Detection method for distinguishing endogenous metabolism from exogenous metabolism based on isotope labeling

Abstract

The invention discloses a detection method for distinguishing endogenous and exogenous metabolism based on isotope labeling, and belongs to the technical field of metabolism detection. The detection method comprises the following specific steps: S1, building a system; s2, debugging a void cabin; s3, experiment preparation; s4, performing initial normalization; s5, detecting experimental data; s6, comparing and analyzing data; s7, calculation of endogenous metabolism and exogenous metabolism; s8, verifying the data; the concentration of 13CO2 and the concentration of 12CO2 are detected through one set of sensor, the total concentration of CO2 and the concentration of O2 are detected through the other set of sensor, and whether energy in the metabolic process comes from exogenous energy consumption or endogenous energy consumption calculation can be distinguished in a food isotope labeling mode. The method is beneficial to research and evaluation of exogenous energy consumption and exogenous consumption ratio rules, has great significance in guiding patients and the public to carry out accurate personalized nutrient supplement, and adds a new detection method for existing metabolism detection.

Description

Technical Field

[0001] The present invention relates to the technical field of metabolic detection, in particular to a metabolic detection method for distinguishing endogenous and exogenous substances based on isotope labeling. Background Art

[0002] Accurate measurement of human energy expenditure (EE) helps to study various issues related to how humans achieve or fail to achieve energy balance. Indirect calorimetry refers to a method of calculating human energy expenditure by measuring the amount of oxygen consumed and the amount of carbon dioxide produced by the human body. The principle is that the process of human energy metabolism is always accompanied by the consumption of oxygen and the production of carbon dioxide. Indirect calorimetry is the gold standard method for evaluating human energy metabolism written into clinical guidelines. The three major energy sources of the human body are sugar (carbohydrates), fat and protein. Among them, the human body will only consume protein for metabolic energy supply when it is extremely hungry. The energy obtained by the human body by consuming the sugar and fat in the external food swallowed is called exogenous energy consumption, and the energy obtained by the human body by consuming the sugar and fat in the body itself is called endogenous energy consumption. The metabolic chamber is a device that uses indirect calorimetry to measure the metabolic expenditure of the human body. It measures the oxygen consumption and carbon dioxide production of the human body in the chamber, calculates energy consumption using the Weir formula, and can deduce the proportion of macronutrient consumption.

[0003] When the existing metabolic chamber is performing metabolic data detection, the oxygen consumption and carbon dioxide generation data used are total data, and only the total energy metabolism flux can be calculated, but it is impossible to distinguish whether the energy source is exogenous or endogenous energy supply substances. Because in the process of metabolism, the human body not only consumes the sugar and fat in the food swallowed, but also consumes the body's own sugar and fat at the same time. The CIM model proposed by a joint study of 17 medical institutions including Harvard Medical School and Weill Cornell Medical College in 2021 shows that different individuals in different states have very different absorption and utilization efficiencies for the same nutrition. Therefore, in medical fields such as endocrinology, physical health (obesity), and clinical nutrition, research and evaluation of exogenous energy consumption and the proportion of exogenous consumption are of great significance for guiding patients and the public to supplement nutrients accurately and individually. Based on this, the present invention designs a method for distinguishing endogenous and exogenous metabolism based on isotope labeling to solve the above problems. Summary of the invention

[0004] The purpose of the present invention is to provide a method for distinguishing endogenous and exogenous metabolism detection based on isotope labeling, so as to solve the problem raised in the above background technology that it is necessary to distinguish whether the energy in the metabolic process comes from exogenous energy consumption or endogenous energy consumption.

[0005] To achieve the above object, the present invention provides the following technical solution: a method for distinguishing endogenous and exogenous metabolism detection based on isotope labeling, the specific steps are as follows:

[0006] S1, system construction: build a metabolic detection system, in which a metabolic chamber for the experimental mice to live is set up, and the metabolic data of the experimental mice are calculated by measuring the concentration of O2 gas consumed and CO2 gas produced by the experimental mice in the metabolic chamber;

[0007] S2, empty cabin debugging: In order to ensure the stable operation of the detection system in step S1, it is necessary to perform empty cabin operation of the system built in step S1 to observe whether there are any defects in the operation of the detection system. After eliminating the operation defects, the measured 13 CO 2环境 concentration, 12 CO 2环境 concentration and total CO 2环境 Concentration and O 2环境 The concentration data is stored in the computer of the system as basic data. 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration data to reduce the impact of ambient gases on experimental results;

[0008] S3, Experimental preparation: After the empty chamber debugging in step S2 is completed, the entire test system runs stably and is stored in the ambient gas 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 After the concentration data are obtained, the preparation of experimental mice before the experiment begins, as follows:

[0009] ① Selection of experimental mice: Select female mice with the same weight, growth cycle, body fat percentage, and good health as experimental subjects. Female mice are less active than male mice, which can reduce the impact of different exercise amounts on the experimental results. The number of experimental mice is one less than the number of metabolic chambers;

[0010] ②Choice of experimental feed: divided into normal corn starch feed and 13 C-labeled corn starch diet, wherein 13 C-labeled corn starch diets were divided into 13 C marked diets and fat 13 C-labeled diets can be used to label glucose13 C mark and on the fat 13 C labeling method can be used to distinguish whether exogenous sugars or exogenous fats are consumed in the mouse experiment. 13 C-labeled corn starch diets in terms of glucose and fat 13 The C-labeled content was determined by elemental analysis isotope mass spectrometry;

[0011] S4, initial normalization: the experimental mice selected according to step S3 are placed in metabolic chambers for adaptive life experiments, so that the mice can adapt to the pressure in the metabolic chambers. One mouse is placed in each metabolic chamber, and there is a blank chamber as a control group, in which no mouse is placed. During the test, the experimental mice are fed with normal corn starch diet;

[0012] During the test, the detection system records the internal flow of each metabolic chamber. 13 CO2 sensor and 12 CO2 sensor and total CO2 sensor and total O2 sensor detect 13 CO 2初始 concentration, 12 CO 2初始 concentration and CO 2初始 Concentration and O 2初始 Concentration data;

[0013] The sensor detects 13 CO2 comes from three sources: 13 C-labeled food comes from the mice themselves 13 C and the environment 13 CO2, and the purpose of this system is to measure the utilization of exogenous nutrients by mice, that is, to focus on the utilization of exogenous nutrients from food. 13 CO2, so it needs to be corrected. The correction formula is as follows:

[0014] 13 CO2 食物 = 13 CO2 总 - 13 CO2 小鼠自身 - 13 CO2 环境 ;

[0015] S5, experimental data detection: When step S4 excludes the external environment and the mouse body 13 C and 12 C After the experimental factors, start feeding the marked 13 C-labeled corn starch diet experiment, in which the experimental group was divided into three groups of equal amount for experiment and one group of blank control experiment, to obtain three groups 13 CO2 concentration,12 CO2 concentration, total CO2 concentration and O2 concentration data and a set of blank control experiments 13 CO2 concentration, 12 CO2 concentration and total CO2 concentration and O2 concentration data;

[0016] S6, comparative analysis of data: First, the blank control experiment obtained in step S5 13 CO2 concentration, 12 The CO2 concentration, total CO2 concentration and O2 concentration data are compared with the ambient gas detected in step S2. 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 The concentration data were compared separately to eliminate the influence of the gas in the environment on the experimental group data, and the average difference formula was used to obtain the control experimental group's 13 CO 2平均 concentration, 12 CO 2平均 concentration and total CO 2平均 Concentration and O 2平均 Concentration data;

