An amitriptyline spray, its preparation method and application

Abstract

The present invention relates to an amitriptyline spray, a preparation method thereof and an application thereof, belonging to the technical field of medicine. The amitriptyline spray of the present invention uses water as a solvent and comprises components with the following concentrations: 30-100 g / L of amitriptyline hydrochloride, 0.5-4 g / L of disodium ethylenediaminetetraacetate, 40-60 mL / L of dimethyl sulfoxide, and 5-15 mL / L of ethylparaben solution. The amitriptyline spray prepared by the present invention can relieve and treat neuropathic pain, and has the advantages of stable properties, good skin absorbability, fast analgesic speed, high analgesic strength, long-lasting analgesic effect, no toxic and side effects, and convenient use.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, and particularly to an amitriptyline spray and its preparation method and application. Background Art

[0002] Neuropathic pain (NP) is a chronic pain caused by primary damage, dysfunction of the somatic nervous system or various diseases. The inducing causes mainly include diabetic neuropathy, drug-induced neuropathy, traumatic nerve injury, herpes zoster, adverse reactions of cancer chemotherapy drugs, etc. According to the initial onset site, it can be divided into central neuropathic pain and peripheral neuropathic pain. The clinical manifestations are spontaneous burning pain, stabbing pain, tingling, and accompanied by induced pain, usually showing persistent and recurrent attacks when gently touched or in the cold, and it is a common symptom of somatosensory nervous system diseases.

[0003] Currently, the clinical treatment of NP is still difficult. The existing treatment methods cannot control the pain symptoms in a short time and improve the recurrent situation of NP. The existing treatment drugs also have defects such as low analgesic strength, poor treatment effect, and more side effects. Therefore, it brings great challenges to the clinical prevention and treatment of NP. Amitriptyline in tricyclic antidepressants (TCA) is applicable to the treatment of diabetic peripheral neuropathy, postherpetic neuralgia and neuropathic pain caused by central stroke. Its mechanism of action is to block the reuptake between serotonin and norepinephrine neurotransmitters. However, studies have found that oral administration of amitriptyline will produce central effects, resulting in the occurrence of many adverse drug events (ADEs) such as severe chest pain, rapid heartbeat, fainting, abnormal bleeding, disturbance of consciousness, delirium, auditory hallucination, visual hallucination, etc., and even cause anticholinergic reactions, orthostatic hypotension and toxic liver damage.

[0004] There is no amitriptyline preparation in the prior art that can stably and effectively relieve and / or treat neuropathic pain and has low toxicity and side effects. Summary of the Invention

[0005] The purpose of the present invention is to provide an amitriptyline spray and its preparation method and application to solve the problems of low analgesic strength, poor treatment effect and more side effects in the prior art during the treatment of neuropathic pain.

[0006] In order to achieve the above-mentioned invention purpose, the present invention provides the following technical solutions:

[0007] The present invention provides an amitriptyline spray, which uses water as a solvent and comprises components with the following concentrations: amitriptyline hydrochloride 30 - 100 g / L, disodium ethylenediaminetetraacetate 0.5 - 4 g / L, dimethyl sulfoxide 40 - 60 mL / L, and ethylparaben solution 5 - 15 mL / L.

[0008] Preferably, the amitriptyline spray uses water as a solvent and comprises components with the following concentrations: amitriptyline hydrochloride 40 - 60 g / L, disodium ethylenediaminetetraacetate 1 - 3 g / L, dimethyl sulfoxide 45 - 55 mL / L, and ethylparaben solution 8 - 12 mL / L.

[0009] Preferably, the amitriptyline spray uses water as a solvent and comprises components with the following concentrations: amitriptyline hydrochloride 50 g / L, disodium ethylenediaminetetraacetate 2 g / L, dimethyl sulfoxide 50 mL / L, and ethylparaben solution 10 mL / L.

[0010] Preferably, the ethylparaben solution uses absolute ethanol as a solvent and comprises components with the following concentrations: ethylparaben 30 - 60 g / L, glycerol 300 - 600 mL / L.

[0011] Preferably, the dosage form of the amitriptyline spray includes one of amitriptyline aqueous spray, amitriptyline oil spray, and amitriptyline gel spray.

