Preparation method of ginsenoside Rb1 and baicalein nano preparation

Abstract

The invention discloses a preparation method of a ginsenoside Rb1 and baicalein nano preparation, and belongs to the technical field of medicine preparation and nano materials. Ginsenoside Rb1 and baicalein are assembled into a nano co-delivery system through a nano precipitation technology, nucleation and growth processes of nano aggregates are regulated and controlled based on dynamics in the nano precipitation technology, the baicalein is wrapped by the ginsenoside Rb1 to form a nano preparation with a small particle size, synthesis operation is simple, and the nano preparation has better targeting performance in drug application. According to the prepared ginsenoside Rb1 and baicalein nano preparation, the good toxoplasma gondii resisting effect of the baicalein can be achieved, the drug solubility of the ginsenoside Rb1 can also be achieved, the drug utilization rate is greatly increased, the service life of the drug is prolonged through a wrapping mode, and the ginsenoside Rb1 and baicalein nano preparation has biological safety and small damage to normal tissue.

Description

Technical Field

[0001] The invention belongs to the technical field of drug preparation, and particularly relates to a method for preparing a ginsenoside Rb1 and baicalein nano preparation. Background Art

[0002] Toxoplasma gondii is an intracellular parasite that causes toxoplasmosis, which is widespread worldwide and can be transmitted between humans, cattle, sheep, pigs, chickens, cats and other animals, causing serious harm to human health and social development. When normal people are infected with Toxoplasma, they are generally latently infected and do not show obvious symptoms. However, if people with low immunity are infected, Toxoplasma may invade the brain, liver, heart and other systems. If pregnant women are unfortunately infected with Toxoplasma, it will cause quite serious consequences, such as spontaneous abortion, fetal malformation and neonatal infection. Up to now, there is no specific treatment drug or strong preventive measures for Toxoplasma infection. In clinical practice, the combination of pyrimethamine and sulfadiazine is generally used to treat toxoplasmosis. Although it has achieved certain therapeutic effects, it has many disadvantages such as adverse reactions, cannot be taken for a long time, and has a high recurrence rate after stopping the drug.

[0003] At present, Chinese herbal medicines and their extracts can be screened as anti-Toxoplasma drugs. For example, reference 1 (Wang Yunchi, Ju Yulin, Yao Lin, Comparative test of the in vitro anti-Toxoplasma effect of three flavonoids of Scutellaria baicalensis [J], Journal of Agricultural Sciences of Yanbian University, 2012, 34, 75-78, doi: 10.13478 / j.cnki.jasyu.2012.01.008) discloses a comparative test of the in vitro anti-Toxoplasma effect of three flavonoids of Scutellaria baicalensis, which proves that baicalein can significantly inhibit the growth of Toxoplasma both in vivo and in vitro. Baicalein also has a good inhibitory effect on protozoan parasites such as Leishmania spp. and Cryptosporidium parvum. However, its own solubility is low and its utilization rate is not high.

[0004] In order to further improve the utilization rate of drugs, prolong the duration of drug effects and optimize the dosage of drugs, finding and selecting a suitable drug delivery system has extremely sufficient scientific research value and practical significance. Ginsenosides show good therapeutic effects in a variety of diseases. The invention application with publication number CN 108888628A discloses the use of ginsenoside Rh2 in the preparation of anti-Toxoplasma compound preparations. Through in vitro drug toxicity experiments, in vitro and in vivo anti-Toxoplasma activity experiments, the anti-Toxoplasma effect of ginsenoside Rh2 was compared. Ginsenoside Rh2 has a good Toxoplasma inhibitory effect and has a protective effect on Toxoplasma infected hosts. However, ginsenoside Rh2 has weak toxic side effects.

