WO2025098365A1 - Levodopa nasal spray, preparation method therefor, and use thereof - Google Patents

Abstract

Disclosed are a levodopa nasal spray, a preparation method therefor, and use thereof. The levodopa nasal spray comprises levodopa or a pharmaceutically acceptable salt thereof, an absorption enhancer, a suspending agent, an antioxidant, a wetting agent, and an antimicrobial agent. The levodopa nasal spray enhances the stability of levodopa, increases olfactory region deposition of the drug, improves nasal-to-brain delivery efficiency, can effectively treat Parkinson's disease while reducing the occurrence of symptom fluctuations, and has suitable viscosity.

Description

A levodopa nasal spray and its preparation method and application Technical Field

The invention belongs to the technical field of biomedicine, and specifically relates to a levodopa nasal spray and a preparation method and application thereof.

Background Art

Parkinson's disease (PD) is a common neurodegenerative disease with a large number of patients. The disease has many causes and mechanisms, and is a comprehensive pathogenic result. The main pathological changes are the loss of dopamine (DA) neurons in the substantia nigra and the formation of Lewy bodies. The main biochemical changes are the lack of dopamine in the striatum, the imbalance of dopamine and acetylcholine transmitters, and the relative hyperfunction of acetylcholine. The main symptoms of clinical patients include motor symptoms such as bradykinesia, resting tremor, muscle rigidity and postural balance disorders, accompanied by non-motor symptoms such as olfactory disorders, constipation, and sleep disorders. Currently, commonly used treatments include drug therapy, surgical treatment, rehabilitation treatment, etc., but it cannot be cured, and can only relieve symptoms and improve the quality of life of patients. Its treatment needs urgent attention.

The drugs used to treat PD mainly include dopamine (levodopa, dopamine receptor agonists, etc.) and choline (benhexyphenidyl hydrochloride, benztropine mesylate, procyclidine hydrochloride, etc.). Among them, levodopa (LDA) is the metabolic precursor of dopamine. Peripherally supplemented levodopa can pass through the blood-brain barrier and be decarboxylated into dopamine in the brain by dopa decarboxylase, thereby playing a role in replacement therapy. It is the standard treatment for Parkinson's disease and the most effective symptomatic drug in Parkinson's disease drug treatment. In addition, levodopa is unstable in a humid state and is prone to oxidation. At present, worldwide, most preparations related to levodopa are solid preparations such as levodopa tablets, capsules, granules, and inhalers that are administered orally. However, after oral administration, only 1% can be transported to the striatum tissue in the brain through neutral amino acid carriers to be converted into dopamine to exert its effect. It has a slow onset of action, low bioavailability, and a large dosage. More than 95% of levodopa is converted to dopamine by decarboxylases widely distributed in peripheral tissues, leading to adverse reactions such as cardiovascular and gastrointestinal diseases. After long-term high-dose use, patients will also experience "On/Off" phenomena. In addition, as motor function deteriorates with the progression of the disease, PD patients will experience dysphagia, which subsequently makes oral intake of levodopa much more difficult in these patients. Therefore, an alternative route for levodopa administration is needed.

In comparison, nasal administration preparations can deliver drugs directly from the olfactory area to the brain, bypassing the blood-brain barrier, and have significant advantages such as rapid onset of action, increased efficiency of levodopa entering the brain, reduced dosage, and avoidance of gastrointestinal reactions. There are a number of nasal formulations on the market for central nervous system diseases, such as zolmitriptan nasal spray for the treatment of migraine. and sumatriptan nasal spray Butorphanol tartrate nasal spray, an opioid used to relieve pain ) and fentanyl citrate nasal spray Diazepam nasal spray for epilepsy treatment Midazolam nasal spray and esketamine nasal spray for depression All of them have been approved by the FDA for marketing, which fully demonstrates the feasibility and clinical prospects of nasal administration of levodopa in the treatment of Parkinson's disease.

Oral levodopa and pramipexole are commonly used drugs in the clinical treatment of Parkinson's disease. Because dopamine has difficulty penetrating the blood-brain barrier, patients need to be supplemented with levodopa, a precursor of dopamine. When levodopa enters the brain and is taken up by dopaminergic neurons, it can be converted into dopamine through a decarboxylation reaction, thereby exerting its pharmacological effects. After levodopa is absorbed by the body, 95% of the drug undergoes a decarboxylation reaction in the periphery to form dopamine, and only 1% of the drug penetrates the blood-brain barrier and enters the brain.

There is a direct anatomical connection between the nasal cavity and the brain tissue: the olfactory mucosal epithelium of the nasal cavity contains bipolar olfactory cells, and the olfactory nerves formed by them pass through the cribriform plate and enter the olfactory bulb of the central nervous system. Therefore, after nasal administration of levodopa, part of the drug can be absorbed through the olfactory mucosa and enter the olfactory bulb or cerebrospinal fluid (CSF), thereby bypassing the blood-brain barrier (BBB) and being directly transported into the brain to exert a central therapeutic effect. Therefore, nasal administration has a fast onset of action, high bioavailability in the brain, and can reduce the dosage of levodopa and reduce adverse reactions.

