hydrophobic solvent. The
different solvents then form a two-layer system, which can be separated using techniques
known in the art. In such an implementation, the hydrophobic solvent
can be any one or more
W0 2012/138438 PCT/U$2012/027537
of an , an ester, a , an aromatic, a haloalkane, an ether,
or a commercial mixture
(e.g., diesel, jet fuel, gasoline).
In some embodiments, the extraction
processes described herein incorporate pH
ion in one or more steps. Such pH excursion is useful for ing proteinaceous
material. In some embodiments, the pH of the extraction
process is acid (e. g., less than about
). In some embodiments, the pH of the extraction
process is alkaline (e.g., greater than about
).
The use of hexane in conventional extraction procedures contaminates algal
biomass such that coproducts may not be used in food products. Embodiments ofthe
present
invention are superior to those known in the art as they require the
use of far less energy and
render products suitable for use as fuels as well as foodstuffs and nutrient supplements.
It is contemplated that any embodiment discussed in this specification
can be
implemented with respect to any method or system of the invention, and vice versa.
Furthermore, s of the invention can be used to achieve methods of the invention.
DESCRIPTION OF EXEMPLARY MENTS
For solvent extraction of oil from algae the best case scenario is
a solvent which
selectively extracts triacylglycerols (TAG) and leaving all polar lipids and non—TAG neutral
lipids such as waxes, sterols in the algal cell with high recoveries. The second option would be
selectively extract polar lipids and then extract purer neutral lipids devoid of polar lipids,
resulting in high ry. The last option would be to extract all the lipids and achieve
very
high recovery in one or two steps.
Referring now to FIG. IA, a flowchart 100 provides an overview of the steps
involved in exemplary ments ofmethods used in the fractionation and
purification of
lipids from an algae-containing biomass. In a first step 110, algal cells are harvested. In a
subsequent step 120, water is d from algal cells to yield a 10-25% solid biomass. In
step 130, a t-based tion is performed on the biomass and the fractions are
collected. In some embodiments, step 130 will also incorporate pH-based extraction and
fraction collection. y, a solid/liquid phase separation, including, but not limited to
techniques such as filtration, decanting, and centrifugation, may be performed in a step 140 to
in order to separate out smaller lipid components.
W0 2012/138438 PCT/U82012/027537
The algae biomass when harvested in step 110 typically consists of about 1-5 g/L of
total solids. The biomass can be partially dewatered in step 120 using ques including,
but not limited to, dissolved air ion, membrane filtration, flocculation, sedimentation,
filter pressing, decantation or centrifiigation. Dewatering is the l of some, most, or all
of the water from a solid or semisolid substance. Embodiments of the
present invention utilize
dewatering techniques to remove water from a harvested algal biomass. Dewatering can be
carried out using any one of or a combination of
any of the methods described herein, as well
as by any other s known to one of skill in the art.
The dewatered algae biomass resulting from step 120 typically consists of about 10—
% solids. This biomass can then be extracted with water miscible slightly nonpolar solvents
(e.g. , alcohols), in a multistage countercurrent solvent extraction s segregating the
fractions at each stage. This type of process can reduce both capital and operating
expenses.
In some embodiments, the biomass also undergoes acid and/or alkaline tion
to fractionate
protein material.
[0122} In some embodiments, ring of an algal biomass
can be carried out by treating
the ted algal biomass with a t such as ethanol. The algal biomass is then d
to settle out of solution and the liquids may then be d by methods such
as, but not
limited to, siphoning. This novel method of dewatering has lower capital and opcrating costs
than known methods, enables solvent recycling, reduces the cost of drying the biomass,
and has
the added benefit of sing the polarity of the algal biomass prior to beginning
extraction
and/or separation of algal components. In fact, it is theorized that the solvent—based
sedimentation processes described herein are effective, in part, due to the fact that organic
solvents reduce or neutralize the negative charge on the algae surface. In some embodiments
ofthe ion, dewatering methods are ed in order to
remove even more water. In
some embodiments, the addition of solvent during the dewatering
process begins the process of
extraction.
shows an illustrative implementation of a dewatering
process 300. An
algal culture 310 having a final dry weight of about 1 g/L to about 10 g/L (226., 0.1-l% w/w) is
subjected to a water tion process 320. Process 320 can include centrifugation, decanting,
settling, or filtration. In one embodiment, a sintered metal tube filter is used to
separate the
algal biomass from the water of the culture. When using such a filter, the recovered water 330
is recycled directed to other algae cultures. Meanwhile, the algal biomass recovered has been
W0 2012/138438 PCT/U52012/027537
concentrated to an “algae paste” with a algae density as high as about 200 g/L (226., 10-20%
w/w). This concentrated algae paste is then treated with a solvent 340 in a solvent-based
sedimentation process 350.
Sedimentation process 350 involves adding solvent 340 to the algae paste to achieve
a mixture having a weight/weight solvent to biomass ratio of between about 1:1 to about 1:10.
The algae is allowed to settle in a settling vessel, and a solvent / water mixture 360 is removed
by, for example, siphoning and/or decanting. The solvent can be recovered and reused by well-
known techniques, such as lation and/or oration. The remaining wet biomass 370 is
expected to have a solids t of about 30% to about 60% w/w in an alcohol and water
solution.
Solvents ideal for dewatering are industrially common water-soluble solvents with
densities over 1.1 g/mL or below 0.9 g/mL. Examples include isopropanol, acetone,
acetonitrile, t-butyl alcohol, ethanol, methanol, 1-propanol, heavy water (D20), ethylene
glycol, and/or glycerin. If the solvent density is over 1.1 g/mL then the algae biomass would
float rather than create a sediment at the bottom of the settling vessel.
is a tic diagram of an exemplary embodiment of an extraction
system
200. The wet or dry algal biomass is transported using s known in the
art, including,
but not limited to a moving belt, a screw
conveyor, or through extraction chambers. The
t for extraction is ulated from a storage tank assigned to each biomass slot position.
The extraction mixture is filtered, returning the biomass solids back into the slot and the
extract
into the storage tank. The solids on the belt move periodically based on the residence time
requirement for extraction. The extracts in each storage tank may either be replenished at
saturation or continuously ed by fresh solvent. This would also reduce the downstream
processing time and cost drastically. This embodiment comprises a y reservoir 210, a
transport ism 220, a ity of separation devices 241-248 (2.g.
, membrane filtration
devices), a plurality of extraction reservoirs 261—268, and a plurality of recycle pumps 281—287.
In this ment, primary reservoir 210 is divided
up into a plurality of inlet reservoirs 211-
218.
During operation, algal s 201 is placed a first inlet reservoir 211 near a first
end 221 of transport ism 220. In addition, solvent 205 is placed into inlet reservoir 218
near a second end 222 of transport mechanism 220. Transport mechanism 220 directs the algal
biomass along transport mechanism 220 from first end 221 towards second end 222. As the
PCT/USZOIZ/027537
algal biomass is transported, it passes through the plurality of separation devices 241-248 and
is separated into fractions of varying polarity. The diffusate portions that
pass through
tion devices 241-248 are directed to reservoirs 261-268.
For e, the diffusate portion of the algal biomass that
passes through the first
separation device 241 (e. g., the portion containing liquid and particles small enough to pass
through separation device 241) is directed to the first reservoir 261. From first reservoir 261,
the ate portion can be recycled back to first inlet reservoir 201. The retentate portion of
the algal biomass that does not pass through first separation device 241 can then be directed by
transport mechanism 220 to second inlet reservoir 212 and second separation device 242,
which can comprise a finer separation or filtration media than the first separation device 241.
The t of the diffusate portion that
passes through second tion device
242 can be directed to second reservoir 262, and then recycled back to second inlet reservoir
212 Via recycle pump 282. The retentate or extracted portion of the algal biomass that does
pass through second separation device 242 can be directed by ort mechanism 220 to third
inlet reservoir 213. This process can be repeated for inlet reservoirs 8 and separation
devices 243-248 such that the retentate portions at each stage are ed to the uent
inlet oirs, while the diffusate portions are directed to the e reservoirs and recycled
back to the current inlet oir.
