Compound amino acid injection as well as stabilizer and preparation method thereof

Abstract

The invention provides a stabilizer for a compound amino acid injection, the compound amino acid injection containing the stabilizer and a preparation method of the compound amino acid injection. The stabilizer is composed of cysteine and cystine according to a weight ratio of (5-80): 1. The compound amino acid injection comprises the stabilizer and amino acid components. The preparation method of the compound amino acid injection comprises the following steps: preparing a first amino acid solution under a nitrogen protection condition; adjusting the pH value of the first amino acid solution to be neutral, adding cysteine and cystine, dissolving, and adding the balance of water for injection to obtain a second amino acid solution; and sterilizing the second amino acid solution to obtain the compound amino acid injection. According to the invention, cysteine and cystine are adopted as stabilizers of the compound amino acid injection, so that the purpose of stabilizing other amino acid impurities is achieved, and the safety is high.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceutical preparations, and particularly relates to a stabilizer for a compound amino acid injection, a compound amino acid injection containing the stabilizer, and a method for preparing the compound amino acid injection. Background Art

[0002] Compound amino acid injection is a parenteral nutrition drug, which plays an important role in nutritional therapy and is widely used in patients who cannot be given through the gastrointestinal tract or whose nutritional needs cannot be met through the gastrointestinal tract. In order to ensure the stability of amino acids in compound amino acid injection products, sulfites, such as sodium bisulfite, are usually added as antioxidants.

[0003] However, sulfites have potential toxic risks, which are manifested by inducing hypersensitivity reactions and damaging tissues and organs. Therefore, long-term use of amino acid injections containing sulfites may cause rashes, itching, and even anaphylactic shock in severe cases. In addition, it may cause damage to organs such as the liver and kidneys.

[0004] At present, some studies have tried to use other substances instead of sulfites to achieve the stability of compound amino acid injection. For example, patent application CN102861011A discloses a pharmaceutical composition and preparation containing 18 kinds of amino acids, in which EDTA and cysteine ​​hydrochloride are selected as antioxidants for compound amino acid injection. However, EDTA is an ion chelator that can bind to calcium ions, resulting in calcium in bones being chelated out, thereby causing osteoporosis in the human body.

[0005] Therefore, providing a stabilizer and a compound amino acid injection with high safety and good stabilization effect against specific amino acid impurities is a technical problem that urgently needs to be solved. Summary of the invention

[0006] In view of the above problems, the object of the present invention is to provide a stabilizer for a compound amino acid injection, a compound amino acid injection containing the stabilizer, and a method for preparing the compound amino acid injection.

[0007] The above object of the present invention is achieved by providing the following technical solutions:

[0008] In a first aspect, the present invention provides a stabilizer for a compound amino acid injection, which is composed of cysteine ​​and cystine, wherein the weight ratio of cysteine ​​to cystine is (5-80):1.

[0009] According to some embodiments of the present invention, the weight ratio of cysteine ​​to cystine is (10-80):1, preferably (10-55):1, and more preferably (15-55):1.

[0010] In a second aspect, the present invention provides a compound amino acid injection, which comprises the stabilizer according to the present invention and amino acid components other than cysteine ​​and cystine.

[0011] According to some embodiments of the present invention, the weight ratio of the stabilizer to the amino acid components other than cysteine ​​and cystine is 1: (60-150); preferably, the weight ratio of the stabilizer to the amino acid components is 1: (70-130).

[0012] According to some embodiments of the present invention, based on a total amount of 1000 ml, the compound amino acid injection contains 0.81 to 1.44 g of the stabilizer; preferably, the compound amino acid injection contains 0.81 to 1.21 g of the stabilizer; more preferably, the compound amino acid injection contains 0.81 to 1.05 g of the stabilizer.

[0013] According to some embodiments of the present invention, based on a total amount of 1000 ml, the compound amino acid injection contains 0.80-1.20 g cysteine ​​and 0.01-0.24 g cystine; preferably, the compound amino acid injection contains 0.80-1.10 g cysteine ​​and 0.01-0.11 g cystine; more preferably, the compound amino acid injection contains 0.80-1.10 g cysteine ​​and 0.02-0.11 g cystine; further preferably, the compound amino acid injection contains 0.80-1.00 g cysteine ​​and 0.02-0.05 g cystine.

[0014] According to some embodiments of the present invention, based on a total amount of 1000 ml, the compound amino acid injection contains 80 to 125 g of amino acid components other than cysteine ​​and cystine; preferably, the compound amino acid injection contains 82 to 123 g of amino acid components other than cysteine ​​and cystine; more preferably, the compound amino acid injection contains 90 to 115 g of amino acid components other than cysteine ​​and cystine.

[0015] According to some embodiments of the invention, the amino acid components other than cysteine ​​and cystine include isoleucine, leucine, lysine acetate, methionine, phenylalanine, threonine, tryptophan, valine, alanine, arginine, aspartic acid, glutamic acid, histidine, proline, serine, tyrosine and glycine.