[0005] In addition, reference 2 (Lixin Du, Huiling Lu, et al. Preparation, characterization and in vivo pharmacokinetic study of ginsenoside Rb 1-PLGA nanopartic1es, Scientific Reports, 2023, 13, 18472, doi: 10.1038 / s41598-023-45858-x) shows that ginsenoside Rb 1 has a binary structure, including a hydrophobic triterpenoid aglycone and a hydrophilic sugar side chain. Nanoprecipitation technology enables ginsenoside Rb 1 to self-assemble into micelles or micelle-like nanoaggregates, with hydrophilic sugar groups aggregated on the outer surface and hydrophobic segments kept isolated inside, thereby forming nanoparticles, enhancing the water solubility of ginsenoside Rb 1, and can be used as a green and non-toxic drug delivery carrier, effectively improving drug efficacy and promoting drug absorption and utilization. Summary of the invention

[0006] The purpose of the present invention is to provide a method for preparing a ginsenoside Rb1 and baicalein nano-preparation, which is used to inhibit the growth of Toxoplasma gondii, improve the utilization rate of traditional anti-Toxoplasma drugs, prolong the service life of drugs, and reduce drug toxicity.

[0007] In order to achieve the above-mentioned object of the invention, the present invention adopts the following technical solutions:

[0008] The present invention provides a method for preparing a ginsenoside Rb1 and baicalein nano preparation, which comprises the following steps:

[0009] (1) preparing ginsenoside Rb1 solution and baicalin solution respectively;

[0010] (2) The ginsenoside Rb1 solution and the baicalein solution prepared in step (1) are mixed, and a nanoprecipitation strategy is adopted to obtain a ginsenoside Rb1 and baicalein nano preparation.

[0011] Preferably, in step (1) of the method for preparing the ginsenoside Rb1 and baicalein nanoformulation, the solvent of the baicalein solution is an organic solvent. The organic solvent can improve the solubility of baicalein and reduce cytotoxicity. More preferably, dimethyl sulfoxide is used as the organic solvent.

[0012] Preferably, in step (2) of the method for preparing the ginsenoside Rb1 and baicalein nanoformulation, the mass ratio of ginsenoside Rb1 to baicalein is 1:6 to 2:1.

[0013] More preferably, in step (2) of the method for preparing the ginsenoside Rb1 and baicalein nanoformulation, the optimal mass ratio of ginsenoside Rb1 to baicalein is 1:1 / 2.

[0014] Preferably, in step (2) of the method for preparing the ginsenoside Rb1 and baicalein nanoformulation, mixing is performed by a vortex and a vertical mixer in a nanoprecipitation. Mixing by a vortex and a vertical mixer can achieve rapid and controllable mixing between solutions, and the nucleation and growth process of nanoaggregates can be regulated by a kinetic method to obtain nanoparticles with smaller particle size and more uniform dispersion.

[0015] Preferably, the vortex mixing is performed for 30 minutes under the action of the vortex.

[0016] Preferably, the vertical mixer is a HS-3 vertical mixer with a rotation speed of 20 rpm for mixing for at least 8 hours.

[0017] Preferably, the preparation method of the ginsenoside Rb1 and baicalein nano-preparation adopts dialysis, and a dialysis bag with a molecular weight cut-off of 1000 is used to obtain the ginsenoside Rb1 and baicalein nano-preparation.

[0018] Organic solvents and other impurities can be removed by dialysis.

[0019] The present invention also provides a ginsenoside Rb1 and baicalin nano preparation prepared by the preparation method.

[0020] The present invention also provides the use of ginsenoside Rb1 and baicalein nano preparation in the preparation of anti-Toxoplasma gondii drugs.

[0021] The present invention also provides an anti-Toxoplasma drug, wherein the active substances comprise the ginsenoside Rb1 and baicalein nano preparation.

[0022] The beneficial effects of the present invention are:

[0023] The preparation method of the ginsenoside Rb1 and baicalein nanoformulation of the present invention adopts nanoprecipitation technology to assemble into a nano co-delivery system, and based on the dynamic regulation of the nucleation and growth process of the nano aggregates in the nanoprecipitation technology, ginsenoside Rb1 encapsulates baicalein to form a nanoformulation with a smaller particle size, the synthesis operation is simple, and it has better targeting in drug applications;

[0024] The prepared ginsenoside Rb1 and baicalin nano-preparation can not only exert the good anti-Toxoplasma effect of baicalin, but also exert the drug solubility of ginsenoside Rb1, greatly improving the drug utilization rate. The encapsulation method prolongs the service life of the drug, and has biosafety and less damage to normal tissues.

