Macerate-loaded polymeric nanoparticle system and in-situ gel phase formulation

Abstract

The invention relates to a two-component topical composite system for the treatment of sensitive tissues such as wounds and burns. It becomes fluid upon prior mechanical stimulation (shearing) and acquires a structure after application, exhibiting a residual gel / film behavior on the skin. The characteristic feature of the invention is that the stability, persistence, and efficacy of the plant-based active ingredients are increased by separately positioning the biodegradable polymeric nanoparticle phase, carrying the lipophilic fraction of Hypericum perforatum (St. John's wort) macerate, and the in-situ gel phase within the packaging. This eliminates problems such as phase separation, runoff, and poor penetration seen in conventional topical formulations.

Description

1 TARIFF Macerate-loaded polymeric nanoparticle system and in-situ gel phase formulation Technical Area 5 The invention is intended for use in the treatment of sensitive tissues such as wounds and burns, prior to use. It becomes fluid upon mechanical stimulation (shearing / applying) and gains structure after application, then adheres to the skin. It relates to a two-component topical composite system exhibiting residual gel / film behavior. The invention specifically aims to increase the stability, persistence, and efficacy of plant-based active ingredients on the skin, 10 Phase separation, runoff, and poor penetration seen in traditional topical formulations To address these problems, Hypericum is individually packaged within the container. Biodegradable polymeric containing lipophilic fraction of St. John's wort perforatum macerate This relates to a topical composite system containing a nanoparticle phase and an in-situ gel phase. State of the Art The integrity of the skin, known as the largest organ of our body, is affected by physical and chemical processes. Wounds and burns can occur due to biological and environmental factors. In known techniques... to accelerate tissue regeneration, reduce inflammation and provide antimicrobial protection 20 topical pharmaceuticals containing various plant extracts, essential oils and natural polymers for this purpose or herbal wound and burn creams consisting of dermocosmetic compositions It is used. Common ingredients in wound and burn creams include Aloe vera, Calendula officinalis, Centella asiatica, 25 Hypericum perforatum, Lavandula angustifolia, Nigella sativa, propolis, olive leaf from components such as extract and pine resin that support wound healing These products generally contain flavonoids to increase antioxidant capacity. Plant fractions containing phenolic compounds, terpenoids, saponins, and alkaloids; moisture retention. and for barrier purposes, beeswax, shea butter, coconut oil, jojoba oil and 30 Carriers such as glycerol; carbomer, xanthan gum, to provide viscosity and stability, It is also used to reduce the risk of infection using natural herbal ingredients. In addition to their antimicrobial effects, zinc oxide, silver ions, honey derivatives, or propolis Synergistic formulations are created with fractions. However, known formulations... A significant portion of the problem is the decrease in the stability of heat-sensitive plant active ingredients, requiring controlled release. 35 inability to achieve this, limited skin penetration, oxidative degradation, active ingredients in the product the amount of the substance decreases over time, leaving an oily layer on the skin surface or insufficient 2 It has problems such as not being able to reach the required level of antimicrobial efficacy, and therefore the current The technique maintains the stability of the active ingredient in multi-component herbal systems, delivered to subcutaneous layers. Effective penetration, long-lasting and controlled release, resistant to oxidative degradation. novel compositions that are resistant and show clinically significant wound / burn healing efficacy It reveals the need that is felt. 5 Such as Hypericum perforatum (St. John's wort) macerate, which is frequently used in the creams in question. Traditional methods are used in the production and integration of lipophilic plant-based ingredients into topical products. There are serious technical problems stemming from (conventional) methods. Traditional The production of St. John's wort macerate involves storing plant parts in carrier oils for extended periods (e.g., 10°C). It is based on being left in the sun for 40 days. Conventional on an industrial scale. Extraction techniques, however, are characterized by long processing times, high solvent consumption, low selectivity, and thermal degradation. It has disadvantages such as degradation (thermal