Antioxidant-free kanamycin injection and preparation method thereof

Abstract

The invention discloses an antioxidant-free kanamycin injection and a preparation method thereof.The injection is composed of kanamycin sulfate, a pH regulator and water for injection, the pH is 4.5-7.0, headspace oxygen is controlled to be smaller than or equal to 1% through nitrogen charging, and the injection is completely free of sodium hydrogen sulfite, cysteine hydrochloride and other antioxidants and can be used for intravenous drip and intramuscular injection; the invention further provides a preparation process and an ion chromatography verification method which are compatible with an existing production line, and the product quality meets the requirements of Chinese Pharmacopoeia 2020 through high temperature, strong light and long-term stability test verification; clinical retrospective analysis shows that the rash occurrence rate is reduced to 3.1% from 27.3%, the safety is remarkably improved while the medicine quality is ensured, and the method has a good industrial application prospect.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical preparations, and in particular to a kanamycin injection solution without antioxidants and its preparation method. Background Technology

[0002] Kanamycin sulfate is an aminoglycoside antibiotic, clinically used for severe infections caused by Gram-negative bacteria. Because the kanamycin molecule contains multiple amino and hydroxyl groups, it is traditionally believed to be easily oxidized in solution. Therefore, antioxidants such as sodium bisulfite and cysteine ​​hydrochloride are commonly added to injectable solutions both domestically and internationally (ChP2020 General Rule 0102 recommends a sodium bisulfite concentration of 0.1%~0.2%). Sodium bisulfite is irritating to the skin and mucous membranes; excessive intake may cause rashes and itching. Therefore, this research group intends to prepare a higher-quality injectable solution by investigating the impact of different antioxidant formulations on quality. However, through systematic research by the inventors of this invention, it was found that:

[0003] 1. The measured values ​​of sodium bisulfite in the injection solutions of four domestic companies ranged from 0.24% to 0.37%, which were significantly higher than the prescription dosage and the upper limit in the pharmacopoeia.

[0004] 2. The original manufacturer (Meiji Seika) injectable solution contains only 0.05% sodium bisulfite, and the instructions do not mention any adverse reaction such as rash; at the same time, the original injectable solution contains antibacterial agents such as methylparaben and ethylparaben, and is only for intramuscular injection in clinical practice.

[0005] 3. Skin rash accounts for 27.3% of adverse reaction reports of kanamycin in China, and is related to excessive sodium bisulfite;

[0006] 4. After removing residual oxygen using a nitrogen purging process, kanamycin itself showed good stability under high temperature (60 ℃, 30 d) and strong light (4500 lx, 10 d) conditions, with no significant changes in related substances, content, pH, and color.

[0007] 5. The sterilization process (100 ℃, 30 min) and long-term storage (121 days after the expiry date) had no significant effect on the antioxidant content, proving that antioxidants are not necessary.

[0008] Therefore, the existing practice of adding antioxidants has drawbacks such as the risk of excessive dosage, increased adverse reactions, and cost waste. There is an urgent need for a kanamycin injection that does not require antioxidants, has stable quality, and is safer. Summary of the Invention

[0009] In view of the above situation and to overcome the shortcomings of the prior art, the purpose of this invention is to provide an antioxidant-free kanamycin injection and its preparation method, which can reduce the incidence of adverse reactions and simplify the prescription while ensuring drug quality.

[0010] The first aspect of the present invention is to provide an antioxidant-free kanamycin injection, wherein each 1 mL of the injection consists of: 250 mg kanamycin sulfate (calculated as kanamycin), water for injection added to 1 mL, and then a pH adjuster is added to make the pH value 4.5-7.0, and the injection is free of sodium bisulfite, cysteine ​​hydrochloride and other antioxidants.

[0011] Preferably, the headspace oxygen content of the injection solution after filling is ≤ 1.0% (volume fraction), which is achieved by nitrogen replacement.

[0012] Preferably, the pH adjuster is sulfuric acid.

[0013] Preferably, the pH value is 5.5-6.5.

[0014] A second aspect of the present invention is to provide a method for preparing an antioxidant-free kanamycin injection, comprising the following steps:

[0015] S1. In a Class 100 clean area, dissolve 700–750g of kanamycin sulfate in water for injection at 25–30℃ and stir under nitrogen protection until completely dissolved.