[0025] Figure 1 The results of ginsenoside Rb1 (GRb1) and baicalein (BAI) nanoformulations (GRb1 / BAI NPs) are shown.

[0026] Figure 2 The graph shows the particle size results of GRb1 / BAI NPs prepared in each example.

[0027] Figure 3 Scanning electron microscopy images of ginsenoside Rb1 (GRb1), baicalein (BAI) and GRb1 / BAI NPs.

[0028] Figure 4 The figure shows the cytotoxicity results of GRb1 / BAI nanoformulation and BAI solution on human foreskin fibroblasts (HFF) in the concentration range of 6.25 μM-400 μM, where Figure 4 The left figure in the figure shows the cytotoxicity of GRb1 / BAI nanoformulation to HFF cells; Figure 4 The right figure in the figure shows the cytotoxicity of BAI solution to HFF cells.

[0029] Figure 5 is the half inhibitory concentration (IC 50 ), where Figure 5 The left figure shows the half inhibitory concentration (IC 50 ) result graph; Figure 5 The right figure shows the half inhibitory concentration (IC 50 ) result graph.

[0030] Figure 6 This is the in vitro release result of GRb1 / BAI nanoformulation.

[0031] Figure 7 This is the result of BAI solution's anti-Toxoplasma effect in mice.

[0032] Figure 8 This is a graph showing the anti-Toxoplasma results of GRb1 / BAI nanoformulation in mice. DETAILED DESCRIPTION

[0033] The specific implementation of the present invention is further described in detail below in conjunction with the accompanying drawings and examples, but the implementation and protection of the present invention are not limited thereto. It should be noted that if there are any processes that are not particularly described in detail below, they can be implemented or understood by those skilled in the art with reference to the prior art. If the manufacturer of the reagents or instruments used is not indicated, they are deemed to be conventional products that can be purchased commercially.

[0034] Example 1

[0035] A method for preparing a nano preparation of ginsenoside Rb1 and baicalin, the preparation method comprises the following steps:

[0036] (1) Preparation of ginsenoside Rb1 (GRb1) and baicalein (BAI) solutions: Weigh 20 mg of ginsenoside Rb1 standard and add 2 mL of deionized water to obtain a 10 mg / mL ginsenoside Rb1 solution, then dilute with deionized water to obtain a 3.5 mg / mL ginsenoside Rb1 solution. Weigh 40 mg of baicalein standard and dissolve in 2 mL of dimethyl sulfoxide (DMSO) to obtain a 20 mg / mL baicalein solution.

[0037] (2) Preparation of GRb1 / BAI nano-solution: 100 μL of 20 mg / mL baicalein solution was taken, and 100 μL of BAI solution was slowly added to 1.14 mL of 3.5 mg / mL ginsenoside Rb1 solution in portions within 10 min, and then deionized water was added to make up the volume to 2 mL. After being fully mixed for 30 minutes under the action of a vortex, the mixture was mixed overnight in a HS-3 vertical mixer at a speed of 20 rpm for more than 8 hours to obtain a GRb1 / BAI nano-solution.

[0038] (3) Dialysis: The next day, the GRb1 / BAI nanoparticle solution was transferred to a MWCO1000 dialysis bag and dialyzed with 50 mL of deionized water for 6 hours, with the dialysate (ddH2O) replaced every 2 hours. After the dialysis, the solution was aspirated to obtain the GRb1 / BAI nanoparticle preparation, which was stored in a 4°C refrigerator for future use.

[0039] The prepared GRb1 / BAI nanoformulation (GRb1 / BAI NPs) is a uniform yellow solution, namely the GRb1 / BAI nanoformulation. Figure 1 shown.

[0040] Example 2

[0041] A method for preparing a ginsenoside Rb1 and baicalein nanoformulation, the other steps are the same as those of Example 1, except that: in step (2), 50 μμL of a baicalein solution with a concentration of 20 mg / mL is slowly dripped into 1.14 mL of a ginsenoside Rb1 solution with a concentration of 3.5 mg / mL, and then deionized water is added to make up the volume to 2 mL.