For nose-to-brain delivery preparations, the amount of drug absorbed in the olfactory area is crucial to its efficacy. The area of the olfactory mucosa is very small (1-5 ^cm2 ), accounting for only 3%-5% of the total area of the nasal cavity. How to deposit the drug in the olfactory area, improve the drug delivery efficiency, and promote the absorption of the drug in the nasal mucosa is the research and development focus and difficulty of levodopa nasal spray. In addition, because levodopa is very easy to oxidize in the air when it is wet, the only levodopa liquid preparation currently on the market is the carbidopa intestinal gel that needs to be frozen for storage. (Continuous Intrajejunal Infusion) One, its stability is another difficulty in the development of L-DOPA nasal spray.

Summary of the invention

The purpose of the first aspect of the present invention is to provide a pharmaceutical composition.

The second aspect of the present invention aims to provide a method for preparing the above composition.

The third aspect of the present invention is to provide the use of the above-mentioned pharmaceutical composition.

The fourth aspect of the present invention aims to provide the use of dodecyl-β-D-maltoside as an absorption enhancer in the preparation of levodopa pharmaceutical preparations.

The technical solution adopted by the present invention is:

The first aspect of the present invention provides a pharmaceutical composition, comprising a drug and an absorption enhancer; the drug is Rotadopa and/or dopamine and/or carbidopa; the absorption enhancer is at least one of dodecyl-β-D-maltoside, tetradecyl-β-D-maltoside, chitosan, sucrose dodecanoate, carboxymethyl chitosan and sodium taurocholate.

Preferably, the absorption enhancer is dodecyl-β-D-maltoside.

Preferably, the pharmaceutical composition further comprises at least one of a suspending agent, an antioxidant, a wetting agent, and an antibacterial agent.

Preferably, the suspending agent is at least one of microcrystalline cellulose-sodium carboxymethyl cellulose RC591 or CL611, carbomer 934, 940.

Preferably, the microcrystalline cellulose-sodium carboxymethyl cellulose includes microcrystalline cellulose-sodium carboxymethyl cellulose RC591, CL611 and the like.

Preferably, the carbomer includes carbomer 934, 940 and the like.

Preferably, the suspending agent is microcrystalline cellulose-sodium carboxymethyl cellulose RC591.

Preferably, the antioxidant is at least one of ascorbic acid, ascorbyl palmitate and sodium metabisulfite.

Preferably, the antioxidant is sodium metabisulfite.

Preferably, the wetting agent may be at least one of polysorbate and poloxamer.

Preferably, the wetting agent may be polysorbate.

Preferably, the polysorbate is polysorbate 80.

Preferably, the antibacterial agent is at least one of benzalkonium chloride, benzalkonium bromide, quaternary ammonium salts, cetrimide, phenoxyethanol, sodium benzoate, and phenylethanol.

Preferably, the antibacterial agent is benzalkonium chloride.

Preferably, the pharmaceutical composition comprises: 0.5% to 20% w/w of the drug, 1% to 3% w/w of the suspending agent; 0.1% to 3% w/w of the antioxidant; 0.1% to 0.5% w/w of the absorption enhancer; 0.01% to 0.03% w/w of the wetting agent; and 0.01% to 0.024% w/w of the antibacterial agent.

Preferably, the pharmaceutical composition (dopasil) comprises: 0.5% to 15% w/w of levodopa or a pharmaceutically acceptable salt thereof, 0.125% to 3% w/w of benserazide or a pharmaceutically acceptable salt thereof; 1.0% to 2% w/w of a suspending agent; 0.2% to 3% w/w of an antioxidant; 0.05% to 0.5% w/w of an absorption enhancer; 0.005% to 0.03% w/w of a wetting agent; and 0.005% to 0.03% w/w of an antibacterial agent.

Preferably, the levodopa composition comprises: 1% to 20% w/w of levodopa or a pharmaceutically acceptable salt thereof, 1% to 3% w/w of a suspending agent; 0.1% to 3% w/w of an antioxidant; 0.1% to 0.5% w/w of an absorption enhancer; 0.01% to 0.03% w/w of a wetting agent; and 0.01% to 0.024% w/w of an antibacterial agent.

Preferably, the pharmaceutical composition comprises: 1% to 15% w/w of or a pharmaceutically acceptable salt thereof, 1.2% to 2% w/w suspending agent; 0.2% to 1% w/w antioxidant; 0.2% to 0.3% w/w absorption promoter; 0.01% to 0.03% w/w wetting agent; 0.015% to 0.02% w/w antibacterial agent.

Preferably, the pharmaceutical composition further comprises a pH adjuster for adjusting the pH of the pharmaceutical composition to 5.0-6.5.

Preferably, the added amount of the pH adjuster is 0.03% to 0.464%.

Preferably, the pH adjuster is at least one of citric acid/sodium citrate and sodium dihydrogen phosphate/disodium hydrogen phosphate.

Preferably, the pH adjuster is citric acid/sodium citrate.

Preferably, the pharmaceutical composition further comprises an osmotic pressure regulator for adjusting the osmotic pressure of the pharmaceutical composition to 270-350 mOsm/kg.