In exemplary embodiments, the first fraction will be extracted with the highest
water to slightly nonpolar solvent ratio, i.e., most polar mixture, while the last fraction will be
extracted with the most pure slightly nonpolar solvent, i.e. the least polar mixture. The
process
therefore extracts components in the order of decreasing polarity with the fraction. The
function ofthe first fraction is to remove the residual water and facilitate the solvent extraction
process. The fractions that follow are rich in polar lipids, while the final fractions are rich in
neutral lipids.
The oil fraction can be fied to liberate the long chain unsaturated fatty acids.
The carotenoids and long chain unsaturated fatty acids can be separated from the oil using
processes such as molecular distillation in conjunction with non-molecular distillation. All of
the fatty acids can be separated from the carotenoids using the molecular distillation. The
distillates can be fractionated using a simple distillation column to te the lower chain
fatty acids for refining. The long chain unsaturated fatty acids remain as high boiling residue in
the column.
W0 2012/138438
In some non-limiting embodiments, the extraction system and methods described
herein incorporate one or more steps to isolate n material from the oleaginous material
(e.g., algal biomass). Such protein extraction steps employ pH adjustment(s) to achieve
isolation and extraction of protein. For example, in one non-limiting embodiment, the pH of
the solvent in the first separation device is optimized for protein extraction, resulting in
a first
fraction that is rich in protein material. The pH of the protein extraction
step is adjusted
depending on the pKa of the proteins of interest. The pKa of a protein of interest may be
ascertained using methods known to one of skill in the art, including, but not limited
to using
the Poisson—Boltzmann on, empirical methods, molecular dynamics based methods,
the use oftitration .
In some embodiments, the solvent pH is alkaline. For example, in some
ments, the solvent pH is greater than about 10. In other embodiments, the solvent pH
ranges from about 10 to about 12. In further embodiments, the solvent pH is about l0, about
11, or about 12. In other embodiments, the t pH is acid. For example, in some
embodiments, the solvent pH is less than about 5. In other embodiments, the solvent pH
ranges
from about 2 to about 5. In further ments, the solvent pH is about 2, about 3, about 4,
about 4.5, or about 5. The extracted portion of the first separation device is then directed
subsequent inlet reservoirs to achieve extraction and onation based on polarity. In another
non—limiting embodiment, protein material is separated in the final separation device by similar
means (i.e., solvent pH ment).
Adjustment of t pH is accomplished in accordance with methods known to
those of skill in the art. For example, acid pH is achieved by e of
an appropriate acid
into the solvent stream. Exemplary acids useful for protein extraction include, without
limitation, phosphoric acid, sulfuric acid, and hydrochloric acid. Similarly, alkaline pH is
achieved by on and mixture of an appropriate base into the solvent
stream. Exemplary
bases useful for n extraction include, without limitation, potassium ide,
sodium ide.
In some embodiments, protein extraction is performed in
a system separate from the
extraction and fractionation system described herein. For example, in some embodiments, an
algal biomass is soaked in a pH-adj usted solvent mixture, followed by ion via an
appropriate separation technique (e.g, centrifugation, or filtration). The remaining solid is then
introduced into an extraction and fractionation system based
on polarity, as described herein.
W0 2012/138438 PCT/U82012/027537
Similarly, in some embodiments, the remaining extract from an extraction and fractionation
process based on polarity is exposed to a pH-adjusted solvent mixture to isolate n
material at the end of the tion process.
As shown in the solvent selection and the theory of fractionation based
polarity were developed by extensive analysis of solvents and the effect on extraction using the
Sohxlet extraction process, which allows the separation of lipids from a solid material. The
Sohxlet extraction system was ed for rapid screening solvents for lipid class selectivity
and recovery. ts from various chemical classes encompassing a wide
range of polarities
such as alkanes, lkane, alkyl halides, esters, ketones, were tested. Prior to the extraction,
the lipid content and composition of the biomass to be extracted
was tested in triplicate using
the standard methods for algae oil estimation such as the Dyer lipid extraction method.
The biomass contained 22.16% total lipid, ofwhich 49.52%
was neutral lipid.
presents the data gathered by extraction of a dry algal mass using various
polar and nonpolar solvents combined with a Sohxlet extraction s. Depending on the
chain length of the alkane t, 60-70% purity ofneutral lipids and 15-45% of total lipid
recovery can be achieved without tion and solvent tion. The longest chain alkane
solvent tested, e, recovered 60% of the neutral lipids and 42% of the total lipid. As
shows, the results of extraction of dry algal mass using solvents and conventional
extraction
methods such as hexane are inefficient, expensive, and result in
poor yields. The systems and
methods discloses herein s these inefficiencies by controlling the proportion of ly
nonpolar solvent to water in order to separate out components of differing polarities with
l loss of components.
[0138} The lower carbon alcohol solvents were more selective for polar lipids. The neutral
lipid purity was 22% for methanol and 45% for ethanol. Isopropyl alcohol did not show any
selectivity between polar and nonpolar lipids, resulting in a 52% pure neutral lipid product.
Methanol recovered 67% of the total lipids and more than 90% of the polar lipids. Therefore,
methanol is an excellent candidate for an embodiment of the
present invention wherein
methanol can be used to selectively t polar lipids from an oleaginous material prior
extracting the neutral lipids using heptane or hexane. The other solvent classes tested did not
show any selectivity towards lipid class since the neutral lipid purity
was close to 49%, similar
to the lipid composition present in the original biomass. Furthermore, the total lipid recovery
W0 2012/138438 2012/027537
achieved with these solvents ranged from about 15—35%, rendering these solvents unsuitable
for the selective extraction of particular lipid classes or total lipid extraction.
The results from the Sohxlct analysis were confirmed using the standard bench scale
batch solvent extraction apparatus described below in Example 1. The solvents selected were
methanol for the first step to recover polar lipids, and petroleum ether in the second
step for
recovery of neutral lipids. All of the extractions were performed with a 1:10 solidzsolvent
ratio. Each extraction step in this experiment was 1 hour long. Other experiments done (data
not shown) te that about 45 minutes or longer is long enough for the extraction
to be
successful. This retention time is dependent on the heat and mass transfer ofthe
The methanol extractions were med at different temperatures, 40 0C, 50 OC,
and 65 °C, in order to determine which was optimal. The petroleum ether extraction
performed at 35°C, close to the boiling point of the solvent. Petroleum ether was chosen
e of its high ivity for neutral lipids, low boiling point, and the product quality
observed after extraction.
shows that the neutral lipid purity in a petroleum ether extraction carried
out after a ol extraction step at 65°C is over 80%, trating that the combination of
these two extraction steps enhanced the neutral lipid content of the final crude oil product.
shows that the total neutral lipid
recovery was low and there was a significant amount
ofneutral lipid loss in the first step.
To minimize the loss of neutral lipids in the methanol extraction
step, the polarity of
the solvent can be sed by adding water to the solvent. and 5B show the results
of extracting the aforementioned biomass with 70% v/v
s ol followed by
extraction with petroleum ether. shows that the neutral lipid purity was much higher
in the petroleum ether extraction than was ed by the
use ofpure methanol. Moreover,
the loss of neutral lipids was y reduced by the use of
aqueous methanol in the first
extraction step. As seen in , methanol extraction at higher
temperatures improved
neutral lipid purity but slightly decreased the total lipid
recovery in the subsequent step.
In some exemplary embodiments the temperature of the extraction
process is
controlled in order to ensure optimal stability of algal ents
present in the algal biomass.
Algal proteins, carotenoids, and chlorophyll are examples of algal components that exhibit
temperature sensitivity. In other embodiments, the ature is increased after the
temperature sensitive algal components have been extracted from the algal biomass.
PCT/U82012/027537
In still other exemplary embodiments, the temperature of the extraction
process is
adjusted in order to optimize the yield of the desired product. Extractions can be run from
ambient temperature up to, but below, the boiling point of the extraction mixture. In still other
embodiments, the temperature ofthe extraction process is changed depending on the solubility
ofthe desired product. In still other embodiments, the extraction temperature is optimized
depending on the algal strain of the biomass to be extracted. Elevated extraction temperatures
increase the solubility of desired compounds and reduce the viscosity of the extraction mixture
enhancing extraction recovery.
In some embodiments, the extraction is run under
pressure to elevate the boiling
point of the tion mixture. In these implementations, the pressure is increased to the
degree necessary to prevent boiling, while maintaining the temperature of the tion
mixture below a temperature at which any of the desired products would begin to degrade,
denature, decompose, or be destroyed.
