A midazolam nanocrystal suspension for needle-free injection, a preparation method thereof and application thereof

Abstract

This invention relates to a midazolam nanocrystal suspension, which, by weight-volume percentage, comprises 5%–45% midazolam, 1%–10% Tween, and 0.1%–10% sodium deoxycholate (SDC). The midazolam nanocrystal suspension of this invention exhibits superior blood-brain barrier permeability and bioavailability, making it suitable for needle-free injection administration.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a midazolam nanocrystal suspension, its preparation method, and its application. Background Technology

[0002] Epilepsy is a chronic neurological disorder that seriously threatens global public health. Data from the "China Antiepileptic Drug Market Assessment and Investment Strategy Report (2019 Edition)" shows that there are over 50 million people with active epilepsy worldwide, with nearly 10 million in my country, and over 400,000 new cases annually. Epilepsy has become the second most common neurological disease in my country, second only to headaches. Epilepsy treatment is a long process, and some patients require lifelong medication; therefore, effectively controlling epileptic seizures has significant clinical value.

[0003] Epilepsy and acute seizures are particularly prominent issues in children aged 0-18. The incidence of acute seizures in children is as high as 3%-4%, 10-15 times higher than in adults, and it easily progresses to frequent seizures or status epilepticus. If an acute seizure lasts longer than 5 minutes and is not effectively controlled, it can cause cerebral ischemia and hypoxia, cerebral edema, and even irreversible brain damage, endangering life or leaving intellectual disabilities. Emergency medications for childhood seizures need to be characterized by rapid onset of action and high absolute bioavailability to seize the "golden 5-minute" treatment window.

[0004] The development of drugs for the nervous system faces a core constraint: the blood-brain barrier. The blood-brain barrier, composed of structures such as the tight junctions of brain capillary endothelial cells, blocks over 99% of small molecule drugs and almost all large molecule drugs from entering the brain, significantly limiting the intrabrain delivery efficiency and bioavailability of neurotherapeutic drugs.

[0005] Midazolam is a benzodiazepine that exerts a potent anticonvulsant effect by enhancing the inhibitory effect of GABAergic nerves, making it a first-line drug for the clinical control of epilepsy and acute seizures. However, its existing formulations have significant limitations: firstly, poor water solubility makes it difficult to prepare high-concentration formulations; secondly, intravenous injections have low accessibility for outpatient emergency use and pose risks of injection pain and tissue irritation; and thirdly, oral formulations exhibit a significant first-pass effect, resulting in a slow onset of action and failing to meet the time-sensitive requirements for emergency treatment of acute seizures.

[0006] Drug nanocrystal technology can effectively improve the solubility and dissolution rate of poorly soluble drugs. Midazolam nanocrystals have advantages such as uniform particle size, resistance to aggregation and stratification, high bioavailability, and low irritation. However, due to their high surface energy, these nanoparticles are prone to spontaneous aggregation, sedimentation, or crystal growth, leading to changes in particle size distribution, which in turn affects dissolution behavior and bioavailability. Furthermore, they are sensitive to environmental factors such as temperature and light, requiring stringent storage conditions. Therefore, it is urgent to screen suitable surfactants and stabilizers to improve the stability of nanocrystal systems.

[0007] Needle-free injection technology provides a feasible solution for rapid outpatient drug administration. It relies on instantaneous high pressure to create an ultrafine fluid stream for subcutaneous or intramuscular drug delivery, offering advantages such as non-invasiveness, convenience, rapid onset of action, and high compliance, making it suitable for emergency situations. However, this technology typically requires a drug delivery volume not exceeding 1 mL, with strict requirements on drug solubility and loading capacity. Therefore, it is necessary to overcome the water solubility limitations of midazolam, achieving high drug loading in a small-volume delivery system to ensure rapid and safe brain entry, meeting the immediate emergency needs for epilepsy and acute seizures. This necessitates the development of midazolam formulations with characteristics such as outpatient emergency care, rapid onset of action, high bioavailability, and ease of use. Summary of the Invention

[0008] The purpose of this invention is to provide a midazolam nanocrystal suspension, which contains 5-45% midazolam, 1-10% Tween and 0.1-10% sodium deoxycholate (SDC) by weight-volume percentage.

