Carthamus tinctorius polysaccharide for lung targeting as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of medicines, and particularly relates to safflower polysaccharide for lung targeting as well as a preparation method and application thereof. The safflower polysaccharide comprises the following monosaccharides in percentage by mole: 0.75% of L-fucose, 36.04% of arabinose, 25.03% of rhamnose, 17.85% of galactose, 4.26% of glucose, 1.33% of xylose, 0.61% of mannose, 12.07% of galacturonic acid and 2.06% of glucuronic acid, and the total percentage is 100%. The molecular weight of the safflower polysaccharide is 16.27 kDa. The safflower polysaccharide disclosed by the invention can be absorbed into blood within 24 hours and almost completely metabolized, and can be used for preparing lung targeting products.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of medicine, and particularly relates to a safflower polysaccharide for lung targeting, a preparation method and application thereof. BACKGROUND

[0002] As one of the important substances constituting life, polysaccharides widely exist in organisms such as higher plants, animals and microorganisms, and exhibit many potential advantages in the field of biological medicine. A large number of studies have confirmed that polysaccharides have immune regulation function, can stimulate the immune system of the body, enhance the activity of immune cells, and improve the defense ability of the body to pathogens, and can be used as a natural immune enhancer to improve the body's resistance to infection, which has important significance for people with low immunity. In terms of anti-inflammatory, polysaccharides can inhibit the production and activation of inflammatory cytokines, and have potential value for the treatment or prevention of inflammatory-related diseases such as rheumatoid arthritis and systemic lupus erythematosus. At the same time, polysaccharides have antioxidant properties, can effectively scavenge free radicals, and neutralize harmful substances produced by oxidative stress, which helps to delay aging and reduce the risk of cardiovascular disease. At present, the research on polysaccharides has made certain progress in extraction and separation and purification. In terms of extraction method, hot water extraction, ultrasonic extraction, acid and alkali extraction, etc. have their own advantages and disadvantages. In terms of separation and purification means, macroporous resin method, anion exchange chromatography, gel chromatography and membrane separation method are widely used. However, although polysaccharides exhibit good efficacy and application potential in many aspects, there are few studies on lung targeting. From the perspective of drug administration route, the lung has a large surface area and rich capillaries, and drugs administered through the lung can directly enter the blood circulation, avoiding the first-pass effect of the liver, which is a very potential drug administration site. However, there are few polysaccharide preparations for lung targeting at present, which limits the application of polysaccharides in the treatment of lung diseases such as asthma, chronic obstructive pulmonary disease and lung infection. From the perspective of immune regulation, the immune environment of the lung is complex, and if polysaccharides can be targeted to the lung, it is expected to precisely regulate the lung immune microenvironment, enhance the immune defense of the lung, and at the same time reduce unnecessary effects on the whole body immune system, but relevant research is still in its infancy, and therefore more polysaccharide drugs suitable for lung targeting need to be developed to open up new ideas for the treatment of lung diseases. SUMMARY

[0003] In order to solve the above problems, the present application provides a safflower polysaccharide for lung targeting, a preparation method and application thereof. The specific technical solutions are as follows.

[0004] The safflower polysaccharide for lung targeting provided by the first aspect of the present application has the following proportions of individual monosaccharides in terms of molar percentage: 0.75% of L-fucose, 36.03% of arabinose, 25.03% of rhamnose, 17.85% of galactose, 4.26% of glucose, 1.33% of xylose, 0.61% of mannose, 12.07% of galacturonic acid, and 2.06% of glucuronic acid, with a total of 100%. The safflower polysaccharide has an alternating connection of galacturonic acid and rhamnose to form a main structure form in a main chain, and the main chain is mainly composed of 1→4 and 1→2 cross-linked glycosidic bonds in the structure of the repeating unit, the 4 position of rhamnose is the starting point of the main branch, and the main component of the branch is arabinose and galactose, part of the arabinose is connected by 1→5 glycosidic bond and arabinose as the terminal sugar, and the other part is connected by 1→4 glycosidic bond and galactose as the terminal sugar.

[0005] In another preferred embodiment, the molecular weight of the safflower polysaccharide is 16.27 kDa.

[0006] The second aspect of the present application provides a preparation method of the safflower polysaccharide for lung targeting, comprising the following steps: The safflower powder is mixed with an alkali solution, and then extracted at 20-60 DEG C for 30-120 min, centrifuged, the supernatant is concentrated, deproteinized, dialyzed to obtain the safflower crude polysaccharide; the safflower crude polysaccharide is subjected to DEAE-FF agarose gel column, eluted with 0.3 mol / L NaCl, the eluate is collected, concentrated, dialyzed, and freeze-dried, and then subjected to S-400 gel column for purification, concentrated, dialyzed with deionized water, and freeze-dried to obtain the safflower polysaccharide.

[0007] In another preferred embodiment, the alkali solution is a sodium hydroxide solution with a concentration of 0.4-0.8 M.

[0008] In another preferred embodiment, the specific process of the S-400 gel column for purification is as follows: eluted with pure water at a flow rate of 0.5 mL / min, the eluate is collected, concentrated, dialyzed, and freeze-dried to obtain the safflower polysaccharide.

[0009] In another preferred embodiment, the concentration refers to concentrating the supernatant to 1 / 3-1 / 5 of the original volume.

[0010] In another preferred embodiment, the reagent used in the alcohol precipitation is 95% ethanol by mass percentage; the reagent used in the deproteinization is Sevag reagent; and the dialysis uses a dialysis bag with a molecular weight of 7-14 kDa.

[0011] The third aspect of the present application provides the use of the safflower polysaccharide for lung targeting in the preparation of a lung targeting product, wherein the product is an oral preparation, the concentration of the safflower polysaccharide in the oral preparation is 10 mg / mL to 12 mg / mL, and the solvent of the oral preparation is water.

[0012] The fourth aspect of the present application provides the use of the safflower polysaccharide for lung targeting in the preparation of a product for relieving lung injury, wherein the product is an injection, and the solvent of the injection is physiological saline.

[0013] The fifth aspect of the present application provides the use of the safflower polysaccharide for lung targeting in the preparation of an immunomodulator product.

[0014] Compared with the prior art, the present application has the following beneficial effects: The present application provides a safflower polysaccharide for lung targeting, and the composition and structure of the safflower polysaccharide are determined, wherein the proportions of the individual monosaccharides are as follows: 0.75% of L-fucose, 36.04% of arabinose, 25.03% of rhamnose, 17.85% of galactose, 4.26% of glucose, 1.33% of xylose, 0.61% of mannose, 12.07% of galacturonic acid, and 2.06% of glucuronic acid, according to the molar percentage, and the total is 100%. After the safflower polysaccharide in the present application is administered by gavage, a strong fluorescence signal can be observed in the lungs of mice within 3h to 24h, which proves that the safflower polysaccharide in the present application has a good absorption effect in the lung and can be absorbed into the blood within 24h and almost completely metabolized, indicating that the safflower polysaccharide can be used for preparing a lung targeting product. In addition, it is found through an acute lung injury model experiment of mice that the safflower polysaccharide can effectively inhibit lung inflammation and immune disorders induced by LPS. BRIEF DESCRIPTION OF DRAWINGS

[0015] Figure 1 The figure is a property analysis result of SPS obtained by different extraction methods; wherein, A is a high-performance anion exchange chromatography analysis result, B is a molecular weight analysis result, C is an FT-IR spectrum, and D is a UV spectrum. Among them, SPS represents a safflower crude polysaccharide.

