Sunflower pollen polysaccharide C and its preparation method and application

Abstract

The present invention relates to the technical field of polysaccharide extraction from pollen, and in particular to a sunflower pollen polysaccharide C and a preparation method and application thereof, wherein the number average molecular weight of the sunflower pollen polysaccharide C is 1.302 kDa, the peak molecular weight is 0.652 kDa, and the weight average molecular weight is 3.265 kDa. The present invention provides a sunflower pollen polysaccharide C, which not only has good antioxidant effects, but also has anti-inflammatory, anti-hypertensive and anti-aging effects, and can be applied to medicines, health products or daily chemical products. The preparation method provided by the present invention is simple and efficient, and can effectively extract sunflower pollen polysaccharide C with anti-inflammatory, anti-hypertensive, anti-aging and antioxidant effects from sunflower pollen, is easy to promote, and is conducive to the comprehensive research of sunflower pollen polysaccharides.

Description

Technical Field

[0001] The invention relates to the technical field of polysaccharide extraction from pollen, in particular to sunflower pollen polysaccharide C and a preparation method and application thereof. Background Art

[0002] Sunflowers have large-scale planting bases in Inner Mongolia and Xinjiang Uygur Autonomous Regions of my country. The utilization value of sunflowers is high. Not only are the seeds edible, but other parts also have edible and medicinal value. The "Compendium of Materia Medica" records that sunflower pollen has the effects of clearing away heat and dampness, relieving qi and dispersing knots. Since sunflower pollen was announced as a common food raw material in 2004, it has received widespread attention.

[0003] Sunflower pollen is an edible pollen of bee pollen. It is rich in nutrients and a variety of active substances and can be used as a natural raw material in the fields of bee products, health foods, cosmetics, and medicine. Sunflower pollen has many active substances, such as flavonoids, polyphenols, and polysaccharides. Previous studies have found that the sugar content of sunflower pollen is greater than 40%. At present, the research on sunflower pollen mainly focuses on the nutritional components of pollen and its antioxidant and uric acid-lowering abilities. The extraction and application of the key functional components of sunflower pollen polysaccharides are not fully studied.

[0004] Therefore, in order to efficiently utilize sunflower pollen and improve the efficacy information of its key ingredients, it is necessary to research and develop the extraction and comprehensive utilization of key factors of polysaccharides in sunflower pollen. Summary of the invention

[0005] The purpose of the present invention is to provide a sunflower pollen polysaccharide C and its preparation method and application in view of the above problems. The sunflower pollen polysaccharide C not only has good antioxidant effect, but also has anti-inflammatory, antihypertensive and anti-aging effects; the preparation method is efficient and stable.

[0006] The present invention provides a sunflower pollen polysaccharide C, wherein the number average molecular weight of the sunflower pollen polysaccharide C is 1.302 kDa, the peak molecular weight is 0.652 kDa, and the weight average molecular weight is 3.265 kDa.

[0007] Preferably, the sunflower pollen polysaccharide C consists of arabinose, galactose and glucose; the molar ratio of the arabinose, the galactose and the glucose is 4:1:15.

[0008] The present invention also provides a method for preparing sunflower pollen polysaccharide C, and the specific steps are as follows:

[0009] S1. extracting the sunflower pollen with acidic water homogenization, and then concentrating, freeze-drying and precipitating the extract with alcohol to obtain crude sunflower pollen polysaccharides;

[0010] The specific method of alcohol precipitation is as follows: weigh the extract and dissolve it in water, the solid-liquid ratio between the extract and water is 1:5 (g / mL), after the extract is completely dissolved, add anhydrous ethanol until an 80% ethanol mixture is obtained, ultrasonicate for 5 minutes, let it stand for 12 hours, and collect the precipitate by centrifugation.

[0011] S2. preparing an aqueous solution of the crude sunflower pollen polysaccharide obtained in step S1, purifying the aqueous solution of the crude sunflower pollen polysaccharide using a bonded phase silica gel separation column, collecting the eluate, and obtaining a preliminarily purified sunflower pollen polysaccharide;

[0012] S3. The preliminarily purified sunflower pollen polysaccharide obtained in step S2 is further purified using a gel chromatography column, the eluate with the largest elution peak is collected, and dried with nitrogen to obtain a yellow solid, which is purified sunflower pollen polysaccharide C.

