Near-infrared two-region fluorescent microsphere as well as preparation method and application thereof

Abstract

The invention relates to a near-infrared two-region fluorescent microsphere as well as a preparation method and application thereof. The near-infrared two-region fluorescent microspheres comprise polymer spheres formed by winding polystyrene chains and fluorescent dye VIX-S distributed among gaps of the polystyrene chains; the structural formula of the fluorescent dye VIX-S is as shown in a formula (1): # imgabs0 #. The near-infrared two-region fluorescent microsphere disclosed by the invention can realize real-time dynamic imaging under the conditions that the obtained fluorescent brightness is relatively high and the signal-to-background ratio is enough.

Description

Technical Field

[0001] This application belongs to the technical field of biochemical analysis, and specifically relates to a near-infrared second-region fluorescent microsphere, its preparation method and application. Background Technique

[0002] The study of intestinal kinetics is of great significance for understanding the mechanisms and treatment strategies of diseases such as gastrointestinal dysfunction, constipation, and intestinal obstruction. Intestinal peristalsis is the basis of gastrointestinal motility function, which affects the passage and excretion of food, liquid, gas, and waste. Changes in intestinal motility are often one of the symptoms of various gastrointestinal diseases, such as chronic constipation, irritable bowel syndrome, gastric emptying disorders, and intestinal obstruction. These diseases are often accompanied by symptoms such as abdominal distension, abdominal pain, and abnormal defecation, seriously affecting the quality of life of patients. Traditional intestinal kinetics monitoring techniques include X-ray imaging, ultrasound examination, magnetic resonance imaging (MRI), and gastrointestinal pressure measurement. Although these monitoring techniques can provide relevant data on intestinal motility, they all have certain limitations. For example, although X-ray and CT scans can provide image information of the intestine, due to their radiation risks, long-term use poses a potential threat to human health; the resolution of ultrasound examination and MRI imaging is relatively low, making it difficult to accurately reflect the dynamic changes in the intestine.

[0003] In recent years, with the progress of biomedical imaging technology, especially the development of near-infrared second-region (NIR-II) fluorescence imaging, more and more studies have begun to use fluorescent probes and markers for real-time monitoring of intestinal kinetics. Compared with traditional imaging techniques, near-infrared second-region fluorescence imaging has become an ideal non-invasive monitoring method due to its lower tissue autofluorescence background, high-depth penetration, and higher imaging resolution. Summary of the Invention

[0004] This application provides a near-infrared second-region fluorescent microsphere, its preparation method and application, which can achieve real-time dynamic imaging under the condition of obtaining sufficient fluorescence brightness and signal-to-background ratio.

[0005] In the first aspect, this application provides a near-infrared second-region fluorescent microsphere, which includes a polymer sphere formed by winding polystyrene chains, and a fluorescent dye VIX-S distributed between the voids of the polystyrene chains; the structural formula of the fluorescent dye VIX-S is shown in Formula (1):

[0006]

[0007] In some embodiments, the average particle size of the near-infrared second-region fluorescent microsphere is 80-120 nm.

[0008] In a second aspect, the present application provides a method for preparing the near-infrared second-region fluorescent microspheres of the first aspect of the present application, including: mixing an aqueous suspension containing polystyrene microspheres with an aqueous solution containing a surfactant to obtain a first dispersion; mixing an organic solution containing the fluorescent dye VIX-S with the first dispersion, and then removing the organic solvent through evaporation treatment to obtain a second dispersion. The structural formula of the fluorescent dye VIX-S is shown in Formula (1):

[0009]

[0010] Performing centrifugation treatment, washing treatment and drying treatment on the second dispersion to obtain the near-infrared second-region fluorescent microspheres.

[0011] In some embodiments, the average particle size of the polystyrene microspheres is 80 nm - 120 nm.

[0012] In some embodiments, the mass concentration of the polystyrene microspheres in the aqueous suspension containing polystyrene microspheres is 2% - 3%.

[0013] In some embodiments, the concentration of the fluorescent dye VIX-S in the organic solution containing the fluorescent dye VIX-S is 0.08 g / L - 0.12 g / L.

[0014] In some embodiments, the volume ratio of the organic solution containing the fluorescent dye VIX-S to the first dispersion is 1:(4 - 6).

[0015] In some embodiments, the mass concentration of the surfactant in the aqueous solution containing the surfactant is 0.2% - 0.3%; the surfactant includes sodium dodecyl sulfate.