Preferably, the content of the osmotic pressure regulator is 0% to 1.5%.

Preferably, the osmotic pressure regulator is glycerol.

Preferably, the pharmaceutical composition further comprises water.

Preferably, the levodopa is levodopa micropowder.

Preferably, the particle size d ₉₀ of the levodopa micropowder is 2 to 20 μm.

Preferably, the pharmaceutical composition is in the form of a spray.

Preferably, the pharmaceutical composition is administered nasally.

Preferably, the acceptable delivery volume of the spray is 100-200 μL, the effective delivery dose of levodopa is 10-40 mg/spray, and the clinical therapeutic dose is 10-1000 mg/day, wherein the delivery volume is determined by the spray device, and the delivery dose is the product of the drug solution concentration and the delivery volume.

In a second aspect, the present invention provides a method for preparing the above-mentioned pharmaceutical composition, comprising the following steps:

S1: Add the wetting agent into water, add the drug and mix until evenly dispersed to obtain liquid 1;

S2: Add the suspending agent to an appropriate amount of water, mix until uniformly dispersed, and then add a pH adjuster to obtain liquid 2;

S3: adding liquid 2, antioxidant, absorption enhancer, antibacterial agent, and osmotic pressure regulator into liquid 1, and stirring until the mixture is uniform, thereby obtaining a pharmaceutical composition;

Preferably, the method further comprises filling the pharmaceutical composition into a nasal spray device to obtain a nasal spray.

The third aspect of the present invention provides use of the pharmaceutical composition of the first aspect of the present invention in the preparation of Parkinson's disease drugs.

The fourth aspect of the present invention provides the use of dodecyl-β-D-maltoside in the preparation of levodopa and/or benserazide and/or carbidopa nasal spray.

The beneficial effects of the present invention are:

The present invention provides a levodopa nasal spray, wherein dodecyl-β-D-maltoside is used as the nasal spray of levodopa. The drug absorption promoter is compounded with a suspending agent, an antioxidant, a wetting agent, an antibacterial agent, etc. to improve the stability of levodopa in the liquid preparation. After administration, the preparation can achieve drug absorption through two pathways: 1) part of the drug is directly delivered to the olfactory area, bypassing the blood-brain barrier, and entering the brain to take effect; 2) part of the drug is quickly absorbed into the blood through the nasal mucosa in the respiratory area, enters the systemic circulation, and enters the brain after passing through the blood-brain barrier to take effect. Among them, it is directly delivered to the olfactory area through the nasal cavity, bypassing the blood-brain barrier, improving the brain drug absorption efficiency, taking effect quickly, reducing the dosage, and is used to treat Parkinson's disease while improving the occurrence of symptom fluctuations. The levodopa nasal spray has a suitable viscosity, can effectively increase the deposition of drugs in the olfactory area, improve the nose-brain delivery efficiency, and effectively treat Parkinson's disease.

BRIEF DESCRIPTION OF THE DRAWINGS

Fig. 1 Release curves of formulations F7-F10 and F11-F14.

Figure 2 shows the appearance of preparations with different sodium metabisulfite concentrations after being placed at 60°C for different periods of time (from left to right: solutions with sodium metabisulfite contents of 0.2%, 0.5%, 1%, 2%, 3%, 0% and no levodopa).

Figure 3 shows the appearance of preparations with different sodium metabisulfite concentrations after exposure to light for different periods of time (from left to right: solutions with sodium metabisulfite contents of 0.2%, 0.5%, 1%, 2%, 3%, 0% and no levodopa).

Figure 4 Droplet size distribution during the spraying process of 30 mm levodopa nasal spray.

Figure 5 Droplet size distribution during the spraying process of 60 mm levodopa nasal spray.

Fig. 6 The intracerebral drug-time curve of dopamine nasal spray.

DETAILED DESCRIPTION

The following will be combined with the embodiments to clearly and completely describe the concept of the present invention and the technical effects produced, so as to fully understand the purpose, characteristics and effects of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, other embodiments obtained by those skilled in the art without creative work are all within the scope of protection of the present invention.

The levodopa used in the following examples is levodopa micropowder, and the particle size D ₉₀ may be 2-20 μm. The pulverization method is as follows:

A JET MILL Lab air flow mill was used with the air inlet pressure and crushing pressure set to 6.5 bar and 7.0 bar respectively. The automatic feeding mode was adopted with a feeding speed of 10 rpm, and the materials were fed for air flow crushing.

Determination of particle size of levodopa micropowder: The particle size of levodopa after micropowder was determined by the Malvern wet method, and the test parameters were as follows: background/sample measurement time 10s, shading 8%-20%, stirring speed 2000rpm, stirring time 5min.

The particle size distribution of levodopa and its micropowder is shown in Table 1:

Table 1 Particle size distribution of levodopa and its micropowder

The present invention provides a nasal spray for treating Parkinson's disease, comprising the following ingredients:

Active pharmaceutical ingredient: 0.5% to 20% w/w;

Suspending agent: 1% to 3% w/w;

Antioxidant: 0.1% to 3% w/w;

Absorption enhancer: 0.1% to 0.5% w/w;

Wetting agent: 0.01% to 0.03% w/w;

pH adjuster: appropriate amount, used to adjust the pH of the spray to 5.0-6.5;

Osmotic pressure regulator: appropriate amount, used to adjust the osmotic pressure of the spray to 270-350mOsm/kg;

Antibacterial agent: 0.01% to 0.024% w/w;

Purified water: add to 100% w/w.

