CN117701661A - Production and preparation process of enzyme modified isoquercitrin - Google Patents
Production and preparation process of enzyme modified isoquercitrin Download PDFInfo
- Publication number
- CN117701661A CN117701661A CN202311744861.6A CN202311744861A CN117701661A CN 117701661 A CN117701661 A CN 117701661A CN 202311744861 A CN202311744861 A CN 202311744861A CN 117701661 A CN117701661 A CN 117701661A
- Authority
- CN
- China
- Prior art keywords
- enzyme
- isoquercitrin
- enzyme modified
- isoquercetin
- modified isoquercitrin
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- OVSQVDMCBVZWGM-QSOFNFLRSA-N quercetin 3-O-beta-D-glucopyranoside Chemical class O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1=C(C=2C=C(O)C(O)=CC=2)OC2=CC(O)=CC(O)=C2C1=O OVSQVDMCBVZWGM-QSOFNFLRSA-N 0.000 title claims abstract description 114
- 108090000790 Enzymes Proteins 0.000 title claims abstract description 94
- 102000004190 Enzymes Human genes 0.000 title claims abstract description 94
- 238000002360 preparation method Methods 0.000 title claims abstract description 18
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 12
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 38
- 238000013375 chromatographic separation Methods 0.000 claims abstract description 25
- OVSQVDMCBVZWGM-IDRAQACASA-N Hirsutrin Natural products O([C@H]1[C@H](O)[C@H](O)[C@H](O)[C@@H](CO)O1)C1=C(c2cc(O)c(O)cc2)Oc2c(c(O)cc(O)c2)C1=O OVSQVDMCBVZWGM-IDRAQACASA-N 0.000 claims abstract description 23
- FVQOMEDMFUMIMO-UHFFFAOYSA-N Hyperosid Natural products OC1C(O)C(O)C(CO)OC1OC1C(=O)C2=C(O)C=C(O)C=C2OC1C1=CC=C(O)C(O)=C1 FVQOMEDMFUMIMO-UHFFFAOYSA-N 0.000 claims abstract description 23
- 239000000243 solution Substances 0.000 claims abstract description 23
- 239000000126 substance Substances 0.000 claims abstract description 23
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 claims abstract description 22
- 239000003456 ion exchange resin Substances 0.000 claims abstract description 22
- 229920003303 ion-exchange polymer Polymers 0.000 claims abstract description 22
- 238000000034 method Methods 0.000 claims abstract description 22
- 239000002994 raw material Substances 0.000 claims abstract description 19
- OVSQVDMCBVZWGM-QCKGUQPXSA-N isoquercetin Natural products OC[C@@H]1O[C@@H](OC2=C(Oc3cc(O)cc(O)c3C2=O)c4ccc(O)c(O)c4)[C@H](O)[C@@H](O)[C@@H]1O OVSQVDMCBVZWGM-QCKGUQPXSA-N 0.000 claims abstract description 17
- 239000007864 aqueous solution Substances 0.000 claims abstract description 14
- 238000006243 chemical reaction Methods 0.000 claims abstract description 14
- 239000000463 material Substances 0.000 claims abstract description 14
- 229920001542 oligosaccharide Polymers 0.000 claims abstract description 14
- 150000002482 oligosaccharides Chemical class 0.000 claims abstract description 14
- 238000007865 diluting Methods 0.000 claims abstract description 9
- 239000004375 Dextrin Substances 0.000 claims abstract description 8
- 229920001353 Dextrin Polymers 0.000 claims abstract description 8
- 235000019425 dextrin Nutrition 0.000 claims abstract description 8
- 238000004458 analytical method Methods 0.000 claims abstract description 7
- 239000011347 resin Substances 0.000 claims abstract description 7
- 229920005989 resin Polymers 0.000 claims abstract description 7
- 238000004821 distillation Methods 0.000 claims abstract description 6
- 238000001035 drying Methods 0.000 claims abstract description 4
- 238000001694 spray drying Methods 0.000 claims abstract description 4
- 238000004587 chromatography analysis Methods 0.000 claims abstract description 3
- 239000007788 liquid Substances 0.000 claims description 15
- 239000007787 solid Substances 0.000 claims description 13
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 12
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 11
- GXMWXESSGGEWEM-UHFFFAOYSA-N isoquercitrin Natural products OCC(O)C1OC(OC2C(Oc3cc(O)cc(O)c3C2=O)c4ccc(O)c(O)c4)C(O)C1O GXMWXESSGGEWEM-UHFFFAOYSA-N 0.000 claims description 6