[0013] Preferably, the specific steps of acid water homogenization extraction in step S1 are: adding sunflower pollen to an acidic solvent for extraction, homogenizing for 5-20 minutes, and separating to obtain an extract. The acidic solvent can be any one of citric acid, malic acid, acetic acid, lactic acid or succinic acid, all with a mass concentration of 0.02%.

[0014] Preferably, the solid-liquid ratio between sunflower pollen and acidic solvent is 1:(30-50) (g / mL).

[0015] Preferably, the specific steps of step S2 are as follows: in the bonded phase silica gel separation column purification, the aqueous solution of the crude sunflower pollen polysaccharide obtained in step S1 is gradient eluted with 1%-5% acetonitrile, and then the eluate of the elution peak is collected after elution with 2% acetonitrile to obtain the preliminary purified sunflower pollen polysaccharide. Preferably, the crude sunflower pollen polysaccharide and the filler are loaded at a mass ratio of 1:50 (g:g), and during the gradient elution, 1% acetonitrile is used for elution for 10 minutes, 2% acetonitrile is used for elution for 10 minutes, 3% acetonitrile is used for elution for 10 minutes, 4% acetonitrile is used for elution for 10 minutes, and 5% acetonitrile is used for elution for 10 minutes, and the elution flow rate is 1mL / min; the filler is a commonly used filler in this field. The bonded phase silica gel separation column adopts a spherical C18 column with a particle size of 20-45μm and a pore size of

[0016] Preferably, pure water is used as the mobile phase for purification in the gel chromatography column purification in step S3. The gel in the gel layer of the gel chromatography column uses dextran coagulation G-25. Preferably, the preliminary purified sunflower pollen polysaccharide and the filler are loaded at a mass ratio of 1:60 (g:g), eluted with pure water, with a flow rate of 0.5 mL / min, and the maximum absorption peak is collected; the filler is a commonly used filler in the field.

[0017] The present invention provides application of sunflower pollen polysaccharide C in preparing medicines, health products or daily chemical products with anti-inflammatory effects.

[0018] The present invention provides application of sunflower pollen polysaccharide C in preparing medicines, health products or daily chemicals with blood pressure lowering effects.

[0019] The present invention provides application of sunflower pollen polysaccharide C in preparing medicines, health products or daily chemical products with anti-aging effects.

[0020] The present invention provides application of sunflower pollen polysaccharide C in preparing medicines, health products or daily chemicals with antioxidant effects.

[0021] Beneficial effects:

[0022] The invention provides a sunflower pollen polysaccharide C. The sunflower pollen polysaccharide C not only has good antioxidant effect, but also has anti-inflammatory, blood pressure-lowering and anti-aging effects, and can be applied to medicines, health products or daily chemical products.

[0023] The preparation method provided by the present invention is simple and efficient, can effectively extract sunflower pollen polysaccharide C with anti-inflammatory, antihypertensive, anti-aging and antioxidant effects from sunflower pollen, is easy to promote, and is conducive to the comprehensive research of sunflower pollen polysaccharides. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 This is the elution pattern of the initially purified helianthus pollen polysaccharide C;

[0025] Figure 2 This is the elution profile of purified sunflower pollen polysaccharide C;

[0026] Figure 3 This is a graph showing the absolute molecular weight test results of sunflower pollen polysaccharide C;

[0027] Figure 4 The test spectra of various monosaccharide standards;

[0028] Figure 5 This is the chromatogram of monosaccharide components of sunflower pollen polysaccharide C;

[0029] Figure 6 This is a graph showing the test results of different concentrations of sunflower pollen polysaccharide C on the cell viability of RAW264.7 cells;

[0030] Figure 7 This is the test result of different concentrations of sunflower pollen polysaccharide C on the secretion of NO by cells;

[0031] Figure 8 This is a graph showing the results of the time series lifespan test on Saccharomyces cerevisiae with different concentrations of sunflower pollen polysaccharide C. DETAILED DESCRIPTION

[0032] It should be noted that the following detailed descriptions are illustrative and are intended to provide further explanation of the present application. Unless otherwise specified, all technical and scientific terms used herein have the same meanings as those commonly understood by those skilled in the art to which the present application belongs.

[0033] It should be noted that the terms used herein are only for describing specific embodiments and are not intended to limit the exemplary embodiments according to the present application. As used herein, unless the context clearly indicates otherwise, the singular form also includes the plural form. In addition, it should be understood that when the terms "comprise" and / or "include" are used in this description, it indicates the presence of features, steps, operations, devices, components and / or combinations thereof.

