阅读说明：本技术 一种甜味敏感的食物配方及其动物模型的构建方法 (Sweet taste sensitive food formula and construction method of animal model thereof ) 是由 王巧平 胡芸 李安琦 于 2021-08-05 设计创作，主要内容包括：本发明公开了一种改善有机体甜味敏感性的食物营养配方,利用果蝇模型和营养几何框架,首次系统性地评价了食物中主要营养素：蛋白和碳水化合物对甜味敏感性的影响,并找到了最佳的改善甜味敏感性的食物营养配方,其中蛋白质与碳水化合物的质量比为1：(1～2)。本发明还提供了一种高甜味敏感性的动物模型的构建方法以及检测蛋白质和碳水化合的比例对改善果蝇甜味敏感性的方法。(The invention discloses a food nutrition formula for improving organism sweet taste sensitivity, which utilizes a fruit fly model and a nutrition geometric framework to systematically evaluate main nutrients in food for the first time: influence of protein and carbohydrate on sweetness sensitivity, and finding an optimal food nutrition formula for improving sweetness sensitivity, wherein the mass ratio of protein to carbohydrate is 1: (1-2). The invention also provides a construction method of the animal model with high sweet taste sensitivity and a method for detecting the ratio of protein and carbohydrate to improve the sweet taste sensitivity of drosophila melanogaster.)

1. A food nutrition formula for improving the sweetness sensitivity of organisms is characterized in that the mass ratio of protein to carbohydrate in the food nutrition formula is (0.8-1): (1-2.5); the organism is preferably a human, mouse or fruit fly.

2. The food nutritional formula according to claim 1, wherein the mass ratio of protein to carbohydrate in the food nutritional formula is (0.9-1): (1.8-2), more preferably 1: 2.

3. A diet method for improving the sweetness sensitivity of an organism is characterized in that the mass ratio of ingested protein to carbohydrate is (0.8-1): (1-2.5); the organism is preferably a human, mouse or fruit fly.

4. A dietary method according to claim 3, wherein the mass ratio of ingested protein to carbohydrate is (0.9-1): (1.8-2), more preferably 1: 2.

5. A method for constructing an animal model with high sweet taste sensitivity is characterized in that the mass ratio of protein to carbohydrate in a food formula of the animal model is (0.8-1): (1-2.5); the organism is preferably a mouse or a fruit fly.

6. The construction method according to claim 5, wherein the mass ratio of protein to carbohydrate in the food formula of the animal model is (0.9-1): (1.8-2); more preferably 1: 2.

7. A method for detecting the sensitivity of the ratio of protein and carbohydrate to improve the sweetness of drosophila, comprising the steps of:

s1, feeding the fruit flies with food with a target index proportion in groups, wherein the index proportion is the proportion of protein and carbohydrate;

s2, detecting the influence of the food with the target index proportion on the sweet taste sensitivity of the fruit flies through a nose-growing kiss reaction test of the fruit flies.

8. The method as claimed in claim 7, wherein the specific method of the rhinoplasty test of drosophila in step S2 is as follows:

transferring the fruit flies to a container without food;

after 4 hours of starvation, the drosophila were anesthetized and mounted on a glass slide;

after 6 hours of hunger, the drosophila melanogaster front legs were touched twice with drops of sucrose solution, with 1 second interval each time, and the number of individuals producing kiss reaction was recorded and the reaction rate was calculated.

9. The method of claim 8, wherein the drosophila forelegs are dabbed with a gradient concentration drop of sucrose solution.

10. The method of claim 9, wherein the gradient concentration sucrose solutions are 6.25, 12.5, 25, 50, 100, 200, 400, 800(mM) sucrose solutions, respectively.

Technical Field

The invention belongs to the technical field of biology, and particularly relates to a sweet taste sensitive dietary formula and a construction method of an animal model thereof.

Background

Sweetness sensitivity refers to the level of body perception of sweetness. Many foods rich in nutrients and energy are sweet in nature, and therefore the sweet taste sensation is very important for guiding humans and various animals in food selection and energy intake. People and various animals naturally like sweet foods, and the weakening or the lack of sweet taste sensitivity can cause the interest of the body to the sweet foods to be reduced, and the sweet foods are often generated in metabolism-related diseases such as inappetence, anorexia, diabetes and the like and depression of the body, so that the improvement of the sweet taste sensitivity of the body is beneficial to enhancing the appetite of the body, promoting the ingestion and helping to maintain the normal nutrition and metabolism level of the body.

The body's sweetness sensitivity affects the perception of food and, at the same time, can be altered by food. Some studies in humans, mice and drosophila now find that food can significantly up-regulate or down-regulate the sweet taste sensitivity of the body, suggesting the potential for the use of dietary modulation in improving the sweet taste sensitivity of the body. It is not clear what food ingredients affect sweetness sensitivity, nor has it been analyzed how the main nutrients in a food affect sweetness sensitivity in combination with the optimal food formulation that affects sweetness sensitivity.

