Urinary tract infection model for simulating antibacterial drug in-vitro pharmacokinetics / pharmacodynamics

Abstract

The invention relates to a urinary tract infection model for simulating in-vitro pharmacokinetics / pharmacodynamics of antibacterial drugs. The urinary tract infection model comprises a kidney chamber, a bladder chamber, a drug feeding tank and a drug discharging tank, wherein a bacterial culture medium is arranged in the kidney chamber. The use method of the model comprises the steps of culture medium preparation, drug preparation, strain preparation, flow velocity setting, drug administration simulation and urination simulation. And continuously carrying out continuous medication simulation for 1-7 days, counting the drug concentration and bacteria, and evaluating the action effects of the drug under different doses according to the change of the colony count and the initial bacteria quantity at the observation end point in 24 hours or in 24 hours. The method can be used as a urinary tract infection model for simulating antibacterial drug in-vitro pharmacokinetics / pharmacodynamics research, and PK / PD index and target value research of antibacterial drug urinary tract infection is carried out; evaluating the effectiveness of the antibacterial agent in treating urinary tract infection under the clinical dosage; the clinical treatment effect is improved according to the clinical treatment scheme of the optimized antibacterial medicine; and evaluating the sterilization condition of the antibacterial agent on the bacteria forming the biological membrane in the catheter.

Description

Technical Field

[0001] The present invention belongs to the technical field of in vitro pharmacokinetics and pharmacodynamics of antibacterial drugs, and particularly relates to a urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs. Background Art

[0002] With the continuous advancement of antimicrobial pharmacokinetics (PK) and pharmacodynamics (PD), PK / PD studies are becoming increasingly important in evaluating the pharmacodynamics of antimicrobial drugs, developing and optimizing dosing regimens, and mitigating bacterial resistance. Currently, numerous in vitro PK / PD study methods exist. In addition to traditional in vitro pharmacodynamic methods (such as minimum inhibitory concentration determination and static kill curves), in vitro PK / PD models are also evolving. These in vitro PK / PD models utilize in vitro devices to simulate the pharmacokinetic processes of antimicrobial drugs in the human body and investigate changes in their efficacy during this process. The key difference between these in vitro PK / PD models and traditional in vitro pharmacodynamic studies is that drug concentrations vary according to human pharmacokinetic laws, rather than being constant, which is more consistent with drug behavior in the human body and can be maintained for 72 hours. Furthermore, single or multiple flow rates can be set, providing high controllability and fully simulating single- or multi-compartment pharmacokinetic behavior in the human body.

[0003] Patent CN106281998B, a state-of-the-art patent, discloses an in vitro dynamic pharmacokinetic / pharmacodynamic model system and construction method for anaerobic bacteria. This system primarily simulates the dynamic pharmacokinetic / pharmacodynamics of anaerobic bacteria using an anaerobic chamber, anaerobic bags, and a closed gas circulation system. The existing technology combines the anaerobic chamber and anaerobic bag gas oxygen consumption methods to ensure that both the external and internal environments of the entire dynamic model achieve good anaerobic conditions. Furthermore, the system can be combined with a computer to control the flow rate of the model in real time to simulate various pharmacokinetic processes. However, this system is not suitable for simulating urinary tract infections, and there are no existing indicators for evaluating the effectiveness of antimicrobial drugs in treating urinary tract infections at clinical doses. Summary of the Invention

[0004] The technical problem to be solved by the present invention is to provide a urinary tract infection model that simulates the in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs, provide experimental support for the simulation of in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs, and can be used to evaluate the effectiveness of antibacterial drugs in treating urinary tract infections at clinical doses.

[0005] The technical solution adopted by the present invention to solve its technical problems is: to provide a urinary tract infection model that simulates the in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs, including a kidney chamber with a bacterial culture medium inside, a bladder chamber, a drug administration tank and a drug discharge tank, the drug administration tank and the drug discharge tank are respectively connected to the kidney chamber through liquid paths provided with infusion pump 1 and infusion pump 3, the kidney chamber is respectively connected to the bladder chamber and the waste liquid tank through liquid paths provided with infusion pump 4 and infusion pump 2, and the bladder chamber is connected to the waste liquid tank through a urinary catheter with a valve.