[0016] Figure 2 The figure is a Congo red experiment, a potassium iodide experiment, and an XRD analysis of SPS obtained by different extraction methods; wherein, A is a Congo red experiment result, B is a potassium iodide experiment result, and C is an XRD analysis.

[0017] Figure 3 The figure is a SEM image of SPS obtained by different extraction methods.

[0018] Figure 4Fig. 1 is a thermogravimetric curve of SPS obtained by different extraction methods; wherein A is a thermogravimetric curve of SPS obtained by hot water extraction, B is a thermogravimetric curve of SPS obtained by ultrasonic extraction, C is a thermogravimetric curve of SPS obtained by acid extraction, and D is a thermogravimetric curve of SPS obtained by alkali extraction.

[0019] Figure 5 Fig. 2 is a functional property and biocompatibility result diagram of SPS obtained by different extraction methods; wherein A is a water holding capacity analysis effect diagram, B is a flocculation activity result analysis diagram, C is a foaming amount and foam stability analysis diagram, and D is an emulsification activity index and emulsion stability index analysis result diagram; in the diagram, WHC represents water holding capacity, FAC represents flocculation activity, FC represents foaming amount, FS represents breaking stability, EAI represents emulsification activity index, and ESI represents emulsion stability index.

[0020] Figure 6 Fig. 3 is a rheological property analysis diagram of SPS obtained by different extraction methods; wherein A is an apparent viscosity analysis diagram, and B is a shear stress analysis diagram. Figure 7 Fig. 4 is a storage modulus and loss modulus curve diagram of SPS obtained by different extraction methods; A is a storage modulus and loss modulus curve diagram of SPS obtained by hot water extraction, B is a storage modulus and loss modulus curve diagram of SPS obtained by ultrasonic extraction, C is a storage modulus and loss modulus curve diagram of SPS obtained by acid extraction, and D is a storage modulus and loss modulus curve diagram of SPS obtained by alkali extraction.

[0021] Figure 8 Fig. 5 is an in vitro antioxidant and anti-inflammatory capacity analysis result diagram of SPS obtained by different extraction methods; wherein A is a reducing power analysis result diagram, B is an ABTS free radical scavenging activity analysis result diagram, C is a DPPH free radical scavenging activity analysis result diagram, and D is an albumin denaturation inhibition and NO generation inhibition capacity analysis result diagram.

[0022] Figure 9 Fig. 6 is a biocompatibility and immune regulation capacity analysis result diagram of SPS obtained by different extraction methods; wherein A is a cell viability analysis result diagram, B is a blood compatibility evaluation analysis result diagram, C is a ROS analysis result diagram, D is a NO generation analysis result diagram, E is a phagocytic capacity analysis result diagram, F is an IL-6 analysis result diagram, G is an IL-1β analysis result diagram, and H is a TNF-α analysis result diagram.

[0023] Figure 10 Fig. 7 is an analysis diagram of the influence of different extraction conditions on the content of safflower polysaccharide; wherein A is an extraction time influence analysis diagram, B is an extraction temperature influence analysis diagram, C is a NaOH concentration influence analysis diagram, and D is a solid-liquid ratio influence analysis diagram.

[0024] Figure 11 Three-dimensional response surface and contour plots of different conditions, wherein A is the contour plot of extraction time and extraction temperature, B is the contour plot of extraction time and NaOH concentration; C is the contour plot of extraction time and solid-liquid ratio, D is the three-dimensional response surface plot of polysaccharide content and extraction temperature, E is the three-dimensional response surface plot of polysaccharide content and NaOH concentration, F is the three-dimensional response surface plot of polysaccharide content and solid-liquid ratio, Figure 12 Three-dimensional response surface and contour plots of different conditions, wherein A is the contour plot of extraction temperature and NaOH concentration; B is the contour plot of extraction temperature and solid-liquid ratio, C is the contour plot of NaOH concentration and solid-liquid ratio, D is the three-dimensional response surface plot of polysaccharide content and NaOH concentration, E is the three-dimensional response surface plot of polysaccharide content and extraction temperature, F is the three-dimensional response surface plot of polysaccharide content and solid-liquid ratio.

[0025] Figure 13 Artificial neural network analysis chart; wherein, A is the analysis chart of the number of hidden layer neurons on DA, B is the artificial neural network model architecture topology analysis chart for AES extraction, C is the regression analysis chart of actual value and artificial neural network model prediction result, D is the training performance chart of artificial neural network, E is the evolution analysis chart of best and average fitness in traditional algorithm.

[0026] Figure 14 Elution curve and structure analysis chart of AES-S; wherein, A is the elution curve chart on DEAE-FF Sepharose gel column, B is the elution curve chart on S-400 gel filtration chromatography column, C is the molecular weight chart of AES-S, D is the infrared spectrum analysis chart of AES-S.

[0027] Figure 15 Structure analysis characterization chart of AES-S; A is 1 H NMR chart, B is 13 C NMR chart, C is the RAMP chart, D is the two-dimensional nuclear magnetic spectrum chart, E is the HSQC chart, F is the HMBC chart, G is the NOESY chart, and H is the TOCSY chart.

[0028] Figure 16 Infrared spectrum analysis chart and digestion experiment chart of AES-S-Cy5.5; wherein, A is the infrared spectrum analysis chart, and B is the digestion experiment chart.

[0029] Figure 17 Dynamic distribution chart of oral AES-S-Cy5.5 and Cy5.5 in vivo; wherein, A is the near-infrared fluorescence imaging chart in vivo within 24 hours after administration, and B is the near-infrared fluorescence imaging chart of the lung and different gastrointestinal tracts within 24 hours.

[0030] Figure 18Figure for the body weight change after gavage of AES-S.

[0031] Figure 19 Figure for the thymus index after gavage of AES-S.

[0032] Figure 20 Figure for the proportion analysis results of T lymphocyte subsets CD4 / CD3 in the spleen of mice.

[0033] Figure 21 Figure for the columnar analysis of Figure 20

[0034] Figure 22 Figure for the proportion analysis results of T lymphocyte subsets CD8 / CD3 in the spleen of mice.

[0035] Figure 23 Figure for the columnar analysis of Figure 22

[0036] Figure 24 Figure for the proportion analysis results of T lymphocyte subsets CD4 / CD8 in the spleen of mice.

