CN109517015B - Method for enriching crocin from crocus sativus - Google Patents
Method for enriching crocin from crocus sativus Download PDFInfo
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- C07H13/00—Compounds containing saccharide radicals esterified by carbonic acid or derivatives thereof, or by organic acids, e.g. phosphonic acids
- C07H13/02—Compounds containing saccharide radicals esterified by carbonic acid or derivatives thereof, or by organic acids, e.g. phosphonic acids by carboxylic acids
- C07H13/04—Compounds containing saccharide radicals esterified by carbonic acid or derivatives thereof, or by organic acids, e.g. phosphonic acids by carboxylic acids having the esterifying carboxyl radicals attached to acyclic carbon atoms
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- C—CHEMISTRY; METALLURGY
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- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
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Abstract
The invention relates to a method for enriching crocin from crocus sativus. The method comprises the following steps: (1) placing stigma croci Sativi in a percolation tank, adding appropriate amount of metal salt solution, completely immersing the medicinal materials, soaking for 1h, adding metal salt solution 8-60 times of the medicinal materials, percolating and extracting, and collecting percolate. (2) Passing the percolate through macroporous adsorbent resin column at flow rate of 0.2-2BV/h, sequentially washing the resin column with purified water, 25% ethanol and 70% ethanol, discarding water and 25% ethanol eluate, and collecting 70% ethanol eluate. (3) Concentrating 70% ethanol eluate under reduced pressure, and freeze drying to obtain crocin refined product. The method can greatly reduce the degradation rate of the crocin in the solution, can be applied to the extraction, concentration, purification and drying processes of the crocin, and improves the transfer rate of the crocin in the technological process by improving the stability of the crocin.
Description
Technical Field
The invention belongs to the technical field of medicines, and particularly relates to a method for enriching crocin from crocus sativus.
Background
Crocin (crocin) is a water-soluble natural pigment, mainly crocin I and crocin II, and is a traditional Chinese medicinal material gardenia (fructus Gardeniae)Gardenia jasminoidesEllis) and saffron (Crocus sativusL.) one of the main active ingredients. Pharmacological studies have shown that: crocin has anti-tumor effect, and can remarkably inhibit cancer cell proliferation and promote cancer cell apoptosis; can improve cognitive ability, enhance neuron activity, resist oxidative stress, and inhibit amyloid oligomer aggregation and entanglement, thereby having the effects of treating Parkinson disease, Alzheimer disease, vascular senile dementia and other neurodegenerative diseases; can be used for treating cardiovascular diseases from aspects of resisting platelet aggregation and apoptosis, resisting oxidative stress and resisting cardiovascular cell apoptosis. Therefore, the crocin has wide drug effect and development potential.
Crocin belongs to diterpenoid components and is composed of crocetin and sugar chains, and has poor stability due to a plurality of conjugated double bonds in a molecular structure, and can be rapidly decomposed in a solution under the action of heat and light. The unstable structure of crocin results in low transfer rate in the industrial extraction, concentration, purification and drying processes, which greatly limits the development of enrichment process. The prior art has few means for improving the stability of the crocin and can only avoid heating and illumination as much as possible in the process.
During the research of the stability of the crocin, we unexpectedly found that a part of metal ions such as Na+、K+、Ca2+、Al3 +The inhibition effect on the degradation of the crocin is strong, and the inhibition effect is probably related to the improvement of the electrolyte strength of the crocin solution. Therefore, a set of extraction method is designed for percolating saffron medicinal material by using metal salt solution as medium, then carrying out macroporous resin column chromatography purification on the percolate containing metal ions, concentrating and drying to obtain crocin extractThe extraction method greatly overcomes the defects of large degradation and low transfer rate of crocin in the traditional process.
Aiming at the problems of easy degradation and poor stability of the existing crocin in process engineering, the invention provides the crocin enrichment method which is simple, convenient and feasible, low in cost, high in transfer rate, environment-friendly and wide in application range.
Disclosure of Invention
The invention aims to overcome the problems of easy degradation and poor stability of the crocin in the process, and provides the crocin enrichment method which is simple, convenient and feasible, low in cost, high in transfer rate, environment-friendly and wide in application range.
In order to achieve the purpose, the technical scheme adopted by the invention is as follows:
the invention provides a method for enriching crocin from crocus sativus, which comprises the steps of adding a metal salt solution into crocus sativus medicinal materials, soaking, and then percolating and extracting.
The other scheme of the invention is that after a metal salt solution is added into the saffron medicinal material for soaking, the metal salt solution with the weight 8-60 times of the medicinal material is added for percolation extraction, and the percolation liquid is collected.
Further, the percolate can be passed through macroporous adsorbent resin column at flow rate of 0.2-2BV/h, sequentially washing the resin column with purified water, 25% ethanol and 70% ethanol, discarding water and 25% ethanol eluate, and collecting 70% ethanol eluate.
Further, the 70% ethanol eluate can be concentrated under reduced pressure, and lyophilized to obtain crocin refined product.
