Composition for resisting allergic inflammation as well as preparation method and application thereof

Abstract

The invention discloses an anti-allergic inflammation composition and a preparation method and application thereof, the active ingredients of the composition comprise sphingomyelin and beta-arbutin, and the concentration ratio of the sphingomyelin to the beta-arbutin is 1: 2-2: 1. The sphingomyelin and beta-arbutin composite formula has a huge application prospect in the development of anti-allergic inflammation products (such as respiratory mucosa protective agents, skin anti-inflammatory care products, allergy prevention dietary supplements and the like).

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to an anti-allergic and anti-inflammatory composition, its preparation method, and its application. Background Technology

[0002] Rhinitis is a common inflammatory disease of the nasal mucosa, with typical symptoms such as nasal congestion, nasal itching, sneezing, and runny nose. In severe cases, it can lead to decreased sense of smell, sleep disorders, and secondary respiratory infections, significantly impacting patients' quality of life and physical and mental health.

[0003] Currently, clinical treatment for rhinitis primarily relies on drug intervention, with core medications including antihistamines, corticosteroids, decongestants, and leukotriene modifiers. While antihistamines can quickly relieve symptoms such as sneezing and nasal itching, their effectiveness in improving nasal congestion is limited, and long-term use can easily lead to adverse reactions such as drowsiness and dry mouth. Corticosteroid nasal sprays are effective for various types of rhinitis, but long-term use may cause local side effects such as nasal mucosal atrophy and nosebleeds, making them particularly unsuitable for children and pregnant women. Decongestants, due to their tendency to cause drug-induced rhinitis, are strictly limited to a duration of 7 days in clinical use. Furthermore, existing medications primarily aim to provide symptomatic relief and are unlikely to fundamentally repair the damaged nasal mucosal barrier, leading to frequent relapses of rhinitis and generally low patient compliance.

[0004] With the development of biomedical technology, rhinitis treatment based on natural active ingredients has gradually become a research hotspot due to its high safety and mild effects. Sphingomyelin, a natural phospholipid component widely found in mammalian cell membranes, possesses excellent biocompatibility and membrane repair functions. In vivo, it can repair damaged mucosal barriers by promoting cell membrane fluidity and enhancing intercellular connection stability. Existing studies have shown that sphingomyelin has good application potential in areas such as skin and mucosal injury repair and inflammation regulation. For example, in the treatment of dermatitis, it can reduce redness and swelling symptoms by inhibiting the release of inflammatory factors. However, research on its application in improving nasal mucosal inflammation has not yet been reported, and its mechanism of action and application value in rhinitis treatment still need further exploration.

[0005] β-Arbutin is a phenolic active ingredient extracted from natural plants such as bearberry and blueberry. Traditionally, it has been widely used in skin whitening and skincare due to its ability to inhibit tyrosinase activity. Recent pharmacological studies have found that β-arbutin also possesses significant anti-inflammatory and antioxidant activities. Compared to synthetic anti-inflammatory ingredients, β-arbutin is naturally derived, has extremely low toxicity, and good water solubility, making it easy to prepare into mucosal delivery formulations. However, its current applications are mainly concentrated in cosmetics and the treatment of skin diseases; no studies have yet combined it with sphingomyelin for the prevention and treatment of rhinitis.

[0006] Based on the current state of technology, there is an urgent need to develop a novel treatment for rhinitis that combines symptom relief, barrier repair, and inflammation regulation with high safety. In existing technologies, research on sphingomyelin and β-arbutin is conducted in independent areas, and no researchers have yet focused on their synergistic potential, let alone proposed a technical solution combining them for rhinitis improvement. Therefore, this invention innovatively combines the mucosal repair function of sphingomyelin with the anti-inflammatory activity of β-arbutin to construct a novel composition. Through the complementary and synergistic effects of their mechanisms of action, it achieves a comprehensive therapeutic effect of "repairing the damaged nasal mucosal barrier to block inflammatory triggers, inhibiting the release of inflammatory factors to relieve symptoms, and enhancing nasal mucosal resistance to prevent recurrence." This solves the technical problems of existing rhinitis treatments, such as significant side effects, high recurrence rates, and insufficient barrier repair capabilities, providing a completely new approach and solution for rhinitis treatment. Summary of the Invention

[0007] The purpose of this section is to outline some aspects of embodiments of the present invention and to briefly describe some preferred embodiments. Simplifications or omissions may be made in this section, as well as in the abstract and title of this application, to avoid obscuring the purpose of these documents; however, such simplifications or omissions should not be construed as limiting the scope of the invention.

