CN117427032A - Bioactive component delivery system, delivery method and application - Google Patents
Bioactive component delivery system, delivery method and application Download PDFInfo
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- 238000002716 delivery method Methods 0.000 title abstract description 4
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- 210000004623 platelet-rich plasma Anatomy 0.000 claims abstract description 43
- HRPVXLWXLXDGHG-UHFFFAOYSA-N Acrylamide Chemical compound NC(=O)C=C HRPVXLWXLXDGHG-UHFFFAOYSA-N 0.000 claims abstract description 27
- 238000002604 ultrasonography Methods 0.000 claims abstract description 25
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/06—Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/14—Blood; Artificial blood
- A61K35/16—Blood plasma; Blood serum
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- A61K35/14—Blood; Artificial blood
- A61K35/19—Platelets; Megacaryocytes
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/32—Macromolecular compounds obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. carbomers, poly(meth)acrylates, or polyvinyl pyrrolidone
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- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0002—Galenical forms characterised by the drug release technique; Application systems commanded by energy
- A61K9/0009—Galenical forms characterised by the drug release technique; Application systems commanded by energy involving or responsive to electricity, magnetism or acoustic waves; Galenical aspects of sonophoresis, iontophoresis, electroporation or electroosmosis
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- A61M37/00—Other apparatus for introducing media into the body; Percutany, i.e. introducing medicines into the body by diffusion through the skin
- A61M37/0092—Other apparatus for introducing media into the body; Percutany, i.e. introducing medicines into the body by diffusion through the skin using ultrasonic, sonic or infrasonic vibrations, e.g. phonophoresis
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- A61M37/00—Other apparatus for introducing media into the body; Percutany, i.e. introducing medicines into the body by diffusion through the skin
- A61M2037/0007—Other apparatus for introducing media into the body; Percutany, i.e. introducing medicines into the body by diffusion through the skin having means for enhancing the permeation of substances through the epidermis, e.g. using suction or depression, electric or magnetic fields, sound waves or chemical agents
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Abstract
The invention discloses a bioactive component delivery system, a delivery method and application, wherein the bioactive component delivery system comprises hydrogel and an ultrasonic therapeutic apparatus, and the preparation raw materials of the hydrogel comprise acrylamide monomers, activated platelet-rich plasma and a crosslinking agent; delivering said activated platelet rich plasma under ultrasound of said ultrasound therapeutic apparatus, said ultrasoundThe intensity of sound was 0.1W/cm 2 ‑2.0W/cm 2 The frequency is 1.0MHz to 1.5MHz. The low-frequency ultrasonic energy in the invention obviously promotes the release of activated platelet-rich plasma in the hydrogel and promotes the action and effect thereof. When the hydrogel is used for wound repair, the hydrogel can remarkably promote skin wound healing under the action of low-frequency ultrasound, and has no scar.
Description
Technical Field
The invention relates to the technical field of biological medicine, in particular to a delivery system, a delivery method and application of a bioactive component.
Background
The skin is the largest organ of the human body and plays a key role in maintaining homeostasis and protecting internal organs from the external environment. Skin injuries, particularly chronic wounds, burns and infected wounds, require long-term, laborious treatment, which creates a significant economic burden on the worldwide healthcare system. Therefore, there is an urgent need for effective wound healing strategies.
Hydrogels are considered ideal dressings for wound repair due to their excellent physicochemical structure and functional properties. The high water content of the hydrogel keeps the granulation and epithelial tissues in a moist environment, its soft texture and low interfacial tension gives them good biocompatibility and non-irritating, and helps reduce inflammatory responses in the adjacent areas. The hydrogel dressing is soft and elastic, and is easy to remove after wound healing without any discomfort to the patient. The hydrogel dressing also reduces the temperature of the wound, thereby reducing pain. However, most of the existing hydrogel dressings have complex preparation processes and the preparation effect needs to be improved.
Platelet Rich Plasma (PRP) is a concentrated platelet rich plasma product containing 4 to 6 times higher number of platelets than normal blood. When PRP is activated, the platelet releases a large amount of bioactive factors, which can promote the repair and growth of human tissues and is widely applicable to surgical treatment. However, since the active factors are released in a large amount in a short time after PRP is activated and are easily degraded by enzymes in the body of the wound surface, the clinical application of PRP is greatly limited.