[0016] According to some embodiments of the present invention, based on a total amount of 1000 ml, the compound amino acid injection comprises 6.40-9.60 g isoleucine, 11.20-16.80 g leucine, 11.84-17.76 g lysine acetate, 3.12-4.68 g methionine, 5.60-8.40 g phenylalanine, 4.56-6.84 g threonine, 1.60-2.40 g tryptophan, and 6.40-9.60 g valine. , 6.40-9.60g alanine, 6.72-10.08g arginine, 0.80-1.20g aspartic acid, 0.80-1.20g glutamic acid, 4.00-6.00g histidine, 4.00-6.00g proline, 2.40-3.60g serine, 0.40-0.60g tyrosine, 4.72-7.08g glycine, 0.80-1.20g cysteine ​​and 0.01-0.24g cystine.

[0017] Preferably, based on a total amount of 1000 ml, the compound amino acid injection comprises 7.20-8.80 g isoleucine, 12.60-15.40 g leucine, 13.32-16.28 g lysine acetate, 3.51-4.29 g methionine, 6.30-7.70 g phenylalanine, 5.13-6.27 g threonine, 1.80-2.20 g tryptophan, 7.20-8.80 g valine, 7.2 0-8.80 g alanine, 7.56-9.24 g arginine, 0.90-1.10 g aspartic acid, 0.90-1.10 g glutamic acid, 4.50-5.40 g histidine, 4.50-5.40 g proline, 2.70-3.30 g serine, 0.45-0.55 g tyrosine, 5.31-6.49 g glycine, 0.80-1.10 g cysteine ​​and 0.01-0.11 g cystine.

[0018] More preferably, based on a total amount of 1000 ml, the compound amino acid injection comprises 7.20-8.80 g isoleucine, 12.60-15.40 g leucine, 13.32-16.28 g lysine acetate, 3.51-4.29 g methionine, 6.30-7.70 g phenylalanine, 5.13-6.27 g threonine, 1.80-2.20 g tryptophan, 7.20-8.80 g valine, 7.2 0-8.80 g alanine, 7.56-9.24 g arginine, 0.90-1.10 g aspartic acid, 0.90-1.10 g glutamic acid, 4.50-5.40 g histidine, 4.50-5.40 g proline, 2.70-3.30 g serine, 0.45-0.55 g tyrosine, 5.31-6.49 g glycine, 0.80-1.10 g cysteine ​​and 0.02-0.11 g cystine.

[0019] Further preferably, based on a total amount of 1000 ml, the compound amino acid injection contains 8.00 g isoleucine, 14.00 g leucine, 14.80 g lysine acetate, 3.90 g methionine, 7.00 g phenylalanine, 5.70 g threonine, 2.00 g tryptophan, 8.00 g valine, 8.00 g alanine, 8.40 g arginine, 1.00 g aspartic acid, 1.00 g glutamic acid, 5.00 g histidine, 5.00 g proline, 3.00 g serine, 0.50 g tyrosine, 5.90 g glycine, 1.00 g cysteine ​​and 0.05 g cystine.

[0020] According to some embodiments of the present invention, the pH value of the compound amino acid injection is 6.5-7.5, preferably 7.0.

[0021] In a third aspect, the present invention provides a method for preparing the compound amino acid injection according to the present invention, which comprises the following steps:

[0022] (1) Under nitrogen protection, adding a portion of water for injection to the amino acid components other than cysteine ​​and cystine, and dissolving at 60 to 80° C. to obtain a first amino acid solution;

[0023] (2) adjusting the pH value of the first amino acid solution to neutral, adding cysteine ​​and cystine, and adding the remaining amount of water for injection after dissolution to obtain a second amino acid solution;

[0024] (3) sterilizing the second amino acid solution to obtain a compound amino acid injection solution;

[0025] Alternatively, it comprises the following steps:

[0026] (1) Under nitrogen protection, adding a portion of water for injection to the amino acid components other than glutamic acid, cysteine ​​and cystine, and dissolving at 60 to 80° C. to obtain a first amino acid solution;

[0027] (2) lowering the temperature of the first amino acid solution to below 40° C. and adjusting its pH value to neutral, adding glutamic acid, cysteine ​​and cystine, and adding the remaining amount of water for injection after dissolution to obtain a second amino acid solution;

[0028] (3) Sterilizing the second amino acid solution to obtain a compound amino acid injection solution.

[0029] According to some embodiments of the present invention, in step (1), the partial water for injection is 70% to 90% by weight of the total water for injection.

[0030] According to some embodiments of the present invention, in step (1), the nitrogen protection condition is to seal and continuously fill with nitrogen to maintain a positive nitrogen pressure.

[0031] According to some embodiments of the present invention, in step (1), the dissolved oxygen of the first amino acid solution is controlled to be less than 300 ppb by continuous stirring and nitrogen filling.

[0032] According to some embodiments of the present invention, in step (2), the pH value of the first amino acid solution is adjusted to 6.5-7.5 using glacial acetic acid.

[0033] According to some embodiments of the present invention, in step (2), the dissolved oxygen of the second amino acid solution is less than 300 ppb.

[0034] According to some embodiments of the present invention, in step (1) and step (2), the water for injection is deoxygenated water for injection.

