Drug-loaded microalgae, preparation method and application thereof

Abstract

The application discloses a kind of medicine microalgae and its preparation method and application, specifically related to biological medicine technical field.The application uses Arthrospira platensis as the carrier of drug, by self-assembly technology, preparation berberine-hope phenol-Arthrospira platensis composite drug system, with the characteristics of natural environmental protection, lower cost, simple operation;The drug delivery system can inhibit intestinal inflammation, regulate intestinal flora homeostasis, most importantly, repair intestinal barrier, improve intestinal permeability, reduce peripheral inflammation level, reduce LPS and intestinal flora metabolites into blood, reduce the damage to liver through regulation of intestinal-liver axis, weaken the oxidative stress caused by alcohol in liver, and can regulate lipid metabolism in liver;In addition, the delivery system can be effectively decomposed, metabolized, and excreted through gastrointestinal tract, without accumulation toxicity and causing tissue damage, with good biocompatibility.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a drug-loaded microalgae, its preparation method, and its application. Background Technology

[0002] Alcoholic liver disease (ALD) is a leading cause of liver-related morbidity and mortality worldwide. The course of ALD typically progresses gradually from early alcoholic liver injury, potentially leading to steatosis, alcoholic hepatitis, liver fibrosis, cirrhosis, and even hepatocellular carcinoma. Therefore, effective control of alcoholic liver injury in the early stages of the disease is crucial for halting ALD progression and reducing the disease burden. However, current research primarily focuses on the liver itself to explore the pathogenesis of ALD, emphasizing intrahepatic pathological processes such as acetaldehyde toxicity from alcohol metabolism, oxidative stress, lipid metabolism disorders, and inflammatory cell infiltration. This strategy has limited clinical efficacy. Therefore, the occurrence and development of ALD may be influenced by extrahepatic factors in addition to intrahepatic mechanisms.

[0003] In recent years, the role of the gut-liver axis in alcoholic liver injury has received increasing attention. The intestine and liver are closely connected through pathways such as the portal vein circulation. Alcohol exposure can damage the intestinal barrier structure, increase intestinal permeability, and cause intestinal microecological imbalance, allowing intestinal substances such as lipopolysaccharide (LPS) to enter the liver, thereby inducing or aggravating liver inflammation and tissue damage.

[0004] Furthermore, some individuals continue or repeatedly drink alcohol even when they have underlying medical conditions. In clinical practice, in individuals with intestinal inflammation or a weakened intestinal barrier, continued alcohol consumption further impairs the intestinal barrier function, making it easier for harmful enterogenic substances to act on the liver through the portal vein circulation, thereby exacerbating alcoholic liver injury. Secondly, alcohol is metabolized in the liver, directly causing liver damage.

[0005] Based on this, regulating alcoholic liver injury from the perspective of the gut-liver axis involves improving intestinal barrier function, reshaping the intestinal microenvironment, and reducing the entry of enterogenic harmful substances into the liver. This reduces the liver's continuous exposure to harmful stimuli, thereby achieving early, continuous, and systemic regulation of alcoholic liver injury and related diseases. Intervening in the gut-liver axis may provide new treatment strategies for alcoholic liver injury.

[0006] Microalgae are abundant, natural, and renewable biological resources, and have attracted widespread attention in biomedical applications in recent years. Among them, Spirulina platensis (… Spirulina platensis*Coptis chinensis* (SP) is a naturally occurring, spiral-shaped microalga, 200-500 μm in length. It is an edible microorganism rich in various nutrients, characterized by low cost, high biocompatibility, and good pharmacological activity. Its cell membrane possesses 14-16 nm aqueous channels and junctional pores, facilitating the transport of specific ions, small molecules, and macromolecules. Huanglian Houpu Decoction, composed of *Coptis chinensis* and *Magnolol* bark, is a famous classic Chinese medicine prescription frequently used to treat gastrointestinal diseases. Berberine (BBR) is a representative active ingredient in *Coptis chinensis*, and magnolol (MAG) is a representative lignan extracted from *Magnolol* bark. The extremely low oral bioavailability of BBR and MAG limits their clinical application prospects in inflammatory diseases.