[0006] The method for using the urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs comprises the following steps:

[0007] S1. Culture medium preparation: adding bacterial culture medium into the kidney chamber, bladder chamber, medication tank, and drug discharge tank, wherein the volume of bacterial culture medium in the kidney chamber is a fixed value V1, and the volume of bacterial culture medium in the bladder chamber is an initial value V2;

[0008] S2. Drug preparation: Add the antimicrobial drug to be verified into the dosing tank according to the experimental settings;

[0009] S3. Strain preparation: Add the test strain into the bladder chamber according to the experimental settings;

[0010] S4, flow rate setting: set the flow rate F1 of infusion pump 1 from the drug administration tank to the kidney chamber according to the simulated antimicrobial drug concentration rising rate; set the flow rate F3 of infusion pump 3 from the drug discharge tank to the kidney chamber according to the simulated drug concentration falling rate; set the flow rate F4 of infusion pump 4 from the kidney chamber to the bladder chamber according to the simulated urine production rate; set the flow rate F2 of infusion pump 2 from the kidney chamber to the waste liquid tank to maintain the volume V1 of the bacterial culture medium in the kidney chamber fixed;

[0011] S5. Simulated drug administration: Based on clinical medication, start infusion pump 1 to simulate drug administration and gradually increase the drug concentration in the kidney, while keeping infusion pump 3 closed. After reaching the peak drug concentration in the kidney, turn off infusion pump 1 and start infusion pump 3 to simulate the gradual decrease of drug concentration in the kidney after drug withdrawal.

[0012] S6. Simulated urination: Keep infusion pump 4 turned on throughout the experiment to simulate the kidneys constantly producing urine into the bladder; divide 24 hours into daytime and nighttime periods, and perform urination simulation at least once every 4 hours and no more than once every hour during the daytime period. The urination simulation refers to starting infusion pump 4 to discharge the bacterial culture medium in the bladder chamber until the remaining volume returns to the initial value V2.

[0013] The evaluation method of the urinary tract infection model for simulating the in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs is as follows: after continuous drug use simulation for 1 to 7 days according to the usage method, drug concentration and bacterial count are performed, and the effect of the drug at different doses is evaluated based on the change in the colony count and the initial bacterial count at 24 hours or the observation endpoint.

[0014] Preferably, the kidney chamber and the bladder chamber are arranged on a magnetic stirrer, and a stirrer matching the magnetic stirrer is arranged inside the kidney chamber and the bladder chamber.

[0015] Preferably, sampling needles are provided on the kidney chamber and the bladder chamber.

[0016] Preferably, the infusion pump 1, the infusion pump 2, the infusion pump 3 and the infusion pump 4 are all peristaltic pumps.

[0017] Preferably, the volume of the bacterial culture medium provided in the kidney chamber is 100 mL to 500 mL; the initial volume of the culture medium provided in the bladder chamber is 1 to 10 mL.

[0018] Preferably, in S3, the amount of the test strain added is 10 5 ~10 9 CFU / mL.

[0019] Preferably, in said S4, the infusion pump flow rate F4 set according to the urine production rate is 0.1-0.5 mL / min.

[0020] In S5, the daytime period is 14 hours long, the nighttime period is 10 hours long, and the urination frequency is once every 2 hours, a total of 7 times.

[0021] Preferably, the temperature in the bladder chamber and the kidney chamber is maintained at 35±2°C.

[0022] Preferably, the bladder chamber is connected to the catheter through a valve, and the other end of the catheter is connected to the waste liquid tank. In this case, the evaluation method also includes observing the growth of biofilm in the catheter and counting bacteria to evaluate the effect of antibacterial drugs on catheter-related urinary tract infections.

[0023] The beneficial effects are as follows: the present invention can be used as a urinary tract infection model for in vitro pharmacokinetic / pharmacodynamic studies of antimicrobial drugs, (1) to conduct PK / PD index and target value studies of urinary tract infections with antimicrobial drugs; (2) to evaluate the effectiveness of antimicrobial drugs in treating urinary tract infections at clinical doses; (3) to optimize clinical treatment plans for antimicrobial drugs based on the studies and improve clinical efficacy; and (4) to evaluate the bactericidal effect of antimicrobial drugs on bacteria that form biofilms in urinary catheters. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1Flowchart of a urinary tract infection model that simulates the in vitro pharmacokinetics / pharmacodynamics of antimicrobial drugs.

[0025] Among them, 1-drug administration tank; 101-infusion pump one; 2-drug discharge tank; 201-infusion pump three; 3-kidney chamber; 301-infusion pump two; 302-infusion pump four; 4-bladder chamber; 401-valve; 402-catheter; 5-waste liquid tank; 6-sampling needle; 7-magnetic stirrer. DETAILED DESCRIPTION

[0026] Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiments are only used to illustrate the present invention and are not used in limiting the scope of the present invention.In addition, should be understood that after reading the content taught by the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms fall equally within the scope limited by the appended claims of the application.

