Soybean seed coat polysaccharide zinc chelate and preparation method thereof

Abstract

The invention discloses a soybean seed coat polysaccharide zinc chelate which is prepared by taking soybean seed coat polysaccharide and zinc sulfate as raw materials in a system with the pH value of 5.5-6.5 in the presence of sodium citrate. The invention discloses a preparation method of a soybean seed coat polysaccharide zinc chelate, which comprises the following steps: dissolving soybean seed coat polysaccharide in water to obtain a soybean seed coat polysaccharide solution, and adjusting the pH value to 6.0-6.2; the method comprises the following steps: dissolving zinc sulfate and sodium citrate in water to obtain a mixed solution of zinc sulfate and sodium citrate, and adjusting the pH value to 5.5-6.0; dropwise adding a mixed solution of zinc sulfate and sodium citrate into the soybean seed coat polysaccharide solution while stirring, continuously stirring after dropwise adding, and maintaining the pH value at 5.5-6.5; and dialyzing in pure water, centrifuging, and freeze-drying supernate to obtain the soybean polysaccharide zinc chelate. The sodium citrate is introduced, so that the irreversible gelation phenomenon caused by excessive zinc at the moment that the zinc-containing compound solution reacts with the polysaccharide is avoided.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology and relates to a soybean seed coat polysaccharide zinc chelate, its preparation method and antioxidant application. Technical Background

[0002] Zinc, as one of the essential trace mineral elements for the human body, plays a vital role in metabolism, growth and development, and immune function. However, the human body cannot synthesize zinc on its own and can only maintain zinc homeostasis through diet or zinc supplements. Traditional zinc supplements, such as zinc sulfate and zinc oxide, have significant drawbacks, including low bioavailability, strong gastrointestinal irritation, and susceptibility to dietary interference. These limitations have prompted researchers to seek safer and more efficient forms of zinc supplementation. In recent years, an increasing number of studies have confirmed that natural polysaccharides, due to their excellent biocompatibility, can serve as ideal organic ligands for zinc supplements. Compared to traditional zinc supplements, natural polysaccharide zinc has advantages such as good biocompatibility, low toxicity, and mild gastrointestinal irritation, enabling it to maintain relevant physiological functions more efficiently and safely.

[0003] Soybean seed coat polysaccharide is an acidic high-molecular-weight polysaccharide extracted and purified from soybean seed coat. Its chemical composition mainly includes galacturonic acid, galactose, rhamnose, and arabinose. Soybean seed coat polysaccharide possesses various physiological functions, including immunomodulation, hypoglycemia, antioxidant effects, and liver protection. Particularly in its intestinal probiotic function, it can provide energy for intestinal microbial metabolism, lower intestinal pH to some extent, and significantly increase the abundance of Proteobacteria and Firmicutes. As a major byproduct of soybean processing, soybean seed coat is abundant and inexpensive. Developing novel zinc supplements using it as the main raw material could not only increase the added value of soybean processing byproducts but also provide new options for the zinc supplement market.

[0004] Zn 2+ It coordinates with active groups such as hydroxyl and carboxyl groups in polysaccharide molecules to form a stable complex, which not only improves the stability and absorption rate of zinc, but also achieves the complementarity of polysaccharides and zinc in physiological functions. Summary of the Invention

[0005] The purpose of this invention is to provide a zinc chelate, which is a zinc chelate of soybean seed coat polysaccharide extracted from soybean seed coat, a by-product of soybean processing, and zinc sulfate. This zinc chelate exhibits no gelation and has high stability.

[0006] To achieve the above objectives, the present invention adopts the following technical solution:

[0007] A zinc chelate of soybean seed coat polysaccharide is prepared by using soybean seed coat polysaccharide and zinc sulfate as raw materials in the presence of sodium citrate in a system with a pH of 5.5 to 6.5.

[0008] Preferably, the soybean seed coat polysaccharide zinc chelate is prepared by the following method: a soybean seed coat polysaccharide solution with a pH of 6.0-6.2 and a mixed solution of zinc sulfate and sodium citrate with a pH of 5.5-6.0 are prepared respectively. The mixed solution of zinc sulfate and sodium citrate is added dropwise to the soybean seed coat polysaccharide solution. The soybean seed coat polysaccharide zinc chelate is formed in a system with a pH of 5.5-6.5. Then, the solution is dialyzed in pure water until the conductivity of the dialysate (pure water) is <5 μm / cm. After centrifugation, the supernatant is freeze-dried to obtain the soybean polysaccharide zinc chelate.

