CN112655873B - Solubilization method of high polymer procyanidine, compound prepared by solubilization method and application of high polymer procyanidine - Google Patents

Abstract

The invention provides a solubilization method of high polymer procyanidine, a compound prepared by the solubilization method and application of the high polymer procyanidine; the solubilization method comprises adding polysaccharides into aqueous solution containing high polymer procyanidin, or adding polysaccharides before dissolving high polymer procyanidin in water, mixing, and stirring. According to the solubilization method of the high polymer procyanidine, provided by the invention, through adding the polysaccharide substance and through the interaction of polysaccharide-polyphenol, the high polymer procyanidine and the polysaccharide substance form a stable composite particle system in an aqueous solution under a certain condition, so that the solubility of the high polymer procyanidine and the polysaccharide substance is improved by about 70 times; the solubilization method of the high polymer procyanidine provided by the invention has simple process, is simple and convenient, and has excellent practical application effect by only adding polysaccharide substances in proportion and stirring and mixing; the compound formed by compounding the high polymer procyanidine and the polysaccharide substance has obvious weight-losing and blood sugar-reducing effects, and the synergistic effect of the high polymer procyanidine and the polysaccharide substance is obvious.

Description

A method for solubilizing polymeric proanthocyanidins and its prepared complex and application

Technical field

The present invention relates to the technical field of dissolving plant functional components, and more specifically, to a method for solubilizing polymeric proanthocyanidins and the composites and applications thereof.

Background technique

Proanthocyanidins (PACs) are the most widespread type of polyphenols found in fruits and vegetables. They mainly exist in the form of polymerized proanthocyanidins (PPACs) in most fruits and vegetables.

At present, research on proanthocyanidins mainly focuses on the evaluation of their health effects and mechanisms. It has been confirmed that proanthocyanidins have many health effects such as antioxidant, antibacterial, anti-inflammatory, and regulation of glucose and lipid metabolism. However, how to widely apply proanthocyanidins in food system, there are fewer studies. The reason is mainly because proanthocyanidins, especially polymerized proanthocyanidins, have poor water solubility, strong astringency, and can easily interact with protein and other components in food, thus affecting the processing characteristics and nutritional functions of food. These properties of proanthocyanidins, especially polymeric proanthocyanidins, seriously hinder their application as food ingredients in food production and processing.

Currently, for high-polymeric proanthocyanidins in fruits and vegetables, oligomeric proanthocyanidins are mainly prepared by cracking to reduce their molecular weight and improve solubility; however, these methods are often accompanied by complex processes, low yields, and high costs.

In summary, how to overcome the technical problem of insolubility of polymeric proanthocyanidins is a key technical problem that those skilled in the art urgently need to solve.

Contents of the invention

The object of the present invention is to provide a method for solubilizing polymeric proanthocyanidins and the composites prepared therefrom and their applications.

In order to achieve the above objects, the present invention adopts the following technical solutions:

A method for solubilizing high-polymerized procyanidins includes adding polysaccharides to an aqueous solution containing high-polymerized proanthocyanidins, or adding polysaccharides before dissolving the high-polymerized proanthocyanidins in water; mixing and stirring.

Specifically, in the above technical solution, the high polymerized proanthocyanidins form a stable composite particle system in the aqueous solution under the action of polysaccharides, which can greatly improve its solubility, and the process is simple and easy to operate.

Further, in the above technical solution, the polysaccharide material is one or more of pectin, carrageenan, sodium alginate, konjac gum and inulin.

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

Further, in the above technical solution, the added mass ratio of the polysaccharide material and the high polymerized proanthocyanidin is 20-40:1.

Further, in the above technical solution, during the mixing and stirring process, the temperature is controlled to be 20-50°C.

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

Further, in the above technical solution, during the mixing and stirring process, the pH is controlled to be 4-9.

