CN112079709B - Method for preparing 3-hydroxydecanoic acid by hydrolyzing rhamnolipid - Google Patents
Method for preparing 3-hydroxydecanoic acid by hydrolyzing rhamnolipid Download PDFInfo
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- FYSSBMZUBSBFJL-UHFFFAOYSA-N 3-hydroxydecanoic acid Chemical compound CCCCCCCC(O)CC(O)=O FYSSBMZUBSBFJL-UHFFFAOYSA-N 0.000 title claims abstract description 161
- FCBUKWWQSZQDDI-UHFFFAOYSA-N rhamnolipid Chemical compound CCCCCCCC(CC(O)=O)OC(=O)CC(CCCCCCC)OC1OC(C)C(O)C(O)C1OC1C(O)C(O)C(O)C(C)O1 FCBUKWWQSZQDDI-UHFFFAOYSA-N 0.000 title claims abstract description 109
- 238000000034 method Methods 0.000 title claims abstract description 60
- 230000003301 hydrolyzing effect Effects 0.000 title description 10
- 238000006243 chemical reaction Methods 0.000 claims abstract description 59
- 238000006460 hydrolysis reaction Methods 0.000 claims abstract description 59
- 230000007062 hydrolysis Effects 0.000 claims abstract description 45
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 32
- 239000002253 acid Substances 0.000 claims abstract description 22
- SHZGCJCMOBCMKK-JFNONXLTSA-N L-rhamnopyranose Chemical compound C[C@@H]1OC(O)[C@H](O)[C@H](O)[C@H]1O SHZGCJCMOBCMKK-JFNONXLTSA-N 0.000 claims abstract description 20
- SHZGCJCMOBCMKK-UHFFFAOYSA-N D-mannomethylose Natural products CC1OC(O)C(O)C(O)C1O SHZGCJCMOBCMKK-UHFFFAOYSA-N 0.000 claims abstract description 19
- PNNNRSAQSRJVSB-UHFFFAOYSA-N L-rhamnose Natural products CC(O)C(O)C(O)C(O)C=O PNNNRSAQSRJVSB-UHFFFAOYSA-N 0.000 claims abstract description 19
- 239000007864 aqueous solution Substances 0.000 claims abstract description 19
- 238000006555 catalytic reaction Methods 0.000 claims abstract description 5
- 108090000623 proteins and genes Proteins 0.000 claims description 33
- 102000004169 proteins and genes Human genes 0.000 claims description 32
- 229920001184 polypeptide Polymers 0.000 claims description 28
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 28
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 28
- 150000001413 amino acids Chemical class 0.000 claims description 25
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 claims description 22
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 18
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 16
- 102000015636 Oligopeptides Human genes 0.000 claims description 16
- 108010038807 Oligopeptides Proteins 0.000 claims description 16
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims description 11
- 239000002994 raw material Substances 0.000 claims description 8
- 230000035484 reaction time Effects 0.000 claims description 6
- 239000000243 solution Substances 0.000 claims description 6
- JOXIMZWYDAKGHI-UHFFFAOYSA-N toluene-4-sulfonic acid Chemical compound CC1=CC=C(S(O)(=O)=O)C=C1 JOXIMZWYDAKGHI-UHFFFAOYSA-N 0.000 claims description 6
- 239000000463 material Substances 0.000 claims description 5
- PSZYNBSKGUBXEH-UHFFFAOYSA-N naphthalene-1-sulfonic acid Chemical compound C1=CC=C2C(S(=O)(=O)O)=CC=CC2=C1 PSZYNBSKGUBXEH-UHFFFAOYSA-N 0.000 claims description 3
- 239000012071 phase Substances 0.000 description 66
- 239000003921 oil Substances 0.000 description 49
- 235000019198 oils Nutrition 0.000 description 49
- FYSSBMZUBSBFJL-VIFPVBQESA-N (S)-3-hydroxydecanoic acid Chemical compound CCCCCCC[C@H](O)CC(O)=O FYSSBMZUBSBFJL-VIFPVBQESA-N 0.000 description 29
- 238000004811 liquid chromatography Methods 0.000 description 20
- 238000004458 analytical method Methods 0.000 description 15
- 239000000047 product Substances 0.000 description 13
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 9
- 238000000605 extraction Methods 0.000 description 9
- 239000008367 deionised water Substances 0.000 description 8
- 229910021641 deionized water Inorganic materials 0.000 description 8
- 239000000203 mixture Substances 0.000 description 8
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 7
- 238000002360 preparation method Methods 0.000 description 7
- 239000012535 impurity Substances 0.000 description 6
- 238000003756 stirring Methods 0.000 description 6
- CLBRCZAHAHECKY-UHFFFAOYSA-N [Co].[Pt] Chemical compound [Co].[Pt] CLBRCZAHAHECKY-UHFFFAOYSA-N 0.000 description 5
- 238000006266 etherification reaction Methods 0.000 description 5
- 238000000855 fermentation Methods 0.000 description 5
- 230000004151 fermentation Effects 0.000 description 5
- 239000005014 poly(hydroxyalkanoate) Substances 0.000 description 5
- 229920000903 polyhydroxyalkanoate Polymers 0.000 description 5
- 239000000539 dimer Substances 0.000 description 4
- 238000010438 heat treatment Methods 0.000 description 4
- 150000002500 ions Chemical class 0.000 description 4
- 238000004519 manufacturing process Methods 0.000 description 4
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 230000001580 bacterial effect Effects 0.000 description 3
- 239000007795 chemical reaction product Substances 0.000 description 3
- NKLPQNGYXWVELD-UHFFFAOYSA-M coomassie brilliant blue Chemical compound [Na+].C1=CC(OCC)=CC=C1NC1=CC=C(C(=C2C=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=2C=CC(=CC=2)N(CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=C1 NKLPQNGYXWVELD-UHFFFAOYSA-M 0.000 description 3