[0016] In some embodiments, the organic solvent in the organic solvent containing the fluorescent dye VIX-S includes one or more of tetrahydrofuran, chloroform and acetone.

[0017] In some embodiments, the preparation method further includes: after mixing the organic solution containing the fluorescent dye VIX-S with the first dispersion, performing ultrasonic treatment; the time of the ultrasonic treatment is 4 min - 6 min.

[0018] In some embodiments, the rotation speed of the centrifugation treatment is 8000 rpm - 12000 rpm, and the time of the centrifugation treatment is 13 min - 17 min.

[0019] In some embodiments, the temperature of the drying treatment is 35°C - 45°C, and the time of the drying treatment is 0.8 - 1.2 h.

[0020] In a third aspect, the embodiments of the present application provide an application of the near-infrared second-region fluorescent microspheres of the first aspect of the present application as an intestinal motility monitoring tool.

[0021] In the embodiments of the present disclosure, by combining polystyrene with the fluorescent dye VIX-S, real-time dynamic imaging can be achieved when obtaining relatively high fluorescence brightness and sufficient signal-to-background ratio. Description of the Drawings

[0022] By reading the following detailed description of the preferred embodiments, various other advantages and benefits will become clear to those of ordinary skill in the art. The drawings are only for the purpose of showing the preferred embodiments and are not considered to limit the scope of the present application. Moreover, throughout the drawings, the same reference numerals are used to represent the same components.

[0023] Figure 1 It is a scanning electron microscope image of polystyrene microspheres in Example 1 of the present application.

[0024] Figure 2 It is a scanning electron microscope image of the second near-infrared region fluorescent microspheres prepared in Example 1 of the present application.

[0025] Figure 3 It is an NIR-II fluorescence image of the second near-infrared region fluorescent microspheres prepared in Example 1 of the present application in a phosphate buffered saline (PBS) solution.

[0026] Figure 4 It is a fluorescence imaging map of the distribution of the second near-infrared region fluorescent microspheres prepared in Example 1 of the present application in the intestines of healthy mice at different time points ( Figure 4 A), the selected fluorescent spots ( Figure 4 B left) and their movement trajectory maps ( Figure 4 B right).

[0027] Figure 5 It is the fluorescence imaging map of the distribution of the second near-infrared region fluorescent microspheres prepared in Example 1 of the present application in constipated mice ( Figure 5 A) and constipated mice recovering from constipation ( Figure 5 B) models at different time points.

[0028] Figure 6 It is the selected fluorescent spots and their movement trajectory maps of the second near-infrared region fluorescent microspheres prepared in Example 1 of the present application in the constipated mouse model ( Figure 6 A), and the selected fluorescent spots and their movement trajectory maps in the constipated mouse recovering from constipation model ( Figure 6 B). Detailed Embodiments

[0029] Hereinafter, embodiments of the present application will be described with reference to the drawings. However, it should be understood that these descriptions are only exemplary and are not intended to limit the scope of the present application.

[0030] Unless otherwise defined, all technical and scientific terms used in this application have the same meaning as commonly understood by those skilled in the technical field to which this application belongs; the terms used in this application are only for the purpose of describing specific embodiments and are not intended to limit this application; the terms "comprising" and "having" and any variations thereof in the specification and claims of this application and the above description of the drawings are intended to cover non-exclusive inclusion.

[0031] Reference to "embodiment" in this application means that a particular feature, structure, or characteristic described in connection with the embodiment can be included in at least one embodiment of this application. The phrase appears in various places in the specification and does not necessarily refer to the same embodiment, nor is it an independent or alternative embodiment mutually exclusive with other embodiments. Those skilled in the art will explicitly and implicitly understand that the embodiments described in this application can be combined with other embodiments.

[0032] In the description of the embodiments of this application, the term "and / or" is merely a description of the association relationship of associated objects, indicating that three relationships can exist. For example, A and / or B can represent three situations: A exists alone, A and B exist simultaneously, and B exists alone.

[0033] In the description of the embodiments of this application, the term "plurality" means two or more (including two).

[0034] Unless otherwise specified, the test temperature of each parameter mentioned in this disclosure is 25 °C.

[0035] To achieve precise and real-time monitoring of intestinal dynamics, a new type of fluorescent marker needs to be designed so that it can perform highly sensitive imaging in the NRI-II band without interfering with the normal physiological functions of the intestine. Fluorescent polystyrene microspheres, as a common type of nanoparticles, have good optical properties, chemical stability, and biocompatibility and have been widely used in biomedical imaging. Especially with the support of NIR-II fluorescence technology, near-infrared second-region fluorescent microspheres can provide deeper tissue imaging and higher imaging resolution. Combining NIR-II fluorescent probes with fluorescent microspheres can provide a new non-invasive monitoring technology for intestinal dynamics.

