OAB-14 suspension

Abstract

Disclosed is an OAB-14 suspension, which comprises: OAB-14, a stabilizer, and an aqueous solution. The stabilizer is a combined stabilizer, and the combined stabilizer comprises an anionic surfactant and a high-molecular polymer. The mass ratio of OAB-14 to the stabilizer, and moisture in the aqueous solution is 1:(0.08-16):(1.5-28); the particle size d(0.5) of OAB-14 is 1-1000 nm, and d(0.9) thereof is 2-3000 nm. The solubility and the lipid solubility of OAB-14 are increased, the purposes of improving the bioavailability and penetrating the blood-brain barrier are achieved, and the key problem in the development of OAB-14 as a drug is solved.

Description

OAB-14 suspension TECHNICAL FIELD

[0001] The present application relates to the technical field of pharmaceutical preparation, in particular to a OAB-14 suspension. BACKGROUND

[0002] Alzheimer's disease (AD) is a progressive neurodegenerative disease. Clinically, it is characterized by memory impairment, aphasia, apraxia, agnosia, visual-spatial skill impairment, executive function disorder, and personality and behavior changes, and is one of the refractory diseases that seriously threaten the health of the elderly.

[0003] The three major theories of the pathogenesis of Alzheimer's disease are Aβ cascade, abnormal phosphorylation of Tau protein, and cholinergic theory. In addition, there are theories of neural inflammation, abnormal excitation of glutamic acid system, and mitochondrial dysfunction. AD has complex and diverse pathological changes, but neuronal loss, synaptic dysfunction (such as synaptic loss and synaptic plasticity defects), extracellular amyloid beta (Aβ) deposition to form amyloid plaques, and abnormal phosphorylation of Tau protein to form intracellular neurofibrillary tangles are common in AD patients.

[0004] Currently, the FDA has approved several cholinesterase inhibitor drugs for AD, such as donepezil and memantine. However, only Biogen's single antibody drug Aducanumab targeting Aβ has been approved for marketing. Currently, the main research drugs are focused on Aβ amyloid protein, Tau protein, and other targets.

[0005] OAB-14 is a new compound based on the structure of Bexarotene, a β-amyloid protein (Aβ) clearance compound reported in Science magazine. It has poor water and lipid solubility, and a high melting point (340℃). Our company's patent CN113754566A discloses the synthesis process of OAB-14, and CN113896659A, CN116924941A, and CN117567324A disclose the preparation methods of crystal forms A, B, and C of OAB-14, respectively.

[0006] The chemical name of OAB-14 is 4-(3-(2-aminoethyl)ureido)-N-(1,1,4,4,5,5,8,8-octamethyl-1,2,3,4,5,6,7,8-octahydroanthracen-9-yl)benzamide.

[0007] The structural formula of OAB-14 is:

[0008] Molecular formula: C32H46N4O2

[0009] Molecular weight: 518.75

[0010] Chemical Abstracts (CAS) No. 2140911-49-3

[0011] Zhang Xian et al. (Neurochemistry International. Volume 171, Issue. 2023. 105633) found that OAB-14 can promote the clearance of Aβ in the brain through the glymphatic system and improve cognitive dysfunction. Zhang Feng et al. (International immunopharmacology. Volume 124, Issue Pt A. 2023. PP 110911-110911) found that OAB-14 can regulate microglial polarization by regulating PPAR-γ signal, thereby reducing neuroinflammation and improving the cognitive function of AD mice. Guo Xiaoli et al. (ACS chemical neuroscience. Volume 12, Issue 21. 2021. 3985-3993) found that OAB-14 can promote lysosomal Aβ clearance by reducing EAL dysfunction in AD mice. Based on these findings, OAB-14 is a promising new candidate drug for treating AD.

[0012] OAB-14 raw material is stable to high temperature, high humidity and light, and has good chemical stability. However, OAB-14 is insoluble in water, dimethyl sulfoxide or acetonitrile, slightly soluble in ethanol, and slightly soluble in methanol; the solubility in pH 1.2 hydrochloric acid aqueous solution, pH 4.5 acetate buffer, pH 6.8 phosphate buffer and water is 0.027 mg / ml, 0.028 mg / ml, 0 mg / ml and 0.001 mg / ml, respectively, and the solubility is poor. The LgPapp of OAB-14 is 0.209, and the lipid solubility is poor.

[0013] The present application is based on the fact that OAB-14 has a large molecular weight and poor water and lipid solubility, and needs to be prepared into a drug for acting on the brain for the purpose of treating Alzheimer's disease. The present application prepares OAB-14 into a microparticle suspension, which improves the dissolution and lipid solubility of OAB-14, and the drug particle size is small, the particle surface area is increased to improve the dissolution rate of the drug; increase its gastrointestinal adhesion, prolong the gastrointestinal transit time, improve the dissolution and absorption; improve the biodistribution, thereby improving the bioavailability. The OAB-14 suspension of the present application is expected to improve the bioavailability by improving the biodistribution, and improve the effect of treating AD. SUMMARY

[0014] The present application mainly aims at OAB-14 which has poor water solubility and fat solubility, and needs to be prepared into a preparation for acting on the brain for treating Alzheimer's disease, and the type and amount of various stabilizers such as surfactants, high molecular polymers, organic solvents and the like are comprehensively investigated, finally the combined stabilizer composed of anionic surfactants and high molecular polymers is selected, and finally the phospholipid-coated suspension is prepared, the solubility and fat solubility of OAB-14 are increased, the purpose of improving the bioavailability and penetrating the blood-brain barrier is achieved. And the OAB-14 suspension is solidified to further improve the stability of the preparation, so that the suspension can be used as a final preparation, and can also be used as an intermediate to further prepare other forms of preparation.

[0015] The present application provides a kind of OAB-14 suspension, comprising: OAB-14, stabilizer, aqueous solution;

[0016] The stabilizer is a combined stabilizer composed of anionic surfactant and high molecular polymer;

[0017] The mass ratio of OAB-14 to stabilizer and water in aqueous solution is 1:(0.08-16):(1.5-28);

[0018] The particle size d(0.5) of OAB-14 is 1-1000 nm, and d(0.9) is 2-3000 nm.

[0019] Preferably, the high molecular polymer is one or a combination of cellulose derivative polymers and N-vinyl pyrrolidone; and the aqueous solution is one or more of purified water, water for injection, glucose water, distilled water and physiological saline.