- 239000013612 plasmid Substances 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- -1 rhamnolipid Chemical compound 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- MYKOKMFESWKQRX-UHFFFAOYSA-N 10h-anthracen-9-one;sulfuric acid Chemical compound OS(O)(=O)=O.C1=CC=C2C(=O)C3=CC=CC=C3CC2=C1 MYKOKMFESWKQRX-UHFFFAOYSA-N 0.000 description 2
- WHBMMWSBFZVSSR-UHFFFAOYSA-N 3-hydroxybutyric acid Chemical compound CC(O)CC(O)=O WHBMMWSBFZVSSR-UHFFFAOYSA-N 0.000 description 2
- 229910017052 cobalt Inorganic materials 0.000 description 2
- 239000010941 cobalt Substances 0.000 description 2
- GUTLYIVDDKVIGB-UHFFFAOYSA-N cobalt atom Chemical compound [Co] GUTLYIVDDKVIGB-UHFFFAOYSA-N 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 235000014113 dietary fatty acids Nutrition 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 229930195729 fatty acid Natural products 0.000 description 2
- 239000000194 fatty acid Substances 0.000 description 2
- 230000002401 inhibitory effect Effects 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 150000007522 mineralic acids Chemical class 0.000 description 2
- 239000000178 monomer Substances 0.000 description 2
- 150000007524 organic acids Chemical class 0.000 description 2
- 101150030205 phaG gene Proteins 0.000 description 2
- 229910052697 platinum Inorganic materials 0.000 description 2
- 239000002244 precipitate Substances 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 239000001117 sulphuric acid Substances 0.000 description 2
- 235000011149 sulphuric acid Nutrition 0.000 description 2
- HPMGFDVTYHWBAG-UHFFFAOYSA-N 3-hydroxyhexanoic acid Chemical compound CCCC(O)CC(O)=O HPMGFDVTYHWBAG-UHFFFAOYSA-N 0.000 description 1
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 101150017040 I gene Proteins 0.000 description 1
- 235000019482 Palm oil Nutrition 0.000 description 1
- 241000589517 Pseudomonas aeruginosa Species 0.000 description 1
- 102000004357 Transferases Human genes 0.000 description 1
- 108090000992 Transferases Proteins 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 238000005904 alkaline hydrolysis reaction Methods 0.000 description 1
- 235000011114 ammonium hydroxide Nutrition 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 239000008346 aqueous phase Substances 0.000 description 1
- 239000012620 biological material Substances 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 239000012295 chemical reaction liquid Substances 0.000 description 1
- 238000010367 cloning Methods 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 239000012043 crude product Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 238000006471 dimerization reaction Methods 0.000 description 1
- 238000004821 distillation Methods 0.000 description 1
- 238000000132 electrospray ionisation Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- RTZKZFJDLAIYFH-UHFFFAOYSA-N ether Substances CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 1
- 150000002170 ethers Chemical class 0.000 description 1
- 150000004665 fatty acids Chemical class 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 239000000413 hydrolysate Substances 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 125000000896 monocarboxylic acid group Chemical group 0.000 description 1
- 238000002552 multiple reaction monitoring Methods 0.000 description 1
- 238000001728 nano-filtration Methods 0.000 description 1
- 239000002540 palm oil Substances 0.000 description 1
- 238000005191 phase separation Methods 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 238000006116 polymerization reaction Methods 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 230000008092 positive effect Effects 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 238000005173 quadrupole mass spectroscopy Methods 0.000 description 1
- 230000007226 seed germination Effects 0.000 description 1
- 238000004704 ultra performance liquid chromatography Methods 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C51/00—Preparation of carboxylic acids or their salts, halides or anhydrides
- C07C51/347—Preparation of carboxylic acids or their salts, halides or anhydrides by reactions not involving formation of carboxyl groups
- C07C51/367—Preparation of carboxylic acids or their salts, halides or anhydrides by reactions not involving formation of carboxyl groups by introduction of functional groups containing oxygen only in singly bound form
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Engineering & Computer Science (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
A method of preparing 3-hydroxydecanoic acid from rhamnolipid hydrolysis, the method comprising: 1) The rhamnolipid aqueous solution is subjected to hydrolysis reaction under the catalysis of acid to obtain a water phase containing rhamnose and an oil phase containing 3-hydroxydecanoic acid; 2) Separating the oil phase containing the 3-hydroxydecanoic acid from the reaction system, and carrying out deep hydrolysis reaction under the catalysis of acid to generate the 3-hydroxydecanoic acid. The method can prepare the 3-hydroxydecanoic acid with high yield and high purity.
Description
Technical Field
The invention belongs to the field of organic chemistry, and particularly relates to a preparation method of 3-hydroxydecanoic acid.