The present invention also provides a method for preparing the above-mentioned nasal spray, comprising the following steps:

S1: The API is crushed by a jet mill, the crushing pressure is 5-10 bar, the feed pressure is 7-10 bar, and the feed speed is 50-65 g/min to obtain drug micropowder;

S2: Add appropriate amount of purified water and wetting agent to the liquid preparation tank, set the stirring speed to 10-20 Hz, stir for 3-8 min, increase the stirring speed to 40-50 Hz, add the prescribed amount of levodopa micropowder under stirring, and continue stirring for 20-40 min after the addition is completed to obtain liquid 1;

S3: Add an appropriate amount of purified water to another liquid preparation tank, set the stirring speed to 40-50 Hz, add the suspending material under stirring, continue stirring for 20-40 minutes, set the emulsification speed to 40-60 Hz, homogenize and emulsify for 80-120 minutes, add the pH adjuster under stirring, and continue emulsification for 20-40 minutes to obtain liquid 2;

S4: Set the stirring speed of the liquid preparation tank to 40-60 Hz, add liquid 2, antioxidant solution, absorption promoter solution, antibacterial agent solution and osmotic pressure regulator solution to liquid 1 in sequence under stirring, set the pressure of the liquid preparation tank to about -0.08 MPa, turn on the vacuum pump, and turn off the vacuum pump after reaching the set pressure. Stir for 20-40 minutes until the mixture is evenly mixed to obtain a levodopa suspension;

S5: Fill the levodopa suspension into a high-density polyethylene bottle and cap the nasal spray pump under a nitrogen environment to obtain the levodopa nasal spray.

Example 1 Levodopa destruction experiment

According to the standards of levodopa in the United States Pharmacopoeia, the USP content determination method is used to destroy the raw material sample The content of levodopa was tested to investigate the stability of the levodopa raw material under the destruction conditions (3.0% hydrogen peroxide oxidation for 0.5, 1 and 2 h, 3.0% hydrogen peroxide oxidation at 60°C for 0.5, 1 and 2 h, and high temperature of 130°C for 1, 2 and 4 h).

Table 2 Levodopa content after destruction under different conditions

The results showed that L-dopa was unstable under high temperature and oxidation. With the extension of oxidation time and high temperature storage time, the L-dopa content gradually decreased. When oxidized at 60℃ for 0.5h, the L-dopa content decreased to 92.66%, and after oxidizing for 2h, the L-dopa content decreased significantly to 74.35%.

Example 2 Antioxidant Screening

The formula (Table 3 and Table 4) was designed with 1% microcrystalline cellulose-sodium carboxymethyl cellulose as suspending agent, ascorbic acid, ascorbyl palmitate, and sodium metabisulfite as candidate antioxidants, and the type and dosage as prescription variables. The formula was placed under high temperature 60°C and high humidity 75% to observe the changes of related substances.

Table 3 Specific composition of antioxidant screening prescription

Table 4 Prescription design of different antioxidants

Table 5 Related substances of each prescription after being placed at high temperature and high humidity for 0 days and 5 days

The results are shown in Table 5. It can be seen that at 60°C, after each prescription was placed for 5 days, the impurity content increased significantly. In addition, the color of the ascorbic acid group, ascorbyl palmitate group and the prescription without antioxidant changed significantly to brown-yellow, and the antioxidant performance was poor. In addition, since ascorbic acid itself is unstable and the color changes greatly after oxidation, sodium pyrosulfite is preferably used as an antioxidant.

Example 3 Screening of suspending agents

Microcrystalline cellulose-sodium carboxymethyl cellulose RC591 and CL611 approved by FDA for nasal administration were used as suspension aids. The formulation designs of the suspending agent and the suspending agent are shown in Tables 6 and 7. The content uniformity and release behavior of the preparations at different suspending agent concentrations were investigated.

Table 6 Specific composition of the suspending agent screening prescription

Table 7 Prescription design of different suspending agents

The content uniformity results are shown in Table 8;

Table 8 Content and content uniformity of different prescriptions

The results show that the RSD of the content of each prescription after preparation is less than 2%, which meets the requirements. After being placed at room temperature for 5 days, the content of each prescription has not changed, and the content uniformity meets the requirements.

The results of the relevant substance contents are shown in Table 9;

Table 9 Contents of related substances in different prescriptions

The results show that the total impurity content of each prescription is far less than 1%, and the content of each single impurity is far less than 0.1%, which meets the requirements.

The release behavior of each prescription was investigated using a transdermal diffusion instrument. The release medium was 9 mL of PBS solution with a pH of 5.8, the medium temperature was 37 ± 0.5 °C, the sampling volume was 9 mL, and the rehydration volume was 9 mL; the release behavior curve is shown in Figure 1.