[0017] According to the control experimental group 13 CO 2平均 concentration, 12 CO 2平均 concentration and total CO 2平均 Concentration and O 2平均 The concentration data were obtained by differential equation to obtain the metabolic parameters of the three groups of experimental mice. 13 CO 2代谢 concentration, 12 CO 2代谢 concentration and total CO 2代谢 Concentration and O 2代谢 Concentration data;

[0018] S7, calculation of endogenous metabolism and exogenous metabolism: First, according to the three groups of experimental mice obtained in step S6, 13 CO 2代谢 concentration, 12 CO 2代谢 concentration and total CO 2代谢 , O 2代谢 Concentration data were used to calculate the proportion of endogenous metabolism and exogenous metabolic consumption in experimental mice;

[0019] S8, data verification: by verifying the data obtained in step S6 ( 13 CO2+ 12The accuracy of this scheme is judged by whether the deviation between the CO2 value and the total CO2 value is less than 1%;

[0020] when( 13 CO2+ 12 When the deviation between the measured CO2 value and the total CO2 value is less than 1%, it indicates that the detection data is accurate, and the data calculated in step S7 is accurate data and can be used as research data;

[0021] when( 13 CO2+ 12 If the deviation between the measured CO2) and the total CO2 value is less than 1%, it means that the detection data is inaccurate, and the data calculated in step S7 cannot be used as research data. At the same time, it is necessary to return to step S2 for a new experiment.

[0022] Preferably, the metabolic detection system described in step S1 includes a plurality of metabolic chambers, and the air inlet ends of the plurality of metabolic chambers are provided with a two-position five-way solenoid valve for controlling the inflow of gas, and the air inlet end of the two-position five-way solenoid valve is provided with a gas buffer chamber, the interior of the gas buffer chamber is a double-chamber design, and one chamber is used for gas buffering at the air inlet end, and the other chamber is used for gas buffering at the exhaust end, and the air inlet end of the gas buffer chamber for the air inlet end is provided with an air intake fan for inputting external gas, and the exhaust end of the plurality of metabolic chambers is also provided with A two-position five-way solenoid valve is used to control gas discharge, and a drying tube for removing water vapor in the exhaust gas is arranged at the output end of the two-position five-way solenoid valve, and an electronic pressure controller EPC for controlling the pressure in the drying tube is arranged at the output end of the drying tube, and the output end of the electronic pressure controller EPC is connected to the air inlet end of the gas buffer chamber for the exhaust end of the gas buffer chamber, and the exhaust end of the gas buffer chamber for the exhaust end of the gas buffer chamber is provided with a ruby ​​gas resistance for controlling the flow rate of the exhaust gas, and the output end of the ruby ​​gas resistance is integrated with 13 CO2 sensor and 12 CO2 sensor, the 13 CO2 sensor and 12 The output end of the CO2 sensor is provided with a total CO2 sensor and a total O2 sensor. 13 CO2 sensor and 12 The data output terminals of the CO2 sensor, the total CO2 sensor, and the total O2 sensor are provided with a control host for collecting the detection data. 13 CO2 sensor and 12 CO2 sensor and total CO2 sensor and total O2 sensor detect 13 CO2 concentration, 12The CO2 concentration, total CO2 concentration and O2 concentration data are collected and sent to the computer wirelessly for data storage and statistics. The gas detected by the total CO2 sensor and the total O2 sensor is directly discharged into the atmosphere.

[0023] Preferably, the empty cabin debugging step in step S2 is that first, multiple groups of metabolic cabins are all set as empty cabins, the power supply of the air intake fan is turned on, so that the air intake fan can run continuously and stably, and then the detection cycle t0 is set by the computer, and at the same time, two groups of two-position five-way solenoid valves are set to synchronously open the air intake and exhaust ends of multiple groups of metabolic cabins in each detection cycle t0, so that each group of metabolic cabins can perform ventilation detection separately, and then during the test, the temperature T of the drying tube and the pressure data P of the electronic pressure controller EPC are synchronously set by the computer to observe whether there are defects in the operation of the detection system;

[0024] The criterion for determining whether the operation is stable is the detected 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 Is the concentration data in a stable fluctuation? Because the gas in the environment is in a stable state, when the detected 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 When the concentration data drifts less than 1%, it indicates that the detection system is running stably. Otherwise, there are defects in the detection system and the hardware in the system needs to be checked and replaced.

[0025] Preferably, the correction step described in step S4 is specifically to collect the detected 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration data, as the environment 13 CO2 and 12 CO2 data. At the same time, for each mouse, there will be a one-hour fasting state before the formal experiment feeding, that is, the t0 state mouse metabolism collection process, detection and storage 13 CO 2小鼠t0 concentration, 12 CO 2小鼠t0 concentration and CO 2小鼠t0 Concentration data, as each mouse's own 13 CO2 and 12 CO2.

[0026] Preferably, the experimental steps described in step S5 are as follows

[0027] The first group was fed with glucose-labeled 13 The second group was fed a diet labeled with corn starch. 13 C-labeled corn starch diet, and the third group was fed with glucose and fat labeled 13 C-labeled corn starch mixed diet, labeled on glucose 13 C-labeled corn starch diets and C-labeled fat 13 The ratio of corn starch feed marked with C is 1:1, and the weight of feed fed to the three groups is the same. After setting the detection cycle detection cycle t2, the drying temperature T2 of the drying tube, and the pressure data P2 of the electronic pressure controller EPC, the experimental test begins;

[0028] First, two groups of two-position five-way solenoid valves are controlled by a computer to control the first group to only feed the glucose marked on the 13 The intake and exhaust of the metabolic chamber of mice fed with C-labeled corn starch were closed for the other experimental groups and the control group. During the detection period t2, 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-a concentration, 12 CO 2-a concentration and total CO 2-a , O 2-a代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation;

[0029] Again, the computer controls two groups of two-position five-way solenoid valves to control the second group to only feed the fat marked on the 13 The intake and exhaust of the metabolic chamber of mice fed with C-labeled corn starch were closed for the other experimental groups and the control group. During the detection period t2, 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-b concentration, 12 CO 2-b concentration and total CO 2-b , O 2-b代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation;

[0030] Again, two groups of two-position five-way solenoid valves are controlled by computers to control the third group of feeding markers on glucose and fat. 13 The intake and exhaust of the metabolic chamber of mice fed with a mixed diet of C-labeled corn starch were closed for the other experimental and control groups. During the detection period t2, 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-c concentration, 12 CO 2-c concentration and total CO 2-c , O 2-c代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation;

[0031] Finally, two groups of two-position five-way solenoid valves are controlled by computer to control the air intake and exhaust of the blank control metabolic chamber, while the air intake and exhaust of other experimental groups are closed. 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-d concentration, 12 CO 2-d concentration and total CO 2-d , O 2-d代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation.