[0034] The technical solution of the present invention will be clearly and completely described below in conjunction with the embodiments. Obviously, the described embodiments are part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0035] Example 1

[0036] A method for preparing sunflower pollen polysaccharide C, the specific steps are as follows:

[0037] S1. Extract the sunflower pollen by acid water homogenization. Weigh 100g of sunflower pollen and add 4L of 0.02% citric acid aqueous solution for extraction. The solid-liquid ratio between the sunflower pollen and the citric acid aqueous solution is 1:40. Homogenize at room temperature for 10 minutes, pass through a 500-mesh sieve, separate and obtain an extract, concentrate the extract to 1.5L at 55°C, precool it in a -80°C refrigerator, and freeze-dry it in a vacuum freeze dryer to obtain an extract. The extract is precipitated with alcohol: weigh the extract and dissolve it in water. The solid-liquid ratio between the extract and water is 1:5 (g / mL). After the extract is completely dissolved, add anhydrous ethanol until an 80% ethanol mixture is obtained. Ultrasonicate for 5min and let it stand for 12h. Centrifuge and collect the precipitate to obtain 12g of crude sunflower pollen polysaccharide.

[0038] S2. The crude polysaccharide from sunflower pollen obtained in step S1 is prepared into an aqueous solution with a concentration of 5 mg / mL. The crude polysaccharide from sunflower pollen and the filler are loaded on a bonded phase silica gel separation column at a mass ratio of 1:50 (g:g). The aqueous solution of the crude polysaccharide from sunflower pollen is gradient eluted at a flow rate of 1 mL / min. The gradient elution is performed in sequence with 1% acetonitrile elution for 10 min, 2% acetonitrile elution for 10 min, 3% acetonitrile elution for 10 min, 4% acetonitrile elution for 10 min, and 5% acetonitrile elution for 10 min. The elution curve is as follows: Figure 1 As shown, Figure 1The horizontal axis in the middle represents the elution volume, the vertical axis represents the detection signal value, and the % value on the right is the acetonitrile percentage indicator in the eluent; then, the eluent of the elution peak obtained after elution with 2% acetonitrile was collected to obtain 8.4 g of preliminarily purified sunflower pollen polysaccharide;

[0039] The bonded phase silica gel separation column uses a spherical C18 column with a particle size of 20-45μm and a pore size of

[0040] S3. The preliminarily purified sunflower pollen polysaccharide obtained in step S2 is further purified by gel chromatography column, and the preliminarily purified sunflower pollen polysaccharide and filler are loaded at a mass ratio of 1:60 (g:g), pure water is used as the mobile phase, the flow rate is 0.5 mL / min, and the gel layer in the gel chromatography column is dextran coagulation G-25. The elution curve is as follows: Figure 2 As shown (the abscissa represents the elution volume, and the ordinate represents the detection signal value), the eluate of the largest elution peak was collected and dried with nitrogen to obtain a yellow solid, which was the purified sunflower pollen polysaccharide C 0.5 g.

[0041] Example 2

[0042] A method for preparing sunflower pollen polysaccharide C, the specific steps are as follows:

[0043] S1. The sunflower pollen is subjected to acid water homogenization extraction. 500g of sunflower pollen is weighed and 15L of 0.02% citric acid aqueous solution is added for extraction. The solid-liquid ratio between the sunflower pollen and the citric acid aqueous solution is 1:30. The mixture is homogenized at room temperature for 20 minutes, passed through a 500-mesh sieve, and an extract is separated. The extract is concentrated to 5L at 55°C, pre-cooled in a -80°C refrigerator, and then freeze-dried in a vacuum freeze dryer to obtain an extract. The extract is subjected to alcohol precipitation: the extract is weighed and dissolved in water. The solid-liquid ratio between the extract and water is 1:5 (g / mL). After the extract is completely dissolved, anhydrous ethanol is added until an 80% ethanol mixture is obtained. After ultrasonication for 5 minutes, the mixture is allowed to stand for 12 hours. The precipitate is collected by centrifugation to obtain 58.9g of crude sunflower pollen polysaccharides.