- 230000008569 process Effects 0.000 claims description 5
- 238000011010 flushing procedure Methods 0.000 claims description 4
- 239000008213 purified water Substances 0.000 claims description 4
- 238000004064 recycling Methods 0.000 claims description 3
- 239000007921 spray Substances 0.000 claims description 3
- 230000005526 G1 to G0 transition Effects 0.000 claims description 2
- 239000004793 Polystyrene Substances 0.000 claims description 2
- 239000011159 matrix material Substances 0.000 claims description 2
- 238000002156 mixing Methods 0.000 claims description 2
- 229920002223 polystyrene Polymers 0.000 claims description 2
- 230000000052 comparative effect Effects 0.000 description 12
- 230000035484 reaction time Effects 0.000 description 9
- 108090000637 alpha-Amylases Proteins 0.000 description 8
- 239000012535 impurity Substances 0.000 description 8
- 108010019077 beta-Amylase Proteins 0.000 description 6
- 102000000340 Glucosyltransferases Human genes 0.000 description 5
- 108010055629 Glucosyltransferases Proteins 0.000 description 5
- 238000002835 absorbance Methods 0.000 description 5
- 239000012295 chemical reaction liquid Substances 0.000 description 4
- 238000005259 measurement Methods 0.000 description 3
- 239000011343 solid material Substances 0.000 description 3
- 238000004090 dissolution Methods 0.000 description 2
- 230000002255 enzymatic effect Effects 0.000 description 2
- 125000002791 glucosyl group Chemical group C1([C@H](O)[C@@H](O)[C@H](O)[C@H](O1)CO)* 0.000 description 2
- 238000006116 polymerization reaction Methods 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 238000001179 sorption measurement Methods 0.000 description 2
- 239000012086 standard solution Substances 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 240000000797 Hibiscus cannabinus Species 0.000 description 1
- 241000219071 Malvaceae Species 0.000 description 1
- 241001529936 Murinae Species 0.000 description 1
- 241000207834 Oleaceae Species 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 230000000259 anti-tumor effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000009144 enzymatic modification Effects 0.000 description 1
- 229930003935 flavonoid Natural products 0.000 description 1
- -1 flavonoid compound Chemical class 0.000 description 1
- 235000017173 flavonoids Nutrition 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 238000002715 modification method Methods 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Saccharide Compounds (AREA)
Abstract
The invention relates to the technical field of preparation of enzyme modified isoquercitrin, and discloses a production and preparation process of enzyme modified isoquercitrin, which comprises the following steps: s1: the preparation method comprises the following steps: taking isoquercetin and dextrin substances as raw materials, and preparing enzyme modified isoquercetin by an enzyme conversion method; s2: removing oligosaccharide: diluting the raw material in the step S1, passing the diluted solution through an ion exchange resin column, analyzing the resin column with the completely adsorbed alcohol, and removing the alcohol in the analysis solution through reduced pressure distillation, wherein the step S3: collecting enzyme modified isoquercetin: separating and extracting enzyme modified isoquercitrin from the aqueous solution after removing the alcohol in the step S2 by chromatography and collecting the enzyme modified isoquercitrin; s4: concentrating: concentrating the material containing the enzyme modified isoquercitrin Pi Sugao collected in the step S3; s5: and (3) drying: spray drying the concentrated material in the step S4; the invention adopts ion exchange resin and chromatographic separation method to separate and purify the enzyme modified isoquercitrin, and can improve the purity of the enzyme modified isoquercitrin to more than 90 percent, thereby ensuring better stability of the product.
