Nanoparticles for contrast agent as well as preparation method and application of nanoparticles

Abstract

The invention discloses nanoparticles for a contrast agent as well as a preparation method and application of the nanoparticles, and relates to the technical field of nano composite materials. According to the nanoparticle for the contrast agent, a shell layer comprises polydopamine and albumin which are connected through a covalent bond, a core structure comprises a complex formed by metal ions and polyphenol, and the metal ions in the core structure and the polydopamine in the shell layer also have a coordination effect. Due to the fact that all the components are tightly connected, the rotation time of the nano-particles is prolonged, the T1 relaxation rate of the nano-particles is greatly increased, meanwhile, due to introduction of polydopamine, release of metal ions in a human body is avoided, the biological safety of the nano-particles is improved, albumin in the nano-particles can further improve the biocompatibility of the particles, and the nano-particles can be used for preparing the nano-particles. Therefore, the nanoparticles provided by the invention also have wide 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, and a preparation method and application 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 radio frequency 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 micro-lesion tissues are similar to those of normal tissues, and the relaxation time of hydrogen protons is also similar, making it difficult to effectively identify them. For this reason, the nuclear magnetic resonance signal can be improved by applying contrast agents to the imaging object. Contrast agents in nuclear magnetic resonance are all paramagnetic substances. After being applied with an external magnetic field, such paramagnetic substances can generate a locally enhanced magnetic field, which accelerates the relaxation of neighboring hydrogen protons, thereby enhancing the nuclear magnetic resonance imaging signal that depends on the relaxation rate of hydrogen protons.

[0003] According to different physicochemical properties, magnetic resonance 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 are difficult to remove, easy to produce artifacts under high magnetic fields, and difficult to identify dark signals, which greatly increases the difficulty of clinical diagnosis. They have now 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 magnetic resonance imaging with higher sensitivity, 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 encapsulate 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 relaxation efficiency is only 0.89mM. -1 s -1 This means that in order to obtain sufficiently clear imaging results, more contrast agents need to be used. Even if the possibility of contrast agent particles releasing metal ions in the human body is reduced, the high concentration of contrast agents used may cause the concentration of metal ions remaining in the human body to be too high. Therefore, in order to reduce the harm of magnetic resonance contrast agents to the human body, two aspects should be considered: improving the relaxation efficiency of nanoparticles and improving the biosafety of particles. Summary of the invention

[0004] In order to solve the problem that the existing metal ion complex magnetic resonance contrast agent is difficult to take into account both high relaxation efficiency and high biosafety, the present invention provides a nanoparticle for a contrast agent, wherein the particle shell includes polydopamine and albumin, the core structure includes a complex formed by metal ions and polyphenols, and the metal ions in the core structure and the polydopamine in the shell also have a coordination effect. The components in the nanoparticles provided by the present invention are closely connected, the rotation time of the nanoparticles is prolonged, and thus the T1 relaxation rate of the nanoparticles is greatly improved, and the introduction of polydopamine can prevent the release of metal ions in the human body, thereby improving the biosafety of the nanoparticles.

[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 above-mentioned nanoparticles for contrast agent.

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

[0008] Another object of the present invention is to provide particles for optical imaging comprising the above-mentioned nanoparticles for contrast agent.

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

[0010] A nanoparticle for contrast agent, comprising an inner core and a shell layer, wherein the shell layer comprises polydopamine and albumin, and the inner 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;

[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 ferric iron, ferrous iron or manganese. Currently, the three metal elements commonly used in the field of contrast agents are iron, manganese and gadolinium, but contrast agents prepared using iron or manganese elements have higher biosafety, so using iron or manganese elements to construct contrast agents has become a mainstream trend.

[0014] In the nanoparticle provided by the present invention, the core is a complex formed by metal ions and polyphenols, wherein the metal ions provide paramagnetism, so that the nanoparticles can produce a locally enhanced magnetic field under external action, and enhance the magnetic resonance signal formed by the relaxation of adjacent hydrogen protons. However, the design of magnetic resonance contrast agents also needs to consider the amount of metal, the exchange rate of contrast agents and water molecules, the rotation-related speed of contrast agent molecules, the toxicity of contrast agents and metabolic efficiency and other factors except the paramagnetism of the metal, so the core structure does not simply include metal ions in the nanoparticle provided by the present invention, but includes a complex of metal ions and polyphenols, wherein polyphenols can also improve the loading efficiency of metal ions, thus improving the effective content of metal ions in the nanoparticles, and then improving the relaxation rate of the nanoparticles. In addition, metal ions easily cause Fenton reaction in the human body and cause excessive reactive oxygen free radicals (ROS) to be produced in the human body, and polyphenols have good antioxidant capacity, therefore the Fenton reaction caused by metal ions can be suppressed, and the biosafety of nanoparticles is improved.