[0042] Example 3

[0043] A method for preparing a ginsenoside Rb1 and baicalein nanoformulation, the other steps are the same as those in Example 1, except that: in step (2), 50 μL of a baicalein solution with a concentration of 20 mg / mL is slowly dripped into 1.71 mL of a ginsenoside Rb1 solution with a concentration of 3.5 mg / mL, and then deionized water is added to make up the volume to 2 mL.

[0044] Example 4

[0045] A method for preparing a ginsenoside Rb1 and baicalein nanoformulation, the other steps are the same as those in Example 1, except that: in step (2), 100 μL of a baicalein solution with a concentration of 20 mg / mL is slowly dripped into 1.71 mL of a ginsenoside Rb1 solution with a concentration of 3.5 mg / mL, and then deionized water is added to make up the volume to 2 mL.

[0046] Example 5

[0047] A method for preparing a ginsenoside Rb1 and baicalein nanoformulation is the same as Example 1 except that: in step (2), 200 μL of a baicalein solution with a concentration of 20 mg / mL is slowly dripped into 1.14 mL of a ginsenoside Rb1 solution with a concentration of 3.5 mg / mL, and then deionized water is added to make up the volume to 2 mL.

[0048] Example 6

[0049] A method for preparing a ginsenoside Rb1 and baicalein nanoformulation, the other steps are the same as those of Example 1, except that: in step (2), 200 μμL of a baicalein solution with a concentration of 20 mg / mL is slowly dripped into 1.71 mL of a ginsenoside Rb1 solution with a concentration of 3.5 mg / mL, and then deionized water is added to make up the volume to 2 mL.

[0050] Test Example 1

[0051] The particle size and Zeta potential of the GRb1 / BAI NPs prepared in each example were detected in the following steps:

[0052] (1) Particle size was measured by the German Partical Metrix Zeta view nanoparticle tracking analyzer: GRb1 / BAI NPs were diluted with deionized water at a concentration of 1:1000, and then the diluted solution was injected into the nanoparticle analyzer using a 1 mL sterile syringe;

[0053] The particle size results are shown as Figure 2As shown, the particle size of GRb1 / BAI NPs is about 100 nm (three repeated independent tests were performed), reaching the nanometer level; among them, the particle size of GRb1 / BAI NPs prepared in Example 1 is the smallest, which is 109.7 nm, and has good targeting in disease treatment.

[0054] (2) Zeta potential was measured by BI-200SM / NanoBrook ZetaPALS / BI-DNDC instrument: GRb1 / BAINPs were placed in a 4°C refrigerator for one week. After one week, GRb1 / BAI NPs were diluted with deionized water at a concentration of 1:100 for Zeta value detection. The total solution volume was 2 mL.

[0055] Zeta potential is an important indicator for characterizing the stability of colloidal dispersions. When the absolute value (positive or negative) of the Zeta potential is higher, the system is more stable. On the contrary, when the absolute value of the Zeta potential is lower, the dispersion is destroyed and coagulation or aggregation occurs, and the system is unstable. In addition, the positive and negative values ​​of the Zeta potential also represent the charge properties of the nanoparticle surface.

[0056] The Zeta potential results of the GRb1 / BAI NPs prepared in each example are shown in Table 1:

[0057] Table 1 GRb1 / BAI NPs prepared in each example and their Zeta potential

[0058]

[0059] The Zeta potential results showed that the GRb1 / BAI NPs prepared in this study were all negatively charged, and the absolute values ​​of the Zeta potential were all around 30 (three repeated independent tests were performed), indicating that the prepared GRb1 / BAI NPs system was stable and maintained a stable morphology after being placed in a 4°C refrigerator for a long time; among them, the absolute value of the Zeta potential of the GRb1 / BAI NPs prepared in Example 1 was the largest, at -33.82±0.85 mV, indicating that it had the most stable morphology compared to other examples.

