Nanoparticles for contrast agents, preparation method and application thereof

Abstract

The present invention discloses nanoparticles for use as contrast agents, as well as their preparation methods and applications, and relates to the technical field of nanocomposite materials. The nanoparticles for use as contrast agents provided by the present invention have a shell comprising polydopamine and albumin linked by covalent bonds, and a core structure comprising a complex formed by metal ions and polyphenols. The metal ions in the core structure also coordinate with the polydopamine in the shell. Because the components are tightly connected, the rotation time of the nanoparticles is prolonged, thereby significantly improving the T1 relaxation rate of the nanoparticles. Simultaneously, the introduction of polydopamine prevents the release of metal ions in the human body, improving the biosafety of the nanoparticles. The albumin in the nanoparticles can also enhance the biocompatibility of the particles. Consequently, the nanoparticles provided by the present invention have broad application prospects in the fields of fluorescence imaging and targeted drug delivery.

Description

Technical Field

[0001] The present invention relates to the technical field of nanocomposite materials, and in particular to nanoparticles for contrast agents, a preparation method thereof, and applications thereof. Background Art

[0002] Magnetic resonance imaging (MRI) is a widely used detection technology in clinical practice. It has the advantages of being non-invasive, non-destructive, high-resolution, and capable of scanning at any angle. It uses radiofrequency pulses applied under a magnetic field to cause hydrogen protons in the human body to resonate and release energy through a relaxation process, thereby obtaining transverse and longitudinal imaging signals. However, the physiological indicators of early-stage microlesions are similar to those of normal tissue, and the relaxation time of hydrogen protons is also similar, making them difficult to effectively identify. To this end, the nuclear magnetic resonance signal can be improved by applying contrast agents to the imaging subject. Contrast agents in nuclear magnetic resonance are all paramagnetic substances. When an external magnetic field is applied to these paramagnetic substances, they can generate a locally enhanced magnetic field, accelerating the relaxation of neighboring hydrogen protons, thereby enhancing the nuclear magnetic resonance imaging signal that depends on the hydrogen proton relaxation rate.

[0003] According to different physicochemical properties, magnetic resonance imaging contrast agents can be divided into T1 contrast agents and T2 contrast agents. T2 contrast agents are mainly superparamagnetic iron oxide nanoparticles (SPIO), which can reduce the transverse relaxation time of hydrogen protons. However, they have problems such as being difficult to remove, prone to artifacts under high magnetic fields, and difficult to identify dark signals, which greatly increases the difficulty of clinical diagnosis. They have now been withdrawn from the market. T1 contrast agents mainly shorten the longitudinal relaxation time of hydrogen protons through the direct interaction between paramagnetic metals and hydrogen protons in the surrounding environment, thereby enhancing the imaging effect. Currently, the T1 contrast agents on the market are basically gadolinium-based contrast agents, including Gd-DTPA, Gd-EOB-DTPA, Gd-DO3A-Butriol, etc. However, the T1-weighted relaxation rate of these gadolinium-based contrast agents is not high, which makes the clinical dose as high as 15.7 mg Gd kg -1 , which leads to the retention of gadolinium in the human body and nephrogenic fibrosis, causing people to worry about the safety and reliability of contrast agents. The use of iron-based or manganese-based contrast agents to replace gadolinium-based contrast agents has become the mainstream trend. However, the existing non-gadolinium-based contrast agents also have the problem of insufficient relaxation rate. Most metal ion complex magnetic resonance contrast agents have a T1 relaxation efficiency of no more than 5mM. -1 s -1This means that in order to obtain higher sensitivity magnetic resonance imaging, it is still necessary to increase the dose of contrast agent. However, even for non-heavy metal contrast agents, excessive levels in the human body can cause damage to the human body. Using substances with good biosafety to stably bind and coat the metals in the contrast agent can effectively prevent the release and residue of metal ions in the human body and reduce the damage caused by metal ions to the human body. For example, the prior art Bsa-assisted synthesis of ultrasmallgallic acid-Fe(III)coordination polymer nanoparticles for cancer theranostics discloses a GA (gallic acid)-Fe@BSA (Bovine Serum Albumin) nanoparticle, which uses bovine serum albumin to improve the biosafety of the Fe-GA complex in the human body, but its relaxivity is only 0.89mM. -1 s -1 This means that to obtain sufficiently clear imaging results, more contrast agent must be used. Even if the possibility of contrast agent particles releasing metal ions in the human body is reduced, the high concentration of contrast agent used may still lead to excessively high concentrations of metal ions remaining in the human body. Therefore, to reduce the harm of magnetic resonance imaging contrast agents to the human body, two aspects should be considered: improving the relaxivity of nanoparticles and enhancing the biosafety of the particles. Summary of the Invention

[0004] To address the difficulty of existing metal ion complex magnetic resonance contrast agents in achieving both high relaxivity and high biosafety, the present invention provides nanoparticles for use as contrast agents. The particle shell comprises polydopamine and albumin, and the core structure comprises a complex formed by a metal ion and a polyphenol. The metal ion in the core structure also coordinates with the polydopamine in the shell. The components of the nanoparticles provided by the present invention are tightly connected, extending the nanoparticle's rotation time, thereby significantly improving the nanoparticle's T1 relaxivity. Furthermore, the introduction of polydopamine prevents the release of metal ions in the human body, thereby improving the nanoparticle's biosafety.