[0008] In view of the problems existing in the above and / or prior art, the present invention is proposed.

[0009] Therefore, the purpose of this invention is to overcome the shortcomings of the prior art and provide an anti-allergic and anti-inflammatory composition, its preparation method, and its application.

[0010] To solve the above-mentioned technical problems, the present invention provides the following technical solution: an anti-allergic and anti-inflammatory composition, wherein the active ingredients of the composition include sphingomyelin and β-arbutin, and the concentration ratio of sphingomyelin to β-arbutin is 1:2 to 2:1.

[0011] In a preferred embodiment of the composition of the present invention, the concentration ratio of sphingomyelin to β-arbutin is 1:2.

[0012] As a preferred embodiment of the composition of the present invention, the concentration of sphingomyelin in the composition is 2.5 μM to 10 μM, and the concentration of β-arbutin is 2.5 μM to 10 μM.

[0013] As a preferred embodiment of the composition of the present invention, the concentration of sphingomyelin in the composition is 2.5 μM and the concentration of β-arbutin is 5 μM.

[0014] Another object of the present invention is to overcome the shortcomings of the prior art and provide a method for preparing an anti-allergic and anti-inflammatory composition, comprising stirring and dissolving sphingomyelin and β-arbutin to obtain the composition.

[0015] In a preferred embodiment of the preparation method described in this invention, the stirring temperature is 25~37℃ and the stirring time is 10~30min.

[0016] Another object of the present invention is to overcome the shortcomings of the prior art and provide the use of the composition in the preparation of a medicament for treating rhinitis.

[0017] Another object of the present invention is to overcome the shortcomings of the prior art and provide a rhinitis treatment drug, characterized in that the drug comprises the above-described composition.

[0018] As a preferred embodiment of the rhinitis treatment drug of the present invention, wherein: the rhinitis treatment drug repairs the barrier and blocks inflammatory signals, including, It inhibits excessive secretion of MUC5AC, restores the mucus barrier, and enhances the physical defense of the mucosa against allergens; Lowering ICAM-1 levels reduces inflammatory cell adhesion and infiltration, thus inhibiting inflammation amplification; It inhibits VEGF-A secretion, improves vascular permeability, and relieves mucosal edema; Targeting key transcripts in the PI3K / Akt / NF-κB pathway, it blocks upstream inflammatory signals.

[0019] As a preferred embodiment of the rhinitis treatment drug of the present invention, the allergens include dust mites, pollen, and IgE cross-linking in immune stimulation.

[0020] Beneficial effects of this invention: This invention provides a synergistic compound formulation of sphingomyelin and β-arbutin, which has a significant effect in relieving allergic inflammation and exhibits outstanding advantages in safety and regulatory efficiency, specifically reflected in: 1. Mast cell (RBL-2H3) level – blocking the initiation and amplification of allergies: inhibiting degranulation, significantly reducing the release of β-hexosidase (β-Hex), cutting off the allergy cascade at its source; strongly inhibiting excessive histamine (HIS) secretion, quickly relieving acute symptoms such as itching and redness; downregulating antigen-specific IgE expression, intervening in the sensitization stage, and reducing the risk of recurrence.

[0021] 2. Nasal mucosal epithelial cell (HNEpC) level – repairing the barrier and blocking inflammatory signals: inhibiting excessive secretion of MUC5AC, restoring the mucus barrier, and enhancing the physical defense of the mucosa against allergens; reducing ICAM-1 levels, reducing inflammatory cell adhesion and infiltration, and curbing inflammation amplification; inhibiting VEGF-A secretion, improving vascular permeability, and relieving mucosal edema; targeting key transcripts of the PI3K / Akt / NF-κB pathway, blocking inflammatory signals upstream, and achieving multi-node synergistic regulation.

[0022] 3. Combination optimization level - synergistic effect and safety improvement: Low dose of sphingomyelin combined with the benchmark dose of β-arbutin can produce maximum synergistic inhibition, with clear concentration dependence and ratio specificity, significantly reducing the dosage of single components, and further broadening the safety window and applicable scenarios.

[0023] Therefore, this sphingomyelin and β-arbutin complex formula has great application potential in the development of anti-allergic and anti-inflammatory products (such as respiratory mucosal protectants, anti-inflammatory skin care products, and dietary supplements for allergy prevention). Attached Figure Description

[0024] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort. Wherein: Figure 1 The figure shows the effect of different samples on the secretion level of β-Hex, a marker enzyme for degranulation of RBL-2H3 cells, in an embodiment of the present invention.