Drosophila melanogaster (Drosophila melanogaster) is an ideal animal model for studying how food improves sweetness sensitivity and sweet taste control mechanisms. The drosophila melanogaster has a taste sensing system and sweet taste similar to those of human, taste receptor neurons are distributed on the lips, legs and wings of the drosophila melanogaster, sweet taste can be sensed, the long lips of the drosophila melanogaster are generally zoomed in and out of the head, and when the drosophila melanogaster is stimulated by food such as sweet substances, the long lips stretch and open, eating behaviors are clear and visible, so that the physiological structure of the drosophila melanogaster is very convenient for researching sweet taste sensitivity and taste preference. In addition, the drosophila has clear genetic background and rich genetic and neurological research tools, and is a mature, controllable and easily obtained animal with a physiological and pharmacological research model.

Disclosure of Invention

The invention aims to utilize a drosophila model and a nutrition geometric framework to uniformly evaluate the influence of main nutrient proteins and carbohydrates in food on sweetness sensitivity, find out an optimal food formula capable of effectively improving the sweetness sensitivity of organisms and provide a construction method of an animal model with high sweetness sensitivity.

The technical scheme adopted by the invention is as follows:

according to the first aspect of the invention, the food nutrition formula for improving the sweetness sensitivity of the organism is provided, and the mass ratio of protein to carbohydrate in the food nutrition formula is (0.8-1): (1-2.5).

Further, the organism is a human, a mouse or a fruit fly.

Preferably, the mass ratio of protein to carbohydrate in the food nutrition formula is (0.9-1): (1.8-2), more preferably 1: 2.

In a second aspect of the present invention, there is provided a dietary method for improving the sweetness sensitivity of an organism, wherein the mass ratio of ingested protein to carbohydrate is (0.8 to 1): (1-2.5).

Further still, the organism is preferably a human, mouse or fruit fly.

Preferably, the mass ratio of protein to carbohydrate in the food nutrition formula is (0.9-1): (1.8-2), more preferably 1: 2.

The third aspect of the invention provides a construction method of an animal model with high sweet taste sensitivity, wherein the mass ratio of protein to carbohydrate in a food formula of the animal model is (0.8-1): (1-2.5).

Further, the organism is a mouse or a drosophila.

Preferably, the mass ratio of protein to carbohydrate in the food formula of the animal model is (0.9-1): (1.8-2); more preferably 1: 2.

In a fourth aspect of the invention, a method for detecting the ratio of protein and carbohydrate to improve the sweet taste sensitivity of drosophila is provided, which comprises the following steps:

s1, feeding the fruit flies with food with a target index proportion in groups, wherein the index proportion is the proportion of protein and carbohydrate;

s2, detecting the influence of the food with the target index proportion on the sweet taste sensitivity of the fruit flies through a nose-growing kiss reaction test of the fruit flies.

Further, the specific method of the rhinoplasty reaction test of the drosophila melanogaster in step S2 is as follows:

transferring the fruit flies to a container without food;

after 4 hours of starvation, the drosophila were anesthetized and mounted on a glass slide;

after 6 hours of hunger, the drosophila melanogaster front legs were touched twice with drops of sucrose solution, with 1 second interval each time, and the number of individuals producing kiss reaction was recorded and the reaction rate was calculated.

Further, the flies were anesthetized by ice.

Further, the method according to claim 8, wherein the drosophila forelegs are dabbed with a gradient concentration of drops of sucrose solution.

Preferably, the sucrose solutions of gradient concentration are sucrose solutions of 6.25, 12.5, 25, 50, 100, 200, 400, 800(mM), respectively.

Furthermore, the front legs of the drosophila are lightly touched by drops of sucrose solution, and individuals who respond to water are first rejected.

The invention has the beneficial effects that:

the method utilizes the fruit fly model and the nutrition geometric framework to systematically evaluate the main nutrients in the food for the first time: the effect of proteins and carbohydrates on sweetness sensitivity and finding the best food nutritional formula to improve sweetness sensitivity. Wherein the food contains protein and carbohydrate components and the ratio of protein content is 1: (1-2). Specifically, for the model organism drosophila melanogaster, the food contains protein and carbohydrate, the protein content is 80-100 g/L, the carbohydrate content is 100-250g/L, the sweet taste sensitivity of the organism can be obviously increased, and particularly, the food formula with the optimal sweet taste sensitivity enhancing effect is provided with the protein-carbohydrate ratio of 1: 2.

Drawings

FIG. 1 the response of Drosophila to sucrose solutions of different concentrations at different carbohydrate/protein ratios.

FIG. 2 shows the distribution of the half-number of sucrose-sensitive concentrations of Drosophila at different carbohydrate/protein ratios.

