A cyclodextrin inclusion compound or phospholipid complex of a traditional Chinese medicine extract of ligustrum robustum and a preparation method and application thereof

Abstract

The present application belongs to the technical field of ethnic medicine preparation and medicine delivery, and particularly relates to a cyclodextrin inclusion compound or phospholipid complex of a Lijia naked flower purple perilla extract, a preparation method and application thereof.

Description

Technical Field

[0001] This invention belongs to the field of ethnic medicine preparations and drug delivery technology, specifically relating to a cyclodextrin inclusion complex or phospholipid complex of the extract of the Li medicine *Callicarpa nudiflora*, its preparation method and application. Background Technology

[0002] Hainan Li medicine is a unique ethnic medicine system in my country, and *Callicarpa nudiflora* is a commonly used local Li medicinal resource in Hainan. *Callicarpa nudiflora* extract is rich in various types of flavonoids and phenolic active components, possessing excellent anti-inflammatory, soothing, and repairing activities, and has great development potential and application value in areas such as topical skin care and feminine hygiene. However, the flavonoid extract of Callicarpa nudiflora has significant shortcomings in formulation: the active flavonoid components have extremely poor water solubility and are rich in phenolic hydroxyl groups in their molecular structure, making them highly susceptible to oxidative degradation due to temperature, oxygen, and light. They are also prone to discoloration and inactivation during storage. The unmodified crude extract of Callicarpa nudiflora has poor flowability and weak solubility. When used directly in topical formulations and care products, it suffers from uneven dispersion of active ingredients, low bioavailability, and insufficient long-term storage stability, which greatly limits the industrial application of this Li medicinal resource. Cyclodextrin inclusion complexation and phospholipid complexation are mainstream methods for modifying active ingredients in pharmaceutical formulations, effectively improving the solubility and stability of natural extracts. Existing technologies have disclosed preparation processes for phospholipid complexes of total flavonoids from common plants such as Ginkgo biloba, Epimedium, and Crataegus pinnatifida. However, these processes are all designed for simple flavonoid systems with single components, similar polarities, and no significant component competition. In contrast, the extract of Callicarpa nudiflora is complex, containing various flavonoids and phenolic components with significantly different structures, polarities, and binding activities. Directly applying existing general phospholipid complexation and cyclodextrin inclusion complexation processes will result in a series of problems, including low inclusion / complexation rates, uneven binding of active components, dissociation and precipitation of small-molecule active ingredients during storage, and poor product stability. Currently, the industry lacks a refined modification process specifically tailored to the complex multi-component extract of Callicarpa nudiflora. The applicant's earlier application, "A Natural Transdermal Permeation-Enhancing Composition Based on Hainan Li Medicine and Its Application" (application number 2026106311990), provided a basic scheme for active ingredients with the ability to open stratum corneum channels. However, the permeation-enhancing active components in this composition still exist in their native form. In actual formulation processing and storage, there are still practical production problems such as poor formulation uniformity due to insufficient water solubility, easy oxidation and degradation of active ingredients in humid environments, and poor compatibility of some active components with aqueous matrices. The root cause of the above problems is that the physicochemical form of the native active ingredients has not been adapted to a refined carrier structure, and it is difficult to fundamentally solve the problem by simply adjusting the composition ratio. On this basis, this application further approaches the issue from the technical dimension of carrier modification. Through molecular-level modification using cyclodextrin inclusion or phospholipid complex, the aforementioned permeation-enhancing active ingredients are structurally upgraded to form modified intermediates that can be stably dispersed in aqueous / oil-based formulations and maintain long-term activity, providing upstream raw materials with better performance for subsequent liposome delivery and end products. Summary of the Invention