[0007] Therefore, this invention is proposed. Summary of the Invention

[0008] The technical problem to be solved by this invention is how to provide an oral microalgae drug delivery system with intestinal targeted retention and slow drug release capabilities, its preparation method, and its application in regulating the gut-hepatic axis to treat alcoholic liver injury.

[0009] The present invention solves the above-mentioned technical problems through the following technical means:

[0010] This invention proposes a drug-loaded microalgae, which includes Spirulina platensis, berberine loaded on Spirulina platensis, and magnolol.

[0011] Preferably, the drug-loaded microalgae consists of Spirulina platensis, berberine loaded on Spirulina platensis, and magnolol.

[0012] Preferably, the ratio of Spirulina platensis, berberine loaded on Spirulina platensis, and magnolol is (2-3) mg: (100-500) μg: (150-190) μg.

[0013] The drug-carrying microalgae described in this invention, Spirulina platensis, contains chlorophyll and has natural fluorescent properties. It does not require any chemical modification and can be used for fluorescence imaging in the gastrointestinal tract after oral administration, enabling non-invasive tracking in vivo.

[0014] The present invention also proposes a method for preparing the above-mentioned drug-loaded microalgae, comprising the following steps: S1: Berberine, Spirulina platensis, and solvent are mixed and incubated to obtain Spirulina platensis loaded with berberine. S2: Add magnolol solution to the berberine-loaded Spirulina platensis obtained in step S1, stir, centrifuge, and the precipitate is Spirulina platensis loaded with berberine and magnolol.

[0015] Preferably, in step S1, the ratio of berberine to Spirulina platensis is (100-500) μg: (2-3) mg, and more preferably 300 μg: 2 mg.

[0016] Preferably, in step S1, the ratio of berberine, Spirulina platensis, and solvent is (100-500) μg: (2-3) mg: 10 mL.

[0017] Preferably, in step S1, the solvent includes, but is not limited to, one or more of methanol and water.

[0018] Preferably, in step S1, the incubation conditions are incubation in the dark for 1-12 hours, and more preferably incubation for 6 hours.

[0019] Preferably, in step S2, the concentration of magnolol is 150-190 μg / mL, more preferably 190 μg / mL.

[0020] Preferably, in step S2, the magnolol solution is a mixed solution of magnolol and methanol.

[0021] Preferably, in step S2, the stirring time is 30 min to 180 min, and more preferably 60 min.

[0022] Preferably, in step S2, the centrifugation conditions are 4500-5000 rpm for 8-10 min, and more preferably 4500 rpm for 10 min.

[0023] The present invention also proposes the application of the above-mentioned drug-loaded microalgae in the preparation of drugs for the prevention and treatment of inflammatory bowel disease.

[0024] This invention also proposes the application of the above-mentioned drug-loaded microalgae in the preparation of drugs for repairing intestinal barrier damage.

[0025] The present invention also proposes the application of the above-mentioned drug-loaded microalgae in the preparation of drugs that regulate intestinal flora homeostasis.

[0026] Further preferred, the regulation is to improve the homeostasis of the gut microbiota.

[0027] The present invention also proposes the application of the above-mentioned drug-loaded microalgae in the preparation of drugs for the prevention and treatment of alcoholic liver injury.

[0028] The drug-loaded microalgae described in this invention can protect drugs from the damage caused by the acidic environment of the stomach, reach the site of intestinal inflammation, achieve a slow drug release effect, and improve the bioavailability of the drugs.

[0029] This invention also proposes a method for constructing an animal model of alcoholic liver injury accompanied by intestinal inflammation and intestinal barrier disruption, comprising the following steps: C57BL / 6 male mice were given free access to 2.5% w / v sodium dextran sulfate (DSS) solution while being administered 100 μL of 30% anhydrous ethanol by gavage at fixed times for 7 consecutive days to obtain an animal model of alcoholic liver injury with intestinal inflammation and intestinal barrier disruption.

[0030] The CAS number of the sodium dextran sulfate is 9011-18-1.

[0031] The present invention also proposes the application of the above-mentioned animal model of alcoholic liver injury with intestinal inflammation and intestinal barrier disruption in the preparation of drugs for the prevention and treatment of inflammatory bowel disease.

[0032] This invention also proposes the application of the above-mentioned animal model of alcoholic liver injury with intestinal inflammation and intestinal barrier disruption in the preparation of drugs for repairing intestinal barrier damage.