Description
Technical Field
The invention relates to the technical field of preparation of enzyme modified isoquercitrin, in particular to a production and preparation process of enzyme modified isoquercitrin.
Background
Isoquercetin is a flavonoid compound which is very rare in nature but has remarkable biological activities such as oxidation resistance, anti-tumor and the like, exists in flowers of malvaceae plants and leaves of oleaceae plants of kenaf, but has poor water solubility and also affects the application, in order to improve the water solubility, people add the isoquercetin into hydrophilic groups by an enzyme modification method so as to increase the water solubility, and most of Enzyme Modified Isoquercetin (EMIQ) on the market at present has the content of 50-60%, and impurities of the enzyme modified isoquercetin are mainly isoquercetin and oligosaccharides, and the impurities influence the stability of an EMIQ aqueous solution, and meanwhile, the finished product of the EMIQ is easier to absorb moisture and agglomerate, so that the invention is specially proposed for solving the problems.
Disclosure of Invention
The invention aims to provide a production and preparation process of enzyme modified isoquercitrin, which aims to solve the problems in the prior art: the content of most Enzyme Modified Isoquercetin (EMIQ) in the market at present is 53-82%, and impurities of the enzyme modified isoquercetin mainly comprise isoquercetin and oligosaccharides, and the impurities influence the stability of an EMIQ aqueous solution, and meanwhile, the finished product of the EMIQ is easier to absorb moisture and agglomerate.
In order to achieve the above purpose, the present invention provides the following technical solutions: a production and preparation process of enzyme modified isoquercitrin comprises the following steps:
s1: the preparation method comprises the following steps: taking isoquercetin and dextrin substances as raw materials, preparing enzyme modified isoquercetin by an enzyme conversion method, wherein the content of the enzyme modified isoquercetin in the prepared solid substances of the reaction liquid is 40-60wt%;
s2: removing oligosaccharide: diluting the raw material in the step S1, passing the diluted solution through an ion exchange resin column, analyzing the resin column completely adsorbed by alcohol, and removing the alcohol in the analysis solution by reduced pressure distillation to obtain the solid substance enzyme modified isoquercitrin with the content of 53-82wt%;
s3: collecting enzyme modified isoquercetin: separating and extracting enzyme modified isoquercitrin from the aqueous solution after removing the alcohol in the step S2 by chromatography and collecting the enzyme modified isoquercitrin;
s4: concentrating: concentrating the material containing the enzyme modified isoquercitrin Pi Sugao collected in the step S3;
s5: and (3) drying: and (3) carrying out spray drying on the concentrated material in the step S4.
According to the technical scheme, the isoquercetin and the dextrin substances are used as raw materials, the enzyme modified isoquercetin is prepared through an enzyme conversion method, the content of enzyme modified isoquercetin in the prepared solid substances in the reaction liquid is 40-60wt%, at the moment, the content of main products (enzyme modified isoquercetin) is low, the content of impurities (isoquercetin is high and the content of oligosaccharides substances) is high, the obtained raw materials are diluted, so that the enzyme modified isoquercetin is better adsorbed through an ion exchange resin column, the enzyme modified isoquercetin is adsorbed in the ion exchange resin column, a small amount of impurities are simultaneously adsorbed in the ion exchange resin column, the ion exchange resin column with complete adsorption is analyzed through alcohol, the enzyme modified isoquercetin is separated out through alcohol analysis, meanwhile, a small amount of impurities are contained in the analysis liquid, the alcohol in the analysis liquid is removed through reduced pressure distillation, and at the moment, the content of the enzyme modified isoquercetin in the solid substances in the obtained liquid is 53-82wt%; the aqueous solution after alcohol removal is subjected to chromatographic separation to extract enzyme modified isoquercitrin and collection, and the purity of the enzyme modified isoquercitrin is improved to more than 90wt%, so that the ion exchange resin and the chromatographic separation method are adopted to separate and purify the enzyme modified isoquercitrin, the purity of the enzyme modified isoquercitrin can be improved to more than 90wt%, the stability of the product is better, the quality of the obtained product is more stable, the water solubility is better, and the dissolution rate of the product in the aqueous solution is mainly embodied to be very fast.