[0005] Another object of the present invention is to provide a method for preparing the above-mentioned nanoparticles for contrast agent.

[0006] Another object of the present invention is to provide a contrast agent comprising the aforementioned nanoparticles for contrast agents.

[0007] Another object of the present invention is to provide a nano drug-loaded particle comprising the above-mentioned nanoparticles for contrast agent.

[0008] Another object of the present invention is to provide particles for optical imaging comprising the aforementioned nanoparticles for contrast agents.

[0009] The above-mentioned purpose of the present invention is achieved through the following technical solutions:

[0010] A nanoparticle for a contrast agent, comprising a core and a shell, wherein the shell comprises polydopamine and albumin, and the core comprises a complex of a metal ion and a polyphenol, wherein the metal ion in the complex is connected to the polydopamine via a coordination bond;

[0011] The molar ratio of the polyphenol in the core to the polydopamine in the shell is (0.2-10):1;

[0012] The mass ratio of polydopamine to albumin in the shell layer is (0.2-5):1.

[0013] In a specific embodiment of the present invention, the metal ion in the complex can be at least one of a ferric ion, a ferrous ion, or a manganese ion. Currently, the three metal elements commonly used in the field of contrast agents are iron, manganese, and gadolinium. However, contrast agents prepared using iron or manganese have higher biosafety, and therefore using iron or manganese to construct contrast agents has become a mainstream trend.

[0014] In nanoparticle provided by the present invention, kernel is the complex compound that metal ion and polyphenol substance form, and wherein metal ion provides paramagnetism, makes nanoparticle can produce the magnetic field of local enhancement under external action, strengthens the magnetic resonance signal that adjacent hydrogen proton relaxation forms.Yet the design of magnetic resonance contrast agent also needs to consider the consumption of metal except considering the paramagnetism of metal, the exchange rate of contrast agent and water molecule, the rotational correlation speed of contrast agent molecule, the toxicity of contrast agent and metabolic efficiency and other factors, therefore in nanoparticle provided by the present invention, core structure does not simply comprise metal ion, but comprises the complex compound of metal ion and polyphenol, wherein polyphenol can also improve the loading efficiency of metal ion, therefore improve the effective content of metal ion in nanoparticle, and then improve the relaxivity of nanoparticle.In addition, metal ion easily causes Fenton reaction in human body and causes to produce too many reactive oxygen free radicals (ROS) in human body, and polyphenol has good antioxidant capacity, therefore can suppress the Fenton reaction that metal ion causes, improve the biosafety of nanoparticle.

[0015] The nanoparticles provided by the present invention have a shell comprising polydopamine and albumin, wherein the dopamine units adjacent to metal ions in the polydopamine can also form coordination bonds with the metal ions. The albumin in the shell has an interface modification effect. The present invention introduces albumin into the nanoparticles mainly to improve the biocompatibility of the particles. The introduction of polydopamine can increase the total molecular weight of the nanosystem, prolong the rotational correlation time of the particles, and thereby improve their T1 relaxation rate. Polydopamine can also improve the structural stability of the nanoparticles, preventing them from releasing metal ions in the human body and causing harm.

[0016] The reason for controlling the ratio of polyphenols in the core to polydopamine in the shell is to ensure that nanoparticles can form. If too little shell component is added, the shell will not be able to wrap around the core, resulting in the inability to form nanoparticles. When used for imaging, the metal ions in the core will be easily released and remain in the body. Controlling the ratio of polydopamine to albumin in the shell can make the shell more stable.

[0017] In a specific embodiment of the present invention, the polydopamine in the shell is obtained by polymerization of dopamine monomers during the preparation of the nanoparticles. Therefore, the ratio of polydopamine to other substances is calculated based on the amount of dopamine monomer added, assuming that the dopamine monomers are completely polymerized.

[0018] Preferably, the polydopamine and albumin in the shell are linked via a covalent bond.

[0019] The polydopamine and albumin in the shell are linked by covalent bonds, which can ensure that the components in the nanoparticles of the present invention are closely linked and the structure of the nanoparticles is stable.

[0020] Preferably, the molar ratio of polyphenols to metal ions in the core is (0.5-2):1.

[0021] The purpose of controlling the above molar ratio is to fully complex and coordinate the metal ions with the polyphenols.

[0022] Preferably, the molar ratio of the polyphenols in the core to the polydopamine in the shell is (0.5-3):1.

[0023] More preferably, the molar ratio of the polyphenols in the core to the polydopamine in the shell is 1:1.