[0015] The nanoparticles provided by the present invention have a shell layer comprising polydopamine and albumin, and the dopamine unit adjacent to the metal ion in the polydopamine can also form a coordination bond with the metal ion, wherein the albumin in the shell layer 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 rotation correlation time of the particles, and further increase the T1 relaxation rate thereof; and polydopamine can also improve the structural stability of the nanoparticles, thereby preventing the nanoparticles from releasing metal ions in the human body and causing harm.

[0016] The reason why we need to control the ratio of polyphenols in the core to polydopamine in the shell is to ensure that nanoparticles can be formed; if the amount of shell components added is too little, the shell cannot wrap the core, resulting in the inability to form nanoparticles. When used for imaging, the metal ions in the core are easily released and remain in the human 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 inner 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, especially controlling the molar ratio to 1:1, nanoparticles with a higher MRI-T1 relaxation rate 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, 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 for contrast agents provided by the present invention have a stable structure and a large total molecular weight of the system, so the rotation 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 polyphenol and metal salt in a solvent to obtain a solution of 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 of 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 to 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 make each component solution carry out high-speed coordination self-assembly with the same surface force, completely solving the defect of uneven particle size prepared by traditional self-assembly preparation method, and has the advantages of fast self-assembly efficiency, continuous large-scale preparation, uniform and controllable particle size. At the same time, according to actual needs, the raw materials and proportions 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, so as to diversify the products and diversify the application fields.

[0045] In a specific embodiment of the present invention, in step S2, each solution is introduced into the multi-channel 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 min. More specifically, the reaction time in step S2 is 0.5 to 1 min. The target nanoparticles can be directly obtained by respectively passing the metal ion-polyphenol complex solution, the dopamine monomer solution and the albumin solution into a multi-channel reactor without deliberately controlling the reaction time.

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

[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 relaxation rate 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-carrying particle, comprising the above-mentioned nano particle for contrast agent and a targeting drug loaded in the shell layer of the nano particle.

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

[0059] The nanoparticles provided by the present invention have high relaxivity and excellent biosafety and compatibility, and the albumin in the particle structure has a unique spatial structure and can be a delivery carrier for drugs and / or fluorescent markers. Therefore, the nanoparticles can be used to prepare magnetic resonance contrast agents and can also be used as carriers to load and transport drugs and / or fluorescent markers, thereby obtaining the synergistic performance of magnetic resonance imaging, drug therapy, and fluorescent imaging. The electronegativity of albumin can also effectively extend the circulation time of the delivery system. Among them, when transporting 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 nano drug-carrying particles, the preparation method of the nano drug-carrying particles comprises 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 targeted drug are reacted under alkaline conditions to obtain nano drug-loaded particles.

[0063] In a specific embodiment of the present invention, the method for preparing the above-mentioned nano drug-carrying particles, except for adding a targeting drug in step S2, the remaining steps are consistent with the method for preparing nano particles for contrast agents.

[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, in step S2, the metal ion-polyphenol complex solution, the dopamine monomer solution, the albumin solution and the targeted drug solution are respectively introduced into a multi-channel 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 the contrast agent nanoparticles are combined with the fluorescent marker to form optical imaging particles, the preparation method of 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, in step S2, the metal ion-polyphenol complex solution, the dopamine monomer solution, the albumin solution and the fluorescent marker solution are respectively introduced into a multi-channel 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 ug / 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 T1 relaxation rate of 12 mM, because the components are closely connected to prolong the rotation time of the nanoparticles. -1 s -1 As mentioned above, the coating of polydopamine 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.

[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 medium ( Figure 6 B).

[0080] Figure 7 The TMB color development and absorption value 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.

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

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

[0084] Example 1

[0085] A nanoparticle for a contrast agent, represented by Mn-GA@BSA@DA, wherein the shell is polydopamine (DA) and bovine serum albumin (BSA) connected by covalent bonds, and the core is a manganese ion (Mn)-gallic acid (GA) complex, wherein the manganese ion in the complex is connected 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 agent in this embodiment comprises the following steps:

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

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

[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 nanoparticle for contrast agent, represented by Fe-GA@BSA@DA, is different 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 is different from that in Embodiment 1 only in that:

[0097] Step S1 replaces manganese chloride with ferrous chloride.

[0098] Example 3

[0099] A nanoparticle for contrast agent, represented by Mn-Quercetin@BSA@DA, is different 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 is different from that in Embodiment 1 only in that:

[0102] Step S1 replaces pyrogallic acid with quercetin.

[0103] Example 4

[0104] A nanoparticle for contrast agent, represented by Mn-TA@BSA@DA, is different 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 is different from that in Embodiment 1 only in that:

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

[0108] Example 5

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

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

[0111] The method for preparing nanoparticles for contrast agents in this embodiment is different from that in Embodiment 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 the concentration and pH are consistent with those of the bovine serum albumin solution.

[0113] Example 6

[0114] A nanoparticle for contrast agent, represented by Mn-GA@HSA@DA, is different 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 is different from that in Embodiment 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 value are consistent with those of the bovine serum albumin solution.

