Method specially for predicting glycemic index of rice

Abstract

A method specially for predicting the glycemic index (GI) of rice. An in-vitro digestion model specially for rice is developed, so as to obtain in-vitro digestion test parameters. Rice with known GI is used as a standard reference object, and a mathematical function correlation between the in-vivo GI index and in-vitro simulated digestion parameters is established, so that accurate prediction of the GI index of rice to be tested is achieved, wherein the linear correlation r is 0.95 or above, and the prediction accuracy is high. Unreasonable or unnecessary steps in a conventional method for determining the GI of rice in vitro are omitted in the method, such that the method involves a simpler operation, and can also be applied to rice variety screening and rice breeding.

Description

A method specifically for predicting the glycemic index of rice Technical Field

[0001] The invention relates to a method specifically used for predicting the glycemic index of rice, belonging to the technical field of food detection. Background Art

[0002] Rice (Oryza sativa L.), primarily composed of starch, protein, and lipids, is a staple food for more than half the world's population. Starch, the primary carbohydrate in rice, plays a crucial role in its digestion, influencing blood sugar responses. Due to its ease of digestion and absorption, rice consumption can lead to a rapid rise in blood sugar levels. Consequently, rice is often considered a food with a high glycemic index (GI). In recent years, public health awareness has grown, and consumer demand for higher-quality dietary nutrition has steadily increased. Consequently, medium- and low-GI rice varieties have emerged with advances in agronomy and rice breeding.

[0003] Human trials are the primary method for determining GI (GI) and are currently the standard method for determining food GI both domestically and internationally. They primarily rely on the concept of GI by measuring blood glucose levels after consuming a meal containing 50g of carbohydrates. GI determination in human trials is influenced by multiple factors, including the subject's health, the number of subjects, pre-test meal, exercise, and medication requirements, the reference food, the effective CHO content of the test food, the frequency of the testing procedure, the glucose monitoring method, and data processing. Furthermore, this method of human GI testing is labor- and time-intensive, requiring the recruitment and screening of human subjects, which results in significant financial and time costs. The reference and test foods must be consumed on different days, and testing typically requires several days. Furthermore, due to the large sample volumes consumed in human trials, newly cultivated rice samples often cannot meet the requirements of human trials. In summary, the high cost, time consumption, large sample requirements, and high personnel requirements of human GI testing significantly limit the development and testing of healthy foods in the food industry.

[0004] Therefore, in vitro simulated digestion methods are increasingly being used to predict and compare the GI of foods due to their advantages of rapidity, reproducibility, and low cost. Currently, the main methods for determining the GI of rice under in vitro conditions are the Englyst and M. Minekus methods. Both utilize enzymes to hydrolyze starch and calculate the amount of starch hydrolyzed by measuring the amount of glucose released. The first type of method is relatively simple, involving enzymatic digestion of foods in a fixed buffer. Many existing methods are modified from the Englyst method. However, due to significant differences between experimental methods, the results between different research groups are not comparable. Furthermore, these methods do not account for multiple digestive stages and various non-starch components, which poses significant limitations. The second type, the M. Minekus method, simulates starch digestion stages in the mouth, stomach, and small intestine. The buffer preparation is extremely complex, involving multiple enzymes such as salivary amylase, pepsin, trypsin, pancreatic amylase, convertase, and bile salts. Furthermore, the enzyme types and concentrations required for different foods vary widely. Given that low-GI rice holds great potential as a healthy staple food, developing a method specifically for predicting the glycemic index of rice is particularly important.