[0016] S2, using 0.1 mol·L -1 Adjust the pH to 4.5–7.0 with sulfuric acid;

[0017] S3. Add water for injection to 2000 ml, and circulate for 5 min under nitrogen protection;

[0018] S4. After being filtered through a 0.22 µm polyethersulfone filter, the ampoule is filled into a brown neutral borosilicate glass ampoule and sealed with nitrogen.

[0019] S5. Sterilize with flowing steam at 100℃ for 30 minutes, then perform leak detection, light inspection, and packaging to obtain the product.

[0020] In S4, the nitrogen flow rate is 1–2 L·min -1 Nitrogen filling time: 2–5 s.

[0021] A third aspect of the present invention is to provide a quality control method for the above-mentioned antioxidant-free kanamycin injection, comprising the following steps for stability testing:

[0022] S1. High temperature test: Place at 60℃ for 30 days, and take samples at 0, 5, 10 and 3 days to determine the content, related substances, pH and color;

[0023] S2. Light test: Place at 4500±500 lx for 10 days, and take samples on days 0, 5, and 10 for testing as above;

[0024] S3. Long-term test: Store at 25℃ ± 2℃ / 60% RH ± 5% RH for 12 months, and take samples at 0, 3, 6, 9 and 12 months for testing as above;

[0025] S4. Sodium bisulfite was quantified using ion chromatography (Dionex ICS-5000⁺, AS11-HC column, potassium hydroxide gradient elution). The detection limit was ≤0.005%. The detection result was lower than the detection limit, confirming that sodium bisulfite antioxidants were not added to the sample.

[0026] The beneficial technical effects of this invention are as follows:

[0027] 1. Antioxidants are completely removed from the prescription, avoiding adverse reactions such as rashes caused by excessive sodium bisulfite, thus significantly improving safety;

[0028] 2. By controlling headspace oxygen to ≤1% through nitrogen filling process, the stability of kanamycin itself can meet the requirements of pharmacopoeia and clinical practice;

[0029] 3. Simplify prescriptions, reduce raw material and quality inspection costs, and lessen environmental pressure;

[0030] 4. The manufacturing process is compatible with existing production lines, requiring no new equipment and is suitable for large-scale production;

[0031] 5. After being tested under high temperature, strong light and long-term conditions, the product has been found to be of stable quality, and its various indicators are comparable to those of the original product with 0.05% sodium bisulfite added, meeting the requirements of the National Pharmacopoeia and ICH guidelines. Attached Figure Description

[0032] Figure 1 This is a box plot of known impurities for antioxidant samples of different formulation amounts in this invention placed under high temperature and strong light irradiation conditions (the 3 points in the box plot correspond to the results of 0 days, high temperature and strong light irradiation conditions, respectively).

[0033] Figure 2 This invention relates to the effects of high temperature and strong light irradiation on related substances in samples without antioxidants.

[0034] Figure 3 The results show the content determination of antioxidant samples with different formulation amounts according to the present invention after being placed under high temperature and strong light irradiation.

[0035] Figure 4 This is a chromatogram of the sodium bisulfite reference solution of the present invention.

[0036] Figure 5 The chromatogram for the determination of sodium bisulfite in kanamycin sulfate injection of this invention is shown.

[0037] Figure 6 X-ray diffraction patterns of kanamycin monosulfate and kanamycin sulfate.

[0038] Figure 7 This is a particle size distribution diagram of kanamycin monosulfate and kanamycin sulfate.

[0039] Figure 8 Typical chromatograms for the determination of residual solvents in kanamycin monosulfate and kanamycin sulfate.

[0040] Figure 9 Comparison of related substance determination results between kanamycin monosulfate and kanamycin sulfate. Detailed Implementation

[0041] The specific embodiments of the present invention will be further described in detail below with reference to the examples and accompanying drawings.

[0042] Example 1

[0043] In a specific implementation of the present invention, a kanamycin injection solution without antioxidants is provided, each vial of which consists of: 500 mg kanamycin sulfate (calculated as kanamycin), 2 mL of water for injection, sulfuric acid to make the pH value 6, and is free of sodium bisulfite, cysteine ​​hydrochloride and other antioxidants.

[0044] The preparation method includes the following steps:

[0045] S1. In a Class 100 clean area, dissolve 700–750g of kanamycin sulfate in water for injection at 25°C and stir under nitrogen protection until completely dissolved.

