Compositions and their use in maintaining healthy blood glucose levels

Abstract

The present application relates to the technical field of health food, and particularly relates to a composition and application thereof in maintaining blood glucose health level. The composition comprises rutin, quercetin, kaempferol and pectin. Experiments show that the composition of the present application significantly reduces the activity of alpha-glucosidase, significantly delays starch digestibility, thereby reduces the postprandial blood glucose level of mice, and the effect is obviously better than that of other flavonoid and pectin compositions. It is shown that the rutin, quercetin, kaempferol and pectin are compounded according to a certain proportion, and obvious synergistic effect is produced. In addition, the composition prepared by the present application reduces the use amount of quercetin, increases the utilization rate of pectin, reduces the production cost, and the raw materials are pure natural, safe and non-toxic, functional, low in cost, and have wide application prospect in the deep processing of starch-based health food.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of health food, in particular to a composition and its application in maintaining a healthy level of blood glucose. BACKGROUND

[0002] Alpha-glucosidase is one of the starch-digesting enzymes that has attracted much attention in research. When starch is consumed, it is first hydrolyzed into disaccharides and oligosaccharides by amylase; then, the non-reducing ends of these molecules are cleaved by alpha-glucosidase on the epithelial cells of the intestine, releasing glucose into the blood circulation, which is then absorbed and utilized by the human body. Therefore, by inhibiting the activity of alpha-glucosidase, the production of glucose in the intestine can be delayed or inhibited, and the blood glucose level in the body can be controlled. Currently, alpha-glucosidase inhibitors are mainly some chemically synthesized drugs, such as acarbose, metformin and miglitol, etc. In clinical practice, they are used to assist in the treatment of diabetes, but long-term use can easily cause side effects such as hypoglycemia, gastrointestinal dysfunction and weight gain. Therefore, it is necessary to find new alpha-glucosidase inhibitors that are natural, have stronger inhibitory activity and fewer side effects.

[0003] It has been reported that flavonoids can block the digestion and absorption of carbohydrates by inhibiting the activity of alpha-glucosidase, achieving the purpose of controlling postprandial hyperglycemia, but the effect is still not ideal. It is of great significance to develop a composition with better blood glucose-lowering effect for the preparation of health products. Pectin is a soluble dietary fiber naturally occurring in fruits and vegetables and is one of the components of plant cell walls, which has been widely used in food processing. Currently, there is no report on the use of pectin and flavonoids for maintaining blood glucose. SUMMARY

[0004] Therefore, the present application provides a composition and its application in maintaining a healthy level of blood glucose.

[0005] The composition comprises rutin, quercetin, kaempferol and pectin.

[0006] In the present application, the mass ratio of rutin, quercetin, kaempferol and pectin is (0.0019-0.0074):(0.0008-0.0032):(0.0011-0.0044):(10-15). In some specific embodiments, the mass ratio of rutin, quercetin, kaempferol and pectin is 0.0019:0.0008:0.0011:2.5, 0.0019:0.0008:0.0011:3.75, 0.0037:0.0016:0.0022:2.5, 0.0037:0.0016:0.0022:3.75, 0.0074:0.0032:0.0044:2.5, 0.0074:0.0032:0.0044:3.75, or any ratio between the above two ratio ranges.

[0007] The present application investigates the activity of α-glucosidase and the hypoglycemic effect of other flavonoids (such as rutin, quercetin, kaempferol) and pectin in combination. The results show that the composition of rutin, quercetin, kaempferol and pectin in the present application can effectively inhibit the activity of α-glucosidase, solve the problem of limited anti-starch digestion and hypoglycemic effect of single active ingredient in the prior art, and significantly improve the inhibition activity of α-glucosidase and the hypoglycemic effect compared with other compositions, with CI less than 0.9. It is shown that the four components of rutin, quercetin, kaempferol and pectin in the composition of the present application are reasonable and have a synergistic effect.

[0008] In the present application, the "composition" is not limited to the physical mixing of active ingredients in the composition, but also includes the combination of active ingredients in any order within a certain time interval. Specifically, in the present application, the composition is the combination of flavonoids and pectin in preventing and treating diabetes or maintaining the healthy level of blood glucose, and the two can exist in a mixed form or can exist independently.