degradation). This situation affects hypericin and production of bioactive components such as hyperforin, which are extremely sensitive to heat / light. This causes it to undergo oxidative degradation during this stage. Obtained by conventional methods 15 Adding these lipophilic macerates directly into classic topical gels or creams, Over time, phase separation occurs within the matrix, affecting the sensory properties (smell, color) of the product. This leads to deterioration and loss of viscosity of the gel matrix. The formula of the active ingredients... The inability to maintain a homogeneous and stable environment severely restricts shelf life. Traditional Phytochemicals present in free form in formulations (ointments, creams, classic gels) are lipid cells 20 They cannot penetrate the membranes of the skin barrier (stratum corneum) due to their inability or poor solubility. They have difficulty crossing the barrier and their intratissue absorption remains quite low. In addition to all this, products that will be applied to sensitive tissues such as wounds and burns Traditional solid gels cause high mechanical friction (resistance) when applied, causing pain to the patient. In contrast, classic oily / liquid products run off quickly from the target area, providing the necessary results. It fails to ensure therapeutic contact time and dose standardization. Consequently, due to the aforementioned drawbacks and shortcomings, the relevant technical... The need for innovation in the field has emerged. 30 Purpose of the Invention The present invention meets the aforementioned requirements and eliminates all disadvantages. a macerate-loaded polymeric nanoparticle system and in- that remove and bring some additional advantages It relates to in situ gel phase formulation. 35 3 The purpose of the invention is to create a device used in the treatment of delicate tissues such as wounds and burns. It becomes fluid with mechanical stimulation (shearing / applying) before use, and after application... A two-component topical product that forms a structure and exhibits a residual gel / film behavior on the skin. The goal is to develop a composite system. The purpose of the invention is to create a container for Hypericum perforatum (yellow) bacteria, individually positioned within a package. biodegradable polymeric nanoparticle phase carrying lipophilic fraction of St. John's wort macerate The goal is to develop a topical composite system containing an in-situ gel phase. The aim of the invention is to increase the stability, persistence, and efficacy of plant-based active ingredients on the skin. 10 The aim of the invention is to address phase separation, runoff, and low viscosity issues observed in traditional topical formulations. The goal is to resolve issues such as penetration. One aim of the invention is the standardization of the macerate with a filter cascade, nanoparticle formulation 15 through design space and quality control of the gel phase via pH / viscosity / rheuology window The goal is to reduce discrimination and performance variability. To achieve the purposes described above, the invention is designed to treat sensitive tissues such as wounds and burns. To increase the stability, persistence, and effectiveness of herbal active ingredients used in treatment, 20 Phase separation, runoff, and poor penetration seen in traditional topical formulations To solve the problems, it packages its two components separately. positioned and mentioned components with prior mechanical stimulation (shearing / drifting) It becomes fluid and acquires structure after application, exhibiting a gel / film behavior on the skin. It is a topical composite system exhibiting the following characteristic: maceration of the mentioned components (30 / 40 / 50 25 mg), poly(D,L-lactide-co-glycolide) (200 / 300 / 500 mg) and poly(vinyl alcohol) (0.5 / 1.0 / 2.0% m / v) macerate-loaded polymeric nanoparticle suspension (phase-1) defined by its levels and It is a mechanically stimulated in-situ gel phase (phase-2) based on cross-linked poly(acrylic acid). To achieve the purposes described above, the invention is intended for the treatment of sensitive tissues such as wounds and burns. To increase the stability, persistence, and effectiveness of herbal active ingredients used in treatment. Phase separation, runoff, and poor penetration seen in traditional topical formulations To solve the problems, it packages its two components separately. positioned and mentioned components with prior mechanical stimulation (shearing / drifting) It becomes fluid and acquires structure after application, exhibiting a gel / film behavior on the skin. 35 It is a production method of a topical composite system that exhibits the following