[0025] Figure 2 The figure shows the effect of different samples on the secretion level of the inflammatory mediator HIS in RBL-2H3 cells in the embodiments of the present invention.

[0026] Figure 3 The figure shows the effect of different samples on the secretion level of immunoglobulin IgE in RBL-2H3 cells in the embodiments of the present invention.

[0027] Figure 4 The figure shows the effect of different samples on the secretion level of HNEpC cell mucin MUC5AC in the embodiments of the present invention.

[0028] Figure 5 The figure shows the effect of different samples on the secretion level of HNEpC cell adhesion factor ICAM-1 in the embodiments of the present invention.

[0029] Figure 6The figure shows the effect of different samples on the secretion level of endothelial growth factor VEGEA in HNEpC cells according to the embodiments of the present invention.

[0030] Figure 7 Figure 1 shows the effects of different samples on the transcriptional levels of the PI3K / Akt / NF-κB inflammatory pathway in HNEpC cells according to embodiments of the present invention. Figure A shows the changes in PI3K transcriptional levels, Figure B shows Akt, and Figure C shows NF-κB. Detailed Implementation

[0031] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the specific embodiments of the present invention will be described in detail below with reference to the examples in the specification.

[0032] Many specific details are set forth in the following description in order to provide a full understanding of the invention. However, the invention may also be practiced in other ways different from those described herein, and those skilled in the art can make similar extensions without departing from the spirit of the invention. Therefore, the invention is not limited to the specific embodiments disclosed below.

[0033] Secondly, the term "one embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in different places in this specification does not necessarily refer to the same embodiment, nor is it a single or selective embodiment that is mutually exclusive with other embodiments.

[0034] Unless otherwise specified, all raw materials used in the embodiments of this invention are commercially available. See Table 1 for details.

[0035] Table 1

[0036] Example 1 The alleviating effect of different groups on degranulation state of rat leukocyte disease cells (RBL-2H3) Frozen rat leukemia cells (RBL-2H3) were rapidly thawed at 37°C and resuspended in three times the volume of cell culture medium. The cells were centrifuged at 500 g for 5 min, the supernatant was discarded, and the cells were resuspended again in cell culture medium and seeded into T25 cell culture dishes.

[0037] After resuscitation, cells were cultured at 37°C with 5% CO2. When approximately 80%-90% of the cell culture dish was covered by growing cells, the cells were passaged. Cells were digested with 1 mL of trypsin-EDTA (0.25%) and observed under a microscope. Once the cells were in suspension, digestion was immediately stopped with three times the volume of complete cell culture medium. After centrifugation, the cells were collected and passaged at a ratio of 1:3. The experimental groups are shown in Table 2.

[0038] Table 2 Grouping Information Table

[0039] After equilibrating the ELISA kit with cell supernatant at room temperature for 30 min, add 100 μL of serially diluted cell supernatant to each well of the microplate and incubate at 37°C for 80 min. After washing three times with washing buffer and patting dry, add 100 μL of biotinylated antibody working solution and HRP enzyme conjugate working solution sequentially, and incubate at 37°C for 50 min each time. Wash after each incubation according to the above conditions. Then wash five times with washing buffer, add 90 μL of TMB substrate solution and incubate at 37°C for 20 min in the dark for color development. Then add 50 μL of stop solution to terminate the reaction. Measure the absorbance (OD value) at 450 nm using a microplate reader and calculate the concentration of different cytokines in the sample by fitting the equation to a standard curve.

[0040] Effects of different groups on the level of β-Hex, a marker of cell degranulation β-Hexase (β-Hex) is a characteristic acidic hydrolase found in the intracellular granules of mast cells and basophils. Stored within the granules at rest, it undergoes degranulation when cells are exposed to allergens (such as dust mites and pollen) or immune stimuli (such as IgE cross-linking), releasing β-Hex inflammatory mediators (such as histamine and leukotrienes) extracellularly. The amount released is positively correlated with the cell degranulation rate, and the assay exhibits high sensitivity and stability. It is considered one of the "gold standards" for accurately quantifying cell activation, replacing easily degradable mediators like histamine, and can directly reflect the initiation and intensity of allergic inflammation.