[0033] This invention also proposes the application of the above-mentioned animal model of alcoholic liver injury with intestinal inflammation and intestinal barrier disruption in the preparation of drugs that regulate intestinal flora homeostasis.

[0034] The present invention also proposes the application of the above-mentioned animal model of alcoholic liver injury with intestinal inflammation and intestinal barrier disruption in the preparation of drugs for the prevention and treatment of alcoholic liver injury.

[0035] The beneficial effects of this invention are as follows: 1. This invention uses *Spirulina platensis* as a drug carrier and prepares a berberine-honokiol-*Spirulina platensis* composite drug system through self-assembly technology. This system is characterized by its natural and environmentally friendly nature, low cost, and simple operation, making it easy to commercialize and translate into clinical applications. The spiral structure of *Spirulina platensis* is easily captured by intestinal villi, significantly prolonging the drug's residence time at the site of inflammation and improving the bioavailability of berberine and honokiol. This drug delivery system can inhibit intestinal inflammation, regulate intestinal flora homeostasis, primarily repair the intestinal barrier, improve intestinal permeability, reduce peripheral inflammation levels, reduce LPS and intestinal flora metabolites entering the bloodstream, alleviate liver damage by regulating the gut-hepatic axis, reduce oxidative stress caused by alcohol in the liver, and regulate lipid metabolism in the liver. Based on the inherent fluorescence of chlorophyll, the drug delivery system can achieve non-invasive tracking and monitoring of drug distribution in the body. Furthermore, this delivery system can be effectively decomposed and metabolized, and excreted through the gastrointestinal tract, exhibiting no cumulative toxicity and no tissue damage, and demonstrating good biocompatibility.

[0036] 2. This invention preferentially loads BBR onto SP, and then forms a stable nano-self-assembled structure in aqueous solution through electrostatic interactions and π–π stacking interactions between BBR and MAG. The drug delivery platform based on microalgae improves the delivery efficiency of BBR and MAG, achieving more effective treatment. BBR, as a classic natural isoquinoline alkaloid, and MAG, as a bisphenol natural product, can both alleviate oxidative stress, inhibit inflammation, regulate intestinal flora, and repair the intestinal barrier. After the intestinal barrier is repaired, the amount of LPS entering the liver via the portal vein circulation is reduced, further alleviating liver damage. It is expected to achieve the treatment of alcoholic liver injury through the gut-liver axis.

[0037] 3. The drug delivery system based on microalgae and self-assembly technology of this invention is a novel oral functional material that is natural and safe in composition. It has the ability to resist oxidation, anti-inflammation, regulate intestinal flora and repair the intestinal barrier, showing the application potential of treating alcoholic liver injury by regulating the gut-liver axis.

[0038] Of course, implementing any product or method of the present invention does not necessarily require achieving all of the advantages described above at the same time. Attached Figure Description