[0024] By controlling the molar ratio of polyphenol to polydopamine within the range of (0.5-3):1, and especially controlling the molar ratio to 1:1, nanoparticles with higher MRI-T1 relaxivity can be obtained.

[0025] Preferably, the molar ratio of polydopamine to albumin in the shell layer is (1-2):1.

[0026] More preferably, the molar ratio of polydopamine to albumin in the shell layer is 1.5:1.

[0027] During the exploration process, it was found that when the molar ratio of polydopamine to albumin was (1-2):1, especially 1.5:1, the MRI-T1 relaxation rate of the nanoparticles was higher.

[0028] In a specific embodiment of the present invention, the T1 relaxation rate of the nanoparticles used as contrast agents is 15 to 27 mM. -1 s -1 .

[0029] The nanoparticles used as contrast agents provided by the present invention have a stable structure and a large total molecular weight, so the rotational correlation time is longer and the T1 relaxation rate is as high as 20mM -1 s -1 The above results are far superior to those reported in existing literature.

[0030] In a specific embodiment of the present invention, the polyphenol is any one or more of tannic acid, catechin, epicatechin, quercetin, and gallic acid.

[0031] In a specific embodiment of the present invention, the albumin is any one or more of human serum albumin, bovine serum albumin, ovalbumin, and chicken serum albumin.

[0032] The present invention also protects a method for preparing the above-mentioned nanoparticles for contrast agent, comprising the following steps:

[0033] S1. dissolving a polyphenol and a metal salt in a solvent to obtain a solution of a metal ion-polyphenol complex;

[0034] S2. The metal ion-polyphenol complex, dopamine monomer and albumin are reacted under alkaline conditions to obtain nanoparticles for contrast agent.

[0035] In a specific embodiment of the present invention, the solvent in step S1 is water.

[0036] In a specific embodiment of the present invention, the metal salt in step S1 can be at least one of manganese chloride, ferric chloride, and ferrous chloride.

[0037] In a specific embodiment of the present invention, the concentration of polyphenols in the metal ion-polyphenol complex solution in step S1 is 2.5-5 mg / ml, and the concentration of metal ions is 1-2 mg / ml.

[0038] In a specific embodiment of the present invention, the alkaline condition in step S2 is a pH of 7.4-10.

[0039] The reason for making the substances react under alkaline conditions is to enable the dopamine monomer to undergo a polymerization reaction and form polydopamine outside the metal ion-polyphenol complex.

[0040] In a specific embodiment of the present invention, the reaction in step S2 is carried out in a solution environment.

[0041] Preferably, step S2 is carried out in a multi-channel reactor.

[0042] In a specific embodiment of the present invention, the multichannel reactor in step S2 is a multichannel vortex static mixer (rapid nanoprecipitation mixer).

[0043] In a specific embodiment of the present invention, the reaction is carried out in a multi-channel reactor as follows: the metal ion-polyphenol complex solution, dopamine monomer solution and albumin solution obtained in step S1 are respectively introduced into the multi-channel reactor and reacted under alkaline conditions to obtain nanoparticles for contrast agent.

[0044] The above preparation method belongs to the flow control micro-area synthesis technology, which can enable the solutions of each component to undergo high-speed coordination self-assembly with the same surface force, completely solving the defect of uneven particle size prepared by traditional self-assembly preparation methods. It has the advantages of fast self-assembly efficiency, continuous large-scale preparation, and uniform and controllable particle size. At the same time, according to actual needs, the raw materials and ratios of self-assembly can be replaced, and the flow rate parameters of the equipment can be changed to obtain nanoparticles with different functions, different morphologies, and different particle sizes, thereby diversifying products and diversifying application fields.

[0045] In a specific embodiment of the present invention, in step S2, each solution is introduced into the multichannel reactor at a flow rate of (1-40) ml / min.

[0046] In a specific embodiment of the present invention, the alkaline condition in step S2 is provided by an albumin solution, that is, the albumin solution is alkaline.

[0047] In a specific embodiment of the present invention, the dopamine monomer solution in step S2 is a solution obtained by dissolving dopamine hydrochloride in water.

[0048] In a specific embodiment of the present invention, the concentration of dopamine monomer in the dopamine monomer solution in step S2 is (0.5-4) mg / ml.

[0049] In a specific embodiment of the present invention, the albumin solution in step S2 is a solution obtained by dissolving albumin in water.

[0050] In a specific embodiment of the present invention, the concentration of albumin in the albumin solution in step S2 is 0.25-5 mg / ml.

[0051] In a specific embodiment of the present invention, the flow rates of the polyphenol-metal ion solution, the dopamine monomer solution and the albumin solution in step S2 are consistent.

[0052] In a specific embodiment of the present invention, the reaction time in step S2 is less than 1 minute. More specifically, the reaction time in step S2 is 0.5 to 1 minute. The target nanoparticles can be directly obtained by separately introducing the metal ion-polyphenol complex solution, the dopamine monomer solution, and the albumin solution into a multichannel reactor without the need to specifically control the reaction time.