[0032] Preferably, the data comparison step described in step S6 is specifically as follows:

[0033] First, the first group of mice in the metabolic chamber in step S5 13 CO 2-a concentration, 12 CO 2-a concentration and total CO 2-a Concentration data, obtained 13 CO 2-a(t) concentration, 12 CO 2-a(t) concentration and total CO 2-a(t) Concentration data, and then the obtained 13 CO 2-a(t) concentration, 12 CO 2-a(t) concentration and total CO 2-a(t) Concentration data and obtained 13 CO 2-d(t) concentration, 12 CO2-d(t) concentration and total CO 2-d(t) The concentration data were obtained by difference equation to obtain the metabolic 13 CO 2-a代谢(t) concentration, 12 CO 2-a代谢(t) concentration and total CO 2-a代谢(t) Concentration data at different times t, where t0 is set to the time just after eating, that is, t0 reflects the mouse's own 13 CO2, 12 CO2 and total CO2 levels;

[0034] Again, the second group of mice in the metabolic chamber in step S5 13 CO 2-b concentration, 12 CO 2-b concentration and total CO 2-b Concentration data, obtained 13 CO 2-b(t) concentration, 12 CO 2-b(t) concentration and total CO 2-b(t) Concentration data, and then the obtained 13 CO 2-b(t) concentration, 12 CO 2-b(t) concentration and total CO 2-b(t) Concentration data and obtained 13 CO 2-d(t) concentration, 12 CO 2-d(t) concentration and total CO 2-d(t) The concentration data were obtained by difference equation to obtain the metabolic 13 CO 2-b代谢(t) concentration, 12 CO 2-b代谢(t) concentration and total CO 2-b代谢(t) Concentration data at different times t, where t0 is set to the time just after eating, that is, t0 reflects the mouse's own 13 CO2, 12 CO2 and total CO2 levels;

[0035] Finally, the third group of mice in the metabolic chamber in step S5 13 CO 2-c concentration, 12 CO 2-c concentration and total CO 2-c Concentration data, obtained 13 CO 2-c(t) concentration, 12 CO 2-c(t) concentration and total CO 2-c(t)Concentration data, and then the obtained 13 CO 2-c(t) concentration, 12 CO 2-c(t) concentration and total CO 2-c(t) Concentration data and obtained 13 CO 2-d(t) concentration, 12 CO 2-d(t) concentration and total CO 2-d(t) The concentration data were obtained by difference equation to obtain the metabolic 13 CO 2-c代谢(t) concentration, 12 CO 2-c代谢(t) concentration and total CO 2-c代谢(t) The concentration data at different times t, where t0 is set to the time when the mouse is fasting and not eating, that is, t0 reflects the mouse's own 13 CO2, 12 CO2 and total CO2 levels.

[0036] Preferably, the calculation formula for the exogenous metabolism ratio in step S7 is:

[0037] ( 13 CO2%)= 13 CO2 / ( 13 CO2+ 12 CO2)*100%;

[0038] In step S7, endogenous metabolism is divided into endogenous sugar metabolism and endogenous fat metabolism;

[0039] The calculation formula for the proportion of endogenous sugar metabolism is:

[0040] Endogenous glucose metabolism ratio = (TEE-FCR glucose(t) *16.18) / TEE;

[0041] The formula for calculating the proportion of endogenous fat metabolism is:

[0042] Endogenous fat metabolism ratio = (TEE-FCR lipid(t) *39.8) / TEE.

[0043] Compared with the prior art, the present invention has the following beneficial effects:

[0044] The present invention detects the 13 CO2 concentration and 12CO2 concentration, the total CO2 concentration and O2 concentration are detected by another set of sensors, and the isotope labeling in food can distinguish whether the energy in the metabolic process comes from exogenous energy consumption or endogenous energy consumption calculation, which is conducive to the study and evaluation of exogenous energy consumption and the proportion of exogenous consumption. It is of great significance for guiding patients and the general public to supplement nutrients accurately and individually, and adds new detection methods to existing metabolic detection.

[0045] Of course, any product implementing the present invention does not necessarily need to achieve all of the advantages described above at the same time. BRIEF DESCRIPTION OF THE DRAWINGS

[0046] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings required for describing the embodiments will be briefly introduced below. Obviously, the accompanying drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other accompanying drawings can be obtained based on these accompanying drawings without paying creative work.

[0047] Figure 1 It is a schematic diagram of the system module of the present invention;

[0048] Figure 2 The figure is a flow chart of the method of the present invention. DETAILED DESCRIPTION

[0049] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0050] When the existing metabolic chamber is testing metabolic data, the oxygen consumption and carbon dioxide generation data used are total data. This is because during the metabolic process, the human body consumes both sugar and fat from the food it swallows and its own sugar and fat. Therefore, it is necessary to distinguish whether the energy during the metabolic process comes from exogenous energy consumption or endogenous energy consumption.

[0051] See also Figure 1-2 The present invention provides a technical solution: a method for distinguishing endogenous and exogenous metabolism detection based on isotope labeling, the specific steps are as follows:

[0052] S1, system construction: follow the Figure 1A metabolic detection system is constructed in the manner shown, which is used for detecting metabolic data of experimental mice. The metabolic data of the experimental mice are calculated by measuring the O2 gas concentration consumed by the experimental mice during breathing and the CO2 gas concentration produced by the experimental mice. The system includes multiple sets of metabolic chambers, wherein the multiple sets of metabolic chambers are divided into experimental chambers for the experimental mice to live in and a set of blank chambers for control. A two-position five-way solenoid valve for controlling the inflow of gas is provided at the air inlet end of the multiple sets of metabolic chambers, and a gas buffer chamber is provided at the air inlet end of the two-position five-way solenoid valve. The interior of the gas buffer chamber is a double-chamber design, and one chamber is used for gas buffering at the air inlet end, and the other chamber is used for gas buffering at the exhaust end, so as to reduce the direct impact of gas on the electrical components in the detection system and cause damage, thereby reducing maintenance costs. An air intake fan for external gas input is provided at the air intake end of the gas buffer chamber for the air inlet end. The air intake fan is 24V DC brushless fan, the flow rate is 100L / min. A two-position five-way solenoid valve for controlling gas discharge is also provided at the exhaust end of the multiple metabolic chambers, and a drying tube for removing water vapor in the exhaust gas is provided at the output end of the two-position five-way solenoid valve. The drying tube is a resistance wire heating drying tube, and a temperature and humidity sensor is provided inside the drying tube. An electronic pressure controller EPC for controlling the pressure in the drying tube is provided at the output end of the drying tube. The output end of the electronic pressure controller EPC is connected to the air inlet end of the gas buffer chamber for the exhaust end of the gas buffer chamber so that the exhaust gas can be collected. A ruby ​​gas resistance for controlling the flow rate of the exhaust gas is provided at the exhaust end of the gas buffer chamber for the exhaust end of the gas buffer chamber. The ruby ​​gas resistance can be used to control the flow rate of the exhaust gas to be tested at 100ml / min. A sensor for detecting the exhaust gas is integrated at the output end of the ruby ​​gas resistance. 13 CO2, 12 CO2 concentration 13 CO2 sensor and 12 CO2 sensor, and 13 CO2 sensor and 12 The CO2 sensor is a URAS26 infrared sensor that uses non-dispersive infrared absorption. 13 CO2, 12 CO2 is analyzed separately. 13 CO2 sensor and 12 The output end of the CO2 sensor is provided with a total CO2 sensor and a total O2 sensor for detecting the total CO2 concentration and O2 concentration in the exhaust gas. The total CO2 sensor is a single-wavelength, single-beam DNIR carbon dioxide sensor, and the total O2 sensor is a zirconium oxide sensor. 13 CO2 sensor and 12 The data output terminals of the CO2 sensor, the total CO2 sensor and the total O2 sensor are provided with a control host for collecting the detection data, and the control host is connected to the13 CO2 sensor and 12 The CO2 sensor, the total CO2 sensor and the total O2 sensor communicate via the 232 interface to obtain the detected 13 CO2 concentration, 12 The CO2 concentration, total CO2 concentration and O2 concentration data are sent to the computer wirelessly for data storage and statistics. At the same time, the computer can also send the switching command of the two sets of two-position five-way solenoid valves, the heating temperature setting of the drying tube and the pressure data of the electronic pressure controller EPC to the control host. The gas detected by the total CO2 sensor and the total O2 sensor is directly discharged into the atmosphere;