[0039] Figure 1 Images of Spirulina platensis (SP) under bright field, fluorescence-field, and scanning electron microscopes (SEM) in Example 1 of this invention, as well as SEM images of SP, SP@BBR, and SP@BM; Figure 2 This is a cumulative release curve of berberine and magnolol in artificial gastric fluid (SGF) and artificial intestinal fluid (SIF) in Example 2 of the present invention; Figure 3 This is a stability graph of the microalgae-based drug delivery system SP@BM in Embodiment 2 of the present invention after being placed in artificial gastric fluid for 3 hours and artificial intestinal fluid for 3 hours. Figure 4 This is a comparison chart of the effects of continuous alcohol consumption on the intestinal barrier damage in mice with intestinal inflammation and intestinal barrier disruption in Example 3 of the present invention (ZO-1, Occludin). Figure 5 This is a comparison chart of liver function (ALT, AST, TC, TG) in mice with intestinal inflammation and intestinal barrier disruption who had been continuously drinking alcohol in Example 3 of the present invention. Figure 6 This is a comparison chart of endotoxin (LPS) levels in the blood of mice with intestinal inflammation and intestinal barrier disruption who continuously consumed alcohol in Example 3 of the present invention. Figure 7These are in vivo fluorescence images of healthy and diseased mice at different time points after oral administration of the microalgae-based drug delivery system SP@BM in Example 4 of this invention. Figure 8 This is a comparison of colon length in diseased mice after treatment using the SP@BM microalgae-based drug delivery system in Example 5 of this invention. Figure 9 The images show a comparison of the effects of the microalgae-based drug delivery system SP@BM on improving intestinal damage and on intestinal barrier repair (ZO-1, Occludin) in Embodiment 5 of this invention. Figure 10 This is a comparative diagram showing the regulation of beneficial and harmful bacteria in the mouse gut microbiota by the microalgae-based drug delivery system SP@BM in Example 6 of the present invention. Figure 11 This is a comparative diagram showing the regulation of mouse gut microbiota structure by the phylogenetic level of the microalgae-based drug delivery system SP@BM in Example 6 of the present invention. Figure 12 This is a comparison chart of endotoxin (LPS) levels in the blood after treatment with the microalgae-based drug delivery system SP@BM in Example 6 of the present invention. Figure 13 This is a comparison chart showing the effects of the microalgae-based drug delivery system SP@BM on liver function (ALT, AST, TC, TG) in Example 7 of the present invention. Figure 14 This is a comparison chart showing the results of blood routine indicators (WBC, white blood cells; RBC, red blood cells; HGB, hemoglobin; MCH, mean corpuscular hemoglobin; MCHC, mean corpuscular hemoglobin concentration; MCV, mean corpuscular volume; PLT, platelets; HCT, hematocrit) and blood biochemical indicators (ALT, alanine aminotransferase; AST, aspartate aminotransferase; UREA, blood urea; CK, creatine kinase) after 30 days of continuous oral administration of the microalgae-based drug delivery system SP@BM in Example 8 of this invention. Figure 15 The image shows H&E staining of tissue sections from important organs (heart, liver, spleen, lung, kidney, and intestine) after 30 days of continuous oral administration of the microalgae-based drug delivery system SP@BM in Example 8 of this invention. The data in the attached figure were analyzed using a one-way ANOVA test to determine the significance of differences between groups. A p-value < 0.001 indicates a highly significant difference, denoted as _____. A highly significant difference is defined as 0.001 ≤ P < 0.01. A significant difference is defined as 0.01 < P ≤ 0.05. If the p-value is greater than 0.05, the difference is not significant and is denoted as ns. Detailed Implementation

[0040] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Unless otherwise defined, the technical terms used below have the same meaning as understood by those skilled in the art.

[0041] Unless otherwise specified, the test materials and reagents used in the following examples are commercially available or prepared by known methods.

[0042] Unless otherwise specified, all techniques or conditions described in the embodiments can be performed in accordance with the techniques or conditions described in the literature in this field or in the product manual. Unless otherwise specified, the quantitative experiments in the following embodiments are all repeated three times or more, and the results are averaged.

[0043] Example 1: Preparation of Spirulina platensis SP@BBR loaded with berberine A drug-loaded microalgae, comprising Spirulina platensis, berberine loaded on Spirulina platensis, and magnolol, in a ratio of 2 mg: 300 μg: 190 μg.

[0044] The preparation method of the above-mentioned drug-loaded microalgae includes the following steps: S1: Berberine and magnolol were dissolved in methanol to prepare a 10 mg / mL stock solution. 2 mg (dry weight) of Spirulina platensis (SP) was suspended in 9 mL of ultrapure water. 1 mL of berberine (BBR) solution with a concentration of 300 μg / mL was added (30 μL of the stock solution was diluted to 1 mL). The mixture was incubated in a constant temperature shaker at 25℃ and 90 rpm for 6 h in the dark. After centrifugation at 4500 rpm for 10 min, the supernatant and precipitate were collected. The precipitate was resuspended in ultrapure water to obtain Spirulina platensis loaded with berberine (SP@BBR). The supernatant was diluted to a certain proportion and the absorbance of BBR was measured by UV-Vis spectrophotometer. The drug loading rate of SP to BBR was found to be 41.03%.

[0045] S2: The prepared SP@BBR was resuspended in 9 mL of ultrapure water, and 1 mL of magnolol solution (190 μg / mL) was added dropwise. The mixture was stirred at room temperature for 60 min at 200 rpm, then centrifuged at 4500 rpm for 10 min. The precipitate, SP@BM, was collected. The drug loading rate of MAG was determined to be 25.50% using a fluorescence spectrophotometer. Microscopic and scanning electron microscope images were taken, and the results are as follows. Figure 1 As shown, Spirulina platensis SP has a spiral morphology and exhibits red fluorescence imaging characteristics due to its chlorophyll content. After being loaded with drugs, the surface of SP becomes rough.

