Method for detecting impurities in compound amino acid injection

Abstract

The invention provides a method for detecting impurities in a compound amino acid injection, and belongs to the field of medicine quality determination. According to the method, impurities in the compound amino acid injection are analyzed and detected by adopting a high performance liquid chromatography carried with a CAD detector, a special amino acid chromatographic column is selected as a chromatographic column, heptafluorobutyric acid is used as a mobile phase A, acetonitrile is used as a mobile phase B, and other impurities such as 4-guanidinobutyric acid in the compound amino acid injection are detected and quantified. According to the detection method provided by the invention, the 4-guanidinobutyric acid can be effectively separated from other components in the compound amino acid injection, the detection precision and accuracy are relatively high, the reproducibility is relatively good, the operation is simple, and the method is environment-friendly; meanwhile, the detection method is high in sensitivity, impurities with small content can be analyzed and detected, the loss of a chromatographic column is small, and the detection cost is reduced; in addition, the detection method can be used for analyzing and detecting various amino acids, impurities introduced by the amino acids, impurities generated by degradation of the amino acids and the like.

Description

Technical Field

[0001] The invention relates to a method for detecting impurities in a compound amino acid injection, and belongs to the field of drug quality detection and analysis. Background Art

[0002] Compound amino acid injection is a compound preparation containing multiple amino acids. It is now widely used in the pharmaceutical industry. Strict quality control is required for compound amino acid injection to ensure its safety and effectiveness. Therefore, it is of great significance to analyze and detect various substances in compound amino acid injection. Compound amino acid injection may produce a variety of amino acid impurities during production and storage, such as arginine oxidation degradation to produce 4-guanidinobutyric acid. However, the composition of amino acids and impurities in compound amino acid injection is complex, and most of them are terminal absorption, with poor ultraviolet response, which increases the difficulty of analyzing and detecting impurities.

[0003] There are few existing detection methods for 4-guanidinobutyric acid, and it is easy to cause the 4-guanidinobutyric acid peak to be unable to be effectively separated from other component peaks due to the selection of mobile phase and changes in elution procedures. At the same time, the ultraviolet detector used in the existing detection methods is not sensitive enough. When the content of 4-guanidinobutyric acid is low, it is impossible to effectively detect and determine the content; in addition, the existing detection methods usually use ordinary chromatographic columns, which cause great loss of chromatographic columns when detecting amino acid preparations. For example: In the article "Determination of 4-guanidinobutyric acid in Clinimix E (Clinimix E)" published by Wu Huibin, the HILIC chromatographic column was used to analyze and detect the content of 4-guanidinobutyric acid in the compound amino acid injection. This determination method has many problems such as being easily interfered by other substances, low detection sensitivity, and large loss of chromatographic columns.

[0004] In view of this, it is indeed necessary to propose improvements to the detection method of impurities in compound amino acid injection to solve the above problems. Summary of the invention

[0005] The object of the present invention is to provide a method for detecting impurities in a compound amino acid injection, so as to solve at least one of the problems of the existing detection method in analyzing and detecting the compound amino acid injection, namely, low sensitivity, large loss of chromatographic column and complicated operation steps.

[0006] To achieve the above object, the present invention provides a method for detecting impurities in a compound amino acid injection, comprising: using a high performance liquid chromatography method equipped with a CAD detector to analyze and detect impurities in the compound amino acid injection, specifically:

[0007] Chromatographic column: amino acid special chromatographic column;

[0008] Mobile phase A: 0.25-0.35% heptafluorobutyric acid solution;

[0009] Mobile phase B: acetonitrile;

[0010] The gradient elution program was:

[0011] Time (min) Mobile phase A (%) Mobile phase B (%) 0 95～100 5～0 10 95～100 5～0 25 80～90 20～10 35 80～90 20～10 55 40～60 60～40 60 40～60 60～40 61 95～100 5～0 80 95～100 5～0 .

[0012] Further, the gradient elution procedure is preferably:

[0013] Time (min) Mobile phase A (%) Mobile phase B (%) 0 95～98 5～2 10 95～98 5～2 25 80～85 20～15 35 80～85 20～15 55 40～50 60～50 60 40～50 60～50 61 95～98 5～2 80 95～98 5～2 .

[0014] The gradient elution program is more preferably:

[0015] Time (min) Mobile phase A (%) Mobile phase B (%) 0 95 5 10 95 5 25 84 16 35 84 16 55 40 60 60 40 60 61 95 5 80 95 5 .