[0046] S2, using 0.1 mol·L -1 Adjust the pH to 5.5 with sulfuric acid;

[0047] S3. Add water for injection to 2000 ml, and circulate for 5 min under nitrogen protection;

[0048] S4. Filtered through a 0.22 µm polyethersulfone filter, filled into brown neutral borosilicate glass ampoules, and purged with nitrogen for 3 s (flow rate 1.5 L·min). -1 Immediately melt and seal;

[0049] S5. Sterilize with flowing steam at 100℃ for 30 minutes, then perform leak detection, light inspection, and packaging. The headspace oxygen level is 0.8%.

[0050] Stability test

[0051] The sample from Example 1 was subjected to high temperature, strong light, and long-term tests according to the above quality control methods. Results:

[0052] Content (HPLC): 104.2% ± 0.4%;

[0053] Total impurities: 4.0%–4.5%;

[0054] pH: 5.8 ± 0.1;

[0055] Color: ≤ YG1;

[0056] Sodium bisulfite: Not detected (< 0.005%).

[0057] Comparative Example 1: Injection containing 0.3% sodium bisulfite

[0058] The prescription is the same as in Example 1, except that 0.3% sodium bisulfite is added, and the rest of the process is the same.

[0059] result:

[0060] Content (HPLC): 104.4% ± 0.3%;

[0061] Total impurities: 4.0%–4.5%;

[0062] pH: 5.8 ± 0.1;

[0063] Color: ≤ YG1;

[0064] Sodium bisulfite: 0.3%.

[0065] This invention provides an injection solution composed of kanamycin sulfate, sulfuric acid, and water for injection, with a pH of 4.5–7.0. Headspace oxygen is controlled to ≤1% through nitrogen purging. It is completely free of antioxidants such as sodium bisulfite and cysteine ​​hydrochloride, and is suitable for intravenous infusion and intramuscular injection. This invention also provides a preparation process and ion chromatography verification method compatible with existing production lines. High temperature, strong light, and long-term stability tests have verified that the product quality meets the requirements of the 2020 edition of the Chinese Pharmacopoeia. A retrospective clinical analysis showed that the incidence of rash decreased from 27.3% to 3.1%. This invention significantly improves safety while ensuring drug quality and has good prospects for industrial application. Relevant experimental data are as follows:

[0066] I. Effects of different formulation amounts of antioxidant samples on key quality attributes such as color, pH value, related substances, and content.

[0067] This invention commissioned an injection manufacturer to prepare samples with different formulation amounts of antioxidants using the same batch of raw materials, following the approved production process: no antioxidant, antioxidants with the original manufacturer's formulation amount, and antioxidants with formulation amounts from four domestic companies, as shown in Table 1. The samples were placed at high temperature (60℃) for 30 days and under strong light irradiation (4500 lx) for 10 days. Color, pH value, related substances, and their content were used as evaluation indicators to compare the mass changes of the samples with different formulation amounts of antioxidants under 0-day and high-temperature / strong-light irradiation conditions. Sodium bisulfite was determined using ion chromatography, with a detection limit ≤ 0.005%.

[0068] Table 1 Simulated preparation of antioxidant samples with different formulation amounts

[0069] prescription Sodium bisulfite Cysteine ​​hydrochloride Prescription 1 No additions No additions Prescription 2 0.05% No additions Prescription 3 0.3% No additions Prescription 4 0.2% 0.2%

[0070] like Figure 1-2 As shown, under conditions of 0 days, high temperature, and strong light irradiation, the colors of the four prescription antioxidant samples were all lighter than the yellow-green No. 1 standard colorimetric solution, indicating that the amount of antioxidant added had no effect on the color of the samples.

[0071] Under conditions of 0 days, high temperature, and strong light irradiation, the pH value of the four antioxidant samples with different formulation amounts did not differ by more than 0.1, and the amount of antioxidant added had no effect on the pH value of the samples.

[0072] Under conditions of 0 days, high temperature, and strong light irradiation, there were no significant differences in the content of various impurities and total impurities in the four formulations of antioxidant samples, indicating that the amount of antioxidant added has essentially no effect on related substances. Formulation 1 did not contain any antioxidant, but the related substances showed no change under high temperature and strong light irradiation conditions, indicating that the injection solution has good stability without the addition of antioxidant.