Preferably, the enzyme is one or a combination of several of glucosyltransferase, pullulanase or beta-amylase.
By adopting the technical scheme, the method is preferably one or a combination of several of glucosyltransferase, pullulanase or beta-amylase, the dextrin is formed by connecting a plurality of glucose structures, the dextrin is cut into glucose units with different polymerization degrees through the biological enzyme cutting action, and then the glucose with different polymerization degrees is connected to the hydroxyl of the isoquercetin through the biological enzyme.
Preferably, the enzymes are glucosyltransferase, pullulanase and beta-amylase.
By adopting the technical scheme, the enzymes are preferably glucosyltransferase, pullulanase and beta-amylase, the enzyme conversion reaction time is reduced by adopting the complex enzyme, and the purity of the obtained enzyme modified isoquercitrin is improved.
Preferably, the S2 feedstock is diluted to an aqueous solution having a solids content of 10-15wt% and the dilution is passed through the ion exchange resin column at a rate of 0.1-0.5 column/hour.
By adopting the technical scheme, the concentration of the diluted raw materials and the speed of the diluted liquid passing through the ion exchange resin column are optimized, so that the ion exchange resin column can absorb the diluted liquid better.
Preferably, the alcohol in S2 is methanol or ethanol, and the alcohol is in the range of 60-80wt%.
By adopting the technical scheme, the alcohol is preferably methanol or ethanol, the methanol or ethanol has good solubility for enzyme modified isoquercitrin, however, the solubility for impurities (isoquercitrin is an oligosaccharide substance) is poor, so that the enzyme modified isoquercitrin in the ion exchange resin column can be resolved, the concentration range of the alcohol is further preferably 60-80vol%, and the resolving performance of the alcohol on the ion exchange resin column with complete absorption is further improved.
Preferably, the specific method of S3 is as follows: diluting the aqueous solution after removing alcohol in S2 to 5-10wt% concentration, performing chromatographic separation feeding, connecting the chromatographic separation system by 8 chromatographic columns in series to form a closed system connected end to end, wherein the diameter-to-diameter ratio of each chromatographic column is 1:10, the feeding speed of each chromatographic column is 1-2 times of column/hour, the feeding speed of chromatographic separation is 5-8 times of column/hour, purified water is used as flushing liquid, the moving speed of enzyme modified isoquercetin in the mobile phase is faster than that of isoquercetin, collecting effluent at the outlet of the last chromatographic column of the chromatographic separation system, collecting after the enzyme modified isoquercetin is found to flow out, measuring the enzyme modified isoquercetin by an ultraviolet spectrophotometer, changing one collecting bottle every half an hour, continuously collecting for 6-8 hours, and measuring the components in each collecting bottle.
Preferably, the stationary phase used in the chromatographic separation system is Rogowski chromatographic separation resin, and the matrix is crosslinked polystyrene.
Preferably, the specific operation method of S4 is as follows: uniformly mixing the material liquid in the collecting bottle with the enzyme modified isoquercitrin in S3 accounting for more than 90 percent by weight of solid substance, and concentrating in vacuum until the concentration of the material is 40 to 60 percent by weight.
Preferably, the spray inlet temperature is 130 ℃ to 140 ℃.
Preferably, S6: recycling: the oligosaccharides eluted in S2 and the isoquercitrin remaining in S3 are reused as raw materials for preparing the enzyme-modified isoquercitrin.
Compared with the prior art, the invention has the beneficial effects that:
according to the invention, the ion exchange resin and the chromatographic separation method are adopted to separate and purify the enzyme modified isoquercitrin, so that the purity of the enzyme modified isoquercitrin can be improved to more than 90wt%, the stability of the product is better, the quality of the obtained product is more stable, the water solubility is better, the dissolution rate of the product in aqueous solution is fast, the conventional product needs to be vigorously stirred for a long time to be completely dissolved, and after the conventional product is placed for a long time, the isoquercitrin contained in the conventional product is separated out, and the product is relatively stable even if placed for a long time due to the low content of the isoquercitrin.