[0012] The present invention also provides a preparation method of the amitriptyline spray, which comprises the following steps: mixing and dissolving amitriptyline hydrochloride, disodium ethylenediaminetetraacetate, dimethyl sulfoxide, ethylparaben solution, and water to obtain the amitriptyline spray.

[0013] The present invention also provides the application of the amitriptyline spray in the preparation of a drug for relieving and / or treating neuropathic pain.

[0014] The present invention has the following technical effects and advantages:

[0015] The amitriptyline spray prepared by the present invention can relieve and treat neuropathic pain, and has the advantages of stable properties, good skin absorbability, fast analgesic speed, high analgesic strength, long-lasting analgesic effect, convenient use, etc., and there is no obvious stain left. Animal experiments found that the amitriptyline spray of the present invention takes effect within 10 minutes after spraying, the analgesic time can last for 3 - 4 hours, and long-term administration helps to promote the repair of damaged nerves. No common side effects were found in all model mice after administering amitriptyline. Description of the Drawings

[0016] Figure 1 It is the standard curve of amitriptyline hydrochloride;

[0017] Figure 2These are the HPLC chromatograms of the reference substance and the test substance, where A is the HPLC chromatogram of the reference substance and B is the HPLC chromatogram of the test substance;

[0018] Figure 3 These are the diagrams of the mouse model of neuropathic pain. The left diagram is a schematic diagram of the mouse model of neuropathic pain, and the right diagram is a real - shot diagram of the mouse model of neuropathic pain;

[0019] Figure 4 These are the results of the determination of the mechanical pain threshold in mice;

[0020] Figure 5 These are the results of the determination of the cold pain response in mice;

[0021] Figure 6 These are the results of the determination of the heat pain response in mice;

[0022] Figure 7 These are the schematic diagrams of the sites for measuring the motor nerve conduction velocity in mice. In the figure, S 1 is the stimulating electrode at the sciatic notch of the mouse, S 2 (R 1 ) is the stimulating electrode at the ankle joint of the mouse or the double - needle recording electrode at the ankle joint of the mouse, R 2 is the double - needle recording electrode at the first interosseous muscle of the mouse toe, and E is the reference electrode.

[0023] Figure 8 These are the results of the determination of the motor nerve conduction velocity in mice. Detailed implementation mode

[0024] The present invention provides an amitriptyline spray. The amitriptyline spray uses water as a solvent and includes components with the following concentrations: amitriptyline hydrochloride 30 - 100 g / L, preferably 40 - 60 g / L, and more preferably 50 g / L; disodium ethylenediaminetetraacetate 0.5 - 4 g / L, preferably 1 - 3 g / L, and more preferably 2 g / L; dimethyl sulfoxide 40 - 60 mL / L, preferably 45 - 55 mL / L, and more preferably 50 mL / L; ethylparaben solution 5 - 15 mL / L, preferably 8 - 12 mL / L, and more preferably 10 mL / L.

[0025] In the present invention, the ethylparaben solution uses absolute ethanol as a solvent and includes components with the following concentrations: ethylparaben 30 - 60 g / L, preferably 50 g / L; glycerol 300 - 600 mL / L, preferably 500 mL / L.

[0026] In the present invention, the dosage form of the amitriptyline spray includes one of amitriptyline water spray, amitriptyline oil spray, and amitriptyline gel spray.

[0027] The present invention also provides a method for preparing the amitriptyline spray, which comprises the following steps: dissolving amitriptyline hydrochloride, disodium edetate, dimethyl sulfoxide, ethylparaben solution and water to obtain the amitriptyline spray.

[0028] In the present invention, the preparation method comprises the following steps: adding amitriptyline hydrochloride, disodium edetate, dimethyl sulfoxide and ethylparaben solution to water in sequence, dissolving and mixing uniformly to obtain the amitriptyline spray.

[0029] The present invention also provides the application of the amitriptyline spray in the preparation of a drug for relieving and / or treating neuropathic pain.

[0030] The technical solutions provided by the present invention are described in detail below with reference to the examples, but they should not be construed as limiting the protection scope of the present invention.

