Sagittaria sagittifolia polysaccharide / vitamin C inclusion compound as well as preparation method and application thereof

Abstract

The invention belongs to the field of preparation and application of natural products, and particularly relates to a sagittaria sagittifolia polysaccharide / vitamin C (VC) inclusion compound and a preparation method and cosmetic application thereof. The sagittaria sagittifolia polysaccharide / VC clathrate compound prepared by the technical scheme is high in comprehensive score (72.93 + / -1.07%); the thermal stability of VC can be obviously improved (the temperature is 212-333 DEG C, and the weight loss rate is only 24.28%); the antioxidant activity of the sagittaria sagittifolia polysaccharide can be improved; in a dry environment, the moisture retention rate is 82.9 + / -2.35% within 48 hours, and the moisture retention rate is obviously higher than that of sagittaria sagittifolia polysaccharide and VC (Vitamin C). The sagittaria sagittifolia polysaccharide / VC clathrate compound prepared by the technical scheme can be used for preparing anti-oxidation and moisturizing cosmetics such as gels, cream, emulsions, film agents and the like.

Description

Technical Field

[0001] The invention belongs to the field of preparation and application of natural products, and specifically relates to a sagittaria polysaccharide / vitamin C inclusion compound and a preparation method thereof and application thereof in cosmetics. Background Art

[0002] Vitamin C (VC) is a water-soluble vitamin that plays an important role in biological redox and cellular respiration, and is widely involved in amino acid metabolism, collagen synthesis, etc. (Journal of Tianjin Medical University, 2001, 7(4):507-508). In addition, VC is also a recognized antioxidant and is widely used in the fields of medicine, food, cosmetics, etc. However, due to the presence of active dienol structures in VC molecules, it is easily oxidized and inactivated, making it less stable, especially unstable to heat (Food and Fermentation Industries, 2018, 44(12):60-65).

[0003] Sagittaria trifolia is a perennial aquatic herb of the genus Sagittaria in the family Alismataceae. It is mainly eaten as a vegetable in my country and is widely planted in the middle and lower reaches of the Yangtze River. Polysaccharides are the main active ingredients of Sagittaria, which have a wide range of pharmacological effects such as lowering blood sugar, enhancing immunity, and protecting the liver (Journal of the Science of Food and Agriculture, 2021, 101: 3085-3098). However, the antioxidant activity of Sagittaria polysaccharides is not high. As reported in the literature, at 5 mg / mL, the scavenging rate of Sagittaria polysaccharides for 1,1-diphenyl-2-trinitrophenylhydrazine (DPPH) free radicals is only 58.12±3.46 mg / mL (Chemistry&Biodiversity 2022, 19: e202200219).

[0004] The technology of encapsulating one molecule in the cavity structure of another molecule to form an inclusion complex is called inclusion technology (Trends in Food Science & Technology, 2021, 112: 667-675). Summary of the invention

[0005] The purpose of the present invention is to provide a preparation method and use of a sagittaria polysaccharide / VC inclusion compound. The inclusion compound prepared by the method can significantly improve the thermal stability of VC, improve the antioxidant activity of sagittaria polysaccharide, and has high moisturizing performance, and can be used in the preparation of antioxidant and moisturizing cosmetics.

[0006] According to the first aspect of the present invention, the present invention provides a sagittaria polysaccharide / VC inclusion complex, in which the content of sagittaria polysaccharide is higher than that of VC, and VC is encapsulated inside the sagittaria polysaccharide (VC is encapsulated inside the sagittaria polysaccharide through hydrogen bonding); the thermal stability of the inclusion complex is better than that of VC.

[0007] Preferably, the moisturizing property of the inclusion compound is better than that of Sagittaria sagittifolia polysaccharide and VC.

[0008] According to the second aspect of the present invention, the present invention provides a method for preparing a sagittaria polysaccharide / VC inclusion complex: VC and sagittaria polysaccharide are placed in a solvent, interacting with each other at a certain temperature and time, and after the reaction is completed, the mixture is allowed to stand, and the precipitate is collected by filtration under reduced pressure, and dried to obtain a sagittaria polysaccharide / VC inclusion complex; in the preparation method, the amount of sagittaria polysaccharide added is greater than the amount of VC added.

[0009] Preferably, the interaction is carried out in a microwave / ultrasonic combined extractor under ultrasonic mode.

[0010] Preferably, the solvent is water or a C1-6 alkyl alcohol, more preferably water.

[0011] Preferably, the preparation method of the Sagittaria polysaccharide / VC inclusion compound specifically comprises the following steps:

[0012] Sagittaria polysaccharide is added into distilled water, and dissolved by shaking in an ultrasonic cleaning machine to prepare a saturated solution (concentration is about 28 mg / mL), and then VC is slowly added according to a certain material ratio, and the solution is put into a microwave / ultrasonic combined extractor. Under the ultrasonic mode, the power is fixed at 300W, and the reaction is carried out at different temperatures for a certain time; after the reaction is completed, it is allowed to stand at 0-10°C for 2-48 hours, and the precipitate is collected by filtration under reduced pressure, and the precipitate is washed with a small amount of distilled water, and vacuum dried at 30-60°C to constant weight to obtain Sagittaria polysaccharide / VC inclusion complex.

[0013] Preferably, in the method, the material ratio of Sagittaria polysaccharide to VC is 3:1 to 8:1, and more preferably 7.5:1;

[0014] Preferably, the reaction temperature is 30-70°C, more preferably 50°C;

[0015] Preferably, the reaction time is 30 to 150 min, more preferably 124 min;

[0016] Preferably, the material ratio is 7.5:1, the reaction time is 124 min, and the reaction temperature is 50°C.

[0017] Preferably, the arrowhead polysaccharide is purified arrowhead polysaccharide; for example, it can be prepared by referring to the teachings in the prior art Chemistry & Biodiversity 2022, 19: e202200219 and patent CN113801248 B.

[0018] Under this condition, the comprehensive score of Sagittaria polysaccharide / vitamin C inclusion complex was 72.93±1.07%, with an encapsulation rate of 79.35±1.23% and a yield of 57.94±0.81%.

[0019] According to the third aspect of the present invention, the present invention provides a sagittaria polysaccharide / VC inclusion compound, which has a loose structure and a wide lamellar morphology; the sagittaria polysaccharide / VC inclusion compound has good thermal stability, and the weight loss rate at 212-333°C is only 24.28%, while the weight loss rate of VC is as high as 71.36%; the sagittaria polysaccharide / VC inclusion compound has good antioxidant activity, and the scavenging rate of superoxide anions at 0.5 mg / mL is 84.7±1.68%, the scavenging rate of DPPH free radicals at 20 μg / mL is 68.4±2.77%, the scavenging rate of hydroxyl free radicals at 0.4 mg / mL is 16.7±1.83%, and the reduction rate of Fe at 0.2 mg / mL is 0. 3+ The absorbance was 1.05±0.06, which was significantly higher than that of the same concentration of sagittaria polysaccharide (P<0.05 or P<0.01); the sagittaria polysaccharide / VC inclusion complex had excellent moisturizing properties. In a dry environment, the moisturizing rate was still as high as 82.9±2.35% after 48 hours, which was significantly higher than that of sagittaria polysaccharide (74.4±1.08%, P<0.01) and VC (43.3±1.71%, P<0.01).

[0020] According to the fourth aspect of the present invention, the present invention provides the use of the Sagittaria polysaccharide / VC inclusion complex in the preparation of antioxidant and moisturizing cosmetics; it can be used to prepare gels, creams, emulsions, films and other products.

[0021] The present invention uses inclusion technology to provide a technical solution for preparing a sagittaria polysaccharide / VC inclusion compound, so as to improve the stability of VC and the antioxidant activity of sagittaria polysaccharide, thereby achieving a "two-way improvement" effect. In addition, the present invention also provides an application of the sagittaria polysaccharide / VC inclusion compound in the preparation of cosmetics.

[0022] The present invention has the following beneficial effects:

[0023] ① The preparation method of the arrowhead polysaccharide / VC inclusion complex provided by the present invention has the advantages of simple operation and high comprehensive score, and has certain potential for industrial application.

[0024] ② The arrowhead polysaccharide / VC inclusion complex provided by the present invention has high thermal stability and has a good protective effect on structurally sensitive VC.

[0025] ③ The antioxidant activity of the arrowhead polysaccharide / VC inclusion complex provided by the present invention is significantly higher than that of arrowhead polysaccharide, which greatly improves the antioxidant effect of arrowhead polysaccharide.

[0026] ④ The arrowhead polysaccharide / VC inclusion compound provided by the present invention also has excellent moisturizing performance, which is significantly higher than that of arrowhead polysaccharide and VC.

[0027] In short, the arrowhead polysaccharide / VC inclusion complex prepared by the technical solution of the present invention can not only improve the stability of VC, but also improve the antioxidant activity of arrowhead polysaccharide, and has a "two-way enhancement" effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 .The impact of material ratio on comprehensive score. Different lowercase letters represent statistical differences.

[0029] Figure 2 .The effect of reaction time on the comprehensive score. Different lowercase letters indicate statistical differences.

[0030] Figure 3 .The effect of reaction temperature on the comprehensive score. Different lowercase letters indicate statistical differences.

[0031] Figure 4 .UV spectra of inclusion complex, arrowhead polysaccharide and VC;

[0032] Figure 5 .Infrared spectra of inclusion complex, arrowhead polysaccharide and VC;

[0033] Figure 6 .Differential scanning calorimetry curves of inclusion complex, arrowhead polysaccharide and VC;

[0034] Figure 7 .Scanning electron micrographs of VC (A), Sagittaria polysaccharide (B) and inclusion complex (C);

[0035] Figure 8 .Thermogravimetric curves of Sagittaria polysaccharide (A), VC (B) and inclusion complex (C);

[0036] Fig. 9 . The scavenging effects of inclusion complex, arrowhead polysaccharide and VC on superoxide anion free radicals. Different lowercase letters represent statistical differences within the groups; different uppercase letters represent differences between groups at the same concentration.

[0037] Fig.10. The scavenging effects of inclusion complex, Sagittaria polysaccharide and VC on DPPH free radicals. Different lowercase letters represent statistical differences within the groups; different uppercase letters represent differences between groups at the same concentration;

[0038] Fig.11 . The scavenging effects of inclusion complex, arrowhead polysaccharide and VC on hydroxyl free radicals. Different lowercase letters represent statistical differences within the groups; different uppercase letters represent differences between groups at the same concentration.

[0039] Fig.12 . Effects of inclusion complex, Sagittaria polysaccharide and VC on Fe 3+ Different lowercase letters represent statistical differences within the groups; different uppercase letters represent differences between groups at the same concentration;

[0040] Fig.13 .The moisturizing effects of inclusion complex, arrowhead polysaccharide and VC. Different lowercase letters represent statistical differences within the groups; different uppercase letters represent differences between groups at the same concentration. DETAILED DESCRIPTION

[0041] Arrowhead was supplied by the vegetable base of Southeast Development Zone, Changshu, Jiangsu Province, and the variety was Suzhou yellow arrowhead; vitamin C (VC, >99.0%), α-amylase (50 U / mg) and cellulase (10,000 U / g) were purchased from Shanghai MacLean Biochemical Technology Co., Ltd.; XH-300B microwave / ultrasonic combined extractor was provided by Beijing Xianghu Technology Development Co., Ltd.; UV spectrum was measured by T600B UV-visible spectrophotometer; infrared spectrum was measured by PerkinElmer Spotlight 200i Fourier transform infrared microscope system; differential scanning calorimetry (DSC) was measured by PerkinElmer DSC 8000 differential scanning calorimeter; thermogravimetry (TG) was measured by PerkinElmer TGA 8000 thermogravimetric analyzer; scanning electron microscopy (SEM) was measured by Regulus 8100HITACHI ultra-high resolution field emission scanning electron microscope; experimental data were expressed as mean ± SD (n = 3), and statistical test was performed using one-way analysis of variance (ANOVA).

[0042] Example 1 Preparation of Sagittaria polysaccharide

[0043] Reference Chemistry & Biodiversity 2022, 19: e202200219 and patent CN113801248B are used to extract and purify arrowhead polysaccharide.

[0044] The arrowhead was washed, sliced, dried at 60°C to constant weight, crushed and passed through a 60-mesh sieve, degreased with petroleum ether in the dark for 12 hours, and filtered under reduced pressure to obtain the arrowhead defatted powder. The arrowhead defatted powder was soaked in water at a solid-liquid ratio of 1:43 g / mL for 4 hours, the pH value was adjusted to 5.0, 2% (based on the weight of the arrowhead defatted powder, w / w) of compound enzyme (mass ratio α-amylase: cellulase = 1:1) was added, incubated in a shaker at 50°C for 2 hours, then inactivated in a boiling water bath for 15 minutes, and the pH value was adjusted to 7.0 after cooling. The enzymatic solution was placed in a microwave / ultrasonic combined extractor, and the extraction was continued at 506W for 8 minutes in microwave mode. After the extraction, it was filtered under reduced pressure, and the filtrate was concentrated to about 20% of the original volume, and then 3 times the volume of anhydrous ethanol was added to precipitate at 4°C for 12 hours, and the precipitate was collected by centrifugation. Redissolve the precipitate with distilled water, add 1 / 4 volume of Sevag reagent (chloroform: n-butanol = 4:1, v / v), mix and shake for 15 minutes and then centrifuge. Take the top clear liquid and repeat the above operation until no obvious protein is produced. The top clear liquid is freeze-dried to obtain the deproteinized polysaccharide. The deproteinized polysaccharide is prepared into a 10 mg / mL deionized water solution and loaded onto a diethylaminoethyl (DEAE) cellulose chromatography column (40×500mm); after the column is balanced, slowly elute with 0.5 mol / L NaCl solution, control the flow rate to about 1 mL / min, collect the eluate, concentrate to 1 / 4 of the original volume, dialyze with deionized water 3 times (each time interval of 12 hours, the dialysis bag has a molecular weight cutoff of 7000Da), and the dialyzate is freeze-dried to obtain the arrowhead polysaccharide. The phenol-sulfuric acid method was used, and the glucose standard curve (Y=11.669X+0.0163, R 2 =0.9982) to calculate the total sugar content of Sagittaria sagittifolia polysaccharide to be 77.04±2.51%.

[0045] Example 2 Preparation of Sagittaria polysaccharide / VC inclusion compound

[0046] 1.4g of arrowhead polysaccharide was added to 50mL of cold distilled water, and the solution was dissolved by shaking at 50℃ using an ultrasonic cleaner to prepare a saturated aqueous solution with a concentration of about 28mg / mL. Then VC was slowly added according to a certain material ratio, and the solution was placed in a microwave / ultrasonic combined extractor. Under the ultrasonic mode, the temperature was fixed at 300W, and the reaction was performed at different temperatures for a certain time. After the reaction was completed, the solution was allowed to stand at 4℃ for 24h, and the precipitate was collected by vacuum filtration, and the precipitate was washed with a small amount of distilled water, and vacuum dried at 35℃ to constant weight to obtain the arrowhead polysaccharide / VC inclusion complex.

[0047] The yield of the inclusion complex Y (%) = M1 / (M2+M3)×100 (Formula 1), wherein M1 is the mass (mg) of the sagittaria polysaccharide / VC inclusion complex, M2 is the mass (mg) of the sagittaria polysaccharide, and M3 is the mass (mg) of the input VC.

[0048] The VC content was determined by 2,6-dichloroindophenol titration, the detection wavelength was 243 nm, and the standard curve was Y = 0.0581X-0.0404, R 2 =0.9992.

[0049] 100 mg of Sagittaria polysaccharide / VC inclusion complex was dissolved in 1% (w / v) oxalic acid solution, stirred at room temperature for 3 h, and centrifuged at 4000×g for 15 min to obtain the supernatant. The VC content in the supernatant was calculated using the 2,6-dichloroindophenol titration method using the VC standard curve, and substituted into formula (2) to obtain the mass (m) of VC in the inclusion complex.

[0050] m=C×V×N (Formula 2), C is the VC content in the supernatant (mg / mL), V is the volume of the supernatant (mL), and N is the dilution multiple.

[0051] The encapsulation rate of the inclusion complex X (%) = m / M3 × 100 (Formula 3), wherein m is the mass of VC in the inclusion complex and M3 is the mass of the input VC.

[0052] The inclusion compound yield (Y) and encapsulation rate (X) were used as process evaluation indicators. Since the encapsulation rate is one of the most important indicators for evaluating the inclusion effect, a weight coefficient of 0.7 was assigned to the encapsulation rate and a weight coefficient of 0.3 was assigned to the yield, and the comprehensive score R (%) = 0.7X + 0.3Y.

[0053] 2.1 Single factor experiment

[0054] 2.1.1 Impact of material ratio on comprehensive score

[0055] When investigating the effect of material ratio on the comprehensive score, the material ratio of arrowhead polysaccharide to VC was changed (3:1-8:1 mg / mg), and other factors were kept constant. Figure 1 As shown in the data, within the range of 3:1 to 7:1 mg / mg, as the amount of arrowhead polysaccharide increased, the comprehensive score increased significantly (P<0.01); when the material ratio exceeded 7:1 mg / mg, as the amount of polysaccharide increased, the comprehensive score decreased significantly (P<0.01). Therefore, the appropriate material ratio of arrowhead polysaccharide to VC is 7:1 mg / mg.

[0056] 2.1.2 Effect of reaction time on comprehensive score

[0057] When investigating the effect of reaction time on the comprehensive score, the reaction time was changed (30 to 150 minutes) and other factors were kept constant. Figure 2 As shown in the data, within the range of 60 to 120 minutes, as the reaction time increased, the comprehensive score increased significantly (P<0.01); when the reaction time exceeded 120 minutes, as the reaction time increased, the comprehensive score decreased significantly (P<0.01). Therefore, the appropriate reaction time is 120 minutes.

[0058] 2.1.3 Effect of reaction temperature on comprehensive score

[0059] When investigating the effect of reaction temperature on the comprehensive score, the reaction temperature was changed (30-70°C) and other factors were kept constant. Figure 3 As shown in the results, within the range of 30-50℃, as the reaction temperature increases, the comprehensive score increases significantly (P<0.05 or P<0.01); when the reaction temperature exceeds 50℃, as the reaction temperature increases, the comprehensive score decreases significantly (P<0.01). Therefore, the appropriate reaction temperature is 50℃.

[0060] 1.2 Response surface optimization

[0061] According to the results of the single factor test, the material ratio is 7:1 mg / mg, the reaction time is 120 min, and the reaction temperature is 50 ° C. In the response surface optimization, three factors, namely material ratio A, reaction time B, and reaction temperature C, are selected as independent variables, and the comprehensive score R is used as the response value. The experimental factors and levels are shown in Table 1.

[0062] Table 1. Factors and levels of response surface optimization

[0063]

[0064] Using Design-Expert software, a three-factor three-level response surface optimization scheme was designed according to Box-Behnken Design (BBD). The results are shown in Table 2.

[0065] Table 2. Experimental plan and results of response surface optimization

[0066]

[0067] Using Design-Expert software BBD, after multiple regression fitting, the quadratic regression model equation of the predicted value of the comprehensive score R on the A, B, and C coding values ​​is obtained as follows:

[0068] R=70.85+9.21×A+1.89×B-0.2687×C+0.8675×AB+0.525×AC+2.23×BC-8.88×A 2 -8.24×B 2 -11.97×C 2 .

[0069] The results of one-way ANOVA of the regression model are shown in Table 3.

[0070] Table 3. One-way ANOVA of regression model

[0071]

[0072]

[0073] * P < 0.05; ** P<0.01; “--” means no difference.

[0074] As shown in Table 3, the P value of the model is less than 0.0001, indicating that the model is extremely significant; the lack of fit term is not significant (P>0.05), indicating that the model fits well and the unknown factors have little interference with the results. 2 =0.9912, indicating that 99.12% of the change in the response value comes from the selected variable; Adj R 2 =0.98, indicating that the model can explain 98% of the variability of the data. According to the F value, the influence of each factor on the comprehensive score is: A>B>C, that is, material ratio>reaction time>reaction temperature.

[0075] The optimization conditions obtained by Design-Expert software are: material ratio 7.52596:1mg / mg, reaction time 124.301min, reaction temperature 50.1412℃, under which the predicted comprehensive score is 73.4027%. Combined with the feasibility of actual operation, the above conditions were adjusted to: material ratio 7.5:1mg / mg, reaction time 124min, reaction temperature 50℃, under which the actual comprehensive score was 72.93±1.07% (relative error with the predicted value -0.64%), of which the inclusion encapsulation efficiency was 79.35±1.23% and the yield was 57.94±0.81%, further confirming that the use of this model to optimize the preparation process of Sagittaria polysaccharide / VC inclusion complex is effective and feasible.

[0076] Example 3 Characterization of Sagittaria polysaccharide / VC inclusion complex

[0077] 3.1 UV spectrum analysis

[0078] Sagittaria polysaccharide / VC inclusion complex, Sagittaria polysaccharide and VC were prepared into 5 mg / mL aqueous solutions respectively, and placed in an ultraviolet spectrophotometer for scanning in the scanning range of 200-800 nm.

[0079] like Figure 4 As shown, the characteristic absorption peak of VC at 265nm disappears after the formation of inclusion complex, while the inclusion complex has an ultraviolet absorption peak similar to that of Sagittaria polysaccharide, indicating that VC has been fully included by Sagittaria polysaccharide.

[0080] 3.2 Infrared spectroscopy analysis

[0081] The dried arrowhead polysaccharide / VC inclusion complex, arrowhead polysaccharide, VC and spectral grade potassium bromide were mixed evenly, pressed into tablets and placed in an infrared spectrometer for scanning in the scanning range of 4000-400 cm -1 .

[0082] like Figure 5 As shown in the infrared spectrum of VC, 3530 cm -1 、3411cm -1 、3315cm -1 and 3028cm -1 The absorption peak at 2917cm is caused by the stretching vibration of -OH in VC; -1 It is the stretching vibration of CH; 1755cm -1 It is the stretching vibration of -C=O; 1676cm -1 It is -C=C- stretching vibration. In the infrared spectrum of arrowhead polysaccharide, 3420cm -1 Represents the stretching vibration of -OH in Sagittaria polysaccharide; 2927cm -1 It is caused by CH stretching vibration; 1642cm -1 and 1400cm -1 The infrared spectrum of the inclusion complex is basically consistent with that of arrowhead polysaccharide, except for the 1755 cm -1 and 1676cm -1 Most of the characteristic peaks of VC were covered by Sagittaria polysaccharide, which further confirmed that VC was included in Sagittaria polysaccharide.

[0083] 3.2 Differential Scanning Calorimetry (DSC) Analysis

[0084] 1.287 mg of sample was accurately weighed and placed in an aluminum crucible. The sample was measured using a PerkinElmer DSC 8000 differential scanning calorimeter under nitrogen. The temperature range was 50-500° C., and the heating rate was 10° C. / min.

[0085] like Figure 6 As shown, at 191°C, the DSC curve of VC has a strong exothermic peak, which disappears after inclusion with Sagittaria polysaccharide, further indicating the successful formation of the inclusion complex.

[0086] 3.3 Scanning electron microscopy analysis

[0087] A small amount of the dried powder of Sagittaria polysaccharide / VC inclusion complex, Sagittaria polysaccharide and VC was respectively dipped with a cotton swab, pasted on a double-sided conductive adhesive stage, placed in an ion sputtering instrument, sprayed with gold under vacuum, and after processing, placed in a scanning electron microscope to observe the morphology.

[0088] like Figure 7As shown in Figure 2, at a magnification of ×600, VC exhibits a strip-shaped or diamond-shaped crystal morphology ( Figure 7 A); The surface structure of arrowhead polysaccharide is relatively loose, showing irregular block or flake morphology ( Figure 7 B); the inclusion complex has a loose structure and wide lamellar morphology. Although there are some small lumps and fine particles scattered, the crystal morphology of VC is no longer observed ( Figure 7 C), further indicating that the inclusion of VC by Sagittaria polysaccharide is not a simple physical stacking, but the formation of an inclusion complex that wraps VC inside the polysaccharide.

[0089] Example 4 Evaluation of the stability of Sagittaria polysaccharide / VC inclusion complex

[0090] Thermogravimetric analysis (TG) is a thermal analysis technique that measures the relationship between the mass and temperature change of the sample under program-controlled temperature. It is mainly used to evaluate the thermal stability of the sample.

[0091] 1.287 mg of sample was accurately weighed and placed in an aluminum crucible. The measurement was performed using a PerkinElmer TGA8000 thermogravimetric analyzer under nitrogen. The temperature range was 30-500°C and the heating rate was 10°C / min. The TG and differential thermogravimetric (DTG) hyperbola was plotted.

[0092] like Figure 8 As shown in A, there are two main weight loss steps for Sagittaria polysaccharide: 168-205℃, the weight loss rate is 9.89%; 280-329℃, the weight loss rate is 5.74%; at 500℃, the remaining amount of Sagittaria polysaccharide is 35.36%. Figure 8 As shown in B, VC has a main weight loss step: 198-225℃, the weight loss rate is 24.28%; at 500℃, the remaining VC is 18.14%. Figure 8 As shown in Figure C, the inclusion complex has a main weight loss step: 212-333°C, with a weight loss rate of 24.28%; while the weight loss rate of VC at 333°C is 71.36% ( Figure 8 B), indicating that the thermal stability of the inclusion complex is significantly higher than that of VC. At 500℃, the residual amount of the inclusion complex is 34.73% ( Figure 8 C), which was significantly higher than VC (18.14%) and close to that of Sagittaria polysaccharide (35.36%), further confirming that the thermal stability of the inclusion complex was higher than that of VC.

[0093] In summary, the inclusion of VC by arrowhead polysaccharide can significantly improve the thermal stability of VC, and the stability of the inclusion complex is almost close to that of arrowhead polysaccharide.

[0094] Example 5 Evaluation of the Antioxidant Activity of Sagittaria Polysaccharide / VC Inclusion Complex

[0095] Scavenging superoxide anion free radicals: Add 1 mL of arrowhead polysaccharide / VC inclusion complex aqueous solution (0.1-0.5 mg / mL) to 3 mL of Tris-HCl solution (50 mM, pH 8.2), mix well, let stand at room temperature for 10 min, add 1 mL of pyrogallol solution (25 mM), mix quickly and evenly, react at room temperature for 4 min, add 0.5 mL of HCl solution (10 M) to terminate the reaction, and measure the absorbance value at a wavelength of 320 nm. The same concentration of vitamin C (VC) and arrowhead polysaccharide were used as the control group. Formula (4) was used to calculate the scavenging rate of the sample for superoxide anion free radicals.

[0096] Superoxide anion radical scavenging rate (%) = [1-(A1-A2) / A0] × 100 Formula (4)

[0097] Wherein, A1 is the absorbance of the sample; A0 is the absorbance of the blank group using distilled water instead of the sample solution; A2 is the absorbance of the control group using distilled water instead of Tris-HCl and pyrogallol solution.

[0098] like Fig. 9 As shown, within the concentration range of 0.1-0.5 mg / mL, the scavenging effect of the inclusion complex on superoxide anion radicals increased significantly with increasing concentration (P<0.01), reaching 84.7±1.68% at 0.5 mg / mL, which was significantly higher than the same concentration of Sagittaria polysaccharide (P<0.01) and lower than VC (P<0.05).

[0099] Scavenging 1,1-diphenyl-2-trinitrophenylhydrazine (DPPH) free radicals: Add 2 mL of Sagittaria polysaccharide / VC inclusion complex aqueous solution (4-20 μg / mL) to 2 mL of DPPH-ethanol solution (0.1 mM), mix well, and stand at room temperature in the dark for 30 minutes, then measure the absorbance at 517 nm. Use VC and Sagittaria polysaccharide of the same concentration as the control group. Use formula (5) to calculate the scavenging rate of the sample for DPPH free radicals.

[0100] DPPH clearance rate (%) = [1-(A1-A2) / A0] × 100 Formula (5)

[0101] Wherein, A1 is the absorbance value of the sample; A2 is the absorbance value of the control group in which anhydrous ethanol is used instead of DPPH solution; A0 is the absorbance value of the blank group in which distilled water is used instead of the sample.

[0102] like Fig.10As shown in the results, within the concentration range of 4-20 μg / mL, the scavenging effect of the inclusion compound on DPPH free radicals increased significantly with the increase of concentration (P<0.01), reaching 68.4±2.77% at 20 μg / mL, which was significantly higher than the same concentration of arrowhead polysaccharide (P<0.01) and lower than VC (P<0.01). In addition, the scavenging effect of the inclusion compound on DPPH free radicals was also higher than that of arrowhead polysaccharide reported in the literature, and its IC 50 The value was 0.012±0.0006mg / mL, which was significantly lower than the IC value of arrowhead polysaccharide in scavenging DPPH free radicals. 50 value (4.04±0.18mg / mL) (Chemistry&Biodiversity, 2022, 19:e202200219).

[0103] Scavenging hydroxyl free radicals: 1 mL of Sagittaria polysaccharide / VC inclusion complex aqueous solution (0.08-0.4 mg / mL), 1 mL of ferrous sulfate solution (6 mM), and 1 mL of salicylic acid-ethanol solution (6 mM) were added to the test tube in sequence, mixed evenly, and then 1 mL of hydrogen peroxide solution (6 mM) was added. The mixture was reacted at 37°C for 30 min, and then the absorbance at 510 nm was measured. The same concentration of VC and Sagittaria polysaccharide was used as the control group. The scavenging rate of the sample for hydroxyl free radicals was calculated using formula (6).

[0104] Hydroxyl radical scavenging rate (%) = [1-(A1-A2) / A0] × 100 Formula (6)

[0105] Wherein, A1 is the absorbance of the sample; A2 is the absorbance of the control group with distilled water instead of hydrogen peroxide solution; A0 is the absorbance of the blank group with distilled water instead of the sample.

[0106] like Fig.11 As shown, in the range of 0.08-0.4 mg / mL, the scavenging effect of the inclusion complex on hydroxyl free radicals increased significantly with increasing concentration (P<0.05 or P<0.01), reaching 16.7±1.83% at 0.4 mg / mL, which was significantly higher than the same concentration of Sagittaria polysaccharide (P<0.05) and lower than VC (P<0.01).

[0107] Fe 3+Reducing power: 1 mL of Sagittaria polysaccharide / VC inclusion complex aqueous solution (0.04-0.2 mg / mL), 2.5 mL of phosphate buffer (200 Mm, pH 6.6), and 2.5 mL of potassium ferricyanide solution (1%, w / v) were added to a test tube in sequence, mixed evenly, reacted at 50°C for 20 min, and 2.5 mL of trichloroacetic acid solution (10%, w / v) was added to terminate the reaction, and then centrifuged at 3000 rpm / min for 10 min; 2.5 mL of supernatant was taken, 2.5 mL of distilled water and 0.5 mL of ferric chloride solution (0.1%, w / v) were added in sequence, mixed evenly, and then the absorbance at 700 nm was measured. The same concentration of VC and Sagittaria polysaccharide was used as the control group.

[0108] like Fig.12 As shown in the figure, within the concentration range of 0.04 to 0.2 mg / mL, the inclusion complex has a strong 3+ The reducing power increased significantly with the increase of concentration (P<0.01). The absorbance reached 1.05±0.06 at 0.2 mg / mL, which was significantly higher than that of Sagittaria polysaccharide at the same concentration (P<0.01) and lower than that of VC (P<0.01).

[0109] In summary, the antioxidant activity of Sagittaria polysaccharide / VC inclusion complex is significantly higher than that of Sagittaria polysaccharide, indicating that inclusion can significantly improve the antioxidant activity of Sagittaria polysaccharide; the antioxidant activity of Sagittaria polysaccharide / VC inclusion complex is lower than that of VC, further confirming that VC is fully included in Sagittaria polysaccharide.

[0110] Example 6 Evaluation of the Moisturizing Performance of Sagittaria Polysaccharide / VC Inclusion Complex

[0111] Weigh 2g of Sagittaria polysaccharide / VC inclusion complex and mix it with 2mL of distilled water, then put it into a weighing bottle and place it in a desiccator filled with color-changing silica gel. Weigh it after 2, 4, 6, 8, 12, 24, and 48 hours. The same concentration of VC and Sagittaria polysaccharide was used as the control group. The moisturizing rate was calculated using formula (7).

[0112] Moisture retention rate (%) = H n / H0×100 formula (7)

[0113] Where, H0 is the weight of water before placement (g), H n is the weight of water after standing (g).

[0114] like Fig.13 As shown in the data, with the extension of time, the moisturizing rates of the inclusion complex, sagittaria polysaccharide and VC gradually decreased, but the decrease of the inclusion complex was relatively slow, still as high as 82.9±2.35% at 48h, which was significantly higher than sagittaria polysaccharide (74.4±1.08%, P<0.01) and VC (43.3±1.71%, P<0.01).

[0115] The above results show that the moisturizing performance of Sagittaria polysaccharide / VC inclusion complex is significantly better than that of Sagittaria polysaccharide and VC, which may be related to its loose and wide surface morphology, which is more conducive to binding with water molecules.

[0116] Example 7 Preparation of Sagittaria polysaccharide / VC inclusion compound gel

[0117] 3 g of Carbomer-980 was added to 500 mL of deionized water and soaked for 12 hours to fully swell; 5 g of Sagittaria polysaccharide / VC inclusion compound was dissolved in 100 mL of deionized water, and slowly added to the fully swollen Carbomer-980 matrix under stirring, and then 5 mL of ethylparaben-ethanol solution (5%, w / v) and 2 mL of triethanolamine were added; the mixture was fully mixed to obtain a Sagittaria polysaccharide / VC inclusion compound gel.

[0118] Example 8 Preparation of Sagittaria polysaccharide / VC inclusion compound cream

[0119] Mix 8g of monostearate glyceryl, 6g of stearic acid, 5g of beeswax, 5g of liquid paraffin and 3g of olive oil under heating conditions to obtain phase A; add 3g of butylene glycol and 4g of glycerol to 100mL of pure water, mix well, add 3g of arrowhead polysaccharide / VC inclusion complex, 0.2g of xanthan gum and 50mg of EDTA-disodium, mix well to obtain phase B; add phase B to phase A while stirring, heat at 85°C for 10min, and then transfer to a mortar and grind to room temperature to obtain a arrowhead polysaccharide / VC inclusion complex cream.

[0120] Example 9 Preparation of Sagittaria polysaccharide / VC inclusion compound emulsion

[0121] 5 g of Sagittaria polysaccharide / VC inclusion complex, 2 g of hydroxyethylpiperazineethanesulfonic acid, 1 g of dipotassium glycyrrhizinate, and 50 mg of EDTA-disodium were dissolved in 100 mL of deionized water (phase A), and then slowly added into 50 mL of linseed oil (phase B); after homogenization at 20000 r / min for 10 min, a Sagittaria polysaccharide / VC inclusion complex emulsion was obtained.

[0122] Example 10 Preparation of Sagittaria polysaccharide / VC inclusion compound film

[0123] Dissolve 5 g of Sagittaria polysaccharide / VC inclusion complex in 100 mL of deionized water, add 5 g of konjac glucomannan, and stir thoroughly to swell to obtain phase A; dissolve 2 g of sorbitol and 6 g of glycerol in 100 mL of deionized water, and stir thoroughly to obtain phase B; while stirring, slowly add phase A to phase B, mix well to obtain Sagittaria polysaccharide / VC inclusion complex film.

[0124] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications made without departing from the spirit and principles of the present invention shall be equivalent replacement modes and shall be included in the protection scope of the present invention.

Claims

1. A sagittaria polysaccharide / VC inclusion complex, wherein the content of sagittaria polysaccharide in the inclusion complex is higher than that of VC, and VC is wrapped inside the sagittaria polysaccharide; and the thermal stability of the inclusion complex is better than that of VC.

2. The arrowhead polysaccharide / VC inclusion compound according to claim 1, wherein the arrowhead polysaccharide / VC inclusion compound has an infrared spectrum substantially as shown in FIG5 ; And / or, the Sagittaria polysaccharide / VC inclusion complex has antioxidant activity; And / or, the Sagittaria polysaccharide / VC inclusion compound has moisturizing properties.

3. The Sagittaria polysaccharide / VC inclusion compound according to claim 1 or 2, wherein the weight loss rate of the Sagittaria polysaccharide / VC inclusion compound is 24.28% at 212-333°C; And / or, the scavenging rate of superoxide anion at 0.5 mg / mL was 84.7±1.68%, the scavenging rate of DPPH free radical at 20 μg / mL was 68.4±2.77%, the scavenging rate of hydroxyl free radical at 0.4 mg / mL was 16.7±1.83%, and the scavenging rate of Fe reduction at 0.2 mg / mL was 1. 3+ The absorbance was 1.05 ± 0.06; And / or, the sagittaria polysaccharide / VC inclusion compound has a moisture retention rate of 82.9±2.35% in a dry environment for 48 hours.

4. A method for preparing a sagittaria polysaccharide / VC inclusion complex: VC and sagittaria polysaccharide are placed in a solvent and allowed to interact at a certain temperature and time. After the reaction is completed, the mixture is allowed to stand, the precipitate is collected by filtration, and the mixture is dried to obtain a sagittaria polysaccharide / VC inclusion complex. In the preparation method, the amount of sagittaria polysaccharide added is greater than the amount of VC added.

5. The method according to claim 4, wherein the interaction is carried out in a microwave / ultrasonic combined extractor under ultrasonic mode; And / or, the solvent is water or C1-6 alkyl alcohol, more preferably water.

6. According to the method of claim 4 or 5, the arrowhead polysaccharide is added to distilled water, and an ultrasonic cleaner is used to shake and dissolve it to prepare a saturated solution, and then VC is slowly added according to a certain material ratio, and the solution is put into a microwave / ultrasonic combined extractor, and reacted at different temperatures for a certain time under ultrasonic mode; after the reaction is completed, the solution is allowed to stand at 0-10°C for 2-48 hours, and the precipitate is collected by filtration, and the precipitate is washed with distilled water, and vacuum dried to constant weight to obtain the arrowhead polysaccharide / VC inclusion complex.

7. The method according to claim 6, wherein the material ratio of sagittaria polysaccharide to VC is 3:1 to 8:1, and more preferably 7.5:1; And / or, the reaction temperature is 30-70°C, more preferably 50°C; And / or, the reaction time is 30 to 150 min, more preferably 124 min; And / or, the material ratio is 7.5:1, the reaction time is 124min, and the reaction temperature is 50°C; And / or, the Sagittaria polysaccharide is purified Sagittaria polysaccharide.

8. Use of the Sagittaria polysaccharide / VC inclusion complex according to claim 1 or the Sagittaria polysaccharide / VC inclusion complex prepared by the method according to claim 4 in the preparation of antioxidant and moisturizing cosmetics.

9. The use according to claim 8, wherein the antioxidant and moisturizing cosmetic is a gel, cream, emulsion or film product.

10. The use according to claim 9, wherein the preparation method of the emulsion, cream, gel or film product is: 3g of Carbomer-980 was added to 500mL of deionized water and soaked for 12h to fully swell; 5g of Sagittaria polysaccharide / VC inclusion compound was dissolved in 100mL of deionized water, and slowly added to the fully swollen Carbomer-980 matrix under stirring, and then 5mL of ethylparaben-ethanol solution (5%, w / v) and 2mL of triethanolamine were added; the mixture was fully mixed to obtain Sagittaria polysaccharide / VC inclusion compound gel; 8 g of glyceryl monostearate, 6 g of stearic acid, 5 g of beeswax, 5 g of liquid paraffin, and 3 g of olive oil were mixed under heating conditions to obtain phase A; 3 g of butylene glycol and 4 g of glycerol were added to 100 mL of pure water, and mixed evenly, and then 3 g of arrowhead polysaccharide / VC inclusion compound, 0.2 g of xanthan gum and 50 mg of EDTA-disodium were added, and mixed evenly to obtain phase B; phase B was added to phase A while stirring, heated at 85° C. for 10 min, and then transferred to a mortar and ground to room temperature to obtain arrowhead polysaccharide / VC inclusion compound cream; 5 g of Sagittaria polysaccharide / VC inclusion compound, 2 g of hydroxyethylpiperazineethanesulfonic acid, 1 g of dipotassium glycyrrhizinate, and 50 mg of EDTA-disodium were dissolved in 100 mL of deionized water (phase A), and then slowly added into 50 mL of linseed oil (phase B); homogenized at 20000 r / min for 10 min to obtain Sagittaria polysaccharide / VC inclusion compound emulsion; Dissolve 5 g of Sagittaria polysaccharide / VC inclusion complex in 100 mL of deionized water, add 5 g of konjac glucomannan, and stir thoroughly to swell to obtain phase A; dissolve 2 g of sorbitol and 6 g of glycerol in 100 mL of deionized water, and stir thoroughly to obtain phase B; while stirring, slowly add phase A to phase B, mix well to obtain Sagittaria polysaccharide / VC inclusion complex film.

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