The metal salts include, but are not limited to: one or more of sodium salt, potassium salt, calcium salt or aluminum salt.
The concentration of the metal salt solution is 0-0.1 mol/L.
Preferably, the concentration of the metal salt solution is 0.0001-0.1 mol/L.
To further clarify the process for the enrichment of crocin from saffron according to the present invention, the following protocol is provided:
(1) placing stigma croci Sativi in a percolation tank, adding appropriate amount of metal salt solution, completely immersing the medicinal materials, soaking for 1h, adding metal salt solution 8-60 times of the medicinal materials, percolating and extracting, and collecting percolate.
(2) Passing the percolate through macroporous adsorbent resin column at flow rate of 0.2-2BV/h, sequentially washing the resin column with purified water, 25% ethanol and 70% ethanol, discarding water and 25% ethanol eluate, and collecting 70% ethanol eluate.
(3) Concentrating 70% ethanol eluate under reduced pressure, and freeze drying to obtain crocin refined product.
In the step (1), those skilled in the art can adjust the soaking time according to experience and specific operation conditions, or can directly perform percolation extraction without soaking.
The elution solvent used in the above step (2) and its concentration can be adjusted by those skilled in the art according to the prior art and experience.
The invention has the beneficial effects that:
1. the invention can greatly reduce the degradation rate of the crocin in the solution.
2. The method can be applied to the extraction, concentration, purification and drying processes of the crocin, and improves the transfer rate of the crocin in the technological process by improving the stability of the crocin.
Drawings
FIG. 1 is a liquid chromatogram of a reference sample of crocin I;
FIG. 2 is a liquid chromatogram of a crocin II control;
FIG. 3 is a liquid chromatogram of a refined crocin product;
FIG. 4 effect of different metal salt solutions on the stability of crocin I;
FIG. 5 effect of different metal salt solutions on the stability of crocin II;
FIG. 6 effect of NaCl solutions of different concentrations on the stability of crocin I;
FIG. 7 the effect of NaCl solutions of different concentrations on the stability of crocin II.
Detailed description of the preferred embodiments
The following is a further illustration of the invention with reference to specific examples. These examples are only illustrative and not intended to limit the scope of the present invention. The experimental methods of the following examples, in which specific experimental conditions are not specified, are generally performed according to conventional conditions.
Example 1:
weighing 500g of saffron medicinal material, adding purified water to completely immerse the medicinal material, soaking for 1h, adding 20L of purified water for percolation extraction, collecting percolate, passing the percolate through a macroporous adsorption resin column at the flow rate of 0.2BV/h, washing the resin column with purified water, 25% ethanol and 70% ethanol in sequence, discarding water and 25% ethanol eluent, collecting 70% ethanol eluent, carrying out reduced pressure concentration on the 70% ethanol eluent, and freeze-drying to obtain 91g of crocin refined product, wherein the content of crocin I is measured to be 26.2%, and the content of crocin II is measured to be 9.3%.
Example 2:
weighing 500g of saffron medicinal material, adding 0.001mol/L NaCl solution to completely immerse the medicinal material, soaking for 1h, adding 30L of 0.001mol/L NaCl solution for percolation extraction, collecting percolate, passing the percolate through a macroporous adsorption resin column at the flow rate of 0.5BV/h, sequentially washing the resin column with purified water, 25% ethanol and 70% ethanol, discarding water and 25% ethanol eluent, collecting 70% ethanol eluent, carrying out reduced pressure concentration on the 70% ethanol eluent, and freeze-drying to obtain 101g of a crocin refined product, wherein the content of the crocin I is measured to be 36.8%, and the content of the crocin II is measured to be 13.5%.
Example 3: weighing 500g of saffron medicinal material, adding 0.01 mol/L KCl solution to completely immerse the medicinal material, soaking for 1h, adding 4L of 0.01 mol/L KCl solution for percolation extraction, collecting percolate, passing the percolate through a macroporous adsorption resin column at the flow rate of 1BV/h, sequentially washing the resin column with purified water, 25% ethanol and 70% ethanol, discarding water and 25% ethanol eluent, collecting 70% ethanol eluent, carrying out reduced pressure concentration on the 70% ethanol eluent, and freeze-drying to obtain 69g of crocin refined product, wherein the content of the crocin I is 35.7%, and the content of the crocin II is 12.1%.
Example 4:
weighing 500g of saffron medicinal material, and adding 0.1mol/L CaCl2Soaking the medicinal materials in the solution for 1 hr, and adding 0.1mol/L CaCl2Solution 10L intoPercolating, collecting percolate, passing the percolate through a macroporous adsorbent resin column at the flow rate of 1.5 BV/h, sequentially flushing the resin column with purified water, 25% ethanol and 70% ethanol, discarding water and 25% ethanol eluent, collecting 70% ethanol eluent, performing reduced pressure concentration on the 70% ethanol eluent, and freeze-drying to obtain 77g of crocin refined product, wherein the content of crocin I is 32.5%, and the content of crocin II is 11.3%.