[0005] In recent years, researchers have improved the M.Minekus method and disclosed a method for in vitro determination of the glycemic index of carbohydrate foods in patent CN 113238010A. This patent uses inorganic salts and enzymes to simulate saliva, gastric juice, and small intestinal digestive fluid, and simulates the digestion of food in the mouth, stomach, and small intestine. The GI value of the tested sample can be quickly evaluated by measuring the area under the curve AUC-20. Although this method has a high correlation, it has the following problems: (1) It needs to simulate the three stages of oral, gastric, and small intestinal digestion, and the process is cumbersome; (2) The composition of the simulated liquid is complex and the types of inorganic salts required are complicated; for example, simulated saliva (SSF) is composed of a mixture of the following components: 30.2mL, 0.5mL, 1mL, 2mL, 3mL, 4mL, 5mL, 6mL, 7mL, 8mL, 9mL, 10mL, 10mL, 20mL, 30mL, 10mL, 20mL, 30mL, 10mL, 40mL, 8mL, 7mL, 8mL, 9mL, 10mL, 10mL, 10mL, 20mL, 30mL, 10mL, 20mL, 30mL, 10mL, 40mL, 8mL mol / L KCl solution + 7.4mL, 0.5mol / L KH2PO4 solution + 13.6mL, 1mol / L NaHCO3 solution + 1.0mL, 0.15mol / L MgCl2·6H2O solution + 0.12mL, 0.5mol / L (NH4)2CO3 solution + 0.18mL, 6mol / L HCl solution; for example, artificial gastric juice (SGF) consists of component A and component B and commercial porcine pepsin (P110928-100g, enzyme activity ≥400unit / mg, purchased from Sigma), wherein component A consists of 0.101mol / L The artificial intestinal fluid is composed of a mixture of KCl, 0.013mol / L KH2PO4, 0.364mol / LNaHCO3, 0.688mol / LNaCl, 0.002mol / L MgCl2, 0.007mol / L(NH4)2CO3 and 0.227mol / L HCl, and the component B is composed of 0.10mL, 0.3mol / L CaCl2·2H2O and 0.90mL, 6mol / L HCl solution; (3) pancreatic enzyme is required in artificial intestinal fluid, and as a mixed enzyme, there is a problem of inconsistent content of different enzymes.

[0006] In summary, many existing analytical methods for predicting rice GI in vitro have significant defects, which can be simply summarized as follows: (1) unreasonable or unnecessary simulated digestion steps lead to cumbersome operations; currently, most in vitro digestion test methods are based on simulating digestion in the mouth, stomach and small intestine; (2) the preparation of digestion simulation fluid is too complicated; the existing digestion simulation fluid contains a variety of inorganic salts, which increases the complexity of the experiment, and uses mixed enzymes, which will introduce new uncertainties.

[0007] Therefore, the development of a relatively simple method for predicting the glycemic index of rice can not only be used for screening low-GI rice varieties, but also for the development of new rice product products.

[0008] Summary of the Invention

[0009] To address the significant differences in in vitro digestion models for different foods, the lack of comparability between methods, and the inherent limitations of existing prediction models, this method aims to develop an in vitro digestion model specifically for rice and specifically for predicting the glycemic index of rice. This method is efficient, simple, accurate, and can specifically measure the GI of rice.

[0010] In the present invention, on the one hand, some unreasonable or unnecessary simulated digestion steps are eliminated; in fact, the inventors have also found that for some food samples, simulating digestion in the human mouth or stomach is unnecessary and should be adjusted according to actual conditions, thereby further streamlining the existing in vitro digestion model; on the other hand, the present invention simplifies the preparation of digestion simulation liquid; in fact, the inventors have found that for some food samples, in a simulation liquid with a relatively simple system, the GI value also has a good correlation with certain digestion parameters.

[0011] The invention provides a method for predicting the glycemic index of rice, comprising the steps of: performing in vitro digestion simulation on the rice to be predicted, obtaining an in vitro digestion parameter AUC-60, and then substituting the AUC-60 value as x into a model y=0.0548x-134.42 to obtain a y value, wherein the y value is the glycemic index (GI) of the rice.

[0012] In one embodiment, the in vitro digestion parameter AUC-60 refers to measuring the reducing sugar release at multiple time points within 0-60 minutes after the start of the enzymatic digestion reaction of the rice to be tested in an in vitro simulated human small intestine, generating a reducing sugar release curve, and converting the reducing sugar release curve into a digestion curve with starch digestibility (value range 0-100) as the vertical axis and reaction time (unit min) as the horizontal axis; performing a definite integral calculation to obtain the area under the in vitro digestion curve (AUC) of the rice to be tested within 0-60 minutes, which is the in vitro digestion parameter AUC-60.