[0003] The purpose of this invention is to overcome the shortcomings of existing technologies, such as poor solubility, easy oxidation and degradation, weak storage stability, and poor formulation compatibility, of *Callicarpa nudiflora* extract. This invention provides a cyclodextrin inclusion complex or phospholipid complex of *Callicarpa nudiflora* extract, its preparation method, and its application. This invention designs two parallel modification schemes: one improves the water solubility and antioxidant stability of the extract through cyclodextrin inclusion, and the other improves the lipid solubility and formulation compatibility through phospholipid complexation. Both schemes target the same raw material, address the same technical pain points, and belong to the same inventive concept, allowing for flexible selection based on different product scenarios. To achieve the above objectives, the present invention adopts the following technical solution: This invention provides a cyclodextrin inclusion complex or phospholipid complex of *Callicarpa nudiflora* extract, wherein the active ingredient of *Callicarpa nudiflora* is *Callicarpa nudiflora* extract; the *Callicarpa nudiflora* extract is a standardized purified product with a total flavonoid content ≥50%, and is determined by UV-Vis spectrophotometry according to the General Chapter IV of the 2020 edition of the Chinese Pharmacopoeia, using rutin as a reference standard. The raw materials are strictly controlled for heavy metal and pesticide residue limits to ensure stable and controllable raw material quality. The cyclodextrin inclusion complex is prepared by compounding extract of Callicarpa nudiflora, modified cyclodextrin, and refined coconut oil; the modified cyclodextrin is selected from one or more of β-cyclodextrin, hydroxypropyl-β-cyclodextrin, and sulfobutyl ether-β-cyclodextrin, preferably hydroxypropyl-β-cyclodextrin, which has better water solubility and biocompatibility; the mass ratio of Callicarpa nudiflora extract to modified cyclodextrin is 1:1 to 1:20, preferably 1:5 to 1:18, and more preferably 1:10 to 1:16. The refined coconut oil conforms to the national standard GB / T 10496-2018, with a lauric acid content ≥40%, an acid value ≤0.5mg / g, and a peroxide value ≤5 meq / kg. The coconut oil exists in the system as micron-sized emulsion droplets, acting as a synergistic interfacial stabilizer. By improving the oil-water interface distribution and reducing the escape of active components, it stabilizes the inclusion system, significantly improving the inclusion rate and product stability. After freeze-drying, the tiny coconut oil droplets are uniformly embedded within the cyclodextrin powder matrix, resulting in a pale yellow, loose powder with no oil separation or agglomeration, exhibiting excellent powder properties. The phospholipid complex is composed of *Callicarpa nudiflora* extract and phospholipids, with the phospholipids selected from at least one of soybean phospholipids and egg yolk lecithin. The mass ratio of *Callicarpa nudiflora* extract to phospholipids is 1:2 to 1:8. For the complex multi-component system of *Callicarpa nudiflora*, different flavonoids and phenolic components exhibit competition for phospholipid binding sites, making efficient and uniform compounding impossible with conventional processes. This invention, through a precise feeding ratio, controlled reflux reaction, and gradient petroleum ether washing, effectively alleviates problems such as low compounding rate, active dissociation, and storage instability caused by competitive binding of multiple components. Unlike existing general phospholipid compounding processes for single flavonoid systems, this invention achieves efficient compounding of complex *Callicarpa nudiflora* flavonoid components, and the product's storage stability is significantly better than that of conventional process products and physical mixtures. Those skilled in the art should understand that the phospholipid complex described in this invention differs fundamentally from conventional phospholipid liposome delivery carriers and should not be confused with them. The phospholipid complex is a stoichiometric complex formed at the molecular level by the active pharmaceutical ingredient and phospholipid molecules through hydrogen bonds and hydrophobic interactions. Its composite morphology is a non-vesicular structure, not characterized by encapsulating an aqueous core, and does not possess the bilayer membrane-closed vesicle morphology of liposomes. Liposomes are closed vesicles formed by a phospholipid bilayer encapsulating an aqueous core. Their association with the active ingredient is mainly physical encapsulation or intermembrane distribution; a stoichiometric complex relationship is not necessarily present between the active ingredient and phospholipids. The two differ fundamentally in structural morphology, formation mechanism, intercomponent interactions, and final performance characteristics. The phospholipid complex of this invention is positioned for the **molecular morphology modification** of the active ingredient (improving lipophilicity and enhancing stratum corneum distribution behavior), rather than for use as a delivery carrier. Further vesicle-based delivery systems require additional vesicle treatment based on the complex of this invention; the two belong to different