[0041] Experimental results are as follows Figure 1As shown, after IgA sensitization and human serum albumin (HSA) stimulation, the β-Hex release level of RBL-2H3 cells was significantly increased by 46.91% compared with the control group, reaching 26.63 ng / mL (P<0.0001), indicating that the cell degranulation model was successfully established. In the single intervention experiment, both sphingomyelin and β-arbutin significantly downregulated the β-Hex release level, with inhibition rates of 56.41% and 45.69%, respectively, and their regulatory effects were comparable to those of the positive control group (P<0.0001). Concentration gradient experiments showed that the low-concentration combination 1 had an inhibition rate of only 21.47% on β-Hex release (P<0.05); as the combination concentration increased, the regulatory activity increased in a concentration-dependent manner, and the inhibitory effects of the medium-concentration combination 2 and the high-concentration combination 3 on β-Hex release reached significant levels compared with the single-substance intervention group. Further optimization experiments revealed differences in the regulatory effects of different formulations on β-Hex release: combination 4 (low-concentration sphingomyelin + baseline concentration β-arbutin) showed an inhibition rate of 43.78%, while combination 5 showed an inhibition rate of 33.94%. This suggests that the ratio of low-concentration sphingomyelin to baseline concentration β-arbutin is more conducive to synergistic inhibition, resulting in better regulation of β-Hex release. In summary, sphingomyelin and β-arbutin, alone or in combination, can effectively inhibit RBL-2H3 cell degranulation, and their combined effect is concentration-dependent and ratio-specific. The combination of low-concentration sphingomyelin and baseline concentration β-arbutin can achieve more efficient degranulation inhibition, providing experimental evidence for further optimization of the sphingomyelin and β-arbutin compound formulation and enhancement of anti-allergic and anti-inflammatory activity.

[0042] Effects of different groups on the level of the cellular inflammatory mediator HIS Histamine (HIS) is the primary inflammatory mediator released from mast cells and basophils during degranulation, possessing extremely strong biological activity. Acting on vascular endothelial cells, it can cause vasodilation and increased permeability, leading to skin erythema and edema (such as eczema and urticaria); it also stimulates respiratory mucosal gland secretion, exacerbating sneezing and runny nose symptoms of allergic rhinitis. Its release level is positively correlated with the severity of allergic inflammatory symptoms in children. Measuring HIS levels can directly quantify the degree of pathological damage caused by inflammation, establishing a direct link between in vitro experimental results and children's clinical symptoms, thus enhancing the clinical translational value of the research.

[0043] Experimental results are as follows Figure 2As shown, after IgA sensitization and human serum albumin (HSA) stimulation, the HIS release level of RBL-2H3 cells was significantly increased by 44.0% compared with the control group, reaching 24.32 ng / mL (P<0.0001), indicating that the allergic inflammatory cell model was successfully constructed. In the single intervention experiment, sphingomyelin and β-arbutin could both significantly downregulate the HIS release level, with inhibition rates of 40.27% and 39.60%, respectively, and their regulatory effects were comparable to those of the positive control group (P<0.001). Concentration gradient experiments showed that the inhibition rate of low-concentration combination 1 on β-Hex release was only 19.08%, which was not statistically different from the model group (P>0.05); as the combination concentration increased, the regulatory activity increased in a concentration-dependent manner, and the inhibitory effects of medium-concentration combination 2 and high-concentration combination 3 on β-Hex release reached significant levels comparable to the single-substance intervention group. Further optimization experiments revealed differences in the regulatory effects of different formulations on β-Hex release: combination 4 (low-concentration sphingomyelin + baseline concentration β-arbutin) showed an inhibition rate of 41.99%, while combination 5 showed an inhibition rate of 32.80%. This suggests that the ratio of low-concentration sphingomyelin to baseline concentration β-arbutin is more conducive to synergistic inhibition, resulting in better regulation of β-Hex release. Therefore, sphingomyelin and β-arbutin, used alone or in combination, can effectively inhibit histamine release from RBL-2H3 cells, and their combined effect is concentration-dependent and ratio-specific. The combination of low-concentration sphingomyelin and baseline concentration β-arbutin achieves more efficient histamine release inhibition, providing direct experimental evidence for further optimization of anti-allergy compound formulations and enhancing the clinical translation potential of the compositions.