[0036] This application provides a near-infrared second-region fluorescent microsphere. The near-infrared second-region fluorescent microsphere includes a polymer sphere formed by winding polystyrene chains and a fluorescent dye VIX-S distributed between the voids of the polystyrene chains; the structural formula of the fluorescent dye VIX-S is shown in Formula (1):

[0037]

[0038] In the embodiments of the present disclosure, by combining polystyrene with the fluorescent dye VIX-S, real-time dynamic imaging can be achieved under the conditions of obtaining relatively high fluorescence brightness and sufficient signal-to-background ratio.

[0039] The longer the exposure time, the higher the fluorescence brightness, but the lower the imaging frame rate. The near-infrared II fluorescent microspheres in the embodiments of the present application have good photochemical properties, which can endow the near-infrared II fluorescent microspheres with excellent fluorescence brightness. Therefore, imaging can be performed under a lower exposure time, that is, real-time dynamic imaging can be achieved under the condition of sufficient fluorescence brightness obtained.

[0040] In addition, the near-infrared II fluorescent microspheres in the embodiments of the present application have good chemical stability and photostability in artificially simulated gastric juice and intestinal juice, and thus can be used for long-term in-vivo imaging; the emission wavelength of the near-infrared II fluorescent microspheres reaches the near-infrared II region, with strong tissue penetration ability and high signal-to-background ratio, which can effectively reduce the influence of the autofluorescence background signal in the living body.

[0041] It should be noted that the fluorescent dye VIX-S shown in formula (1) in the embodiments of the present application can be prepared by referring to the preparation method in the patent document with the publication number CN116041306A, and details will not be elaborated here.

[0042] In some embodiments, the average particle size of the near-infrared II fluorescent microspheres can be 80nm - 120nm, for example, it can be 80nm, 90nm, 100nm, 110nm, 120nm, or a range composed of any of the above values.

[0043] [Preparation Method of Near-Infrared II Fluorescent Microspheres]

[0044] The present application provides a preparation method of near-infrared II fluorescent microspheres, including the following steps:

[0045] Mix a water suspension containing polystyrene microspheres with an aqueous solution containing a surfactant to obtain a first dispersion;

[0046] After mixing an organic solution containing the fluorescent dye VIX-S with the first dispersion, remove the organic solvent through evaporation treatment to obtain a second dispersion. The structural formula of the fluorescent dye VIX-S is as shown in formula (1):

[0047]

[0048] Perform centrifugation treatment, washing treatment, and drying treatment on the second dispersion to obtain near-infrared II fluorescent microspheres.

[0049] The preparation method of the embodiment of the present application has a simple process, can improve the encapsulation efficiency of polystyrene microspheres for the fluorescent dye VIX-S, and obtain near-infrared second-region fluorescent microspheres with stable fluorescence signals and uniform particle size distributions.

[0050] The polystyrene microspheres in the embodiment of the present application have a polymer chain structure, form a spherical shape through winding, and can be swollen by adding an organic solvent to make them relatively loose; by stirring and swelling, the fluorescent dye VIX-S can be uniformly dispersed in the chain segments of the polystyrene microspheres. The fluorescent dye VIX-S can be distributed inside the polystyrene microspheres and on the surface of the polystyrene microspheres. However, usually, the fluorescent dye VIX-S on the surface of the polystyrene microspheres will be removed by washing treatment.

[0051] In some embodiments, the average particle size of the polystyrene microspheres can be 80 nm - 120 nm, for example, it can be 80 nm, 90 nm, 100 nm, 110 nm, 120 nm, or a range composed of any of the above values.

[0052] It should be noted that the morphology and size of the polystyrene microspheres in the embodiment of the present application are comparable to those of the near-infrared second-region fluorescent microspheres obtained by dyeing.

[0053] In some embodiments, the mass concentration of the polystyrene microspheres in the aqueous suspension containing the polystyrene microspheres can be 2% - 3%.

[0054] In some embodiments, the concentration of the fluorescent dye VIX-S in the organic solution containing the fluorescent dye VIX-S can be 0.08 g / L - 0.12 g / L.

[0055] In some embodiments, the volume ratio of the organic solution containing the fluorescent dye VIX-S to the first dispersion can be 1:(4 - 6), for example, it can be 1:4, 1:4.5, 1:5, 1:5.5, 1:6, or a range composed of any of the above values.