Figure 1 shows that prescriptions F7 to F14 exhibit rapid release, with almost complete release at 24 hours. In addition, by comparing the prescriptions, it can be seen that as the viscosity of the prescription increases, the release rate of each prescription increases, and the cumulative release increases. Among them, prescription F7 has the highest release rate, with more than 60% released in 1 hour and a cumulative release of more than 90% in 24 hours. It is preferred as the best suspending agent Proportion.

Example 4 Sodium pyrosulfite concentration screening

Prepare the pharmaceutical solutions according to Table 10, with sodium metabisulfite concentrations of 0.2%, 0.5%, 1%, 2% and 3%, respectively. Fill the above pharmaceutical solutions into vials, place them at a high temperature of 60°C and light (4500Lx±500Lx), observe their appearance, and detect the levodopa content and related substances.

Table 10

The results are shown in Figures 2-3 and Tables 11-14.

Table 11 Levodopa content of different sodium metabisulfite concentrations in prescriptions placed at 60°C for different times

Table 12 Related substances in different sodium metabisulfite concentrations at 60°C for different periods of time

Table 13 Levodopa content in different sodium metabisulfite concentration prescriptions placed under different lighting conditions for different periods of time

Table 14 Related substances in different sodium metabisulfite concentration prescriptions placed under light conditions for different times

The results show:

1) When placed at 60°C, the solution without sodium metabisulfite changed color. When the sodium metabisulfite content was 0.2% to 3%, there was no color change after 30 days of placement, and there was no significant change in the levodopa content and no significant increase in related substances (Figure 2 and Tables 11-12).

2) When placed under light conditions, the solution without sodium metabisulfite changed color. When the sodium metabisulfite content was 0.2% to 3%, there was no color change after 13 days, no significant change in levodopa content, and no significant increase in related substances (Figure 3 and Tables 13-14).

Example 5 Quality Evaluation of Preferred Prescriptions

1. Prepare the liquid medicine according to the prescription in Table 15 and fill it. The preparation method is as follows:

S1: The L-dopa API is crushed by a jet mill at a crushing pressure of 7 bar, a feed pressure of 8 bar, and a feed rate of 56 g/min to obtain L-dopa micropowder;

S2: Add appropriate amount of purified water and wetting agent to the liquid preparation tank, set the stirring speed to 15 Hz, stir for 5 min, increase the stirring speed to 45 Hz, add the prescribed amount of levodopa micropowder under stirring, and continue stirring for 30 min after the addition is completed to obtain liquid 1;

S3: Add an appropriate amount of purified water to another liquid preparation tank, set the stirring speed to 45 Hz, add the suspending material under stirring, continue stirring for 30 minutes, set the emulsification speed to 50 Hz, homogenize and emulsify for 100 minutes, add the pH adjuster under stirring, and continue emulsification for 30 minutes to obtain liquid 2;

S4: Set the stirring speed of the liquid preparation tank to 50 Hz, add liquid 2, antioxidant solution, absorption promoter solution, antibacterial agent solution and osmotic pressure regulator solution to liquid 1 in sequence under stirring, set the pressure of the liquid preparation tank to -0.08 MPa, turn on the vacuum pump, and turn off the vacuum pump after reaching the set pressure. Stir for 30 minutes until the mixture is evenly mixed to obtain a levodopa suspension;

S5: Fill the levodopa suspension into a high-density polyethylene bottle and cap the nasal spray pump under a nitrogen environment to obtain the levodopa nasal spray.

The API content, viscosity, osmotic pressure, pH, sedimentation stability, spray performance, etc. of the levodopa nasal spray were tested. The results are shown in Table 16.

Table 15

Table 16

The results showed that the preparation met the requirements for nasal preparations in the 2020 edition of the Chinese Pharmacopoeia.

2. Investigation of delivery dose uniformity

The above-mentioned drug solution was filled into a high-density polyethylene bottle, and the nasal spray pump was capped under a nitrogen environment. According to the FDA guidelines and the requirements of the Chinese Pharmacopoeia 2020 edition, the delivery dose of 10 bottles of samples from the same batch was tested. The results are shown in Table 17.

Table 17

The results showed that the delivery doses of the first 10 sprays and the last spray (60th spray) were 98.44% and 98.25% of the labeled amount, respectively, which can stably deliver drugs and meet the requirements of the Chinese Pharmacopoeia 2020 edition and FDA guidelines.

3. Droplet size distribution

The spray droplet size of the levodopa nasal spray at 30 mm and 60 mm was measured by a Sympatec laser particle size analyzer (HELOS & SPRAYER ^™ ), with an R5 lens, a trigger pressure of 60 N, and a measuring time of 150 ms.

The spray process of nasal spray can be divided into three stages: formation period, stabilization period and dissipation period: formation period is the spray formation stage, the droplet concentration increases rapidly and the particle size increases rapidly; during the stabilization period, the droplet size reaches a peak and remains stable; during the dissipation period, the droplet concentration decreases rapidly and the particle size fluctuates greatly. As shown in Figures 4 and 5, the stabilization period of levodopa nasal spray is 20% to 50%, so this is the stabilization period of levodopa nasal spray, and the three-stage method is used to detect the droplet size of the nasal spray during the stabilization period.