[0118] Example 7

[0119] A nanoparticle for contrast agent, represented by Mn-GA@BSA@DA-2, is different 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 is different from that in Embodiment 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 nanoparticle for contrast agent, represented by Mn-GA@BSA@DA-3, is different 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 is different from that in Embodiment 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 nanoparticle for contrast agent, represented by Mn-GA@BSA@DA-4, is different 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 is different from that in Embodiment 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 nanoparticle for contrast agent, represented by Mn-GA@BSA@DA-5, is different 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 is different from that in Embodiment 1 only in that:

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

[0138] Embodiment 11

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

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

[0141] The method for preparing nanoparticles for contrast agents in this embodiment is different from that in Embodiment 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 nanoparticle for contrast agent, represented by Mn-GA@BSA@DA-7, is different 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 is different from that in Embodiment 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 structure nanoparticle, represented by Mn@BSA@DA, is different 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 is different from that in Example 1 only in that:

[0152] In step S1, pyrogallic acid is not added.

[0153] Comparative Example 2

[0154] A core-shell structure nanoparticle, represented by Mn-GA@BSA, is different 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 is different 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: A 1.5T magnetic resonance imaging system was used to perform magnetic resonance detection on the nanoparticles obtained in the embodiment and the comparative example 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 Embodiment 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 conducted.

[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 particles are not easy to agglomerate. From the analysis of the data of Examples 1, 7 to 9 in Table 1, it can be seen that when the molar ratio of the polyphenol substance in the core of the nanoparticle 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. From a comprehensive analysis of the data of Examples 1, 10 to 12 in Table 1, it can be seen that when the molar ratio of polydopamine to albumin in the shell of the nanoparticle 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 obtained nanoparticles also have a higher relaxation rate and Zeta potential absolute value, but the subsequent exploration will find that the biosafety and stability of the nanoparticles in Comparative Example 2 are insufficient:

[0169] In order to explore 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 powders, and the particle sizes before and after freeze-drying were 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 re-dissolution, 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 of the particle size stability of the above two particles in 1640 medium. Figure 6 It can be seen that polydopamine can strengthen the stability of particles, thereby increasing the biological safety 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 of 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 is blue, it means that a Fenton reaction has occurred in the system. The experimental results are as follows: 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, and it was found that the positive control group MnCl2 showed an obvious blue color, while the nanoparticles hardly changed color, indicating that after the phenolic hydroxyl groups on the polyphenols were coordinated with metal ions, 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 test

[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 the present application as a nanoparticle delivery system.

[0176] Experimental Example 1

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

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

[0179] The method for preparing the optical imaging particles in this embodiment is different from that in Embodiment 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 ug / ml.

[0181] The nanoparticles were injected into mice to obtain fluorescence imaging results at different times, such as 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 highlighting (circled arrow part) can be seen in the Mn-GA@BSA@DA@IR780 group. 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, is different 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 of Fig. 9 As shown in A.

[0186] The method for preparing the optical imaging particles in this embodiment is different from that in Embodiment 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 ug / ml.

[0188] Experimental Example 3

[0189] A nano drug-carrying particle, represented by Mn-GA@BSA@DA@paclitaxel, is different 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 of Fig. 9 As shown in B.

[0192] The preparation method of the drug-carrying nanoparticles in this embodiment is different 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 fluorescent 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 solution of the present invention rather than to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solution of the present invention can be modified or replaced by equivalents without departing from the essence and scope of the technical solution of the present invention.

Claims

1. A nanoparticle for contrast agent, characterized in that: It comprises an inner core and a shell, wherein the shell comprises polydopamine and albumin, and the inner 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 mass ratio of polydopamine to albumin in the shell layer is (0.2-5):

1.

2. The nanoparticle for contrast agent according to claim 1, characterized in that: The polydopamine in the shell layer is linked to the albumin via a covalent bond.

3. The nanoparticles for contrast agent according to claim 1, characterized in that: The molar ratio of polyphenols to metal ions in the inner core is (0.5-2):

1.

4. The nanoparticle for contrast agent according to claim 1, characterized in that: The molar ratio of the polyphenol in the core to the polydopamine in the shell is (0.5-3):

1.

5. The nanoparticles for contrast agent according to claim 1, characterized in that: The mass ratio of polydopamine to albumin in the shell layer is (1-2):

1.

6. The nanoparticle for contrast agent according to claim 1, characterized in that: The polyphenols include at least one of tannic acid, catechin, epicatechin, quercetin and gallic acid.

7. The method for preparing nanoparticles for contrast agents according to any one of claims 1 to 6, characterized in that: The steps include: S1. dissolving polyphenol and metal salt in a solvent to obtain a solution of 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.

8. A contrast agent, characterized in that The nanoparticles for contrast agent include those described in any one of claims 1 to 6.

9. A drug-carrying nanoparticle, characterized in that: The invention comprises the nanoparticles for contrast agent according to any one of claims 1 to 6 and a targeting drug loaded in the shell of the nanoparticles.

10. A particle for optical imaging, characterized in that: The contrast agent comprises the nanoparticles according to any one of claims 1 to 6 and a fluorescent marker loaded in the shell of the nanoparticles.

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