In some exemplary embodiments, the extraction is performed near the boiling point
ofthe solvent used, at the conditions under which the extraction is med (e.
g., atmospheric
or elevated pressures). In other embodiments, the extraction is performed near the boiling
point of the extraction mixture, again accounting for other extraction conditions. At such
temperatures, vapor phase penetration of the solvent into the algal cells is faster due to lower
mass er resistance. If the extraction temperature is allowed to significantly exceed the
boiling point of the solvent, the solvent—water system can form an azeotrope. Thus,
ining the system at or near the boiling point of solvent would generate enough vapors to
enhance the extraction, while reducing expense. In addition, the solubility of oil is sed at
higher temperatures, which can further increase the effectiveness of tion at temperatures
close to the solvent boiling point. shows the total lipid recovery in the
aqueous
methanol-petroleum ether extraction scheme. Although performing the methanol tion
near its boiling temperature slightly decreases the l lipid recovery
as observed in , it enhances the total lipid recovery.
In other embodiments, the tion is carried out under t lighting
ions. In other embodiments, the tion is d out in an
opaque container such as,
but not limited to, a steel tube or , in order to t light sensitive algal
components
from degradation. Carotenoids are light sensitive algal components.
W0 2012/138438 PCT/U52012/027537
In other exemplary embodiments, the extraction takes place under normal
atmospheric conditions. In still other embodiments, the extraction takes place under a nitrogen
atmosphere in order to protect algal components prone to oxidation. In still other
embodiments, the extraction takes place under an atmosphere of inert gas in order to protect
algal components prone to oxidation. Algal components that might be prone to oxidation
include carotenoids, chlorophyll, and lipids.
In exemplary embodiments, the solvent-to-solid ratio for the extraction is between
3—5 based on the dry weight of the solids in the biomass. The residual algal biomass is rich
carbohydrates (e.g., starch) and can be used as a feed stock to produce the solvent used for
extraction.
shows the effect of the solvent to solid ratio on the total lipid
recovery. As
the solvent to solid ratio was increased, there was a corresponding and drastic increase in
total
lipid recovery. It is ed that this was because of the lower solubility of lipids in methanol
as ed to other commonly used oil extraction solvents such
as hexane.
The solubility of ents is affected by the polarity of t used in
extraction process. The solubility properties can be used to determine the ratio of
wet biomass
to solvent. For example, a 40% w/w wet biomass has 40
g s and 60 g water for every
100 g ofwet s. If 100 g of l is added to this mixture, the ratio of ethanol
to wet
biomass is 1 part wet biomass to 1 part l and the concentration of ethanol in the mixture
is 100/(100+60) equals about 62% w/w of ethanol in the liquid phase. 62% w/w of
ethanol in
l water mixture corresponds to a ty index of 6.6, calculated by weight
averaging the polarities of the components. Ethanol, having a polarity index of 5.2, and water,
having a polarity index of 9, in a mixture containing 62% ethanol and 38% water results in a
polarity index of (0.62*5.2+.38*9) about 6.6. The polarity index of the mixture for extraction
ofpolar lipids and neutral lipids is calculated to be about 5.8 and 5.4 respectively.
In light of
the instant disclosure, one of skill in the art would be able to formulate
a solvent set that can
selectively extract these ents.
In another example, if the extraction solvent is a 1:1 mixture of isopropyl alcohol
and ethanol, the polarity of this t is ((3.9+5.4)/2) which is about 4.65. The
ratio of
solvent to wet biomass would be calculated to match the polarities. To
get a 6.6 polarity index,
we would need to make a 55% w/w of IPA-water mixture calculated by g the following
algebraic equation:
W0 2012/138438 PCT/U$2012/027537
X*4.65+(1—x)*9=6.6
x = (9-6.6)/(9-4.65) = 0.55
55% w/w*ofsolvent mix in solvent mix—water
For a 40% w/w wet biomass, the wet biomass to IPA ratio is (l-0.55)/0.6 ~ 0.75.
With a 40% w/w wet biomass this would pond to a ratio of 100
parts wet
biomass to 75 parts solvent mixture. A 40% W/w wet biomass has 40
g biomass and 60 g water
for every 100 g of wet biomass. If 75 g of solvent mixture is added to this mixture, the
concentration of solvent in the mixture is (75/(75+60)) is about 55% w/w of solvent mixture in
the solvent mixture—water solution. This calculation can be used to obtain the solvent biomass
ratio at each extraction stage and for each product. A few nonlimiting examples of solvent
sets
appear in Table 3.
Table 3
......................Extraction"?
; g s 3 5 §Solvent- solvent
:parts wet2parts EBiomass Parts Dry EParts iwater :polarity
§95% ethanol 5% methanol mixture
395% ethanol 5% IPA e
One step Polar lipids Extraction
EEthanol
gPropanol
21:1 lPA EtOH e
95% l water mixture
i 95% ethanol 5% methanol e
' 5% ethanol 5%lPA mixture
95% ethanol 5% met
§95% ethanol 5%IPA mixture
PCT/U52012/027537
The extraction mixture described in all es, is made
up of a substantially solid
phase and a substantially liquid phase. These phases are then separated post extraction. This
can then be followed by removal of the liquid solvent from the liquid phase, yielding
extraction product. In some embodiments, the solvent is evaporated. In such an
implementation, a liquid-liquid extraction technique can be used to reduce the amount of
solvent that needs to be evaporated. Any solvents used can be recycled if conditions allow.
It was theorized that treatment of the algal biomass prior to tion would
enhance the productivity and ncy of lipid extraction. In this direction an experiment was
done comparing the effect of adding a base or another organic solvent to
an algal biomass to
change the e properties and enhance extraction. A variety of treatments including
aqueous methanol, aqueous sodium hydroxide, and aqueous DMSO were attempted. As
demonstrates, the on of 5% DMSO increases the lipid recovery 3-fold. These extraction
steps may be exploited to dramatically reduce the ol extraction steps. However, the
solutions used in the above experiments may not be ideal for use on larger scales due
to the
high cost, viscosity, and ability to recover and recycle DMSO.
is a chart showing the effect of an eight step ol extraction
on the
cumulative total lipid yield and the purity of the extracted neutral lipid. In this embodiment,
112 grams of wet biomass (25.6% dry weight), was extracted with 350 mL
pure methanol and
heating for 10 minutes at 160 W irradiance power in each step. This resulted in an extraction
temperature of about 75°C, which was near the boiling point of the extraction mixture. Using
this process, it was determined that it is possible to obtain highly
pure neutral lipids from algal
oil once the majority of the polar lipids have been extracted. shows that it is le to
isolate high purity neutral lipid once the polar lipids are all extracted. In this case a 5% yield of
total biomass was achieved with over 90% neutral lipids purity in ol extraction
steps 5
through 8. Furthermore, due to the boiling point of the extraction e, most of the water in
the biomass is completely extracted in the first extraction step, along with ydrates,
proteins and metals.
shows that recovery of lipids can be made more efficient by the
use of
ethanol to extract lipids and protein from wet biomass. By using ethanol, 80% total lipid
ry can be achieved in about 4 steps rather than the 9 lly needed by using
methanol. This increase in recovery may be attributed to greater solubility of lipids in ethanol
as compared to methanol. Furthermore, the boiling point of aqueous ethanol is higher than
W0 2012/138438 PCT/U82012/027537
aqueous methanol, facilitating fiirther recovery of lipids. This is because the higher
temperature renders the oil less viscous, thereby improving diffusability. Another distinct
advantage of this process is using the al ethanol in the oil on for transesterification
as well as lowering the heat load on the biomass drying operation.
Further, FIG. lO demonstrates that the initial fractions are non-lipid rich, containing
proteins and other highly polar molecules, followed by the polar lipid rich fractions and finally
the l lipid ons. Hence with a proper design of the extraction
apparatus, one can
recover all the three products in a single extraction and fractionation s.