Disclosure of Invention
The present invention aims to solve at least one of the above technical problems in the prior art. To this end, the object of the invention is to provide a method.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
in a first aspect of the present invention, a bioactive component delivery system is provided, comprising a hydrogel and an ultrasonic therapeutic apparatus, wherein the hydrogel is prepared from raw materials including acrylamide monomer, activated platelet rich plasma and a crosslinking agent; delivering the activated platelet rich plasma under the ultrasonic of the ultrasonic therapeutic instrument, wherein the frequency of the ultrasonic is 1.0 MHz-1.5 MHz.
In the invention, the low-frequency ultrasonic energy remarkably promotes the release of activated platelet-rich plasma in the hydrogel and promotes the action and effect thereof.
In some embodiments of the invention, the ultrasound has an intensity of 0.1W/cm 2 -2.0W/cm 2 。
In some embodiments of the invention, the ultrasound has an intensity of 0.5W/cm 2 -1.0W/cm 2 The frequency is 1.0MHz to 1.5MHz.
In some embodiments of the invention, the acrylamide monomer and the activated platelet rich plasma are physically crosslinked to form a first network and then crosslinked to the acrylamide monomer via a crosslinking agent to form a second crosslinked network.
In some embodiments of the invention, the physical action is selected from at least one of hydrogen bonding, physical entanglement.
In some embodiments of the invention, the mass ratio of the activated platelet rich plasma to the acrylamide monomer is 15% to 30%.
In some embodiments of the invention, the molar ratio of the acrylamide monomer to the crosslinker is 1: 0.005-1: 0.004.
in some embodiments of the invention, the cross-linking agent is selected from at least one of methylene bisacrylamide, polyethylene glycol dienoate.
In some embodiments of the invention, the activation enrichmentPlatelet plasma is the activated platelet rich plasma supernatant; preferably, ca-containing 2+ And/or thrombin solutions activate platelet rich plasma.
In some embodiments of the invention, the method of preparing the hydrogel comprises the steps of: mixing an aqueous solution of acrylamide monomer with an activated platelet rich plasma solution; and adding a photoinitiator and a crosslinking agent, mixing, and irradiating by ultraviolet light to obtain the hydrogel.
In some embodiments of the invention, the photoinitiator is selected from at least one of 2-hydroxy-2-methyl-1- [4- (2-hydroxyethoxy) phenyl ] -1-propanone, phenyl (2, 4, 6-trimethylbenzoyl) phosphate lithium salt.
In some embodiments of the invention, the photoinitiator is present at a concentration of 0.1% (w/v) to 0.25% (w/v).
In some embodiments of the invention, the concentration of the cross-linking agent is from 0.12% (w/v) to 0.15% (w/v).
In some embodiments of the invention, the ultraviolet light is irradiated with ultraviolet light having a wavelength of 350nm to 380nm for a period of 30s to 1min.
In a second aspect of the invention, a method of drug delivery is presented, comprising delivering activated platelet rich plasma in a hydrogel under ultrasound; the preparation raw materials of the hydrogel comprise acrylamide monomers, activated platelet-rich plasma and a crosslinking agent; the frequency of the ultrasonic wave is 1.0 MHz-1.5 MHz.
In some embodiments of the invention, the ultrasound has an intensity of 0.1W/cm 2 -2.0W/cm 2 。
In some embodiments of the invention, the ultrasound has an intensity of 0.5W/cm 2 -1.0W/cm 2 The frequency is 1.0MHz to 1.5MHz.
In a third aspect of the invention, an application of hydrogel in preparing a wound repair material is provided, wherein the hydrogel is matched with an ultrasonic therapeutic apparatus for use; the preparation raw materials of the hydrogel comprise acrylamide monomers, activated platelet-rich plasma and a crosslinking agent; the ultrasonic frequency of the ultrasonic therapeutic apparatus is 1.0 MHz-1.5 MHz.