[0035] According to some embodiments of the present invention, in step (3), the second amino acid solution is sterilized after filling; preferably, the second amino acid solution is filtered before filling; further preferably, a polyethersulfone filter element is used for filtration; more preferably, the pore size of the polyethersulfone filter element is 0.22 µm.

[0036] According to some embodiments of the present invention, in step (3), the headspace oxygen of the second amino acid injection after filling is controlled within 0.5%.

[0037] According to some embodiments of the present invention, in step (3), the sterilization is performed by spraying superheated water; preferably, the sterilization conditions are a sterilization temperature of 121±2°C, a sterilization time of 8 to 12 minutes, and an F0 value greater than 8.

[0038] In a fourth aspect, the present invention provides use of the compound amino acid injection according to the present invention in the preparation of a medicament for amino acid supplementation in patients with hypoproteinemia, malnutrition and / or pre- and post-operative conditions.

[0039] In a fifth aspect, the present invention provides use of the stabilizer according to the present invention in the preparation of a medicament for improving the stability of a compound amino acid preparation.

[0040] Compared with the prior art, the technical solution of the present invention has the following beneficial effects:

[0041] 1. The present invention uses a combination of cysteine ​​and cystine as a stabilizer for the compound amino acid injection to achieve the effect of stabilizing other amino acids. Since cystine and cysteine ​​are both natural amino acids, they are safer as stabilizers and have a good effect on reducing specific impurities.

[0042] 2. Experimental verification shows that compared with the combination of cysteine ​​and sodium bisulfite, the stabilizer of the present invention can achieve a comparable or even better stabilization effect, while the safety is greatly improved. Compared with other stabilizers, such as the combination of cysteine ​​and EDTA, the stabilizer of the present invention can greatly improve the performance of the compound amino acid injection in reducing several specific impurities.

[0043] 3. Experimental results show that the amino acid compound injection of the present invention has good clarity and transmittance. DETAILED DESCRIPTION

[0044] The present invention is further described in detail below in conjunction with specific examples. The examples given are only for illustrating the present invention, rather than for limiting the scope of the present invention.

[0045] The experimental methods used in the following examples and comparative examples are conventional methods unless otherwise specified. The reagents used in the following examples and comparative examples are commercially available products unless otherwise specified.

[0046]

[0047] Example 1

[0048] The prescription of the compound amino acid injection of this example is shown in Table 1.

[0049]

[0050] The preparation process of the compound amino acid injection of this embodiment is as follows:

[0051] Weigh the prescribed amount of amino acid APIs according to Table 1 and set aside. Add amino acid raw materials (16 kinds of amino acids except glutamic acid, cysteine ​​and cystine) under positive nitrogen pressure, seal and continuously fill with nitrogen for standby, and add deoxygenated water for injection to 90% of the preparation volume in the preparation tank. Maintain the temperature at 65 to 70°C, start stirring, continue to pass nitrogen, and control the dissolved oxygen to <300ppb. Cool down to 40°C, add glacial acetic acid to adjust the pH value (6.5~7.5 (target value at 25°C: 7.0)), and add glutamic acid, cysteine, and cystine. Rinse the tank wall with deoxygenated water at 40°C and stir to dissolve. Control the dissolved oxygen (less than 300ppb), and add deoxygenated water for injection to the preparation tank to the full amount. Use a polyethersulfone filter element (pore size 0.22µm) for filtration. After filling the drug solution, each bottle is vacuumed and filled with nitrogen several times, then plugged and capped, and the headspace oxygen is controlled within 0.5%. Use superheated water spray sterilization, sterilization conditions: sterilization temperature (121℃), sterilization time (12min), F0 value (12). Packaging after light inspection.

[0052] Example 2

[0053] The prescription of the compound amino acid injection of this example is shown in Table 2.

[0054]

[0055] The preparation process of the compound amino acid injection of this embodiment is as follows:

[0056] Weigh the prescribed amount of amino acid APIs according to Table 2 and set aside. Add amino acid raw materials (17 kinds of amino acids except cysteine ​​and cystine) under positive nitrogen pressure, seal and continuously fill with nitrogen for standby use, and add deoxygenated water for injection to 70% of the preparation volume in the preparation tank. Maintain the temperature at 60 to 65°C, start stirring, and continuously pass nitrogen to control the dissolved oxygen to <300ppb. Add glacial acetic acid to adjust the pH value (6.5~7.5 (target value at 25°C: 7.0)), and add cysteine ​​and cystine. Rinse the tank wall with deoxygenated water and stir to dissolve. Control the dissolved oxygen (less than 300ppb), and add deoxygenated water for injection to the full amount in the preparation tank. Use a polyethersulfone filter element (pore size 0.22µm) for filtration. After filling the drug solution, each bottle is vacuumed and filled with nitrogen several times, then plugged and capped, and the headspace oxygen is controlled within 0.5%. Use superheated water spray sterilization, sterilization conditions: sterilization temperature (121℃), sterilization time (12min), F0 value (12). Packaging after light inspection.

[0057] Example 3

[0058] The prescription of the compound amino acid injection of this example is shown in Table 3.