[0031] The test mice of the present invention are C57BL / 6J mice, with an age of 6-8 weeks and a weight of 20-25 g, purchased from Liaoning Changsheng Biotechnology Co., Ltd.

[0032] Among the experimental equipment of the present invention, the Ugo Basile cold / hot combination plate is purchased from Shanghai Yuchun Biotechnology Co., Ltd.

[0033] Example 1: Preparation of amitriptyline spray

[0034] (1) Dissolve 50 g of ethylparaben in an appropriate amount of absolute ethanol, add 500 mL of glycerol and mix uniformly, then make up the volume to 1000 mL with absolute ethanol to obtain an ethylparaben solution with a mass concentration of 5%.

[0035] (2) Dissolve 50 g of amitriptyline hydrochloride in an appropriate amount of pure water to obtain an amitriptyline hydrochloride solution.

[0036] (3) Add 2 g of disodium edetate, 50 mL of dimethyl sulfoxide and 10 mL of the ethylparaben solution with a mass concentration of 5% to the amitriptyline hydrochloride solution in sequence. After complete dissolution and uniform mixing, make up the volume to 1000 mL with pure water, and bottle it to obtain the amitriptyline spray.

[0037] Example 2: Preparation of amitriptyline spray

[0038] (1) Dissolve 60 g of ethylparaben in an appropriate amount of absolute ethanol, add 600 mL of glycerol and mix uniformly, then make up the volume to 1000 mL with absolute ethanol to obtain an ethylparaben solution with a mass concentration of 6%.

[0039] (2) Dissolve 30 g of amitriptyline hydrochloride in an appropriate amount of pure water to obtain an amitriptyline hydrochloride solution.

[0040] (3) Add 0.5 g of disodium ethylenediaminetetraacetate, 60 mL of dimethyl sulfoxide, and 5 mL of a 6% ethylparaben solution to the amitriptyline hydrochloride solution in sequence. After complete dissolution and thorough mixing, make up the volume to 1000 mL with pure water, bottle it, and obtain the amitriptyline spray.

[0041] Example 3: Preparation of Amitriptyline Spray

[0042] (1) Dissolve 30 g of ethylparaben in an appropriate amount of absolute ethanol, add 300 mL of glycerol and mix well, then make up the volume to 1000 mL with absolute ethanol to obtain a 3% ethylparaben solution;

[0043] (2) Dissolve 70 g of amitriptyline hydrochloride in an appropriate amount of pure water to obtain an amitriptyline hydrochloride solution;

[0044] (3) Add 4 g of disodium ethylenediaminetetraacetate, 40 mL of dimethyl sulfoxide, and 15 mL of a 3% ethylparaben solution to the amitriptyline hydrochloride solution in sequence. After complete dissolution and thorough mixing, make up the volume to 1000 mL with pure water, bottle it, and obtain the amitriptyline spray.

[0045] Example 4: Preparation of Amitriptyline Spray

[0046] (1) Dissolve 50 g of ethylparaben in an appropriate amount of absolute ethanol, add 500 mL of glycerol and mix well, then make up the volume to 1000 mL with absolute ethanol to obtain a 5% ethylparaben solution;

[0047] (2) Dissolve 40 g of amitriptyline hydrochloride in an appropriate amount of pure water to obtain an amitriptyline hydrochloride solution;

[0048] (3) Add 1 g of disodium ethylenediaminetetraacetate, 55 mL of dimethyl sulfoxide, and 8 mL of a 5% ethylparaben solution to the amitriptyline hydrochloride solution in sequence. After complete dissolution and thorough mixing, make up the volume to 1000 mL with pure water, bottle it, and obtain the amitriptyline spray.

[0049] Example 5: Preparation of Amitriptyline Spray

[0050] (1) Dissolve 50 g of ethylparaben in an appropriate amount of absolute ethanol, add 500 mL of glycerol and mix well, then make up the volume to 1000 mL with absolute ethanol to obtain a 5% ethylparaben solution;

[0051] (2) Dissolve 60 g of amitriptyline hydrochloride in an appropriate amount of pure water to obtain an amitriptyline hydrochloride solution;

[0052] (3) Add 3 g of disodium ethylenediaminetetraacetate, 45 mL of dimethyl sulfoxide, and 12 mL of a 5% ethylparaben solution to the amitriptyline hydrochloride solution in sequence. After complete dissolution and thorough mixing, make up the volume to 1000 mL with pure water, bottle it, and obtain the amitriptyline spray.