[0053] S2, empty cabin debugging: In order to ensure the stable operation of the detection system in step S1, it is necessary to perform empty cabin operation on the system built in step S1. 13 CO 2环境 concentration, 12 CO 2环境 concentration and total CO 2环境 Concentrated O 2环境 The concentration data is stored in the computer as basic data. 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 Concentration data, reduce the impact of ambient gas on the experimental results, as follows:

[0054] First, multiple metabolic chambers are all set up with empty chambers, and the power supply of the air intake fan is turned on to enable the air intake fan to operate continuously and stably. Then, the detection cycle t0 is set by the computer. At the same time, two sets of two-position five-way solenoid valves are set to synchronously open the air intake and exhaust ends of multiple metabolic chambers in each detection cycle t0, so that each metabolic chamber can perform ventilation detection separately. In the test process, the temperature T of the drying tube and the pressure data P of the electronic pressure controller EPC are synchronously set by the computer to observe whether the detection system has defects during operation (the standard for judging whether the operation is stable is the detected 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 Is the concentration data in a stable fluctuation? Because the gas in the environment is in a stable state, when the detected 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO2环境 Concentration and O 2环境 When the concentration data drifts less than 1%, it indicates that the detection system is running stably. Otherwise, the detection system has defects and the hardware in the system needs to be checked and replaced. After eliminating the operational defects, according to the designed detection cycle t0 and two sets of two-position five-way solenoid valves, the air inlet and exhaust ends of multiple metabolic chambers are opened synchronously in sequence, so that each metabolic chamber can be checked in sequence during the empty chamber process. 13 CO2 sensor and 12 CO2 sensor, total CO2 sensor and total O2 sensor obtain the ambient gas 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 Concentration data, these data are stored in the computer as basic data for backup;

[0055] S3, Experimental preparation: After the empty chamber debugging in step S2 is completed, the entire test system runs stably and is stored in the ambient gas 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 After the concentration data are obtained, the preparation of experimental mice before the experiment begins, as follows:

[0056] ① Selection of experimental mice: Female mice with the same weight, growth cycle, body fat percentage, and good health were selected as experimental subjects. Female mice are less active than male mice, so as to reduce the impact of different exercise amounts on the experimental results. The number of experimental mice is one less than the number of metabolic chambers (because one group of metabolic chambers is a blank chamber without mice);

[0057] ②Choice of experimental feed: divided into normal corn starch feed and 13 C-labeled corn starch diet, wherein 13 C-labeled corn starch diets were divided into 13 C marked diets and fat 13 C-labeled diets can be used to label glucose 13 C mark and on top of fat 13 C labeling method can be used to distinguish whether exogenous sugars or exogenous fats are consumed in the mouse experiment. 13 C-labeled corn starch diets in terms of glucose and fat 13The C-labeled content is determined by elemental analysis isotope mass spectrometry (elemental analysis isotope mass spectrometry is a prior art and will not be described in detail herein);

[0058] S4, initial normalization: the experimental mice selected according to step S3 are placed in a metabolic chamber for an adaptive life experiment, so that the mice can adapt to the pressure in the metabolic chamber, one mouse is placed in each metabolic chamber, and there is a blank chamber as a control group, where no mouse is placed, and the detection cycle t1, the switching logic of the two groups of two-position five-way solenoid valves after each detection cycle t1, the drying temperature T1 of the drying tube, and the pressure data P1 of the electronic pressure controller EPC are set by a computer, and then the air intake fan is connected to the power supply to evenly deliver fresh air to the metabolic chamber. At the same time, during the test, the experimental mice are fed with normal corn starch feed;

[0059] During the test, the computer records the internal flow of each metabolic chamber. 13 CO2 sensor and 12 CO2 sensor and total CO2 sensor and total O2 sensor detect 13 CO 2初始 concentration, 12 CO 2初始 concentration and CO 2初始 Concentration and O 2初始 Concentration data;

[0060] The sensor detects 13 CO2 comes from three sources: 13 C-labeled food comes from the mice themselves 13 C and the environment 13 CO2. The purpose of this system is to measure the utilization of exogenous nutrients by mice, that is, to focus on the 13 CO2, so it needs to be corrected. The correction formula is as follows:

[0061] 13 CO2 总 = 13 CO2 食物 + 13 CO2 小鼠自身 + 13 CO2 环境 (1)

[0062] in 13 CO2 总 By total 13 CO2 sensor detection can be obtained, 13 CO2 小鼠自身 It can be measured when the mouse is fasting (at time t0). 13 CO2 环境Then, for the blank control chamber, that is, the 13 CO2 can be measured when there are no mice in the chamber;

[0063] After transformation, the following formula is obtained:

[0064] 13 CO2 食物 = 13 CO2 总 - 13 CO2 小鼠自身 - 13 CO2 环境 (2)

[0065] The modified experimental process is as follows. Before each exogenous metabolic test experiment, it is necessary to: first collect the detected 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration data, as the environment 13 CO2 and 12 CO2 data. At the same time, for each mouse, before the formal experimental feeding, there will be a one-hour fasting state (t0 state) mouse metabolism collection process, detection and storage 13 CO 2小鼠t0 concentration, 12 CO 2小鼠t0 concentration and CO 2小鼠t0 Concentration data, as each mouse's own 13 CO2 and 12 The above experimental steps are to correct the CO2 in the mouse itself and the environment according to formula (2). 13 The influence of C on the measurement results;

[0066] S5, experimental data detection: When step S4 excludes the external environment and the mouse body 13 C and 12 C After the experimental factors, start feeding the marked 13 C-labeled corn starch diet experiment, in which the experimental group was divided into three groups of equal weight for experiment and one group of blank control experiment;

[0067] The first group was fed with glucose-labeled 13 The second group was fed a diet labeled with corn starch. 13 C-labeled corn starch diet, and the third group was fed with glucose and fat labeled 13 C-labeled corn starch mixed diet, labeled on glucose 13 C-labeled corn starch diets and C-labeled fat13 The ratio of corn starch feed marked with C is 1:1, and the weight of feed fed to the three groups is the same. After setting the detection cycle detection cycle t2, the drying temperature T2 of the drying tube, and the pressure data P2 of the electronic pressure controller EPC, the experimental test begins;

[0068] First, two groups of two-position five-way solenoid valves are controlled by a computer to control the first group to only feed the glucose marked on the 13 The intake and exhaust of the metabolic chamber of mice fed with C-labeled corn starch were closed for the other experimental groups and the control group. During the detection period t2, 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-a concentration, 12 CO 2-a concentration and total CO 2-a , O 2-a代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation;

[0069] Again, the computer controls two groups of two-position five-way solenoid valves to control the second group to only feed the fat marked on the 13 The intake and exhaust of the metabolic chamber of mice fed with C-labeled corn starch were closed for the other experimental groups and the control group. During the detection period t2, 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-b concentration, 12 CO 2-b concentration and total CO 2-b , O 2-b代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation;