Another embodiment of the current invention utilizes aves to assist
extraction. Based on usly gathered data disclosed in this application, it is shown that
methanol is the best single solvent for extraction of all lipids from algae. Hence, a single
solvent multiple step extraction, as described in Example 1 of the instant application, was
performed in order to gather data on the efficacy of a one solvent microwave extraction system.
is a logarithmic plot comparing the extraction time and total lipid
recovery
of conventional extraction and microwave-assisted tion. Based
on the slope of the curve,
it was calculated that the microwave system reduces the extraction time by about five fold
more. While the conventional methods have a higher net lipid
recovery, this is due to higher
recoveries of polar lipids. Based on these results, the conditions for extraction of dry algal
biomass using ts with and without microwave assistance have been optimized. Some
embodiments ofthe invention use traditional microwave apparatus, which emit wavelengths
that excite water molecules. Further embodiments of the invention e ized
microwave apparatus capable of exciting different solvents. Still other embodiments of the
invention utilize custom microwave apparatus capable of exciting the lipids
present in the algal
biomass. In some embodiments, the lipids present in the algal biomass
are excited using
microwaves, thereby enhancing the separation and extraction of the lipid components from the
algal biomass.
Moisture content is another parameter of s that will influence the ncy
of oil tion. In some embodiments of the present invention, dry algal
mass is extracted
and fractionated. In other embodiments, the algal mass is wet. Biomass samples with algae
mass contents of 10%, 25%, and 33% were used to investigate the influence of moisture
extraction performance.
PCT/U52012/027537
A shows an illustrative process 400 for a step—wise extraction of products
from an algae biomass. All units in FlG. 12A are in pounds. A shows a mass balance
ofthe process 400, while the details of the equipment and/or systems for performing the
process are described elsewhere herein. A biomass containing 5 pounds of algae has about
0.63 pounds ofpolar lipids, 1.87 pounds neutral lipids, 1 pound protein, and 1.5 pounds
carbohydrates. The biomass and 1000 pounds of water is sed in a dewatering step 405,
which separates 950 pounds of water from the mixture and
passes 5 pounds of algae in 45
pounds of water to a first extraction step 410. Any of the dewatering techniques disclosed
herein can be used tin dewatering step 405. In the first extraction step 410, 238 pounds of
ethanol and 12 pounds r are combined with the algae and water from the previous
step.
The first tion step 410 has a liquid phase of about 80.9% W/W ethanol. A first liquid
phase of231 pounds of l, 53 pounds of water, and 0.5 pounds of algal proteins are
recovered, from which water and ethanol are removed by, e.g.
, ation, leaving a protein-
rich product 415. Solvent recovered from the evaporation can be recycled to the first extraction
step 410.
A first solid phase from the first extraction step 410 is passed to
a second extraction
step 420; this first solid phase includes 4.5 pounds of algae, 2.6 pounds of water, and 10.9
pounds of ethanol. Eighty-six pounds of ethanol and 4 pounds of water are added to the first
solid phase from the previous step. The second extraction step 420 has
a liquid phase of about
93.6% w/w ethanol. A second liquid phase of 85.9 pounds ethanol, 5.9 pounds
water, and 0.6
pounds polar lipids are recovered, from which water and ethanol are removed by,
e.g.,
evaporation, leaving a polar lipid-rich product 425. Solvent recovered from the evaporation
can be recycled to the second extraction step 420.
A second solid phase from the second extraction step 420 is passed
to a third
extraction step 430; this first solid phase es 3.9 pounds of algae, 0.7 pounds of
water, and
11 pounds of ethanol. Seventy-found and a half pounds of ethanol and 3.5 pounds of water are
added to the second solid phase from the previous step. The third extraction
step 430 has a
liquid phase of about 95.4% w/w l. A third liquid phase of 78.9 pounds l, 3.9
pounds water, and 1.6 pounds neutral lipids are recovered, from which water and ethanol are
removed by, e.g., evaporation, leaving a neutral lipid-rich product 435. Solvent recovered from
the ation can be recycled to the second extraction step 430 A solid phase of
2.3 pounds
algac, 0.3 pounds water, and 6.6 pounds cthanol rcmain.
W0 2012/138438 PCT/U52012/027537
As demonstrated in A, the resulting lipid profile with each sequential
ethanol extraction step was largely influenced by the moisture content in the starting algae.
Models of process 400 were run on three different s collections, each having
a ent
initial water content. As the initial water content decreased, the maximum lipid
recovery step
changed from the third extraction step to a fourth (not . However, the overall lipid
recovery from these three s samples were quite similar, all above 95% of the total lipid
content of the algal biomass.
When algal mass with higher moisture content was used, the ethanol concentration
in the aqueous ethanol mixture was much lower, and consequently the neutral lipid
tage
in the crude t was also lower. It has been ed that dewatering an algae paste with
90% water is a very energy intensive
process. The methods described herein unexpectedly can
be used to successfully extract and fractionate an algal mass containing mostly
water. As
overall lipid ry was not significantly influenced by starting from
an algae paste
containing 90% water (10% algal solids), unlike conventional extraction methods, the methods
disclosed herein do not e the use of an energy intensive drying
step.
B shows an illustrative implementation 500 of one of the extraction
steps of
process 400. An algae biomass and solvent mixture 505 is provided to an extraction vessel
510. After the algae is extracted (as described elsewhere herein), the mixture is provided
to a
coarse filtration system 515, such as a sintered metal tube filter, which separates the mixture
into a liquid phase and a solid phase. The solid phase is passed to
a downstream extraction
step. The liquid phase is passed to a solvent removal system 520, e.g., an evaporator, to reduce
the solvent (e.g., ethanol) content in the liquid phase. The liquid phase remaining after
solvent
removal is, ally, passed to a centrifuge 525. Any solids remaining in the solvent
removal
system are recycled or discarded. Centrifuge 525 assists in separating the d algal product
(e.g., proteins or lipids) from any remaining water and/or solids in the liquid phase.
shows an example of a
process 600 by which an algal mass can be
processed to form or recover one or more algal products. In this example, an algal biomass is
extracted in a ise manner in a front-end
s 605 using the methods disclosed herein.
The extraction and separation steps are followed by an fication
process 610, a hydrolysis
process 615, a hydrotreating process 620, and/or a distillation process 625 to further isolate
components and products. The ents and products include algal lipids, algal proteins,
glycerine, carotenoids, nutraceuticals (e.g., long chain unsaturated oils and/or esters), fuel
2012/027537
esters (generally, the esters having chain lengths of C20 or shorter), fuels, fuel additives,
naphtha, and/or liquid petroleum substitutes. In preferred embodiments the fuel esters are C16
chain s. In others, the fuel esters are C18 chain s. In still other embodiments, fuel
esters are a mixture of chain lengths, C20 or shorter.
The esterification process 610, hydrolysis
process 615, reating process 620,
and distillation process 625 are optional and can be used in various orders. The dashed
arrows
and dotted arrows te some, but not all, of the options for when the hydrolysis,
hydrotreating, and/or distillation processes may be performed in the processing of the lipid
fractions. For example, in some embodiments of the ion, after extraction and/or
separation are carried out, the neutral lipids fraction can be directly hydrotreated in order to
make fuel products and/or additives. Alternatively, in other ments, the l lipid
on can be passed to esterification process 610.
Esterification process 610 can include techniques known in the art, such
as acid /
base sis, and can include transesterification. Although base catalysis is not excluded for
producing some products, acid catalysis is red as those techniques avoid the soaps that
are formed during base catalysis, which can complicated downstream processing. Enzymatic
esterification techniques can also be used. Esterification can process ntially pure lipid
material (over 75% lipid, as used herein). After fication, glycerine byproduct can be
removed. The esterified lipids can then undergo molecular and/or nonmolecular distillation
(process 625) in order to separate esterified lipids of different chain lengths as well as
carotenoids present in the lipid fraction. The esterified lipids can then be passed to
hydrotreating process 620 to generate jet fuel, biodiesel, and other fuel products. Any
hydrotreating process known in the art can be used; such a process adds hydrogen to the lipid
molecules and removes oxygen molecules. Exemplary conditions for hydrotreating comprise
reacting the triglycerides, fatty acids, fatty acid esters with hydrogen under high pressure in the
range of 600 psi and temperature in the range of 600°F. Commonly used catalysts are NiMo or
CoMo.
Hydrotreating the fiiel esters rather than the raw lipids has several advantages.
First, the esterification process 610 reduces the levels of certain phosphorus and metals
compounds present in algal oils. These materials are poisons to catalysts typically used in
hydrotreating processes. Thus, esterification prior to hydrotreating prolongs the life of the
hydrotreating catalyst. Also, fication reduces the molecular weight of the compounds
W0 2012/138438
being hydrotreated, thereby improving the performance of the hydrotreating process 620.
r still, it is advantageous to retain the fuel esters from the distillation
s 625 to be
reated in a vaporous form, as doing so reduces the energy needed for hydrotreating.