[0016] Further, the mobile phase A is a heptafluorobutyric acid solution with a concentration of 0.27-0.33%. Preferably, the mobile phase A is a heptafluorobutyric acid solution with a concentration of 0.29-0.31%. More preferably, the mobile phase A is a heptafluorobutyric acid solution with a concentration of 0.30%.

[0017] Furthermore, the flow rate of the mobile phase is 0.5 to 1.2 ml / min, preferably, the flow rate of the mobile phase is 0.7 to 0.9 ml / min, and more preferably, the flow rate of the mobile phase is 0.8 ml / min.

[0018] Further, the injection volume is 5 to 50 μL, preferably, the injection volume is 5 to 20 μL, and more preferably, the injection volume is 10 μL.

[0019] Furthermore, the amino acid-specific chromatographic column is a chromatographic column equivalent to WelchAmino Acid Plus. Preferably, the model specification of the amino acid-specific chromatographic column is WelchAmino Acid Plus, 4.6×300 mm.

[0020] Furthermore, the column temperature of the amino acid-specific chromatographic column is 25 to 35°C, preferably 28 to 32°C, and more preferably 30°C.

[0021] Furthermore, the atomization temperature of the CAD detector is 30-50°C, preferably 45°C.

[0022] Furthermore, the filtering time of the CAD detector is 5 to 15 seconds, preferably 10 seconds.

[0023] Furthermore, the CAD detector acquisition frequency is 5 to 25 Hz, preferably 20 Hz.

[0024] As a further improvement of the present invention, the method for detecting impurities in the compound amino acid injection comprises: using high performance liquid chromatography equipped with a CAD detector to analyze and detect impurities in the compound amino acid injection, specifically:

[0025] Chromatographic column: amino acid special chromatographic column, model specification is Welch Amino Acid Plus, 4.6×300mm, column temperature is 30℃;

[0026] Mobile phase A: 0.3% heptafluorobutyric acid solution;

[0027] Mobile phase B: acetonitrile;

[0028] The mobile phase flow rate was 0.8 ml / min, and the injection volume was 10 μL;

[0029] The nebulization temperature of the CAD detector was 45°C, the filtering time was 10 s, and the acquisition frequency was 20 Hz; the gradient elution program was:

[0030] Time (min) Mobile phase A (%) Mobile phase B (%) 0 95 5 10 95 5 25 84 16 35 84 16 55 40 60 60 40 60 61 95 5 80 95 5 .

[0031] Furthermore, the impurity is 4-guanidinobutyric acid.

[0032] The present invention adopts the above detection method, which not only solves the above problems well, but also makes the 4-guanidinobutyric acid peak effectively separated from other components, and has high sensitivity, and can effectively detect and measure the content of impurities with low content; at the same time, it can reduce the loss of amino acid preparations to the chromatographic column; in the analysis and detection of the reference substance and the test sample, the retention time of 4-guanidinobutyric acid in the reference substance and the test sample solution is consistent, indicating that the detection precision and accuracy of the method are high, and the repeatability is good. In addition, the detection method of the present invention can also effectively detect and separate amino acids and / or other impurities in the compound amino acid injection, thereby improving the practicality of the detection method.

[0033] The beneficial effects of the present invention are as follows: the detection method provided by the present invention can effectively separate 4-guanidinobutyric acid from other components in the compound amino acid injection, and has high detection precision and accuracy, good reproducibility, simple operation and environmental friendliness; at the same time, the detection method has high sensitivity, can analyze and detect impurities with a small content, and has low loss to the chromatographic column, thereby reducing the detection cost; in addition, the detection method can analyze and detect multiple amino acids in the compound amino acid injection, impurities introduced by amino acids, and impurities generated by amino acid degradation, and has high practicality. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 This is the complete chromatogram of the blank solution in Example 1.

[0035] Figure 2 It is the complete chromatogram of the test solution in Example 1.

[0036] Figure 3 It is a partial chromatogram of the test solution in Example 2.

[0037] Figure 4 This is the complete chromatogram of the reference solution in Example 3.

[0038] Figure 5 It is a partial chromatogram of the mixed solution in Example 3.

[0039] Figure 6 It is a partial chromatogram of the mixed solution in Example 4.

[0040] Figure 7 It is a partial chromatogram of the mixed solution in Example 5.