characteristics: 4  To prepare a macerate-loaded polymeric nanoparticle suspension (phase-1), Dried H. perforatum material is transferred to the olive oil or black cumin oil phase. At ±2 °C, in a dark environment, for 10–14 days, once a day for 5 minutes (300–500 rpm) stirring and letting it stand, and then filtering the resulting macerate,  To prepare the organic phase, a biodegradable polymer is mixed with an organic solvent or solvent 5 Preparing the polymer solution by dissolving it in the mixture,  Addition of H. perforatum macerate to the polymer solution,  To prepare the aqueous phase, a stabilizer is dissolved in distilled / deionized water. to be solved,  The resulting aqueous phase is partially mixed with ethyl acetate to improve emulsification stability. or being completely satisfied,  Adding the organic phase containing the macerate dropwise to the aqueous phase containing PVA, and Emulsification is achieved by mixing the aqueous phase at a range of 400–10,000 rpm during addition. obtaining,  The emulsion created is designed to reduce particle size and narrow the dispersion. probe sonication,  Removal of organic solvent after emulsification of nanoparticles obtaining,  To prepare the in-situ gel phase (phase-2), carbomer and optional aloe vera were distilled in water. dissolved, mixed and homogenized, and then immersed in 0.1 M triethanolamine at pH 20. setting the value to the range of 6.5–7.5,  The resulting gel should be left for 2–24 hours to allow bubbles to dissipate and the network structure to stabilize. being left to stand for a certain period of time (preferably 12 hours),  Phase-1 is added to phase-2 by mixing at 20–25 °C and 300–600 rpm for 3–8 minutes. It includes the steps involved in the process. The structural and characteristic features and all the advantages of the invention are described in detail below. This will make it clearer, and therefore the evaluation will also be based on this detail. This should be done taking the explanation into consideration. 30 Figures that will help understand the invention. Figure 1: SEM image of the nanocarrier obtained within the scope of the invention. Figure 2: Histopathological appearance of wound healing in the skin defect area according to groups. 35 Detailed Description of the Invention This detailed explanation describes a macerate-loaded polymeric nanoparticle system and an in-situ gel phase. the formulation is solely for the purpose of better understanding the subject and has no limiting effects. It is explained in a way that will not create a problem. The invention is intended for use in the treatment of sensitive tissues such as wounds and burns, with a prior examination of 5 years. It becomes fluid upon mechanical stimulation (shearing / applying) and gains structure after application, then adheres to the skin. It relates to a two-component topical composite system exhibiting residual gel / film behavior. The characteristic of the invention is the lipophilic fraction of Hypericum perforatum (St. John's wort) macerate. The biodegradable polymeric nanoparticle phase and the in-situ gel phase are packaged separately. By positioning it correctly, the stability, persistence, and effectiveness of the plant-based active ingredients on the skin are increased by 10%. increasing the risk of phase separation, runoff, and low viscosity seen in traditional topical formulations. The goal is to resolve problems such as penetration. The macerate-loaded polymeric nanoparticle suspension (phase-1) obtained within the scope of the invention, It is based on poly(D,L-lactide-co-glycolide) (50:50; acid-end grade) and has aqueous phase stabilization of 15 It is made with poly(vinyl alcohol) (87–90% hydrolyzed). Pre-formulation design space; macerate (30 / 40 / 50 mg), poly(D,L-lactide-co-glycolide) (200 / 300 / 500 mg) and poly(vinyl alcohol) (0.5 / 1.0 / 2.0%) It is defined by m / v) levels. The mechanically-stimulated in-situ gel phase (phase-2) is cross-linked. It is poly(acrylic acid) based and is defined in two variants (Type-A / Type-B). Type-A: higher structure / resistance (poly(acrylic acid) 0.10–0.90% w / w; optional aloe 0.1–2.5%) w / w) Type-B: more fluid application profile (poly(acrylic acid) 0.10–0.45% w / w; optional aloe 0.1– 2.0% w / w) In both variants, neutralization was performed with a 0.1–1 M triethanolamine solution at a pH of 6.5–7.5. The process is carried out as follows: pre-mixing 10 minutes, homogenization after nanocarrier addition 1000 It is defined as 20 minutes at rpm. The packaging and application scenario is as follows: Phases are kept separate in the packaging; 30 During use, Phase-1:Phase-2 1:1–1:4 (w / w) ratio, 20–25 °C temperature, 300–600 rpm The mixture is combined by stirring at high speed for 3–8 minutes. The final pH target is 6.8–7.4, the final viscosity target is... (100 s⁻¹) is the 15,600–17,800 cP band. Production of the topical composite system that is the subject of the invention: 35 To prepare a macerate-loaded polymeric nanoparticle suspension (phase-1), firstly... Dried H. perforatum material is mixed with olive oil or black cumin oil in a 1:8 ratio. 