[0044] Effects of different groups on the level of cellular immunoglobulin IgE Immunoglobulin E (IgE) can initiate an immune response in allergic inflammation through the "IgE-FcεRI" pathway. When allergens (such as HSA and house dust mite proteins) enter the body, they can induce B cells to produce specific IgA. This antibody binds to FcεRI receptors on the surface of mast cells and basophils, completing "sensitization." Subsequent exposure to the same allergen triggers receptor cross-linking, initiating cell degranulation and the release of inflammatory mediators such as histamine and β-Hex, thus initiating an allergic reaction. Detecting IgE levels in the experimental system can verify the effectiveness of the "IgE sensitization-RBL-2H3 cell degranulation" model (after sensitization, IgE binds to the cell surface, ensuring the specificity of subsequent challenge experiments); at the same time, it quantifies the degree of immune activation in the allergic reaction initiation phase, providing direct evidence for assessing whether the "sensitization process" needs to be intervened in different samples.

[0045] Experimental results are as follows Figure 3As shown, after IgE sensitization and human serum albumin (HSA) stimulation, the IgE release level of RBL-2H3 cells was significantly increased by 46.88% compared with the control group, reaching 40.18 μg / mL (P<0.0001), indicating that the IgE-mediated cellular hypersensitivity model was successfully established. In the single intervention experiment, both sphingomyelin and β-arbutin significantly downregulated IgE release levels (P<0.0001), with inhibition rates of 38.93% and 35.13%, respectively, while the positive control group reached 46.31%. Concentration gradient experiments showed that the low-concentration combination 1 had an inhibition rate of only 26.27% on IgE release (P<0.01), with a relatively mild regulatory effect; as the combination concentration increased, the regulatory activity increased in a concentration-dependent manner, with the medium-concentration combination 2 and the high-concentration combination 3 having inhibition rates of 46.49% and 42.58% on IgE release, respectively, among which the inhibitory effect of combination 2 was close to that of the positive control group. In the ratio optimization experiment, the regulatory effects of different combinations on IgE release showed a high degree of consistency: combination 4 (low concentration of sphingomyelin + baseline concentration of β-arbutin) had an inhibition rate of 41.15%, while combination 5 had an inhibition rate of 43.49%. This suggests that when sphingomyelin and β-arbutin are used in combination, the regulatory effect on IgE release is less affected by the ratio, and efficient inhibition can be achieved within the tested ratio range. Their synergistic regulatory effect has good ratio tolerance. In summary, both sphingomyelin and β-arbutin, alone and in combination, can effectively inhibit IgE-mediated IgE release in RBL-2H3 cells. The combined intervention showed significant concentration dependence and strong ratio adaptability. Among them, the medium concentration combination 2 had the best inhibitory effect (46.49%), which is close to the level of the positive control group. This provides experimental support for subsequent optimization of anti-allergy compound formulations and improvement of the intervention effect of the composition on the allergic sensitization stage.

[0046] Example 2 Effects of different sample groups on improving the mucosal barrier of human nasal mucosal epithelial cells (HNEpC) Frozen rat leukemia cells (RBL-2H3) were rapidly thawed at 37°C and resuspended in three times the volume of cell culture medium. The cells were centrifuged at 500 g for 5 min, the supernatant was discarded, and the cells were resuspended again in cell culture medium and seeded into T25 cell culture dishes.

[0047] After resuscitation, cells were cultured at 37°C with 5% CO2. When approximately 80%-90% of the cell culture dish was covered by the growing cells, the cells were passaged. Cells were digested with 1 mL of trypsin-EDTA (0.25%) and observed under a microscope. Once the cells were in suspension, digestion was immediately stopped with three times the volume of complete cell culture medium. After centrifugation, the cells were collected and passaged at a 1:2 ratio. The experimental groups are shown in Table 3.

[0048] Table 3 Grouping Information Table

[0049] The method for detecting cell supernatant is described in Example 1.

[0050] After washing the cells three times with PBS, RNA extraction reagent was added, and the cells were repeatedly pipetted to accelerate cell lysis. RNA was extracted from the cells according to the RNA extraction kit instructions. After reverse transcription was completed according to the reverse transcription kit instructions, the cDNA sample was diluted 20 times and real-time quantitative PCR was performed according to the q-PCR kit instructions. The primers are shown in Table 4.