[0044] S2. The crude sunflower pollen polysaccharide obtained in step S1 is prepared into an aqueous solution with a concentration of 5 mg / mL. The crude sunflower pollen polysaccharide and filler are loaded on a bonded phase silica gel separation column at a mass ratio of 1:50 (g:g). The aqueous solution of the crude sunflower pollen polysaccharide is gradient eluted at a flow rate of 1 mL / min. During the gradient elution, 1% acetonitrile is used for elution for 10 min, 2% acetonitrile is used for elution for 10 min, 3% acetonitrile is used for elution for 10 min, 4% acetonitrile is used for elution for 10 min, and 5% acetonitrile is used for elution for 10 min. Then, the eluate of the elution peak obtained after elution with 2% acetonitrile is collected to obtain 42.1 g of preliminarily purified sunflower pollen polysaccharide;

[0045] The bonded phase silica gel separation column uses a spherical C18 column with a particle size of 20-45μm and a pore size of

[0046] S3. The preliminarily purified sunflower pollen polysaccharide obtained in step S2 was further purified by gel chromatography column, and the preliminarily purified sunflower pollen polysaccharide and filler were loaded at a mass ratio of 1:60 (g:g), pure water was used as the mobile phase, the flow rate was 0.5 mL / min, and the gel in the gel layer of the gel chromatography column was dextran G-25. The eluate of the maximum elution peak was collected, and after drying with nitrogen, a yellow solid was obtained, which was 2.56 g of purified sunflower pollen polysaccharide C.

[0047] Example 3

[0048] A method for preparing sunflower pollen polysaccharide C, the specific steps are as follows:

[0049] S1. Extract the sunflower pollen by acidic water homogenization. Weigh 50g of sunflower pollen and add 2.5L of 0.02% citric acid aqueous solution for extraction. The solid-liquid ratio between the sunflower pollen and the citric acid aqueous solution is 1:50. Homogenize at room temperature for 5 minutes, sieve through a 500-mesh screen, separate and obtain an extract, concentrate the extract to 0.8L at 55°C, pre-cool in a -80°C refrigerator, and freeze-dry in a vacuum freeze dryer to obtain an extract. The extract is precipitated with alcohol: weigh the extract and dissolve it in water. The solid-liquid ratio between the extract and water is 1:5 (g / mL). After the extract is completely dissolved, add anhydrous ethanol until an 80% ethanol mixture is obtained. Ultrasonicate for 5 minutes and let stand for 12 hours. Centrifuge and collect the precipitate to obtain 6.1g of crude sunflower pollen polysaccharide.

[0050] S2. The crude sunflower pollen polysaccharide obtained in step S1 is configured into an aqueous solution with a concentration of 5 mg / mL. The crude sunflower pollen polysaccharide and filler are loaded in a mass ratio of 1:50 (g:g) for purification on a bonded phase silica gel separation column. The aqueous solution of the crude sunflower pollen polysaccharide is gradient eluted at a flow rate of 1 mL / min. During the gradient elution, 1% acetonitrile is used for elution for 10 min, 2% acetonitrile is used for elution for 10 min, 3% acetonitrile is used for elution for 10 min, 4% acetonitrile is used for elution for 10 min, and 5% acetonitrile is used for elution for 10 min. Then, the eluate obtained after elution with 2% acetonitrile / elution peak is collected to obtain 4.40 g of preliminarily purified sunflower pollen polysaccharide;

[0051] The bonded phase silica gel separation column uses a spherical C18 column with a particle size of 20-45μm and a pore size of

[0052] S3. The preliminarily purified sunflower pollen polysaccharide obtained in step S2 is further purified by gel chromatography column, and the preliminarily purified sunflower pollen polysaccharide and filler are loaded at a mass ratio of 1:60 (g:g), pure water is used as the mobile phase, the flow rate is 0.5 mL / min, and the gel in the gel layer of the gel chromatography column is dextran coagulation G-25. The eluate of the maximum elution peak is collected, and after drying with nitrogen, a yellow solid is obtained, which is 0.26 g of purified sunflower pollen polysaccharide C.

[0053] Determination of Molecular Weight of Sunflower Pollen Polysaccharide C

[0054] 5 mg of the purified sunflower pollen polysaccharide C prepared in Example 1 was weighed, and the molecular weight of the polysaccharide was detected by gel exclusion chromatography columns Ohpak SB-805HQ (300×8 mm) and Ohpak SB-803HQ (300×8 mm) in series.