[0053] In a specific embodiment of the present invention, step S2 further includes a dialysis step after the reaction is completed.

[0054] In a specific embodiment of the present invention, the dialysis is performed at room temperature for 24 to 78 hours.

[0055] The present invention also protects a contrast agent, comprising the above-mentioned nanoparticles for contrast agent.

[0056] The nanoparticles provided by the present invention have a high relaxivity and can improve the quality of magnetic resonance imaging at a low dose when used to prepare magnetic resonance contrast agents.

[0057] The present invention also protects a nano drug-loaded particle, comprising the above-mentioned nano particle for contrast agent and a targeted drug loaded in the shell of the nano particle.

[0058] The present invention also protects a particle for optical imaging, comprising the above-mentioned nanoparticles for contrast agent and a fluorescent marker loaded in the shell of the nanoparticles.

[0059] The nanoparticles provided by the present invention have high relaxivity while also exhibiting excellent biosafety and compatibility. The albumin in the particle structure possesses a unique spatial structure that can serve as a delivery vehicle for drugs and / or fluorescent markers. Therefore, in addition to being used to prepare magnetic resonance contrast agents, the nanoparticles can also be used as a carrier to load and transport drugs and / or fluorescent markers, thereby achieving synergistic performance between magnetic resonance imaging, drug therapy, and fluorescence imaging. The electronegativity of albumin can also effectively extend the circulation time of the delivery system. When delivering drugs, the targeting of the delivery system should be provided by the drug itself.

[0060] In a specific embodiment of the present invention, when contrast agent nanoparticles are combined with targeted drugs to form drug-loaded nanoparticles, the preparation method of the drug-loaded nanoparticles includes the following steps:

[0061] S1. dissolving polyphenol and metal salt in a solvent to obtain a metal ion-polyphenol complex solution;

[0062] S2. The metal ion-polyphenol complex, dopamine monomer, albumin and the targeted drug are reacted under alkaline conditions to obtain drug-loaded nanoparticles.

[0063] In a specific embodiment of the present invention, the method for preparing the drug-loaded nanoparticles is consistent with the method for preparing the nanoparticles for contrast agents except for adding the targeted drug in step S2.

[0064] In a specific embodiment of the present invention, the targeted drug may be at least one of anti-VEGF antibody, paclitaxel or doxorubicin hydrochloride.

[0065] In a specific embodiment of the present invention, step S2 involves introducing the metal ion-polyphenol complex solution, the dopamine monomer solution, the albumin solution, and the targeted drug solution into a multichannel reactor for reaction. More specifically, the targeted drug solution is a solution obtained by dissolving the targeted drug in water, wherein the concentration of the targeted drug is 0.25 to 2 mg / ml.

[0066] In a specific embodiment of the present invention, when contrast agent nanoparticles are combined with fluorescent markers to form optical imaging particles, the method for preparing the optical imaging particles includes the following steps:

[0067] S1. dissolving polyphenol and metal salt in a solvent to obtain a metal ion-polyphenol complex solution;

[0068] S2. The metal ion-polyphenol complex, dopamine monomer, albumin and fluorescent marker are reacted under alkaline conditions to obtain particles for optical imaging.

[0069] In a specific embodiment of the present invention, the method for preparing the above-mentioned particles for optical imaging, except for adding a fluorescent marker in step S2, the remaining steps are consistent with the method for preparing nanoparticles for contrast agents.

[0070] In a specific embodiment of the present invention, the fluorescent marker may be IR780 iodide and / or Nile red.

[0071] In a specific embodiment of the present invention, step S2 involves introducing the metal ion-polyphenol complex solution, the dopamine monomer solution, the albumin solution, and the fluorescent marker solution into a multichannel reactor for reaction. More specifically, the fluorescent marker solution is a solution obtained by dissolving the fluorescent marker in water, wherein the concentration of the fluorescent marker is 5 to 100 μg / ml.

[0072] Compared with the prior art, the present invention has the following beneficial effects:

[0073] The nanoparticles for contrast agents provided by the present invention have a greatly improved T1 relaxation rate of 12 mM due to the close connection of the components, which prolongs the rotation time of the nanoparticles. -1 s -1 As mentioned above, the polydopamine coating can ensure that the metal ions inside the particles are not easily released, and the nanoparticles have excellent biosafety. Therefore, the nanoparticles provided by the present invention have broad application prospects in the field of magnetic resonance imaging. BRIEF DESCRIPTION OF THE DRAWINGS

[0074] Figure 1 Schematic diagram of the preparation process of nanoparticles used as contrast agents in the present invention.

[0075] Figure 2 This is a TEM image of the contrast agent nanoparticles Mn-GA@BSA@DA obtained in Example 1 of the present invention.

[0076] Figure 3 This is a comparison chart of the particle sizes of the contrast agent nanoparticles Mn-GA@BSA@DA obtained in Example 1 of the present invention before and after freeze-drying.