[0037] Figure 25 Figure for the columnar analysis of Figure 24 DETAILED DESCRIPTION

[0038] In order to make the objects, technical solutions and advantages of the present application clearer, the present application will be further described in detail below with reference to examples. It should be understood that the specific examples described herein are only used to explain the present application, and are not used to limit the present application. The experimental methods described in the following examples are conventional methods unless otherwise specified. The reagents and materials described are commercially available unless otherwise specified.

[0039] A safflower polysaccharide for lung targeting, a preparation method thereof and an application thereof are described below.

[0040] Example 1: Determination of an extraction method of a safflower polysaccharide for lung targeting.

[0041] 1. Extraction method: In order to determine the most suitable extraction method, four extraction methods, i.e., hot water extraction (HWES), ultrasonic wave extraction (UAES), acid extraction (HES) and alkali extraction (AES), were used to extract the polysaccharide in safflower. 95% ethanol was used to remove fat and pigment components in safflower powder. The specific process is as follows.

[0042] ​​​Hot water extraction (HWES): 150 g of safflower powder was mixed with 4.5 L of deionized water at a solid-to-liquid ratio of 1:30, and the mixture was extracted at 90 °C for 1 h in a water bath. The safflower residue was recovered, and the extraction was repeated three times. The combined extract was concentrated to 1 / 3 of the original volume, precipitated in 95% ethanol overnight, and the precipitate was treated with Sevag reagent to remove proteins. Finally, the extract was dialyzed against water at room temperature for 48 h and then against distilled water for another 48 h. The hot water extracted safflower crude polysaccharides were obtained by freeze-drying.

[0043] Ultrasonic-assisted extraction (UAES): The only difference was that the ultrasonic extractor was used for extraction, and the ultrasonic power was adjusted to 300 W at 50 °C for 30 min. After centrifugation, concentration, protein removal, and dialysis, the ultrasonic-assisted extracted safflower crude polysaccharides were obtained by freeze-drying.

[0044] Acid extraction (HES): 150 g of safflower powder was mixed with 4.5 L of HCl solution (pH = 3), and the mixture was extracted at 70 °C for 1 h in a water bath. After centrifugation, concentration, protein removal, and dialysis, the acid-extracted safflower crude polysaccharides were obtained by freeze-drying.

[0045] Alkaline extraction (AES): 150 g of safflower powder was mixed with 4.5 L of 0.5 M NaOH solution, and the mixture was mixed uniformly. After centrifugation, concentration, protein removal, and dialysis, the alkaline-extracted safflower crude polysaccharides were obtained by freeze-drying. The safflower crude polysaccharides extracted by the above methods were denoted as SPS.

[0046] 2. Determination of the physicochemical properties of safflower polysaccharides The physicochemical properties of the safflower polysaccharides extracted by the above methods were detected. The contents of polysaccharides, phenols, proteins, and flavonoids were detected by the phenol-sulfuric acid method, the gallic acid method, the Coomassie brilliant blue method, and the quercetin-AlCl3 method, respectively. The results are shown in Table 1. As can be seen from Table 1, the yield and total sugar content of AES were significantly higher than those of the other three SPS.

[0047] Table 1 Extraction yield and chemical composition of polysaccharides extracted from safflower by different methods Note: GAE represents gallic acid, and RE represents rutin acid.

[0048] 3. Determination of the extraction yield of different parts of safflower The sugar content and extraction yield of different parts of safflower were determined by the phenol-sulfuric acid method, and the results are shown in Table 2.

[0049] Table 2 Extraction yield and polysaccharide content of different parts of safflower The polysaccharide content and extraction rate of different safflower wastes were significantly different. The polysaccharide content (152.36 ± 4.12 mg / g) and extraction rate (43.87 ± 0.89%) of safflower filaments were the highest, while the polysaccharide content (57.50 ± 3.10 mg / g) and extraction rate (5.4 ± 0.2%) of safflower leaves were the lowest.

[0050] 4. Monosaccharide composition of crude safflower polysaccharides obtained by different extraction methods SPS was degraded with trifluoroacetic acid (2M) at 121 °C for 2 h, then dried with nitrogen. The residue was redissolved in deionized water and filtered through a 0.22 μm microfiltration membrane for further measurement. The results are shown in Table 3.

[0051] Table 3. Monosaccharide composition of SPS (mol%) Note: Man represents mannose, GlcA represents glucuronic acid, Rha represents rhamnose, GalA represents galacturonic acid, Glc represents glucose, Gal represents galactose, Xyl represents xylose, Ara represents arabinose.

[0052] As can be seen from Table 3, all SPSs contain eight common monosaccharides, including mannose, glucuronic acid, rhamnose, galacturonic acid, glucose, galactose, xylose and arabinose (A in Table 3). This indicates that the overall monosaccharide composition type of these polysaccharides remains consistent. Figure 1

[0053] 5. Molecular weight detection of safflower polysaccharides The molecular weight of SPS was determined by high-performance gel permeation chromatography (HPGPC-ELSD) using an Agilent 1260 Infinity ELSD evaporative light scattering detector, a TSK-gel G-3000 PWXL chromatographic column (7.8 x 300 nm), and the results are shown in B in Figure 1 HES showed the highest molecular weight of 404.38 kDa. The molecular weights of HWES, UAES and AES were 382.77 kDa, 256.46 kDa and 308.31 kDa, respectively.

[0054] 6. Infrared spectrum detection of safflower polysaccharides FT-IR spectra were tested on an infrared spectrometer using a Nexus 470 FT-IR spectrometer in the range of 4000 cm -1 to 400 cm -1 . The results are shown in Figure 1 ​Figure 1 shows the FTIR spectra of SPS obtained by HWE, UAE, HE and AES extraction methods. As shown in Figure 1C, SPS obtained by HWE, UAE, HE and AES extraction methods retained the characteristic absorption patterns of polysaccharides, which indicated that the basic structural integrity of functional groups was largely unaffected by the extraction techniques. Polysaccharides prepared by HWE, UAE and HE showed an additional absorption peak at 1542 cm -1 relative to SPS prepared by the AES method. This additional peak can be attributed to C=C stretching vibration within the sugar chain. These observations indicate that the core structural features of polysaccharides were largely unchanged under different extraction methods.

[0055] 7. UV Spectroscopic Analysis of Safflower Polysaccharides UV spectroscopic analysis was performed in the range of 200 nm to 400 nm to determine the presence of proteins and nucleic acids in SPS using a full wavelength UV spectrophotometer (TECAN-Spark, Switzerland). SPS and potassium bromide powder were pressed into thin sheets. Figure 1 Figure 1D shows that SPS prepared by the AES method exhibited absorption peaks between 250 nm and 300 nm, indicating the presence of impurities such as proteins and nucleic acids in the polysaccharide sample obtained by this method, which is consistent with the higher protein content of AES compared to the other three SPS in Table 1.