[0046] Example 2: Drug release and stability in an in vitro simulated gastrointestinal tract SP@BM (quantified as BBR, concentration 300 μg / mL) prepared in Example 1 was placed in simulated gastric fluid for 2 h and simulated intestinal fluid for 6 h, with a total volume of 10 mL. Supernatant was collected at each time point, and the release rate of BBR was measured using UV-Vis spectrophotometry, while the release rate of MAG was measured using fluorescence spectrophotometry. The results are as follows: Figure 2 As shown, SP@BM releases a small amount of drug under SGF conditions and releases the drug slowly under SIF conditions. After 2 hours of artificial gastric fluid and 6 hours of artificial intestinal fluid, BBR releases only 39.78% of the drug, while MAG releases 53.23%.

[0047] SP@BM was placed in simulated gastric fluid for 3 hours and simulated intestinal fluid for 3 hours, with a total volume of 10 mL. The precipitate was collected by centrifugation at each time point, and bright-field images of SP@BM were taken using an optical microscope. SP@BM showed a complete structure in SGF and SIF, indicating that SP@BM has good stability and can successfully cross gastric fluid to reach the intestinal tract. Results are as follows: Figure 3 As shown.

[0048] Example 3: Establishment of an alcoholic liver injury model with intestinal inflammation and intestinal barrier disruption To investigate the relationship between intestinal barrier disruption and alcoholic liver injury, an alcoholic liver injury model with intestinal inflammation and intestinal barrier disruption was constructed using sodium dextran sulfate (DSS) in combination with anhydrous ethanol. Male C57BL / 6 mice were randomly divided into four groups: a blank control group, an EtOH group, a DSS group, and a DSS+EtOH group (DE group). Mice in the blank control group had free access to water for 7 days. Mice in the EtOH group had free access to water while receiving 100 μL of 30% anhydrous ethanol by gavage at fixed times for 7 consecutive days. Mice in the DSS group had free access to water containing 2.5% w / v sodium dextran sulfate (DSS), while mice in the DE group had free access to water containing 2.5% w / v DSS (i.e., 2.5 g DSS dissolved in 100 mL of ultrapure water) while receiving 100 μL of 30% anhydrous ethanol by gavage at fixed times for 7 consecutive days. Mice were euthanized on day 8, and the degree of intestinal barrier disruption (ZO-1, Occludin), the degree of liver function impairment (ALT, AST, TC, TG), and the level of LPS in the blood were assessed in each group. The results are as follows: Figure 4 , Figure 5 and Figure 6 As shown, DSS treatment can induce intestinal barrier disruption. Continued alcohol consumption on the basis of intestinal inflammation and barrier disruption can exacerbate intestinal barrier damage and significantly increase blood LPS levels, leading to liver injury and abnormal liver function. Experimental results indicate that alcohol intake can further aggravate intestinal barrier dysfunction, subsequently triggering or exacerbating liver inflammation and damage via the gut-hepatic axis.

[0049] Example 4: Dynamic distribution of a drug delivery system based on microalgae in vivo To investigate the distribution of drug-loaded Spirulina platensis in healthy and diseased mice, male C57BL / 6 mice were randomly divided into two groups: a control group (Healthy) and a DE group (Diease). Healthy mice had free access to water for 7 days, while diseased mice received free access to water and were simultaneously administered 100 μL of 30% anhydrous ethanol by gavage at fixed times for 7 consecutive days. 250 μL of SP@BM (quantified by BBR, dose 25 mg / kg) was injected into the mice by gavage. Fluorescence imaging of SP@BM in mice was tested using a small animal in vivo imaging system. Using the chlorophyll channel of SP, whole-body fluorescence imaging was performed on the mice at different time points after administration, and changes in fluorescence signal were recorded. The results are shown below. Figure 7As shown, after SP@BM was administered to mice via gavage, fluorescent signals were detected in the gastrointestinal tract, indicating that Spirulina platensis can be used for non-invasive fluorescence imaging to monitor the drug's distribution dynamics in vivo. Fluorescent signals were still detected in the gastrointestinal tract of healthy mice after 24 hours, indicating that SP@BM has good intestinal retention properties. This retention effect is due to the helical structure of SP, which is easily captured by intestinal villi after oral administration, prolonging the retention time of the delivery system in the intestinal lumen. Furthermore, the fluorescence signal intensity in the gastrointestinal tract of healthy mice was lower than that of diseased mice after 24 hours. Spirulina platensis retained for a longer time in inflamed intestinal regions. This effect is because SP@BM carries a negative charge, and intestinal inflammation leads to the enrichment of positively charged proteins, further prolonging the retention time of SP@BM.