6 and at 25 ±2 °C, in a dark environment, for 10–14 days, once a day for 5 minutes (300–500 rpm) It is stirred and left to stand. Then the resulting macerate is pre-filtered at 250–500 μm, and then filtered at 5 μm. pre-filter, 0.45 μm cellulose nitrate and, if necessary, 0.20 / 0.45 μm hydrophilic PTFE sequence. It is filtered. This invention describes the use of polymeric nanoparticles containing H. perforatum macerate / extract in an oil-in-water 5-degree process. The invention relates to the production of (O / W) emulsification-based products. The method described in the invention is: efficient loading of the macerate onto the polymer matrix, particle size distribution It is defined in a way that ensures control and repeatability of the production process. Preparation of the organic phase: 10 Polymer solution: A biodegradable polymer (preferably PLGA) in a suitable organic solvent or It dissolves in the solvent mixture. The polymer concentration is 0.01–1.00% (w / v) relative to the total organic phase. It is selected within the range; preferably 0.05–0.50% (w / h). Solvent selection: The organic phase solvent can be alone or in a mixture; for example, 15 A chloroform / ethanol mixture can be used. The organic phase volume is 0.5–10 mL depending on the application. It can be selected within a range. Maceration loading: H. perforatum macerate / extract, polymer according to target loading level. It is added to the solution. The macerate amount can be selected, for example, in the range of 10–100 mg. 20 The macerate / polymer ratio and macerate concentration affect the loading amount and physico- These are adjustable parameters that affect chemical properties. Preparation of the aqueous phase (stabilization and pre-saturation): Stabilizer solution: Aqueous phase; a stabilizer (preferably PVA) in distilled / deionized water 25 It is prepared by dissolving. Stabilizer concentration is in the range of 0.1%–1.0% (w / v); preferably 0.2%–0.7%. The aqueous phase volume is in the range of (a / h). The aqueous phase volume can be selected in the range of 10–1000 mL depending on the process scale. Pre-saturation with ethyl acetate (optional but preferred): Aqueous phase improves emulsification stability. It is partially or completely saturated with ethyl acetate to increase emulsification. This step is part of the emulsification process. during which the organic phase is more stably dispersed within the aqueous phase and the droplet / particle 30 It is applied in a way that will contribute to a more controlled formation. Emulsification (formation of O / W emulsion): Addition by drip: The organic phase containing the macerate is added dropwise to the aqueous phase containing PVA. During addition, the aqueous phase is mixed at a speed of 400–10,000 rpm; preferably 800–6,000 rpm. It is in the rpm range. 7 Addition rate and sequence control: Addition rate of the organic phase, emulsion droplet size. It is selected in a way that will affect its distribution. Additional controls are used to prevent local overconcentration. It is done. Energy application (thinning with probe sonication): 5 Probe sonication: The resulting emulsion is used to reduce particle size and narrow the dispersion. For this purpose, the probe is subjected to sonication. The sonication duration is between 1 and 10 minutes in total; Ideally, it should be between 2 and 6 minutes. Amplitude and conversion: Sonication can be applied in the amplitude range of 10%–90%. Sonication, each of which It can be run in 2–10 cycles lasting 1–2 minutes each, with 10–60 seconds between cycles. Waiting intervals can be specified. Heat control: To control heating during sonication, the emulsion should preferably be ice. It is kept in a cooling bath or equivalent arrangement. Solvent removal and nanoparticle solidification: 15 Removal of organic solvent: Organic solvent after emulsification; • Evaporation under vacuum,  rotary evaporation,  freezing at low temperatures (e.g., -80 °C) and / or  lyophilization 20 It is removed from the environment by one or more of these methods. As a result of this process, PLGA The matrix solidifies and PLGA / PVA nanoparticles are formed. Purification and final product preparation (optional but recommended for industrial use): Separation / washing: Nanoparticles can be collected by centrifugation; the supernatant is removed and the particles are separated by 25°C. It can be