[0020] Preferably, the anionic surfactant is SDS; and the high molecular polymer is one or more of HPC, HPMC and PVP.

[0021] Preferably, the combined stabilizer is HPMC and SDS, or HPC and SDS.

[0022] Preferably, the stabilizer is a combined stabilizer composed of anionic surfactant, high molecular polymer and phospholipid and its derivative.

[0023] Preferably, the anionic surfactant is SDS, the high molecular polymer is one or more of HPC, HPMC and PVP, and the phospholipid and its derivative is one or both of soybean phospholipid and dipalmitoyl phosphatidylcholine.

[0024] Preferably, the combined stabilizer is HPC, SDS and soybean phospholipid, or HPMC, SDS and soybean phospholipid, or PVP, SDS, soybean phospholipid and ethanol.

[0025] Preferably, the stabilizer is a combination stabilizer consisting of anionic surfactant, high molecular polymer, phospholipid and its derivatives, and organic solvent.

[0026] Preferably, the anionic surfactant is SDS, the high molecular polymer is one or more of HPC, HPMC and PVP, the phospholipid and its derivatives is one or both of soybean phospholipid and dipalmitoyl phosphatidylcholine, and the organic solvent is one or both of ethanol and glacial acetic acid.

[0027] Preferably, the combination stabilizer is HPC, SDS, soybean phospholipid and glacial acetic acid, or HPMC, SDS, soybean phospholipid and ethanol.

[0028] Preferably, the prepared OAB-14 suspension is used as an intermediate, and a suspending agent is added, the suspending agent being one or more of a combination of sugar alcohols, polysaccharides, gums, cellulose derivative polymers, and N-vinyl pyrrolidones.

[0029] Preferably, the suspending agent is one or more of a combination of HPMC, PVP, gum arabic, gum tragacanth, pregelatinized starch, xanthan gum, sodium carboxymethyl starch, microcrystalline cellulose-sodium carboxymethyl cellulose, glycerol, mannitol, and sorbitol.

[0030] Preferably, a solidification stabilizer is added to the OAB-14 suspension, and a solid powder is prepared after solidification treatment, the solidification stabilizer being one or more of a combination of sugar alcohols, cellulose derivative polymers, and N-vinyl pyrrolidones.

[0031] Preferably, the solidification stabilizer is one or more of a combination of mannitol, lactose, glucose, HPMC, and PVP.

[0032] Preferably, the OAB-14 suspension further comprises one or more of a pH adjuster, a bacteriostatic agent, and an osmotic pressure adjuster.

[0033] Preferably, the pH adjuster is one or more of sodium hydroxide, hydrochloric acid, acetic acid, sodium dihydrogen phosphate, disodium hydrogen phosphate, sodium acetate, citric acid, and sodium citrate.

[0034] The bacteriostatic agent is one or more of phenol, cresol, chlorocresol, benzyl alcohol, and chlorobutanol.

[0035] The osmotic pressure adjuster is one or more of sodium chloride, glucose, and glycerol.

[0036] Preferably, the particle size d(0.5) of the OAB-14 is 1-500 nm, and the particle size d(0.9) is 2-2000 nm.

[0037] In the present application, the amount of the organic solvent and water can be determined according to the actual product requirement. For example, when the required concentration of OAB-14 is 200 mg / ml, 50 mg / ml of the organic solvent and 800 mg / ml of water are required to be correspondingly added.

[0038] The concentration of the solidification stabilizer in the present application can be 0-20% (w / w), preferably 1%-15% (w / w), for example 10% (w / w).

[0039] The present application provides a preparation method of OAB-14 suspension, which comprises the following steps:

[0040] The stabilizer is added to water, and is fully stirred, dissolved or dispersed to obtain an aqueous solution of the stabilizer; the OAB-14 is added to the aqueous solution containing the stabilizer, and is stirred and dispersed to obtain a premix suspension of OAB-14; the premix suspension is poured into a ball mill containing grinding beads, and is ground until the particle size d(0.5) of the OAB-14 raw material is 1-1000 nm and the particle size d(0.9) is 2-3000 nm, and the grinding liquid is collected as the OAB-14 suspension.

[0041] In the present application, the particle size d(0.5) of the OAB-14 raw material is 1-1000 nm, and the particle size d(0.9) is 2-3000 nm; preferably, the particle size d(0.5) is 1-500 nm, and the particle size d(0.9) is 2-2000 nm.

[0042] In the present application, the mass ratio of the OAB-14 to the stabilizer and water is preferably 1:(0.08-16):(1.5-28), more preferably 1:(0.1-12):(1.5-20).

[0043] The rotation speed of the stirring paddle is preferably 100-1000 rpm, for example 200 rpm, and the stirring time is preferably 1-60 minutes, for example 30 minutes.

[0044] In the present application, the ball mill is a wet medium grinding mill, and the grinding process can be realized by simultaneously opening the device circulating cooling water during grinding.

[0045] The grinding rotation speed is 1000-6000 rpm, preferably 1500-4000 rpm, more preferably 2000-3000 rpm, for example 2500 rpm or 3000 rpm.

[0046] The grinding time is 0.5-24 h, preferably 2-16 h, for example 2 h, 6 h, 8 h, 10 h, 12 h or 14 h.

[0047] In the present application, the grinding uses zirconium oxide or stainless steel ball milling beads, the size of the ball milling beads is 0.05-1mm, for example 0.3mm; the volume of the ball milling beads can be 10-10000mL, for example 600mL.

[0048] Compared with the prior art, the present application has the following beneficial technical effects:

[0049] The disclosed OAB-14 medicine exists in a raw material original crystal form or an amorphous form, and a combined stabilizer of an anionic surfactant and a high polymer is used. Compared with a single stabilizer prescription, the particle size d(0.9) of the OAB-14 suspension is greatly reduced, the aggregation speed and the settling speed of the drug particles are also slowed down; compared with the raw material, the dissolution rate is increased by 4 times, and the final dissolution amount is about 20% or so.

[0050] The combined stabilizer of the anionic surfactant, the high polymer and the phospholipid and its derivatives is used, and the dissolution rate of the prepared suspension is increased by 2 times compared with the combined stabilizer of the anionic surfactant and the high polymer, and is increased by 8-9 times compared with the raw material dissolution rate.

[0051] The combined stabilizer of the anionic surfactant, the high polymer, the phospholipid and its derivatives and the organic solvent is used, and the viscosity of the prepared suspension is reduced, the flow state of the suspension is normal in the circulating grinding, and the particle size distribution of the suspension is narrowed.