[0053] Comparative Example 1: Preparation of the amitriptyline spray adjuvant

[0054] (1) Dissolve 50 g of ethylparaben in an appropriate amount of absolute ethanol, add 500 mL of glycerol and mix well, then make up the volume to 1000 mL with absolute ethanol to obtain a 5% ethylparaben solution;

[0055] (2) Dilute 2 g of disodium ethylenediaminetetraacetate, 50 mL of dimethyl sulfoxide, and 10 mL of the 5% ethylparaben solution to 1000 mL with pure water, mix well and bottle it to obtain the amitriptyline spray adjuvant.

[0056] Experimental Example 1: Determination of the active ingredient content of the amitriptyline spray

[0057] (1) Solution preparation:

[0058] Preparation of the methanol-water-triethylamine solution: Mix 60 mL of methanol, 0.3 mL of triethylamine, and 40 mL of water, and adjust the pH to 3.1 with phosphoric acid to obtain the methanol-water-triethylamine solution;

[0059] Preparation of the test sample: Take 1 mL of the amitriptyline spray prepared in Example 1, add the methanol-water-triethylamine solution to make up the volume to 50 mL to obtain the test sample stock solution; then take 1 mL of the test sample stock solution and add the methanol-water-triethylamine solution to make up the volume to 50 mL to obtain the test sample;

[0060] Preparation of the reference substance: Take 10 mg of the amitriptyline hydrochloride reference standard, mix it with methanol and dissolve it by ultrasonic treatment for 10 min, then add methanol to make up the volume to 25 mL to obtain the reference substance stock solution; then take 1 mL of the reference substance stock solution and add the methanol-water-triethylamine solution to make up the volume to 10 mL to obtain the reference substance;

[0061] (2) Preparation of the standard curve of amitriptyline hydrochloride: Dilute the reference stock solution prepared in step (1) with methanol-water-triethylamine solution to obtain reference gradient solutions with concentrations of 0.816 mg / mL, 1.631 mg / mL, 3.263 mg / mL, 6.525 mg / mL, 13.050 mg / mL, and 26.100 mg / mL respectively; perform liquid chromatography detection on the reference gradient solutions at each concentration. The liquid chromatography conditions are as follows: using octadecylsilane chemically bonded silica as the filler, using the methanol-water-triethylamine solution prepared in step (1) as the mobile phase for liquid chromatography, the liquid chromatography wavelength is 240 nm, and the injection volume is 2 μL; calculate the peak areas of the reference gradient solutions at each concentration according to the external standard method, and draw the standard curve of amitriptyline hydrochloride. The results are as Figure 1 shown;

[0062] (3) Determination of the content of amitriptyline hydrochloride in the amitriptyline spray: Perform liquid chromatography detection on the test sample and the reference prepared in step (1) under the liquid chromatography conditions described in step (2), and calculate the peak area and the content of amitriptyline hydrochloride according to the external standard method. The results are as Figure 2 shown.

[0063] The results show that the regression equation of the standard curve of amitriptyline hydrochloride is y = 3.0755x - 0.3957 (R 2 = 0.9999), showing a linear regression relationship, and the linear range is 0.816 - 26.100 mg / mL; according to the calculation results, it can be known that the content of amitriptyline hydrochloride in the amitriptyline spray of the present invention is 5.0157%.