[0073] like Figure 3 As shown, under high temperature and strong light irradiation conditions, there was no significant difference in kanamycin content among the four prescription antioxidant samples, indicating that the amount of antioxidant added had no effect on the content.

[0074] Therefore, under high temperature and strong light irradiation conditions, the amount of different antioxidants added has virtually no effect on the color, pH value, related substances, and content of the samples. Even without adding antioxidants (Formula 1), the quality of the injection solution remains unchanged, demonstrating good stability. The reason for this is that all four formulations use a nitrogen-filling process to remove oxygen from the drug solution, resulting in low residual oxygen levels. Furthermore, kanamycin is an aminoglycoside antibiotic, which is inherently stable and not easily oxidized. Excessive sodium bisulfite can cause rashes, and rashes account for the largest proportion of adverse reactions to injection solutions in China (27.3%). The injection solution of this invention does not contain antioxidants, significantly reducing this risk. After completely removing antioxidants, the quality is essentially the same as the formulation with added antioxidants; key quality attributes such as content, related substances, and pH show no significant changes. However, without antioxidants such as sodium bisulfite, adverse reactions are fewer, and the safety is significantly superior to existing technologies.

[0075] II. Experiment on the selection of raw materials for this invention

[0076] 2.1 Experiment Content

[0077] By examining the key quality attributes of kanamycin monosulfate and kanamycin sulfate, such as crystal form, particle size, solubility, residual solvent, related substances, residual protein, genotoxic impurities, and elemental impurities, the rationality of using kanamycin sulfate as a raw material for injection is evaluated.

[0078] 2.2 Experimental Methods

[0079] (1) The crystal form was determined using a Shimadzu XRD-7000 powder XRD diffractometer with Cu Kα radiation from a graphite monochromator as the light source, tube voltage 40 kV, tube current 30 mA, 2θ scanning range 3°~60°, scanning rate 2° / min, and scanning interval 0.01°.

[0080] (2) Particle size was determined using a Mastersizar 2000 laser particle size analyzer by dry method. The shading was set to 0.5%~6%, the sample refractive index was 1.52, the absorptivity was 0.1, the vibration injection speed was 60%, the dispersion pressure was 0.8 bar, the hopper gap width was 1.5 mm, the background measurement duration was 10 s, and the sample measurement duration was 12 s.

[0081] (3) Solubility: Take appropriate amounts of kanamycin monosulfate and kanamycin sulfate raw materials and place them in 10 ml of water at 25°C. Shake vigorously for 30 seconds every 5 minutes and observe the solubility over 30 minutes.

[0082] (4) Residual solvent was tested using an Agilent 7980B gas chromatograph with a capillary column (DB-624, 30m×0.32mm, 1.8μm or equivalent) using 6% cyanopropylphenyl-94% dimethyl polysiloxane as the stationary phase. The initial temperature was 45℃, maintained for 5 minutes, and then increased to 240℃ at a rate of 20℃ per minute, maintained for 10 minutes. The injection port temperature was 220℃, the detector temperature was 250℃, the split ratio was 50:1, the carrier gas was nitrogen, the flow rate was 1.0 ml per minute, the headspace vial equilibration temperature was 90℃, and the equilibration time was 30 minutes.

[0083] (5) Methods for determining related substances:

[0084] A Dionex ICS-5000+ ion chromatograph with a YMC-Triart C18 ExRS (4.6×250mm, 5μm) column was used. 1.5g of sodium octane sulfonate and 20.0g of anhydrous sodium sulfate were dissolved in 50ml of 0.2mol / L phosphate buffer (pH 3.0, adjusted to pH 3.0 with phosphoric acid) and 900ml of water. 50ml of acetonitrile was added and mixed thoroughly to form mobile phase A. 1.5g of sodium octane sulfonate and 20.0g of anhydrous sodium sulfate were also dissolved in 50ml of 0.2mol / L phosphate buffer (pH 3.0, adjusted to pH 3.0 with phosphoric acid). Dissolve in 800 ml of water, add 150 ml of acetonitrile, mix well, and use as mobile phase B at a flow rate of 1.0 mL / min; perform linear gradient elution according to Table 2; column temperature is 35℃; detection is performed using an integrated pulsed amperometric electrochemical detector, with a gold electrode (3 mm in diameter) as the working electrode, an Ag / AgCl composite electrode as the reference electrode, and a titanium alloy counter electrode; four waveform detection potentials are used, and the table of four waveform detection potentials is shown in Table 3; add alkali (21 g / L sodium hydroxide solution, flow rate 0.5 mL / min) after column loading; injection volume is 20 μl.

