PBTK-TD model of rat acute oral exposure rice yeast acid and construction method thereof

Abstract

The invention provides a method for constructing a PBTK-TD model for acute oral exposure of rice yeast acid in rats, which comprises the following steps: S1, taking rats orally taking rice yeast acid as test samples, and measuring the concentration of rice yeast acid in plasma and tissue samples of the rats; s2, determining an overall structure of the model, wherein the overall structure comprises a core physiological compartment and a target spot of a toxic effect dynamics module; s3, effective parameters are collected; S4, a model program composed of multivariate differential equations is established; and S5, optimizing model program parameters, and constructing a rice yeast bacterial acid PBTK-TD model. The invention also provides a model constructed by the method, which is used for quantitatively evaluating the dynamic change of the rice yeast acid in the body and the generated toxic effect, knowing the harm degree of the rice yeast acid exposed by the body, and achieving the technical purpose of performing health risk evaluation on the rice yeast acid. And a new technical support is provided for a supervision department to formulate a safe dosage standard of the rice yeast acid.

Description

Technical Field

[0001] The present invention relates to the field of food safety, specifically to the assessment of biotoxins and the association of in vivo and in vitro exposures, and particularly to a method for constructing a toxicokinetics and toxicodynamics model (PBTK-TD model) for rats acutely exposed to bongkrekic acid via oral administration. Background Art

[0002] Bongkrekic acid (BA) is a bacterial biotoxin produced by Burkholderia gladioli subsp. cocovenenans in foods, and is the main cause of food poisoning caused by starch products, cereal fermentation products, potato products, and deteriorated tremella, etc. Bongkrekic acid is a small molecule substance of fatty acids, highly toxic and heat-resistant, and difficult to be removed by ordinary cooking methods. The level of exposure of this toxin in the body directly affects the degree of damage to human tissues and organs. Currently, there is no good model for the human health risk assessment of bongkrekic acid.

[0003] Physiological toxicokinetics (PBTK) can quantitatively describe the processes of absorption, distribution, metabolism, and excretion of compounds and biotoxins in organisms, predict the changes in the concentrations of compounds and biotoxins in organisms over time, and associate the environmental exposure concentrations and target concentrations of compounds and biotoxins. It is used for extrapolation of in vivo - in vitro toxicity test data, cross-species extrapolation, etc., and plays an important role in the ecological risk assessment and management of compounds and biotoxins. On this basis, combined with toxicodynamics (TD), it can predict the dose of the toxin prototype or its active metabolite in the target tissue, and use physiological effective parameters to construct a dose-response relationship closer to the actual metabolic process, making the extrapolation of animal experiment results to humans more credible. It can predict the internal exposure dose and toxic effect level of the organism according to the external exposure dose, and is an important technical means in risk assessment.

[0004] Therefore, there is an urgent need for a method for assessing the health risk of bongkrekic acid, which can quantitatively evaluate the dynamic changes of bongkrekic acid in the organism and the resulting toxic effects, understand the harm degree of the organism exposed to bongkrekic acid, provide key technical support for regulatory authorities to formulate bongkrekic acid safety limit standards based on scientific evidence, and further improve the food safety risk monitoring and early warning system. Summary of the Invention

[0005] The object of the present invention is to propose a method for constructing a PBTK-TD model for rats acutely exposed to bongkrekic acid via oral administration, which is used to construct the PBTK-TD model of bongkrekic acid, for quantitatively evaluating the dynamic changes of bongkrekic acid in the organism and the resulting toxic effects, so as to achieve the technical purpose of health risk assessment of bongkrekic acid.

[0006] The object of the present invention can be achieved by the following technical solutions:

[0007] A method for constructing a PBTK-TD model for acute oral exposure of rats to bongkrekic acid, comprising the following steps:

[0008] S1: Using rats orally administered bongkrekic acid as test samples, collecting plasma and tissue samples of the test samples, and measuring the concentration of bongkrekic acid in the samples for analyzing the kinetic processes of absorption, distribution, metabolism, and excretion of bongkrekic acid in rats;

[0009] S2: Establishing the overall structure of the model, including the composition structure of the core physiological compartments and the targets of the toxicokinetics module;