Drawings
FIG. 1 is a flow chart of a production and preparation process of enzyme modified isoquercitrin in the present invention.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Referring to fig. 1, the invention provides a production and preparation process of enzyme modified isoquercitrin, which comprises the following steps:
s1: the preparation method comprises the following steps: taking isoquercetin and dextrin substances as raw materials, adding enzyme into the raw materials, and preparing enzyme modified isoquercetin by an enzyme conversion method, wherein the content of the enzyme modified isoquercetin in the prepared solid substances of the reaction liquid is 40-60wt%;
s2: removing oligosaccharide: diluting the raw material in S1 to 10-15wt% water solution, passing the diluted solution through an ion exchange resin column, wherein the speed of the diluted solution passing through the ion exchange resin column is 0.1-0.5 times of the column/hour, resolving the resin column with 60-80wt% alcohol, and removing alcohol in the resolved solution by reduced pressure distillation to obtain the solid substance enzyme modified isoquercitrin with 53-82wt% in the liquid;
s3: collecting enzyme modified isoquercetin: diluting the aqueous solution after removing alcohol in S2 to 5-10wt% concentration, carrying out chromatographic separation feeding, connecting a chromatographic separation system by 8 chromatographic columns in series to form a closed system connected end to end, wherein the diameter-to-diameter ratio of each chromatographic column is 1:10, the feeding speed is 1-2 times of column/hour, the feeding amount of chromatographic separation is 5-8 times of column/hour, purified water is used as flushing liquid, the moving speed of enzyme modified isoquercetin in a mobile phase is faster than that of isoquercetin, collecting effluent liquid at the outlet of the last chromatographic column of the chromatographic separation system, collecting after the enzyme modified isoquercetin is found to flow out, measuring the enzyme modified isoquercetin by an ultraviolet spectrophotometer, changing a collecting bottle every half an hour, continuously collecting the components in each collecting bottle, and measuring the components in each collecting bottle, wherein the content of the enzyme modified isoquercetin in the solution in the collecting bottle accounts for more than 90% wt% of solid substances;
s4: concentrating: concentrating the material containing the enzyme modified isoquercitrin Pi Sugao collected in the step S3 to obtain a material concentration of 40-60wt%;
s5: and (3) drying: spray drying the concentrated material in the step S4, wherein the spray inlet temperature is 130-140 ℃;
s6: recycling: the oligosaccharides eluted in S2 and the isoquercitrin remaining in S3 are reused as raw materials for preparing the enzyme-modified isoquercitrin.
Examples
Example 1
S1: the preparation method comprises the following steps: taking 1kg of isoquercetin and 1kg of dextrin substances as raw materials, adding 5ml of glucosyltransferase, 3ml of pullulanase and 3ml of beta-amylase into the raw materials, and preparing enzyme modified isoquercetin by an enzyme conversion method, wherein the content of the enzyme modified isoquercetin in the prepared solid substances of the reaction liquid is 60wt%, the content of the isoquercetin is 20wt% and the content of the oligosaccharide substances is 20wt%;
s2: removing oligosaccharide: diluting the raw materials in S1 to a water solution with the concentration of 13wt%, passing the diluted solution through an ion exchange resin column, wherein the speed of the diluted solution passing through the ion exchange resin column is 0.3 times of the speed of the ion exchange resin column per hour, analyzing the resin column completely adsorbed by 70wt% of ethanol, and removing the ethanol in the analysis solution by reduced pressure distillation to obtain the liquid with the solid material enzyme modified isoquercetin content of 82wt%, the isoquercetin content of 15wt% and the oligosaccharide material content of 3wt%;
s3: collecting enzyme modified isoquercetin: diluting the aqueous solution after alcohol removal in S2 to concentration of 8wt%, carrying out chromatographic separation feeding, connecting a chromatographic separation system by 8 chromatographic columns in series to form a closed system connected end to end, wherein the diameter-to-diameter ratio of each chromatographic column is 1:10, the feeding speed is 2 times of column/hour, the feeding amount of chromatographic separation is 7 times of column, purified water is used as flushing liquid, the moving speed of enzyme modified isoquercetin in a mobile phase is faster than that of isoquercetin, collecting effluent at the outlet of the last chromatographic column of the chromatographic separation system, collecting after the enzyme modified isoquercetin is found to flow out, measuring the enzyme modified isoquercetin by an ultraviolet spectrophotometer, changing a collecting bottle every half hour, continuously collecting for 8 hours, measuring the components in each collecting bottle, and the content of enzyme modified isoquercetin in the solution in the collecting bottle is 95wt%.