washed with water / a suitable cotton pad. The washing cycle may be 1-3 times. Drying and storage: The final product can be stored in suspension form or frozen. It can be obtained as a dry powder by lyophilization. Particle integrity must be preserved during lyophilization. The use of suitable cryoprotectants is optional. Critical quality attributes (CQA) and control approach (strongly defended in the patent): The resulting nanoparticles were characterized by their mean size, size distribution (PDI), and surface charge (zeta). It is characterized by criteria such as potential), maceration loading efficiency and redistributability. Process parameters (mixing speed, sonication amplitude / duration, solvent removal) By establishing a relationship between these quality attributes and the conditions (of production), repeatable production is targeted. 35 Table 1 shows the pre-formulation design space for phase 1 (macerate type / carrier oil variant). and / or level bands are shown. 8 Table 1: Pre-formulation design space for Phase 1 Component Level-1 Level-2 Level-3 H. perforatum macerate (Olea europaea carrier) mg 40 mg 50 mg H. perforatum macerate (Nigella sativa carrier) mg 40 mg 50 mg PLGA 50:50 (acidic end grade) 200 mg 300 mg 500 mg Stabilizer (PVA, 87-90% hydrolyzed) 0.5% (m / v) 1.0% (m / v) 2.0% (m / v) To prepare the in-situ gel phase (phase-2), carbomer and optional aloe vera were distilled in water. It is dissolved, homogenized by stirring for 10 minutes. Subsequently, it is mixed with 0.1 M triethanolamine at pH 6.5– It is adjusted to the 7.5 range. Homogenization at 1000 rpm for 20 minutes after nanocarrier addition. 5 is applied. Table 2 shows the composition bands and target pH of the gel phase variants (Type-A / Type-B). The data has been shared. Table 2: Composition bands and target pH of gel phase variants (Type-A / Type-B). Parameter Type-A (high structure / film) Type-B (more fluid) Carbomer (cross-linked poly(acrylic) acid)) 0.30-0.60% (w / w) 0.20-0.45% (w / w) Aloe vera (optional) 0.5-1.5% (w / w) 0.5-1.0% (w / w) Neutralizer (triethanolamine solution) 0.1 M; pH 6.5-7.5 0.1 M; pH 6.5-7.5 Mixing (pre-mixing) 10 min 10 min Homogenization (nanocarrier) post) 1000 rpm, 20 min 1000 rpm, 20 min Carbopol pre-dispersion (pre-hydration): Carbopol 934 shall be present in the total formulation at a concentration of 0.20–3.00% (w / w), It is added to distilled water at room temperature. The mixture is heated at a speed of 300–800 rpm. Hydration of the polymer for at least 30–120 minutes while stirring (preferably at 400 rpm). This is ensured. In this step, to reduce clumping, Carbopol is sprinkled onto the surface gradually for 15 minutes. is added. Preparation of Aloe Vera fraction: Aloe vera powder should be in the range of 0.10–1.00% (w / w) of the total formulation. (preferably 0.25–0.75%) It is pre-wetted with water in a separate container and a homogeneous suspension is obtained. 20 Mix at 200–600 rpm until aloe suspension is mixed with carbopol dispersion. It is added slowly. 9 Preparation of the nanoparticle phase and its integration into the gel matrix: Active ingredient-loaded nanoparticles, for example as a stock suspension at a concentration of 40 mg / mL. The nanoparticle suspension is prepared in the final gel formulation at a concentration of 1–20% (w / v). It is added to the Carbopol / Aloe dispersion (preferably at 10% w / v). During this addition process... The mixture is kept under stirring at 300–600 rpm and the nanoparticles are formed in the gel matrix. Homogeneous distribution is ensured. Initiating gelation by gradually adjusting the pH: Gelation is achieved with a triethanolamine (TEA) solution prepared in the range of 0.05–0.2 M (preferably 0.1 M). This is done by adjusting the pH. 10 pH adjustment is done in two stages to support the homogeneous establishment of the gel network:  First, the pH is brought to the range of 5.0–5.5,  Next, the pH is adjusted to the range of 6.5–7.5 (preferably 6.8–7.2). During pH adjustment, the mixture is stirred continuously to avoid local over-neutralization. TEA solution is added drop by drop. 15 Maturation (network stabilization): The resulting gel is left for 2–24 hours to allow bubbles to dissipate and the network structure to stabilize. It is left to stand for an interval (preferably 12 hours). If necessary, short-term de-aeration under vacuum is performed. It is applied. 20 Characterization / acceptance criteria (example): The gel was evaluated based on its appearance (absence of phase separation), pH (6.5–7.5), homogeneity, viscosity / rheology, and It is evaluated in terms of nanoparticle dispersion. Preferably, under specific storage conditions. The absence of observed phase separation (e.g., at room temperature) is defined as the “acceptance criterion”. 