[0059]

[0060] The preparation process of the compound amino acid injection of this embodiment is as follows:

[0061] Weigh the amino acid APIs in the prescribed amount according to Table 3 and set aside. Add the amino acid raw materials (16 kinds of amino acids except glutamic acid, cysteine ​​and cystine) under positive nitrogen pressure, seal and continuously fill with nitrogen for standby use, and add deoxygenated water for injection to 80% of the preparation amount in the preparation tank. Maintain the temperature at 70 to 75°C, start stirring, and continuously pass nitrogen to control the dissolved oxygen to <300ppb. Add glacial acetic acid to adjust the pH value (6.5~7.5 (target value at 25°C: 7.0)), cool to 30°C, and then add glutamic acid, cysteine, and cystine in turn. Rinse the tank wall with deoxygenated water and stir to dissolve. Control the dissolved oxygen (less than 300ppb), and add deoxygenated water for injection to the full amount in the preparation tank. Use a polyethersulfone filter element (pore size 0.22µm) for filtration. After filling the drug solution, each bottle is vacuumed and filled with nitrogen several times, then plugged and capped, and the headspace oxygen is controlled within 0.5%. Use superheated water spray sterilization, sterilization conditions: sterilization temperature (121℃), sterilization time (8min), F0 value (8). Packaging after light inspection.

[0062] Example 4 to Example 5

[0063] The prescriptions of the compound amino acid injections of Examples 4 and 5 are shown in Table 4.

[0064]

[0065] The preparation process of the compound amino acid injection of Examples 4 and 5 is basically the same as that of Example 1, except that the dosage of various amino acids is adjusted according to the prescription amount in Table 4.

[0066] Comparative Example 1

[0067] The compound amino acid injection of this comparative example uses cysteine ​​+ sodium bisulfite as antioxidant and stabilizer, and the prescription is shown in Table 5.

[0068]

[0069] The preparation process of the compound amino acid injection of this comparative example is as follows:

[0070] Weigh the prescribed amount of amino acid APIs according to Table 5 and set aside. Under nitrogen positive pressure protection, add 16 kinds of amino acid raw materials except glutamic acid and cysteine ​​to the preparation tank, seal and continuously fill with nitrogen for standby, and add 70% of the prepared amount of deoxygenated water for injection. Maintain the temperature at 65 to 70°C, start stirring, continuously pass nitrogen, and control the dissolved oxygen to <300ppb. Stir until completely dissolved. Cool down to 40°C, add glacial acetic acid to adjust the pH value (6.5~7.5 (target value at 25°C: 7.0)), add sodium bisulfite, glutamic acid and cysteine, rinse the tank wall with a small amount of deoxygenated water, and stir to dissolve. Control the dissolved oxygen (less than 300ppb), and add deoxygenated water for injection to the full amount in the preparation tank. Use a polyethersulfone filter element (pore size 0.22µm) for filtration. After filling the drug solution, each bottle is vacuumed and filled with nitrogen several times, then plugged and capped, and the headspace oxygen is controlled within 0.5%. Use superheated water spray sterilization, sterilization conditions: sterilization temperature (121℃), sterilization time (8min), F0 value (8). Packaging after light inspection.

[0071] Comparative Example 2

[0072] The compound amino acid injection of this comparative example uses cysteine ​​+ EDTA as a stabilizer, and the prescription is shown in Table 6.

[0073]

[0074] The preparation process of the compound amino acid injection of this comparative example is as follows:

[0075] Weigh the prescribed amount of amino acid APIs according to Table 6 and set aside. Under positive nitrogen pressure, add 17 kinds of amino acid raw materials except cysteine ​​to the preparation tank, seal and continuously fill with nitrogen for standby use, and add deoxygenated water for injection to 80% of the preparation amount. Maintain the temperature at 70 to 75°C, start stirring, continue to pass nitrogen, control the dissolved oxygen to <300ppb, add glacial acetic acid and EDTA to adjust the pH value (6.5~7.5 (target value at 25°C: 7.0)), and add cysteine. Rinse the tank wall with a small amount of deoxygenated water and stir to dissolve. Control the dissolved oxygen (less than 300ppb), and add deoxygenated water for injection to the full amount in the preparation tank. Use a polyethersulfone filter element (pore size 0.22µm) for filtration. After filling the liquid, each bottle is evacuated and filled with nitrogen several times, then plugged and capped, and the headspace oxygen is controlled within 0.5%. Use superheated water spray sterilization, sterilization conditions: sterilization temperature (121℃), sterilization time (12min), F0 value (12); package after light inspection.

[0076] Comparative Example 3

[0077] The compound amino acid injection of this comparative example only adds cysteine ​​as a stabilizer, and the prescription is shown in Table 7.