[0056] 8. Congo Red Assay of Safflower Polysaccharides SPS solution (2 mg / mL, 1 mL) was mixed with Congo red solution (0.2 mM, 1 mL) and different volumes of NaOH solution (1 M) to a total volume of 4 mL, so that the concentration of NaOH was 0 M to 0.5 M. After 5 min of reaction at room temperature, the maximum absorption wavelength (λmax) of the SPS-Congo red complex was screened on a UV spectrophotometer in the range of 400 nm to 700 nm. The results are shown in Figure 2A. Figure 2 As shown in Figure 2A, when the concentration of sodium hydroxide was about 0.1 M, the Congo red-SPS complex showed the highest absorption, and as the concentration of sodium hydroxide increased, the λmax of SPS decreased significantly. This result indicates that SPS does not have a triple helix structure.

[0057] 9. I2-KI Assay of Safflower Polysaccharides SPS solution (1.0 mg / mL, 2.0 mL) was mixed with iodine solution (0.2 wt% KI and 0.02 wt% I2, 1.2 mL), and the absorbance was measured by a microplate reader in the range of 300 nm to 700 nm. The results are shown in Figure 2B. Figure 2 As shown in Figure 2B, the maximum absorption peak of SPS was around 345 nm, and no absorption peak was detected at 565 nm, indicating that SPS may have a complex chain structure composed of longer and more branched chains.

[0058] 10. XRD Analysis of Safflower Polysaccharides The crystal structure of SPS was determined using X-ray diffraction. The dry sample powder was placed uniformly on a glass plate and scanned at a rate of 2° / min in the 2θ range of 5-90°. The results are shown in Figure 2 As shown in C, all four SPSs showed a broad bread-like peak at 2θ ≈ 20°, which suggested that the components of SPS were mainly in a semi-crystalline state.

[0059] 11. Scanning electron microscope images of Safflower polysaccharides The microstructure of SPS was recorded on a high-resolution field emission scanning electron microscope system. The accelerating voltage was 10 kV, and the magnification was 200x, 500x, 1000x and 3000x. The results are shown in Figure 2 As shown in C, HWES showed a smooth surface and dispersed micropores, indicating that hot water slowly penetrated the cell wall, dissolved polysaccharides and transferred them outside the cell, causing minimal damage to the cell organization of polysaccharides. The pores of HWES were round and more widely distributed, while the pores of HES were closely arranged, round and oval, and the pores of AES were irregular. The pores of UAES were larger than those of the other SPSs, which may be due to the strong cavitation, turbulence and shear force generated by the high-intensity ultrasonic equipment. Therefore, these results indicate that the extraction method affects the morphology of polysaccharides.

[0060] 12. Thermal properties of Safflower polysaccharides All SPSs showed two distinct thermal degradation stages, as shown in Figure 4 The TG curves of SPSs were relatively similar in shape. The initial mass loss of SPSs occurred in the temperature range of 30-110°C. Given the abundance of hydrophilic groups in polysaccharides, the evaporation of combined water was the main reason for mass loss in the first stage. AES showed the most significant water loss (11.60%), indicating that it had a strong water-holding capacity. The second stage weight loss of polysaccharides occurred in the temperature range of 200-350°C, mainly due to the degradation of sugar rings and chains. The weight of SPSs gradually decreased in the temperature range of 350-800°C. Compared with the other three SPSs, AES showed less mass loss in the second stage weight loss, which means that AES is more suitable for development as a drug or nutritional supplement compared with HWES, UAES and HES.

[0061] 13. Functional properties of Safflower polysaccharides The oil holding capacity (OHC), water holding capacity (WHC), foaming capacity and emulsifying capacity of emulsifiers were determined using a modified conventional method, and the results are shown in Figure 5 Different extraction methods had a significant impact on the WHC and OHC of the four different SPSs, with UAES extracted by the UAE method showing the most prominent WHC, 2.55 ± 0.14 g / g (p < 0.05). Figure 5On the other hand, HWES and HES showed the best OHC, which were 15.61±0.59 g / g and 15.17±0.57 g / g, respectively ( Figure 5 Overall, SPS plays a key role in preventing dehydration and improving the texture and viscosity of certain formulated foods due to its high WHC and OHC, and has the potential to be developed as a flavor enhancer and stabilizer. Figure 5 As shown in Figure C, SPS obtained by different extraction methods showed significant differences in foaming performance and foam stability. AES exhibited the best foaming ability, reaching 214.52 ± 8.7%, while HES showed the best foaming stability, at 82.89 ± 0.38%. All SPS exhibited good foaming ability and foam stability. Figure 5 As shown in Figure 3D, AES exhibited the highest EAI, measured at 5.14 ± 0.04 m² / g, significantly higher than the other SPSs. In contrast, HWES exhibited the best ESI, at 77.96 ± 1.39 min. Therefore, both HWES and AES are more suitable for use as stabilizers in food emulsions.

[0062] 14. Rheological properties of safflower polysaccharide The rheological properties of the polysaccharides were determined using an MCR302 rheometer (Anton Paar, Austria). Figure 6 Figures A and B show the relationship between the viscosity and shear rate of SPS extracted using different methods. The apparent viscosity of all four SPS samples decreases with increasing shear rate, showing typical shear thinning behavior. Among them, HES has the highest viscosity, which may be attributed to its higher molecular weight. In contrast, HWES and AES have lower apparent viscosities, which may be due to the disentanglement of their molecular chains in solution under alkaline and high temperature conditions. The dynamic scanning test results of SPS are shown in Figure 1. Figure 7 As shown in Figure 2, the dynamic modulus of SPS increases with the increase of frequency and shows obvious frequency dependence. For HWES and UAES, the storage modulus (G ' ) always exceeds the loss modulus (G '' ), indicating that elastic behavior is dominant in these two SPSs. In contrast, the G'' of HES and AES is higher than the storage modulus G ' , reflecting their more viscous liquid properties. These results highlight the different viscoelastic properties of the four SPSs and demonstrate that the extraction method significantly affects their apparent viscosity and dynamic modulus behavior.