[0056] When the volume ratio of the organic solution containing the fluorescent dye VIX-S to the first dispersion is within the above range, the amount of the organic solvent can be within a suitable range to adjust the ratio of the polystyrene microspheres to the organic solvent, thereby reducing the dissolution of the polystyrene polymer segments and enabling the polystyrene microspheres to be fully swollen.

[0057] In some embodiments, the mass concentration of the surfactant in the aqueous solution containing the surfactant can be 0.2% - 0.3%. By adding the surfactant with the above mass concentration, the polystyrene microspheres can be fully dispersed.

[0058] In some embodiments, the surfactant includes sodium dodecyl sulfate.

[0059] In some embodiments, the organic solvent in the organic solvent containing the fluorescent dye VIX-S may include one or more of tetrahydrofuran, chloroform, and acetone.

[0060] In some embodiments, the preparation method may further include: after mixing the organic solution containing the fluorescent dye VIX-S with the first dispersion liquid, performing ultrasonic treatment. Through ultrasonic treatment, the fluorescent dye VIX-S can be better mixed with the polystyrene microspheres, and the fluorescent dye VIX-S can enter the interior of the polystyrene microspheres better.

[0061] In some embodiments, the time of ultrasonic treatment can be 4 min - 6 min.

[0062] In some embodiments, the preparation method may further include stirring the mixed solution of the organic solution containing the fluorescent dye VIX-S and the first dispersion liquid before removing the organic solvent by evaporation treatment. Thereby, the polystyrene microspheres can be fully swollen to adsorb the fluorescent dye VIX-S.

[0063] Optionally, the temperature of the stirring treatment can be 25°C - 35°C; the time of the stirring treatment can be 2.5 h - 3.5 h.

[0064] In some embodiments, the evaporation treatment method can be rotary evaporation, and the temperature of the evaporation treatment can be 35°C - 45°C. Thereby, the polystyrene microspheres can be deswollen, and then the fluorescent dye VIX-S is encapsulated inside the polymer microspheres.

[0065] The near-infrared second-region fluorescent microspheres can be obtained by centrifugation.

[0066] In some embodiments, the rotation speed of the centrifugation treatment can be 8000 rpm - 12000 rpm, and the time of the centrifugation treatment can be 13 min - 17 min.

[0067] In some embodiments, the detergent used for the washing treatment may include deionized water and ethanol.

[0068] Exemplarily, for the washing treatment, the near-infrared second-region fluorescent microspheres obtained by centrifugation can be washed with deionized water first, and then washed with ethanol.

[0069] In some embodiments, the temperature of the drying treatment can be 35°C - 45°C, and the time of the drying treatment can be 0.8 h - 1.2 h.

[0070] The embodiments of the present application provide an application of a near-infrared second-region fluorescent microsphere as an intestinal motility monitoring tool.

[0071] The near-infrared II region fluorescent microspheres prepared in the embodiments of the present application have excellent fluorescence brightness, are stable in vivo imaging reagents, can successfully illuminate the intestine, and provide accurate information on the intestinal peristalsis frequency for monitoring the intestinal peristalsis frequency. Thus, the near-infrared II region fluorescent microspheres can be used as an intestinal motility monitoring tool to study disease models related to intestinal motility.

[0072] In the embodiments of the present application, by preparing near-infrared II region fluorescent microspheres and using them as an intestinal motility monitoring tool, a new intestinal motility monitoring method is provided, which makes up for the deficiencies of existing monitoring technologies, and can be used to monitor parameters such as intestinal peristalsis, evaluate gastric emptying, and intestinal transit speed in real time, and has great application prospects.

[0073] Example

[0074] The following examples more specifically describe the content of the present application. These examples are only for illustrative purposes, because various modifications and changes within the scope of the content of the present application are obvious to those skilled in the art. Unless otherwise stated, all parts, percentages, and ratios reported in the following examples are based on mass, and all reagents used in the examples are commercially available or synthesized according to conventional methods and can be used directly without further treatment, and the instruments used in the examples are all commercially available.