[0009] In a preferred embodiment of the present invention, the suspension contains 5-30% midazolam, 1-10% Tween, and 0.1-5% sodium deoxycholate.

[0010] In a preferred embodiment of the present invention, the suspension contains 5-20% midazolam, 1-8% Tween, and 0.1-1% sodium deoxycholate.

[0011] In a preferred embodiment of the present invention, the suspension contains 5-10% midazolam, 1-5% Tween, and 0.1-1% sodium deoxycholate.

[0012] In a preferred embodiment of the present invention, the Tween is selected from any one of Tween-20 and Tween-80 or a combination thereof.

[0013] In a preferred embodiment of the present invention, the mass ratio of midazolam: Tween-20: sodium deoxycholate in the suspension is 10-40:1-30:1, preferably 15-35:5-25:1, and more preferably 20-30:10-20:1.

[0014] In a preferred embodiment of the present invention, the suspension contains 5.0% midazolam, 2.5% Tween-20, and 0.2% SDC by weight-volume percentage.

[0015] In a preferred embodiment of the present invention, the suspension contains 10.0% midazolam, 2.5% Tween-20, and 0.2% SDC by weight-volume percentage.

[0016] In a preferred embodiment of the present invention, the average particle size of the midazolam nanocrystals in the suspension is ≤200nm, preferably ≤180nm.

[0017] In a preferred embodiment of the present invention, the PDI of midazolam nanocrystals in the suspension is 0.1-0.3, preferably 0.1-0.25.

[0018] In a preferred embodiment of the present invention, the zeta potential of the midazolam nanocrystals in the suspension is absolutely ≥25mV, preferably ≥30mV.

[0019] In a preferred embodiment of the present invention, the suspension optionally contains any one or a combination of an acid-base regulator, a preservative, and a lyophilization protectant.

[0020] In a preferred embodiment of the present invention, the acid-base regulator is selected from any one or a combination of hydrochloric acid, sulfuric acid, sorbic acid, citric acid, carboxylic acid, hydroxy acid, keto acid, acetic acid, oxalic acid, succinic acid, formic acid, acetic acid, butyric acid, malonic acid, succinic acid, adipic acid, pyruvic acid, glutamic acid, itaconic acid, ascorbic acid, fumaric acid, α-ketoglutarate, fruit acid, sodium hydroxide, potassium hydroxide, ammonium hydroxide, citric acid, potassium citrate, sodium citrate, malic acid, sodium malate, potassium malate, sodium carbonate, sodium bicarbonate, potassium carbonate, phosphoric acid, sodium phosphate, disodium hydrogen phosphate, sodium dihydrogen phosphate, dipotassium hydrogen phosphate, potassium dihydrogen phosphate, diamine hydrogen phosphate, diamine dihydrogen phosphate, monoethanolamine, diethanolamine, triethanolamine, lactic acid, sodium lactate, potassium lactate, propionic acid, sodium propionate, potassium propionate, tartaric acid, sodium tartrate, potassium tartrate, fumaric acid, sodium fumarate, and potassium fumarate.

[0021] In a preferred embodiment of the present invention, the midazolam nanocrystal suspension has a pH of 6.0-8.0, preferably 6.5-7.5.

[0022] In a preferred embodiment of the present invention, the preservative is selected from any one or a combination of phenol, cresol, benzoic acid, sodium benzoate, potassium benzoate, sodium dehydroacetate, chlorobutanol, benzyl alcohol, sorbic acid, potassium sorbate, sodium sorbate, methylparaben, ethylparaben, propylparaben, calcium propionate, sodium diacetate, phenoxyethanol methylparaben, phenoxyethanol methylisothiazolinone, methylparaben, methylparaben, ethylparaben, propylparaben, sodium metabisulfite, chlorhexidine, sodium citrate, butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), ethylenediaminetetraacetic acid, propyl gallate, and quaternary ammonium compounds.

[0023] In a preferred embodiment of the present invention, the preservative contained in the suspension is 0.1-5%, preferably 0.2-4%, and more preferably 0.5-2%.

[0024] In a preferred embodiment of the present invention, the freeze-drying protectant is selected from any one or a combination of sucrose, maltose, lactose, fructose, dextran, trehalose, sorbitol, xylitol, maltitol, oligosaccharide alcohols, glycerol, mannitol, sorbitol, dextran, glucose, glyceryl triethyl acetate (HES), polyethylene glycol, ethylene glycol, phosphate, acetate, citrate, and starch.