technical levels and product forms. This invention further provides a method for preparing the above-mentioned cyclodextrin inclusion complex, the specific steps of which are as follows: S01: Modified cyclodextrin and refined coconut oil are added to deionized water and sheared for 3-10 min using a high-shear homogenizer at 8000-15000 r / min to prepare a uniform and stable emulsion carrier dispersion system without layering within 30 min, ensuring uniform dispersion of coconut oil and playing a role in interface stabilization. S02: Mix the extract of Callicarpa nudiflora with ethanol at a material-to-liquid ratio of 1g:2mL to 1g:10mL to aid dissolution, and prepare a homogeneous active component solution after complete dissolution. S03: Slowly add the active component solution dropwise to the carrier dispersion system, and continuously stir at 400-800 r / min to ensure uniform dispersion of the active component; S04: Stir at 40-50℃ for 2-4 hours to incorporate. After the reaction is complete, filter with medium-speed qualitative filter paper to remove only trace amounts of insoluble impurities. Coconut oil droplets can pass through the filter paper completely without retention or loss. After freeze-drying the filtrate, a loose powdery cyclodextrin inclusion complex is obtained. This invention also provides a method for preparing the above-mentioned phospholipid complex, the specific steps of which are as follows: P01: Dissolve phospholipids in an anhydrous organic solvent and stir thoroughly to obtain a homogeneous phospholipid organic solution; P02: Dissolve the extract of Callicarpa nudiflora in the same anhydrous organic solvent system to prepare an active organic liquid; P03: Mix the active organic liquid with the phospholipid organic solution, reflux and stir at 50-60℃ for 2-4 hours to achieve molecular-level compounding; P04: The organic solvent was removed by vacuum distillation to obtain a solid crude product. This product was then washed three times with 5-10 times its mass of petroleum ether until no absorption peak of the free active component was detected by UV 254 nm, ensuring thorough elution of unrecombined free components. Simultaneously, process screening was used to minimize phospholipid dissolution loss. After drying, a highly stable phospholipid complex was obtained. The phosphomolybdic blue method was used to detect the phospholipid content of the product during preparation, verifying no significant phospholipid loss and ensuring accurate and reliable composite rate data. This invention also provides the application of the modified complex of the above-mentioned nude flower purple artichoke extract in the preparation of topical preparations and functional care products. The topical preparations include skin repair gels, soothing creams, etc., and the care products are preferably functional sanitary napkins. It has a wide range of applications and strong industrialization feasibility. Detailed Implementation Overview of detection methods Definition of total flavonoids in feed: Total flavonoids in feed = Mass of extract from naked flower purpureus × Measured total flavonoid content of the extract in this batch (UV-Vis method). The measured value of the batch shall prevail to ensure accurate calculation and traceability of data. Formula for calculating the inclusion rate of cyclodextrin: Inclusion rate (%) = (Mass of total flavonoids in the feed extract of Callicarpa nudiflora − Mass of free, unincluded flavonoids in the ultrafiltration permeate) ÷ Mass of total flavonoids in the feed extract of Callicarpa nudiflora × 100%; Detection method: The inclusion reaction filtrate was subjected to ultrafiltration centrifugation at 3000 Da, and the ultrafiltration permeate was taken. The content of free total flavonoids was determined by UV-Vis spectrophotometry according to the Chinese Pharmacopoeia with rutin as a reference standard, and the membrane adsorption blank error was deducted. The formula for calculating the phospholipid complexation rate is: Complexation rate (%) = (mass of total flavonoids in the extract of Callicarpa nudiflora - mass of free total flavonoids in the petroleum ether washing solution) ÷ mass of total flavonoids in the extract of Callicarpa nudiflora × 100%; the total flavonoids in the washing solution were also determined by UV-Vis method. Storage stability testing: Using luteolin as a characteristic marker, the active ingredient retention rate was calculated by HPLC external standard method; the residual amount of phospholipid complex was detected by phosphomolybdic blue colorimetric method to monitor process loss. Example 1: Cyclodextrin inclusion complex of Naked Flower Purple Callicarpa extract Raw material ratio: 3 parts of Naked Flower Purple Beauty Flavonoid Extract (total flavonoids ≥50%, UV-Vis method), 45 parts of Hydroxypropyl-β-Cyclodextrin, and 2 parts of Refined Coconut Oil. Preparation process: Hydroxypropyl-β-cyclodextrin was dissolved in deionized water, refined coconut oil was added, and homogenized at 10000 r / min for 6 min under high shear to obtain a stable emulsion system; the extract of Callicarpa nudiflora was slowly added dropwise to the emulsion system at a ratio of 1 g: 5 mL ethanol as a co-dissolving agent, and stirred at 600 r / min until homogeneous; the mixture was incorporated at 45℃ for 3 h, and impurities were removed by qualitative medium-speed filter