[0063] 15. In vitro activity determination of safflower polysaccharide The antioxidant capacity of SPS was determined by ABTS, DPPH and reducing power. Ascorbic acid (Vc) was used as a positive control. The free radical scavenging capacity of SPS increased with increasing concentration (Figure 8 A) in Fig. 2. In terms of reducing power, UAES, HWES and AES showed significant improvement in reducing ability even with a small increase in concentration, while HES only showed a slight improvement under the same conditions. At a concentration of 4 mg / mL, the reducing power of UAES, HWES and AES was 0.89 ± 0.18 μmol / L, 0.68 ± 0.03 μmol / L and 0.62 ± 0.01 μmol / L, respectively. AES showed the highest ABTS radical scavenging rate at 93.66 ± 1.64 % at a concentration of 4 mg / mL, which was significantly higher than other SPS (P < 0.05) (Fig. 2B). With the increase of SPS concentration, DPPH scavenging activity gradually increased (Fig. 2C). At a concentration of 4 mg / mL, the DPPH scavenging ability of HWES, UAES, HES and AES was 35.62 ± 0.81 %, 26.69 ± 1.46 %, 47.25 ± 1.54 % and 65.69 ± 1.21 %, respectively. The potency of AES was significantly higher than other SPS (P < 0.05). In combination with the three antioxidant abilities, SPS extracted by the AE method had relatively the best antioxidant ability. SPS obtained by HWE, UAE, HE and AE methods all showed various antioxidant properties, including reducing power and radical scavenging activity. The anti-inflammatory properties of SPS were evaluated by BSA test and NO, and the results are shown in Fig. 2D. The inhibitory ability of HES was significantly higher than the other three SPS, reaching 75.52 ± 0.74 %. HWES showed the best performance in inhibiting NO production (13.39 ± 0.44 %). Overall, different extraction methods had a significant impact on the albumin denaturation inhibitory ability and NO production inhibitory ability of SPS. In addition, SPS also showed certain in vitro anti-inflammatory activity. Figure 8 Figure 8 Figure 8

[0064] 16. In vitro immune activity determination of safflower polysaccharides DCFH-DA staining technique was used to measure the ROS production induced by SPS at a concentration range of 0-100 μg / mL and lipopolysaccharide (LPS). Neutral red staining method was used to evaluate the effect of SPS on cell phagocytic activity. In addition, the levels of NO, IL-6, IL-1β and TNF-α in cell supernatant were determined according to the manufacturer's instructions. The proliferative effect of SPS on RAW264.7 cells was evaluated by CCK-8 method (Fig. 3A). The results showed that the two SPS significantly enhanced the proliferation of RAW264.7 cells, and no cytotoxicity was observed at a concentration of 100 μg / mL. At the same time, hemolysis test was used to determine the hemolytic activity of SPS at 800 µg / mL on healthy sheep red blood cells, and the results are shown in Fig. 3B. The results showed that SPS had no hemolytic activity on healthy sheep red blood cells. Figure 9 Figure 9 ​​​​As shown in B in Figure 1. According to ASTM classification standards, all SPS are non-hemolytic within the range of 2-5%. Therefore, SPS has good biocompatibility and safety and is suitable for further development. Figure 9 As shown in Figure C, compared with the blank group, all SPS significantly promoted the release of reactive oxygen species in macrophages at a concentration of 100 μg / mL. It is worth noting that AES has a stronger promoting effect on the release of cellular reactive oxygen species than LPS and Ganoderma lucidum polysaccharide groups (P<0.05), and is significantly stronger than the other three SPS. Nitric oxide (NO) is an important effector molecule of macrophages and a key indicator of macrophage activation. Figure 9 As shown in Figure D, the NO production in the SPS group was significantly higher than that in the control group, but its activity was still significantly lower than that in the LPS group (P<0.05). Figure 9 As shown in Figure E, the phagocytic ability of all SPSs was higher than that of the blank group (P<0.05). In addition, the phagocytic activity of AES (179.93±1.33%) was significantly higher than that of the other three SPS and LPS groups, and the phagocytic activity of the 100μg / mL AES-treated group was comparable to that of the LPS group (P<0.05). Therefore, the above results indicate that SPS significantly promoted macrophage proliferation, NO release, and phagocytosis, among which AES may serve as a new immunomodulator with important potential in regulating the immune process. Cytokine levels were measured by ELISA kits, including IL-6 ( Figure 9 F), IL-1β ( Figure 9 G) and TNF-α ( Figure 9 Compared with the control group, SPS significantly increased the release of IL-6, IL-1β, and TNF-α (P < 0.05). However, no significant difference in IL-6 release was observed in the LPS group treated with HWES, UAES, or AES (P > 0.05). These results suggest that SPS can promote the release of TNF-α, IL-6, and IL-1β cytokines, activating macrophages to exert their immune activity.

[0065] Example 2: AES extraction process optimization and safflower polysaccharide composition From the results in Example 1, it can be seen that the SPS obtained by AES has a higher yield and better antioxidant activity and in vitro immune activity than the other three extraction methods. Therefore, the AES extraction process was optimized.

[0066] 1. Single factor experiment and response surface methodology (RSM) were used to optimize the extraction conditions of AES polysaccharides.

[0067] Three factors were selected in the current one-factor experiment: time (30 min, 75 min and 120 min), temperature (20 °C, 40 °C and 60 °C), NaOH concentration (0.4 M, 0.6 M, 0.8 M) and the ratio of material to liquid (1 :30, 1 :50 and 1 :70 g / mL) Figure 10 ). Optimization employed a Box-Behnken design, with each variable coded into three levels (-1, 0, +1). RSM design and regression analysis were performed using Design-Expert 8.0.6. In addition, a feed-forward artificial neural network with backpropagation was used to dynamically model the AES method through MATLAB software. Two different sets of optimal extraction conditions were obtained using two different modeling calculation methods, as shown in Figures 11-13 , which were verified by triplicate experiments under the same conditions. The actual conditions for RSM verification were: time 30 min, temperature 35 °C, NaOH concentration 0.7 M, and the ratio of material to liquid 1 :60, with an optimal extraction rate of 32.48 ± 0.50%, close to the predicted value of 33.27% (P < 0.05). The actual verification conditions for ANN were: time 30 min, temperature 30 °C, NaOH concentration 0.6 M, and the ratio of material to liquid 1 :60, with an optimal extraction rate of 43.87 ± 0.89%, close to the predicted value (44.35%).

[0068] 2. Separation and purification of AES The polysaccharides were eluted from the DEAE-FF Sepharose column with deionized water and different concentrations of NaCl solution (0.1 mol / L, 0.3 mol / L, 0.5 mol / L and 1 mol / L) at a flow rate of 5 mL / min. The eluate was monitored and collected by the phenol-sulfuric acid method. The elution curve was plotted according to the absorbance values at 490 nm, as shown in Figure 14 . The samples of different components were collected, concentrated, dialyzed with deionized water, and freeze-dried. Since the peak was the highest when eluted with 0.3 mol / L NaCl, it was used for subsequent studies and named AES-D0.3. AES-D0.3 was further purified using an S-400 gel column, monitored by the phenol-sulfuric acid method, concentrated, dialyzed with deionized water, and freeze-dried. The further purified safflower polysaccharide was obtained, as shown in Figure 14 , and named AES-S.

[0069] 3. Determination of the monosaccharide composition of AES-S The AES-S monosaccharide composition was determined, and the results are shown in Table 4. AES-S is mainly composed of arabinose and rhamnose, with a total molar percentage content of 61.06%. After purification, the Rha and Ara of AES increased significantly, and GalA, Gal also increased relatively, which indicated that AES-S mainly contains pectin typical sugars, which are the key components of the pectin polysaccharide backbone and branches.