Example 5:
weighing 500g of saffron medicinal material, and adding 0.005 mol/L AlCl3Soaking the medicinal materials in the solution for 1 hr, and adding 0.005 mol/L AlCl3Percolating 25L of the solution, collecting percolate, passing the percolate through a macroporous adsorption resin column at the flow rate of 2BV/h, sequentially flushing the resin column with purified water, 25% ethanol and 70% ethanol, discarding water and 25% ethanol eluent, collecting 70% ethanol eluent, carrying out reduced pressure concentration on the 70% ethanol eluent, and freeze-drying to obtain 97g of a crocin refined product, wherein the content of the crocin I is 28.9% and the content of the crocin II is 9.8%.
Experimental example 1: method for measuring content of crocin I and crocin II in crocin refined product
(1) Chromatographic conditions and System suitability test
Octadecylsilane chemically bonded silica is used as a filling agent; methanol-water (45: 55) is used as a mobile phase; the detection wavelength was 440 nm.
(2) Preparation of control solutions
Taking appropriate amount of reference substances of crocin I and crocin II respectively, precisely weighing, and adding 50% ethanol to obtain solution containing 39 μ g of crocin I per 1m L and solution containing 12 μ g of crocin II per 1m L.
(3) Preparation of test solution
Taking 20mg of crocin refined product, precisely weighing, placing in a 25mL volumetric flask, adding mobile phase, placing in an ice bath for ultrasonic dissolution, fixing the volume, precisely sucking 1mL, placing in a 10mL volumetric flask, and adding the mobile phase for fixing the volume to obtain the crocin.
(4) Assay method
Precisely sucking 10 μ L each of the reference solution of crocin I, the reference solution of crocin II and the test solution, injecting into a liquid chromatograph, and measuring.
Experimental example 2: effect of different metal salt solutions on the stability of crocin.
9 parts of the purified crocin product obtained in example 1 were precisely weighed, and 150mg of each of the purified crocin products were placed in a 100mL volumetric flask, and purified water, 0.001mol/L NaCl and FeCl were added to the flask3、FeCl2、CuSO4，CaCl2、MgSO4、AlCl3And fully dissolving the KCl solution, fixing the volume to a scale, standing at room temperature in a dark place, taking a small amount of solution every 3 hours, filtering, injecting into a liquid chromatogram, measuring the concentration of the crocin I and the concentration of the crocin II in the solution, and calculating the degradation rate (the result is shown in the attached figures 4 and 5 of the specification).
Experimental example 3: effect of NaCl solutions of different concentrations on the stability of crocin.
Precisely weighing 5 parts of the crocin refined product obtained in example 1, adding 150mg of each part into a 100mL volumetric flask, respectively adding purified water, 0.1mol/L, 0.01 mol/L, 0.001mol/L and 0.0001 mol/L NaCl solution to fully dissolve, fixing the volume to the scale, standing at room temperature in a dark place, taking a small amount of solution every 3h, filtering, injecting into a liquid chromatogram, measuring the concentration of crocin I and crocin II in the solution, and calculating the degradation rate (see the results shown in the attached figures 6 and 7 of the specification).
Claims (4)
1. A method for enriching crocin from crocus sativus is characterized in that a metal salt solution is added into crocus sativus medicinal materials for soaking and then percolation extraction is carried out; wherein the content of the first and second substances,
the metal salt is NaCl, KCl, CaCl2Or AlCl3One or more of;
the concentration of the metal salt solution is 0.0001-0.1 mol/L.
2. The method as claimed in claim 1, wherein the crocus sativus is soaked in a metal salt solution, and then a metal salt solution 8-60 times the weight of the crocus sativus is added for percolation extraction, and a percolate is collected.
3. The method according to claim 2, further comprising the steps of passing the percolate through a macroporous adsorbent resin column at a flow rate of 0.2-2BV/h, washing the resin column with purified water, 25% ethanol, 70% ethanol in sequence, discarding the water and 25% ethanol eluate, and collecting the 70% ethanol eluate.
4. The method according to claim 3, further comprising the step of concentrating the 70% ethanol eluate under reduced pressure, and freeze-drying to obtain a crocin-refined product.
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Families Citing this family (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110279640A (en) * | 2019-07-17 | 2019-09-27 | 杭州宗科愿生物科技开发有限公司 | Saffron extract and its application as melanin inhibitor |
| CN111171087B (en) * | 2020-02-17 | 2023-09-08 | 天津理工大学 | Purification method and application of high-purity crocin I |
| CN113134047A (en) * | 2021-04-23 | 2021-07-20 | 瑞阳制药股份有限公司 | Preparation method and application of saffron total glycoside extract |
| CN113144057B (en) * | 2021-06-03 | 2022-09-16 | 上海瀛洲西红花种植专业合作社 | Stigma croci extract and preparation method and application thereof |
| CN114984119A (en) * | 2022-07-28 | 2022-09-02 | 浙江长典药物技术开发有限公司 | Saffron fluid extract and preparation process thereof |
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