[0010] S3: Collecting effective parameters: A. Physiological parameters of rats; B. Toxicokinetic parameters of bongkrekic acid, including absorption rate constant k a , metabolic rate constant K m , tissue-blood partition coefficient; C. Toxicokinetic parameters of bongkrekic acid, including the binding rate constant kon and dissociation rate constant k off of ANT, and median effective concentration EC 50 ;

[0011] S4: Establishing a model program, and according to the law of conservation of mass, writing differential equations for each physiological compartment to describe the concentration changes of bongkrekic acid in each compartment, obtaining a model program composed of multiple differential equations;

[0012] S5: Conducting sensitivity analysis and optimization of the model program parameters to construct a bongkrekic acid PBTK-TD model.

[0013] Furthermore, the method for constructing a PBTK-TD model of the present invention further includes using the bongkrekic acid PBTK-TD model to calculate the dose-response relationship through the model, evaluating the quantitative relationship between the exposure dose and the toxic effect, and using the model to predict the distribution and toxic effect of bongkrekic acid in the body at different doses.

[0014] Optimization scheme: In S2 of the method for constructing a PBTK-TD model of the present invention, determining the core physiological compartments: The physiological compartments included in the model are mainly the heart, liver, kidneys, and intestines, and the compartments are interconnected by blood flow to simulate the absorption, distribution, metabolism, and excretion processes of bongkrekic acid toxin in the test samples. And determining the targets of the toxicokinetics module: For bongkrekic acid, its main toxic target is ANT, and it inhibits ATP generation by binding to ANT.

[0015] Furthermore, the model program composed of multiple differential equations in S4 of the method for constructing a PBTK-TD model of the present invention is as follows:

[0016] (Ⅰ) Absorption equation:

[0017] (Ⅰ-1)

[0018] Represents the rate of change of the concentration of bongkrekic acid toxin in the intestine over time, with the unit of mg / h;

[0019] (Ⅰ-2)

[0020] Represents the rate of change of the concentration of bongkrekic acid toxin in the blood over time, with the unit of mg / h, reflecting the rate at which bongkrekic acid toxin is absorbed from the intestine into the blood;

[0021] C gut is the concentration of bongkrekic acid toxin in the intestine, C blood is the concentration of bongkrekic acid toxin in the blood, k a is the absorption rate constant, F is the bioavailability, V blood is the volume of the blood;

[0022] (Ⅱ) Distribution equation:

[0023]

[0024] Q organ refers to the blood flow rate to the tissue, C organ is the concentration of bongkrekic acid toxin in the tissue, P organ is the tissue-blood partition coefficient, V organ is the volume of the tissue;

[0025] (Ⅲ) Metabolism equation:

[0026]

[0027] C liver is the concentration of bongkrekic acid toxin in the liver, Q liver is the amount of blood flowing to the liver per unit time, V liver is the physiological volume of the liver, P liver is the equilibrium concentration ratio of bongkrekic acid toxin in the liver and blood, V max is the maximum metabolic capacity of the liver metabolic enzyme;

[0028] (Ⅳ) Excretion equation:

[0029]

[0030] C kidney is the concentration of bongkrekic acid toxin in the kidney, Q kidney is the amount of blood flowing to the kidney per unit time,

[0031] V kidneyis the physiological volume of the kidney, P kidney is the equilibrium concentration ratio of bongkrekic acid toxin in the kidney and blood, CL renal is the rate at which the kidney clears bongkrekic acid toxin by filtration or active secretion;

[0032] (V) Binding and dissociation equation of bongkrekic acid toxin and adenine nucleotide translocase ANT on the mitochondrial membrane:

[0033]

[0034] C mito is the concentration of bongkrekic acid toxin bound to ANT in the mitochondrion, C organ is the concentration of free bongkrekic acid toxin in the target organ, ANT total is the total concentration of ANT, k on is the binding rate constant, the binding rate constant of bongkrekic acid toxin and ANT, reflecting the binding efficiency; k off is the dissociation rate constant, the dissociation rate constant of the bongkrekic acid toxin-ANT complex, reflecting the binding stability;

[0035] (VI) Toxicity effect equation:

[0036] Furthermore, in S5 of the PBTK-TD model construction method of the present invention, the optimization method for constructing the bongkrekic acid PBTK-TD model is: by using the said multivariate differential equation, plotting the metabolic kinetics curve of bongkrekic acid toxin, and adjusting the bongkrekic acid toxicokinetics parameters, when the simulated curve tends to be consistent with the experimental result curve, the set of multivariate differential equations corresponding to the simulated curve is the metabolic kinetics model established after optimization training.