In some embodiments ofthe invention, the neutral algal lipids are directly
hydrotreated in order to convert the lipids into fuel products and additives. While in other
implementations, the neutral lipids are cstcrificd and separated into carotenoids, long chain
unsaturated esters, eicosapentaenoic acid (EPA) esters, and/or fuel esters via distillation
process 625. Distillation process 625 can include molecular distillation as well as
any of the
distillation techniques known in the art. For example, the distillates can be fractionated using a
simple distillation column to separate the lower chain fatty acids for refining. The long chain
unsaturated fatty acids remain as high g residue in the column. In some embodiments,
the remaining vapor can then be sent to the hydrotreating
process. Two of the advantages of
the present invention are that it yields pure feed as well as a
vapor product, which favors the
energy intensive hydrotreating reaction, as described above.
In some embodiments ofthe invention, polar lipids (and, optionally, neutral lipids)
are yzed in hydrolysis process 615 before being passed to the esterification
process.
Doing so unbinds the fatty acids of the algal lipids, and enables a greater amount of the algal
lipids to be formed into useful products.
is a flowchart g a
process 700 for producing eutical products
from l lipids. In one implementation ofprocess 700, neutral lipids
are fed to an
adsorption process 705 that separates carotenoids from EPA-rich oil. The neutral lipids can be
from an algae source generated by any of the selective extraction techniques disclosed
herein.
However, the neutral lipids can be from other sources, such as plant sources.
tion process 705 includes contacting the l lipids with an adsorbent to
adsorb the carotenoids, such as beta carotene and xanthophylls. In one implementation, the
adsorbent is Diaion HP2OSS (commercially available from ITOCHU Chemicals
America,
Inc.). The neutral lipids can contact the adsorbent in a type process, in which the neutral
lipid and adsorbent are held in a vessel for a ed amount of time. After the contact time,
the absorbent and liquid are separated using techniques known in the art. In other
implementations, the adsorbent is held in an adsorbent bed, and the neutral lipids are passed
through the adsorbent bed. Upon passing through the adsorbent bed, the noids content of
the neutral lipids is reduced, y producing an oil rich in EPA.
W0 2012/138438 PCT/U52012/027537
The carotenoids can be recovered from the adsorbent material by treating the
adsorbent with an appropriate solvent, including, but not limited to, alcohols such
as ethanol,
isopropyl alcohol , butanol, esters such as ethyl e or butyl acetate, alkanes such as
hexane, and pentane.
In yet another embodiment of the invention, phylls and carotenoids
ed from the neutral . The adsorption
process can be performed using a hobic
adsorbent such as polystyrene-divinylbenzene resin (PS-DVB) or with higher carbon loading
resin. Some examples of the resins are XAD 16, XAD 4 from Rohm and Hass delphia,
PA) or HP20, SP—850 from Mitsubishi Chemical Industries Limited , Japan) buted
by ltochu Chemicals America, Inc, White Plains, NY). The tion fraction dissolved in
aqueous ethanol would be passed over a packed column to selectively retain the chlorophylls
and carotenoids. Upon reaching the adsorption loading capacity, the column would be washed
with a hydrophobic solvent such as hexane. The
pure carotenoids and chlorophyll on
dissolved in hexane is evaporated under mild ions to obtain a concentrate. The
pure
carotenoids and chlorophyll could also be llized and precipitated in the solution.
In yet another embodiment of the invention, extracting l lipids from the algal
biomass, followed by a membrane diafiltration step for the separation of chlorophylls and
carotenoids from the neutral lipids. Membrane diafiltration works based
on the principle of
selectively eluting the product through a membrane and retaining the remaining components of
the feed mixture. In this case, we would use a solvent stable nanofiltration membrane such
SelRO products from Koch membrane systems (Wilmington, MA). The neutral lipids
extract
suspended in aqueous ethanol would be filtered through the membrane to selectively separate
the unbound carotenoids and chlorophylls. The feed is continuously diluted with ethanol
until
all the carotenoids and chlorophylls are washed off and ted in
permeate.
In yet another embodiment of the ion, chlorophylls and carotenoids
isolated from the polar lipids. The adsorption
process can be med using a hydrophobic
adsorbent such as polystyrene-divinylbenzene resin (PS—DVB)
or with higher carbon loading
resin. Some examples of the resins are XAD 16, XAD 4 from Rohm and Hass (Philadelphia,
PA) or HP20, SP—850 from Mitsubishi Chemical Industries Limited (Tokyo, Japan) distributed
by ltochu Chemicals America, Inc, White Plains, NY). The polar lipids extract is dissolved in
aqueous ethanol would be passed over a packed column to selectively retain the chlorophylls
and carotenoids. Upon reaching the adsorption loading capacity, the column would be
washed
W0 2012/138438
with a hydrophobic solvent such as hexane. The
pure carotenoids and chlorophyll fraction
dissolved in hexane is evaporated under mild conditions to obtain a trate. The
pure
carotenoids and phyll could also be crystallized and itated in the solution.
In yet another embodiment of the invention, ting polar lipids from the algal
biomass, followed by a membrane diafiltration step for the separation of chlorophylls and
carotenoids from the polar lipids. Membrane diafiltration works based on the principle of
selectively eluting the product through a membrane and retaining the remaining components of
the feed mixture. In this case, we would use a solvent stable nanofiltration membrane such
SelRO from Koch membrane systems (Wilmington, MA). The polar lipids t suspended in
aqueous ethanol would be filtered h the membrane to selectively separate the unbound
noids and chlorophylls. The feed is continuously diluted with ethanol until all the
carotenoids and chlorophylls are washed off and collected in permeate.
In yet another embodiment of the invention, a method of making biofiiels includes
dewatering ntially intact algal cells to make an algal biomass, extracting neutral lipids
from the algal biomass, ed by an adsorption step for the separation of chlorophylls and
carotenoids from the neutral lipids. The method also includes esterifying the neutral lipids with
a catalyst in the presence of an alcohol, and separating a water soluble fraction comprising
glycerin from a water insoluble fraction comprising fuel esters. The method still r
includes distilling the fuel esters under vacuum to obtain a C16 or shorter fuel
esters fraction, a
C16 or longer fuel ester fraction, and a e comprising omega—3 fatty acids and
hydrogenating and deoxygenating at least one of (i) the C16 or shorter fuel esters to obtain a jet
fuel blend stock and (ii) the C16 or longer fuel esters to obtain
a diesel blend stock.
In yet another embodiment of the invention, a method ofmaking biofuels includes
dewatering substantially intact algal cells to make an algal biomass, extracting l lipids
from the algal biomass, followed by a membrane diafiltration
step for the separation of
chlorophylls and noids from the neutral lipids. The method also includes esterifying the
neutral lipids with a st in the presence of an alcohol, and separating
a water soluble
fraction comprising glycerin from a water insoluble fraction comprising fuel
esters. The
method still further includes distilling the fuel esters under vacuum to obtain
a C16 or shorter
fuel esters fraction, a C16 or longer fuel ester fraction, and a residue comprising
3 fatty
acids and hydrogenating and deoxygenating at least one of (i) the Cl 6
or shorter fuel esters to
obtain a jet fuel blend stock and (ii) the C16 or longer fuel esters to obtain
a diesel blend stock.
W0 2012/138438 PCT/U52012/027537
In yet another embodiment of the invention, a method of making euticals and
health foods includes dewatering substantially intact algal cells to make an algal biomass,
extracting neutral lipids from the algal biomass, followed by an adsorption step for the
separation of chlorophylls and carotenoids from the neutral lipids. The neutral lipids resulting
from this step would be rich in omega-3 fatty acids with a balanced ratio of monounsaturated,
polyunsaturated and saturated fatty acids. This can be sold as high y oil for better health.