[0027] like Figure 1 As shown, the present invention provides a urinary tract infection model for simulating the in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs, comprising a kidney chamber 3 with a bacterial culture medium inside, a bladder chamber 4, a drug administration tank 1 and a drug discharge tank 2, wherein the drug administration tank 1 and the drug discharge tank 2 are respectively communicated with the kidney chamber 3 through a fluid path provided with an infusion pump 1 101 and an infusion pump 3 201, the kidney chamber 3 is respectively communicated with the bladder chamber 4 and the waste liquid tank 5 through a fluid path provided with an infusion pump 4 302 and an infusion pump 2 301, and the bladder chamber 4 is communicated with the waste liquid tank 5 through a urinary catheter with a valve 401.

[0028] The method for using the urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs comprises the following steps:

[0029] S1. Culture medium preparation: Add bacterial culture medium into the kidney chamber 3, bladder chamber 4, medication tank 1 and drug discharge tank 2, wherein the volume of bacterial culture medium in the kidney chamber 3 is a fixed value V1, and the volume of bacterial culture medium in the bladder chamber 4 is an initial value V2;

[0030] S2. Drug preparation: Add the antimicrobial drug to be verified into the dosing tank 1 according to the experimental settings;

[0031] S3. Strain preparation: Add the test strain into bladder chamber 4 according to the experimental settings;

[0032] S4. Flow rate setting: Set the flow rate F1 of infusion pump 101 from drug administration tank 1 to kidney chamber 3 based on the simulated antimicrobial drug concentration increase rate; set the flow rate F3 of infusion pump 3 201 from drug discharge tank 2 to kidney chamber 3 based on the simulated drug concentration decrease rate; set the flow rate F4 of infusion pump 4 302 from kidney chamber 3 to bladder chamber 4 based on the simulated urine production rate; and set the flow rate F2 of infusion pump 2 301 from kidney chamber 3 to waste liquid tank 5 to maintain a constant volume V1 of the bacterial culture medium in kidney chamber 3.

[0033] S5. Simulated drug administration: Based on clinical medication, start infusion pump 101 to simulate drug administration and gradually increase the drug concentration in the kidney, while keeping infusion pump 201 off. After reaching the peak drug concentration in the kidney, turn off infusion pump 101 and simultaneously start infusion pump 201 to simulate the gradual decrease of drug concentration in the kidney after drug withdrawal.

[0034] S6. Simulated urination: The infusion pump 4 302 is kept turned on throughout the experiment to simulate the kidneys constantly producing urine into the bladder; 24 hours are divided into daytime and nighttime periods, and urination simulation is performed at least once every 4 hours and at most once every 1 hour during the daytime period. The urination simulation refers to starting the infusion pump 4 302 to discharge the bacterial culture medium in the bladder chamber 4 until the remaining volume returns to the initial value V2.

[0035] The volume of the bacterial culture medium set in the kidney chamber 3 is 100-500 mL; the initial volume of the culture medium set in the bladder chamber 4 is 1-10 mL. The flow rates F1 and F3 set for infusion pump 1 101 and infusion pump 3 201 should be greater than / equal to the target urine production rate F4. Therefore, in S6, to ensure that the volume in the kidney chamber 3 remains unchanged, when the total bacterial culture medium injected into the kidney chamber 3 by infusion pump 1 and infusion pump 3 exceeds the bacterial culture medium output from the kidney chamber 3 to the bladder chamber, infusion pump 2 is activated to transfer the excess bacterial culture medium to the waste liquid tank.

[0036] The evaluation method of the urinary tract infection model for simulating the in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs is as follows: after continuous drug administration simulation for 1 to 7 days according to the usage method, drug concentration and bacterial count are performed, and the effect of the drug at different doses is evaluated based on the changes in the colony count and the initial bacterial count at the 24-hour or observation end point of the urinary catheter.

[0037] The kidney chamber 3 and the bladder chamber 4 are mounted on a magnetic stirrer 7, and a stirrer matching the magnetic stirrer 7 is mounted inside the kidney chamber 3 and the bladder chamber 4. During the experiment, the magnetic stirrer 7 was turned on, and the stirrer was located below the liquid surface of the bacterial culture medium. The magnetic force rotated clockwise, continuously mixing the bacterial culture medium, ensuring uniform distribution of bacteria and facilitating accurate sampling and counting results.

[0038] The kidney chamber 3 and bladder chamber 4 are equipped with a sampling needle 6, typically a heparin cap and needle combination. During the experiment, the needle was positioned below the surface of the bacterial culture medium and was used to aspirate the culture medium at the sampling time, allowing subsequent drug concentration determination and colony counts to be performed. By sampling and measuring the drug concentration in the kidney chamber 3, it was verified that the infusion pump flow rate was set correctly and could simulate the drug concentration in the human body at clinical doses. The sampling needle 6 in the bladder chamber 4 collected samples before each culture medium drain to determine the drug concentration in the culture medium and perform bacterial counts. By measuring the urine concentration in the bladder chamber 4, the urine excretion rate was calculated and verified for consistency with human data.