[0052] The prepared OAB-14 suspension is used as an intermediate, and then a suspending agent is added, which is beneficial to the properties and the particle size stability of the suspension.

[0053] The solidification stabilizer is added to the OAB-14 suspension, and a solid powder is prepared after solidification treatment. The addition of the solidification stabilizer can effectively control the particle size of the raw material after spray drying, and the OAB-14 medicine can be redispersed to obtain a relatively stable suspension after adding water. The powder after solidification can be used as an intermediate, and is convenient for designing into various dosage forms.

[0054] The present application improves the solubility and the fat solubility of OAB-14, achieves the purpose of improving the bioavailability and penetrating through the blood-brain barrier, and solves the key problem of OAB-14 medicine. BRIEF DESCRIPTION OF DRAWINGS

[0055] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the following will briefly introduce the drawings needed to be used in the embodiment or prior art description.

[0056] Fig. 1 is a comparison diagram of the dissolution curves of the OAB-14 suspensions of two stabilizers of the present application and the raw material;

[0057] Figure 2 is a comparison chart of the dissolution curves of OAB-14 suspensions with two stabilizers and three stabilizers of the present application and the raw material;

[0058] Figure 3 is the effect of OAB-14 of the present application on the 1 h preference index in the novel object recognition test of APP / PS1 mice (6 days after administration) (n = 9-10, mean ± SEM);

[0059] Figure 4 is the effect of OAB-14 of the present application on the 1 h preference index in the novel object recognition test of APP / PS1 mice (two weeks after administration) (n = 9-10, mean ± SEM);

[0060] Figure 5 is the effect of OAB-14 of the present application on the total number of arm entries in the Y maze test of APP / PS1 mice (n = 9-10, mean ± SEM);

[0061] Figure 6 is the effect of OAB-14 of the present application on the spontaneous alternation rate in the Y maze test of APP / PS1 mice (n = 9-10, mean ± SEM);

[0062] Figure 7 is the effect of OAB-14 of the present application on the latency in the dark avoidance test of APP / PS1 mice (n = 9-10, mean ± SEM);

[0063] Figure 8 is the effect of OAB-14 of the present application on the number of errors in the dark avoidance test of APP / PS1 mice (n = 9-10, mean ± SEM);

[0064] Figure 9 is a schematic diagram of the novel object recognition test of the present application;

[0065] Figure 10 is a schematic diagram of the Y maze structure of the present application;

[0066] Figure 11 is a schematic diagram of the Morris water maze structure of the present application. DETAILED DESCRIPTION

[0067] The present application is further described in detail by the following examples, which are intended to illustrate but not to limit the present application. It should be noted that for those skilled in the art, several improvements and modifications can be made to the present application without departing from the principles of the present application, and these improvements and modifications also fall within the scope of protection of the present application.

[0068] The present application is based on the fact that OAB-14 has poor water and fat solubility, and needs to be prepared into an oral preparation for acting on the brain for the treatment of Alzheimer's disease. The OAB-14, stabilizer and water are prepared into a suspension, the particle size of the drug is reduced, the surface area of the particles is increased, the dissolution rate of the drug is improved, the gastrointestinal adhesion of the drug is increased, the absorption rate and amount of the drug are increased, the biodistribution is improved, and the bioavailability is improved. The present application will be further described in conjunction with the examples, but the content of the present application is not limited thereto.

[0069] The particle size d(0.1) of OAB-14 raw material (untreated) used in the present application is 6.091 μm, d(0.5) is 18.6 μm, and d(0.9) is 46.67 μm.

[0070] The particle size of OAB-14 suspension is measured by BT9300S laser particle size distribution instrument. The sample preparation for particle size measurement is as follows: about 50 mg of OAB-14 is taken, the light shielding rate of the detector is in the range of 40%-50%, 5 g of water is added, and ultrasonic treatment is performed for 1-2 minutes (ultrasonic power 150 W, ultrasonic frequency: 53 kHz). The determination method is as follows: the instrument pump speed is set to 40%, the cycle time is 60 s, and the determination time is 120 s.

[0071] Dissolution curve determination method: the dissolution determination method (Chinese Pharmacopoeia 2020 edition four parts 0931 second method) is used for measurement, 500 mL of distilled water is used as the dissolution medium, the rotation speed is 100 revolutions per minute, and the temperature is 37±0.5℃. Samples are taken at 5, 10, 15, 30, 45, 60, 90 and 120 minutes, the absorbance value is determined by ultraviolet spectrophotometry, the cumulative dissolution of the drug at each time point is calculated, and the dissolution curve is drawn with time (min) as the abscissa and dissolution rate (%) as the ordinate.

[0072] Examples 1-10

[0073] In this experiment, single stabilizers are screened. The screened stabilizers include non-ionic surfactants (poloxamer, carbomer, tween, soluplus), anionic surfactants (sodium dodecyl sulfate SDS, sodium docusate) and high molecular polymers (PVP, copolyvidone, HPC, HPMC), and the formula is shown in Table 1.

[0074] Table 1 Formulation of single stabilizer OAB-14 suspension

[0075] Preparation method: the stabilizer was added into purified water to prepare an aqueous solution, and the blank aqueous solution was used as a control. OAB-14 was added, and the drug and the solution were mixed uniformly by magnetic stirring to prepare a crude suspension. The Mini-Easy grinder was used, and zirconium oxide beads with a particle size range of 0.6-0.8 mm were selected as the grinding medium. The fixed rotation speed was 3500 rpm, and the grinding time was 2 h to obtain the OAB-14 suspension. The particle size of the suspension was detected, and the settling speed was observed. After one week, the drug solution was shaken again to form a suspension state, and the particle size was detected. The results are shown in Table 2.

[0076] Table 2 Particle size and property observation results of suspensions prepared by single stabilizer screening

[0077] From the results, compared with the untreated raw material, the suspension prepared in Comparative Example 1 had smaller drug particle size, but it was difficult to achieve and stabilize d(0.9)≤3000 nm. The particle size of the suspension prepared by adding one stabilizer was smaller than that of Comparative Example 1, but it was still difficult to achieve and stabilize d(0.9)≤3000 nm, and the drug particles aggregated quickly during storage, and the physical stability was poor. One stabilizer cannot obtain the required OAB-14 suspension.