[0060] According to the results of Zeta potential value and particle size, the optimal volume ratio of ginsenoside Rb1 and baicalin BAI was comprehensively selected. The results showed that when the ginsenoside Rb1 solution was 1.14mL and the baicalin BAI solution was 100μL, the Zeta potential value and nanoparticle size were both optimal, and this ratio was used in subsequent tests.

[0061] Test Example 2

[0062] The microstructures of GRb1, BAI and GRb1 / BAI NPs prepared in the examples were characterized by transmission electron microscopy (TEM).

[0063] like Figure 3 The results showed that GRb1 alone was round in shape, BAI alone was rod-shaped, and the prepared GRb1 / BAI NPs were round after combination. Under the same transmission electron scanning microscopy conditions, the particle size of GRb1 / BAINPs was significantly larger than that of GRb1 alone, proving that GRb1 encapsulated and combined BAI to prepare GRb 1 / BAI NPs.

[0064] Test Example 3

[0065] The toxicity of the GRb1 / BAI NPs and BAI solution prepared in Example 1 to human foreskin fibrocytes (HFF) cells specifically includes the following steps:

[0066] (1) HFF cells were seeded in a 96-well plate containing 200 μL of DMEM medium containing 10% fetal bovine serum at a seeding density of 2×10 4 The cells were plated at 37°C in a cell culture incubator with 5% CO2 for 24 hours.

[0067] (2) Experimental group: The next day, GRb1 / BAI NPs and BAI solution were diluted from 400 μM to 6.25 μM, respectively, and added to HFF cells in a 96-well plate. 200 μL of GRb1 / BAI NPs or BAI solution was added to each well, and 3 parallel wells were set for each concentration. In addition, a blank group and a negative control group were set up: the blank group contained only 200 μL of culture medium; each well of the negative control group contained only 2×10 4 The experimental group, blank group and negative control group were placed in a cell culture incubator with 5% CO2 at 37°C for 48 hours.

[0068] (3) After 48 hours, the cell viability was determined using a CCK-8 kit. 10 μL of CCK-8 solution was added to each well and incubated at 37°C for 1 hour. The viability of HFF cells was calculated by measuring the absorbance at 450 nm according to the following formula.

[0069]

[0070] (5) The results of the cytotoxicity of GRb1 / BAI NPs and BAI solution to HFF cells were analyzed using GraphPadPrism 8.0 software.

[0071] like Figure 4The results showed that GRb1 / BAINPs had no cytotoxicity to HFF cells in the concentration range of 6.25 μμM to 400 μμM, whereas BAI solution alone had no cytotoxicity to HFF cells in the same concentration range. 50 The (half toxic concentration) was 143.8 μμ M. The experimental results showed that the toxicity of GRb1 / BAI NPs prepared by ginsenoside Rb1 encapsulation combined with BAI to HFF cells was greatly reduced, and the cytotoxic effect of baicalin BAI on GRb1 / BAI NPs of HFF cells was effectively improved, indicating that GRb1 / BAI NPs had almost no toxic side effects on cells.

[0072] Test Example 4

[0073] The half inhibitory concentration (EC 50 ) to measure the drug effectiveness of the GRb1 / BAI NPs and BAI solution prepared in Example 1, specifically comprising the following steps:

[0074] (1) HFF cells were seeded in a 96-well plate containing 200 μL of DMEM medium containing 10% fetal bovine serum at a seeding density of 2×10 4 cells / well and cultured in a cell culture incubator with 5% CO2 at 37°C for 24 hours.

[0075] (2) Experimental group: The next day, the culture medium in the 96-well plate was discarded, and the plate was gently washed 2-3 times with 10 mM PBS (pH = 7.3-7.5), and the RH strain of Toxoplasma gondii (T. gondii) was infected at a multiplicity of infection (MOI) of 1:1. At the same time, 200 μL of GRb1 / BAI NPs and BAI solution diluted from 400 μM to 6.25 μM were added. At the same time, a positive control group, a negative control group and a blank group were set up: in the positive control group, HFF cells were only infected with the RH strain of Toxoplasma gondii (T. gondii), and the culture medium did not contain drugs; the negative control group was the culture medium and cells without any treatment; the blank control group contained only the culture medium. Three wells were set up in each group as parallel control groups. The experimental group, positive control group, negative control group and blank group were placed in a cell culture incubator with 5% CO2 and cultured at 37°C for 48 hours.