paper filtration. The filtrate was freeze-dried to obtain a light yellow loose powder with no oil precipitation and no agglomeration. Test results: In three parallel experiments, the average inclusion rate was 89.3%, RSD=1.5%, indicating excellent process stability. Example 2 Phospholipid Complex from Naked Flower Artemisia capillaris Extract Raw material ratio: 4 parts of Naked Flower Purple Artemisia Flavonoid Extract, 16 parts of Soy Lecithin. Preparation process: Soybean lecithin and Naked Flower Purple Artemisia extract were dissolved in anhydrous ethanol, mixed and refluxed at 55°C for 3 hours; the solvent was removed under reduced pressure to obtain a solid product; the product was washed three times with 8 times its mass of petroleum ether until no UV characteristic absorption was observed at 254 nm, and then vacuum dried to obtain a phospholipid complex. Test results: Composite rate 86.1%; Phosphomolybdic blue method verification showed no significant washing loss of phospholipids; after 3 months of sealed storage at 25℃±2℃ and RH60%±5%, luteolin retention rate ≥95%, peroxide value ≤5meq / kg, with no rancidity or inactivation. Example 3: Topical gel formulation of cyclodextrin inclusion complex The cyclodextrin inclusion complex prepared in Example 1 was uniformly dispersed in a conventional aqueous gel matrix and stirred to obtain a skin-soothing and repairing topical gel with uniform dispersion of active components and excellent stability. Example 4: Phospholipid Complex Topical Gel Formulation The phospholipid complex prepared in Example 2 was dispersed in a gel matrix system and stirred at a constant speed until completely homogeneous to prepare a lipid-soluble, adaptable topical repair gel with good formulation compatibility. Example 5: Preparation of Functional Sanitary Napkins The composite of the present invention is mixed into the sanitary napkin surface fiber finishing liquid at a mass ratio of 0.1%-2%, and applied to the fiber surface layer by spraying, impregnation or roller coating. It is then dried and shaped at a low temperature below 60°C to obtain a functional sanitary napkin with the active ingredient of Callicarpa nudiflora. The entire process is carried out at a low temperature to ensure that the active ingredients do not degrade. Comparative Example 1: Coconut Oil-Free Cyclodextrin Inclusion Complex Compared to Example 1, no refined coconut oil was added, while all other raw materials and process parameters remained identical. Three parallel experiments showed an average inclusion rate of 72.2% and an RSD of 2.1%, significantly lower than Example 1, demonstrating that the interfacial synergistic stabilizing effect of coconut oil can effectively improve the inclusion effect. Comparative Example 2: Physical mixture of Naked Flower Purple Callicarpa extract and phospholipids Compared to Example 2, only simple physical mixing of raw materials was performed, without reflux composite reaction, and the rest of the processing methods were the same. The measured equivalent composite rate was 38.7%, which only came from physical adsorption and entrainment, without molecular composite effect; after 3 months of storage, the activity retention rate was only 67.2%, the peroxide value increased significantly, and the stability was extremely poor, proving the necessity of the proprietary composite process of this invention. Beneficial effects 1. This invention targets the complex multi-component flavonoid system of Callicarpa nudiflora and designs two exclusive modification schemes to overcome the technical bottleneck of poor adaptability of existing general processes. It effectively improves the solubility, powder properties and storage stability of the extract, and solves the core pain points of traditional Callicarpa nudiflora crude extract, such as easy oxidation, easy inactivation and difficulty in formulation. 2. The cyclodextrin inclusion system introduces refined coconut oil as an interface synergistic stabilizer, which can stabilize the inclusion system without the need for port blocking, significantly improves the inclusion rate, and produces a uniform powder of freeze-dried product without oil precipitation and agglomeration. The process is highly reproducible and has high batch stability. 3. The phospholipid complexing process specifically addresses the competitive binding problem of multiple components in Callicarpa nudiflora. Through precise proportioning, controlled reaction, and gradient washing, the entire process achieves high complexing rate and high storage stability. At the same time, it adds phospholipid content quality control measures, ensuring that the data is authentic and verifiable, which is different from simple physical mixing and existing general processes. 4. All process parameters, testing methods, and calculation formulas of this invention are complete and closed-loop, and the raw material quality control, process control, and finished product testing standards are clear. Those skilled in the art can completely replicate it, and there is no defect of insufficient disclosure. 5. The modified product of this invention can be adapted to various industrialization scenarios such as topical gels and functional care products, broadening the high-value application path of Hainan Li medicinal herb Callicarpa nudiflora resources and possessing extremely high industrialization value.