[0051] Table 4 Primer Sequences

[0052] Effects of different groups on the secretion level of cell mucin MUC5AC Mucin 5AC (MUC5AC) is a gel-like mucin secreted by mucosal epithelial cells of the respiratory tract, digestive tract, and conjunctiva. It is an important component of the mucosal barrier, and its abnormal expression is closely related to the pathological process of allergic inflammation. Under normal physiological conditions, MUC5AC forms a mucus layer, preventing allergens and pathogens from penetrating the mucosal epithelium and maintaining barrier integrity. Under allergic conditions, inflammatory factors stimulate mucosal epithelial cells to excessively secrete MUC5AC, leading to thickened mucus and increased viscosity, triggering common allergic symptoms such as airway obstruction (e.g., runny nose and wheezing in allergic rhinitis and asthma) and abnormal gastrointestinal mucus secretion (e.g., diarrhea in allergic enteritis). Its secretion level is positively correlated with the severity of inflammation. Detecting MUC5AC levels can directly quantify the degree of damage to the mucosal barrier and provide a basis for assessing the pathological relevance of allergy models.

[0053] Experimental results are as follows Figure 4As shown, after HIS induction, the MUC5AC release level of HNEpC cells increased significantly by 51.72% compared with the control group, reaching 129.36 pg / mL (P<0.0001), indicating that the HIS-induced abnormal cell mucus secretion damage model was successfully established. In the single intervention experiment, both sphingomyelin and β-arbutin significantly downregulated the MUC5AC secretion level (P<0.0001), with inhibition rates of 42.42% and 36.57%, respectively, while the positive control group reached 47.68%. Concentration gradient experiments showed that the low concentration combination 1 had an inhibition rate of only 25.27% on MUC5AC release (P<0.001); as the combination concentration increased, the regulatory activity increased in a concentration-dependent manner (P<0.0001), with the medium concentration combination 2 and high concentration combination 3 having inhibition rates of 47.42% and 41.77% on MUC5AC secretion, respectively. In the ratio optimization experiment, different combinations showed different regulatory effects on MUC5AC secretion: combination 4 (low concentration sphingomyelin + baseline concentration β-arbutin) had an inhibition rate of 52.32%, while combination 5 had an inhibition rate of 40.84%, suggesting that this ratio is more conducive to the synergistic inhibitory effect of the two, and its effect on regulating abnormal MUC5AC secretion is the best, superior to single substances and the positive control group. In summary, sphingomyelin and β-arbutin, alone or in combination, can effectively inhibit HIS-induced abnormal mucus secretion in HNEpC cells, and the combined intervention has significant concentration dependence and ratio specificity. Among them, the combination of low concentration sphingomyelin and baseline concentration β-arbutin has the strongest synergistic effect, and can more efficiently reverse the mucosal barrier damage caused by excessive MUC5AC secretion, with a regulatory effect superior to single components and the positive control.

[0054] Example 3 Effects of different groups on the secretion level of cell adhesion factor ICAM-1 Intercellular adhesion molecule-1 (ICAM-1) is a transmembrane glycoprotein mainly expressed on the surface of vascular endothelial cells, epithelial cells, and immune cells. Its core role in allergic inflammation is to mediate cell adhesion and recruitment. After allergic activation, inflammatory factors induce high expression of ICAM-1, promoting the adhesion of inflammatory cells such as eosinophils, mast cells, and lymphocytes to vascular endothelial cells, which then infiltrate inflammatory sites (such as the respiratory tract and skin mucosa), exacerbating inflammatory damage. At the same time, high expression of ICAM-1 disrupts the tight junctions between epithelial cells, exacerbating the increase in mucosal barrier permeability and promoting the invasion of allergens, forming a vicious cycle of "inflammatory recruitment - barrier damage - allergic exacerbation." Its expression level is positively correlated with the severity of allergic inflammation. Detecting ICAM-1 can directly quantify the degree of activation of the inflammatory cascade response, providing a basis for assessing the pathological relevance of the model.

[0055] Experimental results are as follows Figure 5 As shown, after HIS induction, the ICAM-1 release level in HNEpC cells increased significantly by 40.81% compared with the control group, reaching 107.09 pg / mL (P<0.0001), indicating that the HIS-induced cell epithelial injury model was successfully established. In the single intervention experiment, both sphingomyelin and β-arbutin significantly downregulated ICAM-1 secretion levels, with inhibition rates of 43.38% (P<0.0001) and 24.84% (P<0.001), respectively, while the positive control group reached 32.47% (P<0.0001). Concentration gradient experiments showed that the low concentration combination 1 had an inhibition rate of only 16.43% on ICAM-1 release (P<0.05); with increasing combination concentration, the regulatory activity increased in a concentration-dependent manner (P<0.0001), and the inhibition rates of medium concentration combination 2 and high concentration combination 3 on ICAM-1 secretion were 36.16% and 35.28%, respectively. In the ratio optimization experiment, different combinations showed different regulatory effects on ICAM-1 secretion: combination 4 (low concentration sphingomyelin + baseline concentration β-arbutin) had an inhibition rate of 39.82%, while combination 5 had an inhibition rate of 24.11%, suggesting that this ratio is more conducive to the synergistic inhibitory effect of the two, and its effect on regulating abnormal ICAM-1 release is the best, and it is superior to the single substance and the positive control group. In summary, sphingomyelin and β-arbutin, alone or in combination, can effectively inhibit HIS-induced abnormal ICAM-1 secretion in HNEpC cells. Among them, sphingomyelin alone has the strongest regulatory activity, while the combined intervention of the two shows a clear concentration dependence and ratio specificity. The combination of low concentration sphingomyelin and baseline concentration β-arbutin achieves a highly efficient ICAM-1 regulatory effect through synergistic action, which not only retains the strong activity advantage of sphingomyelin, but also reduces its concentration, providing experimental evidence for the subsequent development of low-dose, high-safety anti-allergy compound formulations.