[0013] In one embodiment, the in vitro digestion simulation of the predicted rice is performed by simulating human gastric digestion and human small intestinal digestion; specifically:

[0014] (1) simulating rice cooking to obtain a rice mixture;

[0015] (2) In vitro simulation of human stomach enzymatic digestion reaction activity:

[0016] After cooling the mixed solution obtained in step (1), reacting it with simulated gastric digestive juice to simulate the gastric digestion process of rice in the human body, and terminating the gastric digestion process after the time is reached;

[0017] (3) In vitro simulation of enzymatic digestion reaction activities in the human small intestine:

[0018] The mixed solution obtained in step (2) is reacted with simulated small intestinal digestive fluid to simulate the intestinal digestion process of rice in the human body.

[0019] In one embodiment, the method of simulating rice cooking in step (1) comprises the following steps: adding rice grains into a glass bottle, wherein the rice varieties to be measured are required to be consistent, the grains have intact appearance, and there are as few gaps as possible; adding 3-6 mL of deionized water for every 100-300 mg of rice grains; and boiling the rice grains in water while avoiding damaging the intact grain structure of the rice or causing the rice to stick to the wall.

[0020] In one embodiment, the in vitro simulation of the enzymatic digestion reaction activity of the human stomach in step (2) comprises the following steps: preparing a simulated gastric fluid with a pH of 2.0, wherein the simulated gastric fluid contains a hydrochloric acid concentration of 0.02M-0.03M and a pepsin concentration of 2000-3000U / ml; activating the simulated gastric fluid by a water bath to obtain an activated digestive fluid, wherein the temperature of the activation water bath is 37°C, the activation time is 10-20 minutes, and the rotation speed is 200rpm-400rpm; then, adding the activated digestive fluid to the rice in step (1), performing simulated gastric digestion at a rotation speed of 100-200rpm for 30 minutes, and then terminating the gastric digestion.

[0021] In one embodiment, in step (2), the gastric digestion process is terminated by adding NaOH to change the pH; the concentration of the NaOH solution is 0.02M-0.03M; and the digestion termination time is 5-10 minutes.

[0022] In one embodiment, the in vitro enzymatic digestion reaction activity simulating the human small intestine in step (3) is performed using activated simulated small intestinal digestive fluid at 37° C. and 100-200 rpm for 3 hours; wherein the simulated small intestinal digestive fluid contains sodium acetate buffer and digestive enzymes.

[0023] In one embodiment, the sodium acetate buffer has a concentration of 0.2 M, is prepared by diluting 2 M sodium acetate and 0.2 M acetic acid, pH 6.0; it also contains 1-5 mM CaC12 and 0.2-1 mM MgC12; the digestive enzymes are composed of pancreatic α-amylase and glucosidase, wherein the concentration of pancreatic α-amylase is 1-20 U / ml, and the concentration of glucosidase is 1-10 U / ml; the activated simulated small intestinal digestive fluid refers to treating the simulated small intestinal digestive fluid at 37°C and 200 rpm-400 rpm for 10-20 min.

[0024] In one embodiment, the concentration of the glucoside amylase is 10 U / ml.

[0025] In one embodiment, the in vitro digestion parameters are obtained by fitting the digestion curve based on a mathematical model to obtain the in vitro digestion test parameters; specifically, a reducing sugar release curve is prepared based on the reducing sugar concentration released at each reaction time point during in vitro enzymatic digestion, and the reducing sugar release curve is converted into a digestion curve consisting of starch digestion percentage and reaction time; the definite integral is calculated to obtain the area under the in vitro digestion curve of the rice to be tested, which is the in vitro digestion parameter.