As shown in Table 1, examples 1 to 10 are different in that the enzyme used in step S1 is different
Comparative example
Comparative example 1
This comparative example differs from example 1 in that the sequence of step 2 and step 3 was exchanged, chromatographic separation was performed first, and then adsorption was performed by passing through an ion exchange resin column.
Comparative example 2
This comparative example differs from example 1 in that step S3 was omitted, directly concentrated and dried.
Comparative example 3
This comparative example differs from example 1 in that pullulanase was replaced with murine Li Tangmei.
Test method
Determining the content of substances in the collected feed liquid in each step by an ultraviolet spectrophotometer:
1. preparing a sample: dissolving a substance to be tested in a proper solvent to prepare a solution with a certain concentration;
2. drawing a standard curve: preparing standard solutions with different concentrations, measuring the absorbance of the standard solutions on an ultraviolet spectrophotometer, and drawing a relation curve between the absorbance and the concentration;
3. measuring the absorbance of the sample: placing the solution to be measured in a sample cell of an ultraviolet spectrophotometer, and measuring the absorbance of the solution;
4. sample concentrations were calculated from standard curves: comparing the measured absorbance with a standard curve to determine the concentration of the solution to be measured;
it should be noted that in the case of measurement using an ultraviolet spectrophotometer, it is necessary to select an appropriate wavelength and an appropriate solvent, and to pay attention to the stability of the solution, and at the same time, in order to improve the accuracy of measurement, it is possible to perform measurement a plurality of times and take an average value.
The reaction times of the enzymatic conversion process are recorded by the table: the reaction time and the corresponding conversion rate are recorded, and the corresponding conversion rate tends to be balanced along with the change of the reaction time, and the inflection point of the conversion rate is the reaction time of the enzyme conversion method.
The content of the enzyme-modified isoquercetin in the solid substance in the steps corresponding to examples 1 to 10 and comparative examples 1 to 2 and the reaction time of the enzyme conversion method are shown in Table 2.
TABLE 2 reaction time of step S1 and content of enzyme-modified isoquercitrin in solid substance in each step of the reaction
As can be seen from the combination of examples 1 to 10 and comparative example 3 and Table 2, the content of the enzyme-modified isoquercetin in the solid material in the corresponding step in example 1 is greater than that in examples 2 to 10, the content of the enzyme-modified isoquercetin in the solid material in step S3 in example 1 is as high as 95wt%, and the reaction time is also the shortest (24 hours), which means that the effect of combining the enzyme pullulanase with the other two enzymes pullulanase and beta-amylase is better than that of combining the other two enzymes singly or with the other two enzymes, and the three enzymes have a synergistic effect, so that the conversion rate of the enzymatic conversion reaction can be improved and the reaction time can be shortened.
As can be seen from the combination of example 1 and comparative example 1 and the combination of Table 2, after the order of step S2 and step S3 in example 1 was changed, the purity of the enzyme-modified isoquercitrin obtained in example 1 was greater than that in comparative example 1, and the order of step S2 and step S3 in the specification affected the purity of the enzyme-modified isoquercitrin product.
It can be seen by combining example 1 and comparative example 2 in combination with Table 2 that the purity of the enzyme-modified isoquercitrin product can be further enhanced by chromatographic separation techniques and can be increased to 95% by weight.