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

Specifically, in the above technical solution, the polysaccharide is inulin, and the solubility of the polymeric proanthocyanidins in the aqueous solution is increased from 0.15 mg/ml to 10 mg/ml, solubilizing 66.67 times.

On the other hand, the present invention also provides a high-polymerized proanthocyanidin complex, specifically a complex of high-polymerized procyanidins and polysaccharides.

Specifically, in the above technical solution, in the high polymer proanthocyanidin complex, the polysaccharide material is one or more of pectin, carrageenan, sodium alginate, konjac gum and inulin, preferably Inulin.

Preferably, in the above technical solution, the mass ratio of the polysaccharide material and the high polymerized proanthocyanidin is 20-40:1.

The reagents and raw materials used in the present invention are all commercially available.

Compared with the prior art, the beneficial effects of the present invention are:

(1) The solubilization method of high-polymerized proanthocyanidins provided by the present invention is by adding polysaccharides and through the interaction of polysaccharides and polyphenols, so that high-polymerized proanthocyanidins and polysaccharides form a stable solution in an aqueous solution under certain conditions. The composite particle system increases its solubility by about 70 times;

(2) The solubilization method of high-polymerized proanthocyanidins provided by the present invention has a simple process. It only needs to add polysaccharides in proportion and stir and mix to complete the solubilization process of high-polymerized proanthocyanidins. It is simple and convenient, and has excellent practical application effects;

(3) The complex formed by the combination of high-polymerized proanthocyanidins and polysaccharides provided by the present invention has a significant weight loss and hypoglycemic effect, and the two have a significant synergistic effect.

Description of drawings

Figure 1 is the structure of high polymerized proanthocyanidins in the embodiment of the present invention;

Figure 2 is a transmission electron microscope image of high polymerized proanthocyanidins dissolved in water in an embodiment of the present invention;

Figure 3 is a graph showing the solubilization effects of inulin aqueous solutions of different concentrations on polymeric proanthocyanidins in the embodiments of the present invention;

Figure 4 is a diagram showing the particle size results of the complex solution under different concentrations of polymerized proanthocyanidins in the embodiment of the present invention;

Figure 5 is a diagram showing the particle size results of complex solutions with different concentrations of polymerized proanthocyanidins at different pH in the embodiments of the present invention;

Figure 6 is a diagram showing the particle size results of the complex under different pH in the embodiment of the present invention;

Figure 7 is the body weight test results of mice in different groups in the embodiment of the present invention;

Figure 8 shows the adipose tissue test results of different groups of mice in the embodiment of the present invention;

Figure 9 is the blood glucose test results of different groups of mice in the embodiment of the present invention;

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

Detailed ways

The present invention will be further described in detail below in conjunction with specific embodiments to enable those skilled in the art to understand the present invention more clearly.

The following examples are only used to illustrate the content of the present invention, but are not used to limit the scope of the present invention.

Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts should fall within the protection scope of the present invention.

In the embodiment of the present invention, the polymeric proanthocyanidins used are self-made persimmon high-polymerizing proanthocyanidins, and the extraction method is as follows:

50g of persimmon pulp was mixed with 1% hydrochloric acid-methanol mixture at a material-to-liquid ratio of 1:8 (methanol 2000ml, hydrochloric acid 20ml), condensed and refluxed at 80°C for 40 minutes, repeated three times, combined the extracts, and left to stand overnight; suction filtered, 35 °C, rotary evaporate, concentrate to 50ml (per 2000ml), suction filter, apply AB-8 macroporous resin, adsorb for 40min (sealed with appropriate water), wash with water until sugar-free (colorless when detected by phenol-sulfuric acid method, remove soluble Water-soluble components such as sugar, hydrochloric acid, methanol, etc. will not lose tannins during the water washing process), wash with 10% ethanol until colorless (removing small molecular substances such as pigments), elute with 90% ethanol and collect the eluent, 35°C Concentrate, remove ethanol (rotary evaporation), and freeze-dry to obtain persimmon tannin. Molecular structure analysis shows that its average degree of polymerization is 26, making it a typical high-polymerized proanthocyanidin.