[0076] (3) After 48 hours, the results of each group were detected using a CCK-8 kit. 10 μL of CCK-8 solution was added to each well and incubated at 37°C for 1 hour. The inhibitory rate of the drug against the RH strain of Toxoplasma gondii was calculated according to the following formula by measuring the absorbance at 450 nm.

[0077]

[0078] (4) The half inhibitory concentration (IC) of GRb1 / BAI NPs and BAI solution 50)The results were analyzed using GraphPad Prism 8.0 software.

[0079] like Figure 5 As shown in the figure, the IC of BAI was detected by CCK-8 kit in the concentration range of 6.25μμM to 400μμM. 50 was 88.97 μM, while the IC 50 The experimental results showed that BAI encapsulated by ginsenoside Rb1 inhibited Toxoplasma gondii at a lower concentration, indicating that GRb1 / BAI NPs improved the effectiveness of the drug.

[0080] Test Example 5

[0081] The in vitro release experiment of GRb1 / BAINPs prepared in Example 1 specifically includes the following steps:

[0082] (1) In vitro drug release experiments are an important method to evaluate the rate and extent of drug release from drug preparations under simulated in vivo conditions. Dialysis was used to analyze the drug release of GRb1 / BAI NPs. 1 mL of GRb1 / BAI NPs solution (BAI concentration of 1 mg / mL) was placed in a dialysis bag (MWCO 1000).

[0083] (2) Soak the dialysis bag in 250 mL of deionized water and dialyze at 4°C. Periodically remove 3 mL of dialysis fluid (ddH2O) and replenish the dialysis fluid to 250 mL.

[0084] (3) The content of BAI drug in the dialysate was determined by high performance liquid chromatography, and the dialysis curve was drawn using GraphPadPrism 8.0 software. Each stability curve represents the average of three independent runs with the same sampling.

[0085] like Figure 5 The results showed that it took about 400 hours for GRb1 / BAI NPs to completely release BAI, indicating that the binding of ginsenoside Rb1 to GRb1 / BAI NPs prepared after BAI encapsulation was stable and could effectively extend the service life of the drug.

[0086] Test Example 6

[0087] The anti-Toxoplasma effect of the BAI solution prepared in Example 1 in mice was evaluated by using a mouse survival curve, which specifically includes the following steps:

[0088] (1) 6-8 week old ICR female mice were used as a model and infected with Toxoplasma gondii RH strain (T. gondii), specifically including: randomly dividing the mice into 4 groups, using a parallel design, with 5 mice in each group. Preventive administration was performed by gavage, and 75 mg / kg of BAI solution (medium dose BM group), 100 mg / kg of BAI solution (high dose BH group), positive drug sulfadiazine (SDZ group) and 500 μL 10 mM PBS solution (pH = 7.3-7.5) (NC group) were administered for 5 consecutive days to achieve a stable blood drug concentration in the mice, and then 100 Toxoplasma gondii RH strain tachyzoites were injected into the abdominal cavity of each of the 4 groups of mice.

[0089] (2) Four hours after intraperitoneal injection of the RH strain of Toxoplasma gondii, the mice were treated by gavage: the experimental groups were gavaged with 75 mg / kg of BAI solution (medium-dose BM group), 100 mg / kg of BAI solution (high-dose BH group), the positive drug sulfadiazine (SDZ group), and 500 μL of 10 mM PBS (pH = 7.3-7.5) (NC group), once a day for 7 consecutive days.

[0090] (3) The condition of mice in each group was monitored daily, the date of death of mice was recorded, and the survival curve of mice was drawn using GraphPad Prism 8.0 software.

[0091] like Figure 7 The results showed that the survival curve indicated that the mice in the NC group died within 10 days, and the survival period of mice treated with BAI gavage (medium-dose BM group and high-dose BH group) was comparable to that of mice treated with SDZ gavage, confirming that BAI gavage treatment greatly prolonged the survival time of mice, indicating that BAI has a good anti-Toxoplasma efficacy in mice.

[0092] Test Example 7

[0093] The anti-Toxoplasma effect of the GRb1 / BAI NPs prepared in Example 1 in mice was evaluated by using a mouse survival curve, which specifically includes the following steps:

[0094] (1) 6-8 week old ICR female mice were used as a model and infected with Toxoplasma gondii RH strain (T. gondii), specifically including: the mice were randomly divided into 3 groups, with 5 mice in each group, using a parallel design. Preventive administration was given by gavage, and 500 μL of 10 mg / mL GRb1 / BAI NPs and positive drug sulfadiazine SDZ group were respectively administered for 5 consecutive days, and 500 μL of 10 mM PBS solution (pH = 7.3-7.5) was gavaged in the blank group to allow the drug to reach a stable blood drug concentration in the mouse body, and then 100 Toxoplasma gondii RH strain tachyzoites were injected into the peritoneum of each of the three groups of mice.

[0095] (2) Four hours after intraperitoneal injection of the RH strain of Toxoplasma gondii, the mice were treated by gavage: the experimental group was gavaged with 500 μL of 10 mg / mL GRb1 / BAINPs and 270 μL of 10 mg / mL positive drug sulfadiazine, once a day for 7 consecutive days; the control group was gavaged with 500 μL 10 mM PBS (pH = 7.3-7.5).

[0096] (3) The condition of mice in each group was monitored daily, the date of death of mice was recorded, and the survival curve of mice was drawn using GraphPad Prism 8.0 software.

[0097] like Figure 8 The results showed that the survival curve indicated that GRb1 / BAI NPs and the positive drug sulfadiazine SDZ could prolong the survival period of mice. All the mice in the control group died within 7-9 days, and the mortality rate of mice in the experimental group was greatly reduced, confirming that GRb1 / BAI NPs had good anti-Toxoplasma efficacy in mice.

Claims

1. A method for preparing a nano preparation of ginsenoside Rb1 and baicalin, comprising the following steps: (1) preparing ginsenoside Rb1 solution and baicalin solution respectively; (2) The ginsenoside Rb1 solution and the baicalein solution prepared in step (1) are mixed, and a nanoprecipitation strategy is adopted to obtain a ginsenoside Rb1 and baicalein nano preparation.

2. The method for preparing the ginsenoside Rb1 and baicalein nanoformulation according to claim 1, characterized in that: The solvent of the baicalein solution is an organic solvent.

3. The method for preparing the ginsenoside Rb1 and baicalein nanoformulation according to claim 1, characterized in that: The mass ratio of ginsenoside Rb1 to baicalein is 1:6 to 2:

1.

4. The method for preparing the ginsenoside Rb1 and baicalein nanoformulation according to claim 1, characterized in that: The process of preparing ginsenoside Rb1 and baicalein nanoformulations was mixed by a vortex and a vertical mixer in a nanoprecipitation.

5. The method for preparing the ginsenoside Rb1 and baicalein nanoformulation according to claim 4, characterized in that: The vortex mixing is mixing for 30 minutes under the action of the vortex.

6. The method for preparing the ginsenoside Rb1 and baicalein nanoformulation according to claim 4, characterized in that: The vertical mixer was mixed in a HS-3 vertical mixer at a rotation speed of 20 rpm.

7. The method for preparing the ginsenoside Rb1 and baicalein nanoformulation according to claim 1, characterized in that: Ginsenoside Rb1 and baicalin nanoformulations were obtained by dialysis using a dialysis bag with a molecular weight cutoff of 1000.

8. The ginsenoside Rb1 and baicalin nano-preparation prepared by the preparation method according to any one of claims 1 to 7.

9. Use of the ginsenoside Rb1 and baicalein nanoformulation according to claim 8 in the preparation of anti-Toxoplasma gondii drugs.

10. An anti-Toxoplasma drug, characterized in that: The active substances are the ginsenoside Rb1 and baicalin nano preparations as described in claim 8.

Images