Dynamic bionic stomach digestion model and application thereof in microplastic detection

Abstract

The invention discloses a dynamic bionic stomach digestion model and application thereof in micro-plastic detection.The dynamic bionic stomach digestion model comprises a box body, a fixing plate, an artificial stomach model, rolling wheels, a pylorus squeezing plate, a pH meter, a heating mechanism, a gastric acid solution cavity, a gastric juice solution cavity, a peristaltic pump and a micro-dialysis probe, and the rolling wheels are rotationally arranged on the fixing plate and located on the periphery of the outer surface of the artificial stomach model; the pylorus extrusion plate is movably arranged on the fixing plate and located on the outer side of the lower end of the artificial stomach model so as to extrude the lower end of the artificial stomach model, and the gastric acid solution cavity and the gastric juice solution cavity are communicated with the upper end of the artificial stomach model through the peristaltic pump. The method comprises the following steps: adding an aged biofilm-loaded micro-plastic sample or a micro-plastic sample loaded with heavy metal and the like and agar balls into an artificial stomach model, digesting in the artificial stomach model, detecting discharged liquid, solids and the like after digestion is completed, detecting the concentration change of digestive juice, micro-plastic particles and micro-plastic precipitates on agar ball particles, and determining the concentration change of the micro-plastic precipitates on the micro-plastic particles and the agar balls. Product identification and the like.

Description

TECHNICAL FIELD

[0001] The present application relates to a dynamic biomimetic gastric digestion model and its application in microplastic detection. BACKGROUND

[0002] Microplastics (plastic particles with particle size <5 mm) have been widely found in water, soil and the entire food chain as emerging environmental pollutants. Humans mainly ingest microplastics through the oral cavity via diet (such as seafood, salt, drinking water and packaged food), with an estimated annual intake of 39,000-52,000 particles per person. After ingestion, microplastics first enter the gastrointestinal digestion system, and their digestion rules under the acidic environment and mechanical peristalsis in the stomach directly determine the release efficiency and bioavailability of harmful additives (such as plasticizers, bisphenol A) and adsorbed pollutants (such as heavy metals), which is the key first step in assessing the human health risk.

[0003] (1) Over-simplified simulation system: Most studies use static single gastric juice systems (such as hydrochloric acid-pepsin solution), completely ignoring the dynamic digestion process in the human stomach, including gastric emptying timing, continuous mechanical peristalsis physical friction, and complex biological molecule interactions (such as adsorption barriers of gastric mucin layer, and encapsulation effects of food lipids). This over-simplified static model cannot simulate the real pollutant migration interface, resulting in significant deviations in the release kinetics data of additives from the real physiological environment.

[0004] (2) Ignoring the aging state of the material environment: Currently, most studies use original, unaged microplastics as experimental objects. However, microplastics in the environment generally undergo processes such as ultraviolet light aging, mechanical wear, and biofilm attachment, which significantly change their surface physicochemical properties (such as specific surface area, functional groups, and crystallinity), and the migration behavior of additives also intensifies. Existing models fail to incorporate these key environmental aging factors, greatly underestimating the release risk of microplastics in actual exposure scenarios.

[0005] Therefore, developing a dynamic biomimetic gastric digestion model that can simultaneously simulate gastric physical peristalsis, chemical digestion, and dynamic acid-enzyme environmental changes, and can monitor the in-situ release and transformation of pollutants from environmentally aged microplastics, is crucial for advancing the human health risk assessment of microplastics. SUMMARY

[0006] To solve the defects in the prior art, the present application provides a dynamic biomimetic gastric digestion model and its application in microplastic detection.