[0078]

[0079] The preparation process of the compound amino acid injection of this comparative example is as follows:

[0080] Weigh the prescribed amount of amino acid APIs according to Table 7 and set aside. Under nitrogen positive pressure, add 16 kinds of amino acid raw materials except glutamic acid and cysteine ​​to the preparation tank, seal and continuously fill with nitrogen for standby, and add deoxygenated water for injection to 80% of the preparation amount. Maintain the temperature at 70 to 75°C, start stirring, continue to pass nitrogen, and control the dissolved oxygen to <300ppb. Stir until completely dissolved. Cool down to 40°C, add glacial acetic acid to adjust the pH value (6.5~7.5 (target value at 25°C: 7.0)), and add glutamic acid and cysteine. Rinse the tank wall with a small amount of deoxygenated water and stir to dissolve. Control the dissolved oxygen (less than 300ppb), and add deoxygenated water for injection to the full amount in the preparation tank. Sampling and central control. Filter with a polyethersulfone filter element (pore size 0.22µm). After filling the drug solution, each bottle is vacuumed and filled with nitrogen several times, then plugged and capped, and the headspace oxygen is controlled within 0.5%. Use superheated water spray sterilization, sterilization conditions: sterilization temperature (121℃), sterilization time (12min), F0 value (12); package after light inspection.

[0081] Comparative Example 4

[0082] The compound amino acid injection of this comparative example adds cysteine ​​+ acetylcysteine ​​as a stabilizer, and the prescription is shown in Table 8.

[0083]

[0084] The preparation process of the compound amino acid injection of this comparative example is as follows:

[0085] Weigh the prescribed amount of amino acid APIs according to Table 8 and set aside. Under nitrogen positive pressure, add amino acid raw materials (16 kinds of amino acids except glutamic acid, cysteine ​​and acetylcysteine), seal and continuously fill with nitrogen for standby, and add deoxygenated water for injection to 90% of the preparation amount in the preparation tank. Maintain the temperature at 65 to 70°C, start stirring, and continuously pass nitrogen to control the dissolved oxygen to <300ppb. Cool down to 40°C, add glacial acetic acid to adjust the pH value (6.5~7.5 (target value at 25°C: 7.0)), and add glutamic acid, cysteine, and acetylcysteine. Rinse the tank wall with deoxygenated water at 40°C and stir to dissolve. Control the dissolved oxygen (less than 300ppb), and add deoxygenated water for injection to the full amount in the preparation tank. Use a polyethersulfone filter element (pore size 0.22µm) for filtration. After filling the drug solution, each bottle is vacuumed and filled with nitrogen several times, then plugged and capped, and the headspace oxygen is controlled within 0.5%. Use superheated water spray sterilization, sterilization conditions: sterilization temperature (121℃), sterilization time (12min), F0 value (12). Packaging after light inspection.

[0086] Comparative Example 5

[0087] The compound amino acid injection of this comparative example adds cysteine ​​+ cystine as a stabilizer, and the prescription is shown in Table 9.

[0088]

[0089] The preparation process of the compound amino acid injection of this comparative example is as follows:

[0090] Weigh the amino acid APIs in the prescribed amount according to Table 9 and set aside. Add the amino acid raw materials (16 kinds of amino acids except glutamic acid, cysteine ​​and cystine) under positive nitrogen pressure, seal and continuously fill with nitrogen for standby use, and add deoxygenated water for injection to 80% of the preparation amount in the preparation tank. Maintain the temperature at 70 to 75°C, start stirring, and continuously pass nitrogen to control the dissolved oxygen to <300ppb. Add glacial acetic acid to adjust the pH value (6.5~7.5 (target value at 25°C: 7.0)), cool to 30°C, and then add glutamic acid, cysteine, and cystine in turn. Rinse the tank wall with deoxygenated water and stir to dissolve. Control the dissolved oxygen (less than 300ppb), and add deoxygenated water for injection to the full amount in the preparation tank. Use a polyethersulfone filter element (pore size 0.22µm) for filtration. After filling the drug solution, each bottle is vacuumed and filled with nitrogen several times, then plugged and capped, and the headspace oxygen is controlled within 0.5%. Use superheated water spray sterilization, sterilization conditions: sterilization temperature (121℃), sterilization time (8min), F0 value (8). Packaging after light inspection.

[0091] Stability test

[0092] 1. Investigation of influencing factors: Place under high temperature conditions of 50°C for 10 days and 30 days for sampling and testing;

[0093] Accelerated condition test: Store at 40℃ for 1 month and 3 months for sampling and testing.

[0094] 2. Methionine sulfoxide and pyroglutamic acid

[0095] Analytical method: Determined by high performance liquid chromatography (Chinese Pharmacopoeia 2020 Edition, Part IV, General Rules 0512).

[0096] Test solution: Accurately measure 2 ml of the product, place it in a 10 ml volumetric flask, dilute to the scale with water, and shake well.

[0097] Reference solution: Take appropriate amount of methionine sulfoxide and pyroglutamic acid reference substances, weigh accurately, dissolve in water and quantitatively dilute to make a solution containing 4 μg of methionine sulfoxide and 8 μg of pyroglutamic acid per 1 ml.

[0098] Chromatographic conditions: Octadecyl silane bonded silica gel was used as filler. Phosphate buffer (prepared with 0.03 mol / L potassium dihydrogen phosphate and 0.01 mol / L sodium octane sulfonate in water, pH adjusted to 1.5 with phosphoric acid solution)-acetonitrile (volume ratio of 95:5) was used as mobile phase A, acetonitrile was used as mobile phase B, gradient elution was performed according to the table below, flow rate was 1.0 ml per minute; column temperature was 35°C; detection wavelength was 205 nm.

