CN115025220B

CN115025220B - Modification method for improving biocompatibility of photosensitizer

Info

Publication number: CN115025220B
Application number: CN202210710748.5A
Authority: CN
Inventors: 卢春华; 王敏; 何华明; 吕瞳; 杨黄浩
Original assignee: Fuzhou University
Current assignee: Fuzhou University
Priority date: 2022-06-22
Filing date: 2022-06-22
Publication date: 2023-07-18
Anticipated expiration: 2042-06-22
Also published as: CN115025220A

Abstract

The invention discloses a simple modification method for improving biocompatibility of a photosensitizer, which is to jointly react a hydrophobic photosensitizer with an organic molecule containing multiple carboxyl groups and an organic molecule containing multiple amino groups through a solid phase reaction strategy to prepare the modified photosensitizer with high biocompatibility. The obtained modified photosensitizer has good water solubility, has low cytotoxicity even under the condition of high concentration, can effectively avoid side effects caused by the hydrophobic photosensitizer, and improves the biocompatibility of the photosensitizer. Meanwhile, the modified photosensitizer can avoid aggregation and self-quenching, and improves the generation efficiency of singlet oxygen. Therefore, the modification method provided by the invention provides a new opportunity for promoting the application of photodynamic therapy in biomedicine.

Description

Modification method for improving biocompatibility of photosensitizer

Technical Field

The invention belongs to the field of photodynamic therapy, and particularly relates to a modification method for improving biocompatibility of a photosensitizer.

Background

Photodynamic therapy (PDT) is an emerging cancer treatment method with the characteristics of simplicity, non-invasiveness and light controllability on demand. PDT relies on Photosensitizers (PSs) used under light irradiation to convert endogenous oxygen into cytotoxic Reactive Oxygen Species (ROS) in vivo, triggering apoptosis and inhibiting tumor growth. During photodynamic processes, photosensitizers play a critical role in the generation of highly reactive singlet oxygen. Ideal photosensitizers for PDT should have good biocompatibility and phototoxicity, that is, no toxicity to cells in the absence of an external light source, but, on the contrary, be effective in killing cancer cells upon irradiation with an external light source.

Although various photosensitizers are used in clinical treatment at present, the conventional clinically used photosensitizers are hydrophobic, have certain biotoxicity and weak killing ability to tumors, and greatly limit the application of photodynamic therapy of the photosensitizers in clinic. In addition, due to the hydrophobic interactions of the photosensitizers, the stacking aggregate is also prone to form inactive aggregates, producing a self-quenching effect, resulting in low singlet oxygen production, since the stacked molecules release the absorbed energy mainly in the form of heat. Therefore, they can only be used in good solvents and low concentrations, which greatly limits the clinical application of photodynamic therapy. In this sense, good water solubility, while avoiding aggregation caused by superposition, is significant in improving the bioavailability of the photosensitizer and the production of active oxygen.

Disclosure of Invention

Therefore, the invention aims to provide a modification method for improving the biocompatibility of a photosensitizer, which aims to solve the problems of low biocompatibility and poor bioavailability of the existing photosensitizer.

In order to achieve the above purpose, the invention adopts the following technical scheme:

a modifying method for improving the biocompatibility of photosensitiser features that the solid-phase synthesis method is used to make the hydrophobic photosensitiser 1-800-mg react with the organic molecule containing multiple carboxyl 1-800-mg and the organic molecule containing multiple amino 1-800-mg to obtain the modified photosensitiser with high biocompatibility.

Wherein the hydrophobic photosensitizer is 5,10,15, 20-tetra (4-carboxyphenyl) porphyrin or 5,10,15, 20-tetra (4-aminophenyl) porphyrin.

The organic molecule containing multiple carboxyl groups is sodium citrate.

The polyamino-containing organic molecule is urea or melamine.

The reaction temperature is 180-240 ℃ and the reaction time is 1-5 hours.

Preferably, the hydrophobic photosensitizer is used in an amount of 80-120 mg, the preferred amount of the organic molecule containing a polycarboxylic group is 10-50 mg, and the preferred amount of the organic molecule containing a polyamino group is 100-300 mg.

The particle size of the obtained modified photosensitizer with high biocompatibility is below 10 nm.

The invention has the beneficial effects that:

the preparation process is simple, does not need complex equipment, and is green and environment-friendly; the obtained modified photosensitizer is a quantum dot below 10 nm, has good water solubility due to quantum dot effect and a large number of hydrophilic groups (carboxyl or amino) contained on the surface, has low cytotoxicity even under the condition of high concentration, can effectively avoid side effects caused by the hydrophobic photosensitizer, and improves the biocompatibility of the photosensitizer. Meanwhile, the generation of aggregation and self-quenching of the obtained photosensitizer can be avoided after modification, and the generation efficiency of singlet oxygen is improved. Therefore, the modification method provided by the invention provides a new opportunity for promoting the application of photodynamic therapy in biomedicine.

