Polygonatum polysaccharide extraction process

Abstract

The invention relates to the technical field of extraction processes, and discloses a polygonatum sibiricum polysaccharide extraction process, which comprises the following steps: S1, raw material pretreatment: washing a polygonatum sibiricum raw material, drying, crushing, and sieving to obtain polygonatum sibiricum powder; s2, enzymolysis extraction: mixing the rhizoma polygonati powder with distilled water, adjusting the pH value, adding a compound enzyme preparation, and carrying out enzymolysis for 1-3 hours; s3, ultrasonic-assisted extraction: putting the enzymatic hydrolysate into ultrasonic equipment, and extracting for 20-50 minutes; s4, centrifugal separation: centrifuging the extracting solution for 10-20 minutes, and taking supernate; s5, alcohol precipitation and purification: adding 95% ethanol with the volume 3-5 times that of the supernate into the supernate, standing for 8-12 hours, and collecting precipitates; s6, drying: performing freeze drying or vacuum drying on the precipitate to obtain a polygonatum polysaccharide finished product. Compared with the prior art, the method has the following advantages: 1, the polysaccharide activity protection is obviously enhanced; 2, the extraction efficiency and the yield are greatly improved; 3, product purity and safety are optimized; 4, the green, energy-saving and economic benefits are outstanding; and 5, the molecular weight uniformity ensures the functional consistency.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of extraction process, and in particular to a polysaccharide extraction process of rhizoma polygonati BACKGROUND

[0002] As a traditional medicine and food homologous plant, the active ingredient of rhizoma polygonati, polysaccharide, has been proven to have significant antioxidant, immunomodulatory, hypoglycemic and anti-aging biological activities. With the development of functional food and modernization of traditional Chinese medicine industry, the market demand for high-purity and high-activity polysaccharide of rhizoma polygonati continues to grow, so it is crucial to develop an efficient and environmentally friendly extraction process.

[0003] The current mainstream method of industrial extraction of polysaccharide of rhizoma polygonati still has obvious defects: 1. Traditional hot water extraction method (such as CN108623660A): high temperature (80-100℃) and long time (3-6 hours) extraction, leading to polysaccharide structure degradation, active ingredient loss rate up to 30% or more, and high energy consumption, usually less than 8% yield; 2. Organic solvent assisted method (such as CN112300358A): using chloroform, acetone and other organic reagents for degreasing and impurity removal, although the purity can be improved, but the solvent residue is difficult to completely remove, which exists food safety risk and does not meet the requirements of green chemistry; 3. Single enzyme hydrolysis or ultrasonic technology (such as CN113527457A): although it can improve the extraction efficiency, but the cellulose wall is not completely broken, the polysaccharide release rate is limited (usually ≤10%), and the problem of wide polysaccharide molecular weight distribution is not solved.

[0004] In addition, the existing process lacks systematic optimization of key parameters (such as enzyme hydrolysis pH, ultrasonic power, alcohol precipitation ratio), resulting in poor batch stability and restricting industrial application.

[0005] In summary, it is urgent to develop a green extraction process that can synergistically break down the cell wall, protect the activity of polysaccharide, and improve the yield and purity, to meet the large-scale production needs of high-quality polysaccharide of rhizoma polygonati. SUMMARY

[0006] The present application provides a polysaccharide extraction process of rhizoma polygonati to solve the problems in the background art.