4. Droplet size during stable period

Sympatec laser particle size analyzer (HELOS & SPRAYER ^™ ) was used to measure the droplet diameter of levodopa nasal spray at 30 mm and 60 mm during the stable period. The lens was R5, the trigger pressure was 60 N, and the stable period was 20% to 50%. The results are shown in Table 18.

Table 18

The results showed that at distances of 30 mm and 60 mm, the spray particle sizes within and between bottles of this preparation were relatively stable.

5. Fine droplet volume

The drug content of fine droplets of nasal spray was investigated using a new generation pharmaceutical disk impactor (NGI). The air flow rate was 15 L/min, the volume of the glass expansion chamber was 5 L, the injection volume was 4 presses, the interval between each press was 5 s, and the preheating temperature was set at 3.5 °C. Cool for 90 minutes; the results are shown in Table 19.

Table 19

The results showed that the drug content of levodopa nasal spray in droplets smaller than 10 μm per press was extremely low, less than 0.1% of the labeled amount, which is in line with FDA guidelines.

Example 6 Investigation of in vivo pharmacokinetic behavior

1. The pharmacokinetic behavior of three administration routes, namely, nasal administration of levodopa nasal spray, oral administration of commercial preparations, and oral inhalation administration of levodopa inhalation powder, was studied to investigate the absorption and metabolic behavior of levodopa in plasma after administration. The preparation method of homemade levodopa inhalation powder is as follows:

a) Prescription design is shown in Table 20:

Table 20 Self-made levodopa inhalation powder

b) Preparation

The levodopa powder was added three times by layer-by-layer method. 4.4 g lactose ML001 was added to the centrifuge tube, and 0.6 g micronized API was added and mixed evenly. Then 1.2 g lactose ML001 and 0.7 g micronized API were added in sequence. The mixture was mixed with 1.2 g of lactose MLO1 and 0.7 g of micronized API, and then 1.2 g of lactose ML001 was manually mixed for 20 min to obtain levodopa inhalation powder.

Male SD rats (180-220g) were raised in an SPF environment and were free to eat and drink. They were fasted for 12 hours before the experiment. Fifteen rats were randomly divided into five groups, with three rats in each group, and given levodopa tablets (25 mg/kg), medopa tablets (20 mg/kg), homemade levodopa inhalation powder (10 mg/kg), homemade levodopa nasal spray (5 mg/kg) and homemade levodopa nasal spray (10 mg/kg), respectively. Continuous orbital blood sampling was performed at 2min, 5min, 10min, 15min, 30min, 1h, 2h, 4h, and 6h after administration to detect the levels of levodopa and dopamine in plasma. The pharmacokinetic parameter calculation results are shown in Tables 21 and 22;

Table 21 Rat plasma levodopa content

Table 22 Rat plasma dopamine content

The results showed that the time-dependent curves of plasma levodopa and dopamine concentrations and pharmacokinetic parameters of each group of rats after administration were as follows: at 1/5 of the administration dose, the AUC _0-6h of levodopa in plasma after nasal administration of the nasal spray (10 mg/kg) was much higher than that of the levodopa tablet group. The plasma levodopa concentration of the levodopa nasal spray was linearly related to the administration dose, and the Cmax and AUC of levodopa in plasma increased with the increase of the administration dose.

In addition, after nasal administration of L-DOPA nasal spray, the dopamine content in plasma was similar to that of Medopar tablets, but much lower than that of Medopar tablets. Compared with the levodopa tablet group, it is safer and is expected to reduce the side effects after levodopa administration.

2. To study the tissue distribution of L-dopa nasal spray after nasal administration and oral administration of commercially available preparations, and to investigate the absorption and metabolic behavior of L-dopa in brain tissue after administration.

Male SD rats (180-220g) were raised in an SPF environment and were free to eat and drink. They were fasted for 12 hours before the experiment. 15 rats were randomly divided into 5 groups, 3 rats in each group, and given levodopa tablets (25mg/kg), medopar tablets (20mg/kg), homemade levodopa nasal spray (5mg/kg) and homemade levodopa nasal spray (10mg/kg) respectively. The rats were killed at 2min, 5min, 10min, 15min, 30min, 1h, 2h, 4h, and 6h after administration, and the brain tissue was completely removed. The homogenate was added at a ratio of brain tissue: homogenate = 1:4 and homogenized under ice bath. The levodopa and dopamine content in the brain tissue homogenate was detected. The pharmacokinetic parameter calculation results are shown in Tables 23-24;

Table 23 Dopamine content in rat brain tissue (baseline subtracted) (n=3)

Table 24 L-DOPA content in rat brain tissue (n=3)

The results showed that after nasal administration, the L-dopa nasal spray was rapidly absorbed and entered the brain, with a peak time of 10-15 minutes, which was faster than that of the oral administration group of Madopar (T _max = 60 minutes). At 1/2 the dosage, the AUC _0-6h of the effective drug (dopamine) was similar to that of the Madopar tablet group, and it is expected to achieve the same therapeutic effect at 1/2 the dosage, which can reduce the L-dopa The dosage of the drug is increased to improve safety.