FIG. 3 weight and hunger survival time distribution of Drosophila at different carbohydrate/protein ratios.

Detailed Description

In order to clearly understand the technical contents of the present invention, the following embodiments are described in detail with reference to the accompanying drawings. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. The experimental procedures, for which specific conditions are not noted in the following examples, are generally carried out under the conditions described in the conventional conditions or under the conditions recommended by the manufacturers. The various chemicals used in the examples are commercially available.

Examples

Animal model: wild type fruit fly W¹¹¹⁸。

Culturing and amplifying a large amount of the strain of drosophila melanogaster according to a standard process (W)¹¹¹⁸) The constant temperature and humidity incubator for experimental feeding of the fruit flies has the following parameters: temperature: 25 +/-1 ℃; relative humidity 65% ± 5%; day and night circulation: 12h of illumination (7: 30-19: 30)/12h of darkness (19: 30-7: 30).

Experimental reagent:

TABLE 1 Experimental reagent Table

Conventional food formula for raising and breeding experimental drosophila: each 1L of the diet contained water, agar (1%), yeast powder (10%), sucrose (5%), propionic acid (0.6%), 10% m/v methyl paraben-ethanol solution (1.2%).

The sweet taste sensitivity detection method comprises the following steps:

the sweet taste sensitivity of Drosophila melanogaster can be measured by the Probossis Extension Response (PER) (Shiraiwa T, Carlson JR. Probossis Extension Response (PER) assay in Drosophila. journal of visual extensions: JoVE,2007, (3): 193-). PER is an instinctive response of fruit flies extending out of the lips after being specifically stimulated by the outside world and was used in this study to test the sensitivity of fruit flies to sweetness. The level of sweetness sensitivity was quantified by taking the number of tickle responses to each concentration of solution in a group of flies (typically 10-12) and calculating the ratio, and the level of sweetness sensitivity was measured as the ratio, usually expressed as a percentage, after stimulating the flies with a series of graded concentrations of sucrose solution from low to high. A high sensitivity means that a higher proportion of Drosophila produces a kissing response and vice versa.

The effect of protein and carbohydrate sensitivity to sweetness was studied using a nutritional geometry framework:

the total flow is as follows: a plurality of foods with different protein and carbohydrate contents and proportions are prepared, after the flies are fed for a period of time, the sweet taste sensitivity of the flies is detected, and data analysis and response surface mapping are carried out on the results by using R.

In order to investigate the effect of major nutrient proteins (proteins, P) and carbohydrates (C) and the ratio between them (P: C) on the sweet taste sensitivity of Drosophila melanogaster, a two-dimensional nutrient geometric framework food consisting of 28 different Protein and carbohydrate contents and ratios of foods (Lee KP, Simpson SJ, Clissold FJ, et al. Life and reproduction in Drosophila: New instructions from nutritional geometry. proceedings of the National Academy of Sciences of the United States of America,2008,105(7): 2498. 2503.) was used in experiments in which seven Protein and carbohydrate ratios were included (P: C: 0:1, 1:16, 1:8, 1:4, 1:2, 1:1, 9: 90, and four total energy levels (180 g/360). The formulation of these 28 diets is shown in table 2 below.

Table 228 kinds food formula table

Remarking: the yeast contains 45% of protein and 24% of carbohydrate; each 1L of the food contained 1% agar, 1.2% methyl paraben 10% solution, and 0.6% propionic acid.

Healthy male W1118 fruit flies of 4-7 days old, which developed on regular food, were selected, 35 flies in each group, and transferred to 28 groups of food, respectively, and cultured for 6 days. Sweetness sensitivity was performed by the detection-PER test on day seven.

The specific process of performing the sweet taste sensitivity test on the drosophila after each group of food treatment is as follows:

9: 00-10:00 AM: flies were transferred to fly tubes containing no food, only wet paper towels soaked with 2ml water, and started to starve.

1: 00-2:00 PM: after 4h of starvation, the flies were anesthetized with ice and fixed back on a glass slide with nail polish and tweezers. Each row is fixed with 12 pieces, and three rows are fixed. The slide with the adhered fruit fly is then placed in a wet box and returned to the incubator to allow the fruit fly to wake up.

3: 00-4: 00 PM: after 6h of starvation, the same batch of fruit flies was tested for PER within 1 h. The test process is as follows: drosophila were first tested under a microscope with water, and individuals who responded to water were excluded. The drosophila forelegs were then tapped twice with drops of 6.25, 12.5, 25, 50, 100, 200, 400, 800(mM) sucrose solution, each time at 1 second intervals, and the number of individuals producing the kissing reaction was recorded and the rate of reaction calculated. This experiment every group sets up three parallel experiment, and every parallel contains 10 ~ 12 fruit flies, guarantees that every parallel is rejected and is greater than 8 to the individual back quantity of water-sensitive.