In some embodiments of the invention, the ultrasound has an intensity of 0.1W/cm 2 -2.0W/cm 2 。
In some embodiments of the invention, the ultrasound has an intensity of 0.5W/cm 2 -1.0W/cm 2 The frequency is 1.0MHz to 1.5MHz.
The beneficial effects of the invention are as follows:
in the invention, the low-frequency ultrasonic energy remarkably promotes the release of activated platelet-rich plasma in the hydrogel and promotes the action and effect thereof. When the hydrogel is used for wound repair, the hydrogel can remarkably promote skin wound healing under the action of low-frequency ultrasound, and has no scar.
Drawings
FIG. 1 is a schematic illustration of a hydrogel formation process and drug delivery process under ultrasound according to an embodiment of the present invention.
FIG. 2 is a physical diagram showing the preparation process of hydrogels of examples 1 to 2 and comparative examples according to the present invention.
FIG. 3 is SEM image of hydrogel of examples 1-2 and comparative example of the present invention.
FIG. 4 is a schematic diagram showing the wound healing process of rats in test example 2 of the present invention.
FIG. 5 is a graph showing wound healing of a rat model of a blank group and a gel group under the effect of low frequency ultrasound/no low frequency ultrasound in test example 2 of the present invention.
FIG. 6 is a bar graph of wound healing rate for rats in the blank and gel groups under low frequency ultrasound/no low frequency ultrasound in test example 2 of the present invention.
Detailed Description
The present invention will be described in further detail with reference to specific examples. The starting materials, reagents or apparatus used in the examples and comparative examples were either commercially available from conventional sources or may be obtained by prior art methods unless specifically indicated. Unless otherwise indicated, assays or testing methods are routine in the art.
In the following examples or comparative examples, activated platelet rich plasma supernatants were prepared by:
purchased rat platelet rich plasma (PRP, platelets)Plate concentration was 5.5 x 10 6 mu.L, guangzhou Hongquan Biotechnology Co., ltd.) 10mL was added 200. Mu.L 1M CaCl 2 Solution, caCl 2 The final concentration of the solution was 20mM. Adding CaCl into the mixture 2 The PRP solution of (C) was allowed to activate at 37℃for 1 hour, and then placed in a refrigerator at 4℃overnight. Centrifuging (2000 g,5 min), collecting supernatant, filtering with 0.22 μm filter membrane to obtain PRP supernatant, and placing in a refrigerator at-20deg.C for use.
Example 1
The embodiment provides a bioactive component delivery system, which comprises the following specific processes:
preparation of polyacrylamide-activated platelet rich plasma supernatant (PAM-PRP-1) hydrogel:
(1) 3.0g of acrylamide monomer (AAM) was weighed and dissolved in 10mL of PBS solution containing 0.25% (w/v) phenyl (2, 4, 6-trimethylbenzoyl) phosphate lithium salt to prepare a 30% by mass concentration of AAM PBS solution, 10.0mL of supernatant of activated platelet-rich plasma was added, and stirred at room temperature for 10min to prepare a homogeneous solution.
(2) 0.030g of methylenebisacrylamide (concentration: 0.15%, w/v) was added, and the mixed solution was stirred at room temperature for 10 minutes to form a homogeneous solution.
(3) The solution was placed in an ultraviolet box with 365nm ultraviolet light. Irradiating for 30s to obtain the PAM-PRP-1 hydrogel.
Delivering the hydrogel under the action of ultrasound, wherein the ultrasonic intensity of an ultrasonic therapeutic instrument is 1W/cm 2 The ultrasonic frequency was 1MHz.
Example 2
The embodiment provides a bioactive component delivery system, which comprises the following specific processes:
preparation of polyacrylamide-activated platelet rich plasma supernatant (PAM-PRP-2) hydrogel:
(1) 3.0g of acrylamide monomer (AAM) was weighed and dissolved in 10mL of PBS solution containing 0.25% of phenyl (2, 4, 6-trimethylbenzoyl) phosphate lithium salt to prepare a PBS solution with a mass concentration of 30% AAM, 20.0mL of supernatant of activated platelet-rich plasma was added, and stirred at room temperature for 10min to prepare a homogeneous solution.