[0070] Again, two groups of two-position five-way solenoid valves are controlled by computers to control the third group of feeding markers on glucose and fat. 13 The intake and exhaust of the metabolic chamber of mice fed with a mixed diet of C-labeled corn starch were closed for the other experimental and control groups. During the detection period t2, 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-c concentration, 12 CO2-c concentration and total CO 2-c , O 2-c代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation;

[0071] Finally, two groups of two-position five-way solenoid valves are controlled by computer to control the air intake and exhaust of the blank control metabolic chamber, while the air intake and exhaust of other experimental groups are closed. 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-d concentration, 12 CO 2-d concentration and total CO 2-d , O 2-d代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation;

[0072] S6: Comparative analysis of data: First, the data obtained in step S5 13 CO 2-d concentration, 12 CO 2-d concentration and total CO 2-d The concentration data is consistent with the ambient gas detected in step S2. 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 The concentration data are compared to eliminate the influence of the gas in the environment on the experimental group data, and the average difference formula (which is a prior art and can be selected by using the Excel spreadsheet tool on the computer) is used to obtain 13 CO 2-d平均 concentration, 12 CO 2-d平均 concentration and total CO 2-d平均 Concentration data;

[0073] Again, the first group of mice in the metabolic chamber in step S5 13 CO 2-a concentration, 12 CO 2-a concentration and total CO 2-a Concentration data, obtained 13 CO 2-a(t) concentration, 12 CO 2-a(t) concentration and total CO 2-a(t) Concentration data, and then the obtained 13 CO 2-a(t)concentration, 12 CO 2-a(t) concentration and total CO 2-a(t) Concentration data and obtained 13 CO 2-d(t) concentration, 12 CO 2-d(t) concentration and total CO 2-d(t) The concentration data were obtained by difference equation to obtain the metabolic 13 CO 2-a代谢(t) concentration, 12 CO 2-a代谢(t) concentration and total CO 2-a代谢(t) Concentration data at different times t, where t0 is set to the time just after eating, that is, t0 reflects the mouse's own 13 CO2, 12 CO2 and total CO2 levels;

[0074] Again, the second group of mice in the metabolic chamber in step S5 13 CO 2-b concentration, 12 CO 2-b concentration and total CO 2-b Concentration data, obtained 13 CO 2-b(t) concentration, 12 CO 2-b(t) concentration and total CO 2-b(t) Concentration data, and then the obtained 13 CO 2-b(t) concentration, 12 CO 2-b(t) concentration and total CO 2-b(t) Concentration data and obtained 13 CO 2-d(t) concentration, 12 CO 2-d(t) concentration and total CO 2-d(t) The concentration data were obtained by difference equation to obtain the metabolic 13 CO 2-b代谢(t) concentration, 12 CO 2-b代谢(t) concentration and total CO 2-b代谢(t) Concentration data at different times t, where t0 is set to the time just after eating, that is, t0 reflects the mouse's own 13 CO2, 12 CO2 and total CO2 levels;

[0075] Finally, the third group of mice in the metabolic chamber in step S5 13 CO 2-c concentration, 12 CO 2-cconcentration and total CO 2-c Concentration data, obtained 13 CO 2-c(t) concentration, 12 CO 2-c(t) concentration and total CO 2-c(t) Concentration data, and then the obtained 13 CO 2-c(t) concentration, 12 CO 2-c(t) concentration and total CO 2-c(t) Concentration data and obtained 13 CO 2-d(t) concentration, 12 CO 2-d(t) concentration and total CO 2-d(t) The concentration data were obtained by difference equation to obtain the metabolic 13 CO 2-c代谢(t) concentration, 12 CO 2-c代谢(t) concentration and total CO 2-c代谢(t) The concentration data at different times t, where t0 is set to the time when the mouse is fasting and not eating, that is, t0 reflects the mouse's own 13 CO2, 12 CO2 and total CO2 levels;

[0076] S7, calculation of endogenous metabolism and exogenous metabolism: First, according to the experimental mice obtained in step S6, 13 CO 2-a代谢 concentration, 12 CO 2-a代谢 concentration and total CO 2-a代谢 , total oxygen O 2-a代谢 Concentration data, calculate the proportion of glucose consumed by endogenous metabolism and exogenous metabolism in experimental mice EE 葡萄糖 ;

[0077] Since the S4 step has been carried out on the mice and the environment 13 CO2 is baseline corrected, so the measured 13 CO2 has eliminated the existing 13 CO2 and trace amounts of 13 C, so step S6 13 CO 2-a代谢 concentration, 12 CO 2-a代谢 concentration and total CO 2-a代谢 The concentration data 13 C comes only from food (i.e. exogenous metabolism), 12C comes from unlabeled carbon atoms in food and endogenous metabolism, where endogenous metabolism refers to the oxidation of glycogen and fat stored by the mouse itself. 13 C mark, so 12 CO 2-a代谢 It reflects the sum of endogenous metabolism and unlabeled glucose in food. The total energy consumption is equal to the sum of energy supply from endogenous metabolism and exogenous metabolism. 13 CO 2-a代谢 Total CO 2-a代谢 The ratio is the exogenous metabolism ratio, so the following three formulas are established:

[0078] 12 CO 2-a代谢 = 12 CO 2-a代谢内源 + 12 CO 2-a代谢食物 (3)

[0079] CO2 produced by consuming food = 12 CO 2-a代谢食物 + 13 CO 2-a代谢 (4)

[0080] Total CO2= 12 CO 2-a代谢 + 13 CO 2-a代谢 (5)

[0081] Formula (4) shows that food 13 The C marker ratio%makeratio is as follows:

[0082]

[0083] food 13 C labeling ratio %makeratio only depends on the labeling ratio during food preparation and does not affect the effectiveness and accuracy of this method. It can also be seen from formulas (3) and (4) that 13 CO2 comes only from food, and 12 CO2 comes from various sources, so 13 CO2 is used as a characteristic value to distinguish internal and external metabolism, and the following formula is used: Exogenous metabolism ratio

[0084] ( 13 CO2%)= 13 CO2 / ( 13 CO2 + 12 CO2)*100% (6)

[0085] Since the detected 13 CO2 13C comes from food, and 1 13 C atoms combine with two oxygen atoms to form, so 1 mol 13 CO2 means that there is 1 mol of 13 C is consumed, so it can be 13 Estimate the amount of CO2 in food 13 C consumption, due to different food components, contains 13 The ratio of C may vary. In this example, the formula for glucose is C6H 12 O6, that is, consuming 1 mol of glucose will consume 6 mol of C and produce 6 mol of carbon dioxide. If the labeled carbohydrate components in the food are different, the corresponding coefficients need to be adjusted according to the chemical formula of the components, but the adjustment of the coefficients does not affect the effectiveness of this method.