[0078] Examples 11-13

[0079] To further reduce the particle size of the OAB-14 suspension and increase the stability of the particles, anionic surfactant and high molecular polymer were used for experiments. According to the experimental enlightenment of Examples 1-10, high molecular polymers HPC, HPMC and PVP were preferred, and anionic surfactant SDS was used for complex experiments. The formula is shown in Table 3. The preparation method is the same as that of Examples 1-10. After preparation, the particle size of the suspension was detected, and the dissolution curve was compared with the raw material. After one week, the properties and particle size were detected, and the results are shown in Table 4 and Figure 1.

[0080] From the results, compared with the single stabilizer prescription, the particle size d(0.9) of the OAB-14 suspension was greatly reduced, the aggregation speed and settling speed of the drug particles were also slow, and the dissolution rate was increased by 4 times compared with the raw material, and the final dissolution amount was about 20%. Combined with the particle size stability and dissolution curve of the OAB-14 suspension, HPMC / SDS and HPC / SDS were preferred as the stabilizer combination.

[0081] Table 3 Formula of OAB-14 suspension prepared by two stabilizers

[0082] Table 4 Particle size and property observation results of suspensions prepared by two stabilizers

[0083] Examples 14-17

[0084] To further improve the dissolution and dissolution rate of OAB-14 suspension and increase the liposolubility of OAB-14, non-ionic surfactant phospholipid: soybean phospholipid, dipalmitoyl phosphatidylcholine (DPPC) was added in the prescription, see Table 5. The preparation method is the same as Examples 1-10. The prepared OAB-14 suspension was tested for particle size and dissolution curve, and the results are shown in Table 6 and Figure 2.

[0085] Table 5 Formulation of OAB-14 suspension prepared with three stabilizers

[0086] From the results, compared with two stabilizers, the particle size of the suspension prepared with three stabilizers did not change significantly. However, the dissolution of the suspension prepared with three stabilizers was 2 times higher than that of the suspension prepared with two stabilizers, and 8-9 times higher than that of the raw material. The third stabilizer is preferably soybean phospholipid. However, the addition of phospholipid makes the suspension more viscous, and the circulation flowability in the equipment pipeline is poor.

[0087] Table 6 Effect of particle size of suspension prepared with three stabilizers

[0088] Examples 18-20

[0089] To improve the flowability of the suspension during production and further increase the wettability, organic solvents (ethanol, glacial acetic acid) were added for experiment, see Table 7.

[0090] Example 18 Preparation method: Put OAB-14 into glacial acetic acid, stir at 40°C for 10 min, and disperse to prepare the organic phase; Put soybean phospholipid into distilled water, disperse uniformly in a 45°C water bath, and cool to room temperature. Then add hydroxypropyl cellulose, stir uniformly, cool in a 4°C refrigerator, and prepare the water phase; quickly inject the organic phase into the water phase under magnetic stirring, stir uniformly, and prepare the pre-suspension; pour the coarse suspension into a Mini-Easy nano-mill filled with 400g 0.2-0.3mm zirconium oxide beads, and grind at 3500rpm for 2h.

[0091] Example 19-20 Preparation Method: Dissolve hydroxypropyl methyl cellulose in 70-90°C hot purified water with stirring to obtain a binding solution; dissolve soybean phospholipid in ethanol; then add the soybean phospholipid ethanol solution to the binding solution, and then add sodium dodecyl sulfate, stirring to dissolve, to obtain a stabilizer solution. Then add OAB-14 and stir to form a coarse suspension. Use a Sichuan Ruishi PNM05 ball mill, select zirconium oxide beads with a particle size of 0.2-0.3 mm as the grinding medium, grind at a main machine speed of 2000 rpm and a feed rate of 250 g / min for 2 h, and then grind at a main machine speed of 2800 rpm and a feed rate of 500 g / min for 12 h, controlling the temperature of the grinding liquid to not exceed 45°C during grinding, to obtain an OAB-14 suspension.

[0092] Observe the flowability of the suspension during grinding, and after grinding, detect the viscosity of the suspension (use a DV2TLV viscometer, use a No. 31 rotor to detect the viscosity of the sample solution at a speed of 60 rpm) and the particle size of the raw material; then place the suspension at room temperature for 10 days, and detect the particle size of the raw material of Examples 18-19. The detection results are shown in Tables 8-9.

[0093] From the results, it can be seen that after adding the organic solvent, the viscosity of the suspension decreases, and the flow state is normal during circulation grinding; the more the amount of organic solvent, the narrower the particle size distribution. The OAB-14 suspension prepared in Examples 18 and 19 is stable in shape within 7 days at room temperature, but the particle size of Example 18 increases faster. However, too much organic solvent increases the cost, equipment corrosion and safety risk, and the amount added should meet the normal operation of production.

[0094] Table 7 Formulation of OAB-14 suspension compounded with four kinds of stabilizers

[0095] Table 8 Effect of particle size of suspension prepared by compounding four kinds of stabilizers

[0096] Table 9 Stability of OAB-14 suspension placed at room temperature for 10 days

[0097] Examples 21-25

[0098] The OAB-14 suspension is a thermodynamically unstable system, and the particles will increase in size and precipitate after being placed for more than 7 days. To obtain a suspension with better physical stability, the OAB-14 suspension prepared in Example 19 is used as an intermediate, and a suspending agent is added to increase the viscosity to slow down the settling speed and resist the cohesive force of the raw material particles.

[0099] Stabilizers HPMC, microcrystalline cellulose-sodium carboxymethylcellulose, PVP, pregelatinized starch: sodium carboxymethyl starch (weight ratio 9:2), glycerol: mannitol: xanthan gum (weight ratio 50:8.6:1.2) were respectively dissolved in hot water to prepare 12%, 4%, 6%, 22%, 22% solution. Respectively mixed with the OAB-14 suspension prepared in Example 19 at 1:1, stirred and passed through colloid mill to mix evenly, the raw material particle size was detected; placed for 3 months, appearance inspection was carried out and the OAB-14 raw material particle size was detected, and the results are shown in Table 10.

[0100] From the results, it can be seen that the addition of suspending agent to the OAB-14 suspension is beneficial to the properties of the suspension and the stability of the particle size.

[0101] Table 10 Effect of adding suspending agent on the particle size of the suspension

[0102] Examples 26-35

[0103] In order to ensure the long-term stability of the OAB-14 suspension, solidification stabilizers were added, and powders or granules were prepared after solidification treatment.