[0085] Accurately measure an appropriate amount of this product and dilute it quantitatively with water to prepare a solution containing approximately 0.5 mg of kanamycin per ml as the test solution; accurately measure an appropriate amount of the test solution and dilute it quantitatively with water to prepare a solution containing approximately 5 μg of kanamycin per ml as the control solution; calculate the impurity content using the main component self-comparison method.

[0086] Table 2 Gradient elution procedure for related substances determination

[0087] Time (minutes) Mobile phase A (%) Mobile phase B (%) 0 60 40 12 60 40 12.1 35 65 32 35 65 32.1 60 40 50 60 40

[0088] Table 3 Four-waveform detection potential table

[0089] Time (seconds) Potential (V) integral 0.00 +0.10 0.20 +0.10 start 0.40 +0.10 Finish 0.41 -2.00 0.42 -2.00 0.43 +0.60 0.44 -0.10 0.50 -0.10

[0090] (6) Method for determining residual protein:

[0091] 1. Instruments and reagents

[0092] Inifinite M200 Pro microplate reader (Tecan, Germany), Millpore deionized water generator (Merck Millipore), Bradford protein assay kit (batch number: A192250415): contains protein standard solution (5 mg / ml BSA, serial number: P0006-2) and Coomassie Brilliant Blue G250 staining solution (serial number: P0006-1).

[0093] 2. Solution preparation

[0094] Kanamycin monosulfate test solution: Take about 250 mg of kanamycin monosulfate and dilute it with 5 ml of water to 50 mg / ml.

[0095] Kanamycin sulfate test solution: Take about 250 mg of kanamycin sulfate and dilute it with 5 ml of water to 50 mg / ml.

[0096] Kanamycin sulfate injection: Take two vials of kanamycin sulfate injection for direct determination.

[0097] Reference standard: Protein standard solution (5 mg / ml BSA).

[0098] 3. Measurement Method

[0099] - Bring the enzyme-labeled reagent, 96-well plate (strip), samples, and G250 staining solution to room temperature, mix gently, and avoid generating bubbles.

[0100] - Add 50 μl of protein standards of different concentrations to the protein standard wells of a 96-well plate.

[0101] - Add 50 μl of sample to the sample well of the 96-well plate.

[0102] Add 125 μl of G250 staining solution to each well.

[0103] - Immediately measure the absorbance at a detection wavelength of 595nm using an ELISA reader.

[0104] - Calculate the protein concentration in the sample based on the standard curve and the volume of the sample used.

[0105] (7) Methods for determining genotoxic impurities:

[0106] The Thermo Scientific Q Exactive Plus quadrupole Orbitrap™ high-resolution mass spectrometer (Thermo Scientific) was used, and the chromatographic column was GLSciences Inertsil. TMODS-3 (100×3.0mm, 5μm), mobile phase A was a methanol solution containing 0.1% formic acid, and mobile phase B was an aqueous solution containing 0.1% formic acid. Gradient elution was performed according to Table 4. Flow rate was 0.3 ml / min, column temperature was 35℃, injection volume was 5 μl, waste liquid was removed from 0 to 6 minutes, and mass spectrometry was performed from 6 to 40 minutes.

[0107] The mass spectrometer detector used an ESI source (+), with a spray voltage of 3.5 kV, ion transfer tube temperature of 350 °C, auxiliary gas temperature of 350 °C, auxiliary gas flow rate of 10 ml / min, and sheath gas flow rate of 40 ml / min. Scan mode: SIM; ion mode: positive ion; scan start / end time: 6–40 minutes. Precise target ion mass numbers m / z (+H): 117.0659 (NMOR), 103.0866 (NDEA), 115.0865 (NPIP), 75.0552 (NDMA), 117.1022 (NEIPA), 131.1179 (NDPA, NDIPA), 137.0709 (NMPA), 159.1492 (NDBA).