[0025] In a preferred embodiment of the present invention, the preparation of the midazolam nanocrystal suspension includes the following steps: dissolving the required amount of Tween and sodium deoxycholate in water, then adding the required amount of midazolam, and wet grinding to obtain the solution.

[0026] In the preferred embodiment of the present invention, the grinding conditions are (1500-4000rpm*20min-3h), preferably (1800-3500rpm*25min-120min), more preferably (2000-3000rpm*30-60min), and even more preferably (2500-3000rpm*30-40min).

[0027] In a preferred embodiment of the present invention, the diameter of the zirconia grinding beads used in wet grinding is 0.1 mm-0.4 mm, preferably 0.2 mm-0.3 mm.

[0028] In a preferred embodiment of the present invention, the filling rate of the zirconia grinding beads used in wet grinding is 50-80%, preferably 60-70%.

[0029] Another objective of this invention is to provide a method for preparing a midazolam nanocrystal suspension, wherein the suspension contains 5-45% midazolam, 1-10% Tween, and 0.1-10% sodium deoxycholate (SDC) by weight-volume percentage. The preparation method includes the following steps: dissolving the required amounts of Tween and sodium deoxycholate in water, then adding the required amount of midazolam, and wet grinding to obtain the solution.

[0030] In the preferred embodiment of the present invention, the grinding conditions are (1500-4000rpm*20min-3h), preferably (1800-3500rpm*25min-120min), more preferably (2500-3000rpm*30-40min), and even more preferably (2500-3000rpm*30-40min).

[0031] In a preferred embodiment of the present invention, the diameter of the zirconia grinding beads used in wet grinding is 0.1 mm-0.4 mm, preferably 0.2 mm-0.3 mm.

[0032] In a preferred embodiment of the present invention, the filling rate of the zirconia grinding beads used in wet grinding is 50-80%, preferably 60-70%.

[0033] Another object of the present invention is to provide the application of the midazolam nanocrystal suspension of the present invention in the preparation of anticonvulsant drugs.

[0034] In a preferred embodiment of the present invention, the convulsion is selected from any one of the following: febrile convulsion, extracranial febrile convulsion, toxic substance convulsion, neurological damage convulsion, immature brain development convulsion, intracranial infection convulsion, electrolyte imbalance convulsion, metabolic disorder convulsion, neurochemical weapon convulsion, neonatal convulsion, childhood convulsion, and acute convulsive attack.

[0035] In a preferred embodiment of the present invention, the extracranial fever is selected from fever caused by any one or a combination of causes such as chickenpox, influenza, gastroenteritis, otitis media, respiratory tract infection, tonsillitis, or vaccination.

[0036] Another object of the present invention is to provide the use of the midazolam nanocrystal suspension of the present invention in the preparation of antiepileptic drugs.

[0037] In a preferred embodiment of the present invention, the epilepsy is selected from any one of generalized epilepsy, partial / focal epilepsy, epileptic spasms, reflex epilepsy, epilepsy caused by neurochemical weapons, or a complication thereof.

[0038] In a preferred embodiment of the present invention, the generalized epileptic seizure is selected from any one of generalized tonic-clonic seizures, absence seizures, tonic seizures, clonic seizures, myoclonic seizures, atonic seizures, or their complications.

[0039] In a preferred embodiment of the present invention, the partial epileptic seizure is selected from any one of simple partial seizures, complex partial seizures, secondary generalized seizures, or their complications.

[0040] Another object of the present invention is to provide an antiepileptic pharmaceutical composition comprising the midazolam nanocrystal suspension of the present invention and other antiepileptic drugs.

[0041] In a preferred embodiment of the present invention, the other antiepileptic drugs are selected from any one or a combination of phenytoin sodium, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, ethosuximide, sodium valproate, lamotrigine, topiramate, levetiracetam, gabapentin, pregabalin, phenobarbital, phenytoin, fosphenytoin, benzodiazepines, nonaminobutyrates, sodium valproate, topiramate, tiagabine, aminovinyl acetate, perampanel, and zonisamide.