[0037] Furthermore, S5 of the PBTK-TD model construction method of the present invention also includes the verification of the constructed bongkrekic acid PBTK-TD model. The specific verification method is: dividing the data set into a training set and a verification set, checking whether the model fitting residuals are randomly distributed, and evaluating the prediction ability of the bongkrekic acid PBTK-TD model.

[0038] Optimization scheme, after the bongkrekic acid PBTK-TD model constructed by the PBTK-TD model construction method of the present invention passes the verification, the actual exposure value of the internal organs in the body can be deduced inward, and the actual exposure value outside the body can be deduced outward through the associated model.

[0039] A PBTK-TD model for acute oral exposure of rats to bongkrekic acid, which obtains the bongkrekic acid PBTK-TD model by using the above PBTK-TD model construction method.

[0040] The present invention has the following outstanding substantial features and remarkable progress:

[0041] The construction method of the PBTK-TD model for acute oral exposure of rats to bongkrekic acid proposed by the present invention established the PBTK-TD model of bongkrekic acid toxin by studying the absorption, distribution, metabolism and excretion characteristics of bongkrekic acid toxin in rats; and proposed the PBTK-TD model. Through this model, the toxicokinetics and toxicodynamics of bongkrekic acid toxin can be accurately predicted and described. The established PBTK-TD model of bongkrekic acid can deduce the actual exposure values of internal organs in the body inward and the actual exposure values in vitro through the associated model outward, providing a scientific basis for further toxicological research and risk assessment of this toxin. Brief Description of the Drawings

[0042] Figure 1 It is the concentration-time relationship diagram of bongkrekic acid in the plasma of rats of the present invention.

[0043] Figure 2 It is the test result with bongkrekic acid 10 mg / kg single gavage of rats as the test sample and the metabolic kinetic curve of the bongkrekic acid toxin of the present invention.

[0044] Figure 3 It is the EC 50 and PD effect values and simulation curves of bongkrekic acid toxin in the present invention.

[0045] Figure 4 It is the comparison between the predicted value and the observed value of the bongkrekic acid metabolic kinetic model after optimization and training in the present invention.

[0046] Figure 5 It is the time change trend of the content levels of bongkrekic acid in liver, heart and kidney tissues predicted by the PBTK-TD model in the present invention.

[0047] Figure 6 It is the comparison between the predicted population exposure risk and the measured value of the kinetic parameters of bongkrekic acid based on the model program established by MAS software and the simulation in the present invention.

[0048] Figure 7 It is the analysis diagram of the simulation and prediction fitting degree of the evaluation model in the present invention. Detailed Embodiments

[0049] The present invention will be further described below in conjunction with specific embodiments, but the protection scope of the present invention is not limited thereto:

[0050] The experimental methods without specific conditions in the following embodiments are carried out according to conventional methods and conditions.

[0051] Embodiment

[0052] A method for constructing a PBTK-TD model for acute oral exposure of rats to bongkrekic acid, comprising the following steps:

[0053] S1: Using rats orally administered bongkrekic acid as test samples, specifically, rats were intragastrically administered bongkrekic acid at different doses once, and plasma and tissue samples of the test samples were collected for detection from 0 to 24 hours. The concentration of bongkrekic acid toxin in the samples was measured, and the kinetic processes of absorption, distribution, metabolism, and excretion of orally exposed bongkrekic acid in rats were analyzed and determined.

[0054] As Figure 1 shown, the detection results of the concentration of bongkrekic acid toxin in the plasma samples of the test samples from 0 to 24 hours were obtained using rats intragastrically administered 10 mg / kg of bongkrekic acid as test samples.

[0055] S2: Integrate the processes of absorption, distribution, metabolism, excretion of bongkrekic acid toxin in the body and the toxic effect mechanism, determine the PBTK-TD model for rats orally exposed to bongkrekic acid, conduct model framework design, and establish the overall structure of the model, including the composition structure of the core physiological compartments and the targets of the toxicokinetics module.