[0009] In the present application, the composition further comprises a buffer solution. In some embodiments, the buffer solution comprises a sodium phosphate buffer, a potassium phosphate buffer, a potassium phosphate buffer, a sodium acetate buffer or a potassium acetate buffer. In some specific embodiments, the buffer solution is a phosphate buffer. In some embodiments, the concentration of the buffer solution is 0.1 M-0.3 M, and specifically can be 0.1 M, 0.2 M, 0.3 M. In some embodiments, the pH of the buffer solution is 6.0-8.0, and specifically can be 6.0, 6.5, 7.0, 7.5, 8.0.

[0010] The application also provides a preparation method of the composition, which comprises: dissolving the rutin, quercetin, kaempferol and pectin in a phosphate buffer. In some embodiments, the preparation method comprises: dissolving the rutin, quercetin and kaempferol in a phosphate buffer, dissolving the pectin in a phosphate buffer, and then mixing. In other embodiments, the preparation method comprises: dissolving the rutin, quercetin, kaempferol and pectin in a phosphate buffer at the same time.

[0011] The application also provides use of the composition in the preparation of a product for maintaining a healthy blood sugar level.

[0012] In the use of the application, the maintaining a healthy blood sugar level comprises: inhibiting α-glucosidase, delaying starch digestion, reducing the content of RDS, increasing the content of RS and / or increasing the content of SDS.

[0013] In the application, when the flavonoids (rutin, quercetin and kaempferol) and pectin in the composition of the application are used for inhibiting α-glucosidase or for maintaining a healthy blood sugar level, the flavonoids and pectin can be used at the same time or in any order. When the two are used at the same time, there are two cases: the flavonoids and pectin exist in a mixed form, or the flavonoids and pectin exist independently and are used at the same time. Experiments show that whether the two are used at the same time or in different orders, they have a significant inhibitory effect on the activity of α-glucosidase.

[0014] The application also provides use of the composition in the preparation of an α-glucosidase inhibitor.

[0015] The application also provides a product for maintaining a healthy blood sugar level, which comprises the composition of the application and an excipient. In the application, the product comprises a health food. Further, the health food comprises but is not limited to a starch-based health food, a starch-based health care food, etc. In the application, the dosage form of the product comprises but is not limited to an oral liquid, a syrup, a granule, a dripping pill, a capsule, a pill and a tablet.

[0016] The application does not have special restrictions on the type of excipient, and any commonly used in the art can be selected and adjusted according to the type of product, for example, when the product is a health food, the excipients of the health food comprise but are not limited to a filler, a sweetener, a disintegrant, a lubricant, a binder, etc. The specific types of the above excipients are not particularly limited, and any commonly used in the art can be used.

[0017] The application also provides a method for maintaining a healthy blood sugar level, which comprises: administering the composition of the application or the product of the application.

[0018] The present application provides a composition and its application in maintaining a healthy blood sugar level. The composition comprises rutin, quercetin, kaempferol and pectin. Experimental results show that the composition of the present application significantly reduces the activity of alpha-glucosidase, significantly delays starch digestion, thereby reducing the postprandial blood sugar level of mice, and its effect is obviously better than that of other flavonoids and pectin compositions. It is shown that the rutin, quercetin, kaempferol and pectin are compounded in a certain proportion, and a significant synergistic effect is produced. In addition, the composition prepared by the present application reduces the use amount of quercetin, increases the utilization rate of pectin, reduces the production cost, and the raw materials are pure natural, safe and non-toxic, functional and low in cost, and has a wide application prospect in the deep processing of starch-based healthy food. BRIEF DESCRIPTION OF DRAWINGS

[0019] Figure 1 The inhibition rate of flavonoid monomers on the activity of alpha-glucosidase;

[0020] Figure 2 The CI value of different combinations of flavonoids for inhibiting the activity of alpha-glucosidase;

[0021] Figure 3 The inhibition rate of different combinations of flavonoids in the presence of pectin on the activity of alpha-glucosidase;

[0022] Table 1 is the CI value of pectin-flavonoid composition for inhibiting alpha-glucosidase;

[0023] Figure 4 The inhibition rate of pectin-rutin-quercetin-kaempferol mixture on the activity of alpha-glucosidase under different addition sequences;

[0024] Figure 5 The starch digestion and hydrolysis curve in the presence or absence of pectin and rutin-quercetin-kaempferol mixture;