[0026] In one embodiment, the model is constructed by conducting an in vitro digestion simulation experiment on a standard carbohydrate food standard (rice) with a known GI value, and establishing a functional relationship between the glycemic index GI of the standard sample and the in vitro digestion parameter AUC-60.

[0027] In one embodiment, the method for determining the reducing sugar content comprises the following steps:

[0028] At a specific time point, the supernatant is aspirated to terminate the simulated human intestinal digestion of rice and obtain the sample solution to be tested;

[0029] The sample solution to be tested is mixed with the GOPOD reagent in a 96-well plate and incubated. The absorbance value is detected and the reducing sugar content is calculated based on the absorbance value of the standard glucose solution concentration.

[0030] In one embodiment, in the method for determining the reducing sugar content, each time point refers to the enzymatic digestion reaction stage simulating the human small intestine, namely 0, 10, 20, 30, 60, 90, 120, and 180 minutes after adding the enzyme solution.

[0031] In one embodiment, in the method for determining the reducing sugar content, the reagent for terminating the simulated intestinal digestion process of rice by the human body is anhydrous ethanol; optionally, the volume ratio of the test sample liquid to anhydrous ethanol is 1:4-9.

[0032] In one embodiment, in the method for determining the reducing sugar content, the sample solution and the GOPOD reagent are calculated at a volume ratio of 1:20-40; optionally, the incubation time is 10-20 minutes, the incubation temperature is 35-45° C., and the determination condition is absorbance at 510 nm.

[0033] The present invention also provides application of the method in rice variety screening and rice breeding.

[0034] Advantages and effects of the present invention:

[0035] (1) The present invention develops a prediction method specifically for the glycemic index of rice, which is more targeted, more accurate, and more reproducible. The prediction method of the present invention uses rice with known GI as a standard reference to establish a mathematical function relationship between the in vivo GI index and the in vitro simulated digestion parameters, thereby achieving an accurate prediction of the GI index of the rice to be measured, with a linear correlation r above 0.95 and a higher prediction accuracy.

[0036] (2) The present invention reduces the unreasonable or unnecessary steps in the traditional in vitro method for determining the glycemic index of rice, further simplifies the existing in vitro digestion model, and makes the operation simpler; the present invention also simplifies the digestion simulation fluid, is simple to operate, has low cost, and is more suitable for popularization and use.

[0037] (3) The method for determining the glycemic index of rice in vitro of the present invention does not rely too much on professional statistical software for data analysis, which greatly reduces the time for data analysis and fully guarantees the accuracy of the data. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 shows the in vitro digestion curves of eight different rice varieties with known GI values.

[0039] FIG2 is a graph showing the relationship between the GI values ​​of rice varieties with known GI values ​​in Example 1 and their in vitro digestion parameters.

[0040] FIG3 shows the effects of different in vitro digestion model combinations on the in vitro digestion curve of starch of Doongara rice.

[0041] FIG4 shows the relationship between the GI values ​​of rice varieties with known GI values ​​and the predicted GI values ​​in Comparative Example 2.

[0042] FIG5 is the in vitro digestion curve of rice in Comparative Example 2.

[0043] FIG6 is a prediction model constructed after changing the concentration of glucoamylase in Comparative Example 3. DETAILED DESCRIPTION

[0044] Commercial rice varieties with known GI values ​​were selected. Jasime, Doongara, and Akita Komachi rice were purchased from Sunrice (Australia); JK rice was purchased from Jiangnan Rice Road Jiangsu Technology Co., Ltd.; NJ46, NJ9108, and WYG rice were purchased from Jiangsu Nongken Agricultural Development Co., Ltd.; TRF rice was purchased from Anhui Tangzhi Guanjia Technology Co., Ltd.; GQ rice was purchased from Zhejiang Wanxinglai Rice Co., Ltd.; WT rice was purchased from Chengdu Tianjianjun Technology Co., Ltd.; and ST rice was purchased from Tunkmai Biotechnology Research Co., Ltd. Jasime, Akita Komachi, WYG, NJ46, Doongara, JK, GQ, and WT are eight rice varieties that have undergone clinical testing and have clear clinical GI values ​​(see Table 1 below). These rice varieties served as standard rice foods. TRF, NJ9108, and ST rice served as test rice foods.