[0050] Example 5: Repairing effect on intestinal barrier damage The experiment was divided into three groups: a blank control group, a model group, and an SP@BM group. Mice in the blank control group had free access to water for 7 days. Mice in the model group were fed 2.5% w / v DSS in drinking water for 7 days and were also administered 100 μL of 30% anhydrous ethanol daily via gavage to establish an alcoholic liver injury model with intestinal inflammation and intestinal barrier disruption. In the SP@BM group, mice were given 250 μL of SP@BM (quantified as BBR, dose 25 mg / kg) via gavage on days 0, 2, 4, 6, 8, and 10, in addition to drinking 2.5% w / v DSS and being administered 100 μL of 30% anhydrous ethanol via gavage. Mice were euthanized on day 11, and their colons were harvested for measurement. Colon sections were stained with H&E and immunofluorescence (ZO-1, Occludin) to assess intestinal damage and its effect on intestinal barrier repair. Results are as follows: Figure 8 , 9 As shown, mice treated with SP@BM exhibited reduced colonic shortening and increased expression of tight junction proteins, indicating that SP@BM can protect the intestine and repair the intestinal barrier.

[0051] Example 6: Regulatory effect on the gut-liver axis Based on the same grouping and treatment protocol as in Example 4, this embodiment collected fresh feces from different groups of mice before euthanasia on day 11 for 16S rRNA gene sequencing to analyze the regulation of gut microbiota. Compared to the model group, mice treated with SP@BM showed a significantly increased relative abundance of beneficial bacteria and a significantly decreased relative abundance of harmful bacteria. The results are as follows: Figure 10 As shown in the figure. Cluster analysis at the genus level showed that the SP@BM group and the blank control group clustered together, as shown in the figure. Figure 11As shown in the figure, the structure of the gut microbiota in the SP@BM group is more similar to that of healthy mice. Blood samples from mice were collected for endotoxin (LPS) testing to assess the levels of metabolites entering the bloodstream through the damaged intestinal barrier after gut microbiota metabolic dysregulation. Results are as follows... Figure 12 As shown, the experimental results indicate that SP@BM can reduce the proportion of harmful bacteria by increasing the proportion of beneficial bacteria, regulate the imbalanced intestinal flora, reduce the amount of endotoxins and other bacterial metabolites reaching the liver via the portal vein circulation, and weaken liver damage.

[0052] Example 7: Effects on Liver Function Improvement This embodiment, based on the same grouping and treatment scheme as in Example 4, involved collecting mouse blood for blood biochemistry analysis, detecting liver function indicators (ALT, AST, TG, TC), and evaluating the effect of SP@BM on improving liver function. The results are as follows: Figure 13 As shown, ALT, AST, TC, and TG levels in the SP@BM treatment group were lower than those in the model group and close to those in the blank control group. This indicates that SP@BM can improve liver function and reduce alcohol-induced liver damage.

[0053] Example 8 Oral Safety Mice were administered 250 μL of SP@BM (quantified as BBR, dose 25 mg / kg) via gavage every other day. After 30 days of administration, the mice were euthanized, and blood samples were collected for complete blood count and blood biochemistry analysis. Major organs (heart, liver, spleen, lung, kidney, and intestine) were also collected and prepared for H&E sectioning. Results are as follows: Figure 14 , 15 As shown: After SP@BM administration, the main blood routine (n=5) and blood biochemical indicators (n=3) of mice were not different from those of the control group, indicating that the microalgae-based drug delivery system has good oral safety.