[0055] The absolute molecular weight test results of Helianthus annuus pollen polysaccharide C are as follows Figure 3 As shown, the chromatographic data were processed using software ASTRA 6.1 to obtain the purified sunflower pollen polysaccharide C with an Mn molecular weight of 1.302 kDa, an Mp molecular weight of 0.652 kDa, and an Mw molecular weight of 3.265 kDa.

[0056] Determination of monosaccharide composition of sunflower pollen polysaccharide C:

[0057] The purified sunflower pollen polysaccharide C prepared in Example 1 was tested and analyzed, and scanned at wavelengths of 260 nm and 280 nm. It was found that there was no absorption wave at these wavelengths, indicating that it did not contain nucleic acids and proteins.

[0058] 5 mg of the purified sunflower pollen polysaccharide C prepared in Example 1 was weighed, and the monosaccharide components of the polysaccharide were analyzed and detected using a Thermo ICS 5000+ ion chromatography system (ICS 5000+, Thermo Fisher Scientific, USA) and an electrochemical detector.

[0059] The monosaccharide composition chromatogram of the C component of Helianthus annuus pollen polysaccharide is shown in Figure 5 As shown, Figure 5 Compared with the test spectra of various monosaccharide standards ( Figure 4 ), these monosaccharide peaks were identified as arabinose, galactose, and glucose.

[0060] The chromatographic data were processed using the Chromeleon software, and the molar ratio of the three monosaccharides, arabinose, galactose, and glucose, in the purified helianthus pollen polysaccharide C was found to be 4:1:15.

[0061] The sunflower polysaccharide C prepared in Example 1 was tested for the following properties:

[0062] 1. Anti-inflammatory ability test of sunflower pollen polysaccharide C:

[0063] A: Effect of samples on the proliferation of RAW264.7 cells

[0064] Cells in logarithmic growth phase were collected and counted at 2.0×10 5 / mL cell density was inoculated into a 96-well plate, 100μL per well, and cultured for 24h; the supernatant was discarded, and the plates were randomly divided into control wells and sample wells. 100μL of complete culture medium was added to the control wells, and 100μL of sample culture medium containing different concentrations of sunflower pollen polysaccharide C (6.25μg / mL, 12.5μg / mL, 25μg / mL, 50μg / mL, 100μg / mL, 500μg / mL, 1000μg / mL) was added to the sample wells, with 5 replicate wells for each concentration, and cultured for 24h; the culture medium was discarded, and 100μL of culture medium containing 10% CCK-8 reagent was added to each well. After incubation for 2h, the sample absorbance was measured at 450nm. The cell survival rate was calculated according to the following formula:

[0065] Cell viability (%) = OD value of sample well / OD value of control well × 100%.

[0066] The cell viability results were Figure 6 As shown, where C refers to the test results in the control well, sunflower pollen polysaccharide C had no significant effect on the proliferation of RAW264.7 cells in the concentration range of 6.25-1000 μg / mL (P>0.05).

[0067] B: Cellular NO content detection

[0068] Cells in logarithmic growth phase were collected and counted at 2.0×10 5 / mL cell density was inoculated into a 96-well plate, 100 μL per well, and cultured for 24 h; the cells were randomly divided into normal control wells, model wells, and sample wells. The normal control wells were added with 100 μL complete culture medium, the model wells were added with 100 μL culture medium containing 1 μg / ml LPS, and the sample wells were added with culture medium containing 1 μg / ml LPS and different concentrations of sunflower pollen polysaccharide C samples (12.5 μg / mL, 25 μg / mL, 50 μg / mL, 100 μg / mL, 500 μg / mL, 1000 μg / mL) and cultured for 24 h. According to the instructions of the nitric oxide kit (Beijing Biyuntian), the NO concentration in each well was determined and calculated.

[0069] NO concentration Figure 7As shown, C refers to the test results in the normal control wells, and M refers to the test results in the model wells. When stimulated by 1 μg / mL LPS, the NO secretion of RAW 264.7 cells increased significantly (P<0.01). Sunflower pollen polysaccharide C had a 46.0% inhibitory effect on the NO secretion of RAW 264.7 cells induced by LPS at a concentration of 1000 μg / mL. The results showed that sunflower pollen polysaccharide C had a good anti-inflammatory effect at a concentration of 12.5-100 μg / mL.