[0086] Based on the above principles, the formula for calculating the exogenous food consumption rate (Food Consume Rate, FCR) by carbon dioxide production is derived as follows:

[0087] ①Exogenous food consumption (mg / min) = currently monitored 13 CO2 rate (mol / min) *Food marker 13 The proportion of C in total C atoms (%)*total amount of food (mg / mol);

[0088] ②t monitoring at all times 13 The CO2 rate (mol / min) is 13 CO2 rate = (( 13 CO2% (t) - 13 CO2% (t0) )*VCO2 (t) (unit: L / min)) / 22.2966 (unit: L / mol), under standard state STP, 1 mol of CO2 corresponds to a gas volume of 22.2966 liters (L), VCO2 (t) Through the total CO 2-a代谢 The CO2 production rate is obtained by multiplying the concentration by the air flow rate (constant, 5 L / min), unit (L / min);

[0089] ③ Labeling in food 13 The proportion of C in the total C atoms (%) = food 13 C Marker ratio %makeratio (a parameter given during food production, determined according to the food recipe);

[0090] ④ The total amount of food needs to be converted from molar mass to mass in the formula calculation. The formula for glucose in this embodiment is C6H 12O6, corresponding molar mass MW glucose It is 180g / mol.

[0091] Based on principle ①, substituting ②, ③, and ④ into the formula for calculating exogenous food energy consumption (Food Consume Rate, FCR) (unit: mg / min)

[0092]

[0093] The calculation of total energy expenditure (TEE), total glucose consumption rate (TGCR) and total fat consumption rate (TLCR) is based on existing technology (weir formula and Kelly LP and Basset EA (2017)), which can be calculated by VO2 and VCO2, as follows:

[0094] TEE=3.941*VO 2(t) +1.106*VCO 2(t) (8)

[0095] TGCR (t) = -3,226 * VO 2(t) + 4.585 * VCO 2(t) - 0.461 *0.066 (9)

[0096] TLCR (t) = 1.695 * VO 2(t) -1.701 * VCO 2(t) - 0.319 * 0.066 (10)

[0097] Among them, VO 2(t) Through the total O 2-a代谢 The O2 consumption rate, VCO, is obtained by multiplying the concentration by the air flow rate (constant, 5 L / min). 2(t) Through the total CO 2-a代谢 The CO2 production rate is obtained by multiplying the concentration by the air flow rate (constant, 5 L / min);

[0098] Therefore, the endogenous sugar metabolism rate is:

[0099] Endogenous glucose metabolism rate = TGCR (t) -FCR glucose(t) (11)

[0100] Endogenous glucose metabolism accounts for the proportion of total energy = (TEE-FCR glucose(t)*16.18) / TEE(12)

[0101] Among them, 16.18 is the glucose combustion equivalent, that is, consuming 1g of glucose produces 16.18kJ of energy. The reference value comes from "Physiology (9th Edition)".

[0102] This can be done through 13 Detection of CO2 calculates and distinguishes endogenous metabolism and exogenous metabolism, and their real-time rate and energy supply proportion in human energy metabolism.

[0103] exist 13 C-labeled fat and 13 The calculation principle is the same when C is used to label mixed meals, except that the molar mass of the label and the food formula affect 13 The proportion of C is different, so the following formula can be obtained through a similar calculation process;

[0104] According to the experimental mice obtained in step S6 13 CO 2-b代谢 concentration, 12 CO 2-b代谢 concentration and total CO 2-b代谢 The concentration data was used to calculate the proportion of fat consumed by endogenous metabolism and exogenous metabolism in experimental mice. 脂肪 ;

[0105] When labeling fat:

[0106]

[0107] In this embodiment, the labeled fat is palmitate, chemical formula C 16 H 32 O2, so the molar mass MW lipid The molar mass of the marked substance needs to be substituted according to the actual situation, which does not affect the effectiveness of the method of the present invention.

[0108] Endogenous fat metabolism rate = TLCR (t) -FCR lipid(t) (14)

[0109] Endogenous fat metabolism accounts for the proportion of total energy = (TEE-FCR lipid(t) *39.8) / TEE(15)

[0110] Among them, 39.8 is the fat burning equivalent, that is, consuming 1g of fat produces 39.8kJ of energy. The reference value comes from "Physiology (9th Edition)".

[0111] Next, the mixed ratio of endogenous metabolism and exogenous metabolism of glucose and fat consumed by the experimental mice EE is calculated according to the ratio of endogenous metabolism and exogenous metabolism of glucose consumed by the experimental mice obtained in step S6. 葡萄糖和脂肪混合 ;

[0112] When marking mixed meals:

[0113] Exogenous sugar consumption rate:

[0114]

[0115] Meal Mixing Ratio(16)

[0116] Exogenous fat consumption rate:

[0117]

[0118] Endogenous glucose metabolism rate = TGCR (t) -FCR glucose(t) (18)

[0119] Endogenous fat metabolism rate = TLCR (t) -FCR lipid(t) (19)

[0120] Endogenous glucose metabolism accounts for the proportion of total energy = (TEE-FCR glucose(t) *17.2) / TEE(20)

[0121] Endogenous glucose metabolism accounts for the proportion of total energy = (TEE-FCR lipid(t) *39.8) / TEE(21);

[0122] S8: Data verification: By verifying the data obtained in step S6 ( 13 CO2+ 12 The accuracy of this scheme is judged by whether the deviation between the CO2 value and the total CO2 value is less than 1%;

[0123] when( 13 CO2+ 12 When the deviation between the measured CO2 value and the total CO2 value is less than 1%, it indicates that the detection data is accurate, and the data calculated in step S7 is accurate data and can be used as research data;

[0124] when( 13 CO2+ 12 If the deviation between the measured CO2) and the total CO2 value is less than 1%, it means that the detection data is inaccurate, and the data calculated in step S7 cannot be used as research data. At the same time, it is necessary to return to step S2 for a new experiment.

[0125] From the above derivation formula, it can be seen that the calculation of distinguishing exogenous and exogenous in this scheme is based on isotope labeling 13 C, and the main influencing variables of subsequent calculation accuracy are based only on the detected 13 CO2, 12 CO2. 12 CO2 is ubiquitous in the environment and comes from various sources. 13 The concentration of CO2 in the environment and in organisms such as mice is very low, so the method is based on 13 The CO2 detection method has high accuracy and sensitivity.

[0126] The method of the embodiment of the present invention detects the 13 CO2 concentration and 12 CO2 concentration, the total CO2 concentration is detected by another set of sensors. If ( 13 CO2+ 12 CO2) is equal to the total CO2 concentration, which means that during the detection process, 13 The concentration detection of CO2 concentration did not result in missing or erroneous concentration signals. At this time, we believe that 13 CO2 and 12 The correct CO2 ratio and total amount collected are conducive to the study and evaluation of exogenous energy consumption and the proportion of exogenous consumption. It is of great significance for guiding patients and the general public to supplement nutrients accurately and individually, and it also adds new detection methods to existing metabolic detection.

[0127] In the description of this specification, the description with reference to the terms "one embodiment", "example", "specific example", etc. means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representation of the above terms does not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described can be combined in any one or more embodiments or examples in a suitable manner.

[0128] The preferred embodiments of the present invention disclosed above are only used to help illustrate the present invention. The preferred embodiments do not describe all the details in detail, nor do they limit the invention to the specific implementation methods described. Obviously, many modifications and changes can be made according to the content of this specification. This specification selects and specifically describes these embodiments in order to better explain the principles and practical applications of the present invention, so that those skilled in the art can understand and use the present invention well. The present invention is limited only by the claims and their full scope and equivalents.