[0007] To solve the above technical problems, the present application provides the following technical solutions: The first object of the present application provides a dynamic bionic stomach digestion model, which comprises a box body, a fixed plate, an artificial stomach model, a peristalsis jacket layer, a pyloric extrusion plate, a pH meter, a heating mechanism, a gastric acid solution cavity, a gastric juice solution cavity and a peristaltic pump, the fixed plate is vertically arranged in the box body, the upper end and the lower end of the artificial stomach model are arranged on the fixed plate, the peristalsis jacket layer comprises a jacket layer main body, rollers and a pressure sensor, the jacket layer main body is arranged on the fixed plate and is sleeved on the outer surface of the artificial stomach model, the rollers are arranged in the circumferential direction of the jacket layer main body and drive the rollers to move back and forth in the direction towards the center of the artificial stomach model and the direction away from the center of the artificial stomach model through a driving mechanism, the pressure sensor is arranged on the rollers, the pyloric extrusion plate is movably arranged on the fixed plate and is located outside the lower end of the artificial stomach model, so as to extrude the lower end of the artificial stomach model, the pH meter is arranged on the inner wall of the artificial stomach model, and the heating mechanism, the gastric acid solution cavity, the gastric juice solution cavity and the peristaltic pump are arranged on the inner wall of the box body, the gastric acid solution cavity and the gastric juice solution cavity are connected with the upper end of the artificial stomach model through the peristaltic pump.

[0008] Preferably, the box body is made of transparent acrylic material.

[0009] Preferably, the upper end of the artificial stomach model is provided with a sample inlet, the sample inlet is provided with a sample adding port, the lower end of the artificial stomach model is provided with a sample outlet, and the sample outlet is provided with a microdialysis probe for collecting a digestive fluid sample.

[0010] Preferably, the inner wall of the artificial stomach model is of a corrugated structure, the artificial stomach model is made of silica gel material, and the inner wall of the artificial stomach model is provided with sodium alginate-gastric mucin gel.

[0011] Preferably, the jacket layer main body comprises two semicircular arc plates, the two semicircular arc plates are connected through a hinge on one side and a quick lock mechanism on the other side, and form a complete cylindrical jacket; and the jacket layer main body is rigidly fixed on the vertical fixed plate through a support.

[0012] Preferably, the roller comprises an outer rubber layer and a rigid hub, the outer rubber layer is sleeved on the rigid hub, the roller is driven by a micro stepping motor or a servo motor, the radial direction of the roller is consistent with the movement direction of the roller, and the pressure sensor is arranged between the outer rubber layer and the rigid hub.

[0013] Preferably, the heating mechanism comprises a heating unit and a temperature sensor, and the heating unit and the temperature sensor are arranged on the inner wall of the box body.

[0014] The second object of the present application provides an application of the dynamic bionic stomach digestion model in microplastic detection, which comprises the following steps: S1, aging treatment of microplastics under irradiation of an ultraviolet light source for a set time; S2, immerging the microplastics after aging treatment into the culture medium containing E. coli / Vibrio, forming a biofilm, and sterilizing; or co-incubating the microplastics after aging treatment with adsorbed pollutants to form a supported microplastic; S3, adding the microplastics and agar balls obtained in step S2 from the sample port into the artificial stomach model, then adjusting the pH value according to the digestion stage, adjusting the temperature and pepsin concentration according to the linkage of temperature and enzyme activity, and performing peristalsis by roller extrusion of the artificial stomach model; S4, collecting the digestion liquid samples at intervals between the microdialysis probes and performing analysis, so as to detect the microplastics and transformation products.

[0015] Preferably, in the step S1, the wavelength of the ultraviolet light source is 280-316 nm, and the intensity of the ultraviolet light source is 0.8 W / m²; the aging treatment is performed under RH 30%-90% humidity cycle.

[0016] Preferably, in the step S2, the temperature of the culture medium culture is 37℃, and the culture time is 48 h.

[0017] Preferably, in the step S3, the pH value is 3.0 (0-1 h), 2.0 (1-3 h), and 1.5 (3-6 h) in sequence, simulating the change of postprandial acidity; under constant temperature of 37℃, the pepsin concentration is 0 mg / mL at 0 h, 1.5 mg / mL at 2 h, and 3.0 mg / mL at 4 h according to the time-activity curve.