[0104] Take 30 g of the OAB-14 suspension prepared in Example 18 (about equivalent to 1.4 g of OAB-14), 5 parts; add 1.4 g of mannitol, lactose, glucose, HPMC and PVP respectively to 5 parts of the suspension, stir at a magnetic stirring speed of 200 rpm for 2 h, and obtain the solidification liquid, which is marked as Example 26, 27, 28, 29 and 30 respectively.

[0105] Take 50 g of the OAB-14 suspension prepared in Example 19 (about equivalent to 10 g of OAB-14), 5 parts; dissolve 10 g of mannitol, lactose, glucose, HPMC and PVP respectively in 50 g of water, and then add to 5 parts of the suspension, stir at a magnetic stirring speed of 200 rpm for 10 min, and obtain the solidification liquid, which is marked as Example 31, 32, 33, 34 and 35 respectively.

[0106] Examples 26-35 and Examples 18, 19 suspensions each 50 g were spray dried by using Buchi-Mini-B290 spray dryer. The parameters were: inlet temperature 130°C, feed rate 20 rpm, air flow 0.4 m 3 / min. OAB-14 dry powder (i.e. the products of Examples 26-35) was obtained.

[0107] Resuspension operation: take 1 g of the spray-dried powder, add 30 g of water, and ultrasonic for 1-2 minutes (ultrasonic power 150 W, ultrasonic frequency: 53 kHz), to obtain the resuspended product, and sample detection particle size, and the results are shown in Table 11.

[0108] From the results, the OAB-14 suspension directly sprayed dry powder cannot be reconstituted into a uniform cloud-like fine suspension, and the particle size of the raw material becomes large; the addition of solidification stabilizer can effectively control the particle size of the raw material in the powder after spraying, and after adding water, it can be reconstituted, but the effect of the high molecular stabilizer on controlling the particle size is better than that of the sugar alcohol.

[0109] Table 11 Particle size results of OAB-14 suspension with solidification stabilizer after spraying and reconstituting

[0110] Example 36

[0111] Pharmacokinetic and pharmacodynamic studies were conducted on OAB-14 suspension and raw materials, OAB-14 fat emulsion.

[0112] Pharmacokinetic study of comparative experiment 1

[0113] 1) Experimental materials

[0114] Experimental drugs:

[0115] OAB-14 suspension was added with mannitol at a ratio of OAB-14: mannitol = 1:1 and spray dried. An appropriate amount of spray dried powder was weighed, water for injection was added, and it was fully dissolved to prepare a test solution OAB-SNP containing OAB-14 at a concentration of 14 mg / ml.

[0116] Fat emulsion: (1) Preparation of water phase: injectable glycerol (2.5 g) and poloxamer F68 (0.4 g) were dispersed in an appropriate amount of water for injection, heated to 70°C in a magnetic stirrer, and stirred until completely dissolved and dispersed in an appropriate amount of water for injection. (2) Preparation of oil phase: OAB-14 (0.3 g) and phospholipid (3.0 g) were added to 16 ml of anhydrous ethanol (weighed), and a phospholipid complex was prepared by magnetic stirring at 60°C water bath for 0.5 h. Then, oleic acid (0.06 g) and injectable medium-chain triglyceride (MCT 20 g) were added, and anhydrous ethanol was removed by rotary evaporation at 50°C to obtain the oil phase. (3) Preparation of primary emulsion: under high-speed shearing (10000 rpm), the oil phase was slowly added to the water phase, and high-speed shearing was performed for about 2 minutes to obtain the primary emulsion. (4) Preparation of final emulsion: the primary emulsion was adjusted to pH 6.0 with 0.1 mol / L sodium hydroxide, diluted with water for injection to the prescribed volume, and transferred to a high-pressure homogenizer. The primary pressure valve was adjusted to 600 bar, and homogenization was performed for 6 times. (5) Sterilization: bottle, seal the tank with nitrogen. 121°C hot steam sterilization for 8 min, ice water bath rapid cooling, ready. The prepared OAB-Emu has a particle size of 180 nm and a concentration of 3 mg / ml.

[0117] OAB-14 raw material: 7 g of OAB-14 was weighed and placed in a mortar, 20 g of 0.5% CMC-Na was added and ground thoroughly, then transferred to a container, the mortar was washed 4 times with 0.5% CMC-Na, the washing liquid was transferred to the container, and 0.5% CMC-Na was added to make up to 500 ml, shaken well, and prepared into a raw material suspension with a concentration of 14 mg / ml.

[0118] Experimental animals: 24 SPF Sprague-Dawley rats, 7-9 months old, half male and half female, weighing 180±20 g.

[0119] Experimental instruments: Hitachi high-performance liquid chromatograph (UV-VIS detector, L7100 pump, D7000-HSM workstation) from Japan's Tianmei Company, SHIMADZU LCMS-8050 mass spectrometer

[0120] 2) Methods and results

[0121] 24 SD rats, 12 males and 12 females, were randomly divided into 4 groups, 6 rats in each group, and were divided into raw material gavage group, emulsion tail vein injection group, emulsion gavage group and OAB-SNP gavage group. Before and after drug administration 0.083 h (only for emulsion tail vein injection group), 0.25 h, 0.5 h, 1 h, 2 h, 4 h, 6 h (not for emulsion tail vein group), 8 h, 12 h, 24 h, 48 h, 72 h, 96 h, about 0.3 ml of blood was taken from the jugular vein, anticoagulated with EDTA-k2, and within 2 h after collection, centrifuged at 4℃, 2600g for 10 min, the supernatant was collected to obtain plasma samples, and the concentration of OAB-14 in plasma was detected by LC-MS / MS method, and the blood drug concentration was calculated by WinNonlin 6.4, the results are shown in Table 13. From the data analysis, the bioavailability of OAB-14 emulsion (OAB-Emu) and OAB-14 suspension (OAB-SNP) in rats after gavage of raw material was 438.87% and 217.91%, respectively, and the bioavailability of OAB-14 suspension in rats after gavage of raw material was increased by about 2 times. The peak time Tmax of blood drug concentration was OAB-14 emulsion gavage group < raw material gavage group < OAB-14 suspension gavage group, indicating that OAB-14 suspension is more easily absorbed than emulsion when gavaged.

[0122] Table 13 Pharmacokinetic parameters of OAB-14 in SD rats after administration of each test product (x±s, n=6) Note: * indicates n=5.