[0041] Figure 8 It is a partial chromatogram of the mixed solution in Example 6.

[0042] Fig. 9 It is a partial chromatogram of the mixed solution in Example 7.

[0043] Fig.10 It is a partial chromatogram of the mixed solution in Example 8.

[0044] Fig.11 This is a partial chromatogram of the specificity test in Experimental Example 1.

[0045] Fig.12 is the linearity graph of the linear solution in Experimental Example 3. DETAILED DESCRIPTION

[0046] In order to make the purpose, technical scheme and advantages of the present invention clearer, the present invention is described in detail below in conjunction with the embodiments. It is necessary to point out that the following embodiments are only used to explain and illustrate the present invention and are not used to limit the present invention. Some non-essential improvements and adjustments made by those skilled in the art based on the above invention content still belong to the protection scope of the present invention.

[0047] In the present invention, the configuration methods of various solutions are as follows:

[0048] Blank solution: Watsons water.

[0049] Negative blank solution: Prepare according to the concentration of each raw material and excipient in the following prescription (without adding arginine).

[0050] The prescription of 500ml compound amino acid injection is: isoleucine: 4.600g; leucine: 4.725g; lysine acetate: 1.975g; methionine: 0.220g; phenylalanine: 0.150g; threonine: 1.070g; tryptophan: 0.350g; valine: 4.450g; alanine: 4.200g; arginine: 7.685g; aspartic acid: 0.100g; histidine: 1.550g; proline: 2.650g; serine: 1.300g; tyrosine: 0.200g; glycine: 2.700g; cysteine: 0.125g; sodium bisulfite: 0.125g; glacial acetic acid: appropriate amount.

[0051] Test solution: Take the compound amino acid injection solution, the type of compound amino acid injection solution is 17AA-III.

[0052] Reference solution: Accurately weigh 4.6 mg of 4-guanidinobutyric acid reference substance, place it in a 20 ml volumetric flask, dissolve and dilute to the scale with water, accurately transfer 1 ml to a 10 ml volumetric flask, add appropriate amount of water to dissolve and dilute to the scale, shake well.

[0053] 4-Guandinobutyric acid stock solution (230 μg / ml): Accurately weigh about 4.6 mg of 4-guanidinobutyric acid, place it in a 20 ml volumetric flask, dissolve it in Watson's water and dilute it to the scale, and shake well.

[0054] Mixed solution: Accurately pipette 1 ml of 4-guanidinobutyric acid stock solution into a 10 ml volumetric flask, dilute to the mark with the test solution, and shake well.

[0055] Example 1

[0056] Chromatographic conditions: amino acid special chromatographic column, model specification: UltimateAminoAcidPlus4.6×300mm. A 0.3% heptafluorobutyric acid solution was used as mobile phase A, and acetonitrile was used as mobile phase B. Heptafluorobutyric acid included two specifications, one with a purity of 98% and the other with a purity of 99.5%. The flow rate was 0.8ml / min, the column temperature was 30°C, the injection volume was 20μL, the nebulization temperature of the CAD detector was 45°C, the filtering time was 10s, and the acquisition frequency was 20Hz. The gradient elution program was as follows:

[0057] Time (min) Mobile phase A (%) Mobile phase B (%) 0 95 5 10 95 5 25 84 16 35 84 16 55 40 60 60 40 60 61 95 5 80 95 5

[0058] Two specifications of heptafluorobutyric acid were used to perform gradient elution on the blank solution and the test solution, and the chromatograms were obtained, as shown in the figure. Figure 1 and Figure 2 As shown, it can be seen that heptafluorobutyric acid has high purity, the baseline noise in the chromatogram is small, and the sensitivity of each component is also higher.

[0059] Example 2

[0060] Chromatographic conditions: amino acid special chromatographic column, model specification: UltimateAminoAcid Plus4.6×300mm. 0.3% heptafluorobutyric acid solution as mobile phase A, acetonitrile as mobile phase B. Flow rate 0.8ml / min, column temperature 30℃, injection volume 50μL, CAD detector nebulization temperature 45℃, filter time 10s, acquisition frequency 20Hz. Gradient elution program is as follows:

[0061] Time (min) Mobile phase A (%) Mobile phase B (%) 0 95 5 10 95 5 25 84 16 35 84 16 55 40 60 60 40 60 61 95 5 80 95 5

[0062] The test solution was diluted 5 times to 50 μL and injected, and then gradient eluted. The chromatogram was as follows: Figure 3 As shown, it can be seen that 4-guanidinobutyric acid and adjacent components have achieved baseline separation.