[0018] Compared with the prior art, the present application has the following beneficial effects: In the present application, the microplastics samples after aging, loading biofilm, or loading heavy metals, etc. are added into the artificial stomach model together with agar balls, and digestion is performed in the artificial stomach model. After the digestion is completed, the discharged liquid, solid, etc. are detected, the concentration change of microplastic precipitates on the digestion liquid and microplastic particles, agar ball particles, and product identification, etc. are detected. Quantitative detection is performed by liquid, liquid mass, gas mass, etc. instruments, and the change law at different digestion times is detected in real time. BRIEF DESCRIPTION OF DRAWINGS

[0019] Figure 1 is a structural schematic diagram of a dynamic bionic stomach digestion model of the present application; Figure 2 is a structural schematic diagram of a peristaltic jacket layer in the present application. DETAILED DESCRIPTION

[0020] The preferred embodiments of the present application will be described below in conjunction with the accompanying drawings, and it should be understood that the preferred embodiments described herein are only used to illustrate and explain the present application, and are not used to limit the present application.

[0021] In the description of the present application, it should be understood that the terms "upper", "lower", "left", "right", etc. indicate the orientation or positional relationship based on the drawings accompanying the specification Figure 1 The orientation or positional relationship shown is only for the convenience of describing the present application and simplifying the description, and does not indicate or imply that the device or element referred to must have a particular orientation, be constructed and operated in a particular orientation, and therefore cannot be understood as a limitation of the present application.

[0022] In the description of the present application, it should be noted that unless otherwise explicitly specified and limited, the terms "mounting", "connecting", "connecting" should be understood broadly, for example, it can be fixedly connected, or it can be detachably connected, or integrally connected; it can be directly connected, or indirectly connected through an intermediate medium. For those skilled in the art, the specific meaning of the above terms in the present application can be understood according to the specific circumstances.

[0023] As Figures 1 to 2 shown, the embodiment provides a dynamic bionic stomach digestion model, which comprises a box body 1, a fixed plate 2, an artificial stomach model 5, a peristaltic jacket layer, a pyloric extrusion plate 8, a pH meter 9, a heating mechanism, a gastric acid solution cavity 13, a gastric juice solution cavity 14, a peristaltic pump 16, the fixed plate 2 is vertically arranged in the box body 1 and is fixed by a hinge 3, the upper end and the lower end of the artificial stomach model 5 are arranged on the fixed plate 2, the peristaltic jacket layer comprises a jacket layer main body, a roller 6 and a pressure sensor 7, the jacket layer main body is arranged on the fixed plate 2 and is sleeved on the outer surface of the artificial stomach model 5, the rollers 6 are arranged along the circumferential direction of the jacket layer main body and drive the rollers to move back and forth in the direction towards the center of the artificial stomach model and the direction away from the center of the artificial stomach model through a driving mechanism, the pressure sensor 7 is arranged on the roller 6, the pyloric extrusion plate 8 is movably arranged on the fixed plate 2 and is located outside the lower end of the artificial stomach model 5, so as to extrude the lower end of the artificial stomach model 5, the pH meter 9 is arranged on the inner wall of the artificial stomach model 5, the heating mechanism, the gastric acid solution cavity 13, the gastric juice solution cavity 14 and the peristaltic pump 16 are arranged on the inner wall of the box body 1, and the gastric acid solution cavity 13 and the gastric juice solution cavity 14 are connected with the upper end of the artificial stomach model 5 through the peristaltic pump 16.

[0024] In the embodiment, the box body 1 is made of transparent acrylic material, which provides a full artificial stomach environment, facilitates observation and constant temperature. A side door 16 is movably arranged on the left side of the box body, which facilitates the installation of various components.

[0025] In the embodiment, the upper end of the artificial stomach model 5 is provided with a sample inlet 4, the sample inlet 4 is provided with a sample adding port 15, the lower end of the artificial stomach model 5 is provided with a sampling port 10, and the sampling port is provided with a microdialysis probe for collecting digestive fluid samples.

[0026] In the embodiment, the inner wall of the artificial stomach model 5 is a pleated structure, the artificial stomach model 5 is made of silica gel material, and the inner wall of the artificial stomach model 5 is provided with sodium alginate-gastric mucin gel.