(2) 0.045g of methylenebisacrylamide (concentration: 0.15%, w/v) was added, and the mixed solution was stirred at room temperature for 10min to form a homogeneous solution.
(3) The solution is placed in an ultraviolet box, and 365nm ultraviolet light is irradiated for 30s, so that PAM-PRP-2 hydrogel is obtained.
Delivering the hydrogel under the action of ultrasound, wherein the ultrasonic intensity of an ultrasonic therapeutic instrument is 1W/cm 2 The ultrasonic frequency was 1MHz.
Comparative example
The comparative example provides a bioactive ingredient delivery system, which comprises the following specific processes:
preparation of hydrogel Polyacrylamide (PAM):
(1) 3.0g of acrylamide monomer (AAM) was weighed and dissolved in 10mL of PBS solution containing 0.25% of phenyl (2, 4, 6-trimethylbenzoyl) phosphate lithium salt to prepare a PBS solution having a mass concentration of 30% of AAM, 0.015g of methylenebisacrylamide (concentration: 0.15%, w/v) was added, and the mixed solution was stirred at room temperature for 10 minutes to form a uniform solution.
(2) The solution is placed in an ultraviolet box, and 365nm ultraviolet light is irradiated for 30s, so that PAM gel is obtained.
Fig. 1 shows the hydrogel-forming process of the example and the drug delivery process under the effect of ultrasound, and fig. 2 shows a physical diagram of the hydrogel-making process of examples 1 to 2 and comparative example.
Test example 1
The Scanning Electron Microscope (SEM) test methods for the hydrogels of examples 1-2 and comparative example 1 were: the hydrogel was lyophilized on a freeze dryer at-60 ℃, the sample was cut into small pieces and fixed on a scaffold with conductive glue. And then the surface of the sample is sprayed with metal, and conducting treatment is carried out. The morphology of the gel surface was observed.
Analysis of results: as shown in FIG. 3, the PAM gel surface formed a significant pore structure with a pore size of 36.79+ -4.16 μm. The PRP in the PAM-PRP gel is entangled in the network formed by PAM through physical winding, and because the concentration of the AAM monomer added into the PRP solution is diluted, the network pores formed by the PAM-PRP-1 hydrogel and the PAM-PRP-2 hydrogel become larger, namely 52.04 +/-5.18 mu m and 72.91 +/-4.50 mu m respectively.
Test example 2
In this test example, hydrogel is applied to a wound repair material under the action of an ultrasonic therapeutic apparatus, and the wound healing rate is tested, and fig. 4 shows a schematic diagram of a rat wound healing process, which specifically includes:
female SD rats (6-8 weeks, body weight 200-250 g), with circular wounds of approximately 1cm diameter made on the backs. Rats were divided into a blank group, a PAM hydrogel group, a PAM-PRP-1 hydrogel group and a PAM-PRP-2 hydrogel group, and were set as an ultrasound-applied group (ultrasound intensity of 1W/cm) 2 Ultrasonic frequency of 1 MHz) and no ultrasound group. The hydrogel is applied to the wound of the rat, and the ultrasonic group performs ultrasonic action by using an ultrasonic therapeutic instrument for 1 time/day. Wound healing was recorded by photographing on day 0, day 4, day 7 and day 14, and the results are shown in fig. 5. Wound healing rate was calculated using Image J software. Wound healing rate (%) = (W) 0 -W n )/W 0 ×100，W 0 And W is n Wound healing area on day 0 and day n (n= 4,7,14) are indicated, respectively. Each set of samples was provided with 5 replicates and averaged, the results are shown in figure 6.