[0025] Figure 6 The starch digestion fragment content in the presence or absence of pectin and rutin-quercetin-kaempferol mixture;

[0026] Figure 7 The change of postprandial blood sugar level of mice in the presence or absence of pectin and rutin-quercetin-kaempferol mixture. DETAILED DESCRIPTION

[0027] The application discloses a kind of compositions for regulating starch digestion and blood glucose level by inhibiting alpha-glucosidase activity, and those skilled in the art can learn from the content herein, and appropriately improve process parameters to realize.It is particularly pointed out that all similar substitutions and changes are obvious to those skilled in the art, and they are all regarded as included in the present application.The application described in the application has been described by preferred embodiments, and the relevant personnel can obviously make changes or appropriate changes and combinations to the method and application described herein without departing from the content, spirit and scope of the application, to realize and apply the present application technology.

[0028] The application relates to an alpha-glucosidase inhibitory activity determination method.

[0029] Mix 50 μL of sample, 50 μL of alpha-glucosidase (0.33 U / mL) and 50 μL of phosphate buffer (0.2 M) thoroughly, and react at 37°C for 10 min. Then, add 100 μL of p-nitrophenyl-beta-D-glucopyranoside (3 mM) and react at 37°C for 20 min. After the reaction is completed, immediately add 750 μL of 0.2 M sodium carbonate solution to terminate the reaction. PBS is used as a blank control. Finally, the absorbance of the sample at 405 nm is determined by using an enzyme label instrument. The calculation formula (1) of the inhibition rate of alpha-glucosidase is as follows:

[0030] (Formula 1)

[0031] Wherein, A, B, C and D represent the absorbance values of the sample, the sample blank, the control and the control blank respectively.

[0032] The application relates to a combined inhibition determination method of alpha-glucosidase.

[0033] Phosphate buffer (0.2 mol / L, pH 6.9) is used as a solvent to configure a series of sample solutions with different proportions. In the reaction system, the concentration ratio of the sample is IC 50 The dose is the concentration of the flavonoid compound combination, and Fa (0 < Fa < 1) is the effect, that is, the inhibition rate. The Fa-combined index (CI) curve is used to evaluate the combined inhibition of the flavonoid compound on alpha-glucosidase. CI < 0.9, CI = 0.9-1.1 and CI > 1.1 respectively represent synergistic effect, additive effect and antagonistic effect.

[0034] The application relates to a determination method of hydrolysis rate and digestion fragment content of in-vitro starch digestion.

[0035] Cassava starch was dissolved in sodium acetate buffer solution (0.2 mol / L) at 10 mg / mL, and then pasted at 90°C for 20 min and stored at 37°C for standby. 0.75 mL of sample solution with different concentrations and 1.25 mL of mixed enzyme solution (290 U / mL of α-amylase and 60 U / mL of α-glucosidase) were mixed thoroughly, and then incubated at 37°C for 20 min. Sodium acetate buffer solution was used as a control. 1.8 mL of pasted starch was added to the above reaction solution, and then oscillated in a constant temperature water bath at 37°C at a speed of 150 r / min. 150 μL of reaction solution was taken at 0, 20, 40, 60, 90, 120 and 180 min, and then added to 1.5 mL of test tube containing 300 μL of 0.3 M Na2CO3 to stop the reaction. Then, the reaction solution was centrifuged at 7300 rpm for 5 min, and the supernatant and precipitate were stored for standby. Then, 100 μL of supernatant was added to 3 mL of GOPOD reagent, and then water-bathed at 50°C for 20 min. The absorbance value of each sample was measured at 510 nm. 100 μL of 1 mg / mL glucose solution was used as a standard sample. The calculation formulae (2)-(5) of the hydrolysis rate of starch and the content of digestion fragments are as follows:

[0036] Equation 2

[0037] Equation 3

[0038] Equation 4

[0039] Equation 5

[0040] wherein Gt represents the glucose content in the sample at time t; RDS represents the content of fast-digesting starch, %; SDS represents the content of slow-digesting starch, %; RS represents the content of resistant starch, %; G0 represents the amount of free glucose before enzyme hydrolysis, mg; G20 represents the amount of glucose produced after enzyme hydrolysis for 20 min, mg; G120 represents the amount of glucose produced after hydrolysis for 120 min, mg; and S represents the total starch content in the sample, mg. t 20 120