[0042] Another object of the present invention is to provide the use of the midazolam nanocrystal suspension of the present invention in the preparation of any of the anesthetic drugs and sedative drugs.

[0043] In a preferred embodiment of the present invention, the sedation is selected from any one of preoperative sedation, conscious sedation before diagnosis or endoscopic procedures, anesthesia induction, intraoperative sedation during general anesthesia or epidural anesthesia.

[0044] Another object of the present invention is to provide the application of the midazolam nanocrystal suspension of the present invention in the preparation of antidotes for neurochemical weapons.

[0045] In a preferred embodiment of the present invention, the antidote is used to relieve any one of the following or its complications: convulsions, epilepsy, and secondary neurological damage caused by neurochemical weapons.

[0046] Another object of the present invention is to provide the use of the midazolam nanocrystal suspension of the present invention in the preparation of drugs for improving blood-brain barrier permeability.

[0047] Unless otherwise stated, the present invention uses the following method for detection:

[0048] 1. This invention uses a nanoparticle size analyzer (Malvern ZS90, Malvern Panalytical, England) to detect the average particle size, zeta potential, and polydispersity index (PDI).

[0049] 2. This invention employs high-performance liquid chromatography (HPLC) to detect the content of midazolam: an Alphasil ES-C18 (5μm, 2.1100mm, Acchrom, Beijing, China) column was used, with phosphate buffer (0.1mol / L phosphoric acid: 0.3mol / L triethylamine in a 1:1 ratio)-methanol (35:65) as the mobile phase, a flow rate of 0.2mL / min, an injection volume of 10μl, a column temperature of 40℃, and a detection wavelength of 220nm. 500 ng / mL diazepam was used as the internal standard, and a linear relationship (10-400 ng / mL) was established using blank serum and a series of midazolam standard solutions. The limit of detection was 5 ng / mL, and the limit of quantitation was 10 ng / mL.

[0050] Unless otherwise stated, when this invention relates to percentages between liquids, the percentage is volume / volume percentage; when this invention relates to percentages between liquids and solids, the percentage is volume / weight percentage; when this invention relates to percentages between solids and liquids, the percentage is weight / volume percentage; the remainder is weight / weight percentage.

[0051] Compared with existing technologies, this invention achieves the following unexpected technical effects by scientifically screening the components and proportions of midazolam nanocrystal suspension:

[0052] 1. The midazolam nanocrystal suspension of the present invention improves the lipid solubility of vascular endothelial cell membranes, and significantly enhances the blood-brain barrier permeability, intracerebral delivery efficiency and bioavailability of midazolam nanocrystals.

[0053] 2. The midazolam nanocrystal suspension of this invention can be used for needle-free injection, which is convenient to administer and has a rapid onset of action. It significantly reduces skin damage and local irritation, greatly improves the safety, compliance and emergency accessibility of patients (especially children), and can significantly shorten the treatment time for anticonvulsants and antiacute seizures. It can quickly control epilepsy and acute seizures, effectively grasp the "golden 5 minutes" treatment window, significantly reduce the incidence of status epilepticus in children, and reduce the resulting cerebral ischemia-hypoxia, cerebral edema and seizure-related brain injury, thereby reducing disability and mortality rates. It is more suitable for out-of-hospital and home emergency scenarios, reducing seizure-related disability and mortality rates.

[0054] 3. The midazolam nanocrystal suspension of the present invention has uniform particle size distribution, excellent stability, rapid release, rapid onset of action and moderate sustained release, and prolonged residence time in the body. It significantly reduces blood drug concentration fluctuations and improves the delivery efficiency and safety of poorly soluble drugs.

[0055] 4. The preparation method of the midazolam nanocrystal suspension of the present invention has the characteristics of simple operation, short cycle, low cost, no need for complex equipment and harsh conditions, and is suitable for large-scale industrial production. Attached Figure Description

[0056] Figure 1 The stability of the midazolam nanocrystal suspension of the present invention was investigated. Detailed Implementation

[0057] The following description of specific embodiments of the present invention does not limit the present invention. Those skilled in the art can make various changes or modifications based on the present invention, and as long as they do not depart from the spirit of the present invention, they should all fall within the scope of protection of the claims of the present invention.