[0045] Example 1: Establishment of a method specifically for predicting the glycemic index of rice

[0046] Table 1 GI values ​​of different rice varieties

[0047] A method for predicting the glycemic index of rice, comprising the following steps:

[0048] (1) In vitro test simulating rice cooking:

[0049] Weigh 200±10mg of intact rice grains and add them to the reagent bottle;

[0050] Add 2.0 mL of distilled water to the reagent bottle and tighten the cap;

[0051] The mixture was heated in a water bath at 100°C with stirring for 30 min to obtain a simulated cooking sample.

[0052] (2) In vitro tests simulate gastric digestion process:

[0053] 4.0 mL of simulated gastric fluid was added to a reagent bottle and stirred in a 37°C water bath at 200 rpm for 30 min with a magnetic rotor to obtain a simulated gastric digest. The simulated gastric fluid had a pH of 2.0, contained 0.03 M hydrochloric acid, and 2000 U / mL pepsin. The simulated gastric fluid was activated in a water bath at 37°C for 20 min at 400 rpm to obtain an activated digestive solution.

[0054] When the digestion was terminated, 4.0 mL of NaOH solution was added and the reaction was continued for 10 min to obtain a simulated gastric digestion sample.

[0055] (3) In vitro tests simulate intestinal digestion process:

[0056] 10.0 mL of simulated small intestinal digestive fluid was added to a reagent bottle and stirred in a 37°C water bath at 200 rpm with a magnetic rotor for 180 minutes to simulate the human intestinal digestion process of carbohydrate foods, thereby obtaining an intestinal digestion simulator. The simulated small intestinal digestive fluid contained sodium acetate buffer (0.2 M), prepared by diluting 2 M sodium acetate and 0.2 M acetic acid, with a pH of 6.0, and also contained 5 mM CaCl2 and 1 mM MgCl2. The digestive enzymes consisted of pancreatic α-amylase and glucosidase, with the concentrations of pancreatic α-amylase and glucosidase being 20 U / ml and 10 U / ml, respectively. The simulated small intestinal digestive fluid was activated in a water bath at 37°C for 20 minutes at 400 rpm to obtain an activated digestive fluid.

[0057] After the start of the simulated small intestinal digestion stage, the supernatant was collected at 0 min, 10 min, 20 min, 30 min, 60 min, 90 min, 120 min, and 180 min, with 200 μL of sample taken each time. The sample was quickly transferred to a 2 mL centrifuge tube with a cap to which 800 μL of anhydrous ethanol had been added in advance, and the cap was quickly closed and shaken to inactivate the enzyme, which was used as the test sample liquid.

[0058] (4) Digestion product test:

[0059] a. Centrifuge the sample to be tested and add 10 μL of the supernatant to a 96-well plate, with two replicates for each sample.

[0060] b. Prepare standard glucose solutions containing 0.2% w / v sodium azide in the concentration range of 0.2-1 mg / mL. Add 10 μL of the supernatant to a 96-well plate, with two replicates per sample.

[0061] c. Add 200 μL of GOPOD solution (purchased from Megazyme) to a 96-well plate and incubate at 37°C with shaking for 15 minutes. Measure the absorbance at 510 nm. Based on the linear relationship between the concentration of the standard glucose solution and the absorbance, the amount of reducing sugar released by the rice under simulated in vitro digestion at each time point can be determined.