[0070] Table 4 AES-S monosaccharide composition 4. Methylation determination of AES-S After 3 mg of safflower polysaccharide was dissolved in DMSO, methylation reagent A / B was added in turn, and ultrasonic reaction was carried out at 30°C for 60 min. After hydrolysis, TFA treatment, sodium borohydride reduction, acetylation with acetic anhydride, and CH2Cl2 extraction and purification. GC-MS analysis was performed using a HP-INNOWAX column (140-230°C programmed temperature increase), and the helium flow rate was 1 mL / min. The results are shown in Table 5, and AES-S is mainly composed of Gal, Rha, Ara and Gal-UA sugars, which is consistent with the monosaccharide composition results. Rha in AES-S mainly exists in the form of →2)-Rhap-(1→ (12.82%) and →2,4)-Rhap-(1→ (9.5%). Gal mainly exists in the form of Galp-(1→ (14.18%) and →4)-Galp-(1→ (9.83%). Gal-UA mainly exists in the form of →4)-Galp-UA-(1→ (13.79%). Ara mainly exists in the form of →5)-Araf-(1→ (13.17%) and Araf-(1→ (8.53%). Structural analysis shows that the polysaccharide forms a main chain skeleton by alternating →4)-Galp-(1→ and →4)-Galp-UA-(1→, and there are branch structures composed of →5)-Araf-(1→ and rhamnose connection modes (→2)-, →2,4)-, →2,3)-).

[0071] Table 5 Methylation determination results of AES-S 5. NMR analysis AES-S was dissolved in D2O to prepare a polysaccharide solution with a concentration of 10 mg / mL. The dissolved solution was transferred to a nuclear magnetic tube, and the amount of 0.5 mL was added. The nuclear magnetic tube was placed in a nuclear magnetic resonance spectrometer for one-dimensional 1H spectrum analysis. The results show that the structure and composition of the polysaccharide are confirmed by multiple one-dimensional and two-dimensional nuclear magnetic resonance spectra, and are represented in the form of a graph. As shown in Fig. A of Figure 15 1 ​In the anomeric region of H NMR (4.3 ppm~5.5 ppm), in addition to the strong out-of-peak of 4.70 ppm of solvent heavy water protons, a plurality of groups of anomeric hydrogen H1 signals belonging to polysaccharide residues can be found, and after identification, the main out-of-peak positions are 5.15 ppm, 5.06 ppm, 4.99 ppm, 4.92 ppm, 4.53 ppm and 4.33 ppm, confirming that the polysaccharide is composed of multiple residues, and the main anomeric positions of the residues are all in α configuration according to the hydrogen spectrum. It is worth noting that the strong signals at 1.21 ppm and 1.15 ppm in the high field of the hydrogen spectrum are confirmed to be the methyl H6 corresponding to the two main rhamnose residues. Nuclear magnetic carbon spectrum 13 As shown by B in Figure 15 , in the range of 160 ppm~200 ppm, the carbonyl carbon signal is found to exist, and combined with the information of the methylated residues, it is judged that the position of 174.13 ppm is the C6 carbonyl corresponding to the 1→4 connected galacturonic acid α-1,4-GalA-(1→ residue. The anomeric region (90 ppm~110 ppm) of the carbon spectrum can identify a plurality of anomeric carbon C1 signals, which are 107.51 ppm, 107.09 ppm, 103.03 ppm, 98.64 ppm and 97.56 ppm, etc., confirming the diversity of the residues that constitute the polysaccharide; the signal at 61.09 ppm is attributed to the non-glycosidically linked residue ring outside methylene-CH2- carbon; the multiple signals at 65 ppm~85 ppm represent other carbon C2-C5 information in the ring and part of the carbon information that may form a glycosidic bond. The no-distortion polarization transfer enhancement spectrum as shown by C in Figure 15 verified the information of the carbon, and it is judged that the out-of-peak at 66.85 ppm is attributed to the C5 of 1,5-Araf forming a glycosidic bond. As shown by D in Figure 15 , the two-dimensional nuclear magnetic spectrum results show that 5.06 / 107.09 is the H1 / C1 signal of the end sugar α-t-Ara-(1→ of the arabinose residue, 5.00 / 107.51 is the H1 / C1 signal of α-1,5-Ara-(1→, 4.53 / 103.48 is the H1 / C1 signal of β-t-Gal-(1→, and 4.38 / 103.48 is the H1 / C1 signal of β-1,4-Gal-(1→. As shown by E in Figure 15 , in the nuclear magnetic heteronuclear single quantum coherence spectrum, the rhamnose α-1,2-Rha-(1→ residue existing in the structure is found to have a strong correlation signal of 5.15 / 77.39, which is judged to be the 1→4 connected glycosidic bond between α-1,2-Rha-(1→ and α-1,4-GalA-(1→, confirming that the two residues constitute the connection between different residues in the main chain; it is worth noting that the nuclear Overhauser effect correlation spectrum, such asFigure 15 As shown in G of FIG. 6, 4.92 / 4.01 was found in the strong signal, confirming the presence of 1→2 linked glycosidic bond of the residue α-1, 4-GalA-(1→ and α-1, 2-Rha-(1→ (or α-1, 2, 4-Rha-(1→), suggesting the presence of 1→2 linked glycosidic bond of the residue α-1, 4-GalA-(1→ and α-1, 2-Rha-(1→ (or α-1, 2, 4-Rha-(1→), thus judging the above alternating connection of galacturonic acid and rhamnose residues, forming the main structure in the main chain; in the branched structure, various residues of arabinose and galactose constitute the main part, in which the heteronuclear multiple bond correlation spectrum (such as Figure 15 As shown in F of FIG. 6, the signal 5.00 / 83.85 suggests the presence of 1→4 linked glycosidic bond of the residue α-1, 5-Ara-(1→ and α-1, 2, 4-Rha-(1→, judging that the 4 position of the rhamnose forms a branch; 4.38 / 3.94 NOESY signal can also be found, suggesting the presence of 1→4 linked glycosidic bond of the residue β-1, 4-Gal-(1→ and α-1, 2, 4-Rha-(1→. In the end sugar component part of the branched structure, various end sugar residue forms can be found, confirming the richness of the polysaccharide branch; among the residues with higher content, the correlation NOESY signal 5.05 / 3.80 of the arabinose end sugar α-t-Ara-(1→ anomeric proton indicates the correlation with the 5 position of α-t-Ara-(1→ and α-1, 5-Ara-(1→, confirming the presence of glycosidic bond connection of α-t-Ara-(1→ 5)-α-1, 5-Ara-(1→; the heteronuclear Overhauser effect correlation spectrum is as shown in Figure 15 As shown in G of FIG. 6, 4.53 / 3.34 signal suggests the presence of 1→4 connection of the end sugar β-t-Gal-(1→ residue of galactose and β-1, 4-Gal-(1→. Therefore, by comprehensively analyzing the above structure, combined with the total correlation spectrum, as shown in H of FIG. 6, the main structure of AES-S is obtained, which is speculated to be mainly composed of 1→4 and 1→2 cross-linked glycosidic bonds of the acidic galactose residue and the rhamnose residue, and the 4 position of part of the rhamnose residue is the starting point of the branch. The main component of the branch is arabinose and galactose, the main connection of arabinose is 1→5 glycosidic bond, and arabinose is the end sugar, and the other part is 1→4 glycosidic bond of galactose, and galactose is the end sugar. Combined with the overall analysis, the following repeating unit structure is obtained as shown in the following formula (1): Figure 14