[0041] The determination method of postprandial blood glucose response involved in the embodiment of the present application is as follows:

[0042] ​​​Male golden hamsters (initial body weight: 75-85 g) were purchased at 4 weeks of age and housed in a SPF-grade animal room (temperature: 24 ± 2℃; relative humidity: 55 ± 5% humidity; 12 h light / dark cycle), 6 per cage, and fed with ordinary feed, allowing free access to food and water. After 1 week of acclimation, the mice were divided into 4 groups (n = 5) according to body weight, and each group of mice was fasted for 18 h. Then, the fasted mice were administered with normal saline (100 mL / kg body weight (BW)), a flavonoid mixture sample (100 mg / kg BW), and a flavonoid mixture-pectin sample (100 mg / kg BW-200 mg / kg BW, 100 mg / kg BW-300 mg / kg BW) by feeding alone. After about 30 min of feeding, each mouse was administered with a cassava starch sample (2 g / kg BW) by gavage. Blood samples were collected from the tail lateral vein at 0, 30, 60, and 120 min after gavage. Blood glucose levels were measured by a blood glucose meter.

[0043] To further illustrate the present application, a composition for regulating starch digestion and blood glucose levels by inhibiting α-glucosidase activity provided by the present application is described in detail below in conjunction with examples.

[0044] The test materials used in the present application are all ordinary commercially available products and can be purchased in the market.

[0045] The present application is further described below in conjunction with examples:

[0046] Comparative Example 1

[0047] Rutin was dissolved in phosphate buffer (0.3 M, pH 6.5). 50 μL of rutin solution at different concentrations (100, 200, 300, 400, 500 μg / mL) was mixed with 50 μL of α-glucosidase (0.33 U / mL) and 50 μL of phosphate buffer, and incubated at 37℃ for 25 min to measure the inhibitory ability on α-glucosidase activity.

[0048] Comparative Example 2

[0049] Quercetin was dissolved in phosphate buffer (0.2 M, pH 6.9). 50 μL of quercetin solution at different concentrations (50, 100, 150, 200, 250 μg / mL) was mixed with 50 μL of α-glucosidase (0.33 U / mL) and 50 μL of phosphate buffer, and incubated at 37℃ for 20 min to measure the inhibitory ability on α-glucosidase activity.

[0050] Comparative Example 3

[0051] Dissolve kaempferol in phosphate buffer (0.2 M, pH 7.5). Take 50 μL of kaempferol solution with different concentrations (100, 200, 300, 400, 500 μg / mL) and mix with 50 μL of α-glucosidase (0.33 U / mL) and 50 μL of phosphate buffer thoroughly, incubate at 37°C for 15 min, and measure the inhibitory ability on α-glucosidase activity.

[0052] Comparative Example 4

[0053] Dissolve rutin and quercetin in phosphate buffer (0.2 M, pH 6.9) and mix in different proportions. The concentration gradient of rutin and quercetin is set according to IC 50 value. Respectively use 1 / 8 IC 50 , 1 / 4 IC 50 , 1 / 2 IC 50 , IC 50 and 2 IC 50 . The concentration ratio of rutin and quercetin is 74.44:31.78, 148.89:63.57, 297.78:127.13, 595.55:254.26, 1191.11:508.52, respectively. The concentration unit of each component is μg / mL. L Take 50 μL of rutin-quercetin mixture and mix with 50 μL of α-glucosidase and 50 μL of phosphate buffer thoroughly, incubate at 37°C for 15 min, and measure the inhibitory ability on α-glucosidase activity, and calculate the CI value.

[0054] Comparative Example 5

[0055] Dissolve rutin and kaempferol in phosphate buffer (0.3 M, pH 6.5) and mix in different proportions. The concentration gradient of rutin and kaempferol is set according to IC 50 value. Respectively use 1 / 8 IC 50 , 1 / 4 IC 50 , 1 / 2 IC 50 , IC 50 and 2 IC 50 . The concentration ratio of rutin and kaempferol is 74.44:43.79, 148.89:87.59, 297.78:175.17, 595.55:350.35, 1191.11:700.69, respectively. The concentration unit of each component is μg / mL. L Take 50 μL of rutin-kaempferol mixture and mix with 50 μL of α-glucosidase and 50 μL of phosphate buffer thoroughly, incubate at 37°C for 25 min, and measure the inhibitory ability on α-glucosidase activity, and calculate the CI value.