[0099]

[0100] Determination method: Accurately measure the test solution and the reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0101] 3. Tyrosine, phenylalanine, methionine and tryptophan

[0102] Analytical method: Determined by high performance liquid chromatography (Chinese Pharmacopoeia 2020 Edition, Part IV, General Rules 0512).

[0103] Test solution: Accurately measure 1 ml of the product, place it in a 20 ml volumetric flask, add water to dilute to the scale, and shake well.

[0104] Reference solution: Take appropriate amount of tyrosine, phenylalanine, methionine and tryptophan reference substances, weigh accurately, dissolve in water and quantitatively dilute to a mixed solution containing 25 μg tyrosine, 350 μg phenylalanine, 195 μg methionine and 100 μg tryptophan per 1 ml.

[0105] Chromatographic conditions: octadecylsilane bonded silica gel was used as filler; 0.03 mol / L triethylamine aqueous solution (pH adjusted to 2.0 with phosphoric acid)-methanol (volume ratio of 95:5) was used as mobile phase; flow rate was 1.0 ml per minute; column temperature was 30°C; detection wavelength was 280 nm.

[0106] System suitability requirements: The separation of each peak in the chromatogram of the reference solution should meet the requirements, and the number of theoretical plates based on the tryptophan peak should not be less than 2000.

[0107] Determination method: Accurately measure the test solution and the reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0108] 4. Dioxindole Alanine, Kynurenine, 2-Hydroxytryptophan

[0109] Analytical method: Determined by high performance liquid chromatography (Chinese Pharmacopoeia 2020 Edition, Part IV, General Rules 0512).

[0110] Test solution: Accurately measure 2 ml of the product, place it in a 10 ml volumetric flask, dilute to the scale with water, and shake well.

[0111] Reference solution: Take appropriate amount of dioxindolealanine, kynurenine and 2-hydroxytryptophan reference substances, weigh accurately, dissolve in water and quantitatively dilute to make a mixed solution containing 0.8 μg of dioxindolealanine, 0.8 μg of kynurenine and 0.8 μg of 2-hydroxytryptophan per 1 ml.

[0112] Chromatographic conditions: octadecyl bonded silica gel was used as filler; phosphate buffer (prepared with 0.02 mol / l diammonium hydrogen phosphate solution, adjusted to pH 8.0 with ammonia water)-methanol (volume ratio of 95:5) was used as mobile phase A, water-acetonitrile (volume ratio of 50:50) was used as mobile phase B, and gradient elution was performed according to the table below; the flow rate was 1.0 ml / min; the column temperature was 30°C; the detection wavelength was 210 nm.

[0113]

[0114] Determination method: Accurately measure the test solution and the reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0115] 5. Phenylethylamine

[0116] Analytical method: Determined by high performance liquid chromatography (Chinese Pharmacopoeia 2020 Edition, Part IV, General Rules 0512).

[0117] Test solution: Accurately measure 3 ml of the product, place it in a 20 ml volumetric flask, dilute to the scale with water, and shake well.

[0118] Reference solution: Take an appropriate amount of phenethylamine reference substance, weigh accurately, dissolve in water and quantitatively dilute to make a solution containing 1 μg of phenethylamine per 1 ml.

[0119] System suitability solution: Take 3 ml of this product and an appropriate amount of phenylethylamine reference substance, place in 20 ml, add water to dilute to make a mixed solution containing this product and 1 μg / ml phenylethylamine.

[0120] Chromatographic conditions: octadecyl bonded silica gel was used as filler; phosphate buffer (an aqueous solution of 0.03 mol / L dipotassium hydrogen phosphate and 0.01 mol / L sodium octane sulfonate, adjusted to pH 4.0 with phosphoric acid)-methanol (volume ratio of 70:30) was used as mobile phase; flow rate was 0.5 ml / min; column temperature was 30°C; detection wavelength was 210 nm.

[0121] Determination method: Accurately measure the test solution and the reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0122] 6. Clarity and color of compound amino acid injection: Take this product and measure it according to the law (General Rules 0901 Method 1 and General Rules 0902 Method 1 of Part IV of the 2020 Edition of the Chinese Pharmacopoeia). The solution should be clear and colorless. If color is developed, it shall not be darker than the yellow No. 2 standard colorimetric solution.

[0123] 7. Transmittance of compound amino acid injection: Take this product and measure it according to the UV-visible spectrophotometry (General Rules 0401 of Part IV of the Chinese Pharmacopoeia 2020 Edition). The transmittance is measured at a wavelength of 430 nm and shall not be less than 97.0%.

[0124] The experimental results of Examples 1 to 5 and Comparative Examples 1 to 5 are shown in Tables 10 to 19, respectively.

