Celecoxib-vitamin e prodrug nanoparticles based on electrostatic spray and methods of making and using the same

Abstract

The present application relates to a kind of celecoxib-vitamin E prodrug nano drug delivery system preparation based on electrostatic spray.The dithio-bis-acetic acid is used to connect celecoxib (CEL) with vitamin E (VE), and the celecoxib-vitamin E prodrug (CEL-SVE) is prepared.On this basis, the celecoxib-vitamin E prodrug nanoparticle (CEL-SVE NPs) is further prepared.The prodrug nanoparticle has some characteristics: the average particle size is less than 400 nm, is uniformly distributed in the form of spherical shape, and the drug loading can reach 6 mg•mL ‑1 Under acidic conditions, it is gradually hydrolyzed, and is relatively stable under pH 7.4 conditions.The prodrug nanoparticle can increase the solubility of celecoxib, significantly improve the bioavailability of celecoxib, increase the targeting effect of celecoxib on tumor cells, and significantly improve the anticancer and anti-inflammatory analgesic efficacy of celecoxib.It has good stability, high safety, and great market prospect.

Description

Technical Field

[0001] This invention belongs to the field of new excipients and dosage forms for pharmaceutical preparations, including the synthesis of celecoxib-vitamin E prodrug and the construction of celecoxib-vitamin E prodrug nanoparticles based on electrostatic spraying, and their application in drug delivery. Background Technology

[0002] Celecoxib is a second-generation specific cyclooxygenase-2 (COX-2) inhibitor, primarily used clinically to treat osteoarthritis, rheumatoid arthritis, ankylosing spondylitis, and acute pain in adults. Compared to non-selective cyclooxygenase inhibitors (such as aspirin and acetaminophen), celecoxib has less effect on cyclooxygenase-1 (COX-1) and does not affect the synthesis of prostacyclin (PGI2), which has a protective effect on the gastrointestinal tract and kidneys. This significantly reduces the likelihood of gastrointestinal complications and kidney side effects. In addition, celecoxib can significantly reduce the recurrence rate of sporadic colorectal adenomas.

[0003] Prodrugs exert their effects through changes in their molecular structure. Biological prodrugs themselves are inactive; their active components are their metabolites within the body. This avoids the inactivation caused by metabolic reactions, instead utilizing the body's metabolism to generate the active compound. Combining prodrug technology with celecoxib can improve drug properties while simultaneously enhancing solubility and bioavailability. Therefore, preparing a celecoxib-vitamin E prodrug can reduce the gastrointestinal irritation of celecoxib and improve its bioavailability. Thus, leveraging the structural characteristics of celecoxib, celecoxib is linked to vitamin E using dithiocarbamate to synthesize a celecoxib-vitamin E prodrug, which is further prepared into celecoxib-vitamin E prodrug nanoparticles. This reduces the gastrointestinal irritation of celecoxib, improves its bioavailability, and enhances its targeting effect on tumor cells, thereby improving its antioxidant and lipid-lowering effects.

[0004] Electrostatic spraying is a method that uses high-voltage electrostatics to form micro, nanoparticles or fibers from polymer solutions or melts, and it has wide applications in materials science, pharmaceuticals, and biomedical engineering. The principle of electrostatic spraying is to apply a high voltage to the nozzle tip where the polymer solution is added, or directly to the polymer solution. On the surface of the droplets at the nozzle tip, charges concentrate and repel each other, causing the droplets flowing out of the nozzle to gradually form what is called a Taylor cone. Further increasing the electric field force forms a jet. As the solvent evaporates or the melt cools and solidifies, the polymer material finally falls onto a collecting plate, forming nanodroplets. The smaller the diameter of the drug-loaded droplets, the larger the specific surface area, which is beneficial for improving drug dispersion and dissolution rate.

[0005] This invention combines electrostatic spraying technology with prodrug nanodelivery technology to develop a self-assembled nanodelivery system for celecoxib prodrug with dual targeting effects. The aim is to improve celecoxib solubility, accelerate dissolution rate, increase bioavailability, expand administration routes, enhance targeting efficiency, and reduce side effects. This provides a new approach and research reference for addressing the bottleneck issues in the development of celecoxib as an anticancer drug, and also for solving the solubilization and targeted application of celecoxib and other poorly soluble drugs. Summary of the Invention

[0006] One of the objectives of this invention is to provide celecoxib-vitamin E prodrug and its preparation method.

[0007] The second objective of this invention is to provide celecoxib-vitamin E prodrug nanoparticles and their preparation method.

[0008] A third objective of this invention is to provide the application of celecoxib-vitamin E prodrug nanoparticles in the preparation of anti-inflammatory and analgesic drugs, anti-colorectal cancer drugs, and to improve the bioavailability of celecoxib.

[0009] The fourth objective of this invention is to provide the application of celecoxib-vitamin E prodrug nanoparticles in celecoxib oral or topical drug delivery systems.

[0010] The technical solution of the present invention is as follows:

[0011] A celecoxib-vitamin E prodrug, which is formed by linking celecoxib to one molecule of vitamin E with dithiodiacetic acid as the parent drug, and the structural formula of the celecoxib-vitamin E prodrug is as follows:

[0012]

[0013] A method for preparing the above-mentioned celecoxib-vitamin E prodrug, comprising the following steps:

[0014] Step 1: Dissolve 1g of dithionyl diacetic acid in 10-20 mL of acetic anhydride and stir at room temperature for 3-4 hours. After the reaction is complete, evaporate to dryness by rotary evaporation. Dissolve the crude product in 15-40 mL of dichloromethane, add 10-30 mg of 4-dimethylaminopyridine (DMAP) and 500 mg of vitamin E, and stir at room temperature for 2 hours. Purify the reaction solution by silica gel column chromatography. The eluent is petroleum ether-ethyl acetate-glacial acetic acid (95:5:0.1 ~ 80:20:0.1, V / V) to obtain S-VE.

[0015] Step 2: Dissolve 500 mg of S-VE in dichloromethane, add 100-400 mg of DCC and 10-30 mg of DMAP, stir at room temperature for 5 min, add 250 mg of celecoxib, stir at room temperature for 2 hours, purify the reaction solution by silica gel column chromatography, using dichloromethane-methanol (100:0-90:10 V / V) as the eluent. A light yellow viscous semi-solid is obtained, which is the celecoxib vitamin E prodrug.

[0016] The reaction formula is as follows:

[0017]

[0018] A celecoxib-vitamin E prodrug nanoparticle based on electrostatic spraying is prepared by reacting the aforementioned celecoxib-vitamin E prodrug and a carrier in a dichloromethane solution. The celecoxib-vitamin E prodrug nanoparticles have an average particle size of less than 700 nm, exhibit a near-spherical, uniform distribution, and a drug loading of up to 6 mg / mL based on celecoxib. -1 .

[0019] A method for preparing the above-mentioned celecoxib-vitamin E prodrug nanoparticles based on electrostatic spraying involves dissolving the celecoxib-vitamin E prodrug and a carrier together in dichloromethane and stirring until a clear, transparent, and homogeneous solution is formed, which is the precursor solution for electrostatic spraying. The precursor solution is added to a 10 mL syringe, which is then attached to a micro-injection pump. The injection rate is controlled by the micro-injection pump, and the spray voltage and electrostatic spraying distance are adjusted by regulating the high-voltage power supply. The electrostatic spraying experiment is then initiated.

[0020] The electrostatic spraying voltage of the above-mentioned celecoxib-vitamin E prodrug nanoparticles is 15-20kV.

[0021] The electrostatic spray flow rate of the above-mentioned celecoxib-vitamin E prodrug nanoparticles is 0.02-0.08 mm / min.

[0022] The electrostatic spraying distance of the above-mentioned celecoxib-vitamin E prodrug nanoparticles is 10-20cm.

[0023] The above-mentioned celecoxib-vitamin E prodrug nanoparticles are used in the preparation of nanomedicine delivery systems.

[0024] The above-mentioned application of celecoxib-vitamin E prodrug nanoparticles in drug delivery systems.

[0025] The above-mentioned celecoxib-vitamin E prodrug nanoparticles are used in the preparation of antipyretic, analgesic, and anticancer drugs.

[0026] The aforementioned celecoxib-vitamin prodrug nanoparticles can be used to prepare oral formulations for antipyretic, analgesic, and anticancer drugs.

[0027] Beneficial effects

[0028] The prepared celecoxib-vitamin E prodrug nanoparticles have an average particle size of less than 400 nm, exhibit a near-spherical and uniform distribution, and a drug loading capacity of up to 6 mg / mL based on celecoxib. -1 It gradually hydrolyzes under acidic conditions and is relatively stable at pH 7.4. These prodrug nanoparticles can increase the solubility of celecoxib, significantly improve its bioavailability, and enhance its targeting effect on tumor cells. Attached Figure Description

[0029] Figure 1 The celecoxib-vitamin E prodrug in Example 1 of this invention 1 H-NMR spectrum.

[0030] Figure 2 This is a scanning electron microscope image of the celecoxib-vitamin E prodrug nanoparticles of Example 2 of the present invention.

[0031] Figure 3 The in vitro release rate of celecoxib-vitamin E prodrug in different media in Example 6 of this invention.

[0032] Figure 4 The cell proliferation inhibition rate of HCT-116 cells in Example 9 of this invention. Detailed Implementation

[0033] The following examples are intended to help those skilled in the art better understand the invention, but are not intended to limit the invention in any way.

[0034] Example 1. Synthesis of celecoxib-vitamin E prodrug

[0035] Dissolve 1 g of dithionyldiacetic acid in 10 mL of acetic anhydride and stir at room temperature for 3-4 hours. After the reaction is complete, evaporate to dryness at 60 °C using a rotary evaporator. Dissolve the crude product in an appropriate amount of dichloromethane, add 20 mg of DMAP and 500 mg of vitamin E, and stir at room temperature for 2 hours. Purify the reaction solution by silica gel column chromatography, using petroleum ether-ethyl acetate-glacial acetic acid (95:5:0.1 ~ 80:20:0.1, V / V) as the eluent, to obtain approximately 800 mg of S-VE. Dissolve 500 mg of S-VE in an appropriate amount of dichloromethane, add 200 mg of DCC and 14 mg of DMAP, and stir at room temperature for 5 min. Add 250 mg of celecoxib and stir at room temperature for 2 hours. Purify the reaction solution by silica gel column chromatography, using dichloromethane-methanol (100:0~90:10 V / V) as the eluent, to obtain approximately 500 mg of a pale yellow viscous semi-solid, which is the celecoxib vitamin E prodrug.

[0036] The structure of celecoxib-vitamin E prodrug in Example 1 was determined by nuclear magnetic resonance.

[0037] Weigh 20 mg of the monomer component, dissolve it in 0.6 mL of deuterated chloroform, and detect it using a nuclear magnetic resonance analyzer. 1 See 1 for the H-NMR spectrum.

[0038] Example 2. Preparation of celecoxib-vitamin E prodrug nanoparticles

[0039] The preparation process is as follows: Celecoxib-vitamin E prodrug from Example 1 and the carrier were dissolved together in dichloromethane and stirred until a clear, transparent, and homogeneous solution was formed, which is the precursor solution for electrostatic spraying. The precursor solution for electrostatic spraying was added to a 10 mL syringe, which was then attached to a micro-injection pump. The injection speed was controlled by the micro-injection pump, and the spray voltage and electrostatic spraying distance were adjusted by adjusting the high-voltage power supply. The electrostatic spraying voltage was 18 kV, i.e., the electrostatic spraying flow rate was 0.05 mm / min, and the electrostatic spraying distance was 15 cm, thus obtaining celecoxib-vitamin E prodrug nanoparticles.

[0040] Example 3. Morphological observation of celecoxib-vitamin E prodrug nanoparticles

[0041] The morphology of celecoxib-vitamin E prodrug nanoparticles in Example 2 was observed by scanning electron microscopy. Specifically, celecoxib-vitamin E prodrug nanoparticles were prepared by attaching the sample to a conductive adhesive to form a thin layer, and then spraying a metal film onto the sample to make the sample conductive. Figure 2 It is in the form of celecoxib-vitamin E prodrug nanoparticles, and is spherical in shape.

[0042] Example 4. Particle size distribution of celecoxib-vitamin E prodrug nanoparticles

[0043] The particle size distribution and zeta potential of the celecoxib-vitamin E prodrug nanoparticles in Example 2 were determined using a laser particle size analyzer. Specifically, celecoxib-vitamin E prodrug nanoparticles were prepared, diluted with an appropriate amount of distilled water, and their particle size distribution was determined using a Brookhaven dynamic light scattering laser particle size analyzer. The average particle size was measured to be 350 ± 1.55 nm, indicating that this nanoparticle system is relatively stable.

[0044] Example 5. Investigation of drug content

[0045] The theoretical concentration is 4 mg / mL. -1 6 mg·mL -1 8 mg·mL -1 Celecoxib-vitamin E prodrug nanoparticles were prepared. Two mL of each nanoparticle was diluted with double-distilled water and filtered through a 0.45 μm microporous membrane. 0.1 mL of the diluted solution was then diluted to 1 mL with methanol, vortexed, and the celecoxib-vitamin E prodrug content was determined by HPLC. The results showed that the drug content of the three concentrations of celecoxib-vitamin E prodrug nanoparticles was 3.63 ± 0.02 mg∙mL⁻¹, respectively. -1 7.97 ± 0.03 mg∙mL -1 and 10.95 ± 0.12 mg∙mL -1 .

[0046] Example 6. Investigation of in vitro drug release characteristics

[0047] To prepare the celecoxib-vitamin E prodrug nanoparticles in Example 2, 1 mL of each nanoparticle was encapsulated in dialysis tape (1500-2000 Da), tied at both ends, and placed in an Erlenmeyer flask. 100 mL of each release medium was added. The release media were 10 mM, 1 mM, 10 μM, and 1 μM GSH. In vitro drug release experiments were conducted in a water bath shaker at 37 ± 1°C. At specified time points after sample release, 1 mL samples were taken, diluted with an appropriate amount of methanol, and the contents of the celecoxib-vitamin E prodrug nanoparticles and celecoxib were determined by HPLC. The results are shown below. Figure 3 The results showed that celecoxib-vitamin E prodrug nanoparticles were slowly released under water and acidic conditions, and were stable under neutral to slightly alkaline conditions.

[0048] Example 7. Pharmacokinetic Study of Celecoxib-Vitamin E Prodrug Nanoparticles

[0049] Twenty-four healthy male SD rats were randomly divided into four groups of six each. They were administered 50 mg / kg of medication. -1Celecoxib or the celecoxib-vitamin E prodrug nanoparticles of Example 2 were administered orally. Approximately 0.5 mL of blood was collected from the orbital cavity at specified time points, and plasma was separated. The concentration of celecoxib in rat plasma was determined by HPLC.

[0050] After oral administration, the pharmacokinetic parameters were calculated:

[0051]

[0052] The results showed that after oral administration of celecoxib-vitamin E prodrug nanoparticles, the plasma drug concentration, AUC, and C2 were significantly improved. max All significantly increased, T after oral administration max The significant prolongation indicates that celecoxib-vitamin E prodrug nanoparticles significantly improve the bioavailability of celecoxib and have a sustained-release effect.

[0053] Example 8. Study on the tissue distribution of celecoxib-vitamin E prodrug nanoparticles

[0054] Thirty healthy male Kunming mice were randomly divided into four groups of five mice each. The mice were administered medication at a dose of 1.5 mg / kg. -1 The mice were administered celecoxib or celecoxib-vitamin E prodrug nanoparticles as described in Example 2 orally at specified time points. Blood was collected from the eyeballs and various organs (heart, liver, spleen, lung, kidney, and brain). Plasma was separated or homogenized with physiological saline, and the drug concentration of celecoxib in the mouse plasma and various organs was determined by HPLC. The drug concentration of the orally administered celecoxib prodrug nanoparticles in the heart, liver, spleen, lung, kidney, and brain was significantly higher than that of the orally administered celecoxib raw material.

[0055] Example 9. Study on the effect of celecoxib-vitamin E prodrug on apoptosis of colorectal cancer cells

[0056] Colorectal cancer cells (HCT-116) were cultured in an incubator containing 10% fetal bovine serum (FBS) at 37°C, 95% humidity, and 5% CO2. The cells grew in a monolayer and adhered to the culture wall. The medium was changed every 2-3 days, and the cells were passaged at a ratio of 1:3 after digestion with 0.25% (w / v) trypsin.

[0057] Weigh a certain amount of blank solution into a sterile EP tube, dissolve it in complete culture medium, sterilize by ultraviolet irradiation for 10 min, and inoculate the cells at a rate of 5 × 10⁻⁶. 4Cells were seeded at a density of 100 μL / mL in 96-well plates. Once cell growth was active, blank solution, CEL, CEL-SVE, and CEL-SVE NPs were added, and the cells were cultured for 24 h, 48 h, and 72 h, respectively, before detection. Detection method: 200 μL of 5 mg / mL thiazolyl blue solution (MTT) was added to each well, and the cells were cultured for another 4 h. The liquid was then discarded, and 150 μL of dimethyl sulfoxide (DMSO) was added to each well. The plates were gently shaken for 10 min, and the absorbance (OD) value at 570 nm was measured using an automated microplate reader. The MTT assay showed that celecoxib prodrug nanoparticles had strong cytotoxic effects on HCT-116 cancer cells and could inhibit cancer cell growth.

[0058] The in vitro cell proliferation inhibition rate of HCT-116 was calculated as follows:

[0059] .

Claims

1. A celecoxib-vitamin E prodrug, characterized in that: It is a celecoxib-vitamin E prodrug formed by linking celecoxib to a molecule of vitamin E with dithiodiacetic acid as the parent drug. The structural formula of the celecoxib-vitamin E prodrug is as follows: The celecoxib-vitamin E prodrug is prepared by electrostatic spraying to form celecoxib-vitamin E prodrug nanoparticles. The average particle size of the celecoxib-vitamin E prodrug nanoparticles is less than 400 nm, and they are uniformly distributed in a near-spherical shape.

2. A method for preparing the celecoxib-vitamin E prodrug of claim 1, characterized in that it Includes the following steps: Step 1: Dissolve 1g of dithionyldiacetic acid in 10-20mL of acetic anhydride and stir at room temperature for 3-4 hours. After the reaction is complete, evaporate to dryness by rotary evaporation. Dissolve the crude product in 15-40mL of dichloromethane, add 10-30mg of 4-dimethylaminopyridine and 500mg of vitamin E, and stir at room temperature for 2 hours. Purify the reaction solution by silica gel column chromatography. The eluent is petroleum ether-ethyl acetate-glacial acetic acid in a volume ratio of 95:5:0.1-80:20:0.1 to obtain the intermediate dithionyldiacetic acid-vitamin E. Step 2: Dissolve 500 mg of dithionyl diacetic acid-vitamin E in dichloromethane, add 100-400 mg of dicyclohexylcarbodiimide and 10-30 mg of 4-dimethylaminopyridine, stir at room temperature for 5 min, add 250 mg of celecoxib, stir at room temperature for 2 hours, purify the reaction solution with silica gel column chromatography, the eluent is dichloromethane-methanol, volume ratio of 100:0-90:10, to obtain a light yellow viscous semi-solid, which is celecoxib-vitamin E prodrug.

3. A method for preparing celecoxib-vitamin E prodrug nanoparticles based on electrostatic spraying as described in claim 1, characterized in that: The process involves dissolving celecoxib-vitamin E prodrug and a carrier together in dichloromethane and stirring until a clear, homogeneous solution is formed. This solution is then added to a 10mL syringe, which is attached to a microinjection pump. The injection speed is controlled by the microinjection pump, and the spray voltage and electrostatic spray distance are adjusted by regulating the high-voltage power supply. The electrostatic spray experiment is then initiated.

4. The application of the celecoxib-vitamin E prodrug nanoparticles as described in claim 1 in the preparation of nanomedicine delivery systems.

5. The use of the celecoxib-vitamin E prodrug nanoparticles according to claim 1 in the preparation of anti-colorectal cancer drugs.

6. The application of the celecoxib-vitamin E prodrug nanoparticles according to claim 5 in the preparation of anti-colorectal cancer drugs, characterized in that: The drug is an injectable medication.

7. The method for preparing celecoxib-vitamin E prodrug nanoparticles according to claim 3, characterized in that, The electrostatic spray voltage is 12–20 kV.

8. The method for preparing celecoxib-vitamin E prodrug nanoparticles according to claim 3, characterized in that, The electrostatic spray velocity is 0.02–0.08 mm / min.

9. The method for preparing celecoxib-vitamin E prodrug nanoparticles according to claim 3, characterized in that, The electrostatic spraying distance is 10-20cm.

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