[0027] Sodium alginate is a natural polysaccharide extracted from brown algae. Its most famous feature is that it can quickly form a hydrogel when it encounters calcium ions Ca 2+ Ion cross-linking occurs afterwards, rapidly forming a hydrogel. This is the principle behind the fruit caviar in the molecular cuisine. It has a high gel strength, but the biocompatibility is general. Gastric mucin is the main component of gastric mucosa secretion, and is a key functional molecule of the mucus layer of the human digestive tract. It has excellent lubricity, biological adhesion, and protective effects, and can resist the degradation of gastric acid. High-purity and active gastric mucin is usually extracted from animal tissues such as pig stomachs. The combination of the two provides the structural strength and stability of the gel, while the gastric mucin provides excellent biological lubricity, mucosal adhesion, and biocompatibility. This composite material can better simulate the real mucus layer of the human stomach, and is therefore very valuable in the biomedical field. The gel is prepared according to specific purposes, such as the required hardness, lubricity, and drug release rate. The preparation process involves the control of multiple parameters such as the concentration ratio of the two solutions, the concentration of the cross-linking agent such as calcium chloride, the pH value, and the reaction time, and is not limited in particular.

[0028] In the embodiment, the heating mechanism includes a heating unit 11 and a temperature sensor 12, which are arranged on the inner wall of the box body 1.

[0029] In the embodiment, the peristaltic jacket layer is a separate, openable and closable ring structure wrapped outside the artificial stomach model, and simulates the directional peristaltic wave of the stomach from the cardia to the pylorus through the coordinated movement of multiple rollers. The pressure sensor is integrated on the roller to realize in-situ and real-time pressure monitoring.

[0030] Specifically, the jacket layer body adopts light weight, high strength medical grade aluminum alloy or engineering plastics such as PEEK as the frame. These materials have high mechanical strength, corrosion resistance, and good biocompatibility. The jacket layer body includes two semicircular arc plates connected by a hinge on one side and a quick lock mechanism on the other side, forming a complete cylindrical jacket. The jacket layer body is rigidly fixed to the vertical fixed plate 2 through a support. This design makes the jacket layer body easy to open and close, facilitating the installation, replacement, and cleaning of the artificial stomach model. The quick lock mechanism can use any product of existing technology that can achieve this function, and is not limited in particular.

[0031] Specifically, the rollers are arranged in 26 columns and 4 rows along the circumferential direction of the jacket layer body, the rollers 6 are driven by micro stepping motors or servo motors, and the radial direction of the rollers 6 is consistent with the movement direction thereof. The control system controls the rollers in sequence to perform the cyclic action of “extrusion-keeping-release-resetting” according to a preset program, to form a pushing force like a wave from front to back, thereby highly simulating the real gastric peristalsis. The independent control of the rollers can simulate complex and real gastric peristalsis waveforms, which cannot be realized by single-point extrusion or simple rollers, and is closer to the physiological reality.

[0032] The roller 6 comprises an outer rubber layer and a rigid hub, the outer rubber layer is sleeved on the rigid hub, and the outer rubber layer is 2-5 mm thick medical grade silicone or thermoplastic elastomer. Such soft material can simulate the soft touch of fingers, avoid scratching the silicone stomach model, and also provide sufficient friction to push the contents.

[0033] The pressure sensor 7 is embedded between the outer rubber layer and the rigid hub of each roller. This design enables the sensor to directly sense the positive pressure applied to the surface of the stomach model, and the measurement is accurate and does not interfere with the peristalsis function.

[0034] The pressure sensor in each roller is connected to a micro slip ring at the center of the roller shaft through a flexible printed circuit. The slip ring ensures that the electrical signal can be stably transmitted when the roller is continuously rotating, and the wire will not be twisted off. The signal line is then led out from the slip ring and gathered into the main wire harness on the jacket layer frame. The signals of all sensors are transmitted to the data acquisition and control unit outside the box through a shielded cable, and the unit includes: Signal amplifier: amplifies weak sensor signals.

[0035] Analog-to-digital converter: converts analog signals to digital signals.

[0036] Microprocessor (such as ARM or FPGA): real-time processing of all pressure data.

[0037] Communication interface (such as USB or Ethernet): upload data to the host computer (computer) for display, recording and storage.

[0038] Based on real-time pressure data, the system forms a closed loop control. For example, when the pressure at a certain place exceeds the preset safety threshold (simulating discomfort or spasm), the control system can automatically adjust the extrusion force or speed of the corresponding roller, making the simulation process more intelligent and safer.

[0039] The embodiment provides an application of a dynamic bionic stomach digestion model in microplastic detection, and comprises the following steps: S1, irradiate the microplastics under a UV light source for a set time for aging treatment; the wavelength of the UV light source is 280-316 nm, and the intensity of the UV light source is 0.8 W / m²; during the aging treatment, RH 30%-90% humidity cycling is performed; S2, immerse the microplastics after the aging treatment in a culture medium containing E. coli / Vibrio to form a biofilm, and perform sterilization treatment; the temperature of the culture medium culture is 37°C, and the culture time is 48 h; S3, add the microplastics and agar balls obtained in step S2 from the sample port to an artificial stomach model, then adjust the pH value according to the digestion stage, adjust the temperature and pepsin concentration according to the temperature-enzyme activity linkage, and pass through the roller to extrude the artificial stomach model to peristalsis; the pH value is 3.0, 2.0, and 1.5 in turn, simulating the change in acidity after eating; at 37°C constant temperature, the pepsin concentration is 0, (2 h, 1.5 mg / mL), and (4 h, 3.0 mg / mL) in turn according to the time-activity curve; the agar balls are introduced to simulate the food stimulating the peristalsis of the artificial stomach, and simulate the digestion process of the food contaminated by the microplastics in the human body; S4, collect the digestion liquid samples at intervals between the microdialysis probes and analyze them, so as to detect the microplastics and the conversion products. The analysis includes synchronous monitoring of the release and conversion process of the microplastic additives: (a) In-situ real-time monitoring is performed by using a double-channel fluorescence probe: probe A: naphthalene dicarboxamide derivative (specifically binds to free bisphenol A, fluorescence λex / λem=365 / 450 nm) and probe B: zinc(II)-dipyridylamine complex (quenching type probe, restores fluorescence when encountering phthalate hydrolysis product, λex / λem=340 / 411 nm), the fluorescence intensity changes of the two channels are monitored in real time by an optical fiber fluorescence spectrometer, and the release kinetics of the pollutants is fitted based on the two-phase first-order kinetic equation C_t = C_f(1-e^(-k1t)) + C_s(1-e^(-k2t)) (C_f: fast release phase concentration; Cs: slow release phase concentration; k1, k2: rate constant; e is a natural constant (about equal to 2.71828), which is the base number of the natural exponential function; t is the time variable); (b) Non-targeted conversion product tracking is performed based on high-resolution mass spectrometry: after the collected digestion liquid samples are subjected to derivatization (BSTFA+1%TMCS) treatment, high-resolution mass spectrometry (Q-Exactive HF-X) is used for full scan data acquisition, and unknown conversion products are identified by comparing the plasticizer conversion library (phthalic acid ester→phthalic acid monoester→phthalic acid) and the bisphenol A conversion path (BPA→BPA quinone→BPA dimer) through Compound Discoverer software.

[0040] As a preferred embodiment, the rest is the same as the previous embodiment, except that the step S2 of the present embodiment co-incubates the microplastics after aging treatment with adsorbed pollutants (such as mycotoxins, heavy metals, viruses, etc.) to form supported microplastics.

[0041] Finally, it should be noted that: the above only for the preferred embodiments of the present application, and not for limiting the present application, although the foregoing detailed description of the present application is made with reference to the foregoing embodiments, for those skilled in the art, it still can be modified, or part of the technical features of the equivalent replacement of the technical solutions described in the foregoing embodiments. Any modification, equivalent replacement, improvement, etc. within the spirit and principles of the present application shall be included within the scope of protection of the present application.