Claims

1. A cyclodextrin inclusion complex or phospholipid complex of an extract of *Callicarpa nudiflora*, characterized in that, The active ingredient of the Li medicine is extract of Callicarpa nudiflora; The extract of Callicarpa nudiflora is a standardized purified product with a total flavonoid content of ≥50%. Rutin was used as a reference standard, and the content was determined by UV-Vis spectrophotometry according to the General Chapter IV of the 2020 edition of the Chinese Pharmacopoeia. The raw materials for the extract were controlled for heavy metal and pesticide residue limits according to the General Chapter IV of the Chinese Pharmacopoeia. The cyclodextrin inclusion complex comprises Callicarpa nudiflora extract, modified cyclodextrin, and refined coconut oil; the mass ratio of Callicarpa nudiflora extract to modified cyclodextrin is 1:1 to 1:20, preferably 1:5 to 1:18, and more preferably 1:10 to 1:

16. The phospholipid complex comprises extract of Callicarpa nudiflora and phospholipids, with a mass ratio of extract of Callicarpa nudiflora to phospholipids of 1:2 to 1:

8.

2. The cyclodextrin inclusion complex or phospholipid complex according to claim 1, characterized in that, The modified cyclodextrin is selected from one or more of β-cyclodextrin, hydroxypropyl-β-cyclodextrin, and sulfobutyl ether-β-cyclodextrin, with hydroxypropyl-β-cyclodextrin being preferred; The refined coconut oil is refined coconut oil that conforms to the national standard GB / T 10496-2018, with a lauric acid content ≥40%, an acid value ≤0.5 mg / g, and a peroxide value ≤5 meq / kg; the refined coconut oil is used as a synergistic stabilizer in the cyclodextrin inclusion complex system.

3. The cyclodextrin inclusion complex or phospholipid complex according to claim 1, characterized in that, The phospholipid is selected from at least one of soybean phospholipids and egg yolk lecithin.

4. The method for preparing the cyclodextrin inclusion complex according to any one of claims 1-3, characterized in that, Includes the following steps: S01: Modified cyclodextrin and refined coconut oil are added to an aqueous solvent and treated with a high-shear homogenizer at 8000-15000 r / min for 3-10 min to obtain a uniform emulsion carrier dispersion system with no obvious stratification within 30 min. S02: Dissolve the extract of Callicarpa nudiflora with ethanol, with the ratio of the extract to ethanol being 1g:2mL to 1g:10mL, and prepare an active component solution. S03: Slowly add the active component solution dropwise to the carrier dispersion system, stirring at 400-800 r / min, and continue stirring; S04: The inclusion reaction was carried out under constant temperature stirring. After the reaction was completed, the trace amounts of insoluble impurities were removed by qualitative medium-speed filter paper to obtain the filtrate. The pore size of the filter paper was larger than the average particle size of the coconut oil droplets. All the emulsified coconut oil droplets passed through the filter paper with the filtrate, and the filtration operation did not cause the retention or loss of coconut oil components. The filtrate was freeze-dried to obtain the cyclodextrin inclusion complex powder. The obtained powder was a light yellow loose powder with no obvious oil precipitation or agglomeration.

5. A method for preparing the phospholipid complex according to any one of claims 1-3, characterized in that, Includes the following steps: P01: Dissolve phospholipids in an anhydrous organic solvent and stir to obtain an organic phospholipid solution; P02: Dissolve the extract of Callicarpa nudiflora in the same organic solvent system to obtain an active organic liquid; P03: Add the active organic liquid to the phospholipid organic solution and reflux with stirring to react; P04: Remove organic solvent by vacuum distillation to obtain solid product; The solid product was washed with petroleum ether at a rate of 5-10 times the mass of the solid product each time, and the washing was repeated three times until the washing solution showed no characteristic absorption peak under 254 nm UV light. The product was then dried to obtain the phospholipid complex. The washing conditions were selected through process screening to minimize the dissolution and loss of phospholipids while ensuring sufficient elution of free active ingredients. After washing, the phospholipid content of the product was tested to confirm that the complex did not experience significant phospholipid loss.

6. The preparation method according to claim 4, characterized in that, The inclusion reaction temperature in SO4 is 40-50℃, and the reaction time is 2-4 h.

7. The preparation method according to claim 5, characterized in that, The PO3 reflux reaction temperature is 50-60℃, and the reaction time is 2-4 h.

8. The use of the cyclodextrin inclusion complex or phospholipid complex of the extract of Callicarpa nudiflora according to any one of claims 1-3 in the preparation of topical preparations and care products.

9. The application according to claim 8, characterized in that, The nursing product in question is a functional sanitary napkin.