[0123] Pharmacodynamic study of comparative experiment 2

[0124] Pharmacodynamics The AD model of APP / PS1 transgenic mice was used to preliminarily investigate the anti-AD effect of OAB-14 raw material, OAB-14 emulsion and OAB-14 suspension by Y maze, new object discrimination, Morris water maze and passive avoidance (step-through) experiments.

[0125] 1) Experimental materials

[0126] Experimental drug: same as Comparative Experiment 1

[0127] Experimental animals: 70-month-old APP / PS1 transgenic AD model mice, 10 wild-type mice from the same litter as blank controls, half male and half female, all clean grade, body weight 20-40 g.

[0128] Experimental instruments: same as Comparative Experiment 1

[0129] Methods and results

[0130] The APP / PS1 mice were randomly divided into a model group, a raw material OAB-14 200 mg / kg (prepared with peanut oil as a solvent to the required concentration) group, an OAB-14 suspension 80 mg / kg group (prepared with double distilled water as a solvent to the required concentration), an OAB-14 emulsion 80 mg / kg group, a tail vein injection OAB-14 emulsion 1 mg / kg, 5 mg / kg and 25 mg / kg dose group, 10 in each group, half male and half female. After 21 days of adaptive feeding of the animals in each group, the drug administration was started, and the administration was continued for one month. The animals were subjected to a new object discrimination experiment on the 6th day after drug administration to preliminarily observe the degree of cognitive impairment of the model animals and the effectiveness of each administration group; the Y maze experiment was performed on the 18th day after drug administration; the new object discrimination experiment was performed on the 19th-21st day; the Morris water maze experiment was performed on the 22nd-26th day; the passive avoidance (step-through) experiment was performed on the 27th-28th day; the drug administration was continued during the behavior experiments until the animals were sacrificed.

[0131] Sampling: all mice were divided into two parts, and blood was taken by enucleation first, and then sampling was performed by two different methods, among which 30 mice were perfused for brain and 28 mice were directly decapitated for brain.

[0132] I. Blood sampling and plasma preparation: each mouse was anesthetized by intraperitoneal injection of 0.5% sodium pentobarbital (50 mg / kg) according to body weight. The eyeball was quickly removed with tweezers, and the blood flowed from the orbit into an EP tube coated with heparin. The heart was pressed to speed up the blood pumping to obtain more blood. The blood was centrifuged at 4°C, 4000 rpm for 10 min, and the supernatant was collected and stored in a -80°C refrigerator.

[0133] Ⅱ. Perfusion and brain collection: After the blood collection, the mice were fixed on their back, and the chest cavity was opened from the xiphoid. The infusion needle was inserted into the left heart apex immediately after the right auricle was cut off. The blood was flushed with pre-cooled PBS. After 50 mL was injected, the tail tip was checked to determine if the blood was flushed. After the blood was flushed, the mice were decapitated, and the brains were immersed in Golgi staining solution for Golgi staining.

[0134] Ⅱ. Direct brain collection: The mice were directly decapitated, and the hippocampus and cortex were quickly separated on the ice box after the brain tissue was removed. The liver, kidney, and intestine of the mice were removed and stored in a -80°C refrigerator.

[0135] As shown in FIG. 9, the experimental method and results of the novel object recognition test are as follows:

[0136] The experimental device is a wooden open square open field with a side length of 44 cm and a height of 13 cm. The objects used in the experiment are all new to the animals. It takes advantage of the natural exploratory nature of rodents in new and unfamiliar environments. The ability of image recognition and memory is investigated by comparing the exploration time of new and old objects. The novel object recognition test is divided into an adaptation stage and a test stage. In the adaptation stage, two mice are placed in the open field at a time, and they are allowed to freely explore the open field for 3 minutes to adapt to the environment, twice a day for two days. On the test day, one mouse is placed in the open field and allowed to freely explore for 3 minutes, then the mouse is removed, and two identical objects (A1, A2) are placed in the open field. The mouse is placed in the middle of the two objects, and the time spent exploring the two objects (tA1, tA2) is recorded within 5 minutes. After 1 hour, object A2 is replaced with a new object B, and the mouse is placed again. The time spent exploring the two objects (tA1', tB) is recorded. The preferential index (Preferential index) and discrimination index (Discrimination index) of the new object are calculated.

[0137] The preferential index calculation formula is as follows: Preferential index (1h) = tB / (tA1'+tB)

[0138] The discrimination index calculation formula is as follows: Discrimination index (1h) = (tB-tA1') / (tA1'+tB)

[0139] The results of novel object recognition tests (6 days after administration) showed that, compared with the blank group, the preference index and discrimination coefficient of the model group mice for new objects in the 1 h test stage were significantly reduced, suggesting that the model group mice had decreased visual discrimination memory ability; compared with the model group mice, the preference index and discrimination coefficient of the oral original drug type, suspension type, emulsion type, and tail vein injection of 5 mg / kg and 25 mg / kg OAB-14 group mice for new objects in 1 h were significantly increased (see Figure 3), suggesting that oral administration of OAB-14 and tail vein injection of OAB-14 both improved the visual discrimination memory disorder of APP / PS1 mice, and the effect of oral administration of OAB-14 on improving the visual discrimination memory disorder of APP / PS1 mice was stronger than that of tail vein injection of OAB-14.

[0140] The results of novel object recognition tests (two weeks after administration) showed that, compared with the blank group, the preference index and discrimination coefficient of the model group mice for new objects in the 1 h test stage were significantly reduced, suggesting that the model group mice had significantly decreased visual discrimination memory ability; compared with the model group mice, the preference index and discrimination coefficient of the oral original drug type, suspension type, emulsion type, and tail vein injection of 5 mg / kg and 25 mg / kg OAB-14 group mice for new objects in 1 h were significantly increased (see Figure 4), suggesting that oral administration of OAB-14 and tail vein injection of OAB-14 both improved the visual discrimination memory disorder of APP / PS1 mice, and the effect of oral administration of OAB-14 on improving the visual discrimination memory disorder of APP / PS1 mice was stronger than that of tail vein injection of OAB-14.