The inulin used was commercially available natural inulin.

Example

Figure 1 shows the structure of polymeric proanthocyanidins.

An aqueous solution of persimmon high-polymerized procyanidins with a concentration of 1 mg/ml was prepared, and a transmission electron microscope was used to observe the existence form of persimmon high-polymerized proanthocyanidins in the aqueous solution.

The results are shown in Figure 2. It can be seen from Figure 2 that the polymeric proanthocyanidins are basically insoluble in water and form agglomerates. Its water-insoluble characteristics make it difficult to be used as a food ingredient in beverages and other food systems.

Prepare inulin aqueous solutions of 0.1, 0.2, 0.25, 0.3 and 0.4g/ml respectively, add 10mg/ml of polymeric procyanidins respectively, vortex for 5 minutes to dissolve, and measure the particle size using a Malvern Nano ZS laser nanoparticle sizer at 25±0.1°C. , each sample was measured three times in parallel.

The results are shown in Figure 3. It can be seen from Figure 3 that high polymerized proanthocyanidin-inulin formed stable nanoparticles, and as the inulin concentration increased, the particle size of the high polymerized proanthocyanidin-inulin complex first decreased. It is small and then increases. At 0.25g/ml, the particle size is the smallest. The particle size of the nanocomposite is a reflection of the existence state of the persimmon polymer proanthocyanidin-inulin complex. Generally, the smaller the particle size, the higher the clarity of the complex and the more stable the complex under the same conditions.

Prepare a 0.25g/ml inulin aqueous solution, dissolve it with ultrasound, add 5, 10, 15, 20, 25, and 30 mg/ml of polymeric proanthocyanidins respectively, vortex for 5 minutes to dissolve, and use a Malvern Nano ZS laser nanoparticle size analyzer to measure the temperature at 25±0.1 The particle size was measured at ℃, and each sample was measured three times in parallel.

The results are shown in Figure 4. It can be seen from Figure 4 that as the concentration of high polymerized proanthocyanidins increases, the particle size of the high polymerized proanthocyanidins-inulin complex gradually increases; in the 0.25g/ml inulin solution, The maximum solubility of polymeric proanthocyanidins is 10 mg/ml. Once exceeded, the particle size of the system will gradually increase and the entire complex system will become unstable.

Prepare inulin solutions of 0.1, 0.2, 0.25, 0.3 and 0.4g/ml respectively, dissolve them with ultrasound, add 10mg/ml of polymeric proanthocyanidins respectively, vortex for 5 minutes to dissolve, and dilute 2, 4, 8, 16, 32 with distilled water in sequence. times, the particle size was measured using a MalvernNano ZS laser nanoparticle sizer at 25±0.1°C, and each sample was measured three times in parallel. This experiment was used to examine its stability under different dilution conditions.

The results are shown in Figure 5. It can be seen from Figure 5 that when the formed stable complex solution is diluted, its particle size does not change much at different dilution multiples. This result shows that the high polymerized proanthocyanidin-chrysanthemum The powder composite is relatively stable, and dilution has little effect on its particle size.

Prepare inulin solutions with concentrations of 0.1, 0.2, 0.25, 0.3 and 0.4g/ml from water with pHs of 2, 4, 7, 8 and 9 respectively, dissolve them with ultrasound, and add 10mg/ml of polymeric proanthocyanidins respectively. Ultrasonicate for 5 minutes, vortex for 15 minutes to dissolve, and measure the particle size using a Malvern Nano ZS laser nanoparticle sizer at 25±0.1°C. Each sample is measured three times in parallel.

The results are shown in Figure 6. It can be seen from Figure 6 that the pH environment has a greater impact on the high-polymerized proanthocyanidin-inulin complex. Under strong acidic conditions, the complex is prone to aggregation and the particle size of the complex becomes larger. This in turn causes the complex to settle and reduce its stability. However, the complex is relatively stable in an environment with a pH of 4-9 and can meet the needs of different food environments.

Effect verification

A mouse model of diet-induced obesity was selected to evaluate the hypoglycemic and lipid-lowering efficacy of the complex. The specific process is as follows.

Fifty C57 mice weighing about 20g were selected and randomly divided into five groups, including a control group, a model group, a high-polymerized proanthocyanidin treatment group, an inulin-treated group, and a high-polymerized proanthocyanidin-inulin complex group, with 10 mice in each group.

in:

Control group: fed 12450J feed;

Model group: fed RD12492 high-fat feed;

The high-polymerized procyanidins treatment group: fed RD12492 high-fat feed and high-polymerized proanthocyanidins at a daily dosage of 20 mg/kg (mouse body weight);

Inulin treatment group: fed RD12492 high-fat feed and inulin, the feeding amount was 500mg/kg (mouse body weight) per day;

Highly polymerized proanthocyanidin-inulin complex group: fed RD12492 high-fat feed and fed high-polymerized proanthocyanidin-inulin complex. The feeding amount was 20 mg/kg high-polymerized proanthocyanidin (mouse body weight) + 500mg/kg per day. Inulin (mouse body weight).

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

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

It mainly includes the weight of blood sugar, blood lipids, and adipose tissue. The blood glucose and blood lipid contents were measured using the kit method; adipose tissue was divided into epididymal fat pad, perirenal fat pad and inguinal fat pad. After the mice were anesthetized and killed, they were taken out and weighed to obtain the weights of the three fat pads.

The results are shown in Figure 7-10.

Figure 7 shows the body weight test results of different groups of mice in the embodiment of the present invention.

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

Figure 8 shows the test results of adipose tissue of mice in different groups in the embodiment of the present invention.

As can be seen from Figure 8, high-fat diet significantly promotes the proliferation of major adipose tissues (epididymal fat pad, perirenal fat pad and inguinal fat pad) in mice, thereby inducing obesity, while high-polymerized proanthocyanidin-inulin complexes can significantly inhibit Hyperplasia of adipose tissue.

Figure 9 shows the blood glucose test results of different groups of mice in the embodiment of the present invention.

As can be seen from Figure 9, the blood sugar of mice induced by high-fat diet increased significantly, and the high-polymerized proanthocyanidin-inulin complex can significantly reduce the increase of blood sugar of mice induced by high-fat meal; at the same time, both showed Significant synergy.

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

The liver and adipose tissue sections shown in Figure 10 show that high-fat diet can significantly induce the occurrence of fatty liver. HE staining of the liver found that a large number of fat bubbles appeared in the liver tissue of the high-fat diet-induced group, while the inulin-high polymerized proanthocyanidin complex treatment group showed It can significantly inhibit the appearance of fat bubbles, thereby inhibiting the occurrence of fatty liver; adipose tissue sections and HE staining also show that high-fat diet can significantly promote the growth of adipocytes, and the diameter of adipocytes is larger, while the high-polymerized proanthocyanidin-inulin complex Things have a strong improvement effect.

From Figures 7-10, it can be found that the inulin-highly polymerized proanthocyanidin complex exhibits 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 used to limit the protection scope of the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention shall be included in the protection scope of the present invention.

Claims (1)

1. A solubilization method of high polymer procyanidine is characterized in that 0.25g/mL of inulin aqueous solution is adopted, ultrasonic dissolution is carried out, 10mg/mL of high polymer procyanidine is added, vortex oscillation is carried out for 5min for dissolution, so that the solubility of the high polymer procyanidine is improved from 0.15mg/mL to 10mg/mL, and solubilization is carried out for 66.67 times, thus obtaining a compound solution of a small-particle-size stable system.