[0009] More preferably, the soybean seed coat polysaccharide zinc chelate is prepared by the following method: a soybean seed coat polysaccharide solution with a pH of 6.0-6.2 and a mixed solution of zinc sulfate and sodium citrate with a pH of 5.5-6.0 are prepared respectively. The mixed solution of zinc sulfate and sodium citrate is added dropwise to the soybean seed coat polysaccharide solution. The soybean seed coat polysaccharide zinc chelate is formed in the system with a pH of 6.0-6.2. Then, the solution is dialyzed in pure water until the conductivity of the dialysate (pure water) is <5 μm / cm. After centrifugation, the supernatant is freeze-dried to obtain the soybean polysaccharide zinc chelate.

[0010] A method for preparing the soybean seed coat polysaccharide zinc chelate includes the following steps:

[0011] Step (1): Dissolve soybean seed coat polysaccharide in water, centrifuge, discard the precipitate, and obtain soybean seed coat polysaccharide solution; adjust the pH of soybean seed coat polysaccharide solution to 6.0-6.2;

[0012] Step (2): Dissolve zinc sulfate and sodium citrate in water to obtain a mixed solution of zinc sulfate and sodium citrate, and adjust the pH of the mixed solution of zinc sulfate and sodium citrate to 5.5-6.0;

[0013] Step (3): At room temperature, while stirring, slowly add the mixed solution of zinc sulfate and sodium citrate obtained in step (2) to the soybean seed coat polysaccharide solution obtained in step (1). After the addition is complete, continue stirring for 1.5 to 2 hours, and maintain the pH at 5.5 to 6.5 throughout the reaction process (during the addition process and the stirring process after the addition is complete) to obtain the reaction solution.

[0014] Step (4): Dialyze the reaction solution obtained in step (3) in pure water at a temperature of 4-8℃. Replace the dialysate (pure water) every 6 hours until the conductivity of the dialysate is <5μm / cm. After dialysis, centrifuge the dialysate, take the supernatant, freeze dry it, and obtain soybean polysaccharide zinc chelate.

[0015] In step (1), the ratio of soybean seed coat polysaccharide to water is 0.25:100 to 1.25:100 g / mL, preferably 0.5 g:100 mL.

[0016] The centrifugation speed was 3000 r / min, and the centrifugation time was 5 min.

[0017] Preferably, the pH of the soybean seed coat polysaccharide solution is adjusted to 6.0–6.2 using 0.1 mol / L NaOH solution and / or 0.1 mol / L HCl.

[0018] In step (2), the mass ratio of soybean seed coat polysaccharide to zinc sulfate is 0.25:0.12 to 1.25:0.12, preferably 0.25:0.12 to 0.75:0.12, and more preferably 0.5:0.12.

[0019] The mass ratio of zinc sulfate to sodium citrate is 1.2:1 to 1.5:1, preferably 1.2:1.

[0020] The concentration of zinc sulfate solution in the mixed solution of zinc sulfate and sodium citrate is 4.8 mg / mL.

[0021] Preferably, the pH of the mixed solution of zinc sulfate and sodium citrate is adjusted to 5.5–6.0 using 0.1 mol / L NaOH and / or 0.1 mol / L HCl.

[0022] In step (3), the stirring speed is 600 r / min. The stirring can be magnetic stirring.

[0023] The dropping rate of the mixed solution of zinc sulfate and sodium citrate is at least slower than 5 s / drop, preferably 5 to 10 s / drop.

[0024] Preferably, the pH is maintained at 6.0 to 6.2 throughout the reaction process.

[0025] Preferably, the pH of the reaction system is maintained using 0.1 mol / L NaOH solution and / or 0.1 mol / L HCl.

[0026] In step (4), preferably, the dialysis temperature is 4°C.

[0027] The molecular weight cutoff of the dialysis bag is 35 kDa.

[0028] Generally, the dialysis time is 48–72 hours, during which the conductivity of the dialysate is <5 μm / cm, indicating that small molecules such as sodium ions and sulfate ions have been removed.

[0029] The centrifugation speed was 3000 r / min, the centrifugation time was 3 min, and the centrifugation temperature was 25℃.

[0030] The soybean seed coat polysaccharide is prepared by the following method, including the following steps:

[0031] Step (1): Crush the soybean seed coat to obtain soybean seed coat powder;

[0032] Step (2): Mix soybean seed coat powder and 1-2% ethanol solution at a material-to-liquid ratio of 1:10 to 1:15 g / mL, stir at room temperature to decolorize, filter, and dry to obtain decolorized soybean seed coat powder;

[0033] Step (3): Mix the decolorized soybean seed coat powder with sodium citrate and distilled water according to a mass ratio of 50:2 to 50:6 and a material-to-liquid ratio of 1:3 to 1:10 g / mL, and perform microwave pretreatment. After microwave pretreatment, add distilled water according to a material-to-liquid ratio of 1:15 to 1:25 g / mL, and perform water bath extraction to obtain the extract. The microwave power of the microwave pretreatment is 160 to 640 W, and the microwave pretreatment time is 10 to 20 min. The water bath extraction temperature is 80 to 90 °C, and the water bath extraction time is 145 to 240 min.

[0034] Step (4): Filter the extracted liquid, centrifuge the filtrate, concentrate the supernatant to 1 / 3 to 1 / 4 of the original volume to obtain a concentrated solution, adjust the pH of the concentrated solution to 3.0 to 5.0 with hydrochloric acid, add double the amount of anhydrous ethanol, and carry out alcohol precipitation at a temperature of 0 to 4℃. Dry the precipitate to obtain soybean seed coat polysaccharide.

[0035] The application of the soybean seed coat polysaccharide zinc chelate described in this invention in the preparation of zinc supplements.

[0036] The present invention relates to the application of the soybean seed coat polysaccharide zinc chelate in the preparation of drugs for diseases caused by oxidative damage or health foods that help with antioxidation.

[0037] The room temperature described in this invention is 25±5℃.

[0038] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0039] Soybean seed coat polysaccharide is an acidic high-molecular-weight polysaccharide containing numerous active groups such as hydroxyl and carboxyl groups. In preliminary experiments, the inventors used a traditional synthesis method to directly react soybean seed coat polysaccharide with a zinc-containing compound to prepare a soybean seed coat polysaccharide-zinc chelate. As an acidic polysaccharide (pH=3.0-4.0), soybean seed coat polysaccharide reacts with Zn... 2+Chelation reactions are often vigorous and can lead to gelation, reducing solubility. This invention differs from traditional polysaccharide zinc synthesis methods by introducing sodium citrate, effectively avoiding irreversible gelation caused by excess zinc during the reaction of zinc-containing compound solutions with polysaccharides. Zinc sulfate first reacts with sodium citrate to generate an intermediate. This intermediate, under suitable pH conditions, chelates more gently with soybean seed coat polysaccharides. The entire preparation process is simple and easy to operate. The resulting soybean seed coat polysaccharide zinc chelate not only retains the solubility and bioavailability of soybean seed coat polysaccharides, increasing the dual health benefits of the polysaccharide zinc chelate, but also ensures the bioavailability of zinc in the soybean seed coat polysaccharide zinc chelate.

[0040] The inventors investigated the stability of soybean polysaccharide zinc chelates during dialysis at room temperature and lower temperatures, finding that the soybean polysaccharide zinc chelates may gel within a short time during dialysis at room temperature. This invention reduces the gelation phenomenon of soybean polysaccharide zinc chelates during dialysis at lower temperatures. Attached Figure Description

[0041] Figure 1 Infrared spectra of soybean seed coat polysaccharide and soybean seed coat polysaccharide zinc chelate.

[0042] Figure 2 XPS images of soybean seed coat polysaccharide and soybean seed coat polysaccharide zinc chelate; where a: full spectrum, b: SHP1s, c: O1s, d: Zn Auger peak, e: SHP-ZnC1s, f: O1s, g: Zn2p peak.

[0043] Figure 3 XRD patterns of soybean seed coat polysaccharide and soybean seed coat polysaccharide zinc chelate.

[0044] Figure 4 Electron micrographs of soybean seed coat polysaccharide (a, scale bar 500 nm) and soybean seed coat polysaccharide zinc chelate (b, scale bar 500 nm).

[0045] Figure 5 Atomic force diagrams of soybean seed coat polysaccharide and soybean seed coat polysaccharide zinc chelate are shown; where a: SHP2D, b: SHP-Zn, c: SHP3D, d: SHP-Zn3D.

[0046] Figure 6 The degree of gelation of the products of Example 2 and Comparative Example 1.

[0047] Figure 7 The in vitro DPPH free radical scavenging activity of soybean seed coat polysaccharide and soybean seed coat polysaccharide zinc chelate was evaluated. Detailed Implementation

[0048] The present invention will be further explained below with reference to specific embodiments, but the embodiments do not limit the present invention in any way.

[0049] Table 1: Composition and content of soybean seed coat (mass fraction, %)

[0050]

[0051] Soybean seed coat polysaccharide (SHP) was prepared according to Example 3 of invention CN 117924534 A, specifically including the following steps:

[0052] S1. Remove impurities from soybean seed coats, crush them, and collect the solid powder that passes through a 60-mesh sieve;

[0053] S2. Weigh a certain amount of sieved soybean seed coat powder, add 1% ethanol solution for decolorization, the ratio of soybean seed coat powder to ethanol solution is 1:10 g / mL, stir at room temperature for 30 min, filter, dry to obtain decolorized soybean seed coat powder.

[0054] S3. Take 50g of decolorized soybean seed coat powder obtained from S2 treatment, add 6g of sodium citrate, add distilled water at a material-liquid ratio of 1:5g / mL and microwave treatment, microwave power 320W, microwave time 20min, to obtain pretreated material solution.

[0055] S4. Cool the pretreated solution, add distilled water to make the solution-to-solid ratio 1:20 g / mL, and extract in an 85℃ water bath for 4 hours to obtain the extract.

[0056] S5. Cool the extracted liquid, filter it, centrifuge the filtrate at 4000 r / min for 10 min, and take the supernatant; evaporate the supernatant by rotary evaporation to 1 / 3 of its original volume, and adjust the pH value to 4.0 with dilute hydrochloric acid;

[0057] S6. Slowly add double the volume of anhydrous ethanol to S5 and stir continuously to precipitate the ethanol. Place at 4°C for 12 hours, centrifuge to obtain the precipitate, freeze dry, and obtain soybean seed coat polysaccharide.

[0058] Soybean seed coat polysaccharide contains 46.59% galacturonic acid, 17.95% galactose, 14.77% rhamnose and 13.97% arabinose.

[0059] Infrared spectrum of soybean seed coat polysaccharides ( Figure 1 The following characteristic peak can be observed: 3263 cm⁻¹ -1 (-OH), 1735cm -1 (-CHO), 1595cm -1 (-C=O), 1143cm -1 (-COC), 1095cm -1 1016cm -1 (-COH), 956cm -1 (-CH). 1143cm-1 1095cm -1 1016cm -1 The peak represents the characteristic absorption peak of the pyran ring.

[0060] Example 1

[0061] I. Effects of pH under different reaction environments on zinc chelates of soybean seed coat polysaccharides

[0062] Weigh 0.5g of soybean seed coat polysaccharide, dissolve it in 100mL of pure water, centrifuge at 3000r / min for 5min, discard the precipitate, and adjust the pH to 6.1 using 0.1 mol / L NaOH solution (hereinafter referred to as Solution A).

[0063] Weigh 0.12g of zinc sulfate heptahydrate, dissolve the zinc sulfate heptahydrate in 25mL of pure water to obtain a zinc sulfate heptahydrate solution, and adjust the pH to 5.8 using 0.1mol / L HCl (hereinafter referred to as solution B).

[0064] At room temperature, while stirring (magnetic stirring at 600 r / min), slowly add solution B to solution A, with a dropping rate of at least 5 s / drop. After the dropping is completed, continue stirring for 1.5 minutes. Monitor the pH throughout the process and adjust the pH to 3, 4, 5, 6, and 7 using 0.1 mol / L HCl to obtain the reaction solution.

[0065] The reaction solution was transferred to a dialysis bag (dialysis molecular weight cutoff of 35 kDa). The reaction solution was dialyzed in pure water at 4°C. The dialysate (pure water) was replaced every 6 hours. After 72 hours of dialysis, the conductivity of the dialysate was <5 μm / cm. The dialysate was centrifuged (3000 r / min, 3 min, 25°C) and freeze-dried to obtain soybean seed coat polysaccharide zinc chelate.

[0066] Table 2: Effect of pH on the zinc content of soybean seed coat polysaccharide zinc chelate

[0067]

[0068] Observation of appearance changes and zinc content revealed that soybean seed coat polysaccharide zinc chelates prepared at different pH levels all exhibited varying degrees of gelation. The soybean seed coat polysaccharide zinc chelate prepared at pH 6 showed less gelation, while those prepared at pH 3, 4, and 5 had higher zinc content. However, the increased chelation rate was accompanied by increased gelation, resulting in the loss of zinc's inherent solubility and thus reduced utilization value. The soybean seed coat polysaccharide zinc chelate obtained at pH 7 contained a large amount of Zn(OH)₂ precipitate, rendering it useless.

[0069] Based on the above, in order to avoid gelation, the inventors attempted to introduce sodium citrate.

[0070] II. Effects of Different Soybean Seed Coat Polysaccharide Addition Amounts on Soybean Seed Coat Polysaccharide Zinc Chelate

[0071] Weigh out 0.25, 0.75, 1.0 and 1.25 g of soybean seed coat polysaccharide respectively, dissolve the soybean seed coat polysaccharide in 100 mL of pure water, centrifuge at 3000 r / min for 5 min, discard the precipitate, and adjust the pH of the soybean seed coat polysaccharide solution to 6.1 using 0.1 mol / L NaOH solution (hereinafter referred to as solution A).

[0072] Weigh 0.12g of zinc sulfate heptahydrate and 0.1g of sodium citrate. Dissolve the zinc sulfate heptahydrate and sodium citrate in 25mL of pure water to obtain a mixed solution of zinc sulfate heptahydrate and sodium citrate. Adjust the pH to 5.8 using 0.1mol / L HCl (hereinafter referred to as solution B).

[0073] At room temperature (25℃), while stirring (magnetic stirring at 600 r / min), solution B was slowly added dropwise to solution A. The dropwise addition rate of solution B was at least 5 s / drop. After the addition was completed, the reaction was stirred continuously for 1.5 h. The pH was monitored throughout the process, and the pH was adjusted to 6.1 using 0.1 mol / L HCl to obtain the reaction solution.

[0074] The reaction solution was transferred to a dialysis bag (dialysis molecular weight cutoff of 35 kDa) and dialyzed in pure water at 4°C. The dialysate (pure water) was replaced every 6 hours. After 72 hours of dialysis, the conductivity of the dialysate was <5 μm / cm. The dialysate was centrifuged (3000 r / min, 3 min, 25°C) and freeze-dried to obtain soybean seed coat polysaccharide zinc chelate (denoted as SHP-Zn).

[0075] Table 3: Effect of soybean seed coat polysaccharide dosage on the zinc content of soybean seed coat polysaccharide zinc chelate

[0076]

[0077] Table 3 shows the effect of the mass ratio of SHP to zinc sulfate on the zinc content in SHP-Zn. The zinc content in SHP-Zn first increases and then decreases with increasing soybean seed coat polysaccharide mass, reaching a maximum of 43.03 mg / g at a mass ratio of 0.5:0.12. At higher mass ratios, the zinc content decreases significantly, leading to raw material waste; at lower mass ratios, the zinc content increases. 2+ At excessively high concentrations, the electrostatic interaction between SHP and zinc ions increases, and the coordinating groups such as hydroxyl and carboxyl groups in SHP interact more with Zn. 2+ Reduced contact between them results in a lower zinc content in SHP-Zn.

[0078] III. Effects of Different Zinc Sulfate Addition Amounts on Zinc Chelates of Soybean Seed Coarse Polysaccharides

[0079] Weigh 0.5g of soybean seed coat polysaccharide, dissolve it in 100mL of pure water, centrifuge at 3000r / min for 5min, discard the precipitate, and adjust the pH to 6.1 using 0.1 mol / L NaOH solution (hereinafter referred to as Solution A).

[0080] Weigh 0.12g, 0.15g of zinc sulfate heptahydrate and 0.1g of sodium citrate. Dissolve zinc sulfate heptahydrate and sodium citrate in 25mL of pure water to obtain a mixed solution of zinc sulfate heptahydrate and sodium citrate. Adjust the pH to 5.8 using 0.1mol / L HCl (hereinafter referred to as solution B).

[0081] At room temperature, while stirring (magnetic stirring at 600 r / min), slowly add solution B dropwise to solution A. The dropwise addition rate should be at least 5 s / drop. After the addition is complete, continue stirring for 1.5 minutes. Monitor the pH throughout the process and adjust it to 6.1 using 0.1 mol / L HCl to obtain the reaction solution.

[0082] The reaction solution was transferred to a dialysis bag (dialysis molecular weight cutoff of 35 kDa) and dialyzed in pure water at 4°C. The dialysate (pure water) was replaced every 6 hours. After 72 hours of dialysis, the conductivity of the dialysate was <5 μm / cm. The dialysate was centrifuged (3000 r / min, 3 min, 25°C) and freeze-dried to obtain soybean seed coat polysaccharide zinc chelate with zinc contents of 43.03 and 35.26 mg / g.

[0083] Table 4: Effect of zinc sulfate dosage on zinc content of soybean seed coat polysaccharide zinc chelate

[0084]

[0085] The effect of the mass ratio of zinc sulfate to sodium citrate on the zinc content in SHP-Zn is shown in Table 4. When the mass ratio of zinc sulfate to sodium citrate is 1.2:1, the zinc content in SHP-Zn is 43.03 mg / g; when the mass ratio is 1.5:1, the zinc content is 35.26 mg / g. This may be because when the mass ratio of zinc sulfate to sodium citrate is 1.2:1, the citrate concentration is higher, making the reaction system more stable, and the coordinating groups in SHP interact more effectively with Zn. 2+ Increased contact leads to increased zinc content.

[0086] Example 2

[0087] Weigh 0.5g of soybean seed coat polysaccharide and dissolve it in 100mL of pure water. Centrifuge at 3000r / min for 5min, discard the precipitate, and adjust the pH of the soybean seed coat polysaccharide to 6.1 using 0.1M NaOH solution (hereinafter referred to as Solution A). Weigh 0.12g of zinc sulfate heptahydrate and 0.1g of sodium citrate, dissolve them in 25mL of pure water to obtain a mixed solution of zinc sulfate heptahydrate and sodium citrate. Adjust the pH of the mixed solution to 5.8 using 0.1mol / L HCl (hereinafter referred to as Solution B). At room temperature, slowly add Solution B dropwise to Solution A while stirring (magnetic stirring at 600r / min), with a dropping rate of at least 5s / drop. After the addition is complete, continue stirring for 1.5h to obtain the reaction solution. Monitor the pH throughout the process and adjust it to 6.1 using 0.1mol / L NaOH solution and / or 0.1mol / L HCl. The reaction solution was transferred to a dialysis bag (dialysis molecular weight cutoff of 35 kDa) and dialyzed in pure water at 4°C to remove small molecules such as sodium ions and sulfate ions. The dialysis fluid (pure water) was replaced every 6 hours. Dialysis was performed for 72 hours to reach the dialysis endpoint (conductivity of the osmotic fluid < 5 μm / cm). The dialysis solution was centrifuged (3000 r / min, 3 min, 25°C) and freeze-dried to obtain soybean seed coat polysaccharide zinc chelate (denoted as SHP-Zn, white flocculent solid) with a zinc content of 43.05 mg / g.

[0088] Figure 1 The infrared spectrum of soybean seed coat polysaccharide zinc chelate shows a characteristic peak at 3305 cm⁻¹. -1 (-OH), 1739cm -1 (-CHO), 1608cm -1 (-C=O), 1143cm -1 (-COC), 1095cm -1 1016cm -1 (-COH), 958cm -1 (-CH). 1143cm -1 1095cm -1 1016cm -1 The peaks are characteristic absorption peaks of the pyran ring. This indicates that after soybean seed coat polysaccharide is chelated with zinc, the spectral shape of the soybean seed coat polysaccharide and its zinc chelate is roughly the same, and some major absorption peaks show a significant red shift, proving that Zn... 2+ It coordinated with soybean seed coat polysaccharides but did not alter the basic structure or functional groups of the polysaccharides. The maximum absorption peak of the OH group was observed at 3263 cm⁻¹ for soybean seed coat polysaccharides. -1 Moved to 3305 cm of its zinc chelate -1 Furthermore, the weakening of the stretching vibration intensity indicates the formation of a coordinate bond between OH and zinc. At 1739 cm⁻¹-1 and 1608cm -1 The absorption peak at 1735 cm⁻¹ belongs to the stretching vibration of C=O, compared to the peak at 1735 cm⁻¹ for soybean seed coat polysaccharides. -1 and 1595cm -1 The absorption peak of the zinc chelate showed a significant red shift, indicating that the C=O group may interact with Zn. 2+ Coordination makes Zn 2+ Electrostatic interaction occurs with soybean seed coat polysaccharides; at 1143 cm⁻¹ -1 and 1095cm -1 1016cm- 1 The absorption peaks at this location belong to the stretching vibrations of COC and COH. Compared to soybean seed coat polysaccharides, the zinc chelates show a significant decrease in absorption peaks at multiple locations, indicating that Zn... 2+ Coordination bonds may have formed with these groups, leading to a weakening of the absorption peaks. These changes in peak shape prove that zinc has been successfully chelated with soybean seed coat polysaccharide, resulting in soybean seed coat polysaccharide zinc chelate.

[0089] To determine the valence state of Zn in the samples, X-ray photoelectron spectroscopy was used to perform a full-spectrum scan of the SHP and SHP-Zn samples. The results are as follows: Figure 2 As shown, the C / O ratio of SHP is approximately 1.4, and that of SHP-Zn is approximately 1.5. In addition to the C1S and O1S peaks, the X-ray photoelectron spectrum of SHP-Zn also shows Zn2p3 and Zn2p1 peaks at binding energies of approximately 1021.79 eV and 1044.13 eV, respectively. The 2p spectrum fitting results for SHP-Zn with a zinc content of 0.47% Zn show the following... Figure 2 The binding energy difference between the main peaks of Zn2p1 / 2 and Zn2p3 / 2 is 23.06 eV, proving that zinc in SHP-Zn mainly exists as Zn. 2+ exist, Figure 2 g is the Auger peak spectrum of ZnLM2, showing typical Auger transition peak positions characteristic of zinc, further proving that zinc is mainly produced in Zn. 2+ It exists in SHP-Zn.

[0090] SHP and SHP-Zn samples were scanned using X-ray diffraction, with a scanning range of 2θ from 5 to 40°. The results are shown in [Figure number missing]. Figure 3SHP exhibits distinct "bun-like" peaks at 2θ of 13.17° and 21.86°, typical of polysaccharides, indicating that SHP has an amorphous or non-crystalline structure. Compared to SHP, the SHP-Zn diffraction characteristics change significantly. The SHP diffraction peak at 2θ of 13.17° disappears, and the 21.86° diffraction peak shifts to 21.54° with a significantly broadened intensity. Furthermore, typical metallic zinc salts (31.77°, 15.5°, 19.8°) are not observed. This confirms that Zn... 2+ The SHP-Zn complex coordinates with the hydroxyl or carboxyl groups in SHP, and does not exist in the form of an inorganic salt, which is consistent with the XPS results. SHP-Zn exhibits a typical amorphous structure, which is beneficial to improving the original solubility and bioavailability of SHP.

[0091] The morphological characteristics of SHP and SHP-Zn under scanning electron microscopy are shown in the figure. Figure 4 Under high magnification, the surface of SHP appears as smooth, tightly packed sheets. Figure 4 a) indicates the presence of hydrogen bonds between SHP molecules. Compared to SHP, SHP-Zn exhibits a significant morphological change, displaying a rough, irregularly shaped aggregate of nanoparticles. Figure 4 b) The reason may be Zn 2+ Coordination with hydroxyl or carboxyl groups in SHP shortens the distance between polysaccharide molecular chains and causes the molecular chains to coil and stretch, further verifying the chelation of SHP-Zn.

[0092] SHP and SHP-Zn differ in molecular size and shape. SHP exhibits a uniform overall distribution and shows no aggregation. Figure 5 a, c). The surface of SHP-Zn is noticeably rough and uneven. Figure 5 b) and d) explain the introduction of Zn 2+ Subsequently, the aggregation behavior of SHP-Zn molecules changed, with enhanced aggregation and a significant increase in molecular size, which may be attributed to Zn. 2+ Coordination with hydroxyl or carboxyl groups in SHP leads to cross-linking between polysaccharide molecular chains, resulting in distinct nanoparticle aggregates. This result validates the SEM analysis findings.

[0093] Figure 6 The image shows the appearance of SHP-Zn. In Example 2, sodium citrate was added, and the sample appeared as a homogeneous water sample with no obvious precipitate or suspended matter. In Comparative Example 1, no sodium citrate was added, and the sample appeared heterogeneous, with gel formation and layering during synthesis. Sodium citrate acts as a buffer during synthesis, forming an intermediate complex with zinc sulfate, slowly releasing Zn... 2+ Release from binding sites on polysaccharide molecular chains, preventing Zn 2+A strong reaction with SHP leads to gel formation. Simultaneously, citrate ions may enhance the stability of the system through electrostatic repulsion, preventing excessive aggregation of SHP-Zn and subsequent gelation. Visual comparison confirms that the introduction of sodium citrate effectively improves the water solubility and stability of SHP-Zn, thereby enhancing its bioavailability.

[0094] Example 3

[0095] In vitro DPPH free radical scavenging activity of soybean seed coat polysaccharide and soybean seed coat polysaccharide zinc chelate

[0096] Preparation of sample solutions: Soybean seed coat polysaccharide solutions and soybean seed coat polysaccharide zinc chelate solutions (Example 2) with concentrations of 2, 4, 6, 8 and 10 mg / mL were prepared with distilled water, as well as strong reducing agent vitamin C (Vc) solutions with concentrations of 0.01, 0.02, 0.03, 0.04 and 0.05 mg / mL.

[0097] Take 1 mL of sample solution and vortex mix it with 0.4 mL of 0.2 mmol / L DPPH solution and 1.6 mL of distilled water to make a total volume of 3 mL. Place the mixture in the dark and incubate at room temperature for 30 min. Measure the OD value at 517 nm (denoted as A2). The OD value at 517 nm measured under the same conditions using the sample solution without DPPH (with the DPPH solution replaced by an equal volume of distilled water) is A0, and the OD value at 517 nm measured under the same conditions using DPPH without sample solution is A1. Calculate the DPPH free radical scavenging rate using the following formula:

[0098]

[0099] like Figure 7 As shown, the results indicate that the DPPH free radical scavenging ability increases with increasing concentrations of soybean seed coat polysaccharide and its zinc chelate. Compared to soybean seed coat polysaccharide, the chelate exhibits superior antioxidant capacity. At a concentration of 10 mg / mL, the DPPH free radical scavenging rate of soybean seed coat polysaccharide was 86.17%. The introduction of Zn... 2+ Subsequently, the DPPH free radical scavenging rate of its chelate increased to 89.68%. This may be due to the synergistic effect of the antioxidant capacity of polysaccharides and zinc ions, which enhances the antioxidant capacity of the chelate.

[0100] Comparative Example 1

[0101] The difference between Comparative Example 1 and Example 2 is that sodium citrate was not added; all other parameters and steps were the same as in Example 2. During the experiment, solution A and solution B gelled immediately after mixing, and the gel was insoluble. After low-temperature ultrasonic treatment (temperature 4°C, ultrasonic power 500W) for 5 minutes, the liquid appeared as a uniform milky white, but after being placed at 4°C for 1 hour, obvious solid-liquid separation occurred (see...). Figure 6 The freeze-dried product is white and flaky, with a zinc content of 86.41 mg / g, and it has lost the original solubility of soybean seed coat polysaccharides.

[0102] The above detailed description is a specific description of one feasible embodiment of the present invention. This embodiment is not intended to limit the patent scope of the present invention. All equivalent implementations or modifications that do not depart from the present invention should be included in the scope of the technical solution of the present invention.

Claims

1. A soybean seed coat polysaccharide zinc chelate, characterized in that: It is a chelate prepared from soybean seed coat polysaccharide and zinc sulfate in the presence of sodium citrate in a pH system of 5.5–6.

5.

2. The soybean seed coat polysaccharide zinc chelate according to claim 1, characterized in that: The soybean seed coat polysaccharide zinc chelate was prepared by the following method: a soybean seed coat polysaccharide solution with a pH of 6.0-6.2 and a mixed solution of zinc sulfate and sodium citrate with a pH of 5.5-6.0 were prepared respectively. The mixed solution of zinc sulfate and sodium citrate was added dropwise to the soybean seed coat polysaccharide solution. The soybean seed coat polysaccharide zinc chelate was formed in a system with a pH of 5.5-6.5, preferably 6.0-6.

2. The solution was then dialyzed in pure water until the conductivity of the dialysate was <5 μm / cm. After centrifugation, the supernatant was freeze-dried to obtain the soybean polysaccharide zinc chelate.

3. A method for preparing the soybean seed coat polysaccharide zinc chelate according to claim 1, characterized in that: Includes the following steps: Step (1): Dissolve soybean seed coat polysaccharide in water, centrifuge, discard the precipitate, and obtain soybean seed coat polysaccharide solution; adjust the pH of soybean seed coat polysaccharide solution to 6.0-6.2; Step (2): Dissolve zinc sulfate and sodium citrate in water to obtain a mixed solution of zinc sulfate and sodium citrate, and adjust the pH of the mixed solution of zinc sulfate and sodium citrate to 5.5-6.0; Step (3): At room temperature, while stirring, add the mixed solution of zinc sulfate and sodium citrate obtained in step (2) dropwise to the soybean seed coat polysaccharide solution obtained in step (1). After the addition is complete, continue stirring for 1.5 to 2 hours, and maintain the pH at 5.5 to 6.5 throughout the reaction process to obtain the reaction solution. Step (4): Dialyze the reaction solution obtained in step (3) in pure water at a temperature of 4-8℃. Replace the dialysate every 6 hours until the conductivity of the dialysate is <5μm / cm. After dialysis, centrifuge the dialysate, take the supernatant, freeze dry it, and obtain soybean polysaccharide zinc chelate.

4. The method for preparing the soybean seed coat polysaccharide zinc chelate according to claim 3, characterized in that: In step (1), the ratio of soybean seed coat polysaccharide to water is 0.25:100 to 1.25:100 g / mL, preferably 0.5 g:100 mL.

5. The method for preparing the soybean seed coat polysaccharide zinc chelate according to claim 3, characterized in that: In step (2), the mass ratio of soybean seed coat polysaccharide to zinc sulfate is 0.25:0.12 to 1.25:0.12, preferably 0.25:0.12 to 0.75:0.12, and more preferably 0.5:0.

12.

6. The method for preparing the soybean seed coat polysaccharide zinc chelate according to claim 3, characterized in that: In step (2), the mass ratio of zinc sulfate to sodium citrate is 1.2:1 to 1.5:1, preferably 1.2:

1.

7. The method for preparing the soybean seed coat polysaccharide zinc chelate according to claim 3, characterized in that: In step (2), the pH is maintained at 6.0 to 6.2 throughout the reaction process.

8. The method for preparing the soybean seed coat polysaccharide zinc chelate according to claim 3, characterized in that: In step (4), the dialysis temperature is 4°C.

9. The method for preparing the soybean seed coat polysaccharide zinc chelate according to claim 3, characterized in that: The molecular weight cutoff of the dialysis bag is 35 kDa.

10. The application of the soybean seed coat polysaccharide zinc chelate according to claim 1 in the preparation of zinc supplements, in the preparation of drugs for diseases caused by oxidative damage, or in health foods that help with antioxidation.

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