[0056] Comparative Example 6

[0057] Quercetin and Kaempferol were dissolved in phosphate buffer (0.3 M, pH 7.5) and mixed in different proportions. The concentration gradient of quercetin and kaempferol was set according to IC 50 values, and 1 / 8 IC 50 , 1 / 4 IC 50 , 1 / 2 IC 50 , IC 50 and 2 IC 50 were used respectively. The concentration ratio of quercetin to kaempferol was 31.78:43.79, 63.57:87.59, 127.13:175.17, 254.26:350.35, 508.52:700.69 respectively. The concentration of each component was μg / mL. L 50 μL of the quercetin-kaempferol mixture was mixed with 50 μL of α-glucosidase and 50 μL of phosphate buffer, and incubated at 37°C for 20 min. The inhibitory ability on α-glucosidase activity was measured, and the CI value was calculated.

[0058] Example 1

[0059] Rutin, quercetin and kaempferol were dissolved in phosphate buffer (0.2 M, pH 6.9) and mixed in different proportions. The concentration gradient of rutin, quercetin and kaempferol was set according to IC 50 values, and 1 / 8 IC 50 , 1 / 4 IC 50 , 1 / 2 IC 50 , IC 50 and 2 IC 50 were used respectively. The concentration ratio of rutin, quercetin and kaempferol was 74.44:31.78:43.79, 148.89:63.57:87.59, 297.78:127.13:175.17, 595.55:254.26:350.35, 1191.11:508.52:700.69 respectively. The concentration of each component was μg / mL. L 50 μL of the rutin-quercetin-kaempferol mixture was mixed with 50 μL of α-glucosidase and 50 μL of phosphate buffer, and incubated at 37°C for 20 min. The inhibitory ability on α-glucosidase activity was measured, and the CI value was calculated.

[0060] The experimental results of Example 1 and Comparative Examples 1-6 are shown in Figure 1 and Figure 2 . From Figure 1It can be seen that the concentrations of rutin, quercetin and kaempferol are positively correlated with the ability to inhibit α-glucosidase. Quercetin has the strongest ability to inhibit α-glucosidase, followed by kaempferol, and rutin has the weakest. When the concentration of quercetin is 250 μg / mL, the inhibition rate of α-glucosidase activity can reach 52.71±1.71%. From the above results, it can be seen that the ability of rutin to inhibit α-glucosidase is weaker than that of quercetin and kaempferol. Therefore, the combination of rutin and quercetin or rutin and kaempferol is used to further study the synergistic effect of the combination of flavonoids on the inhibition of α-glucosidase. Figure 2 It can be seen that, compared with the two two combinations of flavonoids, the CI value of the combination of rutin, quercetin and kaempferol is the lowest. Among them, when the concentration ratio of rutin-quercetin-kaempferol mixture is 74.44:31.78:43.79, 148.89:63.57:87.59, 297.78:127.13:175.17, 595.55:254.26:350.35, 1191.11:508.52:700.69, the CI value is less than 0.9 (CI value is between 0.891±0.009~0.307±0.010). This shows that the combination of rutin, quercetin and kaempferol has the strongest synergistic effect on the inhibition of α-glucosidase.

[0061] Comparative Example 7

[0062] Rutin and quercetin were dissolved in phosphate buffer (0.2 M, pH 6.9) at concentration ratios of 37.22:15.89, 74.44:31.78 and 148.89:63.57, respectively, to obtain different rutin-quercetin solutions. Among them, the concentration unit of each component is μg / mL. L Take 250 μL pectin (10 mg / mL~15 mg / mL) dissolved in 0.2 M pH 6.5 phosphate buffer and mix with 50 μL rutin-quercetin mixture at 37°C for 15 min. Among them, the mass ratio of rutin-quercetin and pectin is 0.0027:2.5, 0.0027:3.75, 0.0053:2.5, 0.0053:3.75, 0.0106:2.5, 0.0106:3.75. Then, 50 μL of α-glucosidase solution and 50 μL of phosphate buffer are added, and the mixture is incubated at 37°C for another 15 min. Then, the ability to inhibit α-glucosidase activity is measured.