[0077] Figure 4 The particle size distribution diagrams of Fe-GA@BSA@DA obtained in Example 2, Mn-Quercetin@BSA@DA obtained in Example 3, Mn-TA@BSA@DA obtained in Example 4, Mn-GA@OVA@DA obtained in Example 5, and Mn-GA@HSA@DA obtained in Example 6 are shown.

[0078] Figure 5 This is a comparison chart of the relaxation rates of Mn-GA@BSA@DA in Example 1 of the present invention and Mn@BSA@DA in Comparative Example 1.

[0079] Figure 6 This is a comparison chart of the particle size stability of Mn-GA@BSA in comparative example 2 of the present invention and Mn-GA@BSA@DA in example 1 in aqueous solution ( Figure 6 A), and the particle size stability comparison chart in 1640 culture medium ( Figure 6 B).

[0080] Figure 7 The TMB color development and absorption values ​​at 652 nm of Mn-GA@BSA in Comparative Example 2 of the present invention and Mn-GA@BSA@DA in Example 1 under different H2O2 concentrations are shown.

[0081] Figure 8 These are near-infrared in vivo imaging images of Mn-GA@BSA@DA@IR780 at different time points (1h, 24h, 48h) in Experimental Example 1 of the present invention.

[0082] Figure 9 Mn-GA@BSA@DA@NileRed( Figure 9 A) and Mn-GA@BSA@DA@paclitaxel obtained in Experimental Example 3 ( Figure 9 B) Particle size distribution diagram. DETAILED DESCRIPTION

[0083] The present invention will be further described below in conjunction with specific embodiments, but the examples do not limit the present invention in any form. Unless otherwise specified, the raw materials and reagents used in the examples of the present invention are conventionally purchased raw materials and reagents.

[0084] Example 1

[0085] A contrast agent nanoparticle, represented by Mn-GA@BSA@DA, has a shell composed of polydopamine (DA) and bovine serum albumin (BSA) linked by covalent bonds, and a core composed of a manganese ion (Mn)-gallic acid (GA) complex. The manganese ion in the complex is linked to the polydopamine in the shell by a coordination bond.

[0086] The molar ratio of gallic acid to manganese ion in the manganese ion-gallic acid complex is 1.16:1;

[0087] The molar ratio of gallic acid in the manganese ion-gallic acid complex to polydopamine in the shell is 1:1;

[0088] The mass ratio of polydopamine to albumin in the shell is 1.5:1.

[0089] The method for preparing nanoparticles for contrast agents in this embodiment includes the following steps:

[0090] S1. The pyrogallic acid and manganese chloride were dissolved in water to obtain a manganese ion - gallic acid complex solution, wherein the concentration of pyrogallic acid was 2.5 mg / ml, the concentration of manganese chloride was 1 mg / ml;

[0091] S2. The manganese ion-gallic acid complex solution obtained in step S1, the dopamine hydrochloride solution, the bovine serum albumin solution, and pure water were introduced into a rapid nanoprecipitation mixer at a flow rate of 5 ml / min for reaction. The volume ratio of the four liquids introduced into the rapid nanoprecipitation mixer was 1:1:1:1. After the reaction was completed, the nanoparticles for contrast agent were obtained by dialyzing at room temperature for 24 hours.

[0092] In step S2, the dopamine hydrochloride solution is a solution obtained by dissolving dopamine hydrochloride in water, with a concentration of 3.75 mg / ml, and the bovine serum albumin solution is a solution obtained by dissolving bovine serum albumin in water, with a concentration of 2.85 mg / ml and a pH of 8.5.

[0093] Example 2

[0094] A contrast agent nanoparticle, represented by Fe-GA@BSA@DA, differs from that in Example 1 only in that:

[0095] The metal ion in the core structure is ferrous ion.

[0096] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0097] In step S1, manganese chloride is replaced with ferrous chloride.

[0098] Example 3

[0099] A contrast agent nanoparticle, represented by Mn-Quercetin@BSA@DA, differs from that in Example 1 only in that:

[0100] The polyphenol in the core structure is quercetin.

[0101] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0102] Step S1 replaces pyrogallic acid with quercetin.

[0103] Example 4

[0104] A contrast agent nanoparticle, represented by Mn-TA@BSA@DA, differs from that in Example 1 only in that:

[0105] The polyphenolic substance in the core structure is tannic acid TA.

[0106] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0107] Step S1 replaces pyrogallic acid with tannic acid.

[0108] Example 5

[0109] A contrast agent nanoparticle, represented by Mn-GA@OVA@DA, differs from that in Example 1 only in that:

[0110] The albumin in the shell is ovalbumin OVA.

[0111] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0112] In step S2, the bovine serum albumin solution is replaced with an ovalbumin solution. The ovalbumin solution is a solution obtained by dissolving ovalbumin in water, and its concentration and pH are consistent with those of the bovine serum albumin solution.