[0058] Nanowet mill (DYNO®-MILL RESEARCH LAB, Willy A. Bachofen AG, German).

[0059] Example 1 Composition and preparation of midazolam nanocrystal suspension

[0060] Dissolve 2.5g Tween-20 and 0.2g SDC in 100mL of water, then add 5g midazolam. Place the mixture in a wet mill (zirconia grinding beads with a diameter of 0.2mm and a filling rate of 60%) and grind (3000rpm*30min) to obtain the final product.

[0061] The prepared nanocrystalline suspension was diluted 200 times with a 0.2% SDC solution. According to the method of the present invention, the average particle size of the midazolam nanocrystals was 146.81 nm, the polydispersity index (PDI) was 0.173, and the zeta potential was -40.54 mV.

[0062] Example 2 Composition and preparation of midazolam nanocrystal suspension

[0063] Dissolve 2.5g Tween-20 and 0.2g SDC in 100mL of water, then add 5g midazolam. Place the mixture in a wet mill (zirconia grinding beads with a diameter of 0.2mm and a filling rate of 70%) and grind (3000rpm*40min) to obtain the final product.

[0064] The prepared nanocrystalline suspension was diluted 200 times with a 0.2% SDC solution. According to the method of the present invention, the average particle size of the midazolam nanocrystals was 150.5 nm, the polydispersity index (PDI) was 0.112, and the zeta potential was -34.57 mV.

[0065] Example 3 Preparation of midazolam nanocrystal suspension

[0066] Dissolve 2.5g Tween-20 and 0.2g SDC in 100mL of water, then add 5g midazolam. Place the mixture in a wet mill (zirconia grinding beads with a diameter of 0.2mm and a filling rate of 60%) and grind (2500rpm*30min) to obtain the final product.

[0067] The prepared nanocrystalline suspension was diluted 200 times with a 0.2% SDC solution. According to the method of the present invention, the average particle size of the midazolam nanocrystals was 163.73 nm, the polydispersity index (PDI) was 0.155, and the zeta potential was -31.34 mV.

[0068] Example 4 Composition and preparation of midazolam nanocrystal suspension

[0069] Dissolve 2.5g Tween-20 and 0.2g SDC in 100mL of water, then add 10g midazolam. Place the mixture in a wet mill (zirconia grinding beads with a diameter of 0.2mm and a filling rate of 60%) and grind (3000rpm*30min) to obtain the final product.

[0070] Comparative Example 1 Preparation of midazolam nanocrystal suspension

[0071] Dissolve 2.5g PVPk25 and 0.2g SDC in 100mL of water, then add 5g midazolam. Place the mixture in a wet mill (zirconia grinding beads with a diameter of 0.2mm and a filling rate of 60%) and grind (3000rpm*30min) to obtain the final product.

[0072] The prepared nanocrystalline suspension was diluted 200 times with 0.2% SDC solution. According to the method of the present invention, the average particle size of midazolam nanocrystals was 186.27 nm, the polydispersity index (PDI) was 0.204, and the zeta potential was -40.9 mV.

[0073] Experimental Example 1 Stability Study of the Midazolam Nanocrystal Suspension of the Present Invention

[0074] The nanocrystalline suspensions prepared in Examples 1-3 and the nanocrystalline suspension prepared in Comparative Example 1 were placed at 25°C (room temperature) for 30 days. Samples were taken on days 0, 10, 20, 30, 40, 50, and 60, and diluted 200 times with 0.2% SDC solution. The particle size was then determined according to the method of this invention. The results are shown in […]. Figure 1 .

[0075] Experimental Example 2 Blood-brain barrier permeability study of the midazolam nanocrystal suspension of this invention

[0076] Ten two-week-old male SD rats (purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.) were randomly divided into an experimental group and a control group, with five rats in each group.

[0077] 1. Isolation and primary culture of brain microvascular endothelial cells

[0078] (1) After the experimental animals were acclimatized for 1 day, they were sacrificed, their heads were clamped with hemostatic forceps, and their heads were immersed in 75% ethanol solution for 2 min. After being immersed in D-Hank's solution, the brain tissue was immediately removed and the cerebral cortex was prepared at 0℃. The cerebral cortex was washed 3 times with D-Hank's solution, transferred to serum-free DMEM and cut into pieces. 4 mL of 0.1% type II collagenase (containing 120 μD Nasel) was added and digested in a 37℃ water bath for 1.5 h. The centrifuge tube was shaken every 20 min and centrifuged (1000 rpm * 8 min). The cells were collected, resuspended in 20% BSA, and centrifuged (14000 rpm * 20 min, 4℃) to remove the middle layer tissue and large blood vessels. The bottom precipitate was collected.