[0108] Table 4 Gradient elution procedure for genotoxic impurity assay

[0109] time min Mobile phase A% mobile phase B% 0 97 3 10 97 3 15 60 40 30 60 40 35 97 3 4 97 3

[0110] (8) Methods for determining elemental impurities:

[0111] A new ICP-MS (inductively coupled plasma mass spectrometry) method was established to determine 12 elemental impurities in raw materials and injection solutions: arsenic (As), cadmium (Cd), mercury (Hg), lead (Pb), cobalt (Co), lithium (Li), chromium (Cr), copper (Cu), antimony (Sb), barium (Ba), iron (Fe), and zinc (Zn). A PerkinElmer NexION 300X inductively coupled plasma mass spectrometer was used, with KED mode as the measurement setting. The gas flow rate in the reaction cell was 1.5 ml / min, the plasma gas flow rate was 18 ml / min, the ICP RF power was 1500 W, the pulse phase voltage was 1200 V, the KED mode reaction cell inlet voltage was -8 V, and the KED mode reaction cell outlet voltage was -26 V.

[0112] 2.3 Experimental Results

[0113] (1) Crystal form

[0114] The X-ray diffraction patterns of kanamycin monosulfate and kanamycin sulfate powders are inconsistent. Kanamycin monosulfate powder exhibits sharp X-ray diffraction angles, indicating it is a crystalline powder, while kanamycin sulfate powder shows no obvious diffraction angles, indicating it is an amorphous powder. Figure 6 As shown.

[0115] (2) Particle size and particle size distribution

[0116] The volume average particle size of kanamycin monosulfate ranged from 35.425 to 65.212 μm, and the particle size distribution diagram showed that some batches had uneven particle size distribution. The volume average particle size of kanamycin sulfate ranged from 37.955 to 33.026 μm, smaller than that of kanamycin monosulfate, and the particle size distribution diagram showed good batch-to-batch homogeneity. Figure 7 As shown in the literature, small volume-average particle size and uniform particle size distribution both contribute to increased dissolution rate.

[0117] (3) Solubility

[0118] Both ChP2000 and ChP2020 standards for kanamycin monosulfate and kanamycin sulfate describe the product as "easily soluble in water." However, the solubility of both raw materials in water was determined. The results showed that the water solubility of kanamycin monosulfate was significantly lower than that of kanamycin sulfate. The water solubility of kanamycin monosulfate converted to kanamycin was 0.13 mg / ml, lower than the specification of the sampled product: 0.25 g / ml (calculated as kanamycin). This low solubility limits its ability to be formulated into the currently available injectable solution, as shown in Table 5. Therefore, current domestic injectable solution production processes require the addition of sulfuric acid to convert kanamycin monosulfate to kanamycin sulfate, thereby increasing solubility.

[0119] Table 5. Solubility of different raw materials in water

[0120] raw material solubility Solubility (Kanamycin) Solubility Kanamycin monosulfate 0.16g / ml 0.13g / ml Easily soluble in water Kanamycin sulfate greater than 1g / ml greater than 0.68g / ml Very soluble in water

[0121] (4) Residual solvent

[0122] Both kanamycin monosulfate and kanamycin sulfate production processes utilize ethanol recrystallization. The ethanol content of the two raw materials was determined. Results showed that the ethanol content of kanamycin monosulfate was 0.10%–0.13%, while no ethanol was detected in kanamycin sulfate. The residual ethanol content in kanamycin monosulfate was higher than that in kanamycin sulfate. Figure 8 As shown.

[0123] (5) Related substances

[0124] Related substances of kanamycin sulfate were determined by HPLC-PAD method, and the results were compared with those of kanamycin monosulfate. The results showed that the contents of all individual impurities and total impurities in kanamycin monosulfate were higher than those in kanamycin sulfate. Figure 9 As shown.

[0125] (6) Residual protein

[0126] Kanamycin monosulfate is produced by fermentation, and residual protein is generated during the fermentation process. The residual protein content of the two raw materials was measured. The residual protein content of kanamycin monosulfate was 0.038%~0.055%, which was higher than that of kanamycin sulfate, which was 0.032%~0.036%.

[0127] (7) Genotoxic impurities

[0128] Kanamycin monosulfate is produced by fermentation, during which N-nitrosamine genotoxic impurities may be generated. Genotoxic impurities in the two raw materials were measured, and none of the nine N-nitrosamine genotoxic impurities were detected in either kanamycin monosulfate or kanamycin sulfate.

[0129] (8) Elemental impurities

[0130] The elemental impurity content of kanamycin monosulfate and kanamycin sulfate is basically the same, and is far below the limit, as shown in Table 6.