[0072] Formula (1); ​R is α-t-Ara-(1→5)-α-Ara-(1→4)- or β-t-Gal-(1→4)-β-Gal-(1→4)-, wherein n represents the number of repeating units, and the molecular weight of the polysaccharide is 16.27 kDa, and the number of repeating units is a constant value.

[0073] 6. Determination of the molecular weight of AES-S The molecular weight of AES-S was determined, and the results are shown as C in Figure 14 Compared with AES, the molecular weight of AES-S decreased to 16.27 kDa, and thus AES-S can have better biological activity. The infrared spectrum of AES-D0.3 was analyzed, and the results are shown as D in Figure 14 After purification of AES, the core structure characteristics of AES did not change.

[0074] 7. Infrared spectrum detection of AES-S 150 mg of AES-S was reacted with salicylic acid (SA) and 4-dimethylaminopyridine (DMAP) in dimethyl sulfoxide solution at 45°C for 12 h to obtain an intermediate product. After dialysis and freeze-drying, AES-D0.3-SA was obtained. It was dissolved in a phosphate buffer solution (PBS; pH = 7.2) with N-hydroxysuccinimide (NHS) and reacted at 25°C for 15 min, and then the fluorescent dye Cy5.5 was added to the reaction, and the reaction was continued for 3 h. After dialysis of the mixture, freeze-drying was performed, and it was named AES-S-Cy5.5. The infrared spectrum of AES-S-Cy5.5 was detected in the same way as in the fourth step of the experiment, and the results are shown as A in Figure 16 Compared with AES-S, the core structure characteristics of AES-S-Cy5.5 did not change.

[0075] 8. Stability experiment of AES-S-Cy5.5 The stability was evaluated in in vitro gastric and intestinal digestion. The results are shown as B in Figure 16 The simulated gastrointestinal analysis of AES-S-Cy5.5 showed that the fluorescence purity was 91.54 ± 1.32% (PH = 1.8) and 92.34 ± 0.83% (PH = 7.5) after 24 h, respectively, indicating that the surface AES-S-Cy5.5 has good stability in gastric and intestinal juice.

[0076] Example 3, Lung targeting effect To investigate the in vivo distribution of AES-S-Cy5.5 in mice following oral administration, 50.0 mg of AES-S was reacted with 5.0 mg of SA and 2.0 mg of DMAP in a dimethyl sulfoxide (DMSO) solution at 45°C for 12 hours to yield an intermediate. After dialysis and freeze-drying, the lyophilized product, 5.0 mg of EDC, and 7.5 mg of NHS were dissolved in 2.0 mL of phosphate buffered saline (PBS; pH 7.2) and reacted at 25°C for 15 minutes. The Cy5.5 DMSO solution was then added, and the reaction continued for 3 hours. The mixture was dialyzed and lyophilized to yield the lyophilized product, AES-S-Cy5.5. Mice were anesthetized with isoflurane and gavage administered AES-S-Cy5.5 at a dose of 10 mg / mL. In vivo imaging was performed using an IVIS Lumina XR system at intervals of 1, 3, 6, 12, and 24 hours. The mice were then immediately sacrificed, and NIR imaging of the major organs, including the heart, liver, spleen, kidney, lung, and intestine, was performed. Compared with the control group, the fluorescence signal mainly accumulated in the abdomen of the experimental group mice. Quantification of the fluorescence intensity in the lungs confirmed that the fluorescence intensity reached a maximum at 6 hours and then gradually weakened with time ( Figure 17 To further elucidate the biodistribution of AES-S-Cy5.5 in mice, intestinal segments, heart, liver, spleen, lung, kidney, and pancreas were isolated and imaged ex vivo at pre-coordinated time intervals. Ex vivo intestinal imaging revealed that fluorescent signals were visible in all intestinal segments at early stages, indicating that each intestinal segment participated in the absorption of AES-S-Cy5.5 to varying degrees. Figure 17 (B) ROI values ​​demonstrate that fluorescence intensity reaches its maximum at 3 hours, with the duodenum showing the highest accumulation, followed by the jejunum and ileum. Notably, strong fluorescence signals were observed in the lungs between 3 and 24 hours, indicating that AES-S-Cy5.5 is well absorbed into the lungs. These findings suggest that AES-S-Cy5.5 can be absorbed into the bloodstream and almost completely metabolized within 24 hours, demonstrating the potential for the preparation of lung-targeted products using safflower polysaccharides.

[0077] Example 4: Experiment on LPS-induced acute lung injury model in mice To further evaluate the immune regulation and lung protection of AES-S, a mouse model of acute lung injury (ALI) induced by lipopolysaccharide was established. Sixty mice were randomly divided into a blank group, a lipopolysaccharide model group (LPS), a positive control group, and AES-S low-, medium-, and high-dose groups. The blood leukocyte parameters, including white blood cell (WBC), percentage of neutrophils (NE%), percentage of lymphocytes (LY%), platelet (PLT), and lung index (lung wet weight / body weight), were detected. The positive control group was administered 2 mg / kg of dexamethasone, the AES-S low-dose group was administered 50 mg / kg of AES-S, the AES-S medium-dose group was administered 100 mg / kg of AES-S, and the AES-S high-dose group was administered 200 mg / kg of AES-S. The results, as shown in Tables 6 and 7, showed that the WBC (18.4 ± 2.3 × 10 9 / L) and lung index (14.9 ± 1.3 mg / g) of the LPS model group were significantly increased (P < 0.01), and neutrophil infiltration (82.6 ± 6.4%) and lymphocyte reduction (12.8 ± 2.5%) were obvious; however, the AES-S high-dose group dose-dependently reversed the above changes, with WBC of 9.6 ±.3 × 10 9 / L, lung index of 7.6 ± 0.7 mg / g, NE of 42.1 ± 4.9%, and LY of 51.3 ± 5.2%, which was close to the effect of the positive control group. The results showed that AES-S can effectively inhibit LPS-induced lung inflammation and immune disorders.