[0141] Y maze test experimental methods and results:

[0142] As shown in Figure 10, the experimental device is a Y-shaped wooden maze, and any two arms form a 120° angle. The white area in the middle of the maze is the boundary of the white area, and when the mouse's body crosses the white area boundary, it is considered that the mouse enters the arm. Each arm is marked with A, B, and C for easy memory. Like the novel object recognition test, it also takes advantage of the natural habit of rodents to explore new and unfamiliar environments, and does not require animals to learn any rules to seek benefits and avoid harm, and can effectively reflect the animal's recognition and memory ability for new and unfamiliar environments. During the experiment, the mouse was placed at the end of arm A with its back to the central part of the maze, and allowed to freely enter and exit the three arms. The sequence of entering the three arms in 5 minutes was recorded, and a correct alternation was recorded once the mouse entered three different arms in succession. The number of correct alternations and the total number of arm entries (N) were recorded. The spontaneous alternation rate was calculated to reflect the mouse's spatial working memory ability.

[0143] Spontaneous alternation response rate (%) = number of correct spontaneous alternation responses / (total number of entries into arms - 2) x 100%.

[0144] The experimental results show that the total number of times that the mice entered the three arms of the Y maze did not show significant differences between the groups (see Figure 5), indicating that the drugs did not affect the spontaneous activity of the mice. The results of the spontaneous alternation response rate showed that, compared with the sham operation group, the spontaneous alternation response rate of the mice in the model group was significantly reduced, indicating that the working memory ability of the mice in the model group was weakened; compared with the model group, the spontaneous alternation response rates of the mice in the oral original drug type, suspension type, emulsion type, and tail vein injection of 5 mg / kg and 25 mg / kg OAB-14 groups were significantly increased (see Figure 6), indicating that oral administration of OAB-14 and tail vein injection of OAB-14 both have the ability to improve the working memory impairment of APP / PS1 mice, and the effect of oral administration of OAB-14 on improving the working memory impairment of APP / PS1 mice is stronger than that of tail vein injection of OAB-14.

[0145] Morris water maze experimental method and results:

[0146] As shown in Figure 11, the experimental device is a circular stainless steel pool, the bottom and walls of the pool are black, and the pool is divided into four virtual quadrants. A circular black platform with a diameter of 10 cm and a height of 23.5 cm is placed in the center of the target quadrant (fourth quadrant), and its position remains unchanged during directional navigation. The water level in the pool is about 1.5 cm higher than the top of the platform, and the water temperature is controlled at (22±1) C. The reference objects around the maze remain unchanged during the experiment, a camera is installed above the maze to record the movement trajectory of the mouse synchronously, and the experiment is divided into two parts:

[0147] (1) Directional navigation experiment: the animal is placed in the water facing the pool wall at the beginning of the experiment, and the time it takes to find the platform is recorded, which is the escape latency. The animal automatically stops collecting after 10 seconds on the platform. If the animal does not find the platform within 60 seconds, the escape latency is recorded as 60 seconds, and the mouse is guided to the platform to stay for 10 seconds. Directional navigation is trained twice a day.

[0148] (2) Probe test: after the directional navigation experiment, the platform is removed, and the animal is placed in the pool to freely explore for 60 seconds. The time, distance, and number of times the animal swims in the original safe quadrant within the specified time are recorded.

[0149] The results of the directional navigation experiment showed that there was no significant difference in swimming speed among the mice in each group in the first experiment, which could rule out the influence of inconsistent swimming speed on escape latency and total swimming distance. Compared with the sham operation group, the escape latency and total swimming distance of the model group mice significantly increased from the second day to the fourth day of directional navigation, indicating that the spatial memory ability of the model group mice was weakened. Compared with the model group, the escape latency of the mice in the oral administration of the original drug type, suspension type and emulsion type OAB-14 groups to reach the platform significantly decreased on the third day of directional navigation; the total swimming distance of the mice in the oral administration of the emulsion type OAB-14 group to reach the platform significantly decreased; the escape latency and total swimming distance of the mice in the tail vein injection of OAB-14 group to reach the platform showed no significant difference. On the fourth day of directional navigation, the escape latency and total swimming distance of the mice in the original drug type, suspension type and emulsion type OAB-14 groups to reach the platform significantly decreased; the total swimming distance of the mice in the tail vein injection of OAB-14 5mg / kg group to reach the platform significantly decreased, and the escape latency showed no significant difference; the escape latency and total swimming distance of the mice in the tail vein injection of OAB-14 1mg / kg and 25mg / kg groups to reach the platform showed no significant difference.

[0150] The results of the spatial exploration experiment showed that compared with the sham operation group, the platform quadrant swimming time, platform quadrant swimming distance percentage and platform crossing times of the model group mice significantly decreased, indicating that the spatial memory reproduction ability of the model group mice was significantly reduced. Compared with the model group, the platform quadrant swimming time, swimming distance percentage and platform crossing times of the mice in the oral administration of the original drug type, suspension type, emulsion type and tail vein injection of 25mg / kg OAB-14 groups significantly increased. It is indicated that oral administration of OAB-14 and tail vein injection of OAB-14 both have the effect of improving the spatial memory impairment of APP / PS1 mice, and the effect of oral administration of OAB-14 on improving spatial memory impairment is stronger than that of tail vein injection of OAB-14.

[0151] Mouse passive avoidance experiment (dark avoidance experiment) experimental method and results

[0152] The experimental device has a light room and a dark room, and there is an arched door between the two rooms. The bottom of the two rooms is paved with copper grids, and the rear part of the dark room can be electrified, and the voltage intensity is controlled by a voltage stabilizer. The experiment is divided into training and testing two parts. In the training stage, the mouse face is placed in the light room with the door facing away, and it is adapted to the environment for 3 minutes, and then electrified with a voltage of 36V. The mouse enters the dark room immediately and is shocked, then escapes from the door back to the light room, and such training is carried out for 5 minutes, and the number of times the mouse is shocked, i.e. the number of errors, is recorded as the learning score. 24 hours later, the test is recorded. The latency period for the first time to enter the dark room and the number of errors within 5 minutes are recorded as the memory score.

[0153] The experimental results show that, compared with the sham operation group, the model group mice have a significantly reduced latency period for entering the dark room and a significantly increased error number, indicating that the long-term memory ability of the model group mice is decreased. Compared with the model group, the oral administration of the OAB-14 raw material type, the suspension type, the emulsion type and the tail vein injection of 25 mg / kg OAB-14 significantly increases the latency period for the mice to enter the dark room and significantly reduces the error number (see Figures 7 and 8). This indicates that the oral administration of OAB-14 and the tail vein injection of OAB-14 both have the ability to improve the long-term memory disorder of APP / PS1 mice, and the oral administration of OAB-14 has a stronger effect on improving the long-term memory disorder of APP / PS1 mice than the tail vein injection of OAB-14.