[0079] (2) Add 2 mL of a mixed solution of 0.1% collagenase / dispersase and 40 μDNase I to the collected bottom precipitate. After digestion for 1 h, centrifuge (1000 rpm * 8 min). Add 3 mL of rat endothelial cell separation solution 1, then slowly add 1 mL of separation solution 2 to the upper layer. Then add 2 mL of FBS-free DMEM to the top layer to resuspend the cell solution, ensuring that the liquid surfaces of each layer are clearly separated. Centrifuge (1000 g * 20 min, 4 °C).

[0080] (3) Take out the middle endothelial cells (BMECs), place them in DMEM medium without FBS, centrifuge (1000 rpm*5 min) and wash twice;

[0081] (4) After resuspending the collected cells in ECM culture medium, seed them in a culture flask for 30 min, then transfer them to a culture flask coated with FN, add puromycin (4 μg / mL), and incubate them at 37℃ and 5% CO2 until the culture flask is fully colonized.

[0082] (5) Aspirate the culture medium, wash 3 times with PBS, add 0.25% trypsin to digest for 1.5 min, and then add culture medium to stop digestion;

[0083] (6) Centrifuge the digested cells (1000 rpm for 5 min), wash, collect the cells, resuspend them in ECM culture medium, and adjust the cell density to 1 x 10⁻⁶ cells / min. 5 The cells were cultured at 37°C and 5% CO2, with the medium changed daily. Once the cells had reached a full confluence, the cells were passaged again.

[0084] (7) Remove the culture medium, wash three times with PBS, then add 0.25% trypsin for 1-2 min to digest, add culture medium to stop digestion, centrifuge (1000 rpm * 5 min), collect the cells and resuspend them in ECM culture medium at 1 * 10 6 Cells were seeded at a density of [number] cells / well in Transwell 24-well plates (FN incubation) and cultured at 37°C and 5% CO2 for 2-3 days. After cell confluence was observed, TEER assays were performed. Electrophysiological resistance was measured from day 1 after seeding and continuously cultured and monitored for 7 days. A TEER value exceeding 200 Ω·cm was achieved. 2 The in vitro blood-brain barrier (BBB) ​​model was successfully constructed.

[0085] (8) Take a culture flask filled with brain microvascular epithelial cells, wash it 3 times with PBS, digest it with 0.25% trypsin for 2 min, observe the cells in suspension under a microscope, and immediately add ECM medium containing 10% FBS to stop the digestion.

[0086] (9) Take a 24-well plate, add 1 mL of culture medium to the lower layer, place a Transwell chamber in the upper layer, and add 200 μL of cells at a density of 1*10⁻⁶. 6 Add 1 / mL of culture medium, then add 200 μL of ECM culture medium to the lower chamber, keep the liquid level constant, and measure the resistance value after 2-4 hours, and calculate the resistance value (Ω·cm). 2 = (R cells - R blank) × S membrane (cm) 2 );

[0087] (10) Aspirate the ECM culture medium from the upper and lower chambers. Add 1200 μL of D-Hanks to the lower chamber and 200 μL of drug-containing culture medium to the upper chamber. The experimental group is the midazolam nanocrystal suspension (10 μg / mL) of Example 1, and the control group is the midazolam nanocrystal suspension (10 μg / mL) of Comparative Example 1. At 5 min, 10 min, 20 min, 30 min, 45 min, and 60 min after administration, take 200 μL of the drug from the lower chamber and add it to the injection vial. After taking it out, replenish the fluid.

[0088] (11) The concentration of midazolam that crosses the blood-brain barrier (BBB) ​​was determined by HPLC. The mobile phase consisted of 0.1 mol / L phosphate buffer (9.81 g phosphate dissolved in 1000 mL water, 4 mL triethylamine added, and pH adjusted to 3.5 with 2 mol / L NaOH): methanol in a volume ratio of 35:65. The column temperature was 30 °C. max 220 nm; flow rate 1.0 mL / min, injection volume 10 μL. Results are shown in Table 1.