In addition, after administration of levodopa nasal spray, at 1/5 of the administered dose, the AUC _0-6h of dopamine in the brain was much higher than that of the levodopa tablet group, and its effectiveness was significantly improved.

Example 7 Study on the in vivo absorption promoting effect of DDM and investigation of its in vivo pharmacokinetic behavior

The pharmacokinetic study of L-DOPA nasal spray with and without dodecyl-β-D-maltoside (DDM) was conducted to evaluate the absorption-enhancing effect of DDM on the drug.

The prescription design of nasal spray (without DDM) is shown in Table 25.

Table 25 DDM-free levodopa nasal spray prescription design

The DDM-free levodopa nasal spray was prepared according to the prescription in the above table, and the preparation method was the same as the preparation method of the levodopa nasal spray in the specific embodiment.

The experimental animals and administration methods were the same as above, and the results are shown in Tables 26 to 29;

Table 26 Dopamine content in rat brain tissue (subtracting base) (n=3)

Table 27 L-DOPA content in rat brain tissue (n=3)

Table 28 Rat plasma levodopa content

Table 29 Rat plasma dopamine content

The results showed that the addition of DDM (dodecyl-β-D-maltoside) significantly promoted the absorption of levodopa. The plasma peak time of levodopa was increased from 120min to 15min, and the AUC _0-6h of dopamine in the brain was increased from 3875ng/mL·min to 14189ng/mL·min.

The above specific implementations have been described in detail for the present invention, but the present invention is not limited to the above embodiments. Various changes can be made within the knowledge of ordinary technicians in the relevant technical field without departing from the purpose of the present invention. In addition, the embodiments of the present invention and the features in the embodiments can be combined with each other without conflict.

Example 8 Dopamine hydrazine nasal spray

1. Preparation of dopamine nasal spray

The proportion range of each component in dopamine nasal spray is shown in Table 30:

Table 30 Dobase hydrazine nasal spray prescription composition

The formulation shown in Table 31 was used to prepare dopamine hydrazine nasal spray for subsequent experiments:

Table 31 Preferred dopamine nasal spray prescription composition

The specific preparation steps are as follows:

S1: crushing the levodopa and benserazide APIs by a jet mill, with a crushing pressure of 7 bar, a feed pressure of 8 bar, and a feed rate of 56 g/min to obtain levodopa and benserazide micropowders with a d90 of 2-20 μm;

S2: Add appropriate amount of purified water and wetting agent to the liquid preparation tank, set the stirring speed to 15 Hz, stir for 5 min, increase the stirring speed to 45 Hz, add the prescribed amount of levodopa micropowder and benserazide micropowder under stirring, and continue stirring for 30 min after the addition is completed to obtain liquid 1;

S3: Add an appropriate amount of purified water to another liquid preparation tank, set the stirring speed to 45 Hz, add the suspending material under stirring, continue stirring for 30 minutes, set the emulsification speed to 50 Hz, homogenize and emulsify for 100 minutes, add the pH adjuster under stirring, and continue emulsification for 30 minutes to obtain liquid 2;

S4: Set the stirring speed of the preparation tank to 50 Hz, add liquid 2, antioxidant solution, absorption promoter solution and antibacterial agent solution to liquid 1 in sequence under stirring, set the pressure of the preparation tank to -0.08 MPa, turn on the vacuum pump, and turn off the vacuum pump after reaching the set pressure. Stir for 30 minutes until the mixture is uniform, and then obtain the dobase hydrazine suspension;

S5: Fill the dobase hydrazine suspension into a high-density polyethylene bottle and cap the nasal spray pump under a nitrogen environment to obtain the dobase hydrazine nasal spray.

2. In vitro evaluation of dopamine nasal spray

1) The content of the prepared dobase hydrazine nasal spray was detected, and the viscosity, osmotic pressure, pH, sedimentation stability, spray performance, etc. were examined. The results are shown in Table 32.

Table 32 Quality evaluation of dopamine nasal spray

The results showed that the preparation met the requirements for nasal preparations in the Chinese Pharmacopoeia 2020 edition.

2) Investigation of delivery dose uniformity

According to the FDA guidelines and the requirements of the Chinese Pharmacopoeia 2020, the delivery dose uniformity of the levodopa + benserazide compound nasal spray was investigated, and the delivery doses of 10 bottles of samples from the same batch were tested. The results are shown in Tables 33-34;

Table 33 L-DOPA detection amount in dopamine hydrazine nasal spray

Table 34 Benserazide content per spray of dobase hydrazide nasal spray

The results showed that the delivery doses of the first 10 sprays and the last spray (60th spray) were 98.23% and 98.48% of the labeled amount, respectively, which can stably deliver drugs and meet the requirements of the Chinese Pharmacopoeia 2020 edition and FDA guidelines.

3. Pharmacokinetic study of dopamine nasal spray

The pharmacokinetic study of dopamine nasal spray without dodecyl-β-D-maltoside (DDM) was carried out to evaluate the brain delivery performance of dopamine nasal spray and the absorption enhancement effect of DDM on the drug.