[0062] (5) Fit the digestion curve based on the mathematical model to obtain the in vitro digestion test parameters:

[0063] a. Based on the reducing sugar concentration released by in vitro enzymatic digestion at various reaction time points, a reducing sugar release curve was prepared and converted into a digestion curve consisting of starch digestion percentage and reaction time; wherein, using 8 different rice varieties with known GI values ​​in Table 1, the digestion curve consisting of starch digestion percentage and reaction time obtained by the above method is shown in Figure 2;

[0064] b. Conduct in vitro digestion experiments on standard carbohydrate food samples with known GI values ​​to clarify and establish the functional relationship between the glycemic index (GI) of the standard samples and in vitro digestion parameters;

[0065] The starch digestibility curves for the eight rice varieties listed in Table 1 (Figure 1) were fitted identically, and the area under the in vitro digestibility curve (AUC-60) for each rice variety was calculated using a definite integral over the 0-60 min timeframe. For the eight rice varieties with known GI values, a functional relationship (y = 0.0548x - 134.42) was established between the glycemic index (GI) of the rice sample and the AUC-60, where GI is y and AUC-60 is x. The results are shown in Figure 2.

[0066] As shown in Figure 2, for 8 rice standards, AUC-60 as a key in vitro digestion parameter showed a linear relationship with the GI value, and R 2 =0.9594. This indicates that the relationship between the in vitro digestion parameters obtained by the method of the present invention and the GI value is linearly correlated above 0.95, indicating a higher prediction accuracy.

[0067] Example 2: Application of a method specifically for predicting the glycemic index of rice

[0068] For rice whose GI value is to be measured, an in vitro digestion simulation is performed on the rice to be predicted, referring to the method of Example 1. During the simulated small intestinal digestion stage, a reducing sugar release curve is generated based on the amount of reducing sugar released at different time periods within 0-60 minutes. This is then converted into a digestion curve consisting of starch digestion percentage and reaction time. The area under the in vitro digestion curve (AUC-60) at 60 minutes is obtained, i.e., x. Then, x is substituted into the model y=0.0548x-134.42 to obtain the value y, which is the predicted glycemic index of the rice. Specifically, based on the in vitro digestion parameters of the rice to be measured, the glycemic index of the sample to be measured, i.e., the predicted GI value, is predicted by comparing the functional relationship between the GI value of the standard sample established in Example 1 and its in vitro digestion test parameters.

[0069] As shown in Table 2, the predicted GI values ​​of multiple samples were compared with the clinical GI values. The results showed that the GI values ​​determined by the method of the present invention were within 4% of the GI values ​​obtained in human experiments, indicating that the method of the present invention is accurate and reliable.

[0070] Table 2 AUC-60 and predicted GI values ​​of rice digestibility parameters tested

[0071] Control Example 1: Effects of different in vitro digestion simulation methods on the in vitro digestion curve of starch

[0072] As shown in Figure 3, the effects of different in vitro digestion simulation methods on the in vitro digestion curve of Doongara rice starch were compared.

[0073] The specific method of scheme 1 (mouth + stomach + intestine) is:

[0074] Compared with the method of Example 1, an oral simulation step is added before the gastrointestinal digestion treatment is performed; the specific oral treatment simulation steps are: preparing simulated saliva with a pH of 6.9, wherein the simulated saliva contains a phosphate concentration of 0.1 M and a salivary amylase concentration of 30 U / ml; activating the simulated saliva in a water bath to obtain an activated digestive solution, wherein the temperature of the activation water bath is 37° C., the time is 20 minutes, and the rotation speed is 400 rpm; then, 2 mL of the activated digestive solution is added to the rice in step (1), and simulated oral digestion is performed at a rotation speed of 400 rpm for 10 minutes.

[0075] The specific method of scheme 2 (stomach + intestine) is:

[0076] That is, steps (1)(2)(3) of Example 1

[0077] The specific method of scheme 3 (stomach (no enzyme) + intestine) is:

[0078] On the basis of Example 1, pepsin was not added, and other steps and methods were consistent with Example 1.

[0079] The specific method of scheme 4 (intestinal) is:

[0080] On the basis of Example 1, the enzymatic digestion reaction process of simulating human stomach in vitro was omitted (i.e., step (2) was omitted). The other steps and methods were consistent with those of Example 1.