[0089] Table 1. Cumulative blood-brain barrier permeability of midazolam nanocrystal suspension

[0090]

[0091] Experimental Example 3 Pharmacokinetic Study of Midazolam Nanocrystal Suspension of the Present Invention

[0092] Twenty-four male rats weighing 250-270g (purchased from Spiford (Beijing) Biotechnology Co., Ltd.) were randomly divided into four groups of six each.

[0093] NF-1 group: needle-free injection of midazolam nanocrystal suspension of Comparative Example 1 (dose of 1 mg / kg).

[0094] Group N-1: Midazolam nanocrystal suspension of Comparative Example 1 was injected via needle (dose was 1 mg / kg).

[0095] NF-2 group: needle-free injection of midazolam nanocrystal suspension from Example 1 (dose 1 mg / kg).

[0096] Group N-2: Midazolam nanocrystal suspension of Example 1 was injected via needle (dose of 1 mg / kg).

[0097] The experimental animals were housed at 25 ± 2℃ and 60% RH, with a 12-hour light / 12-hour dark cycle. During the experiment, the animals had free access to food and water, and were allowed to acclimatize to the environment for one week. Food and water were restricted for 12 hours prior to the experiment. The animals were anesthetized with isoflurane before drug administration and blood collection.

[0098] Blood samples of 0.3 mL were collected from the orbital venous plexus at 2 min, 5 min, 15 min, 30 min, 1 h, 1.5 h, 3 h, 6 h, 9 h, and 12 h after administration to the experimental animals and placed in heparinized test tubes. The samples were centrifuged at 4°C (6000 rpm for 10 min), and the collected serum was used for protein precipitation with 0.1% formate acetonitrile solution. After vortexing for 30 s, the samples were centrifuged again (14000 rpm for 10 min), and the supernatant was collected for HPLC analysis. The results are shown in Table 2.

[0099] Table 2

[0100]

[0101] The above description of specific embodiments of the present invention does not limit the present invention. Those skilled in the art can make various changes or modifications based on the present invention, and as long as they do not depart from the spirit of the present invention, they should all fall within the scope of protection of the claims of the present invention.

Claims

1. A midazolam nanocrystal suspension, comprising, by weight-volume percentage, 5-45% midazolam, 1-10% Tween and 0.1-10% sodium deoxycholate (SDC).

2. The nanocrystalline suspension as described in claim 1, wherein the Tween is selected from any one of Tween-20 and Tween-80 or a combination thereof.

3. The nanocrystalline suspension according to any one of claims 1-2, wherein the mass ratio of midazolam: Tween-20: sodium deoxycholate in the suspension is 10-40:1-30:1, preferably 15-35:5-25:1, and more preferably 20-30:10-20:

1.

4. The nanocrystalline suspension according to any one of claims 1-3, wherein the average particle size of the midazolam nanocrystals in the suspension is ≤200nm, preferably ≤180nm.

5. The nanocrystalline suspension according to any one of claims 1-4, wherein the PDI of the midazolam nanocrystals in the suspension is 0.1-0.3, preferably 0.1-0.

25.

6. The nanocrystalline suspension according to any one of claims 1-5, wherein the pH of the suspension is 6.0-8.0, preferably 6.5-7.

5.

7. The nanocrystalline suspension according to any one of claims 1-6, wherein the suspension optionally contains any one or a combination of an acid-base regulator, a preservative, and a lyophilization protectant.

8. The method for preparing the midazolam nanocrystal suspension according to any one of claims 1-7, comprising the following steps: dissolving the required amount of Tween and sodium deoxycholate in water, then adding the required amount of midazolam, and wet grinding to obtain the solution.

9. The use of the midazolam nanocrystal suspension as described in any one of claims 1-7 in the preparation of any one of anticonvulsant drugs, antiepileptic drugs, anesthetic drugs, sedative drugs, antidotes for neurochemical weapons, and drugs that improve blood-brain barrier permeability.

10. An antiepileptic pharmaceutical composition comprising midazolam nanocrystal suspension as described in any one of claims 1-7 and other antiepileptic drugs.

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