[0113] Example 6

[0114] A contrast agent nanoparticle, represented by Mn-GA@HSA@DA, differs from that in Example 1 only in that:

[0115] The albumin in the shell is human serum protein HSA.

[0116] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0117] In step S2, the bovine serum albumin solution is replaced with a human serum albumin solution. The human serum albumin solution is a solution obtained by dissolving human serum albumin in water, and the concentration and pH are consistent with those of the bovine serum albumin solution.

[0118] Example 7

[0119] A contrast agent nanoparticle, represented by Mn-GA@BSA@DA-2, differs from that in Example 1 only in that:

[0120] The molar ratio of gallic acid in the manganese ion-gallic acid complex to polydopamine in the shell is 3:1.

[0121] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0122] The concentration of the dopamine hydrochloride solution in step S2 is 1.27 mg / ml.

[0123] Example 8

[0124] A contrast agent nanoparticle, represented by Mn-GA@BSA@DA-3, differs from that in Example 1 only in that:

[0125] The molar ratio of gallic acid in the manganese ion-gallic acid complex to polydopamine in the shell is 0.5:1.

[0126] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0127] The concentration of the dopamine hydrochloride solution in step S2 is 7.5 mg / ml.

[0128] Example 9

[0129] A contrast agent nanoparticle, represented by Mn-GA@BSA@DA-4, differs from that in Example 1 only in that:

[0130] The molar ratio of gallic acid in the manganese ion-gallic acid complex to polydopamine in the shell is 1:3.

[0131] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0132] The concentration of the dopamine hydrochloride solution in step S2 is 11.2 mg / ml.

[0133] Example 10

[0134] A contrast agent nanoparticle, represented by Mn-GA@BSA@DA-5, differs from that in Example 1 only in that:

[0135] The mass ratio of polydopamine to albumin in the shell is 0.5:1.

[0136] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0137] The concentration of the dopamine hydrochloride solution in step S2 is 1.4 mg / ml.

[0138] Example 11

[0139] A contrast agent nanoparticle, represented by Mn-GA@BSA@DA-6, differs from that in Example 1 only in that:

[0140] The mass ratio of polydopamine to albumin in the shell is 1:1.

[0141] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0142] The concentration of the dopamine hydrochloride solution in step S2 is 2.85 mg / ml.

[0143] Example 12

[0144] A contrast agent nanoparticle, represented by Mn-GA@BSA@DA-7, differs from that in Example 1 only in that:

[0145] The mass ratio of polydopamine to albumin in the shell is 2:1.

[0146] The method for preparing nanoparticles for contrast agents in this embodiment differs from that in Example 1 only in that:

[0147] The concentration of the dopamine hydrochloride solution in step S2 is 5.69 mg / ml.

[0148] Comparative Example 1

[0149] A core-shell structured nanoparticle, represented by Mn@BSA@DA, differs from that in Example 1 only in that:

[0150] The core structure contains only metal ions Mn 2+ .

[0151] The method for preparing core-shell structured nanoparticles in this embodiment differs from that in Example 1 only in that:

[0152] No pyrogallic acid is added in step S1.

[0153] Comparative Example 2

[0154] A core-shell structured nanoparticle, represented by Mn-GA@BSA, differs from that in Example 1 only in that:

[0155] The shell does not contain polydopamine.

[0156] The method for preparing core-shell structured nanoparticles in this embodiment differs from that in Example 1 only in that:

[0157] In step S2, the dopamine hydrochloride solution is replaced with water.

[0158] Performance Testing

[0159] 1. Basic performance test of particles

[0160] Particle size test: The nanoparticles obtained in the examples and comparative examples were tested using a nanoparticle size-potential detector (NanoBrook, Malvern, America).

[0161] Potential test: The nanoparticles obtained in the examples and comparative examples were tested using a nanoparticle size-potential detector (NanoBrook, Malvern, America).

[0162] T1 relaxation rate test: The nanoparticles obtained in the embodiment and the comparative example were subjected to magnetic resonance imaging using a 1.5T magnetic resonance imaging system to measure the T1 relaxation rate.

[0163] In addition, TEM images of the nanoparticles obtained in the examples and comparative examples were obtained by observation using a TEM electron microscope (Tecnai G2 Spirit T120 kV, Thermo, America).