[0125]

[0126]

[0127]

[0128]

[0129]

[0130]

[0131]

[0132]

[0133]

[0134]

[0135] The above results show that when cysteine ​​+ sodium bisulfite is used as an antioxidant, the tryptophan impurity is significantly increased and exceeds the limit, the content of other impurities is maintained at a low level, and the color of the amino acid injection changes slightly; when cysteine ​​+ EDTA is used as an antioxidant, some tryptophan impurities are significantly increased and exceed the limit, the pyroglutamic acid content is slightly increased, and the color of the amino acid injection changes slightly; when cysteine ​​is used alone as an antioxidant, the tryptophan and methionine impurities are significantly increased and exceed the limit, the pyroglutamic acid content is significantly increased and exceeds the limit, and the color of the amino acid injection changes significantly, becoming a yellow liquid; when cysteine ​​+ acetylcysteine ​​is used as an antioxidant, the tryptophan impurity is significantly increased and exceeds the limit, the pyroglutamic acid content is significantly increased, and the color of the amino acid injection changes significantly, becoming a yellow liquid; when cysteine ​​and cysteine ​​are used as antioxidants, but the ratio of the two components is not within the protection scope of the present invention, the tryptophan impurity is significantly increased and exceeds the limit, the pyroglutamic acid content is significantly increased, and the color of the amino acid injection changes significantly, becoming a yellow liquid.

[0136] The compound amino acid injection using the stabilizer of the present invention always maintains a colorless transparent liquid state under accelerated conditions, and the pH and transmittance remain stable. The content of easily degradable amino acids such as methionine, tyrosine, phenylalanine and tryptophan remains stable, and is greater than 98% by weight. At 3 months under accelerated conditions, the contents of impurities such as methionine sulfoxide, dioxindole alanine, kynurenine, 2-hydroxytryptophan, phenylethylamine and pyroglutamic acid all meet the standard requirements, and compared with the comparative examples using cysteine+sodium bisulfite, cysteine+EDTA, cysteine+acetylcysteine ​​and cysteine, the impurity content is significantly lower. These results all show that the stabilizer of the present invention can achieve a better effect of reducing impurities and improving stability.

[0137] The above are only several exemplary embodiments of the present invention, and are not intended to limit the present invention in any form. Although the present invention is disclosed as above with preferred embodiments, they are not intended to limit the present invention. Any technician familiar with the profession, without departing from the scope of the technical solution of the present invention, can make some changes or modifications to obtain equivalent or equivalent embodiments using the above disclosed technical content, all of which belong to the scope of the present invention.

Claims

1. A stabilizer for compound amino acid injection, which consists of cysteine ​​and cystine, wherein: The weight ratio of cysteine ​​to cystine is (5-80):

1.

2. The stabilizer according to claim 1, wherein The weight ratio of cysteine ​​to cystine is (10-80):1, preferably (10-55):1, and more preferably (15-55):

1.

3. A compound amino acid injection, comprising the stabilizer according to claim 1 or 2 and an amino acid component other than cysteine ​​and cystine; Preferably, the weight ratio of the stabilizer to the amino acid components other than cysteine ​​and cystine is 1:(60-150); more preferably, the weight ratio of the stabilizer to the amino acid components other than cysteine ​​and cystine is 1:(70-130).

4. The compound amino acid injection according to claim 3, wherein: Based on a total amount of 1000 ml, the compound amino acid injection contains 0.81 to 1.44 g, preferably 0.81 to 1.21 g, and more preferably 0.81 to 1.05 g of the stabilizer; Further preferably, based on a total amount of 1000 ml, the compound amino acid injection contains 0.80-1.20 g cysteine ​​and 0.01-0.24 g cystine, preferably 0.80-1.10 g cysteine ​​and 0.01-0.11 g cystine, more preferably 0.80-1.10 g cysteine ​​and 0.02-0.11 g cystine, further preferably 0.80-1.00 g cysteine ​​and 0.02-0.05 g cystine.

5. The compound amino acid injection according to claim 3 or 4, wherein: Based on a total amount of 1000 ml, the compound amino acid injection contains 80 to 125 g, preferably 90 to 115 g of amino acid components other than cysteine ​​and cystine; Further preferably, the amino acid components other than cysteine ​​and cystine include isoleucine, leucine, lysine acetate, methionine, phenylalanine, threonine, tryptophan, valine, alanine, arginine, aspartic acid, glutamic acid, histidine, proline, serine, tyrosine and glycine.