Background
With more and more attention paid to environmental protection, degradable materials become a trend of new material development. The polyhydroxy fatty acid is used as a high-performance polymer, is biodegradable, is derived from organisms, and belongs to a material with a full life cycle and environment friendliness. The 3-hydroxydecanoic acid can be used as a monomer of biological material Polyhydroxyalkanoates (PHA), and can be polymerized or copolymerized with other hydroxyfatty acids such as 3-hydroxybutyric acid or 3-hydroxyhexanoic acid, so that the polyhydroxyalkanoates with excellent performance and commercial prospect can be obtained. In addition, the 3-hydroxydecanoate is also proved to have the functions of inhibiting bacteria, inhibiting pollen growth, seed germination and the like, and has some applications in the aspect of crop protection.
At present, the source of the 3-hydroxydecanoic acid mainly comprises PHA which is obtained by degrading polyhydroxyalkanoate containing the 3-hydroxydecanoic acid to obtain a monomer, or 3-hydroxydecanoic acid is obtained by hydrolyzing a chemical containing the 3-hydroxydecanoic acid, such as rhamnolipid, or is directly prepared by a fermentation method.
Chinese patent CN108587989 provides a strain for producing 3-hydroxydecanoic acid and a production method thereof, the strain is Pseudomonas aeruginosa, the rhamnosyl transferase I gene (rh IB) of the strain is inactivated, and the rh IA gene of the strain is utilized to catalyze 3-hydroxyester acyl-CoA/ACP to synthesize hydroxy fatty acid dimer (HAA, 3- (3-hydroxyakanoyloxy) aldolate). The engineering strain provided by the invention is fermented and cultured in a culture medium which comprises palm oil and/or glycerol as a carbon source, and the 3-hydroxydecanoic acid can be obtained after alkaline hydrolysis treatment.
Chinese patent CN1379110 also provides a preparation method of 3-hydroxydecanoic acid, and the scheme of the invention is to obtain 3-hydroxydecanoic acid by fermenting and culturing a recombinant microorganism strain containing gene phaG. Firstly, cloning a gene phaG into a plasmid to construct a new plasmid, then transferring the constructed plasmid into a bacterial strain to construct a DNA recombinant bacterial strain, and culturing the recombinant bacterial strain to obtain the 3-hydroxydecanoic acid.
The preparation method of 3-hydroxydecanoic acid is also mentioned in the US patent 4,933,281, which adopts a method of hydrolyzing rhamnolipid to obtain rhamnose and simultaneously produce a byproduct of 3-hydroxydecanoic acid.
From the prior preparation technology of 3-hydroxydecanoate, the problems of low yield and low production efficiency exist in the production of 3-hydroxydecanoate by adopting a direct fermentation method, and the separation and purification difficulty of 3-hydroxydecanoate from fermentation liquor is large. The hydrolysis of rhamnolipids to obtain 3-hydroxydecanoic acid is a more feasible approach, but is less described in the prior art.
Rhamnolipids are compounds having the following structure:
wherein R is 1 Is H or:
wherein R is 2 Is H or:
CH[(CH 2 ) 6 CH 3 ]CH 2 COOH
the 3-hydroxydecanoic acid is a structure contained in rhamnolipid, and different rhamnolipid structures contain 3-hydroxydecanoic acid in different proportions:
defining: 3-hydroxydecanoic acid structure average mass fraction = ∑ x i *m i
Wherein x is i The i-th structure rhamnolipid accounts for the mass fraction of all rhamnolipids; m is i Is the proportion of the molecular weight of the 3-hydroxydecanoate structure in the rhamnolipid with the i-th structure to the overall molecular weight of the rhamnolipid.
The hydrolysis of rhamnolipids to rhamnose and 3-hydroxydecanoic acid is as follows:
although the hydrolysis preparation of 3-hydroxydecanoic acid from rhamnolipids is chemically feasible, it was found in the actual preparation process that the hydrolysis preparation of 3-hydroxydecanoic acid from rhamnolipids presents the following problems:
1) The 3-hydroxydecanoate product contains two products (dimers) of the polymerization of the 3-hydroxydecanoate, the hydrolysis of the rhamnolipid of which is incomplete, and the impurities reduce the purity of the 3-hydroxydecanoate product on one hand, and the existence of the dimers reduces the yield of the 3-hydroxydecanoate prepared by the hydrolysis of the rhamnolipid on the other hand.
2) Hydroxyl in the hydrolysate 3-hydroxydecanoate and hydroxyl on rhamnose can generate etherification reaction, and the generated ether bond is difficult to be hydrolyzed again, so that the yield of rhamnose and 3-hydroxydecanoate prepared by rhamnolipid hydrolysis is reduced. The structure shown in the following chemical formula is one of ethers, a rhamnose structure contains 4 hydroxyl groups, and each hydroxyl group can be subjected to etherification reaction with the hydroxyl group of 3-hydroxydecanoic acid to generate an etherification product.
The yield of 3-hydroxydecanoic acid prepared by hydrolyzing rhamnolipid is low due to the reasons, and the purity of the obtained 3-hydroxydecanoic acid is poor, so that the high-end application of the 3-hydroxydecanoic acid is limited.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention provides a method for preparing 3-hydroxydecanoate by efficiently hydrolyzing rhamnolipid with great industrial prospect, and 3-hydroxydecanoate can be prepared with high yield and high purity.