[0056] Effects of different groups on the secretion level of endothelial growth factor VEGEA Vascular endothelial growth factor A (VEGF-A) is a key factor regulating angiogenesis and permeability. In allergic states, histamine and IL-4 released by mast cells induce high expression of VEGF-A in mucosal epithelial / endothelial cells, leading to vasodilation and increased permeability (causing mucosal edema). Simultaneously, it promotes angiogenesis, providing a "channel" for inflammatory cell infiltration and exacerbating allergic symptoms in children such as rhinitis and asthma. Its level is positively correlated with the severity of inflammation.

[0057] Experimental results are as follows Figure 6As shown, after HIS induction, the VEGF-A release level of HNEpC cells increased significantly by 45.85% compared with the control group, reaching 59.97 ng / mL (P<0.0001), indicating that the HIS-induced cell epithelial barrier damage model was successfully established. In the single intervention experiment, both sphingomyelin and β-arbutin significantly downregulated VEGF-A secretion levels (P<0.0001), with inhibition rates of 52.30% and 56.46%, respectively, while the positive control group reached 53.44%. Concentration gradient experiments showed that the low concentration combination 1 had an inhibition rate of only 23.14% on VEGF-A release (P<0.01); as the combination concentration increased, the regulatory activity increased in a concentration-dependent manner (P<0.0001), with the medium concentration combination 2 and high concentration combination 3 having inhibition rates of 48.56% and 54.46% on VEGF-A secretion, respectively. In the ratio optimization experiment, different combinations showed different regulatory effects on VEGF-A secretion: combination 4 (low concentration sphingomyelin + baseline concentration β-arbutin) had an inhibition rate of 54.45% (P<0.0001), while combination 5 had an inhibition rate of 26.37% (P<0.001), suggesting that this ratio is more conducive to the synergistic inhibitory effect of the two, and its effect on regulating abnormal VEGF-A release is the best, superior to single substances and the positive control group. In summary, sphingomyelin and β-arbutin, alone or in combination, can effectively inhibit HIS-induced abnormal VEGF-A secretion in HNEpC cells, and the combined intervention shows a clear concentration dependence and ratio specificity. The combination of low concentration sphingomyelin and baseline concentration β-arbutin achieves a highly efficient and low-dose VEGF-A regulatory effect through synergistic action, which not only reduces the concentration of single components to improve safety, but also effectively inhibits VEGF-A-mediated increase in vascular permeability and epithelial barrier damage.

[0058] Effects of different groups on the transcriptional levels of the PI3K / Akt / NF-κB cellular inflammatory pathway The PI3K / Akt / NF-κB pathway is a "key signaling hub" in the development of allergic inflammation. Its abnormal activation can cascade the entire chain of "sensitization-release of inflammatory mediators-mucosal damage." Allergen-induced IgE / FcεRI crosslinking and histamine stimulation can activate PI3K, promote downstream Akt phosphorylation, and thus activate NF-κB. After entering the cell nucleus, activated NF-κB can directly regulate the transcriptional expression of target genes such as inflammatory mediators, adhesion molecules (ICAM-1), mucosal functional molecules (MUC5AC), and vasoactive factors (VEGF-A), triggering an inflammatory cascade response. The degree of activation of this pathway is positively correlated with the severity of allergic inflammation. Detecting the transcriptional level of its key molecules can quantify the inflammatory activation state at the "upstream signaling" level, overcoming the limitations of only detecting downstream effector molecules (such as histamine and MUC5AC).