[0070] 2. Blood pressure reduction ability test:

[0071] The ACE Kit-WST kit (Tongren Chemical) for detecting the activity of angiotensin converting enzyme (ACE) inhibitors was used to detect the inhibitory ability of different concentrations of sunflower pollen polysaccharide C (0 mg / mL, 0.01 mg / mL, 0.03 mg / mL, 0.05 mg / mL, 0.07 mg / mL, 0.1 mg / mL), and then the ACE enzyme IC50 value (half inhibition concentration value, the concentration value of sunflower pollen polysaccharide C at 50% inhibition rate) was calculated. The positive group was captopril. The test results are shown in Table 1.

[0072] Table 1

[0073] sample ACE enzyme inhibition ability (IC50) (mg / mL) Positive Control <![CDATA[0.03×10 -3 ]]> Sunflower Pollen Polysaccharide C 0.0368

[0074] Among them, the ACE enzyme inhibitory ability of 0.03 mg / mL sunflower pollen polysaccharide C is 41.8%. The IC50 value of helianthus annuus pollen polysaccharide C calculated according to the measurement results is shown in Table 1. Sunflower pollen polysaccharide C has ACE enzyme inhibitory ability, and the ACE enzyme inhibitory ability IC50 value is 0.0368 mg / mL. It can effectively dilate cardiovascular and cerebrovascular vessels and has a good antihypertensive effect.

[0075] 3. Test on the ability to extend the life span of brewer's yeast:

[0076] The cerevisiae yeast cultured to the logarithmic phase was 600 The inoculum of 0.01 was inoculated into YPD medium divided into blank group, low-dose group (sunflower pollen polysaccharide C content 1 μg / mL), medium-dose group (sunflower pollen polysaccharide C content 10 μg / mL) and high-dose group (sunflower pollen polysaccharide C content 50 μg / mL), and cultured in a 30°C constant temperature incubator at 220 r / min. The bacteria cultured for 3, 5, 7, 9, and 11 days were collected, washed 3 times with sterile water, and resuspended to adjust the OD 600 0.5, then diluted to 1×10 5 times, aspirate 100 μL to spread on solid YPD plates, place them in a constant temperature culture at 30°C for 3 days, and determine the survival rate (chronological life span) of the strain by the viable bacteria counting method.

[0077] The results of strain survival test are as follows Figure 8 As shown, by comparing the chronological lifespan with the blank group, it was found that the low-dose group, the medium-dose group and the high-dose group all had different degrees of beneficial effects on the chronological lifespan of yeast, among which the survival rate of the chronological lifespan experiment of the medium-dose group was still 55.1% on the 11th day. This shows that sunflower pollen polysaccharide C can prolong the chronological lifespan of yeast and has a good anti-aging effect.

[0078] 4. In vitro antioxidant capacity test

[0079] Determination of DPPH free radical antioxidant capacity: Accurately weigh 2.5 mg of DPPH and dilute to 100 mL with anhydrous ethanol to prepare a mass concentration of 2.5 × 10 -2 The standard solution of 1000 mg / mL was stored in the refrigerator for future use (prepared before use). 2.0 ml of sunflower pollen polysaccharide C solution of different concentrations (0 mg / mL, 0.5 mg / mL, 1.0 mg / mL, 1.5 mg / mL, 2 mg / mL, 2.5 mg / mL) was taken respectively and placed in a 10 ml centrifuge tube, and 2.0 ml of DPPH solution was added. After vortex mixing, the mixture was reacted at room temperature in the dark for 30 minutes, and the absorbance was detected at 517 nm. Anhydrous ethanol + DPPH was used as the blank group, anhydrous ethanol + sample was used as the background group, sample + DPPH was used as the experimental group, and Vc + DPPH was used as the positive group.

[0080] The calculation formula of DPPH free clearance rate is as follows:

[0081] Clearance rate (%) = [blank group - (experimental group - background color group)] / blank group * 100%;

[0082] In the above formula, blank group, experimental group, and background color group all refer to the corresponding absorbance values.

[0083] ABTS+ scavenging ability determination: accurately prepare 200mM phosphate buffer pH7.4, 3mM·ABTS solution, 1mM myoglobin solution, and 2mM·H2O2 solution. Sequentially add 100μL phosphate buffer, 50μL sunflower pollen polysaccharide C solution (0mg / mL, 0.5mg / mL, 1.0mg / mL, 1.5mg / mL, 2mg / mL, 2.5mg / mL), 10μL ABTS, and 10μL myoglobin to the 96-well plate, and then add 10μL H2O2 solution to the sample in front of the microplate reader. Under constant temperature conditions of 37°C, at 734nm, measure the absorbance every 1min for 15min. Distilled water and Vc are used as blank and positive groups. After the determination, a dynamic absorption graph is drawn based on the data, and the IC50 value is calculated based on the absorbance value of samples of different concentrations at 15min. The calculation formula for ABTS+ scavenging rate is as follows:

[0084] ABTS+ clearance rate (%) = (blank group - sample group) / blank group × 100%;

[0085] In the above formula, blank group and sample group both refer to the corresponding absorbance values.