In order to solve the technical problems, the technical scheme adopted by the invention is as follows:
a method of preparing 3-hydroxydecanoic acid from rhamnolipid hydrolysis, the method comprising:
1) The rhamnolipid aqueous solution is subjected to hydrolysis reaction at a certain temperature under the catalysis of acid to generate rhamnose and 3-hydroxydecanoate; 3-hydroxydecanoic acid is in oil phase, rhamnose is in water phase, and unreacted rhamnolipid is in the lowest precipitation phase;
2) Separating the oil phase containing the 3-hydroxydecanoate from the reaction system, reacting with a certain amount of acid, and carrying out deep hydrolysis reaction at a certain temperature to generate the 3-hydroxydecanoate.
The rhamnolipids sold on the market are usually derived from fermentation processes, and the products usually contain proteins, polypeptides and the like, and these impurities are also subjected to hydrolysis reactions in a hydrolysis process to generate amino acids, polypeptides and the like. In the research process, the inventor finds that amino acid, polypeptide and the like can generate browning reaction with rhamnose, and the browning reaction product and 3-hydroxydecanoic acid are difficult to separate by the conventional method.
According to the method, the rhamnolipid raw material needs to control the contents of protein, polypeptide, oligopeptide and amino acid. The content of protein in the impurities is not more than 0.1% of the weight of rhamnolipid, and the sum of protein, polypeptide, oligopeptide and amino acid in the rhamnolipid raw material is preferably 0.01-0.1% of the weight of rhamnolipid. The content of protein, polypeptide, oligopeptide and amino acid is usually tested by Coomassie Brilliant blue method, and the content of total protein is used to represent the content of protein, polypeptide, oligopeptide and amino acid in the sample.
Generally, organic membranes with different pore diameters can be used for intercepting proteins, polypeptides and the like with different molecular weights, for example, a nanofiltration membrane with the diameter of 1-10nm is adopted, so that the polypeptides, the proteins and the like can be separated; the protein and the target product may be separated from each other by using the difference in solubility between the protein and the target product in two phases, such as extraction. The protein, the polypeptide, the oligopeptide and the amino acid are thoroughly separated from the rhamnolipid, so that the loss of the rhamnolipid in the separation process is overlarge; in the research process, when the content is controlled to be below 0.01%, the browning reaction is not obvious, the production cost is higher, and the economical efficiency is poor.
According to the method, the rhamnolipid in the rhamnolipid aqueous solution has the mass fraction of 5-40%, and preferably 7.5-25%.
According to the method, the acid added in the rhamnolipid hydrolysis process is inorganic acid such as sulfuric acid, phosphoric acid or hydrochloric acid, and can also be organic acid such as naphthalene sulfonic acid and p-toluenesulfonic acid. Preferably one or more of sulphuric acid, phosphoric acid and hydrochloric acid.
According to the method of the present invention, the reaction temperature of the rhamnolipid hydrolysis step in step 1) is 100 ℃ to 150 ℃, preferably 120 ℃ to 140 ℃.
According to the method of the present invention, the amount of acid added in the rhamnolipid hydrolysis step in step 1) (if the acid is an aqueous solution, the amount of acid refers to the mass of solute) is about 0.5% -5%, preferably 1% -2% of the mass of rhamnolipid solution, and preferably the acid is concentrated sulfuric acid with a concentration of 98% or concentrated phosphoric acid with a concentration of 85%.
According to the method of the present invention, the reaction time of the rhamnolipid hydrolysis step in step 1) is 1-10h, preferably 2-4h.
According to the method, the rhamnolipid is subjected to hydrolysis reaction, an oil phase containing 3-hydroxydecanoic acid is generated, and the oil phase is separated for further deep hydrolysis by a phase separation method before the mass of the oil phase accounts for 0.5% of the mass of the rhamnolipid aqueous solution raw material. Preferably, the oil phase is separated before the mass of the oil phase reaches 0.1% of the mass of the rhamnolipid aqueous solution raw material. 3-hydroxy decanoic acid generated by the reaction is transferred away in time to prevent the further reaction; in the experimental process, researchers surprisingly find that when the mass of the oil phase accounts for more than 0.5 percent of that of the rhamnolipid aqueous solution raw material, the dimerization reaction and etherification reaction of the 3-hydroxydecanoate are increased, and the yield of the product is reduced.
According to the method of the invention, the acid added in the deep hydrolysis process of the oil phase containing the 3-hydroxydecanoic acid in the step 2) is inorganic acid such as sulfuric acid, phosphoric acid or hydrochloric acid, and can also be organic acid such as naphthalenesulfonic acid and p-toluenesulfonic acid. Preferably one or more of sulphuric acid, phosphoric acid and hydrochloric acid.
According to the process of the invention, the reaction temperature in the step of deep hydrolysis of the oil phase in step 2) is from 140 ℃ to 180 ℃, preferably from 150 ℃ to 170 ℃.
According to the process of the invention, the acid is added in the step of deep hydrolysis of the oil phase in step 2) in an amount of about 1% to about 5%, preferably about 2% to about 4% of the mass of the oil phase.
According to the method of the present invention, the reaction time of the rhamnolipid hydrolysis step in step 2) is 0.5-5h, preferably 1-2h.
By adopting the technology of the invention, the positive effects are as follows:
1) The method has the advantages that the probability of etherification reaction between the hydroxyl of 3-hydroxydecanoate and the hydroxyl of rhamnose is reduced and the yield of rhamnose and 3-hydroxydecanoate in the hydrolysis process is improved by separating the oil phase containing 3-hydroxydecanoate in time in the hydrolysis reaction process of rhamnolipid.