[0059] Experimental results are as follows Figure 7 As shown, after HIS induction, the levels of PI3K / Akt / NF-κB transduction pathways were significantly upregulated, indicating the successful establishment of the inflammation model. After single-component intervention, each signaling molecule was downregulated to varying degrees. Notably, sphingomyelin alone failed to significantly inhibit Akt transcription, with its expression level being 0.84 times that of the model group (P>0.05); while β-arbutin showed significant inhibitory effects, reducing Akt transcription to 0.46 times that of the model group (P<0.0001), which was superior to the 0.60 times (P<0.0001) in the positive control. In the compound formulation, combinations 2, 3, and 4 showed similar effects, all significantly downregulating Akt transcription levels (P<0.0001), at 0.57, 0.53, and 0.56 times that of the model group, respectively; the regulatory efficacy of combinations 1 and 5 was significantly weaker than the first three. Furthermore, the transcriptional levels of PI3K and NF-κB p65 showed a regulatory pattern consistent with that of Akt: among single components, β-arbutin was significantly more effective than sphingomyelin in inhibiting the transcription of PI3K and NF-κB p65; in the combination, the regulatory effect increased with increasing concentration; and in the ratio optimization experiment, combination 4 (low concentration of sphingomyelin + baseline concentration of β-arbutin) was significantly more effective than combination 5 in inhibiting the transcription of PI3K and NF-κB p65, suggesting that this ratio is more conducive to synergistically downregulating the overall activation level of the pathway. This indicates that the PI3K / Akt / NF-κB signaling pathway is the core molecular target of β-arbutin in exerting its anti-allergic and anti-inflammatory effects, and it can block inflammatory signal transduction upstream by strongly inhibiting the transcription of key molecules in the pathway (especially Akt); while sphingomyelin alone has weak regulatory activity on this pathway and can only play an auxiliary role at specific concentrations. When used in combination, the two drugs achieve highly efficient synergistic regulation of the PI3K / Akt / NF-κB pathway through a "β-arbutin-dominant pathway inhibition + low-concentration sphingomyelin synergistic enhancement" model. This synergistic effect depends on the appropriate concentration and ratio combination. Specifically, the combination of low-concentration sphingomyelin and the baseline concentration of β-arbutin (combination 4) not only maintains the potent pathway-inhibiting activity of β-arbutin alone but also optimizes the regulatory effect through synergistic action, while reducing the concentration of each individual component to improve safety.

[0060] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all such modifications or substitutions should be covered within the scope of the present invention.

Claims

1. An anti-allergic and anti-inflammatory composition, characterized in that: The active ingredients of the composition include sphingomyelin and β-arbutin, wherein the concentration ratio of sphingomyelin to β-arbutin is 1:2 to 2:

1.

2. The anti-allergic and anti-inflammatory composition according to claim 1, characterized in that: The concentration ratio of sphingomyelin to β-arbutin is 1:

2.

3. The anti-allergic and anti-inflammatory composition according to claim 1 or 2, characterized in that: The concentration of sphingomyelin in the composition is 2.5 μM to 10 μM, and the concentration of β-arbutin is 2.5 μM to 10 μM.

4. The anti-allergic and anti-inflammatory composition according to claim 3, characterized in that: The composition contains 2.5 μM sphingomyelin and 5 μM β-arbutin.

5. A method for preparing the anti-allergic and anti-inflammatory composition according to any one of claims 1 to 4, characterized in that: The method includes stirring and dissolving sphingomyelin and β-arbutin to obtain the composition.

6. The preparation method according to claim 5, characterized in that: The stirring temperature is 25~37℃, and the stirring time is 10~30min.

7. Use of the composition according to claim 1 in the preparation of a medicament for treating rhinitis.

8. A rhinitis treatment drug, characterized in that: The drug comprises the composition of claim 1.

9. The rhinitis treatment drug as described in claim 8, characterized in that: The rhinitis treatment medication repairs the skin barrier and blocks inflammatory signals, including, It inhibits excessive secretion of MUC5AC, restores the mucus barrier, and enhances the physical defense of the mucosa against allergens; Lowering ICAM-1 levels reduces inflammatory cell adhesion and infiltration, thus inhibiting inflammation amplification; It inhibits VEGF-A secretion, improves vascular permeability, and relieves mucosal edema; Targeting key transcripts in the PI3K / Akt / NF-κB pathway, it blocks upstream inflammatory signals.

10. The rhinitis treatment drug as described in claim 9, characterized in that: The allergens include dust mites, pollen, and IgE cross-linking in immune stimulation.

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