Although embodiments of the present invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made therein without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (8)
1. The production and preparation process of the enzyme modified isoquercitrin is characterized by comprising the following steps:
s1: the preparation method comprises the following steps: taking isoquercetin and dextrin substances as raw materials, and preparing enzyme modified isoquercetin by an enzyme conversion method;
s2: removing oligosaccharide: diluting the raw materials in the step S1, passing the diluted solution through an ion exchange resin column, analyzing the resin column completely adsorbed by alcohol, and removing the alcohol in the analysis solution by reduced pressure distillation;
s3: collecting enzyme modified isoquercetin: separating and extracting enzyme modified isoquercitrin from the aqueous solution after removing the alcohol in the step S2 by chromatography and collecting the enzyme modified isoquercitrin;
s4: concentrating: concentrating the material containing the enzyme modified isoquercitrin Pi Sugao collected in the step S3;
s5: and (3) drying: and (3) carrying out spray drying on the concentrated material in the step S4.
2. The process for producing and preparing the enzyme-modified isoquercitrin according to claim 1, wherein the raw material in S2 is diluted to a 10-15wt% aqueous solution, and the speed of passing the diluted solution through the ion exchange resin column is 0.1-0.5 times the column speed per hour.
3. The process for producing enzymatically modified isoquercitrin of claim 1 wherein said alcohol in S2 is methanol or ethanol and said alcohol is present in a concentration range of 60-80vol%.
4. The production and preparation process of the enzyme modified isoquercitrin, according to claim 1, characterized in that the specific method of S3 is as follows: diluting the aqueous solution after alcohol removal in S2 to 5-10wt% concentration, carrying out chromatographic separation feeding, connecting 8 chromatographic columns in series by a chromatographic separation system to form a closed system connected end to end, wherein the diameter-to-height ratio of each chromatographic column is 1:10, the feeding speed is 1-2 times of column/hour, the feeding amount of chromatographic separation is 5-8 times of column/hour, taking purified water as flushing liquid, collecting effluent at the outlet of the last chromatographic column of the chromatographic separation system, starting collecting after the enzyme-modified isoquercetin is found, measuring the enzyme-modified isoquercetin by an ultraviolet spectrophotometer, changing one collecting bottle every half hour, continuously collecting for 6-8 hours, and measuring the components in each collecting bottle.
5. The process for preparing enzymatically modified isoquercitrin in accordance with claim 4, wherein said stationary phase used in said chromatographic separation system is Rogowski chromatographic resin and said matrix is crosslinked polystyrene.
6. The production and preparation process of the enzyme modified isoquercitrin, according to claim 1, characterized in that the specific operation method of S4 is as follows: uniformly mixing the material liquid in the collecting bottle with the enzyme modified isoquercitrin in S3 accounting for more than 90 percent by weight of solid substance, and concentrating in vacuum until the concentration of the material is 40 to 60 percent by weight.
7. The process for producing and preparing enzyme-modified isoquercitrin in accordance with claim 1, wherein said spray inlet temperature is 130 ℃ to 140 ℃.