[0056] In S2, according to the physiological structure of rats and the toxicokinetic process of bongkrekic acid, the model framework design was carried out, specifically:

[0057] Determine the core physiological compartments: The model includes multiple physiological compartments, mainly the heart, liver, kidney, and intestine. These compartments are interconnected by blood flow to simulate the processes of absorption, distribution, metabolism, and excretion of bongkrekic acid toxin in the test samples;

[0058] Determine the targets of the toxicokinetics module: For bongkrekic acid, its main toxicity target is mitochondrial adenine nucleotide translocase (ANT), which inhibits ATP generation by binding to ANT.

[0059] S3: Collect effective parameters: A. Physiological parameters of rats; B. Toxicokinetic parameters of bongkrekic acid, including absorption rate constant k a 、metabolic rate constant K m 、tissue-blood partition coefficient; C. Toxicokinetic parameters of bongkrekic acid, including binding rate constant k on and dissociation rate constant k off 、half maximal effective concentration EC 50 ; The specific values of the relevant parameters are shown in Table 1 and Table 2.

[0060]

[0061] S4: Establish a model program. According to the law of conservation of mass, differential equations were written for each physiological compartment to describe the change in the concentration of bongkrekic acid in each compartment, and a model program consisting of multiple differential equations was obtained.

[0062] The model program composed of the partial differential equations is specifically as follows:

[0063] (Ⅰ) Absorption equation:

[0064] (Ⅰ-1)

[0065] represents the rate of change of the concentration of bongkrekic acid toxin in the intestine with time, with the unit of mg / h;

[0066] (Ⅰ-2)

[0067] represents the rate of change of the concentration of bongkrekic acid toxin in the blood with time, with the unit of mg / h, reflecting the rate at which bongkrekic acid toxin is absorbed from the intestine into the blood;

[0068] C gut is the concentration of bongkrekic acid toxin in the intestine, C blood is the concentration of bongkrekic acid toxin in the blood, k a is the absorption rate constant, F is the bioavailability, V blood is the volume of the blood;

[0069] (Ⅱ) Distribution equation:

[0070]

[0071] Q organ refers to the blood flow rate to the tissue, C organ is the concentration of bongkrekic acid toxin in the tissue, P organ is the tissue-blood partition coefficient, V organ is the volume of the tissue; the tissues include the heart, liver, kidney, intestine, etc.

[0072] (Ⅲ) Metabolism equation:

[0073]

[0074] C liver is the concentration of bongkrekic acid toxin in the liver, Q liver is the amount of blood flowing to the liver per unit time, V liver is the physiological volume of the liver, P liver is the equilibrium concentration ratio of bongkrekic acid toxin in the liver and blood, V max is the maximum metabolic capacity of the liver metabolic enzyme;

[0075] (Ⅳ) Excretion equation:

[0076]

[0077] C kidney is the concentration of bongkrekic acid toxin in the kidney, Q kidney is the amount of blood flowing to the kidney per unit time,

[0078] V kidney is the physiological volume of the kidney, P kidney is the equilibrium concentration ratio of bongkrekic acid toxin between the kidney and blood, CL renal is the rate at which the kidney clears bongkrekic acid toxin by filtration or active secretion;

[0079] (Ⅴ) Equation for the binding and dissociation of bongkrekic acid toxin with the adenine nucleotide translocase ANT on the mitochondrial membrane:

[0080]

[0081] C mito is the concentration of bongkrekic acid toxin bound to ANT in the mitochondria, C organ is the concentration of free bongkrekic acid toxin in the target organ, ANT total is the total concentration of ANT, k on is the binding rate constant, the binding rate constant of bongkrekic acid toxin with ANT, reflecting the binding efficiency; k off is the dissociation rate constant, the dissociation rate constant of the bongkrekic acid toxin-ANT complex, reflecting the binding stability;

[0082] (Ⅵ) Toxicity effect equation:

[0083] S5: Sensitivity analysis and optimization of the model program parameters to construct the bongkrekic acid PBTK-TD model. The specific method is as follows:

[0084] (S5-1) Software tools and numerical solution;

[0085] Tool selection: Maspectra, using ode45 to solve the differential equation system.