[0075] Example 1

[0076] Preparation of Near-Infrared-II Fluorescent Microspheres

[0077] Mix 1 mL of an aqueous suspension of polystyrene microspheres with a mass concentration of 2.5% with 10 mL of a sodium dodecyl sulfate (SDS) solution with a mass concentration of 0.25%, and ultrasonicate for 5 min to make the mixture uniform to obtain a first dispersion, where the average particle size of the polystyrene microspheres is 100 nm. Mix a tetrahydrofuran solution of NIR-II fluorescent dye VIX-S with a concentration of 0.1 g / L with the first dispersion in a volume ratio of 1:5. Stir the above mixture at room temperature for 3 h, and then perform evaporation treatment to remove tetrahydrofuran to obtain a second dispersion. Centrifuge the second dispersion at a speed of 10000 rpm for 15 min. The microspheres obtained by centrifugation are washed 3 times with water and ethanol respectively, and then dried at 40 °C for 1 h to obtain near-infrared II region fluorescent microspheres.

[0078] The prepared near-infrared II region fluorescent microspheres are used in Examples 2, 3, 4, and 5.

[0079] Example 2

[0080] Morphology and Size Characterization of Near-Infrared-II Fluorescent Microspheres

[0081] Figure 1This is the scanning electron microscope image of the polystyrene microspheres in Example 1 of this application. Figure 2 This is the scanning electron microscope image of the near-infrared II fluorescent microspheres prepared in Example 1 of this application. From Figure 1 and Figure 2 it can be seen that the morphology of the near-infrared II fluorescent microspheres is regular circular, and the average particle size is about 100 nm. The near-infrared II fluorescent microspheres still maintain the same morphology and size as before staining.

[0082] Example 3

[0083] Fluorescence Intensity Characterization of Near-Infrared-II Fluorescent Microspheres in PBS Solution

[0084] Add 0.5 mg of near-infrared II fluorescent microspheres to a centrifuge tube, and then add 1 mL of PBS buffer to make the concentration of the near-infrared II fluorescent microspheres 0.1 mg / mL. Mix well by ultrasound, place it in a NIR-II fluorescence imager (In-vivo MasterSystem), select 808 nm laser as the excitation wavelength, a 1000 nm long-pass filter, and an exposure time of 33 ms to obtain the NIR-II fluorescence image of the near-infrared II fluorescent microspheres.

[0085] Figure 3 This is the NIR-II fluorescence image of the near-infrared II fluorescent microspheres prepared in Example 1 of this application in phosphate buffered saline (PBS) solution. From Figure 3 it can be seen that the fluorescence intensity of the near-infrared II fluorescent microspheres is 2500 a.u., that is, the near-infrared II fluorescent microspheres can emit strong fluorescence in an aqueous environment.

[0086] Example 4

[0087] Distribution Imaging of Near-Infrared-II Fluorescent Microspheres in the Intestines of Mice at Different Time Points

[0088] Wrap 0.5 mg of near-infrared II fluorescent microspheres in bread and let the mice take it orally. Anesthetize the mice with isoflurane and place them in a NIR-II fluorescence imager (In-vivo Master System) for imaging. Irradiate the whole body of the mice with an 808 nm laser, and use a 1000 nm long-pass filter and an exposure time of 33 ms.

[0089] Figure 4 This is the fluorescence imaging map of the distribution of the near-infrared II fluorescent microspheres prepared in Example 1 of this application in the intestines of healthy mice at different time points ( Figure 4 A), the selected fluorescent spots ( Figure 4 B left) and their movement trajectory map ( Figure 4 B right). As Figure 4 shown, it can be observed that the near-infrared II fluorescent microspheres can successfully light up the intestine ( Figure 4 A), and by tracking one of the fluorescent spots and its movement trajectory (Figure 4 B), the peristaltic frequency of the intestine and the average velocity of the speckle movement can be obtained. The peristaltic frequency in the intestine of healthy mice is 0.51 bps, and the average velocity of the speckle movement is 1.3 mm / s. Therefore, the near-infrared second-region fluorescent microspheres can be well applied to the research of intestinal motility.

[0090] Example 5

[0091] NIR-II Imaging of the Abdomen of Near-Infrared-II Fluorescent Microspheres at Different Time Points in Constipated Mice and Constipation Recovery Mouse Models II Imaging

[0092] (1) Mice were gavaged with loperamide hydrochloride solution (50 mg / kg × 1 mL) twice a day. A constipation model of mice was established. After 7 days, imaging was performed on them according to the same methods and steps as in Example 3.

[0093] (2) After the imaging in step (1) was completed, the constipated mice were given a two-week recovery period. After their defecation returned to normal, imaging was continued on them according to the same methods and steps as in Example 3.

[0094] Figure 5 The near-infrared second-region fluorescent microspheres prepared in Example 1 of this application in constipated mice ( Figure 5 A) and constipated recovered mice ( Figure 5 B), fluorescence imaging maps of intestinal distribution at different time points. Figure 6 The selected fluorescent speckles and their movement trajectories of the near-infrared second-region fluorescent microspheres prepared in Example 1 of this application in the constipated mouse model ( Figure 6 A), and the selected fluorescent speckles and their movement trajectories in the constipated recovered mouse model ( Figure 6 B).

[0095] As Figure 5 and Figure 6 shown, it can be clearly seen that 5 h after the mice took the near-infrared second-region fluorescent microspheres, there was still more fluorescence residue in the abdomen of the constipated mice than that of the constipated recovered mice ( Figure 5 ), indicating that the peristaltic frequency of the constipated mice is much lower than that of the constipated recovered mice. By tracking the movement trajectory of one of the fluorescent speckles, the peristaltic frequency of the constipated mice was calculated to be 0.16 bps, and the average velocity of the fluorescent speckle movement was 0.6 mm / s ( Figure 6 A); the peristaltic frequency of the constipated recovered mice was 0.49 bps, and the average velocity of the fluorescent speckle movement was 1.4 mm / s ( Figure 6 B). These results show that the near-infrared second-region fluorescent microspheres prepared in the examples of this application can be used in the research related to intestinal motility.

[0096] The above embodiments are only used to illustrate the technical solutions of the present application, rather than to limit them; although the present application has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that: they can still modify the technical solutions described in the foregoing embodiments, or perform equivalent replacements on some or all of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present application, and they should all be covered within the scope of the claims and the description of the present application. In particular, as long as there is no structural conflict, the technical features mentioned in each embodiment can be combined in any way. The present application is not limited to the specific embodiments disclosed in the text, but includes all technical solutions that fall within the scope of the claims.

Claims

1. A near-infrared second-region fluorescent microsphere, characterized in that, The near-infrared second-region fluorescent microspheres include polymer spheres formed by winding polystyrene chains, and a fluorescent dye VIX-S distributed among the voids of the polystyrene chains; The structural formula of the fluorescent dye VIX-S is as shown in formula (1):

2. The near-infrared second-region fluorescent microspheres according to claim 1, wherein The average particle size of the near-infrared second-region fluorescent microspheres is 80 nm - 120 nm.

3. A method for preparing the near-infrared second-region fluorescent microspheres according to claim 1 or 2, characterized in that, Including: Mixing an aqueous suspension containing polystyrene microspheres with an aqueous solution containing a surfactant to obtain a first dispersion; After mixing an organic solution containing the fluorescent dye VIX-S with the first dispersion, removing the organic solvent through evaporation treatment to obtain a second dispersion, and the structural formula of the fluorescent dye VIX-S is as shown in formula (1): Performing centrifugation treatment, washing treatment, and drying treatment on the second dispersion to obtain near-infrared second-region fluorescent microspheres.

4. The preparation method according to claim 3, wherein, The average particle size of the polystyrene microspheres is 80 nm - 120 nm.

5. The preparation method according to claim 3, characterized in that, The mass concentration of the polystyrene microspheres in the aqueous suspension containing polystyrene microspheres is 2% - 3%; and / or, The concentration of the fluorescent dye VIX-S in the organic solution containing the fluorescent dye VIX-S is 0.08 g / L - 0.12 g / L.

6. The preparation method according to claim 3, characterized in that, The volume ratio of the organic solution containing the fluorescent dye VIX-S to the first dispersion is 1:(4 - 6).

7. The preparation method according to claim 3, wherein The mass concentration of the surfactant in the aqueous solution containing the surfactant is 0.2% - 0.3%; the surfactant includes sodium dodecyl sulfate; and / or, The organic solvent in the organic solvent containing the fluorescent dye VIX-S includes one or more of tetrahydrofuran, chloroform, and acetone.

8. The preparation method according to claim 3, characterized in that, The preparation method further includes: after mixing the organic solution containing the fluorescent dye VIX-S with the first dispersion, performing ultrasonic treatment; The time of the ultrasonic treatment is 4 min - 6 min.

9. The preparation method according to claim 3, characterized in that, The rotation speed of the centrifugation treatment is 8000 rpm - 12000 rpm, and the time of the centrifugation treatment is 13 min - 17 min; and / or, The temperature of the drying treatment is 35 °C - 45 °C, and the time of the drying treatment is 0.8 h - 1.2 h.

10. Application of the near-infrared second-region fluorescent microspheres according to claim 1 or 2 as an intestinal motility monitoring tool.

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