Claims

1. A method for distinguishing endogenous and exogenous metabolism based on isotope labeling, characterized in that: The specific steps are as follows: S1, system construction: build a metabolic detection system, in which a metabolic chamber for the experimental mice to live is set up, and the metabolic data of the experimental mice are calculated by measuring the concentration of O2 gas consumed and CO2 gas produced by the experimental mice in the metabolic chamber; S2, empty cabin debugging: the system built in step S1 is operated in an empty cabin of the metabolic cabin. After eliminating the operating defects, the measured 13 CO 2环境 concentration, 12 CO 2环境 concentration and total CO 2环境 Concentration and O 2环境 The concentration data is stored in the system as basic data. 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration data to reduce the impact of ambient gases on experimental results; S3, Experimental preparation: After the empty cabin debugging in step S2 is completed, the entire test system runs stably, and the preparations before the process experiment mouse experiment begin, as follows: ① Selection of experimental mice: Select healthy female mice with the same weight, growth cycle, body fat percentage as the experimental subjects. The number of experimental mice should be one less than the number of metabolic chambers. ②Choice of experimental feed: divided into normal corn starch feed and 13 C-labeled corn starch diet, wherein 13 C-labeled corn starch diets were divided into 13 C marked diets and fat 13 C marked diet, 13 C-labeled corn starch diets in terms of glucose and fat 13 The C-labeled content was determined by elemental analysis isotope mass spectrometry; S4, initial normalization: the experimental mice selected according to step S3 are placed in metabolic chambers for adaptive life experiments, one mouse is placed in each metabolic chamber, and there is a blank chamber as a control group, where no mice are placed. During the test, the experimental mice are fed with normal corn starch diet; During the test, the detection system records the internal flow of each metabolic chamber. 13 CO2 sensor and 12 CO2 sensor and total CO2 sensor and total O2 sensor detect 13 CO 2初始 concentration, 12 CO 2初始 concentration and CO 2初始 Concentration and O 2初始 Concentration data; S5, experimental data detection: When step S4 excludes the external environment and the mouse body 13 C and 12 C After the experimental factors, start feeding the marked 13 C-labeled corn starch diet experiment, in which the experimental group was divided into three groups of equal amount for experiment and one group of blank control experiment, to obtain three groups 13 CO2 concentration, 12 CO2 concentration, total CO2 concentration and O2 concentration data and a set of blank control experiments 13 CO2 concentration, 12 CO2 concentration and total CO2 concentration and O2 concentration data; S6: Comparative analysis of data: First, the blank control experiment obtained in step S5 13 CO2 concentration, 12 The CO2 concentration, total CO2 concentration and O2 concentration data are compared with the ambient gas detected in step S2. 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 The concentration data were compared separately to eliminate the influence of the gas in the environment on the experimental group data, and the average difference formula was used to obtain the control experimental group's 13 CO 2平均 concentration, 12 CO 2平均 concentration and total CO 2平均 Concentration and O 2平均 Concentration data; According to the control experimental group 13 CO 2平均 concentration, 12 CO 2平均 concentration and total CO 2平均 Concentration and O 2平均 The concentration data were obtained by differential equation to obtain the metabolic parameters of the three groups of experimental mice. 13 CO 2代谢 concentration, 12 CO 2代谢 concentration and total CO 2代谢 Concentration and O 2代谢 Concentration data; S7, calculation of endogenous metabolism and exogenous metabolism: First, according to the three groups of experimental mice obtained in step S6, 13 CO 2代谢 concentration, 12 CO 2代谢 concentration and total CO 2代谢 , O 2代谢 Concentration data were used to calculate the proportion of endogenous metabolism and exogenous metabolic consumption in experimental mice; S8: Data verification: By verifying the data obtained in step S6 ( 13 CO2+ 12 The accuracy is judged by whether the deviation between the measured CO2 and the total CO2 value is less than 1%.

2. The method for distinguishing endogenous and exogenous metabolism based on isotope labeling according to claim 1, characterized in that: The sensor in S4 detects 13 CO2 comes from three sources: 13 C-labeled food comes from the mice themselves 13 C and the environment 13 CO2, and the purpose of this system is to measure the utilization of exogenous nutrients by mice, that is, to focus on the utilization of exogenous nutrients from food. 13 CO2, so it needs to be corrected. The correction formula is as follows: 13 CO2 食物 = 13 CO2 总 - 13 CO2 小鼠自身 - 13 CO2 环境 。 3. The method for distinguishing endogenous and exogenous metabolism based on isotope labeling according to claim 1, characterized in that: S8 data verification specifically includes: when( 13 CO2+ 12 When the deviation between the measured CO2 value and the total CO2 value is less than 1%, it indicates that the detection data is accurate, and the data calculated in step S7 is accurate data and can be used as research data; when( 13 CO2+ 12 If the deviation between the measured CO2) and the total CO2 value is less than 1%, it means that the detection data is inaccurate, and the data calculated in step S7 cannot be used as research data. At the same time, it is necessary to return to step S2 for a new experiment.

4. The method for distinguishing endogenous and exogenous metabolism based on isotope labeling according to claim 1, characterized in that: The metabolic detection system described in step S1 includes a plurality of metabolic chambers, and the air inlet ends of the plurality of metabolic chambers are provided with a two-position five-way solenoid valve for controlling the inflow of gas, and the air inlet ends of the two-position five-way solenoid valves are provided with a gas buffer chamber, the interior of the gas buffer chamber is a double-chamber design, and one chamber is used for gas buffering at the air inlet end, and the other chamber is used for gas buffering at the exhaust end, and the air inlet end of the gas buffer chamber for the air inlet end is provided with an air intake fan for inputting external gas, and the exhaust ends of the plurality of metabolic chambers are also provided with a fan for A two-position five-way solenoid valve for controlling gas discharge is provided, and a drying tube for removing water vapor in the exhaust gas is provided at the output end of the two-position five-way solenoid valve, and an electronic pressure controller EPC for controlling the pressure in the drying tube is provided at the output end of the drying tube, and the output end of the electronic pressure controller EPC is connected to the air inlet end of the gas buffer chamber for the exhaust end of the gas buffer chamber, and the exhaust end of the gas buffer chamber for the exhaust end of the gas buffer chamber is provided with a ruby ​​gas resistance for controlling the flow rate of the exhaust gas, and the output end of the ruby ​​gas resistance is integrated with 13 CO2 sensor and 12 CO2 sensor, the 13 CO2 sensor and 12 The output end of the CO2 sensor is provided with a total CO2 sensor and a total O2 sensor. 13 CO2 sensor and 12 The data output terminals of the CO2 sensor, the total CO2 sensor, and the total O2 sensor are provided with a control host for collecting the detection data. 13 CO2 sensor and 12 CO2 sensor and total CO2 sensor and total O2 sensor detect 13 CO2 concentration, 12 The CO2 concentration, total CO2 concentration and O2 concentration data are collected and sent to the computer wirelessly for data storage and statistics. The gas detected by the total CO2 sensor and the total O2 sensor is directly discharged into the atmosphere.

5. The method for distinguishing endogenous and exogenous metabolism based on isotope labeling according to claim 4, characterized in that: The empty cabin debugging procedure described in step S2 is as follows: first, multiple groups of metabolic cabins are all set as empty cabins, the power supply of the air intake fan is turned on, so that the air intake fan can run continuously and stably, and then the detection cycle t0 is set by the computer, and at the same time, two groups of two-position five-way solenoid valves are set to synchronously open the air intake and exhaust ends of multiple groups of metabolic cabins in each detection cycle t0, so that each group of metabolic cabins can perform ventilation detection separately, and then during the test, the temperature T of the drying tube and the pressure data P of the electronic pressure controller EPC are synchronously set by the computer to observe whether the detection system has defects during operation; The criterion for determining whether the operation is stable is the detected 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 Is the concentration data in a stable fluctuation? Because the gas in the environment is in a stable state, when the detected 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration and O 2环境 When the concentration data drifts less than 1%, it indicates that the detection system is running stably. Otherwise, there are defects in the detection system and the hardware in the system needs to be checked and replaced.