[0063] Comparative Example 8

[0064] Rutin and kaempferol were dissolved in phosphate buffer (0.3 M, pH 6.9) at concentration ratios of 37.22:21.90, 74.44:43.79 and 148.89:87.59, respectively, to obtain different rutin-kaempferol solutions. Among them, the concentration unit of each component is μg / mL. LExample 1

[0065] Comparative Example 9

[0066] Quercetin and kaempferol were dissolved in phosphate buffer (0.2 M, pH 7.5) at concentration ratios of 15.89:21.90, 31.78:43.79 and 63.57:87.59, respectively, to obtain different quercetin-kaempferol solutions. The concentration of each component was μg / mL. L Example 1

[0067] Example 2

[0068] Rutin, quercetin and kaempferol were dissolved in phosphate buffer (0.2 M, pH 6.9) at concentration ratios of 37.22:15.89:21.90, 74.44:31.78:43.79 and 148.89:63.57:87.59, respectively, to obtain different rutin-quercetin-kaempferol solutions (i.e. flavonoid solutions). The concentration of each component was μg / mL. L250 μL of pectin solution (pectin concentration 10 mg / mL–15 mg / mL) dissolved in 0.2 M pH 6.5 phosphate buffer was mixed with 50 μL of flavonoid solution and incubated at 37 °C for 10 min. The mass ratios of rutin-quercetin-kaempferol and pectin were 0.0038:2.5, 0.0038:3.75, 0.075:2.5, 0.0075:3.75, 0.0150:2.5, and 0.0150:3.75. Then, 50 μL of α-glucosidase solution and 50 μL of phosphate buffer were added, and the mixture was incubated at 37 °C for another 20 min. The inhibitory effect on α-glucosidase activity was then measured.

[0069] Example 3

[0070] Rutin, quercetin, and kaempferol were dissolved in phosphate buffer (0.2 M, pH 6.9) at concentration ratios of 37.22:15.89:21.90, 74.44:31.78:43.79, and 148.89:63.57:87.59 to obtain different rutin-quercetin-kaempferol solutions. The concentration of each component is expressed in μg / mL. 50 μL of the rutin-quercetin-kaempferol mixture was incubated with 50 μL of α-glucosidase solution and 50 μL of phosphate buffer at 37°C for 20 min. Then, 250 μL of pectin solution (10 mg / mL–15 mg / mL, dissolved in 0.2 M pH 7.5 phosphate buffer) was added. The mass ratios of rutin-quercetin-kaempferol and pectin were 0.0038:2.5, 0.0038:3.75, 0.075:2.5, 0.0075:3.75, 0.0150:2.5, and 0.0150:3.75. The mixture was then incubated at 37°C for 10 min, and its inhibitory activity against α-glucosidase was measured.

[0071] The experimental results obtained in Examples 2-3 and Comparative Examples 7-9 are as follows: Figure 3 and Figure 4 As shown. By Figure 3 It was found that the addition of pectin enhanced the inhibition of α-glucosidase by different flavonoid combinations. At the same concentration, the pectin-rutin-quercetin-kaempferol combination showed the strongest synergistic effect on α-glucosidase inhibition (Table 1). Specifically, when the pectin concentration was 10–15 mg / mL, the inhibition rate of α-glucosidase by the 0.075 mg / mL rutin-quercetin-kaempferol mixture increased from 26.69 ± 0.74% to 37.38 ± 1.50%–42.57 ± 1.17%. Furthermore, from… Figure 4It can be seen that compared with the mixture of rutin-quercetin-kaempferol and pectin mixed first (order 1) before adding α-glucosidase, the effect of adding order 2 (mixing the mixture of rutin-quercetin-kaempferol and α-glucosidase first before adding pectin) on the inhibition of α-glucosidase by the mixture of rutin-quercetin-kaempferol is consistent with the trend produced by order 1, that is, pectin and the mixture of rutin-quercetin-kaempferol synergistically inhibit α-glucosidase. Among them, under order 2, when the concentration of pectin is 10-15 mg / mL and the concentration of the mixture of rutin-quercetin-kaempferol is 0.15 mg / mL, the inhibition rate of α-glucosidase is 68.27±0.67%-77.68±1.36%. The inhibition rate of order 1 is 58.33±1.02%-48.86±0.97% under the same concentration. It can be seen that the protective effect of order 1 on α-glucosidase is stronger than that of order 2.