2) Deeply hydrolyzing an oil phase obtained by hydrolyzing the rhamnolipid, so that a dimer of the 3-hydroxydecanoic acid which is difficult to separate from the 3-hydroxydecanoic acid can be further hydrolyzed into the 3-hydroxydecanoic acid, thereby improving the yield of the 3-hydroxydecanoic acid and simultaneously improving the purity of the 3-hydroxydecanoic acid.
3) The inventor finds that protein, oligopeptide, polypeptide and amino acid impurities can perform browning reaction with reaction products such as rhamnose and the like in the hydrolysis process to generate dark substances, and the generation of the reaction products reduces the quality yield of the rhamnose in the reaction process on one hand, and on the other hand, the inventor finds that the color number of the 3-hydroxydecanoic acid cannot be improved by a conventional means in the research process. The invention can keep the color number of the 3-hydroxydecanoate product generated by the reaction at a lower level and have lighter color by controlling the content of impurities such as protein, oligopeptide, polypeptide, amino acid and the like in the rhamnolipid raw material.
Description of the drawings:
FIG. 1 is a schematic diagram of the reaction scheme of the example; wherein 1, rhamnolipid solution is fed; 2. rhamnolipid hydrolysis acid feed; 3. a rhamnolipid hydrolysis reactor; 4. rhamnolipid hydrolysis reaction liquid; 5. a first oil-water separator; 6. a rhamnolipid hydrolysis water phase; 7. the rhamnolipid hydrolysis oil phase; 8. deeply hydrolyzing an acid feed by using an oil phase; 9. an oil phase deep hydrolysis reactor; 10. deeply hydrolyzing the reaction solution; 11. a second oil-water separator; 12.3-hydroxydecanoic acid crude product; 13. the aqueous phase is extensively hydrolyzed.
Detailed Description
The invention will now be further illustrated by the following examples, but is not limited thereto.
The analysis method comprises the following steps:
the analysis of the reaction mass in the process provided by the invention was carried out using ultra high performance liquid tandem triple quadrupole mass spectrometry (Xevo-TQD) as follows:
a chromatographic column: ACQUITY UPLC BEH Amide (1.7 μm, 3.0X 150 mm);
column temperature: 35 ℃;
flow rate: 0.3ml/min
Mobile phase and gradient: a is 0.1% ammonia water; b: acetonitrile;
0min-90％B，4min-50％B，7min-50％B，7.1min-90％B，12min-90％B
an ionization mode: electrospray ionization, negative ion mode;
the detection mode is as follows: multiple Reaction Monitoring (MRM)
Electrospray voltage: 2.8kv
Ion source temperature: 150 ℃ C
Temperature and flow rate of atomizing gas: 400 ℃ and 600L/h
And (3) quantitative ion pair: 163.06/102.97; taper hole voltage 16v and collision voltage 6v
And (3) qualitative ion pair: 163.06/59 (88.98); cone voltage 16v, impact voltage 6v.
The rhamnolipid concentration is determined by adopting an anthrone sulfuric acid method, the protein content is determined by adopting a Coomassie brilliant blue method, and the polypeptide, the oligopeptide and the amino acid are converted into the content of the protein.
And (3) rhamnolipid purification:
the rhamnolipids sold on the market contain a large amount of protein (protein, polypeptide, oligopeptide, amino acid) impurities because the rhamnolipids are products obtained by fermentation. The proteins in rhamnolipids can be isolated by, but not limited to, the following methods. For example, 500g of crude rhamnolipid having a protein, polypeptide, oligopeptide, and amino acid content of 15% was dissolved in 1000g of deionized water to adjust pH to 2.5, thereby obtaining 300g of rhamnolipid-containing precipitate, wherein the rhamnolipid content was 47.5% by the anthrone sulfuric acid method and the total protein, polypeptide, oligopeptide, and amino acid content was 5% by the Coomassie Brilliant blue method. And extracting the obtained rhamnolipid precipitate with 2000g of ethyl acetate to obtain an ethyl acetate solution containing rhamnolipid, and removing ethyl acetate by distillation to obtain 135g of rhamnolipid product with rhamnolipid content of 95%, wherein the total content of protein, polypeptide, oligopeptide and amino acid is 0.075%. Repeating the above extraction for 1 time to obtain rhamnolipid product with content of about 95%, and total content of protein, polypeptide, oligopeptide, and amino acid of 0.05%. Repeating the extraction for 2 times to obtain rhamnolipid product with content of about 95%, and total content of protein, polypeptide, oligopeptide, and amino acid of 0.035%.
Example 1
500g of rhamnolipid aqueous solution obtained by diluting the above purified rhamnolipid with deionized water, with a rhamnolipid concentration of 7.5wt%, and a total of proteins, polypeptides, amino acids, etc. accounting for 0.05wt% of the rhamnolipid mass, was added into a reactor 3 (1L) shown in FIG. 1. 7.5g of concentrated sulfuric acid with a concentration of 98% are added and stirred to 500 rpm. The temperature in the kettle was raised to 140 ℃ by means of an external heater. And (3) extracting a stream 4 at the beginning of the reaction, wherein the extraction rate of the stream 4 is 100ml/min, and controlling the oil phase content in the reaction system by controlling the velocity of the circulating stream. And (3) separating the oil phase containing the 3-hydroxydecanoic acid and the water phase containing the rhamnose, which are generated by the reaction, by a first oil-water separator 5, returning the water phase to the reactor 3, and weighing the oil phase content in the extracted stream 4 in the reaction process to be about 0.05wt%. The reaction was carried out for a total of 3 hours, and 18.0g of the oil phase was collected and analyzed by liquid chromatography external standard, wherein the content of 3-hydroxydecanoic acid was 81%.