[0081] As can be seen from Figure 3, the in vitro digestion curves for Schemes 1 and 2 are highly consistent, with the digestion rate for Scheme 2 being significantly higher than that for Scheme 3, and the digestion rate for Scheme 3 being significantly higher than that for Scheme 4. This suggests that the simulated oral stage has little effect on rice digestion, while both gastric acid and pepsin in the simulated gastric digestion stage significantly promote in vitro rice digestion. Therefore, Example 1 of the present invention simplifies the protocol and directly uses a gastric + intestinal simulated digestion method.

[0082] Control Example 2: Comparison of prediction models constructed using different methods of simulating gastrointestinal digestion

[0083] The test method is the same as that of Example 1, except that the in vitro digestion simulation method is based on the classic Englyst method.

[0084] (1) In vitro test simulating rice cooking:

[0085] Weigh 200 ± 10 mg of intact rice grains, add 5 glass beads and 50 mg of guar gum powder, and place them in a reagent bottle. Add 2.0 mL of distilled water to the reagent bottle and tighten the cap.

[0086] (2) In vitro tests simulate intestinal digestion process:

[0087] A mixed enzyme solution was prepared, consisting of 25 mL of sodium acetate buffer and digestive enzymes; the sodium acetate buffer had a concentration of 0.2 M and a pH of 5.2; the digestive enzymes consisted of pancreatic α-amylase and glucosidase, wherein the pancreatic α-amylase had a concentration of 144 U / ml and the glucosidase had a concentration of 15 U / ml; and the simulated small intestinal digestive fluid was activated in a water bath to obtain an activated digestive fluid, wherein the activation water bath temperature was 37° C., the activation time was 20 minutes, and the rotation speed was 160 rpm.

[0088] After the start of the simulated small intestinal digestion stage, the supernatant was collected at 0, 10, 20, 30, 60, 90, 120, and 180 min, with 200 μL of sample taken each time. The sample was quickly transferred to a 2 mL centrifuge tube with a cap to which 800 μL of anhydrous ethanol had been added in advance, and the cap was quickly closed and shaken to inactivate the enzyme, which was used as the test sample liquid.

[0089] The results are shown in Table 3, Figure 4 and Figure 5.

[0090] Table 3 and Figure 4 show the predicted GI values ​​and clinical rice GI values. The results show that the predicted GI values ​​are far from the clinical rice GI values, and there is no discrimination between different samples.

[0091] Table 3 Predicted GI values ​​of rice obtained using the Englyst method

[0092] Control Example 3: Effect of Glucosidase Concentration on Prediction Model Accuracy

[0093] On the basis of Example 1, the concentration of glucoside amylase in step (3) was replaced with 5 U / mL at 10 U / mL, and the other steps and parameters were the same as those in Example 1.

[0094] Referring to the method of Example 1, the relationship between the in vitro digestion parameters of standard rice varieties and their predicted GI values ​​was constructed, y = 0.0451x - 5.9275 (R 2 =0.5311), where x is AUC-60 and y is the predicted GI value.

[0095] The results are shown in Table 4 and Figure 6.

[0096] Table 4 Clinical GI values ​​and AUC-60 of rice after changing conditions

[0097] The results showed that the GI value predicted by AUC-60 after adjusting the glucosidase concentration was far from the clinical GI value, indicating that the glucosidase concentration had a great influence on the accuracy of the model.

[0098] The above embodiment is merely one embodiment of the present invention, and its description is relatively specific and detailed, but it should not be construed as limiting the scope of the present invention. It should be noted that a person of ordinary skill in the art may make various modifications and improvements without departing from the scope of the present invention, and these modifications and improvements fall within the scope of protection of the present invention. Therefore, the scope of protection of the present invention shall be based on the appended claims.