[0063] Example 3

[0064] Chromatographic conditions: amino acid special chromatographic column, model specification: WelchAmino Acid Plus4.6×300mm. 0.3% heptafluorobutyric acid solution as mobile phase A, acetonitrile as mobile phase B. Flow rate is 0.8ml / min, column temperature is 30℃, injection volume is 10μL, nebulization temperature of CAD detector is 45℃, filtering time is 10s, acquisition frequency is 20Hz. Gradient elution program is as follows:

[0065] Time (min) Mobile phase A (%) Mobile phase B (%) 0 95 5 10 95 5 25 84 16 35 84 16 55 40 60 60 40 60 61 95 5 80 95 5

[0066] The reference solution and the mixed solution were sampled separately, and the contents of 4-guanidinobutyric acid in the reference solution and the mixed solution were detected by peak area according to the external standard method. The detection results of the reference solution are shown in Figure 4 The test results of the mixed solution are shown in Figure 5 .

[0067] Among them, the retention time RT of 4-guanidine butyric acid in the reference solution is 32.125, the retention time RT of 4-guanidine butyric acid in the mixed solution is 31.888, and the separation degree is 1.70.

[0068] Example 4

[0069] The only difference between this embodiment and embodiment 3 is that the concentration of heptafluorobutyric acid in mobile phase A is 0.29%, and only the mixed solution is injected.

[0070] The test results of the mixed solution are shown in Figure 6Among them, the retention time RT of 4-guanidinobutyric acid in the mixed solution is 31.885, and the separation degree is 1.71.

[0071] Example 5

[0072] The only difference between this embodiment and embodiment 3 is that the concentration of heptafluorobutyric acid in mobile phase A is 0.31%, and only the mixed solution is injected.

[0073] The test results of the mixed solution are shown in Figure 7 Among them, the retention time RT of 4-guanidinobutyric acid in the mixed solution is 31.912, and the separation degree is 1.72.

[0074] Example 6

[0075] The only difference between this embodiment and embodiment 3 is that the column temperature is 28° C. and only the mixed solution is injected.

[0076] The test results of the mixed solution are shown in Figure 8 Among them, the retention time RT of 4-guanidinobutyric acid in the mixed solution is 32.107, and the separation degree is 1.57.

[0077] Example 7

[0078] The only difference between this embodiment and embodiment 3 is that the column temperature is 32° C. and only the mixed solution is injected.

[0079] The test results of the mixed solution are shown in Fig. 9 Among them, the retention time RT of 4-guanidinobutyric acid in the mixed solution is 31.049, and the separation degree is 2.00.

[0080] Example 8

[0081] The only difference between this embodiment and embodiment 3 is that in this embodiment, only the mixed solution is sampled to detect all amino acids and impurities in the mixed solution.

[0082] The test results of the mixed solution are shown in Fig.10 It can be seen that the detection method of the present invention can detect amino acids and impurities in a mixed solution with a high degree of separation. The retention time RT and degree of separation of each amino acid and impurity in the mixed solution are shown in the following table:

[0083] Serial number name Retention time min Peak area pA*min Separation 1 Sodium bisulfite 8.280 0.2664 3.48 2 Glycine 8.855 0.4103 2.20 3 Serine 9.294 0.3981 6.40 4 Aspartic acid 10.708 0.2610 13.76 5 Threonine 14.522 0.3161 1.24 6 Alanine 14.914 0.3670 7.83 7 Cysteine 17.877 0.2983 28.39 8 Proline 27.762 0.7470 14.07 9 Ornithine 32.160 1.1319 9.68 10 4-Guandinobutyric acid 35.470 1.2254 6.36 11 Valine 37.604 0.8490 4.35 12 Lysine 39.284 0.7582 1.25 13 Histidine 39.768 0.8622 5.82 14 Methionine 41.965 0.7412 8.90 15 Tyrosine 44.807 1.0765 5.39 16 Arginine 46.155 1.7756 7.14 17 Isoleucine 47.783 1.7573 5.29 18 Leucine 48.795 1.6302 10.29 19 Phenylalanine 50.557 1.5434 12.42 20 Tryptophan 52.503 1.1721 na

[0084] Test Example 1

[0085] Specificity test

[0086] The difference between this embodiment and embodiment 3 is that the blank solution, negative blank solution, reference solution, test solution and mixed solution are injected separately.