[0072] Table 1 is the CI value of pectin-flavonoid composition inhibiting α-glucosidase

[0073]

[0074] Note: R, Q and K represent rutin, quercetin and kaempferol respectively, and P represents pectin.

[0075] Comparative Example 10

[0076] Mix 1.95 mL of sodium acetate buffer solution (0.2 mol / L, pH 5.4) and 1.25 mL of mixed enzyme solution thoroughly. Measure the hydrolysis rate and digestion fragment content of the sample at different time periods.

[0077] Comparative Example 11

[0078] Dissolve the mixture of rutin, quercetin and kaempferol in sodium acetate buffer solution (0.3 mol / L, pH 5.4) at a concentration ratio of 595.55:254.26:350.35. Take 1.20 mL of sodium acetate buffer solution (0.2 mol / L, pH 5.4) and 0.75 mL of the mixture of rutin-quercetin-kaempferol and incubate at 37°C for 20 min. Measure the hydrolysis rate and digestion fragment content of the sample at different time periods.

[0079] Example 4

[0080] A mixture of rutin, quercetin and kaempferol was dissolved in sodium acetate buffer (0.2 mol / L, pH 5.2) at a concentration ratio of 595.55:254.26:350.35. 1.20 mL of pectin solution (2.5 mg / mL, dissolved in 0.2 M pH 5.2 phosphate buffer) and 0.75 mL of the mixture of rutin-quercetin-kaempferol were incubated at 37°C for 10 min. The hydrolysis rate and the content of digested fragments of the sample were determined at different time periods.

[0081] Example 5

[0082] A mixture of rutin, quercetin and kaempferol was dissolved in sodium acetate buffer (0.3 mol / L, pH 5.4) at a concentration ratio of 595.55:254.26:350.35. 1.20 mL of pectin solution (5 mg / mL, dissolved in 0.3 M pH 5.4 phosphate buffer) and 0.75 mL of the mixture of rutin-quercetin-kaempferol were incubated at 37°C for 15 min. The hydrolysis rate and the content of digested fragments of the sample were determined at different time periods.

[0083] Example 6

[0084] A mixture of rutin, quercetin and kaempferol was dissolved in sodium acetate buffer (0.2 mol / L, pH 5.6) at a concentration ratio of 595.55:254.26:350.35. 1.20 mL of pectin solution (10 mg / mL, dissolved in 0.2 M pH 5.6 phosphate buffer) and 0.75 mL of the mixture of rutin-quercetin-kaempferol were incubated at 37°C for 20 min. The hydrolysis rate and the content of digested fragments of the sample were determined at different time periods.

[0085] The effects of Examples 4-6 and Comparative Examples 10-11 on the hydrolysis rate of starch and the content of digested fragments are shown in Tables 1-3. Figure 5 and Figure 6 As can be seen from Tables 1-3, Figure 5 the hydrolysis rate of starch increased rapidly within 0-20 min, slowed down within 20-60 min, and tended to be flat within 60-180 min. After the addition of the mixture of rutin-quercetin-kaempferol, the hydrolysis rate of starch at 180 min decreased from 59.18% to 30.93%. After the addition of pectin at different concentrations, the hydrolysis rate of starch at 180 min further decreased to 28.17%-24.26%, indicating that the combination of pectin and flavonoids enhanced the ability of flavonoids to inhibit starch hydrolysis. Figure 6It can be seen that after the addition of the rutin-quercetin-kaempferol mixture, the RDS content of the starch decreased from 49.27% to 16.92%, and the SDS and RS contents increased from 8.56% and 42.17% to 14.23% and 68.85%, respectively. When different concentrations of pectin were combined with the rutin-quercetin-kaempferol mixture, the RDS content further decreased with the increase of the concentration of pectin, and the SDS and RS contents showed opposite trends. This indicates that pectin helps to enhance the conversion of RDS in starch to SDS and RS by the rutin-quercetin-kaempferol mixture.

[0086] Comparative Example 12

[0087] The cassava starch sample was administered to each mouse by gavage (2 g / kg BW). Blood samples were collected from the lateral tail vein at 0, 30, 60, and 120 minutes after gavage, and blood glucose levels were measured.

[0088] Comparative Example 13

[0089] The mice were administered a feed of the rutin-quercetin-kaempferol mixture (100 mg / kg BW, concentration ratio IC 50 The cassava starch sample was administered to each mouse by gavage (2 g / kg BW) about 30 minutes after the administration of the feed. Blood samples were collected from the lateral tail vein at 0, 30, 60, and 120 minutes after gavage, and blood glucose levels were measured.