The prescription design of nasal spray (without DDM) is shown in Table 35.

Table 35 DDM-free dopamine hydrazine nasal spray prescription design

The DDM-free dopamine hydrazine nasal spray was prepared according to the above prescription, and the preparation method was the same as the preparation method of the dopamine hydrazine suspension nasal spray in the specific embodiment.

2) The tissue distribution of dopamine nasal spray and DDM-free dopamine nasal spray after nasal administration was studied to investigate the absorption and metabolic behavior of dopamine in brain tissue after administration.

Male SD rats (180-220g) were raised in an SPF environment and were free to eat and drink. They were fasted for 12 hours before the experiment. Fifteen rats were randomly divided into two groups, 3 in each group, and given homemade dopamine hydrazide nasal spray (20 mg/kg) and homemade DDM-free dopamine hydrazide nasal spray (20 mg/kg) respectively. The rats were killed at 2min, 5min, 10min, 15min, 30min, 1h, 2h, 4h, and 6h after administration, and the brain tissue was completely removed. The homogenate was added at a ratio of brain tissue: homogenate = 1:4 and homogenized under ice bath. The amine content in the brain tissue homogenate was detected. The pharmacokinetic parameter calculation results are shown in Figure 6 and Table 36;

Table 36 L-DOPA content in rat brain tissue (n=3)

The results showed that the addition of DDM (dodecyl-β-D-maltoside) significantly promoted the absorption of levodopa, increasing the peak concentration of levodopa from 209 ng/mL to 227 ng/mL, and increasing the AUC0-6h of levodopa in the brain from 45462 ng/mL·min to 50464 ng/mL·min.

Claims (10)

A pharmaceutical composition, comprising a drug and an absorption enhancer; the drug is levodopa and/or benserazide and/or carbidopa or a pharmaceutically acceptable salt thereof; The absorption enhancer is at least one of dodecyl-β-D-maltoside, tetradecyl-β-D-maltoside, chitosan, sucrose dodecanoate, carboxymethyl chitosan and sodium taurocholate. The pharmaceutical composition according to claim 1, characterized in that the pharmaceutical composition further comprises at least one of a suspending agent, an antioxidant, a wetting agent, and an antibacterial agent. The pharmaceutical composition according to claim 2, characterized in that the suspending agent is at least one of microcrystalline cellulose-sodium carboxymethyl cellulose and carbomer; Preferably, the antioxidant is at least one of ascorbic acid, ascorbyl palmitate, and sodium metabisulfite; Preferably, the wetting agent may be at least one of polysorbate and poloxamer; Preferably, the antibacterial agent is at least one of benzalkonium chloride, benzalkonium bromide, quaternary ammonium salts, cetrimide, phenoxyethanol, sodium benzoate, and phenylethanol. The pharmaceutical composition according to claim 2, characterized in that the pharmaceutical composition comprises: 0.5% to 20% w/w of the drug, 1% to 3% w/w of the suspending agent; 0.1% to 3% w/w of the antioxidant; 0.1% to 0.5% w/w of the absorption promoter; 0.01% to 0.03% w/w of the wetting agent; 0.01% to 0.024% w/w of the antibacterial agent; Preferably, the pharmaceutical composition comprises: 1% to 15% w/w of the drug, 1.2% to 2% w/w of the suspending agent; 0.2% to 1% w/w antioxidant: 0.1% to 0.3% w/w absorption promoter; 0.01% to 0.03% w/w wetting agent; 0.015% to 0.02% w/w antibacterial agent. The pharmaceutical composition according to claim 1, characterized in that the pharmaceutical composition further comprises a pH regulator for adjusting the pH of the pharmaceutical composition to 5.0-6.5; preferably, the pharmaceutical composition further comprises an osmotic pressure regulator for adjusting the osmotic pressure of the pharmaceutical composition to 270-350 mOsm/kg; preferably, the pharmaceutical composition further comprises water. The pharmaceutical composition according to claim 1, characterized in that the drug is a drug micropowder; preferably, the particle size D ₉₀ of the drug micropowder is 2 to 20 μm. The pharmaceutical composition according to any one of claims 1 to 6 is characterized in that the dosage form of the pharmaceutical composition is a spray; preferably, the pharmaceutical composition is administered via nasal administration. The method for preparing the pharmaceutical composition according to claim 7, comprising the following steps: S1: Add the wetting agent to water, add the drug, and mix until evenly dispersed to obtain liquid 1; S2: Add the suspending agent to an appropriate amount of water, mix until uniformly dispersed, and then homogenize and emulsify, and add a pH adjuster to obtain liquid 2; S3: adding liquid 2, antioxidant, absorption enhancer, antibacterial agent, and osmotic pressure regulator into liquid 1, and stirring until the mixture is uniform, thereby obtaining a pharmaceutical composition; Preferably, the method further comprises filling the pharmaceutical composition into a nasal spray bottle device to obtain a nasal spray. Use of the pharmaceutical composition according to any one of claims 1 to 7 in the preparation of Parkinson's disease drugs. Application of dodecyl-β-D-maltoside in the preparation of levodopa and/or benserazide and/or carbidopa nasal spray.