[0087] Test results are shown in Fig.11 Among them, in the chromatograms of the blank solution and the negative blank solution, there is no interference peak at the 4-guanidinobutyric acid peak position; the 4-guanidinobutyric acid peak positions in the reference solution and the mixed solution are close, and in the mixed solution, the separation degree between the 4-guanidinobutyric acid peak and the adjacent component peak is 1.7, which meets the standard requirement of a separation degree greater than 1.5. This shows that the detection method has good specificity.

[0088] Test Example 2

[0089] Limit of Quantitation and Limit of Detection Tests

[0090] Quantitation limit solution: Take an appropriate amount of reference solution and continue to dilute until the S / N (signal-to-noise ratio) is between 10 and 30.

[0091] Detection limit solution: Take an appropriate amount of reference solution and continue to dilute until the S / N (signal-to-noise ratio) is between 3 and 10.

[0092] Six portions of the quantitative limit solution and one portion of the detection limit solution were sampled and tested. The chromatographic conditions were the same as those in Example 3, and the 4-guanidinobutyric acid peak area and solution concentration were recorded. The test results are shown in the following table:

[0093]

[0094] Among them, the percentage of quantitative limit concentration to limit concentration is 10.5%, which meets the standard of less than 30%; the S / N of the six quantitative limit solutions is between 11.8 and 15.4, which meets the standard of S / N in the range of 10-30; the RSD of the peak area of ​​the quantitative limit solution is 8.2%, which meets the standard of no more than 10%; at the same time, it can be concluded that the quantitative limit concentration is 2.4176μg / ml and the detection limit concentration is 1.2088μg / ml. This shows that the sensitivity of the detection method is good.

[0095] Test Example 3

[0096] Linearity test

[0097] Linear solution-1 (limit of quantitation): Accurately measure 1.0 ml of the 4-guanidinobutyric acid stock solution and dilute it with water to make a solution containing 2.4176 μg of 4-guanidinobutyric acid per 1 ml.

[0098] Linear solution-2 (50%): Accurately measure 1.0 ml of 4-guanidinobutyric acid stock solution, place it in a 20 ml volumetric flask, dilute to the mark with Watson's water, and shake well.

[0099] Linear solution-3 (80%): Accurately measure 2.0 ml of 4-guanidinobutyric acid stock solution, place it in a 25 ml volumetric flask, dilute to the mark with Watson's water, and shake well.

[0100] Linear solution-4 (100%): Accurately measure 1.0 ml of 4-guanidinobutyric acid stock solution, place it in a 10 ml volumetric flask, dilute to the mark with Watson's water, and shake well.

[0101] Linear solution-5 (150%): Accurately measure 3.0 ml of 4-guanidinobutyric acid stock solution, place it in a 20 ml volumetric flask, dilute to the mark with Watson's water, and shake well.

[0102] Linear solution-6 (200%): Accurately measure 2.0 ml of 4-guanidinobutyric acid stock solution, place it in a 10 ml volumetric flask, dilute to the mark with Watson's water, and shake well.

[0103] According to the detection conditions in Example 3, the above linear solution was injected and analyzed, and the obtained peak areas are shown in the following table:

[0104] name Concentration (μg / ml) Peak area Linear-1 2.4176 0.0980 Linear-2 12.0879 0.7567 Linear-3 19.3406 1.2173 Linear-4 24.1758 1.5094 Linear-5 36.2637 2.2617 Linear-6 48.3516 2.9047

[0105] Linear graph Fig.12 As shown, R 2 That is r 2 In the concentration range of the quantitative limit to 200% limit, the peak area of ​​4-guanidinobutyric acid has a good linear relationship, the linear equation is y=0.0611x+0.0044, and the correlation coefficient r is 0.9991.

[0106] Test Example 4

[0107] Accuracy test

[0108] 4-Guandinobutyric acid recovery rate stock solution (230μg / ml): Accurately weigh 4.6mg of 4-guanidine butyric acid standard, place it in a 20ml volumetric flask, dissolve and dilute to the scale with the test solution, and shake well.

[0109] Low concentration solution (50%): Take 1 ml of 4-guanidinobutyric acid recovery rate stock solution, place it in a 20 ml volumetric flask, dilute to the mark with the test solution, and shake well. Prepare 3 portions in the same way.