In yet another ment of the invention, a method of making nutraceuticals and
health foods es dewatering substantially intact algal cells to make an algal biomass,
extracting neutral lipids from the algal s, ed by a membrane diafiltration step for
the separation of chlorophylls and carotenoids from the neutral lipids. The l lipids
resulting from this step would be rich in omega-3 fatty acids with a balanced ratio of
monounsaturated, polyunsaturated and saturated fatty acids. This can be sold as high quality oil
for better health.
is a flowchart showing a process 800 for ing fuel products 830 from
neutral lipids 805. The neutral lipids can be from an algae source generated by
any of the
selective extraction ques disclosed herein. However, the neutral lipids can be from other
sources, such as plant sources. The l lipids are d in a degumming process 810, in
which the lipids are acid washed to reduce the levels of metals and phospholipids in the neutral
lipids. In some implementations, a relatively dilute solution of phosphoric acid is added
to the
neutral lipids, and the mixture is heated and agitated. The precipitated phospholipids and
metals are then separated from the remaining oil, for example, by centrifuge.
The treated oil is then passed to bleaching
process 815 to remove chlorophylls and
other color compounds. In some implementations, bleaching
process 815 includes contacting
the oil with clay and or other adsorbent material such as bleaching clay (Le. bentonite
fuller’s earth), which reduce the levels of chlorophylls and other color nds in the oil.
The d oil then is passed to hydrotreating
process 820, which hydrogenates and
deoxygenates the components of the oil to form fuels products, for e, jet fuel mixtures,
diesel fuel additive, and propane. In addition, the hydrotreating process 820 also causes some
cracking and the creation of smaller chain compounds, such as LPG and naptha. Any of the
hydrotreating processes described herein can be used for hydrotreating process 820.
The mixture of compounds created in the hydrotreating
process 820 are passed to a
distillation process 825 to separate them into various fuel products 830. Distillation process
W0 38438 PCT/U82012/027537
825 can include any of the molecular and non-molecular distillation techniques described
herein or known in the art for tion of fuel compounds.
In some embodiments of the instant invention, ns
may be selectively ted
from an algal biomass. Extraction of proteins using the disclosed methods offers many
advantages. In particular, algal cells do not need to be lysed prior to extracting the desired
proteins. This simplifies and reduces costs of extraction. The methods of the instant ion
exploit the solubility s of different classes of proteins in order to selectively extract and
onate them from an algal culture, biomass, paste, or cake.
For e, an algal biomass may be subjected to heating and mixing to extract
water and salt soluble proteins called albumins and ins. This mixture can then be
subjected to a change in pH to recover the alkali soluble proteins called the glutelins. This step
can then be followed by a solvent-based separation of the alcohol soluble proteins called
prolamins. The remaining biomass would be rich in carbohydrates and .
Proteins can be extracted from both saltwater and freshwater algal cells, as shown in
FIGS. 17 and 18. The presence of salt in the saltwater algal culture or biomass affects the
extraction of different classes of protein, but the methods disclosed herein enable
one to
selectively extract proteins from either fresh or saltwater algae.
In some embodiments, extraction of proteins from ater algal cells is
accomplished by the novel process shown in . Freshwater algal cells or a freshwater
algal biomass are heated and mixed. Mixing can be accomplished by a variety of s
known in the art such as, but not limited to, stirring, ion, and rocking. This process
generates a first heated extraction mixture or slurry, comprised of a first substantially liquid
phase and a first substantially solid phase. The solid and liquid phases are then separated.
Separation can be accomplished by a variety of methods known in the art including, but not
limited to, eentrifugation, decantation, flotation, sedimentation, and filtration. This first
substantially liquid phase is enriched in albumin proteins.
The first ntially solid phase is then mixed with salt water and heated to
generate a second heated extraction mixture or slurry, comprised of a second substantially
liquid phase and a second substantially solid phase. The salt water may be natural seawater or
may be an aqueous salt solution. An example of such a solution would se about
typically 35 g/L comprising mainly ofNaCl. The solid and liquid phases are then separated.
This second substantially liquid phase is enriched in globulin proteins.
PCT/U52012/027537
The second substantially solid phase is then mixed with water and heated to
generate a third heated extraction mixture or , comprised of a third substantially liquid
phase and a third substantially solid phase. The pH of this third extraction mixture or slurry is
then raised to about 9 or greater, enriching the third substantially liquid phase with glutelin
ns. The solid and liquid phases are then ted, the third substantially liquid phase
being enriched in glutelin proteins.
The third substantially solid phase is then mixed with a solvent set and heated
generate a fourth heated extraction mixture or slurry, comprised of a fourth substantially liquid
phase and a fourth substantially solid phase. In one preferred embodiment, the solvent set
comprises l. In other non-limiting embodiments, the t set comprises
one or more
ofthe following solvents: methanol, isopropanol, acetone, ethyl
acetate, and acetonitrile. The
solid and liquid phases are then separated. This fourth substantially liquid phase is enriched in
prolamin proteins. The remaining fourth substantially solid phase may be enriched in lipids,
depending on the composition of the starting algal biomass.
In some embodiments, extraction of proteins from saltwater algal cells is
accomplished by the novel process shown in . ter algal cells or a saltwater algal
biomass are heated and mixed. Mixing can be accomplished by a variety of methods known in
the art such as, but not limited to, stirring, agitation, and rocking. This process generates a first
heated extraction e or , comprised of a first substantially liquid phase and
a first
ntially solid phase. The solid and liquid phases are then separated. Separation can be
accomplished by a variety of methods known in the art including, but not limited to,
centrifugation, ation, flotation, sedimentation, and filtration. This first substantially
liquid phase is enriched in in proteins.
The first substantially solid phase is then mixed with water and heated to
generate a
second heated extraction mixture or slurry, comprised of a second substantially liquid
phase
and a second substantially solid phase. The solid and liquid phases
are then separated. This
second substantially liquid phase is enriched in albumin proteins.
The second substantially solid phase is then mixed with water and heated
generate a third heated extraction mixture or slurry, sed of a third substantially liquid
phase and a third substantially solid phase. The pH of this third extraction mixture or slurry is
then raised to pH 9 or greater, enriching the third substantially liquid phase with glutelin
W0 2012/138438 PCT/U52012/027537
proteins. The solid and liquid phases are then separated, the third substantially liquid phase
being enriched in glutelin proteins.
The third substantially solid phase is then mixed with a solvent set and heated to
generate a fourth heated extraction mixture or , comprised of a fourth substantially liquid
phase and a fourth substantially solid phase. In one preferred ment, the solvent set
comprises ethanol. In other non-limiting ments, the solvent set comprises one or more
ofthe following solvents: ol, isopropanol, acetone, ethyl acetate, and acetonitrile. The
solid and liquid phases are then separated. This fourth substantially liquid phase is enriched in
prolamin proteins. The remaining fourth substantially solid phase may be ed in lipids,
depending on the composition of the starting algal biomass.
The sed s also provide for the selective extraction of different types of
proteins, as shown in -20. Any of the steps of the aforementioned extraction process
can be performed separately from the rest of the steps in order to selectively extract
a single
protein product. Two examples of this appear in and 18, as the as demonstrated by the
dashed box around extraction step la.
In a non-limiting example, globulin proteins can be selectively extracted from
freshwater algal biomass by mixing said biomass with salt water and heating to
generate a
heated extraction e or slurry, comprised of a substantially liquid phase and
substantially solid phase. The solid and liquid phases can then be ted. The liquid phase
is enriched in globulin proteins. See , extraction step la.
In another non—limiting example, albumin proteins can be selectively extracted from
a saltwater algal biomass by mixing said biomass with water and heating to generate
a heated
extraction mixture or , comprised of a ntially liquid phase and a substantially solid
phase. The solid and liquid phases can then be separated. The liquid phase is enriched in
globulin proteins. See , extraction step la.
In a r non~limiting example, prolamin proteins can be selectively extracted
from either a ater or saltwater algal biomass as shown in . The selective
extraction is accomplished by mixing the algal biomass with a solvent set and heating to
generate a heated extraction mixture or slurry, comprised of a substantially liquid phase and a
substantially solid phase. The solid and liquid phases can then be separated. The liquid phase
is enriched in prolamin proteins.
W0 2012/138438 PCT/U52012/027537
In yet another non-limiting example, a protein fraction can be selectively extracted
from either a freshwater or saltwater algal biomass as shown in . The selective
extraction is accomplished by mixing the algal biomass with a solvent set to
generate an
extraction mixture or slurry and effecting a pH change in the e. The mixture is
comprised of a substantially liquid phase and a substantially solid phase. The solid and liquid
phases can then be separated. The liquid phase is enriched in proteins.