25 Assembly and use of the topical composite system described in the invention (combining of phases) (and its implementation) Phase-1 is added to Phase-2 by mixing at 20–25 °C and 300–600 rpm for 3–8 minutes; Phase-1:Phase-2 It is selected in the 1:1–1:4 (w / w) range. Final pH 6.8–7.4, final viscosity (100 s⁻¹) 15,600–17,800 cP 30 This is verified in the band. Table 2 shows the bonding window and final band of the topical composite system. Target performance has been given. Table 3: Topical composite system bonding window and end-target performance. Parameter Range / Target Phase-1:Phase-2 (w / w) 1:1 - 1:4 Integration temperature 20-25 °C Integration mixing speed 300-600 rpm Mixing time: 3-8 minutes Final pH 6.8-7.4 The ultimate viscosity target (100 s^-1) is in the 15,600-17,800 cP range. Visual stability: No color change; no phase separation. Visual stability control of the topical composite system obtained within the scope of the invention. When tested, color change and phase separation control (25 °C, n=6) was reported as “none”. pH and viscosity: Measurement after equilibration at 25 °C; viscosity for NFK-NP gel samples. pH values ​​have been reported. In-vitro release: dialysis bag method; medium pH 7.4 PBS; 37 ±0.5 5 °C; dialysis membrane 3–5 kDa; soak in pure water for 30 minutes before use. It has been defined. The effectiveness of the system obtained within the scope of the invention was demonstrated in an in vivo debridement wound model. It has been evaluated. 10 Experimental design: In one example application, a full-thickness skin wound was created, and the developed formulations were used to treat the wound. Its effect on recovery has been evaluated comparatively. In the experimental model. The animals were divided into multiple 15 groups according to the treatments to be applied to the 1 cm × 1 cm full-thickness wound defect created. divided into groups; the tested formulations were designed to completely cover the defect area. It was applied at the specified frequency. Clinical evaluation: During the postoperative period, the wound area is measured with calipers on specified days and recorded. 20 The percentage of wound healing was calculated based on the initial wound area (AWi initial wound area). (wound area on day AWn n). Histopathological evaluation: At the end of the experiment, skin samples taken from the wound area were fixed with 10% buffered formaldehyde. The samples were processed; after routine tissue processing, they were embedded in paraffin and 5 µm thick sections were prepared. Sections were stained with hematoxylin-eosin (HE) for general histopathological examination; connective tissue Picrosirius Red staining was applied to assess the healing process. Immunohistochemical evaluation: 30 11 Sections for immunohistochemical analysis were placed on poly-L-lysine coated slides and treated with streptavidin- stained by biotin peroxidase method; cytokeratin-1 as an indicator of epithelialization, VEGF expression as an indicator of angiogenesis was scored semi-quantitatively (0–3) and Intergroup comparisons were statistically analyzed. Summary of findings:  In the control group, wound closure and epithelialization were weakest; angiogenesis and It has been reported that collagen synthesis remains more limited.  In the group treated only with carrier hydrogel, improvement was similar to the control group. It has been reported. 10  In applications involving nanoparticles, especially “nanoparticle + plant macerate + Hydrogel combinations enhance epithelialization and connective tissue healing; It has been shown that cytokeratin and VEGF scores were increased compared to the control group (example). Control: cytokeratin 1.16±0.40, VEGF 0.33±0.21; nanoparticle form- BCO / HPM+H: cytokeratin 2.50±0.54, VEGF 2.33±0.51). 15  The highest scores were obtained in the standard treatment group used as a reference. It has been reported (Madecassol: cytokeratin 3.00±0.00, VEGF 3.00±0.00). Morphological verification: SEM images The morphology of the nano-carriers obtained within the scope of the invention was determined using scanning electron microscopy 20 (SEM) analysis was performed; the provided images show that the particles generally have a spherical morphology. It has been observed that it shows and that there are populations of different sizes (on the image (Information regarding EHT=10 kV, SE1 detector, different magnifications and scale bar is included. Figure-) The SEM image of the nanocarrier obtained within the scope of the invention is given in Figure 1. Figure 2 shows the histopathological appearance of wound healing in the skin defect area according to the groups. (A) Olive oil (OO) group: where epithelialization is not complete and in connective