18.0g of the oil phase obtained by the hydrolysis is added into a deep hydrolysis reactor 9, 0.5g of concentrated phosphoric acid with the concentration of 85wt% is added, the stirring speed is 500 r/min, the temperature is raised to 170 ℃, and the reaction is carried out for 1 hour. After the reaction, 15.4g of oil phase is collected, and the content of the 3-hydroxydecanoic acid is 95.4 percent by the analysis of external standard of liquid chromatography.
The color number of the platinum and the cobalt of the 3-hydroxydecanoate is analyzed to be 50 (GB/T3143-1982).
The rhamnolipid adopted in the reaction is a mixture of a plurality of rhamnolipid structures, and the average mass fraction of the 3-hydroxydecanoate structure is 40% through liquid chromatography external standard analysis.
The yield of 3-hydroxydecanoic acid was 98.1% of the theoretical yield.
Example 2
500g of rhamnolipid aqueous solution diluted with deionized water was added to a reactor 3 (1L) shown in FIG. 1, the concentration of rhamnolipid was detected to be 15%, and the total amount of protein, polypeptide, amino acid, etc. was 0.035% of the rhamnolipid mass. 10g of 85% concentrated sulfuric acid was added thereto, and the mixture was stirred to 500 rpm. The temperature in the kettle was raised to 130 ℃ by means of an external heater. The extraction rate of the stream 4 is 75ml/min, the oil phase containing 3-hydroxydecanoic acid and the water phase containing rhamnose which are generated by the reaction are separated by a first oil-water phase separator 5, the water phase returns to the reactor 3, and in the reaction process, the content of the oil in the stream 4 is detected to be 0.045wt%. The reaction was carried out for a total of 3 hours, and 36.0g of oil phase was collected and analyzed by liquid chromatography external standard, wherein the content of 3-hydroxydecanoic acid was 76%.
Adding 36.0g of oil phase obtained by hydrolysis into a deep hydrolysis reactor 9, adding 1.4g of 98% concentrated sulfuric acid, stirring at the rotating speed of 500 r/min, heating to 150 ℃, and reacting for 1.5 hours. 30.4g of oil phase is collected after the reaction is finished, and the content of the 3-hydroxydecanoic acid is 96.1 percent by the analysis of a liquid chromatography external standard. The platinum cobalt color number for 3-hydroxydecanoic acid was analyzed to be 45.
The rhamnolipid adopted in the reaction is a mixture of a plurality of rhamnolipid structures, and the average mass fraction of the 3-hydroxydecanoate structure is 40% through liquid chromatography external standard analysis.
The yield of 3-hydroxydecanoic acid was 97.5% of theory.
Example 3
500g of rhamnolipid aqueous solution diluted by deionized water is added into a reactor 3 (1L) shown in the attached figure, the concentration of the rhamnolipid is detected to be 25%, and the total amount of protein, polypeptide, amino acid and the like accounts for 0.075% of the mass of the rhamnolipid. 10g of concentrated hydrochloric acid having a concentration of 36.5% are added and stirred to 500 rpm. The temperature in the kettle was raised to 120 ℃ by an external heater. The extraction rate of the stream 4 is 50ml/min, the oil phase containing 3-hydroxydecanoic acid and the water phase containing rhamnose which are generated by the reaction are separated by an oil-water phase separator 5, the water phase returns to the reaction kettle 3, and the content of the oil in the stream 4 is detected to be about 0.1wt% in the reaction process. The reaction was carried out for a total of 4 hours, and 60.0g of the oil phase was collected and analyzed by liquid chromatography external standard, wherein the content of 3-hydroxydecanoic acid was 84%.
60.0g of the oil phase obtained by the hydrolysis is added into a deep hydrolysis reactor 9, 1.8g of concentrated hydrochloric acid with the concentration of 36.5 percent is added, the stirring speed is 500 r/min, the temperature is raised to 160 ℃, and the reaction is carried out for 2 hours. 51.5g of oil phase is collected after the reaction is finished, and the content of the 3-hydroxydecanoic acid is 94.3 percent and the color number of the platinum and the cobalt of the 3-hydroxydecanoic acid is 55 percent through the external standard analysis of liquid chromatography.
The rhamnolipid adopted in the reaction is a mixture of a plurality of rhamnolipid structures, and the average mass fraction of the 3-hydroxydecanoate structure is 40% through liquid chromatography external standard analysis.
The yield of 3-hydroxydecanoic acid was 97.1% of theory.
Example 4
500g of rhamnolipid aqueous solution diluted by deionized water is added into a reactor 3 (1L) shown in the attached figure, the concentration of the rhamnolipid is detected to be 7.5%, and the sum of protein, polypeptide, amino acid and the like accounts for 0.075% of the mass of the rhamnolipid. 7.5g of concentrated sulfuric acid with a concentration of 98% are added and stirred to 500 rpm. The temperature in the kettle was raised to 140 ℃ by means of an external heater. The extraction rate of the stream 4 is 100ml/min, the oil phase containing 3-hydroxydecanoic acid and the water phase containing rhamnose which are generated by the reaction are separated by an oil-water phase separator 5, the water phase returns to the reaction kettle 3, and the content of the oil in the stream 4 is detected to be about 0.05wt% in the reaction process. The reaction was carried out for a total of 3 hours, and 17.9g of oil phase was collected and analyzed by liquid chromatography external standard, wherein the content of 3-hydroxydecanoic acid was 82%.