[0110] Medium concentration solution (100%): Measure 1 ml of 4-guanidinobutyric acid recovery rate stock solution, place it in a 10 ml volumetric flask, dilute to the mark with the test solution, and shake well. Prepare 3 portions in the same way.

[0111] High concentration solution (150%): Take 3 ml of 4-guanidinobutyric acid recovery rate stock solution, place it in a 20 ml volumetric flask, dilute to the mark with the test solution, and shake well. Prepare 3 portions in the same way.

[0112] According to the analysis conditions in Example 3, the above solutions were sampled and analyzed respectively, and the recovery rate of each solution was calculated by the external standard method. The statistical results are shown in the following table:

[0113]

[0114] As can be seen from the table, the recovery rates of 4-guanidinobutyric acid in samples of three concentrations are between 92.4% and 105.3%, the average recovery rate is between 93.0% and 105.1%, and the maximum RSD is 1.4%; the average recovery rate of nine samples is 99.0%, and the RSD is 5.4%, which are in line with the standards of a recovery rate between 90.0% and 110.0% and an RSD of less than 10.0%, indicating that the recovery rate of the detection method is good and the accuracy meets the requirements.

[0115] In summary, the detection method provided by the present invention can effectively separate 4-guanidinobutyric acid from other components in the compound amino acid injection, and has high detection precision and accuracy, good reproducibility, simple operation and environmental friendliness; at the same time, the detection method has high sensitivity, can analyze and detect impurities with a small content, and has low loss to the chromatographic column, thereby reducing the detection cost; in addition, the detection method can analyze and detect multiple amino acids in the compound amino acid injection, impurities introduced by amino acids, and impurities generated by amino acid degradation, etc., and has high practicality.

[0116] The above embodiments are only used to illustrate the technical solution of the present invention rather than to limit it. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solution of the present invention may be modified or replaced by equivalents without departing from the spirit and scope of the technical solution of the present invention.

Claims

1. A method for detecting impurities in a compound amino acid injection, characterized in that: include: The impurities in the compound amino acid injection were analyzed and detected by high performance liquid chromatography equipped with a CAD detector, specifically: Chromatographic column: amino acid special chromatographic column; Mobile phase A: 0.25-0.35% heptafluorobutyric acid solution; Mobile phase B: acetonitrile; The gradient elution program was: 。 2. The method for detecting impurities in the compound amino acid injection according to claim 1, characterized in that: The gradient elution procedure is: 。 3. The method for detecting impurities in the compound amino acid injection according to claim 1, characterized in that: The mobile phase A is heptafluorobutyric acid with a concentration of 0.29-0.31%.

4. The method for detecting impurities in the compound amino acid injection according to claim 1, characterized in that: The amino acid-specific chromatographic column is Welch Amino Acid Plus, with a specification of 4.6×300 mm.

5. The method for detecting impurities in the compound amino acid injection according to claim 1, characterized in that: The column temperature of the amino acid-specific chromatographic column is 25-35°C.

6. The method for detecting impurities in the compound amino acid injection according to claim 1, characterized in that: The flow rate of the mobile phase is 0.5-1.2 ml / min.

7. The method for detecting impurities in the compound amino acid injection according to claim 1, characterized in that: The injection volume is 5 to 50 μL.

8. The method for detecting impurities in the compound amino acid injection according to claim 1, characterized in that: The atomization temperature of the CAD detector is 30-50°C; And / or, the CAD detector filtering time is 5 to 15 seconds; And / or, the CAD detector has an acquisition frequency of 5 to 25 Hz.

9. The method for detecting impurities in the compound amino acid injection according to claim 1, characterized in that: The detection method is: Chromatographic column: amino acid special chromatographic column, model specification is Welch Amino Acid Plus, 4.6×300mm, column temperature is 30℃; Mobile phase A: 0.3% heptafluorobutyric acid solution; Mobile phase B: acetonitrile; The mobile phase flow rate was 0.8 ml / min, and the injection volume was 10 μL; The CAD detector had a nebulization temperature of 45 °C, a filter time of 10 s, and an acquisition frequency of 20 Hz; The gradient elution program was:

10. The method for detecting impurities in the compound amino acid injection according to claim 1, characterized in that: The impurity is 4-guanidinobutyric acid.

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