[0164] The specific performance test data are shown in Table 1. Figures 1 to 7 As shown:

[0165] Table 1. Particle size, potential and relaxation rate data of the particles obtained in the examples and comparative examples

[0166] Group Particle size nm Potential mV <![CDATA[T1 relaxation rate mM -1 s -1 > Example 1 18.43±2.69 -26.93±2.04 21.144 Example 2 15.21±4.73 -29.93±2.87 15.6 Example 3 20.69±3.67 -21.27±3.07 12.6 Example 4 27.23±7.65 -13.06±3.02 16.1 Example 5 25.71±1.059 -42.44±1.69 18.5 Example 6 33.42±7.77 -22.59±7.96 - Example 7 17.28±2.49 -13.32±4.34 16.82 Example 8 11.25±2.39 -19.06±2.39 16.11 Example 9 8.39±0.62 -13.32±4.34 15.74 Example 10 11.61±0.80 -19.09±1.60 12.51 Example 11 19.44±1.79 -12.42±4.13 14.133 Example 12 31.65±1.59 -21.53±2.48 14.67 Comparative Example 1 35.73±1.01 -12.05±4.37 4.082 Comparative Example 2 23.47±1.059 -22.59±2.03 20.57

[0167] Note: The data in Table 1 above with “-” means that no test was performed.

[0168] As shown in Table 1 above, the nanoparticles provided by the present invention have a -1 s -1 The above T1 relaxation rates are much better than those reported in most existing literatures, and are also much better than the nanoparticles without polyphenols in the core (Comparative Example 1). The T1 relaxation rate of Mn-GA@BSA@DA in Example 1 (21.144 [Mn] mM -1 s -1 ) is better than the Mn@BSA@DA particles in Comparative Example 1 (4.082[Mn]mM -1 s -1 ) is 5.18 times higher, indicating that the presence of gallic acid can improve the manganese loading efficiency and prolong the rotation efficiency of the particles, thereby greatly improving the T1 relaxation rate. At the same time, the absolute value of the Zeta potential of the nanoparticles provided by the present invention is also higher than 12mV, indicating that the system is relatively stable and it is not easy for particles to agglomerate. Analysis of the data of Examples 1, 7 to 9 in Table 1 shows that when the molar ratio of the polyphenol substance in the nanoparticle core to the polydopamine in the shell is the preferred (0.5-3) of the present invention: 1 (Examples 1, 7 to 8), the resulting particles have a more excellent T1 relaxation rate. Comprehensive analysis of the data of Examples 1, 10 to 12 in Table 1 shows that when the molar ratio of polydopamine to albumin in the nanoparticle shell is the preferred (1 to 2) of the present invention: 1 (Examples 1, 11 to 12), the resulting nanoparticles also have a more excellent T1 relaxation rate. According to Table 1 above, when the nanoparticle shell does not contain polydopamine (Comparative Example 2), the resulting nanoparticles also have a higher relaxivity and absolute value of Zeta potential. However, subsequent research shows that the biosafety and stability of the nanoparticles in Comparative Example 2 are insufficient:

[0169] In order to investigate the stability of the nanoparticles obtained in Example 1 and Comparative Example 2, the nanoparticles obtained in Example 1 were freeze-dried in a vacuum freeze dryer to form a powder, and the particle size before and after freeze-drying was measured using a nanoparticle size detector. At the same time, the nanoparticles obtained in Example 1 and Comparative Example 2 were dissolved in water and 1640 culture medium, respectively, and the particle size changes over time were measured. The experimental results are shown in FIG. Figure 3 and Figure 6 As shown:

[0170] according to Figure 3The particle size of the nanoparticles in Example 1 did not change much before and after freeze-drying, indicating that the particles can be stored at low temperatures for a long time. At the same time, the advantage of high hydrophilicity enables rapid resolubility, indicating that the particles have good stability and have the prospect of being used to prepare drug-loaded particles and particles for fluorescence imaging.

[0171] Figure 6 A is a comparison chart of the particle size stability of Mn-GA@BSA in comparative example 2 of the present invention and Mn-GA@BSA@DA in example 1 in aqueous solution, Figure 6 B is a comparison chart of the particle size stability of the above two particles in 1640 culture medium. Figure 6 It can be seen that polydopamine can strengthen the stability of particles, thereby increasing the biosafety of particles.

[0172] In order to explore the biosafety of the nanoparticles obtained in Example 1 and Comparative Example 2, the following test was also conducted: the nanoparticles obtained in Example 1 and Comparative Example 2 were mixed with H2O2 at different concentrations, and TMB (3,3',5,5'-tetramethylbenzidine) was used as a chromogenic substrate to detect whether a Fenton reaction occurred in the reaction system. If the reaction system turns blue, it indicates that a Fenton reaction has occurred in the system. The experimental results are shown in FIG. Figure 7 As shown:

[0173] Figure 7 The TMB color development and absorption values ​​at 652 nm of Mn-GA@BSA in Comparative Example 2 and Mn-GA@BSA@DA in Example 1 at different H2O2 concentrations are shown. Figure 7 As shown in the figure, the catalytic behavior of Mn-GA@BSA and Mn-GA@BSA@DA particles towards different H2O2 concentrations was tested. It was found that the positive control group MnCl2 showed a distinct blue color, while the nanoparticles hardly changed color. This indicates that after the phenolic hydroxyl groups on the polyphenols were coordinated with the metal ions, the hydroxyl radicals and H2O2 could not obtain electrons from the polyphenol-metal complex, thereby inhibiting the Fenton reaction, further proving the high biosafety of the particles.