[0007] The present application provides a polysaccharide extraction process of rhizoma polygonati, comprising the following steps: S1: raw material pretreatment: washing, drying and crushing the rhizoma polygonati raw material, passing through a 40-100 mesh sieve to obtain rhizoma polygonati powder; S2: enzyme hydrolysis extraction: mixing the rhizoma polygonati powder with distilled water at a mass ratio of 1:15-1:30, adjusting the pH to 4.5-6.0, adding a composite enzyme preparation, and hydrolyzing at 45-60℃ for 1-3 hours; S3: ultrasonic-assisted extraction: the enzymatic hydrolysis solution is placed in an ultrasonic device, and extraction is carried out at a power of 200-500 W and a temperature of 50-70 DEG C for 20-50 min; S4: centrifugal separation: the extraction solution is centrifuged at 8000-12000 rpm for 10-20 min, and the supernatant is taken; S5: alcohol precipitation purification: 3-5 times the volume of 95% ethanol is added to the supernatant, and the mixture is placed at 4-10 DEG C for 8-12 h, and the precipitate is collected; S6: drying: the precipitate is freeze-dried or vacuum-dried to obtain the finished product of polygonatum sibiricum polysaccharide.

[0008] Preferably, the drying in S1 is hot air drying at 50-60 DEG C to a water content of ≤8%.

[0009] Preferably, the compound enzyme preparation in S2 is composed of cellulase, pectinase and neutral protease at a mass ratio of 2:2:1, and the addition amount is 0.5%-2.0% of the mass of polygonatum sibiricum powder.

[0010] Preferably, the ultrasonic extraction in S3 adopts an intermittent mode, and the working cycle is 5 s on / 5 s off.

[0011] Preferably, the supernatant is filtered through a 0.45 μm microporous filter before alcohol precipitation in S5.

[0012] Preferably, the freeze-drying conditions in S6 are: pre-freezing temperature -40 DEG C, cold trap temperature -50 DEG C, vacuum degree 10-30 Pa, and drying time 24-48 h.

[0013] Preferably, it further comprises: S7: the dried polygonatum sibiricum polysaccharide is separated and purified by ultrafiltration membrane, and the polysaccharide component with a molecular weight of 3000-10000 Da is intercepted.

[0014] Preferably, the ultrafiltration membrane is a polyether sulfone (PES) membrane, and the operating pressure is 0.2-0.5 MPa and the temperature is 25-40 DEG C.

[0015] The enzymatic hydrolysis-ultrasonic-assisted extraction process for polygonatum sibiricum polysaccharide provided by the application has the following outstanding advantages compared with the prior art: 1. The activity protection of polysaccharide is significantly enhanced: the mild wall breaking (≤60 DEG C) of the compound enzyme (cellulase+pectinase+protease) is synergized with the ultrasonic cavitation effect, and the damage of high temperature to the structure of polysaccharide is avoided. Experiments show that the DPPH free radical scavenging rate of the obtained polysaccharide is as high as 91.2%, and the antioxidant activity is increased by 16.3%.

[0016] 2. Extraction efficiency and yield are greatly improved: composite enzyme degrades cell wall polysaccharide-protein complex, combined with ultrasonic to accelerate mass transfer, so that the polysaccharide dissolution rate is increased to 12-15% (traditional process ≤8%). Pilot test verification: 100 kg of Huangjing raw material is put into the machine, and 12.8 kg of polysaccharide product is obtained, and the extraction time is shortened to less than 2 hours (traditional process needs about 5 hours).

[0017] 3. Product purity and safety are optimized: three-stage purification of microfiltration (0.45 μm membrane) → alcohol precipitation → ultrafiltration (3000-10000 Da cutoff) is adopted, the polysaccharide purity is stably at 85-92%, HPLC detection has no organic solvent residue (in line with the standard of Chinese Pharmacopoeia 2020 edition), and the heavy metal content is less than 5 ppm.

[0018] 4. Green, energy-saving and economic benefits are outstanding: the whole process uses water / ethanol as medium, avoids toxic reagents such as chloroform, and reduces waste discharge by about 40%; ultrasonic assistance reduces heat energy consumption, and energy consumption per unit output is reduced by about 50%.

[0019] 5. Molecular weight uniformity guarantees functional consistency: ultrafiltration precisely controls the molecular weight distribution of polysaccharide (concentrated in 5000-8000 Da), and the activity difference of polysaccharide between batches is ≤5%, which meets the requirements of drug-grade raw materials. DETAILED DESCRIPTION

[0020] The application will be further described below in combination with specific examples.

[0021] Example 1 The application provides a Huangjing polysaccharide extraction process, which comprises the following steps: S1: raw material pretreatment: the Huangjing raw material is cleaned, dried and crushed, and then sieved through a 60-mesh sieve to obtain Huangjing powder; wherein the drying is hot air drying at 53°C until the water content is ≤8%.

[0022] S2: enzyme extraction: the Huangjing powder is mixed with distilled water at a mass ratio of 1:20, the pH is adjusted to 4.5, and a composite enzyme preparation is added, and then the mixture is subjected to enzyme hydrolysis at 52°C for 1.5 hours; wherein the composite enzyme preparation is composed of cellulase, pectinase and neutral protease at a mass ratio of 2:2:1, and the addition amount is 1.3% of the mass of the Huangjing powder.

[0023] S3: ultrasonic-assisted extraction: the enzyme hydrolysis liquid is placed in an ultrasonic device, and extracted at a power of 400W and a temperature of 57°C for 45 minutes; the ultrasonic extraction adopts an intermittent mode, and the working cycle is 5s on / 5s off.

[0024] S4: centrifugal separation: the extraction liquid is centrifuged at 8000-12000 rpm for 18 minutes, and the supernatant is taken; S5: alcohol precipitation purification: 3.5 times the volume of 95% ethanol is added to the supernatant, and the precipitate is collected after standing at 4-10℃ for 9 hours; the supernatant before alcohol precipitation is filtered through a 0.45μm microporous filter membrane.

[0025] S6: drying: the precipitate is freeze-dried or vacuum-dried to obtain the finished product of polygonatum sibiricum polysaccharide; the freeze-drying conditions are: pre-freezing temperature-40℃, cold trap temperature-50℃, vacuum degree 20Pa, and drying time 40 hours.

[0026] S7: the dried polygonatum sibiricum polysaccharide is separated and purified by ultrafiltration membrane, and the polysaccharide component with a molecular weight of 3000-10000Da is intercepted; the ultrafiltration membrane is a polyether sulfone (PES) membrane, the operating pressure is 0.5MPa, and the temperature is 30℃.

[0027] The final product of polygonatum sibiricum polysaccharide has a purity of ≥85% and a polysaccharide yield of ≥12%.

[0028] Example 2 The present application provides a polygonatum sibiricum polysaccharide extraction process, comprising the following steps: S1: raw material pretreatment: the polygonatum sibiricum raw material is cleaned, dried and crushed, and then sieved through an 80-mesh sieve to obtain polygonatum sibiricum powder; wherein the drying is hot air drying at 58℃ until the water content is ≤8%.

[0029] S2: enzyme extraction: the polygonatum sibiricum powder is mixed with distilled water at a mass ratio of 1:18, the pH is adjusted to 5, a composite enzyme preparation is added, and enzyme hydrolysis is carried out at 60℃ for 2.5 hours; wherein the composite enzyme preparation is composed of cellulase, pectinase and neutral protease at a mass ratio of 2:2:1, and the addition amount is 1.3% of the mass of the polygonatum sibiricum powder.

[0030] S3: ultrasonic-assisted extraction: the enzyme hydrolysis liquid is placed in an ultrasonic device, and extracted at a power of 300W and a temperature of 60℃ for 40 minutes; the ultrasonic extraction adopts an intermittent mode, and the working cycle is 5s on / 5s off.

[0031] S4: centrifugal separation: the extract is centrifuged at 8000-12000rpm for 15 minutes, and the supernatant is taken; S5: alcohol precipitation purification: 3.5 times the volume of 95% ethanol is added to the supernatant, and the precipitate is collected after standing at 4-10℃ for 9 hours; the supernatant before alcohol precipitation is filtered through a 0.45μm microporous filter membrane.

[0032] S6: drying: the precipitate is freeze-dried or vacuum-dried to obtain the finished product of polygonatum sibiricum polysaccharide; the freeze-drying conditions are: pre-freezing temperature-40℃, cold trap temperature-50℃, vacuum degree 20Pa, and drying time 40 hours.

[0033] S7: The dried polygonatum sibiricum polysaccharide is separated and purified by an ultrafiltration membrane, and polysaccharide components with a molecular weight of 3000-10000 Da are intercepted; the ultrafiltration membrane is a polyether sulfone (PES) membrane, the operating pressure is 0.4 MPa, and the temperature is 33 DEG C.

[0034] The purity of the final product, polygonatum sibiricum polysaccharide, is greater than or equal to 83%, and the polysaccharide yield is greater than or equal to 13%.

[0035] The above merely describes a preferred specific embodiment of the present application, but the protection scope of the present application is not limited to this. Any person skilled in the art, according to the technical solution and the inventive concept of the present application, can make equivalent replacements or changes within the technical range disclosed by the present application, which should be covered within the protection scope of the present application.

Claims

1. A process for extracting polysaccharides from Polygonatum sibiricum, characterized in that, Comprising the following steps: S1: raw material pretreatment: the raw material of polygonatum sibiricum is washed, dried, crushed, and sieved through a 40-100 mesh sieve to obtain polygonatum sibiricum powder; S2: enzymatic extraction: the polygonatum sibiricum powder is mixed with distilled water at a mass ratio of 1:15-1:30, the pH is adjusted to 4.5-6.0, a composite enzyme preparation is added, and enzymatic hydrolysis is carried out at 45-60°C for 1-3 hours; S3: ultrasonic-assisted extraction: the enzyme hydrolysate is placed in an ultrasonic device and extracted at a power of 200-500W and a temperature of 50-70°C for 20-50 minutes; S4: centrifugal separation: the extract is centrifuged at 8000-12000rpm for 10-20 minutes, and the supernatant is collected; S5: alcohol precipitation purification: 3-5 times the volume of 95% ethanol is added to the supernatant, which is placed at 4-10°C for 8-12 hours, and the precipitate is collected; S6: drying: the precipitate is freeze-dried or vacuum-dried to obtain polygonatum sibiricum polysaccharide finished product.

2. The extraction process of polygonatum sibiricum polysaccharides according to claim 1, characterized in that, The drying in S1 is hot air drying at 50-60°C to a moisture content of ≤8%.

3. The extraction process of the polygonatum sibiricum polysaccharide according to claim 1, characterized in that, The composite enzyme preparation in S2 is composed of cellulase, pectinase, and neutral protease at a mass ratio of 2:2:1, and the addition amount is 0.5%-2.0% of the mass of the polygonatum sibiricum powder.

4. The extraction process of the polygonatum sibiricum polysaccharide according to claim 1, characterized in that, The ultrasonic extraction in S3 uses an intermittent mode with a working cycle of 5s on / 5s off.

5. The extraction process of the polygonatum sibiricum polysaccharide according to claim 1, characterized in that, The pre-alcohol precipitation supernatant in S5 is filtered through a 0.45μm microporous filter membrane.

6. The extraction process of the polygonatum sibiricum polysaccharide according to claim 1, characterized in that, The freeze-drying conditions in S6 are: pre-freezing temperature -40°C, cold trap temperature -50°C, vacuum degree 10-30Pa, and drying time 24-48 hours.

7. The extraction process of the polygonatum sibiricum polysaccharide according to claim 1, characterized in that, Further comprising: S7: the dried polygonatum sibiricum polysaccharide is separated and purified by ultrafiltration membrane, and the polysaccharide component with a molecular weight of 3000-10000Da is intercepted.

8. The extraction process of the polygonatum sibiricum polysaccharide according to claim 7, characterized in that, The ultrafiltration membrane is a polyether sulfone (PES) membrane, the operating pressure is 0.2-0.5 MPa, and the temperature is 25-40°C.