Having been informed of these aspects of the invention, one of skill in the art would
be able to ively extract a desired protein from either a freshwater
or saltwater algal
biomass by either a single step extraction process, or a step extraction
s. In light of
the instant disclosure, one of skill in the art would be able to interchange the order of the above
disclosed multi—step extraction schemes, provided that the protein content of the algal
mass and
the lity properties of the ns of interest are taken into account. Other embodiments
ofthe disclosed methods may incorporate a wash step between each extraction
step.
For any of the disclosed protein extraction methods, the extraction mixture/slurry
may be maintained at a heated temperature for a period of time. In some embodiments, the
extraction e is maintained at a heated temperature for between about 20 s to about
90 minutes. In some aspects, the extraction mixture is maintained at a heated temperature for
n about 20 s and about 60 minutes. In other aspects, the extraction mixture is
maintained at a heated ature for between about 45 s to about 90 minutes.
In some embodiments, the tion mixture/slurry
may be heated to temperatures
less than about 50°C. In some aspects, the albumin, globulin, and glutelin proteins
extracted at temperatures of less than about 50°C. In other embodiments the extraction
mixture/slurry is heated to a temperature close to the boiling point of extraction mixture/slurry.
In some aspects, the prolamin proteins are extracted at temperatures close to the boiling point
ofthe extraction mixture/slurry. In other embodiments, the pressure is increased above
atmospheric pressure, up to and including, 5Opsi, during the heating and mixing steps to
e extraction.
Example 1
Green microalgae Scena’esmus dz'morphus (SD) were cultured in outdoor panel
photobioreactors. SD samples of varying lipid contents were harvested, After removal of bulk
water by centrifugation, the algal samples were stored as 3-5 cm algae cakes at —80°C until
use.
W0 2012/138438
A pie-calculated amount ofwet algal biomass (15
grams dry algae weight equivalent) and 90
mL of ethanol solvent was added into a three-neck flask equipped with ser, mechanical
stirring and a thermocouple. In one experiment, the mixture was d for 10 min under
microwave irradiance. In a second, the mixture was refluxed for 1h with electronic heating.
Afterwards, the mixture was cooled to room ature and separated into a diffusate and
retentate by filtration.
The total lipids of algal samples were analyzed using a chloroforrn—methanol—water
system ing to Bligh and Dyer’s lipid extraction method. This total lipid value was used
as nce for the lipid recovery calculation. Total lipids were further separated into neutral
lipids and polar lipids by standard column chromatography method using 60—200 mesh silica
gel (Merck Corp, Germany). Each lipid fraction was transferred into a pre-weighed vial,
initially evaporated at 30 °C using a rotary evaporator (Biichi, Switzerland) and then dried
under high vacuum. The dried retentates were placed under nitrogen and then weighed. The
fatty acid profile of each sample was fied by GC-MS after derivatization into fatty acid
methyl esters using heptadecanoic acid (C17:0) as the internal standard.
The results (data not shown) indicated that microwave assisted extraction
was best
for removal of the polar lipids in the first extraction step, and somewhat less effective for
separation of neutral lipids. Electronic heating is more consistent in extraction effectiveness.
The final yield is comparable between microwave assisted tion and onic heating
assisted extraction, but, microwave assisted extraction is significantly faster.
Example 2
Protein extraction from algal biomass
(1) Acid Leaching: Algal biomass was soaked in water at pH 4.5 for 1 hour. The
samples were then centrifuged at 3000 rpm for three minutes, and the supernatant removed.
The remaining solids were washed 3 times with dilute acid (pH 4.5) and freeze dried.
(2) Alkaline tion: Algal biomass was soaked in water at pH 11 for 1 hour.
following the addition of pH-adjusted water. The samples were then centrifuged at 3000
for three minutes, and the supernatant removed. The supernatant
was neutralized with acid (pH
4.5) following the fugation. The remaining solids were washed 3 times with dilute acid
(pH 4.5) and freeze dried.
The results of acid leaching and ne extraction are shown below in Table 4.
W0 2012/138438
LIME!
Process Protein Yield Protein Purity
(% weight) (% weight of
n yield)
Alkaline tion 16 45
Acid Leaching 70 32.5
Protein yield was calculated on a weight basis, comparing the weight of the freeze
dried solids to the weight of the algal biomass prior to soaking in pH-adjusted
water. Protein
purity was determined by the Official Method of the American Oil Chemists' Society (Ba-2a-
38), ing the amount of nitrogen in the freeze dried solids of each process As proteins
are an ant product that adds to the value of algal product extraction, this information
allows for the use of feedstocks with varying levels of protein in the
systems and methods
disclosed herein.
Extraction of Proteins from Saltwater Algal Biomass
The saltwater algal culture initially made
up of about 1—10% w/w solids in saltwater
was heated to 50°C and maintained at this temperature for 1 hr. The resulting slurry was
centrifuged to separate the liquid phase from the solid phase. The liquid extract was
determined to be rich in globulin proteins (about 10% of the total proteins
present in the
original algal biomass).
The solids were then suspended in fiesh water and heated to about 50°C and
maintained for about 1 hour. The resulting slurry was centrifuged again to
te the liquid
from the solid phase. The liquid phase was determined to be rich in albumin
proteins (about
% of the total proteins present in the original algal biomass).
The solids were then ded in ethanol to achieve a 70% w/w mixture. This
mixture was heated to about 75°C and maintained at that temperature for about 1 hour. The
resulting slurry was centrifuged to separate the liquid from the solid phase The liquid phase
was ined to be rich in albumin proteins (about 30% of the total proteins t in the
original biomass).
The solids were then suspended in alkali solution (aqueous NaOH, pH 9) and heated
to about 50°C and maintained at that temperature for about 1 hour. The resulting slurry
centrifuged to separate the liquid from the solid phase. The liquid phase was determined to be
rich in glutelin proteins (about 50% of the total ns present in the original biomass).
Example 4
Step Fractionation and tion of Algal Biomass by Ethanol
One thousand pounds ofNaimochloropsz's biomass red from strain 202.0,
obtained from Arizona State sity, Laboratory for Algae Research and Biotechnology,
ATCC Deposit Number PTA-l 1048), was ted and dewatered until algae comprised
about 35% w/w and then finally frozen.
The extraction steps were performed in a 400 gallon jacketed kettle with hinged
lids. The lids were tied down with straps and sealed with silicone. The
system also contained a
mixer with a 2 horsepower explosion proofmotor with a two blade shaft. The frozen algae
al was emptied into the tank and an equal weight of ethanol
was pumped in using a
pneumatic drum pump. The material was stirred for 15 s and the jacket heated with
steam to obtain the desired temperature at each extraction step. The desired
temperature is
near, meaning within 3°C of the boiling point of the mixture, but not boiling. This desired
temperature is different at each extraction step as the boiling point ofthe mixture changes as
the proportion of ethanol is changed. Upon reaching the desired
temperature, the system was
stirred continuously held at the desired temperature for 60 minutes to
ensure that the contents
ofthe kettle were uniformly heated.
{0220] The contents of the kettle were then pumped out of the extraction vessel and into
es decanter centrifuge, using a pneumatic Viking vane pump at about 1 gallon per
minute. The er centrifuge rotor speed was set to about 6000
rpm. The solids were
collected in an enclosed plastic drum and consisted of about 50% w/w solids
to liquids. These
solids were returned to the kettle, where the aforementioned tion
steps were repeated.
The liquid stream from the decanter was collected into a feed tank
was and then fed to the
membrane filtration system. The membrane used was a 0.375 ft2 SS membrane manufactured
by Graver Technologies. The operating conditions were 60°C i 5°C and with an
average
W0 2012/138438 PCT/U52012/027537
pressure gradient of 40 psi. The membrane system was backwashed about every 15 minutes
with compressed air to maintain the flux. The permeate collected from the membrane
system
was free of any ulate matter. The retentate was collected and recycled to the decanter.