tissue (B) Black cumin oil (BCO) group: better connective tissue where mild proliferation is observed. (G) Hydrogel group: very mild epithelialization observed along with proliferation and connective tissue development is observed, (H) Nanoparticle form – Olive oil + Hypericum 30 Perforatum macerate + hydrogel (NPOOHPM+H) group: with marked epithelialization. (I) Nanoparticle form – Black cumin oil + slight improvement in connective tissue development was observed. Hypericum perforatum macerate + hydrogel (NPBCOHPM+H) group: heme epithelialization (J) Hypermix Gel group: epithelium, where a significant increase in connective tissue development was observed. (K) Madecassol ointment 35 showed slight closure of the tissue and significant improvement in the connective tissue. 12 Group: complete closure of the defect with complete closure of the epithelial and connective tissue, near normal. (L) Terramycin ointment group: mild but improvement appearance in epithelial tissue (M) Control group: Significant closure and marked improvement in connective tissue were observed. Epithelialization and connective tissue healing are very mild, and the defect remains open. Arrows indicate defects. It shows the area. HE, scale bars = 200 µm. 5 15 25 35

Claims

13 REQUESTS 1. The invention describes a blend of plant-based active ingredients used in the treatment of sensitive tissues such as wounds and burns. to increase its stability, persistence on the skin and effectiveness, in traditional topical formulations To address issues such as phase separation, flow, and poor penetration, it has 5 The two components are positioned separately within the packaging, and the aforementioned components... It becomes fluid with mechanical stimulation (shearing / applying) before use, and after application... A topical composite system that forms a structure and exhibits residual gel / film behavior on the skin. Its characteristic feature is that the mentioned components are macerated (30 / 40 / 50 mg), poly(D,L-lactide-co-glycolide) macerate 10 defined by (200 / 300 / 500 mg) and poly(vinyl alcohol) (0.5 / 1.0 / 2.0% m / v) levels. charged polymeric nanoparticle suspension (phase-1) and cross-linked poly(acrylic acid) based It is a mechanically-stimulated in-situ gel phase (phase-2).

2. The invention describes a blend of plant-based active ingredients used in the treatment of sensitive tissues such as wounds and burns. To increase its stability, persistence on the skin, and effectiveness, 15 in traditional topical formulations To address issues such as phase separation, flow, and poor penetration, it has The two components are positioned separately within the packaging, and the aforementioned components... It becomes fluid with mechanical stimulation (shearing / applying) before use, and after application... a topical composite system that acquires a structure and exhibits residual gel / film behavior on the skin It is a production method, and its characteristic is; 20  To prepare a macerate-loaded polymeric nanoparticle suspension (phase-1), Dried H. perforatum material is transferred to the olive oil or black cumin oil phase. At ±2 °C, in a dark environment, for 10–14 days, once a day for 5 minutes (300–500 rpm) stirring and letting it stand, then filtering the resulting macerate, 25  To prepare the organic phase, a biodegradable polymer is mixed with an organic solvent or solvent. Preparing the polymer solution by dissolving it in the mixture,  Addition of H. perforatum macerate to the polymer solution,  To prepare the aqueous phase, a stabilizer is dissolved in distilled / deionized water. 30 to be solved  The resulting aqueous phase is partially mixed with ethyl acetate to improve emulsification stability. or being completely satisfied,  Adding the organic phase containing the macerate dropwise to the aqueous phase containing PVA, and Emulsification is achieved by mixing the aqueous phase at a range of 400–10,000 rpm during addition. obtaining, 35  the emulsion created is designed to reduce particle size and narrow the dispersion. probe sonication, 14  Removal of organic solvent after emulsification of nanoparticles obtaining,  To prepare the in-situ gel phase (phase-2), carbomer and optional aloe vera were distilled in water. dissolved, homogenized by mixing, and then pH adjusted with 0.1 M triethanolamine. Setting the value to the range of 6.5–7.5, 5  The resulting gel should be left for 2–24 hours to allow bubbles to decompose and the network structure to stabilize. being left to stand for a certain period of time (preferably 12 hours),  Phase-1 is added to phase-2 by mixing at 20–25 °C and 300–600 rpm for 3–8 minutes. It includes the steps of the process. 15 25 35

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