[0086] Parameter fitting: Nonlinear least squares method, fitting parameters according to experimental animal data (plasma concentration, tissue concentration, toxicity effect).

[0087] Bayesian method: Combining prior distribution to optimize parameter uncertainty.

[0088] (S5-2) Experimental data requirements;

[0089] Toxicokinetics data: Specifically, the plasma concentration-time curve. After rats are orally administered bongkrekic acid, plasma samples are collected at different time points.

[0090] Tissue distribution data: Specifically, after sacrificing the test article, the concentrations of bongkrekic acid in tissues such as blood, heart, liver, and kidney were measured.

[0091] Toxicokinetics data: Specifically, the mitochondrial ATP production rate, which was determined by an in vitro mitochondrial exposure experiment.

[0092] (S5-3) Experimental results;

[0093] First, the measurement results of the test article with bongkrekic acid at a single dose of 10 mg / kg by gavage in rats in this example. The bongkrekic acid toxin metabolic kinetics curve includes:

[0094] (1) Figure 2 The measured values and simulated curves of the bongkrekic acid toxin concentration-time in the rat liver shown in Liver, and the simulated curve corresponds to the above (II, III) equations;

[0095] (2) Figure 2 The measured values and simulated curves of the bongkrekic acid toxin concentration-time in the rat heart shown in Heart, and the simulated curve corresponds to the above (II) equation;

[0096] (3) Figure 2 The measured values and simulated curves of the bongkrekic acid toxin concentration-time in the rat intestine shown in Intestinal tissue, and the simulated curve corresponds to the above (I-1) equation;

[0097] (4) Figure 2 The measured values and simulated curves of the bongkrekic acid toxin concentration-time in the rat adipose tissue shown in Adipose tissue, and the simulated curve corresponds to the above (II) equation;

[0098] (5) Figure 2 The measured values and simulated curves of the bongkrekic acid toxin concentration-time in the rat kidney shown in Kidney, and the simulated curve corresponds to the above (II, IV) equations;

[0099] (6) Figure 2 The measured values and simulated curves of the bongkrekic acid toxin concentration-time in the rat plasma shown in Plasma, and the simulated curve corresponds to the above (I-2) equation;

[0100] (7) Figure 3 The bongkrekic acid toxin EC 50 and PD effect values and simulated curves, and the simulated curve corresponds to the above (V, VI) equations.

[0101] The method for optimizing the sensitivity of the model program parameters is as follows: by means of the multivariate differential equation, draw the metabolic kinetic curve of bongkrekic acid toxin, and adjust the toxicokinetic parameters of bongkrekic acid to make the simulated curve tend to be consistent with the experimental result curve. When Figure 4 as shown, the set of multivariate differential equations corresponding to the simulated curve is the metabolic kinetic model established after optimized training.

[0102] Figure 4 Analysis conclusion: In this study, we comprehensively evaluated the 10 mg dose bongkrekic acid model to ensure its accuracy and reliability in predicting the kinetic behavior of the toxin. We used the nonlinear mixed effects model (NLME) and verified the model through a series of diagnostic plots, including the comparison of individual predictions with observed values, conditional weighted residual (CWRES) analysis, and the examination of the distribution of residuals. The circles represent the actual observed values (Observations), the solid lines represent the individual predicted values (IPRED), and the dashed lines represent the population predicted values (PRED). In most subplots, there is a high degree of consistency between the observed values and the predicted values, indicating that the model can better capture the toxin kinetic characteristics of individuals. The individual predicted values (solid lines) are all distributed near the observed values (circles), while the population predicted values (dashed lines) provide an average prediction trend. By observing the CWRES plot, we found that the residuals are roughly randomly distributed near the zero line, indicating that the model has no obvious systematic bias. The histogram of the residuals shows that the residuals are roughly normally distributed with a mean close to zero, further verifying the prediction accuracy of the model. Therefore, the quantitative toxicology model constructed in this study shows good performance in predicting the kinetic behavior of 10 mg dose bongkrekic acid. The model evaluation results show that the model can effectively capture the trend of the toxin concentration changing over time and has no obvious systematic bias.