[0090] Example 7

[0091] The mice were administered a feed of the rutin-quercetin-kaempferol mixture-pectin sample (100 mg / kg BW-200 mg / kg BW), and the cassava starch sample was administered to each mouse by gavage (2 g / kg BW) about 30 minutes after the administration of the feed. Blood samples were collected from the lateral tail vein at 0, 30, 60, and 120 minutes after gavage, and blood glucose levels were measured.

[0092] Example 8

[0093] The mice were administered a feed of the rutin-quercetin-kaempferol mixture-pectin sample (100 mg / kg BW-300 mg / kg BW), and the cassava starch sample was administered to each mouse by gavage (2 g / kg BW) about 30 minutes after the administration of the feed. Blood samples were collected from the lateral tail vein at 0, 30, 60, and 120 minutes after gavage, and blood glucose levels were measured.

[0094] The experimental results obtained in Examples 7-8 and Comparative Examples 12-13 are shown in Table 1. Figure 6

[0095] ​The blank control group was given the cassava starch sample of Comparative Example 12.

[0096] The R-Q-K group was given the sample of Comparative Example 13.

[0097] The R-Q-K+ 200mg / kg BW P group was given the sample of Example 7.

[0098] The R-Q-K+ 300mg / kg BW P group was given the sample of Example 8.

[0099] The results show that the postprandial blood glucose level of the mice fed with starch first increased and then decreased, and reached the highest at 30 min. After being fed with the rutin-quercetin-kaempferol mixture, the postprandial blood glucose level of the mice at 30 min decreased from 288.7 mg / dL to 231.1 mg / dL. After being fed with the combination of the rutin-quercetin-kaempferol mixture and pectin, the starch digestion of the mice was further delayed, and the postprandial blood glucose level was reduced (see the R-Q-K+ 200mg / kg BW P group and the R-Q-K+ 300mg / kg BW P group). Among them, the postprandial blood glucose levels of the mice in the R-Q-K+ 300mg / kg BW P group at 30 min and 60 min were 198.8 mg / dL and 144.2 mg / dL, respectively, and the blood glucose-lowering effect was the best. The above results show that the pectin-rutin-quercetin-kaempferol composition has great advantages in reducing blood glucose.

[0100] The above only describes the preferred embodiments of the present application, and it should be noted that those skilled in the art can make several improvements and refinements without departing from the principles of the present application, and these improvements and refinements should also be considered as the protection scope of the present application.

Claims

1. A composition for helping to maintain healthy blood sugar levels, characterized in that, It is composed of rutin, quercetin, kaempferol, pectin, and buffer solution; The mass ratio of rutin, quercetin, kaempferol, and pectin is: 0.0019：0.0008：0.0011：2.5、0.0019：0.0008：0.0011：3.75、0.0037：0.0016：0.0022：2.5、0.0037：0.0016：0.0022：3.75、0.0074：0.0032：0.0044：2.5、0.0074：0.0032：0.0044：3.75。 2. The composition of claim 1, wherein, The buffer solution includes at least one of sodium phosphate buffer, potassium phosphate buffer, potassium phosphate buffer, sodium acetate buffer, or potassium acetate buffer.

3. The composition according to claim 2, characterized in that, The concentration of the buffer solution is 0.10–0.30 M, and the pH is 6.00–8.

0.

4. A method for preparing the composition according to any one of claims 1 to 3, characterized in that, The rutin, quercetin, kaempferol, and pectin were dissolved in phosphate buffer.

5. The use of the composition according to any one of claims 1 to 3 or the composition prepared by the preparation method according to claim 4 in the preparation of health foods that help maintain healthy blood sugar levels.

6. The application according to claim 5, characterized in that, Maintaining healthy blood glucose levels includes: inhibiting α-glucosidase activity, delaying starch digestion, reducing rapidly digestible starch content, increasing resistant starch content, and / or increasing slowly digestible starch content.

7. A health food product that helps maintain healthy blood sugar levels, characterized in that... Includes the composition and excipients as described in any one of claims 1 to 3.

8. The health food according to claim 7, characterized in that, The dosage forms of the health food include oral liquids, syrups, granules, capsules, pills, and tablets.

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