Claims

1. A dynamic in-vivo gastric digestion model, characterized in that, The utility model relates to a kind of artificial stomach model, including box (1), fixed plate (2), artificial stomach model (5), peristalsis jacket layer, pyloric extrusion plate (8), pH meter (9), heating mechanism, gastric acid solution cavity (13), gastric juice solution cavity (14), peristaltic pump (16), the fixed plate (2) vertically is set in box (1), the upper end of the artificial stomach model (5), lower end is set on fixed plate (2), the peristalsis jacket layer includes jacket layer main body, idler (6) and pressure sensor (7), the jacket layer main body is set on fixed plate (2) and is set on the outer surface of artificial stomach model (5), the idler (6) is arranged along the circumferential direction of jacket layer main body and drives idler to move back and forth in the direction towards the center of artificial stomach model and the direction away from the center of artificial stomach model by driving mechanism, the pressure sensor (7) is set on idler (6), the pyloric extrusion plate (8) is movably set on fixed plate (2) and is located the lower end outside of artificial stomach model (5), to extrude the lower end of artificial stomach model (5), the pH meter (9) is set on the inner wall of artificial stomach model (5), the heating mechanism, gastric acid solution cavity (13), gastric juice solution cavity (14), peristaltic pump (16) are set on the inner wall of box (1), the gastric acid solution cavity (13), gastric juice solution cavity (14) are connected with the upper end of artificial stomach model (5) by peristaltic pump (16), the idler (6) is rotatably set on fixed plate (2) and is located the four around of outer surface of artificial stomach model (5).

2. A dynamic in-vitro bionic gastric digestion model according to claim 1, wherein, The box (1) is made of transparent acrylic material.

3. A dynamic in-vitro bionic gastric digestion model according to claim 2, wherein, The upper end of the artificial stomach model (5) is provided with a sample inlet (4), and the sample inlet (4) is provided with a sample adding port (15). The lower end of the artificial stomach model (5) is provided with a sampling port (10), and the sampling port is provided with a microdialysis probe for collecting digestive fluid samples.

4. A dynamic in-vitro bionic gastric digestion model according to claim 3, wherein, The inner wall of the artificial stomach model (5) is of a wrinkled structure, and the artificial stomach model is made of silica gel. The inner wall of the artificial stomach model (5) is provided with a sodium alginate-gastric mucin gel.

5. A dynamic in-vitro bionic gastric digestion model according to claim 4, wherein, The jacket layer main body includes two semicircular arc plates, which are connected by a hinge on one side and a quick lock mechanism on the other side to form a complete cylindrical jacket. The jacket layer main body is rigidly fixed on the vertical fixed plate (2) by a support.

6. A dynamic in-vitro bionic gastric digestion model according to claim 5, wherein, The idler (6) includes an outer rubber layer and a rigid hub. The outer rubber layer is sleeved on the rigid hub. The idler (6) is driven by a micro stepping motor or a servo motor. The radial direction of the idler (6) is consistent with its movement direction. The pressure sensor (7) is arranged between the outer rubber layer and the rigid hub.

7. A dynamic in-vitro bionic gastric digestion model according to claim 6, wherein, The heating mechanism includes a heating unit (11) and a temperature sensor (12), which are arranged on the inner wall of the box (1).

8. Use of a dynamic in-vitro gastric digestion model according to claim 7 for the detection of microplastics, characterized in that, The utility model includes the following steps: S1, irradiate microplastics under ultraviolet light source for a set time for aging treatment; S2, immerse the microplastics after aging treatment into the culture medium containing E. coli / Vibrio to form a biofilm, and sterilize; or co-incubate the microplastics after aging treatment with adsorbed pollutants to form a loaded microplastic; S3, add the microplastics and agar balls obtained in step S2 from the sample port into an artificial stomach model, then adjust the pH value according to the digestion stage, adjust the temperature and pepsin concentration according to the linkage of temperature and enzyme activity, and make the artificial stomach model peristalsis by roller extrusion; S4, collect the digestion liquid samples at intervals by microdialysis probe and analyze to detect the microplastics and transformation products.

9. Use according to claim 8, characterized in that, In the step S1, the wavelength of the ultraviolet light source is 280-316 nm, and the intensity of the ultraviolet light source is 0.8 W / m²; the aging treatment is performed under RH 30%-90% humidity cycle; In the step S2, the temperature of the culture medium is 37℃, and the culture time is 48 h; In the step S3, the pH value is 3.0 (0-1 h), 2.0 (1-3 h), and 1.5 (3-6 h) in sequence to simulate the change of acidity after eating; under constant temperature of 37℃, the pepsin concentration is 0 mg / mL at 0 h, rises to 1.5 mg / mL at 2 h, and rises to 3.0 mg / mL at 4 h according to the time-activity curve.

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