Adding 17.9g of the oil phase obtained by hydrolysis into a deep hydrolysis reactor 9, adding 0.5g of concentrated phosphoric acid with the concentration of 85 percent, stirring at the rotating speed of 500 revolutions per minute, heating to 170 ℃, and reacting for 1 hour. 15.4g of oil phase is collected after the reaction is finished, and the content of the 3-hydroxydecanoic acid is 95.5 percent by the analysis of a liquid chromatography external standard. The color number of the platinum cobalt of the 3-hydroxydecanoic acid is analyzed to be 60. The rhamnolipid adopted in the reaction is a mixture of a plurality of rhamnolipid structures, and the average mass fraction of the 3-hydroxydecanoate structure is 40% through liquid chromatography external standard analysis.
The yield of 3-hydroxydecanoic acid was 97.8% of theory.
Comparative example 1
500g of rhamnolipid aqueous solution diluted with deionized water was added to a reactor 3 (1L) shown in FIG. 1, and the concentration of rhamnolipid was detected to be 7.5%, and the total amount of protein, polypeptide, amino acid, etc. was 0.05% of the mass of rhamnolipid. 7.5g of concentrated sulfuric acid with a concentration of 98% are added and stirred to 500 rpm. The temperature in the kettle was raised to 140 ℃ by means of an external heater. The reaction was carried out for a total of 3 hours and 18.5g of oil phase was collected and analyzed by liquid chromatography external standard to have a 3-hydroxydecanoic acid content of 65%. The rhamnolipid adopted in the reaction is a mixture of a plurality of rhamnolipid structures, and the average mass fraction of the 3-hydroxydecanoate structure is 40% through liquid chromatography external standard analysis. The color number of the platinum cobalt of the 3-hydroxydecanoic acid was analyzed to be 75. The yield of 3-hydroxydecanoic acid was 80.2%.
Example 5
500g of rhamnolipid aqueous solution diluted with deionized water was added to a reactor 3 (1L) shown in FIG. 1, the rhamnolipid concentration was 7.5%, and the sum of proteins, polypeptides, amino acids, etc. was 0.5% of the rhamnolipid mass. 7.5g of concentrated sulfuric acid with a concentration of 98% are added and stirred to 500 rpm. The temperature in the kettle was raised to 140 ℃ by means of an external heater. The extraction rate of the stream 4 is 100ml/min, the oil phase containing 3-hydroxydecanoic acid and the water phase containing rhamnose which are generated by the reaction are separated by an oil-water phase separator 5, the water phase returns to the reaction kettle 3, and the content of the oil in the stream 4 is detected to be about 0.05wt% in the reaction process. The reaction was carried out for a total of 3 hours and 18.8g of oil phase was collected and analyzed by liquid chromatography external standard, wherein the content of 3-hydroxydecanoic acid was 82%.
Adding 18.8g of the oil phase obtained by hydrolysis into a deep hydrolysis reactor 9, adding 0.6g of concentrated phosphoric acid with the concentration of 85 percent, stirring at the rotating speed of 500 revolutions per minute, heating to 170 ℃, and reacting for 1 hour. 15.3g of oil phase is collected after the reaction is finished, and the content of the 3-hydroxydecanoic acid is 95.6 percent by the analysis of a liquid chromatography external standard. The color number of the platinum cobalt of the 3-hydroxydecanoic acid is analyzed to be 450.
The rhamnolipid adopted in the reaction is a mixture of a plurality of rhamnolipid structures, and the average mass fraction of the 3-hydroxydecanoate structure is 40% through liquid chromatography external standard analysis.
The yield of 3-hydroxydecanoic acid was 97.6% of theory.
Example 6
500g of rhamnolipid aqueous solution diluted with deionized water was added to a reactor 3 (1L) shown in FIG. 1, and the concentration of rhamnolipid was detected to be 7.5%, and the total amount of protein, polypeptide, amino acid, etc. was 0.05% of the mass of rhamnolipid. 7.5g of concentrated sulfuric acid with a concentration of 98% are added and stirred to 500 rpm. The temperature in the kettle was raised to 140 ℃ by means of an external heater. The extraction rate of the stream 4 is 100ml/min, the oil phase containing 3-hydroxydecanoic acid and the water phase containing rhamnose which are generated by the reaction are separated by an oil-water phase separator 5, the water phase returns to the reaction kettle 3, and the content of the oil in the stream 4 is detected to be about 1wt% in the reaction process. The reaction was carried out for a total of 3 hours and 17.3g of oil phase was collected and analyzed by liquid chromatography external standard to show that the 3-hydroxydecanoic acid content was 65%.
Adding 17.3g of the oil phase obtained by hydrolysis into a deep hydrolysis reactor 9, adding 0.5g of concentrated phosphoric acid with the concentration of 85 percent, stirring at the rotating speed of 500 revolutions per minute, heating to 170 ℃, and reacting for 1 hour. 13.3g of oil phase is collected after the reaction is finished, and the content of the 3-hydroxydecanoic acid is 85.0 percent by the analysis of external standard of liquid chromatography. The 3-hydroxydecanoic acid was analyzed for a platinum cobalt color number of 60.