This extraction and onation is due to the change in polarity of the solvent
through the process in each extraction. in the extraction shown in F1G. 13, the process began
with about 1000 lbs. ofwet algal biomass containing about 65%
pure water (35% w/w algal
solids). This was mixed with 860 lbs. of denatured ethanol (95% ethanol and 5% methanol),
resulting in a mixture containing about 55% aqueous ethanol. The solids and liquids were
ted using a decanter as described above. The wet solid portion weighed 525 lbs. and was
40% dry mass. A total of 525 lbs. of 95% the denatured ethanol was added to the solids,
resulting in a mixture made up of about 85% aqueous ethanol. The solids and liquids were
separated using a decanter as described above. The solid portion d 354.5 lbs. and was
40% dry mass. To this mass, r 700 lbs. of denatured ethanol was added, resulting in
mixture of about 95% aqueous ethanol. The solids and liquids were separated using
a decanter
as described above. The resulting solids were about 40% dry mass. This s es
60% less energy to dry, calculated based on the latent heat ofwater and ethanol.
In some experiments (data not shown) other types of denatured ethanol
were tried.
Denatured ethanol containing 95% ethanol and 5% isopropyl alcohol
was used in an extraction,
but was found not to be as effective as 95% l and 5% ol. Use of 100% ethanol is
a preferred embodiment of the present invention, but is generally not available due to cost
constraints.
The permeate stream from the membrane system was ated using
an in-house
fabricated batch still. The ing conditions were about 80°C during the
vacuum distillation.
All of the l in the permeate was evaporated. These extraction
steps were repeated three
times, resulting in four product pools, as shown in . This is because with each
extraction step, the polarity changed with the addition of water to the mixture, allowing for the
extraction of different components with each step. Product 1 contained the algal proteins, and
as a result, retained excess water in the system that could not be vaporized under the operating
conditions. Product 2 contained the polar lipids. Product 3 contained the neutral lipids.
Finally, Product 4 was the residual biomass, containing potential coproducts such as
carotenoids.
W0 2012/138438
ring and Extraction of Algal Biomass by Ethanol
Upon harvesting, algal biomass typically contains between about 0.1 to 0.5 %
(w/w) solids. This can be dewatered using any of the methods known in the algae industry,
including, but not limited to membrane filtration, fugation, heating, sedimentation or
flotation. Flocculation can either assist in flotation or sedimentation. The typical result of such
methods is an algae slurry containing about 10% w/w solids. To dewater further, another
dewatering method may be used to remove some of the remaining free water to get the
concentration of solids closer to 40% w/w. However, the cost of dewatering increases
exponentially after the first dewatering is carried out. An advantage of the s and
methods disclosed herein is that the allow for the extraction and fractionation of
an algal mass
that has undergone only one round of dewatering.
An example of such a process might be that in the first extraction round, following
the protocol described in Example 3, 1000 lbs. of wet s containing 90%
pure water and
is mixed with 1000 lbs. of denatured ethanol (95% EtOH and 5% MeOH), resulting in
a solvent
mixture of about 50% aqueous ethanol. The resulting biomass (350 lbs.) is 40% dry. The
t composition of these wet solids is 50%
aqueous ethanol. With another 350 lbs. of
denatured ethanol, the composition of the mixture would be about 81%
s ethanol. The
resulting biomass (235 lbs.) is 40% dry. The solvent composition of these wet solids is 81%
aqueous ethanol. With another 470 lbs. of denatured ethanol, the composition of the mixture
would be about 95% aqueous l. The resulting solids would be 40% dry with about 95%
ethanol. This wet biomass requires 60% less energy to dry based on the latent heat of
water
and ethanol. In this case, 100 lbs. of algae would have been extracted using 1820 lbs. ethanol.
When compared with Example 3, wherein the starting material
was 40% algal solids, 350 lbs.
ofthe dry algae equivalent was extracted with 2085 lbs. ethanol.
PCT/U52012/027537
References
The following references are herein incorporated by reference in their ty:
US. Patent 7,148,366
, Science Progress, 39—90, 2009. Generic review on using algae to produce
biodieselChisti, Y. (2007). Biodiesel from microalgae. Biotechnol Adv 25, 294-306. -
Generic review on using algae to produce biodiesel
Amin, Energy Convers. Manage, 50:1834—1840, 2009. Generic review on using algae to
produce biofuel and gas
Catchpole et al., J. of ritical Fluids, 47:591—597, 2009. SCF C02 based extraction of
specialty lipids
Bligh E G & Dyer W J. A rapid method of total lipid extraction and purification. Can. J.
Biochem. Physiol. 37: 911-917, 1959.
Christie, W. W., Lipid Analysis, 3rd ed., Oily Press, Bridgewarer, UK, 2003, 416.
Approved Methods of the AACC, 9th ed, American Association of Cereal Chemists. St. Paul,
MN, 1995 AACC Method 58-19.
Claims (3)
1. A method of isolating chlorophylls and omega-3 rich oil from algae, sing: a. dewatering substantially intact algal cells to make an algal biomass; b. adding a first ethanol fraction to the algal biomass in a ratio of about 1 part ethanol to about 1 part algal biomass by weight; c. separating a first substantially solid biomass fraction from a first substantially liquid fraction comprising proteins; d. combining the first substantially solid biomass fraction with a second ethanol on in a ratio of about 1 part ethanol to about 1 part solids by weight; e. separating a second substantially solid biomass fraction from a second substantially liquid fraction sing polar lipids; f. combining the second substantially solid biomass fraction with a third ethanol solvent fraction in a ratio of about 1 part ethanol to about 1 part substantially solid biomass by weight; g. separating a third substantially solid biomass fraction from a third substantially liquid on comprising neutral , including omega-3 fatty acids, carotenoids, and chlorophyll, wherein the third substantially solid biomass fraction comprises carbohydrates; and h. isolating at least one of carotenoids, chlorophyll, and omega-3 fatty acids from the third substantially liquid fraction.
2. The method of claim 1, further comprising esterifying the neutral lipids with a catalyst in the presence of an alcohol, and separating a water soluble fraction comprising glycerin from a water ble fraction comprising fuel esters.
3. The method of claim 2, further sing distilling the fuel esters under vacuum to obtain a C16 or shorter fuel ester fraction, a C16 or longer fuel ester fraction, and a e comprising omega-3 fatty acids. . The method of claim 3, further comprising deoxygenating the C16 or shorter filel ester fraction to obtain a jet fuel blend stock and/or the C16 or longer fuel fraction to obtain a diesel blend stock. . The method of any one of the preceding claims, wherein the isolating is carried out by adsorption with an ent material. . The method of claim 5, wherein the isolating is carried out by adsorption with a clay. . The method of claim 6, wherein the clay is ed from the following group consisting ofbleaching clay, ite, and fuller’s earth. . The method of claim 1, substantially as herein described with reference to any one of the Examples and/or
Applications Claiming Priority (7)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US13/081,221 | 2011-04-06 | ||
US13/081,221 US8084038B2 (en) | 2010-04-06 | 2011-04-06 | Methods of and systems for isolating nutraceutical products from algae |
US13/149,595 | 2011-05-31 | ||
US13/149,595 US8115022B2 (en) | 2010-04-06 | 2011-05-31 | Methods of producing biofuels, chlorophylls and carotenoids |
US13/274,201 | 2011-10-14 | ||
US13/274,201 US8242296B2 (en) | 2010-04-06 | 2011-10-14 | Products from step-wise extraction of algal biomasses |
PCT/US2012/027537 WO2012138438A1 (en) | 2011-04-06 | 2012-03-02 | Methods of producing biofuels, chlorophylls and carotenoids |
Publications (2)
Publication Number | Publication Date |
---|---|
NZ615225A NZ615225A (en) | 2015-11-27 |
NZ615225B2 true NZ615225B2 (en) | 2016-03-01 |
Family
ID=
Similar Documents
Publication | Publication Date | Title |
---|---|---|
US8242296B2 (en) | Products from step-wise extraction of algal biomasses | |
EP2556162B1 (en) | Sequential solvent extraction of oil and proteinaceous material from oleaginous material by using solvents of decreasing polarity | |
US8313648B2 (en) | Methods of and systems for producing biofuels from algal oil | |
SG193374A1 (en) | Extraction of neutral lipids by a two solvent method | |
MX2013011453A (en) | Extraction of proteins from algae. | |
AU2012240608A1 (en) | Methods of producing biofuels, chlorophylls and carotenoids | |
US20130217904A1 (en) | Method of extracting polar lipids and neutral lipids with two solvents | |
WO2013142694A1 (en) | Method of extracting neutral lipids with two solvents | |
NZ615225B2 (en) | Methods of producing biofuels, chlorophylls and carotenoids | |
WO2013052598A1 (en) | Method of producing biofuels from algal cell culture |