Claims

1. A method for predicting the glycemic index of rice, characterized in that: The method comprises: performing in vitro digestion simulation on the rice to be predicted, obtaining an in vitro digestion parameter AUC-60, and then substituting the AUC-60 value as x into a model y=0.0548x-134.42 to obtain a y value, wherein the y value is the glycemic index of the rice; Among them, in vitro digestion simulation simulates human stomach digestion and human small intestine digestion, but does not simulate oral digestion; The simulated human gastric digestion is performed by using a simulated gastric fluid of pH 2.0, wherein the simulated gastric fluid is composed of 0.02M-0.03M hydrochloric acid and 2000-3000U / ml pepsin; The small intestinal digestive fluid used to simulate human small intestinal digestion consists of 0.2M sodium acetate buffer, digestive enzymes, 1-5mM CaC12 and 0.2-1mM MgC12; the digestive enzymes consist of pancreatic α-amylase and glucosidase, specifically pancreatic α-amylase with a concentration of 1-20U / ml and glucosidase with a concentration of 10U / ml.

2. The method according to claim 1, characterized in that The in vitro digestion parameter AUC-60 refers to measuring the reducing sugar release at multiple time points within 0-60 min after the start of the enzyme digestion reaction of the rice to be tested in vitro simulating the human small intestine, making a reducing sugar release curve, and converting it into a digestion curve with starch digestibility as the vertical axis and reaction time as the horizontal axis; performing definite integral calculation to obtain the area under the in vitro digestion curve within 0-60 min of the rice to be tested, which is the in vitro digestion parameter AUC-60.

3. The method according to claim 1, characterized in that In vitro digestion simulation of the predicted rice was performed, including: (1) simulating rice cooking to obtain a rice mixture; (2) In vitro simulation of human stomach enzymatic digestion reaction activity: After cooling the mixed solution obtained in step (1), reacting it with simulated gastric digestive juice to simulate the gastric digestion process of rice in the human body, and terminating the gastric digestion process after reaching the time; (3) In vitro simulation of enzymatic digestion reaction activities of the human small intestine: The mixed solution obtained in step (2) is reacted with simulated small intestinal digestive fluid to simulate the intestinal digestion process of rice in the human body.

4. The method according to claim 3, characterized in that The method for simulating rice cooking in step (1) comprises the following steps: adding rice grains into a glass bottle, wherein the rice varieties to be measured are required to be consistent, the grains have a complete appearance, and there are no gaps as much as possible; adding 3-6 mL of deionized water for every 100-300 mg of rice grains; and boiling the rice grains while avoiding destroying the complete grain structure of the rice or causing the rice to stick to the wall.

5. The method according to claim 3, characterized in that: The step (2) of simulating the enzymatic digestion reaction of the human stomach in vitro comprises the following steps: preparing simulated gastric juice with a pH of 2.0; activating the simulated gastric juice by water bath to obtain activated digestive juice, wherein the temperature of the activated water bath is 37° C., the time is 10-20 min, and the rotation speed is 200 rpm-400 rpm; then adding the activated digestive juice to the rice of step (1), performing simulated gastric digestion for 30 min at a rotation speed of 100-200 rpm, and then terminating the gastric digestion.

6. The method according to claim 3, characterized in that The in vitro simulation of the enzymatic digestion reaction activity of the human small intestine in step (3) is performed by using activated simulated small intestinal digestive fluid at 37° C. and 100-200 rpm for 3 hours; wherein the simulated small intestinal digestive fluid contains sodium acetate buffer and digestive enzymes.

7. The method according to claim 6, characterized in that The sodium acetate buffer has a concentration of 0.2M, which is prepared by diluting 2M sodium acetate and 0.2M acetic acid, with a pH of 6.0; the activated simulated small intestinal digestive fluid refers to the simulated small intestinal digestive fluid being treated at 37°C and 200rpm-400rpm for 10-20min.

8. The method according to claim 1, characterized in that The model is constructed by conducting an in vitro digestion simulation experiment on a rice standard with a known GI value, and establishing a functional relationship between the glycemic index GI of the standard sample and the in vitro digestion parameter AUC-60.

9. The method according to claim 8, characterized in that When establishing the model, the in vitro digestion simulation method of claim 1 is used, that is, simulating human gastric digestion and simulating human small intestinal digestion.

10. Application of the method according to any one of claims 1 to 9 in rice variety screening and rice breeding.

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