[0054] Example 9: The difference between this embodiment and Example 1 is that the ratio of berberine to Spirulina platensis is 100 μg: 2 mg. The incubation time is 1 hour. The remaining steps are the same as in Example 1.

[0055] Example 10: The difference between this embodiment and Example 1 is that the ratio of berberine to Spirulina platensis is 500 μg: 3 mg. The incubation time is 12 h. The remaining steps are the same as in Example 1.

[0056] Example 11: The difference between this embodiment and Example 1 is that the concentration of magnolol is 150 μg / mL, the stirring time is 30 min, the centrifugation rate is 5000 rpm, and the centrifugation time is 8 min. The remaining steps are the same as in Example 1.

[0057] Example 12: The difference between this embodiment and Example 1 is that the concentration of magnolol is 180 μg / mL, the stirring time is 180 min, the centrifugation rate is 4800 rpm, and the centrifugation time is 9 min. The remaining steps are the same as in Example 1.

[0058] Comparative Example 1: The difference between this comparative example and Example 1 is that when the amount of berberine is 800ug, the structural integrity of Spirulina platensis is damaged when berberine is co-incubated with Spirulina platensis for 6 hours, resulting in a decrease in carrier stability; and under the condition of extending the incubation time, the drug loading rate is not significantly improved, indicating that the binding of berberine and Spirulina platensis has reached a saturation state.

[0059] Comparative Example 2: The difference between this comparative example and Example 1 is that the stirring time in the SP@BM system was extended to 360 min, which significantly damaged the spiral structure of Spirulina platensis, reduced the binding law of the drug loading system, and did not significantly improve the drug loading rate.

[0060] 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 the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A drug-loaded microalgae, characterized in that, The drug-loaded microalgae include Spirulina platensis, berberine loaded on Spirulina platensis, and magnolol.

2. The method for preparing drug-loaded microalgae according to claim 1, characterized in that, Includes the following steps: S1: Berberine, Spirulina platensis, and solvent are mixed and incubated to obtain Spirulina platensis loaded with berberine. S2: Add magnolol solution to the berberine-loaded Spirulina platensis obtained in step S1, stir, centrifuge, and the precipitate is Spirulina platensis loaded with berberine and magnolol.

3. The preparation method according to claim 2, characterized in that, In step S1, the ratio of berberine to Spirulina platensis is (100-500)ug: (2-3)mg.

4. The preparation method according to claim 2, characterized in that, In step S1, the solvent is one or more of methanol and water; the incubation conditions are incubation in the dark for 1-12 h.

5. The preparation method according to claim 2, characterized in that, In step S2, the concentration of magnolol is 150-190 μg / mL.

6. The preparation method according to claim 2, characterized in that, In step S2, the stirring time is 30 min-180 min.

7. The preparation method according to claim 2, characterized in that... In step S2, the centrifugation conditions are 4500-5000 rpm for 8-10 min.

8. The application of the drug-loaded microalgae according to claim 1 or the drug-loaded microalgae prepared by any one of claims 2-7, characterized in that, The application is at least one of the following: (1) Application in the preparation of drugs for the prevention and treatment of inflammatory bowel disease; (2) Application in the preparation of drugs for repairing intestinal barrier damage; (3) Application in the preparation of drugs for regulating intestinal flora homeostasis; (4) Application in the preparation of drugs for the prevention and treatment of alcoholic liver injury.

9. A method for constructing an animal model of alcoholic liver injury accompanied by intestinal inflammation and intestinal barrier disruption, characterized in that, Includes the following steps: C57BL / 6 male mice were given free access to 2.5% w / v sodium dextran sulfate solution while being administered 100 μL of 30% anhydrous ethanol by gavage at fixed times for 7 consecutive days to obtain an animal model of alcoholic liver injury with intestinal inflammation and intestinal barrier disruption.

10. The application of the animal model of alcoholic liver injury with intestinal inflammation and intestinal barrier disruption as described in claim 9, characterized in that, The application is at least one of the following: (1) Application in the preparation of drugs for the prevention and treatment of inflammatory bowel disease; (2) Application in the preparation of drugs for repairing intestinal barrier damage; (3) Application in the preparation of drugs for regulating gut microbiota homeostasis; (4) Application in the preparation of drugs for the prevention and treatment of alcoholic liver injury.

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