The rhamnolipid adopted in the reaction is a mixture of a plurality of rhamnolipid structures, and the average mass fraction of the 3-hydroxydecanoate structure is 40% through liquid chromatography external standard analysis.
The yield of 3-hydroxydecanoic acid was 75.5% of theory.
Claims (18)
1. A method for preparing 3-hydroxydecanoic acid by hydrolysis of rhamnolipids, the method comprising:
1) The rhamnolipid aqueous solution is subjected to hydrolysis reaction under the catalysis of acid to obtain a water phase containing rhamnose and an oil phase containing 3-hydroxydecanoic acid;
2) Separating the oil phase containing the 3-hydroxydecanoic acid from the reaction system, and carrying out deep hydrolysis reaction under the catalysis of acid to generate the 3-hydroxydecanoic acid;
wherein, the rhamnolipid in the step 1) is hydrolyzed to generate an oil phase containing 3-hydroxydecanoic acid, and the step 2) is carried out before the mass of the oil phase accounts for 0.5 percent of the mass of the rhamnolipid aqueous solution raw material.
2. The method according to claim 1, wherein the total amount of proteins, polypeptides, oligopeptides and amino acids in the rhamnolipid material is not more than 0.1% by mass of rhamnolipid.
3. The method as claimed in claim 2, wherein the sum of the proteins, polypeptides, oligopeptides and amino acids in the rhamnolipid material is 0.01-0.1% by mass of rhamnolipid.
4. The method as claimed in claim 1, wherein the rhamnolipid in the rhamnolipid aqueous solution has a mass fraction of 5-40%.
5. The method as claimed in claim 4, wherein the rhamnolipid in the rhamnolipid aqueous solution has a mass fraction of 7.5% -25%.
6. The method of claim 1, wherein the acid in steps) 1 and 2) is one or more of sulfuric acid, phosphoric acid, hydrochloric acid, naphthalene sulfonic acid, and p-toluenesulfonic acid.
7. The method of claim 6, wherein the acid in steps) 1 and 2) is one or more of sulfuric acid, phosphoric acid and hydrochloric acid.
8. The method according to claim 1 or 6, wherein the acid is added in step 1) in an amount of 0.5 to 5% by mass of the rhamnolipid solution.
9. The method as claimed in claim 8, wherein the acid is added in step 1) in an amount of 1-2% by mass of the rhamnolipid solution.
10. The method according to claim 1, wherein the reaction temperature of the rhamnolipid hydrolysis step in the 1) step is 100 ℃ to 150 ℃; the hydrolysis reaction time is 1-10h.
11. The method as claimed in claim 10, wherein the reaction temperature of the rhamnolipid hydrolysis step in the 1) step is 120 ℃ to 140 ℃; the hydrolysis reaction time is 2-4h.
12. The method according to any one of claims 1 to 7, wherein step 2) is carried out before the oil phase has a mass of 0.1% of the mass of the rhamnolipid aqueous solution raw material.
13. The method according to claim 12, wherein the reaction temperature of the deep oil phase hydrolysis step of step 2) is 140 ℃ to 180 ℃.
14. The method of claim 13, wherein the reaction temperature of the deep oil phase hydrolysis step of step 2) is 150 ℃ to 170 ℃.
15. The method of claim 13, wherein the acid is added in the step of deep oil phase hydrolysis in step 2) in an amount of 1% to 5% by mass of the oil phase.
16. The method of claim 15, wherein the acid is added in the step of deep oil phase hydrolysis in step 2) in an amount of 2% to 4% by mass of the oil phase.
17. The method of claim 15, wherein the hydrolysis reaction time in step 2) is 0.5 to 5 hours.
18. The method of claim 17, wherein the hydrolysis reaction time in step 2) is 1 to 2 hours.
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| US4933281A (en) * | 1987-03-17 | 1990-06-12 | The University Of Iowa Research Foundation | Method for producing rhamnose |
| US5501966A (en) * | 1992-06-25 | 1996-03-26 | Hoechst Aktiengesellschaft | Pseudomonas aeruginosa and its use in a process for the biotechnological preparation of L-rhamnose |
| WO2000029604A1 (en) * | 1998-11-18 | 2000-05-25 | The University Of Akron | Production of biological materials by simultaneous aerobic and anaerobic respiration |
| CN108587989A (en) * | 2018-03-21 | 2018-09-28 | 中国科学院微生物研究所 | A kind of bacterial strain and production method for producing 3- hydroxydecanoic acids |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| US4933281A (en) * | 1987-03-17 | 1990-06-12 | The University Of Iowa Research Foundation | Method for producing rhamnose |
| US5501966A (en) * | 1992-06-25 | 1996-03-26 | Hoechst Aktiengesellschaft | Pseudomonas aeruginosa and its use in a process for the biotechnological preparation of L-rhamnose |
| WO2000029604A1 (en) * | 1998-11-18 | 2000-05-25 | The University Of Akron | Production of biological materials by simultaneous aerobic and anaerobic respiration |
| CN108587989A (en) * | 2018-03-21 | 2018-09-28 | 中国科学院微生物研究所 | A kind of bacterial strain and production method for producing 3- hydroxydecanoic acids |
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