[0131] Table 6. Comparison of elemental impurity results between kanamycin monosulfate and kanamycin sulfate (μg / g)

[0132] element Kanamycin monosulfate Kanamycin sulfate limit Arsenic (As) Not detected ~ 0.0018 0.006～0.007 10.0 CadmiumCd 0.001～0.002 0.001 1.3 Mercury (Hg) Not detected ~ 0.009 Not detected 2.0 Lead (Pb) 0.007～0.083 0.010～0.013 3.3 Cobalt Co 0.0005～0.001 0.001～0.002 3.3 Lithium (Li) 0.002～0.015 0.01～0.02 166.7 Chromium (Cr) 0.08～0.16 0.17～0.22 733.3 Copper Cu 0.008～0.021 0.006～0.01 200.0 Antimony Sb Not detected ~ 0.004 0.005～0.008 60.0 Barium 0.009～0.02 0.13～0.16 466.7 Iron (Fe) 0.05～0.17 0.04～0.10 866.7 Zinc (Zn) 0.10～1.10 0.8～2.5 866.7

[0133] In summary, compared with kanamycin monosulfate, kanamycin sulfate has fewer related substances and residual proteins, lower ethanol residue, and better safety. At the same time, kanamycin sulfate has higher water solubility, more uniform particle size distribution, and faster dissolution rate. It does not require the addition of sulfuric acid to aid dissolution when preparing injection solutions, which simplifies the production process and reduces the safety risks that may be introduced by adding sulfuric acid.

[0134] It should be noted that the above are merely preferred embodiments of the present invention and are not intended to limit the present invention in any way. Any modifications or alterations made by those skilled in the art without departing from the scope of the present invention to create equivalent embodiments based on the disclosed technical content shall fall within the protection scope of the present invention.

Claims

1. A kanamycin injection solution without antioxidants, characterized in that, Each 1 mL of injection solution consists of: 250 mg kanamycin sulfate (calculated as kanamycin), water for injection to 1 mL, and a pH adjuster to make the pH value 4.5-7.

0. It does not contain sodium bisulfite, cysteine ​​hydrochloride, or other antioxidants.

2. The antioxidant-free kanamycin injection solution according to claim 1, characterized in that, The headspace oxygen content of the injection solution after filling is ≤1.0%, which is achieved by nitrogen replacement.

3. The antioxidant-free kanamycin injection solution according to claim 1, characterized in that, The pH adjuster is sulfuric acid.

4. The antioxidant-free kanamycin injection according to claim 1, characterized in that, The pH value is 5.5-6.

5.

5. The method for preparing the antioxidant-free kanamycin injection solution according to claim 1, characterized in that, Includes the following steps: S1. In a Class 100 clean area, dissolve 700-750g of kanamycin sulfate in water for injection at 25-30℃ and stir under nitrogen protection until completely dissolved. S2, using 0.1 mol·L -1 Adjust the pH to 4.5–7.0 with sulfuric acid; S3. Add water for injection to 2000 ml, and circulate for 5 min under nitrogen protection; S4. After being filtered through a 0.22 µm polyethersulfone filter, the ampoule is filled into a brown neutral borosilicate glass ampoule and sealed with nitrogen. S5. Sterilize with flowing steam at 100℃ for 30 minutes, then perform leak detection, light inspection, and packaging to obtain the product.

6. The method for preparing the antioxidant-free kanamycin injection according to claim 5, characterized in that, In S4, the nitrogen flow rate is 1–2 L·min -1 Nitrogen filling time: 2–5 s.

7. The quality control method for the antioxidant-free kanamycin injection solution according to any one of claims 1-6, characterized in that, Includes the following steps: S1. High temperature test: Place at 60℃ for 30 days, and take samples at 0, 5, 10 and 3 days to determine the content, related substances, pH and color; S2. Light test: Place at 4500±500 lx for 10 days, and take samples on days 0, 5, and 10 for testing as above; S3. Long-term test: Store at 25℃ ± 2℃ / 60% RH ± 5% RH for 12 months, and take samples at 0, 3, 6, 9 and 12 months for testing as above; S4. The sodium bisulfite was quantified by ion chromatography, and the detection limit was ≤0.005%. The detection result was lower than the detection limit, which confirmed that no sodium bisulfite antioxidant was added to the sample.

Images