This gave data on the effect of 28 different foods of carbohydrate and protein content and ratio on the sweet taste sensitivity of drosophila (statistics see table 3). Finally, PER experimental data were processed by Excel and R (3.6.3) to generate a protein/carbohydrate nutrient geometry-sweetness sensitivity response profile, and the results are shown in fig. 1 and 2.

TABLE 328 Effect of different carbohydrate and protein content and ratio of foods on Drosophila sweetness sensitivity

In addition, the inventor also measured the body weight of drosophila after culturing for 6 days with 28 kinds of food (6 repeated samples, statistical results are shown in table 4) and the hunger-tolerant survival time in the state without food (12 repeated samples, statistical results are shown in table 5), and obtained a protein/carbohydrate nutrition geometry-body weight response surface graph and a protein/carbohydrate nutrition geometry-hunger survival time response surface graph, and the results are shown in fig. 1 and 2.

Table 428 weight of fruit fly after 6 days of food culture

Body weight (g)	M1	M2	M3	M4	M5	M6	Total up to	Mean value of
									1#	0.0077	0.0074	0.008	0.0079	0.0076	0.0078	0.0464	0.007733
2#	0.0081	0.008	0.0082	0.0081	0.008	0.008	0.0484	0.008067
									3#	0.0084	0.0085	0.0083	0.0084	0.0082	0.0087	0.0505	0.008417
4#	0.0086	0.0086	0.0085	0.0083	0.0085	0.0083	0.0508	0.008467
									5#	0.0092	0.0087	0.0085	0.0083	0.0085	0.0085	0.0517	0.008617
6#	0.0086	0.0087	0.0084	0.0083	0.0076	0.0078	0.0494	0.008233
									7#	0.0079	0.0081	0.0081	0.0073	0.007	0.007	0.0454	0.007567
8#	0.0077	0.008	0.0081	0.0079	0.0076	0.0082	0.0475	0.007917
									9#	0.0083	0.0081	0.0082	0.0079	0.0081	0.0081	0.0487	0.008117
10#	0.0082	0.0084	0.0085	0.0084	0.0083	0.0082	0.05	0.008333
									11#	0.0087	0.0088	0.0086	0.0082	0.0086	0.0085	0.0514	0.008567
12#	0.0088	0.0085	0.009	0.009	0.0085	0.0088	0.0526	0.008767
									13#	0.0086	0.0087	0.0085	0.0082	0.0086	0.0085	0.0511	0.008517
14#	0.0081	0.0082	0.0082	0.008	0.0077	0.0076	0.0478	0.007967
									15#	0.008	0.0079	0.0081	0.0078	0.0078	0.0077	0.0473	0.007883
16#	0.0086	0.0081	0.0084	0.0082	0.0084	0.008	0.0497	0.008283
									17#	0.0084	0.0084	0.0084	0.0084	0.0084	0.0082	0.0502	0.008367
18#	0.0086	0.0086	0.0086	0.0086	0.0079	0.008	0.0503	0.008383
									19#	0.0088	0.0083	0.0085	0.0085	0.0082	0.0087	0.051	0.0085
20#	0.0087	0.0086	0.0084	0.0087	0.0085	0.0086	0.0515	0.008583
									22#	0.008	0.0079	0.0079	0.0077	0.008	0.0075	0.047	0.007833
23#	0.0082	0.0084	0.0083	0.0081	0.0082	0.008	0.0492	0.0082
									24#	0.0087	0.0087	0.0085	0.0081	0.0086	0.0084	0.051	0.0085
25#	0.0088	0.0085	0.0087	0.0087	0.0086	0.0087	0.052	0.008667
									26#	0.0087	0.0089	0.0088	0.0075	0.0096	0.0088	0.0523	0.008717

TABLE 5 statistics of starvation-tolerant survival time in the diet free state

As can be seen from FIGS. 1-2, the six gradient sucrose solution stimulations of 6.26-200mM, the food containing different contents and proportions of protein and carbohydrate have a significant differential effect on the sweet taste sensitivity of Drosophila. High protein and low carbohydrate foods contribute to increased sweetness sensitivity. Particularly, when the protein content is 80-100 g/L and the carbohydrate content is within the range of 100-250g/L, the sweetness sensitivity can be obviously increased; furthermore, when the protein content is 90-100 g/L and the carbohydrate content is within the range of 180-200g/L, the sweetness sensitivity can be increased more remarkably; when the ratio of P to C is 1:2, the sweetness sensitivity induced by the food was the strongest (p < 0.001). By way of assistance, it can be seen from figure 3 that there is a relatively optimal weight accumulation and starvation-tolerant survival at this range, i.e. optimal ratio.

The above-mentioned embodiments only express several embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the present invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention.