CN-121987807-A - Dendrobium officinale exosome delivery system loaded with active oxygen responsive pterostilbene, and preparation method and application thereof

Abstract

The invention provides an active oxygen-loaded responsive pterostilbene dendrobium candidum exosome delivery system, and a preparation method and application thereof, and belongs to the technical field of biological medicines, wherein the preparation method of the dendrobium candidum exosome delivery system comprises the following steps of 1) mixing 4- (hydroxymethyl) phenylboronic acid pinacol ester with N, N' -carbonyl diimidazole, and obtaining a phenylboronic acid ester derivative with a cleavable H 2 O 2 through a first reaction; 2) mixing pterostilbene with the phenylboronate derivative, performing a second reaction to obtain borate-pterostilbene, and 3) mixing and co-incubating an dendrobium candidum exosome with the borate-pterostilbene to obtain an active oxygen responsive pterostilbene-loaded dendrobium candidum exosome. According to the invention, the pterostilbene is modified by the boric acid ester bond, the ROS sensitive prodrug (Bor-PTE) is constructed, the Bor-PTE is entrapped by utilizing the natural carrier characteristic of the dendrobium candidum exosome, and the problems of bioavailability and targeting of the pterostilbene are cooperatively solved.

Inventors

• CHEN YUEWEN
• CHEN YONGQIANG
• FU JINGJING
• CAI WENQIANG
• YUAN YANWEI
• LONG YONGJIE

Assignees

• 浙江工商大学

Dates

Publication Date

20260508

Application Date

20260402

Claims (10)

1. 1. The preparation method of the dendrobium candidum exosome delivery system loaded with the active oxygen responsive pterostilbene is characterized by comprising the following steps of: 1) Mixing 4- (hydroxymethyl) phenylboronic acid pinacol ester and N, N' -carbonyl diimidazole, and carrying out a first reaction to obtain a phenylboronic acid ester derivative CDB with a cleavable H 2 O 2 ; 2) Mixing pterostilbene with the phenylboronate derivative CDB obtained in the step 1), and carrying out a second reaction to obtain borate-pterostilbene Bor-PTE; 3) Mixing and co-incubating the dendrobium candidum exosome with the borate-pterostilbene Bor-PTE in the step 2) to obtain the dendrobium candidum exosome DEV@bor-PTE loaded with active oxygen responsive pterostilbene.
2. 2. The preparation method according to claim 1, wherein the molar ratio of the 4- (hydroxymethyl) phenylboronic acid pinacol ester to the N, N' -carbonyldiimidazole in the step 1) is 1:1-3, and the time of the first reaction is 4-16 h.
3. 3. The preparation method according to claim 1, wherein in the step 2), the molar ratio of pterostilbene to phenylboronate derivative CDB is 1:1-5, and the second reaction time is 24-96 h.
4. 4. The preparation method of claim 1 or 3, wherein after the second reaction in the step 2) is finished, the method further comprises a step of purifying, wherein silica gel column chromatography is adopted in the purification, petroleum ether and ethyl acetate are used as eluent, and the volume ratio of the petroleum ether to the ethyl acetate is 6:1-2.
5. 5. The preparation method of the dendrobium candidum according to claim 1, wherein the mass ratio of the dendrobium candidum exosome to the borate-pterostilbene Bor-PTE in the step 3) is (1-3) (3-1), the co-incubation temperature is 20-40 ℃, and the co-incubation time is 1-3 h.
6. 6. The method according to claim 5, wherein after the co-incubation in step 3) is completed, ultra-high speed centrifugation is used for purification, the centrifugal force of the ultra-high speed centrifugation is 100000-150000 g, and the time of the ultra-high speed centrifugation is 1-3 h.
7. 7. The preparation method according to claim 1, wherein the dendrobium candidum exosomes are obtained by extracting by the following method: s1) mixing dendrobium candidum with a buffer solution, juicing and filtering to obtain filtrate; S2) carrying out differential centrifugation on the filtrate, collecting supernatant, carrying out ultracentrifugation on the supernatant, and collecting precipitate and re-suspending to obtain the dendrobium candidum extracellular fluid solution.
8. 8. The preparation method of claim 7, wherein the buffer solution is PBS or physiological saline, and the ratio of the dendrobium candidum to the buffer solution is 1g (1-3) ml.
9. 9. The dendrobium candidum exosome delivery system loaded with the active oxygen responsive pterostilbene is characterized by comprising an dendrobium candidum exosome and an active oxygen responsive pterostilbene prodrug, wherein the active oxygen responsive pterostilbene prodrug is embedded in the dendrobium candidum exosome, and the active oxygen responsive pterostilbene prodrug is borate modified pterostilbene, namely borate-pterostilbene Bor-PTE; The mass ratio of the dendrobium candidum exosome to the borate-pterostilbene Bor-PTE is (1-3) (3-1); The structural formula of the borate-pterostilbene Bor-PTE is shown in formula II: Formula II.
10. 10. The application of the dendrobium candidum exosome delivery system prepared by the preparation method of any one of claims 1-8 or the dendrobium candidum exosome delivery system of claim 9 in preparing targeted anti-inflammatory drugs, drugs for treating cardiovascular diseases or drugs for treating metabolic diseases.

Description

Dendrobium officinale exosome delivery system loaded with active oxygen responsive pterostilbene, and preparation method and application thereof Technical Field The invention belongs to the technical field of biological medicines, and particularly relates to an active oxygen responsive pterostilbene-loaded dendrobium candidum exosome delivery system, and a preparation method and application thereof. Background Pterostilbene (Pterostilbene, PT for short) is a stilbene polyphenol compound naturally existing in plants such as blueberries, grapes and pterocarpus santalinus, and has obvious pharmacological activity and application potential in the fields of antioxidation, anti-aging, anti-tumor, immunoregulation and the like by virtue of the unique molecular structure. Compared with resveratrol with similar structure, pterostilbene has more excellent fat solubility, bioavailability and in vivo stability, and can penetrate cell membrane more efficiently and play a role in vivo. Pharmacological mechanism researches show that pterostilbene can exert biological activity through multiple ways, namely, on one hand, through directly removing oxidative stress substances such as Reactive Oxygen Species (ROS) and hydroxyl free radicals in vivo and simultaneously up-regulating the expression and activity of endogenous antioxidant enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and the like to maintain the oxidation-antioxidant balance of the organism, and on the other hand, through regulating key inflammation and signal paths such as NF-kappa B, MAPK, PI K/Akt and the like, release of pro-inflammatory factors (TNF-alpha, IL-6 and IL-1 beta) is inhibited, so that unique advantages are shown in prevention and treatment of cardiovascular diseases (such as atherosclerosis and hypertension), metabolic diseases (such as type 2 diabetes mellitus and obesity) and various tumors (lung cancer, breast cancer and liver cancer), and the pterygoid process becomes a research hot spot in the field of natural drug research in recent years. Although pterostilbene has definite pharmacological value, clinical transformation and wide application still face two core bottlenecks, namely low bioavailability, high inherent fat solubility and extremely poor water solubility even though compared with resveratrol, so that the pterostilbene has incomplete gastrointestinal absorption after oral administration, obvious first pass effect and difficult in-vivo circulating concentration to reach an effective treatment threshold, and insufficient targeting, and the traditional administration mode has the advantages of non-specific in-vivo distribution of medicines, low effective enrichment concentration of focus parts, and potential untargeted toxic and side effects on normal tissues, thereby seriously restricting the widening of the treatment window and the exertion of clinical curative effects. To solve the above problems, various drug delivery systems have been developed in the prior art for improving the delivery efficiency of pterostilbene, including liposomes, polymer nanoparticles, microcapsules, cyclodextrin inclusion compounds, and the like. However, the delivery systems still have a plurality of inherent defects that partial carriers (such as synthetic polymer nanoparticles) are poor in biocompatibility, in vivo immune reaction or accumulated toxicity is easy to cause, secondary damage is likely to occur to degradation products, the preparation process of most of the delivery systems is complex, the steps are complicated, the problems of large organic solvent consumption, severe reaction conditions and the like are involved, the large-scale production and clinical transformation are not facilitated, and more importantly, the existing systems are limited in targeting efficiency, depend on passive targeting, have insufficient active targeting capability, are difficult to realize accurate enrichment and controllable release of medicines at focus positions, and cannot fundamentally solve the targeting problem of pterostilbene. Under pathological conditions such as tumors and inflammations, the focus tissue is enhanced due to abnormal metabolism and oxidative stress, the level of the ROS (including hydrogen peroxide, superoxide anions, hydroxyl radicals and the like) in the focus tissue is obviously higher than that of normal tissues, and the characteristic of the pathological microenvironment provides important targets and opportunities for designing an intelligent response type drug delivery system. The ROS responsive drug delivery system can construct a prodrug-carrier integrated structure by introducing ROS sensitive groups (such as thioether bonds, boric acid ester bonds, acetal/ketone bonds and the like), the structure has good stability in a low ROS environment of normal tissues, the drug activity is inhibited, and after entering a tumor or inflammatory focus area, the sensitive groups are subjected to abnormal elevat