[0154] Conclusion of the behavioral experiment of Example 36

[0155] The Y maze, new object discrimination, Morris water maze and dark avoidance and a series of behavioral experiment results show that the oral administration of the OAB-14 raw material type, the suspension type, the emulsion and the tail vein injection of 25 mg / kg OAB-14 can significantly improve the working memory disorder, spatial learning and memory disorder and non-spatial learning and memory disorder such as image discrimination of APP / PS1 mice.

[0156] The efficacy of the oral administration of each group is better than that of the intravenous injection group; the efficacy of the oral administration of 80 mg / kg OAB-14 suspension and emulsion is equivalent to that of the oral administration of 200 mg / kg OAB-14 raw material.

[0157] According to the results of the pharmacokinetic study, the blood drug concentration of the intravenous injection of OAB-14 1, 5 and 25 mg / kg is much higher than that of the oral administration. However, the effect of the oral administration of the OAB-14 raw material type, the suspension and the emulsion on improving the learning and memory disorder of APP / PS1 mice is stronger than that of the tail vein injection of OAB-14, and the oral administration of OAB-14 is a reasonable administration route.

[0158] The results of the pharmacodynamics and pharmacokinetic study of Example 36 show that OAB-14 should be prepared into an oral preparation. The small particle size of the drug particles in the suspension increases the rapid dissolution of the drug in the gastrointestinal tract and promotes absorption; the high adhesion of the drug particles prolongs the retention of the drug in the gastrointestinal tract, which not only increases the dissolution amount and improves the bioavailability, but also increases the opportunity to enter the lymphatic system and improves the biological distribution. To achieve the same therapeutic effect, the oral dose of the OAB-14 suspension is 0.4 times that of the raw material.

[0159] The above specific embodiments are only several optional embodiments of the present application, and based on the technical solutions of the present application and the related inspiration of the above embodiments, those skilled in the art can make various alternative improvements and combinations on the above specific embodiments.

Claims

1. [Amended according to Rule 26 06.08.2025] OAB-14 suspension characterized in that, It comprises OAB-14, stabilizer, aqueous solution; The stabilizer is a combined stabilizer comprising an anionic surfactant and a high molecular polymer; The mass ratio of the OAB-14 to the stabilizer and water in the aqueous solution is 1:(0.08-16):(1.5-28); The particle size d(0.5) of the OAB-14 is 1-1000 nm, and the particle size d(0.9) is 2-3000 nm.

2. The OAB-14 suspension of claim 1, wherein, The high molecular polymer is one or a combination of a cellulose derivative polymer and an N-vinyl pyrrolidone.

3. The OAB-14 suspension of claim 2, wherein, The anionic surfactant is SDS, and the high molecular polymer is one or a combination of HPC, HPMC and PVP.

4. The OAB-14 suspension of claim 3, wherein, The combined stabilizer is HPMC and SDS, or HPC and SDS.

5. The OAB-14 suspension of claim 1, wherein, The stabilizer is a combined stabilizer comprising an anionic surfactant, a high molecular polymer and a phospholipid and derivatives thereof.

6. The OAB-14 suspension of claim 5, wherein, The anionic surfactant is SDS, the high molecular polymer is one or a combination of HPC, HPMC and PVP, and the phospholipid and derivatives thereof is one or both of soybean phospholipid and dipalmitoyl phosphatidyl choline.

7. The OAB-14 suspension of claim 6, wherein, The combined stabilizer is HPC, SDS and soybean phospholipid, or HPMC, SDS and soybean phospholipid.

8. The OAB-14 suspension of claim 5, wherein, The stabilizer is a combined stabilizer comprising an anionic surfactant, a high molecular polymer, a phospholipid and derivatives thereof and an organic solvent.

9. The OAB-14 suspension of claim 8, wherein, The anionic surfactant is SDS, the high molecular polymer is one or a combination of HPC, HPMC and PVP, the phospholipid and derivatives thereof is one or both of soybean phospholipid and dipalmitoyl phosphatidyl choline, and the organic solvent is one or both of ethanol and glacial acetic acid.

10. The OAB-14 suspension of claim 9, wherein, The combined stabilizer is HPC, SDS, soybean phospholipid and glacial acetic acid, or HPMC, SDS, soybean phospholipid and ethanol, or PVP, SDS, soybean phospholipid and ethanol.

11. The OAB-14 suspension of any one of claims 1-10, wherein, The prepared OAB-14 suspension is used as an intermediate, and a suspending agent is added, wherein the suspending agent is one or a combination of a sugar alcohol, a polysaccharide, a gum, a cellulose derivative polymer and an N-vinyl pyrrolidone.

12. The OAB-14 suspension of claim 11, wherein, The suspending agent is one or a combination of HPMC, PVP, gum arabic, gum tragacanth, pregelatinized starch, xanthan gum, sodium carboxymethyl starch, microcrystalline cellulose-sodium carboxymethyl cellulose, glycerol, mannitol and sorbitol.

13. The OAB-14 suspension of any one of claims 1-10, wherein, A solidifying stabilizer is added to the OAB-14 suspension, and a solid powder is prepared after solidification treatment, wherein the solidifying stabilizer is one or a combination of a sugar alcohol, a cellulose derivative polymer and an N-vinyl pyrrolidone.

14. The OAB-14 suspension of claim 13, wherein, The solidifying stabilizer is one or a combination of mannitol, lactose, glucose, HPMC and PVP.

15. The OAB-14 suspension of any one of claims 1-10, wherein, The OAB-14 suspension further comprises one or more of a pH regulator, a bacteriostatic agent and an osmotic pressure regulator.

16. The OAB-14 suspension of claim 15, wherein, The pH regulator is one or more of sodium hydroxide, hydrochloric acid, acetic acid, sodium dihydrogen phosphate, disodium hydrogen phosphate, sodium acetate, citric acid and sodium citrate. The bacteriostat is one or more of phenol, cresol, chlorocresol, benzyl alcohol, and chlorobutanol; The osmotic pressure regulator is one or more of sodium chloride, dextrose, and glycerol.

17. The OAB-14 suspension of claim 1, wherein, The particle size d(0.5) of the OAB-14 is 1-500 nm, and the particle size d(0.9) is 2-2000 nm.

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