8. The process for producing and preparing the enzyme-modified isoquercitrin of claim 1, further comprising S6: recycling: the oligosaccharides eluted in S2 and the isoquercitrin remaining in S3 are reused as raw materials for preparing the enzyme-modified isoquercitrin.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211691437.5A CN115976140A (en) | 2022-12-27 | 2022-12-27 | Production and preparation process of enzyme-modified isoquercitrin |
| CN2022116914375 | 2022-12-27 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN117701661A true CN117701661A (en) | 2024-03-15 |
Family
ID=85964438
Family Applications (2)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211691437.5A Pending CN115976140A (en) | 2022-12-27 | 2022-12-27 | Production and preparation process of enzyme-modified isoquercitrin |
| CN202311744861.6A Pending CN117701661A (en) | 2022-12-27 | 2023-12-18 | Production and preparation process of enzyme modified isoquercitrin |
Family Applications Before (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211691437.5A Pending CN115976140A (en) | 2022-12-27 | 2022-12-27 | Production and preparation process of enzyme-modified isoquercitrin |
Country Status (1)
| Country | Link |
|---|---|
| CN (2) | CN115976140A (en) |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101525956B1 (en) * | 2012-10-19 | 2015-06-08 | 한국생명공학연구원 | preparation method of quercetin or isoquecitrin using β-glucosidase |
| CN110305925A (en) * | 2019-07-12 | 2019-10-08 | 天津宏顺科生物科技有限公司 | Preparation process of enzyme modified rutin mixture |
| CN115011654A (en) * | 2022-03-31 | 2022-09-06 | 成都欧康医药股份有限公司 | Method for preparing isoquercitrin by carrying out enzymolysis on sophora flower buds |
| CN114686549A (en) * | 2022-04-29 | 2022-07-01 | 陕西嘉禾生物科技股份有限公司 | Method for preparing enzyme modified isoquercitrin by using rutin |
-
2022
- 2022-12-27 CN CN202211691437.5A patent/CN115976140A/en active Pending
-
2023
- 2023-12-18 CN CN202311744861.6A patent/CN117701661A/en active Pending
Also Published As
| Publication number | Publication date |
|---|---|
| CN115976140A (en) | 2023-04-18 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN1320122C (en) | Process for extracting xylose and xylitol from a xylose mother liquor or a xylose digest | |
| CN110283226B (en) | Method for extracting antioxidant component in rosemary | |
| CN101792822B (en) | Method for separating and purifying xylose and arabinose from hemicellulose acid hydrolysis liquid | |
| US9926613B2 (en) | Method of producing sugar solution | |
| CN101322758B (en) | Method for producing spina date seed extract using complex enzyme | |
| CN103274992B (en) | A kind of method of combining membrane sepn and column chromatography technology and preparing high purity 1-DNJ | |
| CN108642117B (en) | Method for extracting limonin and nomilin | |
| US20240025937A1 (en) | Preparation method of peonidin-acylated anthocyanin and malvidin-acylated anthocyanin | |
| CN104892717B (en) | A kind of technical grade preparative liquid chromatography separation method of momordica glycoside V | |
| CN111574639B (en) | Method for separating and purifying nitraria tangutorum bobr polysaccharide | |
| CN113686990A (en) | Method for extracting hesperidin from citrus to prepare hesperetin | |
| CN117701661A (en) | Production and preparation process of enzyme modified isoquercitrin | |
| CN106226426A (en) | A kind of high performance liquid chromatography splits the method for canagliflozin five-membered ring impurity enantiomer | |
| QIU et al. | Apple pectin behavior separated by ultrafiltration | |
| CN101366829A (en) | Method for synchronously extracting flavone and alkaloid from folium nelumbinis | |
| JP2023542661A (en) | Method for separating and purifying p-coumaroylated anthocyanin | |
| Li et al. | Determination of seventeen sugars and sugar alcohols in fruit juice samples using hydrophilic interaction liquid chromatography-tandem mass spectrometry combining response surface methodology design | |
| CN102590210B (en) | Method for measuring content of alpha-glucosin cane juice | |
| Oriez et al. | Sugarcane bagasse mild alkaline fractionation and production of purified fractions by pulse chromatography with water | |
| US20240115971A1 (en) | Method for Extracting and Separating Various Components from Flaxseed Meal based on Subcritical Composite Solvent | |
| JP2007520195A (en) | A method for recovering galactose from a solution derived from plant-based biomass using a chromatographic fractionation step and crystallization. | |
| CN103145771A (en) | Method for extracting D-ribose from fermentation liquor by ultrafiltration and ion exchange technologies | |
| CN112079886A (en) | Method for improving purity of xylose and arabinose by chromatographic separation | |
| CN112500442A (en) | Method for extracting raspberry glycoside by using functional polymer resin | |
| CN105130878B (en) | A kind of method that DNJ is extracted from mulberry leaf |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |