阅读说明：本技术 一种高聚原花青素的增溶方法及其制得的复合物和应用 (Solubilization method of high polymeric proanthocyanidins, compound prepared by solubilization method and application of compound ) 是由 李凯凯 张亚杰 李春美 侯焘 李晓芳 陈平 余颖 于 2020-12-12 设计创作，主要内容包括：本发明提供了一种高聚原花青素的增溶方法及其制得的复合物和应用；其增溶方法为,往含有高聚原花青素的水溶液中加入多糖类物质,或在将高聚原花青素溶解于水之前,加入多糖类物质,混合搅拌,即可。本发明所提供的高聚原花青素的增溶方法通过加入多糖类物质,通过多糖-多酚的相互作用,在一定条件下使高聚原花青素与多糖类物质在水溶液中形成稳定的复合微粒体系,使其溶解度提高了70倍左右；本发明所提供的高聚原花青素的增溶方法工艺简单,仅需按比例加入多糖类物质,搅拌混合即可,简单方便,实际应用效果优异；本发明所提供的高聚原花青素与多糖类物质复配形成的复合物具有显著的减肥降糖作用,且两者协同效果显著。(The invention provides a solubilization method of high polymeric proanthocyanidins, a compound prepared by the solubilization method and application of the solubilization method; the solubilization method comprises adding polysaccharide into water solution containing high polymer procyanidin, or adding polysaccharide before dissolving high polymer procyanidin in water, mixing, and stirring. The solubilization method of the high polymeric proanthocyanidins provided by the invention is characterized in that the polysaccharide substance is added, and the polysaccharide-polyphenol interaction is adopted, so that the high polymeric proanthocyanidins and the polysaccharide substance form a stable composite particle system in an aqueous solution under a certain condition, and the solubility of the high polymeric proanthocyanidins is improved by about 70 times; the solubilization method of the high polymeric proanthocyanidins provided by the invention has simple process, only needs to add the polysaccharide substances in proportion and stir and mix, is simple and convenient, and has excellent practical application effect; the compound formed by compounding the high polymeric proanthocyanidins and the polysaccharide substance provided by the invention has obvious weight-losing and blood sugar-reducing effects, and the synergistic effect of the two is obvious.)

1. A method for solubilizing high polymeric proanthocyanidin is characterized in that,

adding polysaccharide substance into water solution containing high polymer procyanidin, or adding polysaccharide substance before dissolving high polymer procyanidin in water; mixing and stirring to obtain the product.

2. The solubilization method according to claim 1,

the polysaccharide substance is one or more of pectin, carrageenan, sodium alginate, konjac glucomannan and inulin, preferably inulin.

3. The solubilization method according to claim 1,

the adding mass ratio of the polysaccharide substance to the high polymeric proanthocyanidin is 20-40: 1.

4. the solubilization method according to claim 1,

in the process of mixing and stirring, the mixture is stirred,

controlling the temperature to be 20-50 ℃, preferably 20-30 ℃;

and/or, the pH is controlled to be 4 to 9, preferably 4 to 7.

5. The solubilization method according to any one of claims 1 to 4,

the polysaccharide substance is inulin, and the solubility of the high polymeric procyanidin in the aqueous solution is improved from 0.15mg/ml to 10mg/ml, and the solubilization is 66.67 times.

6. A high polymeric proanthocyanidin complex characterized by,

is a compound of high polymeric proanthocyanidin and polysaccharide.

7. The procyanidin conjugate of claim 6,

the polysaccharide substance is one or more of pectin, carrageenan, sodium alginate, konjac glucomannan and inulin, preferably inulin.

8. The proanthocyanidin polyparagonide complex as claimed in claim 6 or 7,

the mass ratio of the polysaccharide substance to the high polymeric proanthocyanidin is 20-40: 1.

9. the use of the proanthocyanidin polyplexes of claim 6 for preparing hypoglycemic and hypolipidemic foods.

Technical Field

The invention relates to the technical field of dissolution of plant functional components, in particular to a solubilization method of high polymeric proanthocyanidins, a prepared compound and application thereof.

Background

Proanthocyanidins (PACs) are the most widely occurring polyphenols in fruits and vegetables, and are mainly present in most fruits and vegetables in the form of Polymerized Proanthocyanidins (PPACs).

At present, researches on procyanidine mainly focus on evaluating health effects and mechanisms of procyanidine, and it is proved that procyanidine has various health effects of resisting oxidation, inhibiting bacteria, diminishing inflammation, regulating and controlling glycolipid metabolism and the like, but few researches are carried out on how procyanidine is widely applied to a food system. The reason for this is mainly because procyanidins, especially high polymeric procyanidins, have poor water solubility and strong astringency, and easily interact with components such as proteins in food, thereby affecting the processing characteristics and nutritional functions of food. These properties of procyanidins, particularly polymeric procyanidins, severely hamper their use as food ingredients in food production and processing.

At present, aiming at high polymeric proanthocyanidins in fruits and vegetables, oligomeric proanthocyanidins are mainly prepared in a cracking mode to reduce the molecular weight and improve the solubility; however, these methods are accompanied by the characteristics of complicated process, low yield, high cost, and the like.

In conclusion, how to overcome the technical problem that the high polymeric procyanidins are difficult to dissolve is a key technical problem which needs to be solved urgently by the technical personnel in the field.

Disclosure of Invention

The invention aims to provide a method for solubilizing high polymeric proanthocyanidins, a compound prepared by the method and application of the compound.

In order to achieve the purpose, the invention adopts the following technical scheme:

a method for solubilizing polymeric procyanidin comprises adding polysaccharide to water solution containing polymeric procyanidin, or adding polysaccharide before dissolving polymeric procyanidin in water; mixing and stirring to obtain the product.

Specifically, in the technical scheme, the high polymeric proanthocyanidins form a stable composite particle system in the aqueous solution under the action of the polysaccharide substances, so that the solubility of the high polymeric proanthocyanidins can be greatly improved, and the preparation method is simple in process and convenient to operate.

Further, in the above technical scheme, the polysaccharide substance is one or more of pectin, carrageenan, sodium alginate, konjac glucomannan and inulin.

Preferably, in the above technical solution, the polysaccharide substance is inulin.

Further, in the above technical scheme, the adding mass ratio of the polysaccharide substance to the high polymeric proanthocyanidin is 20-40: 1.

further, in the technical scheme, the temperature is controlled to be 20-50 ℃ in the process of mixing and stirring.

Preferably, in the above technical scheme, the temperature is controlled to be 20-30 ℃ during the mixing and stirring process.

Further, in the technical scheme, the pH is controlled to be 4-9 in the process of mixing and stirring.

Preferably, in the above technical scheme, the pH is controlled to be 4-7 during the mixing and stirring process.

Specifically, in the technical scheme, the polysaccharide substance is inulin, and the solubility of the high polymeric procyanidin in the aqueous solution is improved from 0.15mg/ml to 10mg/ml, namely 66.67 times of solubilization.

The invention also provides a high polymeric proanthocyanidin compound, in particular a compound of the high polymeric proanthocyanidin and polysaccharide substances.

Specifically, in the above technical solution, in the high polymeric proanthocyanidin complex, the polysaccharide substance is one or more of pectin, carrageenan, sodium alginate, konjac glucomannan and inulin, and is preferably inulin.

Preferably, in the above technical solution, the mass ratio of the polysaccharide substance to the high polymeric proanthocyanidin is 20-40: 1.

the reagents and starting materials used in the present invention are commercially available.

The invention also provides the application of the high polymeric proanthocyanidin compound in preparing food and medicine for reducing blood sugar and lipid.

Compared with the prior art, the invention has the beneficial effects that:

(1) the solubilization method of the high polymeric proanthocyanidins provided by the invention is characterized in that the polysaccharide substance is added, and the polysaccharide-polyphenol interaction is adopted, so that the high polymeric proanthocyanidins and the polysaccharide substance form a stable composite particle system in an aqueous solution under a certain condition, and the solubility of the high polymeric proanthocyanidins is improved by about 70 times;

(2) the solubilization method of the high polymeric proanthocyanidins provided by the invention has a simple process, can complete the solubilization process of the high polymeric proanthocyanidins only by adding the polysaccharide substances in proportion and stirring and mixing, and is simple and convenient, and has excellent practical application effect;

(3) the compound formed by compounding the high polymeric proanthocyanidins and the polysaccharide substance provided by the invention has obvious weight-losing and blood sugar-reducing effects, and the synergistic effect of the two is obvious.

Drawings

FIG. 1 shows the structure of a polymeric procyanidin in an embodiment of the invention;

FIG. 2 is a transmission electron microscope image of high polymeric proanthocyanidin dissolved in water in an embodiment of the present invention;

FIG. 3 is a graph showing the results of solubilization of polymeric procyanidins by inulin aqueous solutions of different concentrations in the examples of the present invention;

FIG. 4 is a graph showing the particle size results of compound solutions at different high procyanidin concentrations in accordance with the examples of the invention;

FIG. 5 is a graph showing the particle size results of composite solutions of different concentrations of polymeric procyanidins at different pH's in accordance with an embodiment of the invention;

FIG. 6 is a graph of particle size results for composites of the present examples at various pH's;

FIG. 7 shows the results of weight tests on mice of different groups in the examples of the present invention;

FIG. 8 shows the results of adipose tissue testing of mice of different groups in an example of the present invention;

FIG. 9 shows the results of blood glucose tests of different groups of mice in the examples of the present invention;

FIG. 10 shows the results of liver and adipose tissue sections and HE staining of mice of different groups in the examples of the present invention.

Detailed Description

The present invention is further described in detail below with reference to specific examples so that those skilled in the art can more clearly understand the present invention.

The following examples are provided only for illustrating the present invention and are not intended to limit the scope of the present invention.

All other embodiments obtained by a person of ordinary skill in the art based on the embodiments of the present invention without any creative effort shall fall within the protection scope of the present invention.

In the embodiment of the invention, the high polymeric proanthocyanidins are self-made persimmon high polymeric proanthocyanidins, and the extraction method comprises the following steps:

50g of persimmon pulp, mixing with a 1% hydrochloric acid-methanol mixed solution in a weight ratio of 1: 8 (methanol 2000ml, hydrochloric acid 20ml), condensing and refluxing at 80 ℃ for 40min, repeating for three times, combining the extracting solutions, and standing overnight; suction filtering, rotary steaming at 35 deg.C, concentrating to 50ml (per 2000ml), suction filtering, loading onto AB-8 macroporous resin, adsorbing for 40min (sealing with appropriate water), washing with water until it is colorless (removing water soluble components such as soluble sugar, hydrochloric acid, and methanol, and no tannin loss during washing), washing with 10% ethanol until it is colorless (removing small molecular substances such as pigment), eluting with 90% ethanol, collecting eluate, concentrating at 35 deg.C, removing ethanol (rotary steaming), and lyophilizing to obtain persimmon tannin. Molecular structure analysis shows that the average polymerization degree is 26, and the proanthocyanidin is typical high polymeric proanthocyanidin.

The inulin is commercially available natural inulin.

Examples

The structure of the polymeric procyanidin is shown in figure 1.

Preparing 1mg/ml aqueous solution of persimmon high polymeric procyanidin, and observing the existence form of the aqueous solution of persimmon high polymeric procyanidin by adopting a transmission electron microscope.

The results are shown in FIG. 2, and it can be seen from FIG. 2 that the polymeric proanthocyanidins are substantially insoluble in water and are agglomerated; the water-insoluble characteristic of the water-soluble chitosan makes the water-soluble chitosan-chitosan copolymer difficult to be used as a food ingredient in food systems such as beverages.

Preparing 0.1, 0.2, 0.25, 0.3 and 0.4g/ml inulin aqueous solution respectively, adding 10mg/ml high polymeric procyanidin respectively, dissolving by vortex oscillation for 5min, determining the particle size at 25 +/-0.1 ℃ by using a Malvern Nano ZS laser nanometer particle size analyzer, and determining each sample for 3 times in parallel.

As shown in FIG. 3, it can be seen from FIG. 3 that the polymeric procyanidin-inulin forms stable nanoparticles, and the particle size of the polymeric procyanidin-inulin complex decreases and increases first and then as the inulin concentration increases, and is minimized at 0.25 g/ml. The particle size of the nano-composite is the reaction of existence state of the persimmon high polymeric proanthocyanidin-inulin composite, and generally, the smaller the particle size is, the higher the clarity of the composite is under the same condition is, and the more stable the composite is.

Preparing 0.25g/ml inulin water solution, dissolving by ultrasonic, adding 5, 10, 15, 20, 25 and 30mg/ml high polymeric procyanidin respectively, dissolving by vortex oscillation for 5min, determining the particle size at 25 +/-0.1 ℃ by using a Malvern Nano ZS laser nanometer particle size analyzer, and determining each sample for 3 times in parallel.

As shown in fig. 4, it can be seen from fig. 4 that the particle size of the polymeric procyanidin-inulin complex gradually increases as the concentration of the polymeric procyanidin increases; in the inulin solution of 0.25g/ml, the maximum solubility of the high polymeric proanthocyanidin is 10mg/ml, and after the maximum solubility is exceeded, the particle size of the system is gradually increased, and the whole complex system tends to be unstable.

Preparing 0.1, 0.2, 0.25, 0.3 and 0.4g/ml inulin solutions respectively, dissolving by ultrasonic, adding 10mg/ml polymeric procyanidin respectively, dissolving by vortex oscillation for 5min, diluting with distilled water for 2, 4, 8, 16 and 32 times in sequence, measuring the particle size by a Malvern Nano ZS laser nanometer particle size analyzer at 25 +/-0.1 ℃, and measuring each sample for 3 times in parallel. This experiment was used to investigate its stability under different dilution conditions.

The results are shown in fig. 5, and it can be seen from fig. 5 that the particle size of the formed stable complex solution does not change greatly under different dilution factors, which indicates that the polymeric procyanidin-inulin complex is relatively stable and the dilution has little influence on the particle size.

Preparing inulin solutions with the concentrations of 0.1, 0.2, 0.25, 0.3 and 0.4g/ml by water with the pH values of 2, 4, 7, 8 and 9 respectively, ultrasonically dissolving, adding 10mg/ml of high polymeric procyanidin respectively, ultrasonically dissolving for 5min, carrying out vortex oscillation for 15min, measuring the particle size by using a Malvern Nano ZS laser nanometer particle size analyzer at the temperature of 25 +/-0.1 ℃, and carrying out parallel measurement on each sample for 3 times.

As shown in fig. 6, it can be seen from fig. 6 that the high polymeric proanthocyanidin-inulin complex is greatly affected by the pH environment, the complex is easily aggregated under a strong acidic condition, the particle size of the complex becomes large, and the complex is precipitated, thereby lowering the stability. But the compound is stable in the environment with the pH value of 4-9, and can meet the requirements of different food environments.

Effect verification

A diet-induced obesity mouse model is selected to evaluate the blood glucose and lipid reducing efficacy of the compound, and the specific process is as follows.

About 20g of about 50C 57 mice were selected and randomly divided into five groups, including a control group, a model group, a high polymeric proanthocyanidin treatment group, an inulin treatment group and a high polymeric proanthocyanidin-inulin complex group, each of which contained 10 mice.

Wherein:

control group: feeding 12450J feed;

model group: feeding RD12492 high-fat feed;

high polymeric procyanidin treatment group: feeding RD12492 high fat feed and high polymeric procyanidin at a daily feeding amount of 20mg/kg (mouse body weight);

inulin treatment group: feeding RD12492 high-fat feed and inulin at a daily feeding amount of 500mg/kg (mouse body weight);

high polymeric procyanidin-inulin complex group: feeding RD12492 high fat feed and feeding high polymeric proanthocyanidin-inulin complex at a feeding amount of 20mg/kg high polymeric proanthocyanidin (mouse body weight) +500mg/kg inulin (mouse body weight) per day.

The treatment time was 8 weeks, and the animal body weight and feed were weighed weekly.

After the experiment was completed, blood, adipose tissue, liver and large intestine contents of the mice were collected and subjected to the next stage of experiment and analysis.

Mainly contains blood sugar, blood fat and weight of fat tissue. The contents of blood sugar and blood fat are measured by a kit method; adipose tissues were classified into epididymal fat pad, perirenal fat pad and inguinal fat pad, and after the mice were sacrificed under anesthesia, they were taken out and weighed to obtain the weights of the three fat pads.

The results are shown in FIGS. 7-10.

FIG. 7 shows the results of the weight test of mice in different groups in the examples of the present invention.

As can be seen from fig. 7, the group of high polymeric procyanidin-inulin complex can significantly inhibit the occurrence of high fat diet-induced obesity in mice, and there is a significant synergistic effect between the two.

FIG. 8 shows the results of adipose tissue testing of mice in different groups in the examples of the present invention.

As can be seen from fig. 8, the high fat diet significantly promoted the proliferation of the main adipose tissues (epididymal fat pad, perirenal fat pad and inguinal fat pad) of the mice, thereby inducing obesity, and the high polymeric procyanidine-inulin complex can significantly inhibit the proliferation of the adipose tissues.

FIG. 9 shows the results of blood glucose tests of different groups of mice in the examples of the present invention.

As can be seen from fig. 9, the blood sugar of the mice induced by high fat showed a significant increase, while the high polymeric procyanidin-inulin complex could significantly reduce the blood sugar of the mice induced by high fat diet; and simultaneously, the two show remarkable synergistic effect.

FIG. 10 shows the results of liver and adipose tissue sections and HE staining of mice of different groups in the examples of the present invention.

Fig. 10 shows that liver and adipose tissue sections show that high-fat diet can significantly induce the occurrence of fatty liver, liver HE staining finds that a large number of fatty bubbles appear in the liver tissue of the group induced by high-fat diet, and inulin-procyanidin complex treated group can significantly inhibit the occurrence of fatty bubbles, thereby inhibiting the occurrence of fatty liver; the adipose tissue section and HE staining also show that high-fat diet can obviously promote the growth of fat cells, the diameter of the fat cells is larger, and the high-polymer procyanidin-inulin complex has extremely strong improvement effect.

From fig. 7-10, it can be found that the inulin-proanthocyanidin polymer complex shows a strong synergistic effect, and can significantly improve obesity in mice induced by high fat diet.

Finally, the method of the present invention is only a preferred embodiment and is not intended to limit the scope of the present invention. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.