[0174] 2. Nanoparticle Delivery System Performance Testing

[0175] Next, by adjusting the preparation method, the contrast agent nanoparticles prepared in Example 1 are loaded with fluorescent markers or targeted drugs to explore the possibility of using the contrast agent nanoparticles in this application as a nanoparticle delivery system.

[0176] Experimental Example 1

[0177] An optical imaging particle, represented by Mn-GA@BSA@DA@IR780, differs from that in Example 1 only in that:

[0178] The fluorescent marker IR780 is also included in the shell.

[0179] The method for preparing optical imaging particles in this embodiment differs from that in Example 1 only in that:

[0180] In step S2, pure water is replaced with IR780 iodide solution, which is a solution obtained by dissolving IR780 iodide in DMSO, wherein the concentration of IR780 is 50 μg / ml.

[0181] The nanoparticles were injected into mice to obtain fluorescence imaging results at different times, as shown in Figure 2. Figure 8 As shown. Figure 8 It can be seen that Mn-GA@BSA@DA@IR780 particles can achieve tumor targeting. At 24 hours, obvious tumor highlights can be seen in the Mn-GA@BSA@DA@IR780 group (circled arrow part). In contrast, the fluorescence imaging results of the experimental group injected with IR780 only were not obvious.

[0182] Experimental Example 2

[0183] An optical imaging particle, represented by Mn-GA@BSA@DA@NileRed, differs from that in Example 1 only in that:

[0184] The shell also contains the fluorescent marker Nile Red.

[0185] The nanoparticles were characterized to be spherical with an average particle size of 25.62±4.72nm and a particle size distribution as shown in Figure 9 As shown in A.

[0186] The method for preparing optical imaging particles in this embodiment differs from that in Example 1 only in that:

[0187] In step S2, pure water is replaced with a Nile red solution, which is a solution obtained by dissolving Nile red in DMSO, wherein the concentration of Nile red is 50 μg / ml.

[0188] Experimental Example 3

[0189] A nano drug-loaded particle, represented by Mn-GA@BSA@DA@paclitaxel, differs from that in Example 1 only in that:

[0190] The shell also includes the targeted drug paclitaxel.

[0191] The nanoparticles were characterized to be spherical with an average particle size of 34.64±7.63nm and a particle size distribution as shown in Figure 9 As shown in B.

[0192] The preparation method of drug-loaded nanoparticles in this embodiment differs from that in Example 1 only in that:

[0193] In step S2, pure water is replaced with a paclitaxel solution, which is a solution obtained by dissolving paclitaxel in DMSO, wherein the concentration of paclitaxel is 100 ug / ml.

[0194] According to the above experimental examples, the nanoparticles provided by the present invention can still maintain the particle structure after loading fluorescent markers and targeted drugs, and can therefore be applied to the fields of fluorescence imaging and nano-drug delivery, and can also improve the targeting of fluorescent markers and drugs during application.

[0195] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A nanoparticle for a contrast agent, characterized in that: The invention comprises a core and a shell, wherein the shell comprises polydopamine and albumin, and the core comprises a complex of metal ions and polyphenols, wherein the metal ions in the complex are connected to the polydopamine via a coordination bond; The molar ratio of the polyphenol in the core to the polydopamine in the shell is (0.2-10):1; The molar ratio of polyphenols to metal ions in the core is (0.5-2):1; The polyphenols include at least one of tannic acid, catechin, epicatechin, quercetin, and gallic acid; The mass ratio of polydopamine to albumin in the shell layer is (0.2-5):1; The metal ion is provided by one of manganese chloride, ferric chloride and ferrous chloride.

2. The nanoparticles for contrast agents according to claim 1, wherein The polydopamine in the shell is connected to the albumin via a covalent bond.

3. The nanoparticles for contrast agents according to claim 1, wherein The molar ratio of the polyphenols in the core to the polydopamine in the shell is (0.5-3):

1.

4. The nanoparticles for contrast agents according to claim 1, wherein The mass ratio of polydopamine to albumin in the shell layer is (1-2):

1.

5. The method for preparing nanoparticles for contrast agents according to any one of claims 1 to 4, characterized in that: The steps include: S1. dissolving a polyphenol and a metal salt in a solvent to obtain a solution of a metal ion-polyphenol complex; S2. The metal ion-polyphenol complex, dopamine monomer and albumin are reacted under alkaline conditions to obtain nanoparticles for contrast agent.

6. A contrast agent, characterized in that The nanoparticles for contrast agents according to any one of claims 1 to 4 are included.

7. A drug-loaded nanoparticle, characterized in that: The invention comprises the nanoparticles for contrast agents according to any one of claims 1 to 4 and a targeting drug loaded in the shell of the nanoparticles.

8. A particle for optical imaging, characterized in that: The invention comprises the nanoparticles for contrast agents according to any one of claims 1 to 4 and a fluorescent marker loaded in the shell of the nanoparticles.

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