[0039] In the step S3, about 10 5 ~10 9 The CFU / mL of the tested strains ranged from 1:1 to 1:1, simulating uncomplicated urinary tract infections of varying severity.

[0040] In said S4, the flow rate F4 of the infusion pump four 302 is set to 0.1-0.5 mL / min according to the urine production rate.

[0041] The infusion pump 1 101, infusion pump 2 301, infusion pump 3 201 and infusion pump 4 302 are all peristaltic pumps. The peristaltic pumps are commercially available products and their specific structures will not be described in detail. They are mainly used to adjust the flow rate according to the above experimental design requirements.

[0042] The specific calculation method of the flow rate of each infusion pump in the present invention is as follows: the fixed volume of the kidney chamber 3 is V1 (or Vc), the drug concentration is C1, the initial volume of the bladder chamber 4 is V2, and the drug concentration is C2, wherein the drug concentration in the kidney chamber 3 at a certain time point i is C1i, and the concentration in the bladder chamber 4 is C2 i , the drug concentration in renal compartment 3 at the previous time point is C1 i-1 , the drug concentration in bladder compartment 4 is C2 i-1 , the time interval between time points i-1 and i is Δt, and the drug concentration in the dosing tank 1 is C drug The flow rate of the medication tank 1 pumping into the kidney chamber 3 is F1, the flow rate of the kidney chamber 3 pumping into the waste liquid tank 5 is F2, the flow rate of the drug discharge tank 2 pumping into the kidney chamber 3 is F3, and the flow rate of the kidney chamber 3 pumping into the bladder chamber 4 is F4.

[0043] When simulating the gradual increase of drug concentration in the renal chamber 3, the drug concentration in urine begins to rise. At this time, infusion pump 101 is turned on and infusion pump 3 201 is turned off. At this time, the drug concentration in the renal chamber 3 changes:

[0044] By: (C1 i -C1 i-1 )×V C =Δt×F1×(Cdrug -C1 i-1 )

[0045] Among them, F1=F2+F4 (must satisfy F1≥F4)

[0046] have to:

[0047] The drug concentration and volume pumped into the bladder chamber 4 at time i are:

[0048] (C2 i -C2 i-1 )×V 2i =Δt×F4×(C1 i-1 )

[0049] V2 i =V2 i-1 +F4×Δt

[0050] The urine output during the period t1 to t2 is:

[0051] m2 t1-t2 =V2 t2 ×C2 t2 -V 20 ×C2 t1 The urine excretion rate during the time period t1 to t2 is:

[0052] f% t1-t2 =m2 t1-t2 / (C drug ×V2 t2 )

[0053] According to the urine excretion volume and excretion rate data obtained from the human body during the t1 to t2 period, m2 t1-t2 and f% t1-t2 The target urine output rate and target urine output are calculated by assuming that F1 is input into the renal compartment 3 at a constant rate.

[0054] The drug concentration gradually decreases in the kidney chamber 3: At this time, the drug concentration in the urine begins to decrease. At this time, infusion pump 101 is turned off and infusion pump 3 201 is turned on. Since there is no drug in the drug discharge tank 2, F1 is replaced by F3 in the formula for calculating the concentration in the kidney chamber 3 and the bladder chamber 4, and the rest remains unchanged.

[0055] During the non-urination phase, the volume of V2 gradually increases; after urination, the volume of V2 decreases to the initial volume, and then gradually increases again according to the F4 flow rate setting until the next urination process.

[0056] In one embodiment, the daytime period is 14 hours long, and the nighttime period is 10 hours long. During the daytime period, urination is simulated once every two hours for a total of seven times, and at the end of the nighttime simulation, urination is performed once. The temperature within the bladder chamber 4 and the kidney chamber 3 is maintained at 35±2°C, simulating the ambient temperature within the human body.

[0057] In another embodiment, the bladder chamber 4 is connected to the urinary catheter 402 through a valve 401, and the other end of the urinary catheter 402 is connected to the waste liquid tank 5. In this case, the evaluation method also includes observing the growth of biofilm in the urinary catheter 402 and counting bacteria to evaluate the effect of antibacterial drugs on urinary tract infections related to the urinary catheter 402.