As can be seen in fig. 5, the ultrasound group healed more rapidly than the non-ultrasound group, and the PAM-PRP-1 hydrogel in the ultrasound group healed more rapidly on days 4,7, and 14 than the PAM, PAM-PRP-2, and the placebo group. The wound healing rates of the PAM-PRP-1 hydrogel group under the low-frequency ultrasonic effect on the 4 th day, the 7 th day and the 14 th day are 25.79+/-2.64%, 73.1+/-3.44% and 99.86+/-2.11 respectively; the wound healing rates of the blank control group on the 4 th day, the 7 th day and the 14 th day under the low-frequency ultrasonic effect are 16.64+/-1.93%, 35.39+/-2.5% and 95.34+/-2.79% respectively. The wound healing rates of the PAM group under the low-frequency ultrasonic effect on the 4 th day, the 7 th day and the 14 th day are respectively 20.22+/-4.14%, 65.89+/-1.8 and 97.10 +/-1.86. The wound healing rates of the PAM-PRP-2 group on the 4 th day, the 7 th day and the 14 th day under the low-frequency ultrasonic effect are 15.94+/-2.02, 45.68+/-1.87 and 91.34 +/-1.12 respectively.
Fig. 6 is a bar graph of healing rate, which shows that the healing rate of the ultrasonic group PAM-PRP-1 and PAM-PRP-2 hydrogel groups is significantly higher than that of the ultrasonic group without ultrasonic treatment, so that the release of PRP is faster under the action of low-frequency ultrasonic treatment in the ultrasonic group, and the healing speed of the rat wound is promoted.
The above examples are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above examples, and any other changes, modifications, substitutions, combinations, and simplifications that do not depart from the spirit and principle of the present invention should be made in the equivalent manner, and the embodiments are included in the protection scope of the present invention.
Claims (10)
1. A bioactive ingredient delivery system characterized by: the preparation method comprises the steps of preparing hydrogel and an ultrasonic therapeutic apparatus, wherein the preparation raw materials of the hydrogel comprise acrylamide monomers, activated platelet-rich plasma and a crosslinking agent; delivering the activated platelet rich plasma under the ultrasonic of the ultrasonic therapeutic instrument, wherein the frequency of the ultrasonic is 1.0 MHz-1.5 MHz.
2. The bioactive ingredient delivery system of claim 1, wherein: in the hydrogel, an acrylamide monomer and activated platelet-rich plasma are crosslinked through physical action to form a first network, and then crosslinked with the acrylamide monomer through a crosslinking agent to form a second crosslinked network.
3. The bioactive ingredient delivery system of claim 1, wherein: the mass ratio of the activated platelet-rich plasma to the acrylamide monomer is 15% -30%.
4. The bioactive ingredient delivery system of claim 1, wherein: the molar ratio of the acrylamide monomer to the crosslinking agent is 1: 0.005-1: 0.004.
5. the bioactive ingredient delivery system of claim 1, wherein: the activated platelet rich plasma is an activated platelet rich plasma supernatant.
6. The bioactive ingredient delivery system of claim 1, wherein: the preparation method of the hydrogel comprises the following steps: mixing an aqueous solution of acrylamide monomer with an activated platelet rich plasma solution; and adding a photoinitiator and a crosslinking agent, mixing, and irradiating by ultraviolet light to obtain the hydrogel.
7. A method of drug delivery, characterized by: comprising delivering activated platelet rich plasma in a hydrogel under ultrasound; the preparation raw materials of the hydrogel comprise acrylamide monomers, activated platelet-rich plasma and a crosslinking agent; the frequency of the ultrasonic wave is 1.0 MHz-1.5 MHz.
8. An application of hydrogel in preparing a wound repair material, which is characterized in that: the hydrogel is matched with an ultrasonic therapeutic instrument for use; the preparation raw materials of the hydrogel comprise acrylamide monomers, activated platelet-rich plasma and a crosslinking agent; the ultrasonic frequency of the ultrasonic therapeutic apparatus is 1.0 MHz-1.5 MHz.
9. The bioactive ingredient delivery system according to claim 1, the drug delivery method according to claim 7 or the use according to claim 8, characterized in that: the intensity of the ultrasonic wave is 0.1W/cm 2 -2.0W/cm 2 。
10. The bioactive ingredient delivery system according to claim 1, the drug delivery method according to claim 7 or the use according to claim 8, characterized in that: the intensity of the ultrasonic wave is 0.5W/cm 2 -1.0W/cm 2 The frequency is 1.0MHz to 1.5MHz.
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