[0078] Table 6 Effect on blood leukocyte count in mice Note: “#” indicates significant difference compared with the model group (“#” for p < 0.05, “##” for p < 0.01), and “*” indicates significant difference compared with the blank group (“*” for p < 0.05, “**” for p < 0.01).

[0079] Table 7 Effect on lung index Note: “#” indicates significant difference compared with the model group (“#” for p < 0.05, “##” for p < 0.01), and “*” indicates significant difference compared with the blank group (“*” for p < 0.05, “**” for p < 0.01).

[0080] Example 5: Immune regulation of AES-S on cyclophosphamide-mediated immunosuppressed mice The immunomodulatory effect of AES-S was studied using a cyclophosphamide (CTX)-induced immunosuppressive mouse model. Animal experimental design: The mice were randomly divided into 6 groups (n=8). They were divided into a blank group (CON), a model group (MOD), a positive drug group (LH), a low-dose administration group (AES-SL), and a high-dose administration group (AES-SH). Except for the blank group, the other groups were injected with CTX 80 mg / kg for 3 consecutive days, and the blank group was also given 0.9 g / 100 g of normal saline. After modeling, the LH group was given levamisole 40 mg / kg, the AES-SL group was given AES-S 200 mg / kg, and the AES-SH group was given AES-S 400 mg / kg. The blank group and the model group were given an equal amount of 0.9 g / 100 g of normal saline for 14 consecutive days by gavage. After gavage on the last day, the mice were fasted for 12 hours, weighed and killed, and the body weight change rate was calculated. The thymus of the mice was collected and weighed, and the thymus index was calculated. The results are as follows Figure 18 As shown in the results, the weight change rate of mice in the MOD group was significantly reduced compared with the CON group (P<0.05). Compared with the MOD group, the LH group and different doses of AES-S intervention significantly reversed the decrease in weight change rate caused by CTX (P<0.05), indicating that AES-S can improve the weight loss caused by cyclophosphamide. As a major immune organ, the thymus will change in size and mass when the immune system is damaged. Figure 19 As shown in the figure, compared with the thymic index of the CON group, the thymic index of the MOD group decreased significantly after CTX injection, indicating that cyclophosphamide severely damages the immune organs of mice. The thymic index of mice in the LH group and at different doses of AES-S showed a significant upward trend (P<0.05). These results indicate that AES-S can ameliorate CTX-induced damage to immune organs.

[0081] Example 6: Proportions of T lymphocyte subsets in mouse spleen Flow cytometry was used to analyze T lymphocyte subsets in mouse spleens. The antibodies used for spleen cell staining were CD8a (53-6.7) from PE Anti-Mouse, CD3e (145-2C11) from APC Anti-Mouse, and CD4 from PerCP-Cyanine5.5 Anti-Mouse. Figures 20-25As shown, in the case of immune system damage, CTX induced a significant decrease in the proportion of CD3 / CD4 and CD3 / CD8 cells and the CD4 / CD8 T cell ratio in the mouse spleen (P < 0.05). However, compared with the MOD group, the AES-S group significantly up-regulated the CD3 / CD4 / T cell, CD3 / CD8 / T cell proportion and CD4 / CD8 / T cell ratio (P < 0.05), and the effect of the high-dose group (400 mg / kg) was significantly better than that of the low-dose group (200 mg / kg) (P < 0.05). These results show that AES-S can reverse the decrease in T lymphocytes induced by CTX in mice.

[0082] While embodiments of the present application have been shown and described with reference to particular embodiments thereof, it will be understood by those of ordinary skill in the art that various changes in form and details can be made therein without departing from the spirit and scope of the present application as defined by the following claims and their equivalents.

Claims

1. A safflower polysaccharide for lung targeting, characterized in that: In terms of molar percentage, the proportions of the monosaccharides in the safflower polysaccharide are as follows: L-fucose 0.75%, arabinose 36.04%, rhamnose 25.03%, galactose 17.85%, glucose 4.26%, xylose 1.33%, mannose 0.61%, galacturonic acid 12.07%, glucuronic acid 2.06%, total 100%; The safflower polysaccharide has an alternating connection of galacturonic acid and rhamnose to form a main structural form in the main chain. In the structure of its repeating unit, the main chain is composed of acidic galactose and rhamnose with 1→4 and 1→2 cross-linked glycosidic bonds as the main components, the 4 position of rhamnose is the main starting point for forming a branch, and the main components of the branch are arabinose and galactose, part of the arabinose is connected with a 1→5 glycosidic bond, with arabinose as the terminal sugar, and the other part is connected with galactose with a 1→4 glycosidic bond, with galactose as the terminal sugar.

2. The safflower polysaccharide for lung targeting according to claim 1, characterized in that The molecular weight of the safflower polysaccharide is 16.27 kDa.

3. A method for preparing safflower polysaccharide for lung targeting according to claim 2, characterized in that: The following steps are involved: The safflower powder is mixed with alkali solution, extracted at 20-60°C for 30-120 minutes, centrifuged, and the supernatant is concentrated, deproteinized, and dialyzed to obtain crude safflower polysaccharide. The crude safflower polysaccharide was loaded onto a DEAE-FF agarose gel column and eluted with 0.3 mol / L NaCl. The eluate was collected, concentrated, dialyzed, and freeze-dried. The eluate was then purified using an S-400 gel column, concentrated, dialyzed, and freeze-dried to obtain the safflower polysaccharide.

4. The method for preparing safflower polysaccharide for lung targeting according to claim 3, characterized in that: The alkali solution is a sodium hydroxide solution with a concentration of 0.4M to 0.8M.

5. The method for preparing safflower polysaccharide for lung targeting according to claim 3, characterized in that: The specific process of purification by the S-400 gel column is as follows: Water was used as the eluent for elution at a flow rate of 0.5 mL / min, and the eluent was collected, concentrated, dialyzed, and freeze-dried to obtain the safflower polysaccharide.

6. The method for preparing safflower polysaccharide for lung targeting according to claim 3, characterized in that: The concentration refers to concentrating the supernatant to 1 / 3 to 1 / 5 of the original volume.

7. The method for preparing safflower polysaccharide for lung targeting according to claim 3, characterized in that: The reagent used for the alcohol precipitation is 95% ethanol by mass; the reagent used for the protein removal is Sevag reagent; and the dialysis uses a 7kDa~14kDa dialysis bag.

8. Use of the safflower polysaccharide for lung targeting according to claim 2 in preparing a lung targeting product, characterized in that: The product is an oral preparation, wherein the concentration of safflower polysaccharide in the oral preparation is 10 mg / mL to 12 mg / mL, and the solvent of the oral preparation is water.

9. A use of the safflower polysaccharide for lung targeting according to claim 2 in preparing a product for alleviating lung damage, characterized in that: The product is an injection, and the solvent of the injection is physiological saline.

10. Use of the safflower polysaccharide according to claim 2 in the preparation of an immunomodulatory product.

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