[0064] Experimental Example 2: Mouse model establishment and drug administration

[0065] (1) Model establishment: Select 28 adult male C57BL / 6J mice and divide them into 4 groups, namely experimental group A, experimental group B, control group, and blank group, with 7 mice in each group; use the intraperitoneal injection method to anesthetize the mice in experimental group A, experimental group B, and the control group with pentobarbital sodium at 50 mg / kg of mouse body weight. Then place the mice in the prone position, clean the surface mouse hair near the right femur of the mice, disinfect with iodophor, make an 8-mm longitudinal incision in the middle of the posterior right thigh to expose the subcutaneous muscle after disinfection, use a hemostatic forceps to bluntly separate the muscle to expose the sciatic nerve, and use four chromic catgut to loosely ligate around the sciatic nerve, with a chromic catgut interval of 1 mm, to obtain a neuropathic pain model (CCI model), as Figure 3As shown; the sciatic nerve of the mice in the blank group was not ligated. The main characteristics of the CCI model are as follows: By moderately ligating the sciatic nerve, a certain degree of damage to the sciatic nerve is caused, which in turn affects the distal nerve fibers. The mechanical compression of the sciatic nerve by the chromic intestine forms a chronic inflammatory response, thereby simulating the characteristics of neuropathic pain and inflammatory pain, accompanied by endoneural edema, focal ischemia, and axonal degeneration. After the operation, the sciatic nerve was repositioned, the muscles were arranged, and the wound was sutured;

[0066] (2) Administration: Spray the amitriptyline spray of Example 1 near the right femur of the mice in Experimental Group A and Experimental Group B, spray 3 times a day, 4 sprays each time (1.5 mL), for a total of 2 weeks. Among them, the mice in Experimental Group A were sprayed with amitriptyline spray again 0.5 h before the determination of behavioral and nerve conduction indexes, and the mice in Experimental Group B were not administered 0.5 h before the determination of behavioral and nerve conduction indexes; Spray the amitriptyline spray adjuvant of Comparative Example 1 near the right femur of the control group mice, spray 3 times a day, 4 sprays each time (1.5 mL), for a total of 2 weeks; The mice in the blank group were not administered.

[0067] Experimental Example 3: Determination of mouse behavioral indexes

[0068] (1) Determination of mechanical pain threshold in mice: Before modeling, on the 7th day and the 14th day after modeling and administration in each group of mice in Experimental Example 2, place each group of mice in an acrylic cage with a metal mesh bottom (10×10×10 cm). When the mice adapt to the environment for 15 min and are in a quiet state, use a 37450 type Ugo basile paw tactile tester to measure the mechanical pain threshold of each group of mice. Specifically: Control the mechanical needle to vertically stimulate the skin in the middle of the plantar surface of each group of mice from bottom to top. Set the stimulation intensity to gradually increase from 0 g to 10 g within 20 s. When each group of mice shows a rapid paw withdrawal response, the stimulation stops and the stimulation pressure value is recorded. A total of 3 measurements are taken, with an interval of 5 min each time. The average value of the 3 stimulation pressure values is used as the mechanical pain threshold. The results are as Figure 4 shown;

[0069] (2) Determination of cold pain response in mice: After the administration of each group of mice in Experimental Example 2, use a Ugo Basile cold / hot combination plate to perform a cold pain response. Specifically: Lower the temperature of the cold plate to 5±0.5 °C, and isolate the cold plate from the external environment with a fiberglass plate to maintain a constant temperature; Place each group of mice on the cold plate. The mice will lick or lift their feet due to pain. Observe and record the time when the mice perform licking or paw withdrawal movements. The results are as Figure 5 shown;

[0070] (3) Mouse thermal pain response measurement: After the administration of drugs to each group of mice in Experimental Example 2, the Ugo Basile cold / hot combined plate was used to measure the thermal pain response. Specifically: The temperature of the hot plate was heated to 55 ± 0.5 °C, and the hot plate was isolated from the external environment with a fiberglass plate to maintain a constant temperature; each group of mice was placed on the hot plate, and the mice would lick their feet or lift their feet due to pain. Observe and record the time when the mice showed licking or paw-withdrawal movements. The results are as Figure 6 shown;

[0071] (4) Mouse motor nerve conduction velocity measurement (MNCV): After the administration of drugs to each group of mice in Experimental Example 2, each group of mice was subjected to pulse stimulation. Specifically: Under the conditions of a room temperature of 20 ± 0.5 °C and a mouse body temperature of 37 °C, at the sciatic notch and ankle joint of each group of mice, a stimulating electrode S 1 / S 2 was used for stimulation, and at the ankle joint and the first interosseous muscle of the toe, a double-needle recording electrode R 1 / R 2 was used for stimulation and recording. The reference electrode E was placed between electrode S 1 and electrode S 2 / R 1 as well as between electrode S 2 / R 1 and electrode R 2 , and at a distance of 1 cm from electrode S 2 / R 1 , as Figure 7 shown; A single-pulse square wave stimulation with a pulse width of 0.1 ms was given to each group of mice. First, gradually increase the stimulation intensity, and record the biphasic compound action potential from electrode R 1 / R 2 . Then, gradually decrease the stimulation intensity, and the recorded biphasic compound action potential gradually disappears. Repeatedly adjust the stimulation intensity, and use the stimulation intensity when the compound action potential appears or disappears as the stimulation threshold and record it; Use 1.5 times the stimulation threshold as the stimulation intensity, with a 5 s interval between every 2 stimulations. Take the potential value between the peak and valley of the biphasic compound action potential as the wave amplitude, and record the time from the start of stimulation to the appearance of the action potential as the conduction time of the excitation signal. Repeat the measurement 10 times and calculate the average value; Straighten the hind legs of each group of mice obliquely backward at an angle of 45° with the spine in a natural limb state, and measure the distance from the stimulating electrode to the recording electrode of each group of mice along the position and direction of the sciatic nerve, and calculate the MNCV. The results are as Figure 8 shown;

[0072] The MNCV calculation formula is: MNCV = distance from the stimulating electrode to the recording electrode / conduction time.

[0073] The results showed that spraying amitriptyline spray of the present invention on the mice with neuropathic pain model could improve the symptoms of neuropathic pain in mice. The analgesic time could last for 3 to 4 hours, and there were no any toxic and side effects. It indicated that administering amitriptyline spray of the present invention to the mice with neuropathic pain model could rapidly exert analgesic effect, and long-term administration was helpful for promoting the repair of damaged nerves. No possible skin damage caused by amitriptyline spray was found during the administration process, nor were the related side effects that might occur with oral amitriptyline mentioned in the product instruction manual of amitriptyline disclosed in the prior art.

[0074] As can be seen from the above embodiments, the present invention provides an amitriptyline spray and its preparation method and application. The amitriptyline spray prepared by the present invention can relieve and treat neuropathic pain, and has the advantages of stable property, good skin absorbability, fast analgesic speed, high analgesic intensity, long-lasting analgesic effect, no toxic and side effects, etc.

[0075] The above are only the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. An amitriptyline spray, characterized in that, The amitriptyline spray uses water as a solvent and includes the following components in concentrations: 30-100 g / L amitriptyline hydrochloride, 0.5-4 g / L disodium edetate, 40-60 mL / L dimethyl sulfoxide, and 5-15 mL / L ethylparaben solution; The ethyl hydroxybenzoate solution uses anhydrous ethanol as a solvent and comprises components with the following concentrations: 30-60 g / L of ethyl hydroxybenzoate and 300-600 mL / L of glycerol.

2. The amitriptyline spray according to claim 1, characterized in that The amitriptyline spray uses water as a solvent and comprises components in the following concentrations: 40-60 g / L amitriptyline hydrochloride, 1-3 g / L disodium edetate, 45-55 mL / L dimethyl sulfoxide, and 8-12 mL / L ethylparaben solution.

3. The amitriptyline spray according to claim 1, characterized in that The amitriptyline spray uses water as a solvent and includes components in the following concentrations: 50 g / L amitriptyline hydrochloride, 2 g / L disodium edetate, 50 mL / L dimethyl sulfoxide, and 10 mL / L ethylparaben solution.

4. The amitriptyline spray according to any one of claims 1 to 3, characterized in that The dosage form of the amitriptyline spray includes one of an amitriptyline water spray and an amitriptyline gel spray.

5. The method for preparing the amitriptyline spray according to any one of claims 1 to 4, characterized in that: The method comprises the following steps: mixing and dissolving amitriptyline hydrochloride, disodium ethylenediaminetetraacetate, dimethyl sulfoxide, ethyl hydroxybenzoate solution and water to obtain amitriptyline spray.

6. Use of the amitriptyline spray according to any one of claims 1 to 4 in the preparation of a medicament for relieving and / or treating neuropathic pain.

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