Drawings

FIG. 1 is an electron microscope scan of the modified photosensitizer obtained in example 1;

FIG. 2 is an atomic force microscope image of the modified photosensitizer obtained in example 1;

FIG. 3 is a graph showing the photodynamic properties of the modified photosensitizer obtained in example 1;

FIG. 4 is a graph showing the comparison of photodynamic properties of the modified photosensitizer and porphyrin materials obtained in example 1;

FIG. 5 is a diagram showing the biocompatibility of the modified photosensitizer and the raw material mixture obtained in example 1;

FIG. 6 is a graph showing the biocompatibility of the modified photosensitizer with respect to different cells obtained in example 1;

FIG. 7 is a graph showing phototoxicity of the modified photosensitizer obtained in example 1.

Detailed Description

In order to make the contents of the present invention more easily understood, the technical scheme of the present invention will be further described with reference to the specific embodiments, but the present invention is not limited thereto.

Example 1

(1) 5,10,15, 20-tetra (4-carboxyphenyl) porphyrin (110.7 mg), citric acid (26.7 mg) and urea (201.6 mg) were mixed and ground in an agate mortar for 30 minutes;

(2) Adding the ground mixed powder into a reaction kettle, heating for 1 hour at 180 ℃ in an oven, cooling to room temperature, taking out, dispersing with water, and dialyzing to obtain the modified photosensitizer.

FIG. 1 is an electron microscope scan of the modified photosensitizer obtained in this example. As can be seen, the modified photosensitizer has good dispersibility, relatively uniform size, and a size of about 5 a nm a.

FIG. 2 is an atomic force microscope image of the modified photosensitizer obtained in this example. As shown in the figure, the height of the modified photosensitizer is 5-6 nm.

Further, the modified photosensitizer obtained in example 1 was mixed with a solution of an amount of active oxygen probe (DCFH-DA) at 100 mW/cm with a laser of 660nm ² Is irradiated for 10 min. Excitation spectrum of the solution was measured by light excitation of 488 and nm, and the result is shown in fig. 3. As can be seen from fig. 3, the fluorescence intensity of the active oxygen probe increases with increasing illumination time under the irradiation of 660nm laser light, indicating that the modified photosensitizer is capable of continuously generating active oxygen under the irradiation of near infrared light.

Further, the modified photosensitizer obtained in example 1 and 5,10,15, 20-tetrakis (4-carboxyphenyl) porphyrin were mixed with a solution of a certain amount of active oxygen probe (DCFH-DA), respectively, and laser light of 660nm was used at 100 mW/cm ² Is irradiated for 10 min. Excitation spectrum of the solution was measured by light excitation of 488 and nm, and the result is shown in fig. 4. As can be seen from fig. 4, the photodynamic properties of the modified photosensitizer are superior to those of the porphyrin raw material.

Further, the modified photosensitizer obtained in example 1 and the mixed powder of the raw materials were added to a 96-well plate in which MCF-7 cells were cultured, respectively, washed with PBS after 24-h incubation, mixed solution of CCK-8 and a medium was added, incubated for 30 minutes, absorbance values were measured at 450nm in the well plate, and the viability of the cells was obtained after conversion, and the results are shown in FIG. 5. As can be seen from fig. 5, the modified photosensitizer has lower biotoxicity and better biocompatibility.

Further, the modified photosensitizer obtained in example 1 was added to a 96-well plate in which different cells (MCF-7, hela, A549, L02) were cultured, washed with PBS after 24-h incubation, a mixed solution of CCK-8 and a medium was added, and after 30 min incubation, the absorbance value of the well plate was measured at 450nm, and the viability of the cells was obtained after conversion, and the results are shown in FIG. 6. As can be seen from fig. 6, the modified photosensitizer has only low cytotoxicity even at high concentration, and this high biocompatibility has no cell difference.

At the same time, the modified photosensitizer obtained was added to a 96-well plate in which MCF-7 cells were cultured, incubated 4. 4 h, washed off with PBS, and irradiated with 660. 660nm laser at 100 mM/cm for each well ² After incubation for 24 min h, a mixed solution of CCK-8 and medium was added, after incubation for 30 min, the absorbance value was measured at 450nm in the well plate, and the cell viability was obtained after conversion, and the results are shown in fig. 7. As can be seen from fig. 7, the photosensitizer exhibits a high cell killing ability under irradiation of near infrared light.

Example 2

(1) 5,10,15, 20-tetra (4-aminophenyl) porphyrin (94.6 mg), citric acid (26.7 mg) and urea (201.6 mg) were mixed and added to an agate mortar for grinding for 30 minutes;

(2) The ground mixed powder was added to the reaction vessel and heated in an oven at 180℃for 1 hour. After cooling to room temperature, taking out, dispersing with water, and dialyzing to obtain the modified photosensitizer, wherein the size of the modified photosensitizer is 5 nm.

Example 3

(1) 5,10,15, 20-tetrakis (4-carboxyphenyl) porphyrin (110.7 mg), citric acid (26.7 mg) and melamine (252.2 mg) were mixed and added to an agate mortar for grinding for 30 minutes;

The foregoing description is only of the preferred embodiments of the invention, and all changes and modifications that come within the meaning and range of equivalency of the claims are therefore intended to be embraced therein.

Claims

1. A modification method for improving biocompatibility of a photosensitizer is characterized by comprising the following steps: the method comprises the following steps:

(1) 110.7mg 5,10,15,20-tetra (4-carboxyphenyl) porphyrin, 26.7 mg citric acid and 201.6 mg urea were mixed and added to an agate mortar for grinding for 30 minutes;