6. The compound amino acid injection according to claim 5, wherein: Based on a total volume of 1000 ml, the compound amino acid injection contains 6.40-9.60 g isoleucine, 11.20-16.80 g leucine, 11.84-17.76 g lysine acetate, 3.12-4.68 g methionine, 5.60-8.40 g phenylalanine, 4.56-6.84 g threonine, 1.60-2.40 g tryptophan, 6.40-9.60 g valine, 6.40-9.70 g valine, .60g alanine, 6.72-10.08g arginine, 0.80-1.20g aspartic acid, 0.80-1.20g glutamic acid, 4.00-6.00g histidine, 4.00-6.00g proline, 2.40-3.60g serine, 0.40-0.60g tyrosine, 4.72-7.08g glycine, 0.80-1.20g cysteine ​​and 0.01-0.24g cystine; Preferably, based on a total amount of 1000 ml, the compound amino acid injection comprises 7.20-8.80 g isoleucine, 12.60-15.40 g leucine, 13.32-16.28 g lysine acetate, 3.51-4.29 g methionine, 6.30-7.70 g phenylalanine, 5.13-6.27 g threonine, 1.80-2.20 g tryptophan, 7.20-8.80 g valine, 7.2 0-8.80 g alanine, 7.56-9.24 g arginine, 0.90-1.10 g aspartic acid, 0.90-1.10 g glutamic acid, 4.50-5.40 g histidine, 4.50-5.40 g proline, 2.70-3.30 g serine, 0.45-0.55 g tyrosine, 5.31-6.49 g glycine, 0.80-1.10 g cysteine ​​and 0.01-0.11 g cystine; More preferably, based on a total amount of 1000 ml, the compound amino acid injection comprises 7.20-8.80 g isoleucine, 12.60-15.40 g leucine, 13.32-16.28 g lysine acetate, 3.51-4.29 g methionine, 6.30-7.70 g phenylalanine, 5.13-6.27 g threonine, 1.80-2.20 g tryptophan, 7.20-8.80 g valine, 7.2 0-8.80 g alanine, 7.56-9.24 g arginine, 0.90-1.10 g aspartic acid, 0.90-1.10 g glutamic acid, 4.50-5.40 g histidine, 4.50-5.40 g proline, 2.70-3.30 g serine, 0.45-0.55 g tyrosine, 5.31-6.49 g glycine, 0.80-1.10 g cysteine ​​and 0.02-0.11 g cystine; Further preferably, based on a total amount of 1000 ml, the compound amino acid injection comprises 8.00 g isoleucine, 14.00 g leucine, 14.80 g lysine acetate, 3.90 g methionine, 7.00 g phenylalanine, 5.70 g threonine, 2.00 g tryptophan, 8.00 g valine, 8.00 g alanine, 8.40 g arginine, 1.00 g aspartic acid, 1.00 g glutamic acid, 5.00 g histidine, 5.00 g proline, 3.00 g serine, 0.50 g tyrosine, 5.90 g glycine, 1.00 g cysteine ​​and 0.05 g cystine; Further preferably, the pH value of the compound amino acid injection is 6.5-7.5, preferably 7.

0.

7. The method for preparing the compound amino acid injection according to any one of claims 3 to 6, comprising the following steps: (1) Under nitrogen protection, adding a portion of water for injection to the amino acid components other than cysteine ​​and cystine, and dissolving at 60 to 80° C. to obtain a first amino acid solution; (2) adjusting the pH value of the first amino acid solution to neutral, adding cysteine ​​and cystine, and adding the remaining amount of water for injection after dissolution to obtain a second amino acid solution; (3) Sterilizing the second amino acid solution to obtain a compound amino acid injection solution.

8. The method for preparing the compound amino acid injection according to any one of claims 3 to 6, comprising the following steps: (1) Under nitrogen protection, adding a portion of water for injection to the amino acid components other than glutamic acid, cysteine ​​and cystine, and dissolving at 60 to 80° C. to obtain a first amino acid solution; (2) lowering the temperature of the first amino acid solution to below 40° C. and adjusting its pH value to neutral, adding glutamic acid, cysteine ​​and cystine, and adding the remaining amount of water for injection after dissolution to obtain a second amino acid solution; (3) Sterilizing the second amino acid solution to obtain a compound amino acid injection solution.

9. The preparation method according to claim 7 or 8, wherein: In step (1), the partial water for injection is 70% to 90% by weight of the total water for injection; Preferably, in step (1), the nitrogen protection condition is to seal and continuously fill with nitrogen to maintain a positive nitrogen pressure; Preferably, in step (1), the dissolved oxygen of the first amino acid solution is controlled to be less than 300 ppb by continuous stirring and nitrogen filling; Preferably, in step (2), the pH value of the first amino acid solution is adjusted to 6.5-7.5 using glacial acetic acid; Preferably, in step (2), after adding cysteine ​​and cystine, water for injection is first used for rinsing; Preferably, in step (2), the dissolved oxygen of the second amino acid solution is less than 300 ppb; Preferably, in step (1) and step (2), the water for injection is deoxygenated water for injection.

10. The preparation method according to claim 7 or 8, wherein: In step (3), the second amino acid solution is sterilized after filling; preferably, the second amino acid solution is filtered before filling; further preferably, a polyethersulfone filter element is used for filtration; more preferably, the pore size of the polyethersulfone filter element is 0.22 µm; Preferably, in step (3), the headspace oxygen of the second amino acid injection after filling is controlled within 0.5%; Preferably, in step (3), the sterilization is performed by spraying superheated water; preferably, the sterilization conditions are a sterilization temperature of 121±2°C, a sterilization time of 8 to 12 minutes, and an F0 value greater than 8.

11. Use of the compound amino acid injection according to any one of claims 3 to 9 in the preparation of a medicament for amino acid supplementation in patients with hypoproteinemia, malnutrition and / or pre- and post-operative conditions.

12. Use of the stabilizer according to claim 1 or 2 in the preparation of a medicament for improving the stability of a compound amino acid preparation.

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