Claims

1. A urinary tract infection model that simulates the in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs, characterized in that: The device comprises a kidney chamber with a bacterial culture medium therein, a bladder chamber, a medication tank and a drug discharge tank. The medication tank and the drug discharge tank are respectively connected to the kidney chamber via a fluid path provided with an infusion pump 1 and an infusion pump 3. The kidney chamber is respectively connected to the bladder chamber and the waste liquid tank via a fluid path provided with an infusion pump 4 and an infusion pump 2. The bladder chamber is connected to the waste liquid tank via a urinary catheter with a valve. The method for using the urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs comprises the following steps: S1. Culture medium preparation: adding bacterial culture medium into the kidney chamber, bladder chamber, medication tank, and drug discharge tank, wherein the volume of bacterial culture medium in the kidney chamber is a fixed value V1, and the volume of bacterial culture medium in the bladder chamber is an initial value V2; S2. Drug preparation: Add the antimicrobial drug to be verified into the dosing tank according to the experimental settings; S3. Strain preparation: Add the test strain into the bladder chamber according to the experimental settings; S4, flow rate setting: set the flow rate F1 of infusion pump 1 from the drug administration tank to the kidney chamber according to the simulated antimicrobial drug concentration rising rate; set the flow rate F3 of infusion pump 3 from the drug discharge tank to the kidney chamber according to the simulated drug concentration falling rate; set the flow rate F4 of infusion pump 4 from the kidney chamber to the bladder chamber according to the simulated urine production rate; set the flow rate F2 of infusion pump 2 from the kidney chamber to the waste liquid tank to maintain the volume V1 of the bacterial culture medium in the kidney chamber fixed; S5. Simulated drug administration: Based on clinical medication, start infusion pump 1 to simulate drug administration and gradually increase the drug concentration in the kidney, while keeping infusion pump 3 closed. After reaching the peak drug concentration in the kidney, turn off infusion pump 1 and start infusion pump 3 to simulate the gradual decrease of drug concentration in the kidney after drug withdrawal. S6. Simulated urination: The infusion pump 4 was kept on throughout the experiment to simulate the kidneys constantly producing urine into the bladder. 24 hours were divided into daytime and nighttime periods. During the daytime period, urination simulation was performed at least once every 4 hours and at most once every 1 hour. The urination simulation involved activating the infusion pump 4 to discharge the bacterial culture medium in the bladder chamber until the remaining volume returned to the initial value V2. The evaluation method of the urinary tract infection model simulating the in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs is as follows: After 1 to 7 days of continuous drug administration simulation according to the method, drug concentration and bacterial count are performed, and the effects of the drug at different doses are evaluated based on the changes in the colony count and the initial bacterial count after 24 hours or at the end point of the urinary catheter.

2. The urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs according to claim 1, characterized in that: The kidney chamber and the bladder chamber are arranged on a magnetic stirrer, and the insides of the kidney chamber and the bladder chamber are arranged with a stirrer matched with the magnetic stirrer.

3. The urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs according to claim 1, characterized in that: Sampling needles are provided on the kidney chamber and the bladder chamber.

4. The urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs according to claim 1, characterized in that: The infusion pump 1, infusion pump 2, infusion pump 3 and infusion pump 4 are all peristaltic pumps.

5. The urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs according to claim 1, characterized in that: The volume of the bacterial culture medium set in the kidney chamber is 100mL~500mL; the initial volume of the culture medium set in the bladder chamber is 1~10mL. In S6, when the total bacterial culture medium injected into the kidney chamber by infusion pump 1 and infusion pump 3 is greater than the bacterial culture medium output from the kidney chamber to the bladder chamber, infusion pump 2 is started to output the excess bacterial culture medium to the waste liquid tank.

6. The urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs according to claim 1, characterized in that: In S3, the amount of the test strain added was 10 5 ~10 9 CFU / mL.

7. The urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs according to claim 1, characterized in that: In the above-mentioned S4, the infusion pump flow rate F4 is set to 0.1-0.5 mL / min according to the urine production rate.

8. The urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs according to claim 1, characterized in that: In S5, the daytime period is 14 hours long, the nighttime period is 10 hours long, and the urination frequency is once every 2 hours, a total of 7 times.

9. The urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs according to claim 1, characterized in that: The temperature in the bladder chamber and the kidney chamber was maintained at 35±2°C.

10. The urinary tract infection model for simulating in vitro pharmacokinetics / pharmacodynamics of antibacterial drugs according to claim 1, characterized in that: The bladder chamber is connected to the catheter through a valve, and the other end of the catheter is connected to the waste liquid tank. At this time, the evaluation method also includes observing the growth of biofilm in the catheter and counting bacteria to evaluate the effect of antibacterial drugs on catheter-related urinary tract infections.

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