[0103] (S5-4) Model validation;

[0104] Cross-validation: Divide the data set into a training set and a validation set to evaluate the model's prediction ability.

[0105] Residual analysis: Check whether the model fitting residuals are randomly distributed.

[0106] Validation experiment: Use the experimental data of intragastric administration of 10 mg / kg bongkrekic acid to rats to verify the model extrapolation ability, see Figure 6 .

[0107] After the metabolic kinetic model is verified, it shows that the constructed bongkrekic acid PBTK-TD model can deduce the actual exposure values of internal organs in the body inward and can deduce the actual exposure values in vitro through the correlation model outward.

[0108] Figure 6 Analysis conclusion: FromFigure 6 As can be seen, the solid line (—) represents IPRED (Initial Prediction), that is, the initial predicted value. The circle (o) represents the observations, that is, the data obtained from in vivo experiments on rats. The initial predicted value of the model is consistent with the actual observations in most cases. This indicates that the constructed PBTK-TD model for bongkrekic acid has good extrapolation ability.

[0109] 1. Model prediction accuracy: Figure 6 shows the agreement between the initial predicted value of the model and the actual observations at multiple time points. In most cases, the predicted value and the observed value have the same trend, indicating that the model can well describe the metabolic process of bongkrekic acid in rats. It verifies the prediction of the concentration-time change of bongkrekic acid in vivo by the model.

[0110] 2. Model extrapolation ability: The dashed line (---) represents PRED (Population Prediction), that is, the population predicted value. The model is not only applicable to the current experimental conditions (10 mg / kg gavage in rats), but also can be applicable to other doses and exposure conditions in the population by adjusting parameters. This enhances the practicality and flexibility of the model, indicating that the model has good extrapolation ability.

[0111] 3. Derivation of in vivo organ exposure values: The actual exposure values of each organ in vivo can be derived through the model. This is of great significance for the distribution and metabolism of bongkrekic acid in different organs.

[0112] 4. Derivation of in vitro exposure values: Through the correlation model, the actual in vitro exposure values can be derived. This has reference value for evaluating the behavior and potential risks of bongkrekic acid under different environmental conditions.

[0113] In summary, the constructed PBTK-TD model for bongkrekic acid performs excellently in terms of prediction and extrapolation ability, and can provide a scientific basis for the toxicity assessment and risk management of bongkrekic acid.

[0114] The following is a specific description of the application of the PBTK-TD model for bongkrekic acid constructed in the present invention:

[0115] 1. Toxicity risk assessment;

[0116] Prediction of target organ exposure: Simulate the concentration of bongkrekic acid in liver and heart tissues and evaluate the dose-response relationship with the ATP inhibition rate.

[0117] 2. Calculation of safety thresholds: Determine the no-observed-adverse-effect level (NOAEL) and the lowest-observed-adverse-effect level (LOAEL).

[0118] 3. Human prediction: By adjusting physiological parameters (such as blood flow, organ volume), the PBTK-TD model of bongkrekic acid was extended to human toxicity assessment.

[0119] Specifically as follows:

[0120] Bongkrekic acid is mainly absorbed through the digestive tract mucosa and rapidly distributes to all tissues of the body after entering the blood circulation. Research shows that bongkrekic acid is rapidly absorbed in rats. 15 minutes after gavage, the radioactivity specific activity in plasma and major tissues reaches the peak, especially with the highest concentration in the liver. Therefore, it is rapidly absorbed through the intestine after oral administration (ka≈0.3h - 1), and the bioavailability is about 20% (significant first-pass effect). Bongkrekic acid has high lipid solubility (LogP≈5.0) and is widely distributed in fat (Pfat≈3.0), liver (Pliver≈1.8), and heart (Pheart≈1.5), with a volume of distribution Vd≈2.0L / kg. Then, it is mainly oxidized by CYP3A4 / CYP2C9 in the liver to form hydroxylated products (with reduced toxicity), and some form epoxides (potential toxicity). Through the conjugation metabolism of glutathione (GSH), the clearance is accelerated. Finally, it is excreted through the kidneys, with a low renal clearance rate (CLrenal≈0.15mL / min / kg), a half-life of about 12 hours, and partial excretion through bile (enterohepatic circulation). After simulating a single oral dose of 10mg in humans using the bongkrekic acid PBTK-TD model, the plasma peak concentration Cmax≈1.8μM (reaching the peak at 2 - 4 hours). There is significant accumulation in the liver and heart (Cliver≈2.7μM, Cheart≈2.0μM), the caspase-3 activity increases, and the peak inhibition rate of ATP production reaches 50%. Continuous exposure for 24 hours induces apoptosis. Based on this data, the model parameters were optimized and verified, enabling the bongkrekic acid PBTK-TD model to fully simulate the toxicokinetics process after human ingestion of bongkrekic acid. The predicted content-time change curves of bongkrekic acid in liver, heart, and kidney tissues are basically consistent with the measured values, and the changing trend of the content level over time coincides with the measured values. As Figure 5 shown, the predicted peak concentrations of bongkrekic acid in liver, kidney, and heart tissues obtained by the bongkrekic acid PBTK-TD model are 14.47mg / L, 11.27mg / L, and 10.91mg / L respectively, and the measured peak concentrations are 13.50mg / L, 11.62mg / L, and 9.23mg / L respectively, that is, the predicted peak concentrations are 0.97 - 1.18 times the measured levels, and the model prediction is accurate.

[0121] In addition, in this example, based on the MAS software, the model program and the kinetic parameter simulation of bongkrekic acid were used to predict the population exposure risk and compare it with the measured values, as shown in Figure 6 .

[0122] In this study, we constructed and evaluated a quantitative toxicology model for bongkrekic acid, aiming to predict the kinetic behavior of different doses of bongkrekic acid. The model was fitted by the nonlinear mixed effects method, with time (TIME) as the independent variable and the observed value (DV) as the dependent variable. We used visual predictive check (VPC) as the main tool for model evaluation, and evaluated the predictive performance of the model by comparing the concentration-time curves predicted by the model with the actual observed data. Figure 7 The concentration-time curves of different doses of the toxin are shown. Figure 7 In, the blue dots represent the actual observed values, the red dots represent the individual predicted values, the blue and red solid lines represent the overall predicted values, and the blue and red shaded areas represent the 95% confidence intervals of the overall and individual predictions, respectively. The concentrations predicted by the model are basically consistent with the actual observed values, and most of the actual observed values fall within the predicted 95% confidence interval, indicating that the model has good predictive accuracy and reliability.

Claims

1. Method for constructing PBTK-TD model for acute oral exposure of rats to bongkrekic acid, characterized in that It includes the following steps: S1: Using rats orally administered with bongkrekic acid as test samples, collecting the plasma and tissue samples of the test samples, and measuring the concentration of bongkrekic acid in the samples for analyzing the kinetic processes of the absorption, distribution, metabolism and excretion of bongkrekic acid in rats. S2: Establishing the overall structure of the model, including the compositional structure of the core physiological compartments and the targets of the toxicokinetics module. S3: Collect valid parameters: A. Rat physiological parameters; B. Toxicokinetics parameters of bongkrekic acid, including absorption rate constant k a , metabolic rate constant K m , tissue-blood partition coefficient; C. Toxicodynamics parameters of bongkrekic acid, including binding rate constant k of ANT on and dissociation rate constant k off , median effective concentration EC 50 ; S4: Establishing the model program, and according to the law of conservation of mass, writing differential equations for each physiological compartment to describe the concentration change of bongkrekic acid in each compartment, so as to obtain the model program composed of multivariate differential equations. S5: Conducting sensitivity analysis and optimization of the parameters of the model program to construct the bongkrekic acid PBTK-TD model.

2. The method for constructing a PBTK-TD model of acute oral exposure to bongkrekic acid in rats according to claim 1, characterized in that: Using the bongkrekic acid PBTK-TD model, calculating the dose-response relationship through the model to evaluate the quantitative relationship between the exposure dose and the toxic effect, and predicting the distribution and toxic effect of bongkrekic acid in vivo at different doses by using the model.

3. The method for constructing a PBTK-TD model for acute oral exposure of rats to bongkrekic acid according to claim 1, wherein: In S2, determining the core physiological compartments: The physiological compartments included in the model mainly include the heart, liver, kidney and intestine, and the compartments are interconnected by blood flow to simulate the absorption, distribution, metabolism and excretion processes of bongkrekic acid toxin in the test samples.

4. The method for constructing a PBTK-TD model for acute oral exposure of rats to bongkrekic acid according to claim 3, characterized in that: In S2, determining the targets of the toxicokinetics module: For bongkrekic acid, its main toxic target is ANT, and it inhibits ATP generation by binding to ANT.

5. The method for constructing a PBTK-TD model for acute oral exposure of rats to bongkrekic acid according to claim 4, characterized in that: The model program composed of multivariate differential equations in S4 is as follows: (Ⅰ) Absorption equation: Indicates the rate of change of the concentration of bongkrekic acid toxin in the intestine over time, with the unit of mg / h; Indicates the rate of change of the concentration of bongkrekic acid toxin in the blood over time, with the unit of mg / h, reflecting the rate at which bongkrekic acid toxin is absorbed from the intestine into the blood; C gut is the concentration of bongkrekic acid toxin in the intestine, C blood is the concentration of bongkrekic acid toxin in the blood, k a is the absorption rate constant, F is the bioavailability, V blood is the volume of blood; (Ⅱ) Distribution equation: Q organ refers to the blood flow rate flowing into the tissue, C organ is the concentration of bongkrekic acid toxin in the tissue, P organ is the tissue-blood partition coefficient, V organ is the volume of the tissue; (Ⅲ) Metabolism equation: C liver is the concentration of bongkrekic acid toxin in the liver, Q liver is the blood flow to the liver per unit time, V liver is the physiological volume of the liver, P liver is the equilibrium concentration ratio of bongkrekic acid toxin in the liver and blood, V max is the maximum metabolic capacity of liver metabolic enzymes; (Ⅳ) Excretion equation: C kidney is the concentration of bongkrekic acid toxin in the kidney, Q kidney is the amount of blood flowing to the kidney per unit time, V kidney is the physiological volume of the kidney, P kidney is the equilibrium concentration ratio of bongkrekic acid toxin between the kidney and the blood, CL renal is the rate at which the kidney clears bongkrekic acid toxin by filtration or active secretion; (Ⅴ) Binding and dissociation equations of bongkrekic acid toxin and adenine nucleotide translocase ANT on the mitochondrial membrane: C mito is the concentration of bongkrekic acid toxin bound to ANT in mitochondria, C organ is the concentration of free bongkrekic acid toxin in the target organ, ANT total is the total concentration of ANT, k on is the binding rate constant, the binding rate constant of bongkrekic acid toxin to ANT, reflecting the binding efficiency; k off is the dissociation rate constant, the dissociation rate constant of the bongkrekic acid toxin-ANT complex, reflecting the binding stability; (Ⅵ) Toxicity effect equation:

6. The method for constructing a PBTK-TD model for acute oral exposure of rats to bongkrekic acid according to claim 5, wherein: The optimization method for constructing the bongkrekic acid PBTK-TD model in S5 is: Through the multivariate differential equations, plotting the metabolic kinetic curve of bongkrekic acid toxin, and adjusting the toxicokinetic parameters of bongkrekic acid, when the simulation curve tends to be consistent with the experimental result curve, the set of multivariate differential equations corresponding to the simulation curve is the metabolic kinetic model established after optimization training.

7. The method for constructing a PBTK-TD model for acute oral exposure of rats to bongkrekic acid according to claim 6, wherein: S5 also includes the verification of the constructed bongkrekic acid PBTK-TD model. The specific verification method is: Dividing the data set into a training set and a verification set, checking whether the model fitting residuals are randomly distributed, and evaluating the prediction ability of the bongkrekic acid PBTK-TD model.

8. The method for constructing a PBTK-TD model for acute oral exposure of rats to bongkrekic acid according to claim 7, characterized in that: After the constructed bongkrekic acid PBTK-TD model passes the verification, the actual exposure values of internal organs in vivo can be deduced inward, and the actual exposure values in vitro can be deduced outward through the associated model.

9. PBTK-TD model for acute oral exposure of rats to bongkrekic acid, characterized in that: It is the bongkrekic acid PBTK-TD model obtained by using the construction method described in any one of claims 1 to 8.

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