[0086] The results of the antioxidant capacity (IC50) test are shown in Table 2.

[0087] Table 2

[0088]

[0089] The ABTS+ scavenging rate of 1.5 mg / mL sunflower pollen polysaccharide C solution was 50.7%. The IC50 value of sunflower pollen polysaccharide C was calculated based on the test results as shown in Table 2. The IC50 value of DPPH scavenging ability was 1.0 mg / mL, and the IC50 value of ABTS scavenging ability was 1.48 mg / mL. Sunflower pollen polysaccharide C has excellent antioxidant capacity.

[0090] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A sunflower pollen polysaccharide C, characterized in that: The number average molecular weight of the sunflower pollen polysaccharide C is 1.302 kDa, the peak molecular weight is 0.652 kDa, and the weight average molecular weight is 3.265 kDa. The sunflower pollen polysaccharide C is composed of arabinose, galactose and glucose, and the molar ratio of arabinose, galactose and glucose is 4:1:

15. The steps of preparing the sunflower pollen polysaccharide C are as follows: S1. extracting the sunflower pollen with acidic water homogenization, and then concentrating, freeze-drying and precipitating the extract with alcohol to obtain crude sunflower pollen polysaccharides; S2. preparing an aqueous solution of the crude sunflower pollen polysaccharide obtained in step S1, purifying the aqueous solution of the crude sunflower pollen polysaccharide using a bonded phase silica gel separation column, collecting the eluate, and obtaining a preliminarily purified sunflower pollen polysaccharide; S3. The preliminarily purified sunflower pollen polysaccharide obtained in step S2 is further purified using a gel chromatography column, the eluate with the largest elution peak is collected, and dried with nitrogen to obtain a yellow solid, which is purified sunflower pollen polysaccharide C.

2. The method for preparing sunflower pollen polysaccharide C according to claim 1, characterized in that: The specific steps are as follows: S1. extracting the sunflower pollen with acidic water homogenization, and then concentrating, freeze-drying and precipitating the extract with alcohol to obtain crude sunflower pollen polysaccharides; S2. preparing an aqueous solution of the crude sunflower pollen polysaccharide obtained in step S1, purifying the aqueous solution of the crude sunflower pollen polysaccharide using a bonded phase silica gel separation column, collecting the eluate, and obtaining a preliminarily purified sunflower pollen polysaccharide; S3. The preliminarily purified sunflower pollen polysaccharide obtained in step S2 is further purified using a gel chromatography column, the eluate with the largest elution peak is collected, and dried with nitrogen to obtain a yellow solid, which is purified sunflower pollen polysaccharide C.

3. The preparation method according to claim 2, characterized in that: The specific steps of the acidic water homogenization extraction in step S1 are: adding sunflower pollen into an acidic solvent for extraction, homogenizing for 5-20 minutes, and separating to obtain an extract.

4. The preparation method according to claim 3, characterized in that: The solid-liquid ratio between sunflower pollen and acidic solvent is 1:(30-50).

5. The preparation method according to claim 2, characterized in that: The specific steps of step S2 are: in the bonded phase silica gel separation column purification, the aqueous solution of the crude sunflower pollen polysaccharide obtained in step S1 is eluted with a gradient of 1%-5% acetonitrile, and then the eluate of the elution peak obtained after elution with 2% acetonitrile is collected to obtain the preliminary purified sunflower pollen polysaccharide.

6. The preparation method according to claim 2, characterized in that: In the step S3, the gel chromatography column purification uses pure water as the mobile phase for purification.

7. Use of the sunflower pollen polysaccharide C according to claim 1 in the preparation of medicines, health products or daily chemicals with anti-inflammatory effects.

8. Use of the sunflower pollen polysaccharide C according to claim 1 in the preparation of medicines, health products or daily chemicals with antihypertensive effects.

9. Use of the sunflower pollen polysaccharide C according to claim 1 in the preparation of medicines, health products or daily chemicals with anti-aging effects.

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