6. The method for distinguishing endogenous and exogenous metabolism based on isotope labeling according to claim 4, characterized in that: The correction step described in step S4 is specifically to collect the detected 13 CO 2环境 concentration, 12 CO 2环境 concentration and CO 2环境 Concentration data, as the environment 13 CO2 and 12 CO2 data. At the same time, for each mouse, there will be a one-hour fasting state before the formal experiment feeding, that is, the t0 state mouse metabolism collection process, detection and storage 13 CO 2小鼠t0 concentration, 12 CO 2小鼠t0 concentration and CO 2小鼠t0 Concentration data, as each mouse's own 13 CO2 and 12 CO2.

7. The method for distinguishing endogenous and exogenous metabolism based on isotope labeling according to claim 4, characterized in that: The experimental steps described in step S5 are as follows The first group was fed with glucose-labeled 13 The second group was fed a diet labeled with corn starch. 13 C-labeled corn starch diet, and the third group was fed with glucose and fat labeled 13 C-labeled corn starch mixed diet, labeled on glucose 13 C-labeled corn starch diets and C-labeled fat 13 The ratio of corn starch feed marked with C is 1:1, and the weight of feed fed to the three groups is the same. After setting the detection cycle detection cycle t2, the drying temperature T2 of the drying tube, and the pressure data P2 of the electronic pressure controller EPC, the experimental test begins; First, two groups of two-position five-way solenoid valves are controlled by a computer to control the first group to only feed the glucose marked on the 13 The intake and exhaust of the metabolic chamber of mice fed with C-labeled corn starch were closed for the other experimental groups and the control group. During the detection period t2, 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-a concentration, 12 CO 2-a concentration and total CO 2-a , O 2-a代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation; Again, the computer controls two groups of two-position five-way solenoid valves to control the second group to only feed the fat marked on the 13 The intake and exhaust of the metabolic chamber of mice fed with C-labeled corn starch were closed for the other experimental groups and the control group. During the detection period t2, 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-b concentration, 12 CO 2-b concentration and total CO 2-b , O 2-b代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation; Again, two groups of two-position five-way solenoid valves are controlled by computers to control the third group of feeding markers on glucose and fat. 13 The intake and exhaust of the metabolic chamber of mice fed with a mixed diet of C-labeled corn starch were closed for the other experimental and control groups. During the detection period t2, 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-c concentration, 12 CO 2-c concentration and total CO 2-c , O 2-c代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation; Finally, two groups of two-position five-way solenoid valves are controlled by computer to control the air intake and exhaust of the blank control metabolic chamber, while the air intake and exhaust of other experimental groups are closed. 13 CO2 sensor and 12 The CO2 sensor and the total CO2 sensor detected the 13 CO 2-d concentration, 12 CO 2-d concentration and total CO 2-d , O 2-d代谢 Concentration data, the detected data is sent to the computer through the control host and saved to facilitate the next step of comparison and calculation.

8. The method for distinguishing endogenous and exogenous metabolism based on isotope labeling according to claim 7, characterized in that: The data comparison step described in step S6 is as follows: First, the first group of mice in the metabolic chamber in step S5 13 CO 2-a concentration, 12 CO 2-a concentration and total CO 2-a Concentration data, obtained 13 CO 2-a(t) concentration, 12 CO 2-a(t) concentration and total CO 2-a(t) Concentration data, and then the obtained 13 CO 2-a(t) concentration, 12 CO 2-a(t) concentration and total CO 2-a(t) Concentration data and obtained 13 CO 2-d(t) concentration, 12 CO 2-d(t) concentration and total CO 2-d(t) The concentration data were obtained by difference equation to obtain the metabolic 13 CO 2-a代谢(t) concentration, 12 CO 2-a代谢(t) concentration and total CO 2-a代谢(t) Concentration data at different times t, where t0 is set to the time just after eating, that is, t0 reflects the mouse's own 13 CO2, 12 CO2 and total CO2 levels; Again, the second group of mice in the metabolic chamber in step S5 13 CO 2-b concentration, 12 CO 2-b concentration and total CO 2-b Concentration data, obtained 13 CO 2-b(t) concentration, 12 CO 2-b(t) concentration and total CO 2-b(t) Concentration data, and then the obtained 13 CO 2-b(t) concentration, 12 CO 2-b(t) concentration and total CO 2-b(t) Concentration data and obtained 13 CO 2-d(t) concentration, 12 CO 2-d(t) concentration and total CO 2-d(t) The concentration data were obtained by difference equation to obtain the metabolic 13 CO 2-b代谢(t) concentration, 12 CO 2-b代谢(t) concentration and total CO 2-b代谢(t) Concentration data at different times t, where t0 is set to the time just after eating, that is, t0 reflects the mouse's own 13 CO2, 12 CO2 and total CO2 levels; Finally, the third group of mice in the metabolic chamber in step S5 13 CO 2-c concentration, 12 CO 2-c concentration and total CO 2-c Concentration data, obtained 13 CO 2-c(t) concentration, 12 CO 2-c(t) concentration and total CO 2-c(t) Concentration data, and then the obtained 13 CO 2-c(t) concentration, 12 CO 2-c(t) concentration and total CO 2-c(t) Concentration data and obtained 13 CO 2-d(t) concentration, 12 CO 2-d(t) concentration and total CO 2-d(t) The concentration data were obtained by difference equation to obtain the metabolic 13 CO 2-c代谢(t) concentration, 12 CO 2-c代谢(t) concentration and total CO 2-c代谢(t) The concentration data at different times t, where t0 is set to the time when the mouse is fasting and not eating, that is, t0 reflects the mouse's own 13 CO2, 12 CO2 and total CO2 levels.

9. The method for distinguishing endogenous and exogenous metabolism based on isotope labeling according to claim 1, characterized in that: The calculation formula for the exogenous metabolism ratio in step S7 is ( 13 CO2%)= 13 CO2 / ( 13 CO2+ 12 CO2)*100%; Among them, the detected 13 CO2 13 C comes from food, representing the proportion of exogenous, 12 CO2 is based on the oxidation of the mouse's own stored glycogen and fat, not 13 The C mark represents the endogenous proportion.

10. The method for distinguishing endogenous and exogenous metabolism based on isotope labeling according to claim 9, characterized in that: In step S7, endogenous metabolism is divided into endogenous sugar metabolism and endogenous fat metabolism; The calculation formula for the proportion of endogenous glucose metabolism is: Endogenous glucose metabolism proportion = (TEE-FCR glucose(t) *16.18) / TEE And TEE is total energy expenditure, FCR glucose(t) is the energy consumption of sugar in exogenous food, 16.18 is the glucose combustion equivalent, that is, consuming 1g of glucose produces 16.18kJ of energy; The formula for calculating the endogenous fat metabolism ratio is: Endogenous fat metabolism ratio = (TEE-FCR lipid(t) *39.8) / TEE And TEE is total energy expenditure, FCR lipid(t) is the fat energy consumption in exogenous food, and 39.8 is the fat burning equivalent, that is, consuming 1g of fat will produce 39.8kJ of energy.

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