Method for determining lysine hydrochloride content by high performance liquid chromatography

Abstract

The present application relates to the technical field of amino acid detection, and particularly relates to a method for determining the content of lysine hydrochloride by high performance liquid chromatography. The method comprises the following steps: taking amino acid-silica gel as a filler, and using a mobile phase composed of 0.04-0.06 mol / L potassium dihydrogen phosphate solution and acetonitrile solution; the volume ratio of acetonitrile and water in the acetonitrile solution is 9-11:1; the volume ratio of the potassium dihydrogen phosphate solution and the acetonitrile solution is 25-40:60-80. The method is suitable for determining the content of lysine hydrochloride in calcium zinc gluconate oral solution, can solve the problem of interference of auxiliary materials, and has the advantages of simple sample pretreatment method, high detection sensitivity, high separation efficiency, strong method reproducibility, low cost and the like.

Description

Technical Field

[0001] This invention relates to the field of amino acid detection technology, and in particular to a method for determining the content of lysine hydrochloride by high performance liquid chromatography. Background Technology

[0002] Currently reported methods for determining lysine hydrochloride mainly include the Kjeldahl method, high-performance liquid chromatography (HPLC), and ion chromatography. The Kjeldahl method is too time-consuming, requiring at least two hours to complete; ion chromatography has limited applicability in production and is easily affected by ion interference.

[0003] High-performance liquid chromatography (HPLC) offers advantages such as short detection time and high efficiency. However, for the detection of lysine hydrochloride in calcium gluconate-zinc oral solution, the solution contains calcium gluconate, zinc gluconate, lysine hydrochloride, sodium benzoate, aspartame, acesulfame potassium, lactic acid, and flavorings. Lysine hydrochloride has a maximum absorption wavelength of 200–210 nm (terminal absorption), and the presence of peaks from lysine hydrochloride, sodium benzoate, aspartame, acesulfame potassium, lactic acid, and flavorings can easily interfere with the detection results.

[0004] Chinese patent (publication number CN109596747A) uses high performance liquid chromatography to determine the lysine hydrochloride content in calcium zinc gluconate oral solution. It uses a C18 column and sodium octanesulfonate (ion-pairing reagent) for determination, which results in high column wear and poor reproducibility.

[0005] Chinese patent (publication number CN111443149) describes a high-performance liquid chromatography (HPLC) method for determining the lysine hydrochloride content in a sodium hyaluronate injection composite solution. A C18 column is used, and the sample requires derivatization with a derivatizing reagent. The mobile phase consists of a weakly alkaline solution (phosphate borate solution): methanol at a volume ratio of 90:10-98:2; the weakly alkaline solution has a pH of 8.0-8.8. This method utilizes derivatization techniques to convert substances that are difficult to analyze into those with similar chemical structures but are easier to analyze, facilitating quantification and separation. However, the steps involving derivatization are complex and can easily lead to poor accuracy in the detection results.

[0006] The Chinese literature, "Simultaneous Determination of Lysine Hydrochloride and Sodium Benzoate Content in Calcium Gluconate and Zinc Gluconate Oral Solution by HPLC," discloses a method using a C18 column with a mobile phase of potassium hydrogen phosphate buffer-acetonitrile (90:10). The 0.01 mol / L potassium hydrogen phosphate buffer solution consists of 2.2822 g of potassium hydrogen phosphate, 1000 mL of water, pH adjusted to 4.8 with phosphoric acid, and then 1.08 g of sodium octane sulfonate. Reproducing this method revealed that the blank solvent interfered with the detection of lysine hydrochloride content, and the method also used ion-pairing reagents, which caused significant damage to the column.

[0007] In view of this, the present invention is hereby proposed. Summary of the Invention

[0008] To address the aforementioned technical problems, this invention provides a method for determining the content of lysine hydrochloride using high-performance liquid chromatography (HPLC). This method has advantages such as simple sample pretreatment, high separation efficiency, good selectivity, and high detection sensitivity, and can accurately quantify lysine hydrochloride.

[0009] Specifically, the technical solution of the present invention is as follows:

[0010] In a first aspect, the present invention provides a method for determining the content of lysine hydrochloride by high performance liquid chromatography, wherein the method uses amino-bonded silica gel as a packing material, and the mobile phase consists of a 0.04-0.06 mol / L potassium dihydrogen phosphate solution and an acetonitrile solution; wherein the volume ratio of acetonitrile to water in the acetonitrile solution is 9.0-11.0:1; and the volume ratio of potassium dihydrogen phosphate solution to acetonitrile solution is 25-40:60-80.

[0011] The method provided by this invention utilizes high-performance liquid chromatography (HPLC) to determine the content of lysine hydrochloride. It employs amino-bonded silica gel as the packing material and uses potassium dihydrogen phosphate solution and acetonitrile solution of specific concentrations and ratios as the mobile phase. This method effectively solves the interference problem of components such as calcium gluconate, zinc gluconate, lysine hydrochloride, sodium benzoate, aspartame, acesulfame potassium, lactic acid, and fragrance in the sample on the determination results of lysine hydrochloride content, ensuring accurate testing.

[0012] The method provided by this invention first prepares acetonitrile into an acetonitrile solution, and then mixes it with potassium dihydrogen phosphate solution to obtain a mobile phase. Compared with the mobile phase obtained by directly mixing acetonitrile with potassium dihydrogen phosphate solution, the mobile phase prepared by this method has better anti-interference ability and more accurate detection results.

[0013] In this invention, the concentration of the potassium dihydrogen phosphate solution is preferably 0.045-0.055, more preferably 0.05 mol / L. In the acetonitrile solution, the volume ratio of acetonitrile to water is preferably 9.5-10.5:1, more preferably 10:1. The volume ratio of the potassium dihydrogen phosphate solution to the acetonitrile solution is preferably 30:65-75, more preferably 30:70. The measurement results obtained by the above preferred embodiments are more accurate. This invention does not require further adjustment of the pH of the mobile phase; in the specific embodiments provided by this invention, the pH value of the mobile phase is 7.0 ± 0.5.

[0014] In a preferred embodiment of the present invention, the chromatographic column is packed with amino-bonded silica gel, with dimensions of 4.6 mm × 250 mm and 5 m; preferably, an amino column is used, and more preferably, a Kromasil amino column of 4.6 mm × 250 mm and 5 m is used. The chromatographic mode is reverse separation mode.

[0015] In the method described in this invention, the preferred flow rate of the mobile phase during the determination process is 0.9-1.1 ml per minute, more preferably 1.0 ml per minute. The preferred injection volume is 15-25 μl, more preferably 20 μl. The preferred column temperature is 25-35℃, more preferably 30℃. The preferred detection wavelength is 200 nm-206 nm, more preferably 203 nm. The preferred chromatographic run time is 30-50 min, more preferably 35 min. This invention uses high-performance liquid chromatography (HPLC), and under the above-mentioned preferred determination conditions and parameters, it achieves high separation efficiency, better selectivity, higher detection sensitivity, and more accurate determination results. Furthermore, the method described in this invention can be performed via direct injection, which is simple to operate, time-saving, highly reproducible, and can be automated.

[0016] Furthermore, in this invention, the diluent is composed of the potassium dihydrogen phosphate solution and the acetonitrile solution; the volume ratio of the potassium dihydrogen phosphate solution to the acetonitrile solution in the diluent is preferably 50-70:40, more preferably 60:40. Using the diluent of this invention, the sample to be tested can be diluted and prepared in a one-step method, which has the advantages of simple sample pretreatment (using a one-step dilution method), low solvent consumption, and high detection efficiency.

[0017] This invention employs high-performance liquid chromatography (UV) to determine the content of lysine hydrochloride, particularly effective in determining the lysine hydrochloride content in calcium gluconate zinc oral solutions. The method is simple, easy to operate, and highly reproducible. The chromatographic column used is a common amino column, and the mobile phase consists of common buffer salts and organic phases, offering broad applicability and advantages in terms of cost and widespread adoption.

[0018] Secondly, the present invention provides a method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution, wherein the method described above is used to determine the lysine hydrochloride content in the calcium gluconate zinc oral solution.

[0019] The present invention does not specifically limit the source of the calcium gluconate zinc oral solution. Any calcium gluconate zinc oral solution obtained from conventional commercial channels in the art can be used to determine lysine hydrochloride using the method described in the present invention.

[0020] In a more specific embodiment provided by the present invention, in addition to calcium gluconate, zinc gluconate, and lysine hydrochloride, the calcium gluconate zinc oral solution also contains at least one of sodium benzoate, aspartame, acesulfame potassium, lactic acid, and flavoring.

[0021] In this invention, the calcium gluconate zinc oral solution needs to be diluted with the diluent before loading. The concentration of the diluted sample is preferably 0.8 mg / ml-1.2 mg / ml, more preferably 1.0 mg / ml.

[0022] In a more specific embodiment provided by the present invention, the method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution is as follows:

[0023] The raw materials tested were: calcium gluconate zinc oral solution, with a lysine hydrochloride content of 0.1g / 10ml. The total content of sodium benzoate, aspartame, acesulfame potassium, and flavoring was all lower than that of lysine hydrochloride, while the lactic acid content was higher than that of lysine hydrochloride.

[0024] Liquid chromatography (UV) conditions:

[0025] Chromatographic column: Kromasil amino column (4.6 mm x 250 mm, 5 m);

[0026] Mobile phase: potassium dihydrogen phosphate solution-acetonitrile solution, with a volume ratio of potassium dihydrogen phosphate solution to acetonitrile solution of 30:70. The concentration of potassium dihydrogen phosphate solution is 0.05 mol / L; the volume ratio of acetonitrile to water in the acetonitrile solution is 10:1.

[0027] Diluent: 0.05 mol / L potassium dihydrogen phosphate solution - acetonitrile solution, with a volume ratio of potassium dihydrogen phosphate solution to acetonitrile solution of 60:40;

[0028] Detection wavelength: 203nm;

[0029] Column temperature: 30°C;

[0030] Flow rate: 1.0 ml per minute;

[0031] Injection volume 20 μL;

[0032] Chromatographic mode: Reverse separation mode.

[0033] Beneficial effects:

[0034] This invention provides a high-performance liquid chromatography (HPLC) method for determining the content of lysine hydrochloride. The method uses amino-bonded silica gel as the packing material, and the mobile phase consists of a 0.04-0.06 mol / L potassium dihydrogen phosphate solution and an acetonitrile solution. The volume ratio of acetonitrile to water in the acetonitrile solution is 9-11:1; the volume ratio of the potassium dihydrogen phosphate solution to the acetonitrile solution is 25-40:60-80. This method is suitable for determining the lysine hydrochloride content in calcium gluconate zinc oral solution, and can solve problems such as excipient interference. It also has advantages such as simple sample pretreatment, high detection sensitivity, high separation efficiency, strong reproducibility, and low cost. Attached Figure Description

[0035] To more clearly illustrate the technical solutions in this invention or the prior art, the accompanying drawings used in the description of the embodiments or the prior art will be described below.

[0036] Figure 1 This is a superimposed high-performance liquid chromatogram of the specificity of lysine hydrochloride detection in Example 1.

[0037] Figure 2 This is a superimposed chromatogram of the detection of lysine hydrochloride in Example 2.

[0038] Figure 3 This is a superimposed chromatogram of the detection of lysine hydrochloride in Example 3.

[0039] Figure 4 The chromatogram for detecting lysine hydrochloride in Comparative Example 1 is shown.

[0040] Figure 5 This is a superimposed chromatogram of the detection of lysine hydrochloride in Comparative Example 2. Detailed Implementation

[0041] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention. Unless otherwise specified, the experimental methods used in the embodiments are all conventional methods; the materials and reagents used are all commercially available.

[0042] The samples used in the following examples and comparative examples were: calcium gluconate and zinc oral solution, Aono (China) Pharmaceutical Co., Ltd., batch number: A2203001, specification: each 10ml contains 0.6g of calcium gluconate, 0.03g of zinc gluconate, and 0.1g of lysine hydrochloride.

[0043] Example 1

[0044] This embodiment provides a method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution, the specific operation of which is as follows:

[0045] 1. Instruments and chromatographic conditions

[0046] Shimadzu LC-20AT high performance liquid chromatograph, quaternary pump, UV detector, autosampler.

[0047] Amino-bonded silica gel was used as the filler (Kromasil amino column 4.6mm×250mm, 5μm).

[0048] The mobile phase consisted of 0.05 mol / L potassium dihydrogen phosphate solution-acetonitrile solution (acetonitrile-water volume ratio in the acetonitrile solution was 10:1) or (potassium dihydrogen phosphate solution-acetonitrile solution volume ratio was 30:70).

[0049] The mobile phase flow rate was 1.0 ml per minute, the detection wavelength was 203 nm, the column temperature was 30 °C, the injection volume was 20 μl, and the run time was 35 min.

[0050] 2. Experimental Procedure

[0051] Blank solvent (also known as diluent): 0.05 mol / L potassium dihydrogen phosphate solution - acetonitrile (volume ratio 60:40).

[0052] Test solution: Accurately measure an appropriate amount of calcium gluconate zinc oral solution and dilute it with blank solvent to prepare a solution containing approximately 1 mg of lysine hydrochloride per 1 ml.

[0053] Preparation of blank excipient solution: Prepare a mixed solution of lysine hydrochloride-free according to the prescription ratio. Accurately measure 1 ml of blank excipient solution, place it in a 10 ml volumetric flask, dilute to the mark with blank solvent, and shake well.

[0054] Reference solution: Weigh an appropriate amount of lysine hydrochloride reference standard accurately, dissolve and quantitatively dilute it with a blank solvent to prepare a reference solution containing approximately 1.0 mg per ml.

[0055] Assay: Accurately measure blank solvent, blank excipient, test solution, and reference solution, and inject them separately into the liquid chromatograph, recording the chromatograms. Results are shown below. Figure 1 .

[0056] 3. Results

[0057] Figure 1 The blank solvent and blank excipient did not produce a peak at the retention time of lysine hydrochloride, meaning they did not interfere with the detection of lysine hydrochloride content. The lysine hydrochloride content determination result was 100.0%, which meets the requirements of the Chinese Pharmacopoeia.

[0058] Example 2

[0059] 1. Instruments and chromatographic conditions

[0060] Shimadzu LC-20AT high performance liquid chromatograph, quaternary pump, UV detector, autosampler.

[0061] Amino-bonded silica gel was used as the filler (Kromasil amino column 4.6mm×250mm, 5μm).

[0062] The mobile phase consisted of 0.05 mol / L potassium dihydrogen phosphate solution-acetonitrile solution (acetonitrile-water volume ratio in acetonitrile solution was 10:1) or (potassium dihydrogen phosphate solution-acetonitrile solution volume ratio was 28:72).

[0063] The mobile phase flow rate was 1.0 ml per minute, the detection wavelength was 203 nm, the column temperature was 30 °C, the injection volume was 20 μl, and the run time was 50 min.

[0064] 2. Experimental Procedure

[0065] Blank solvent: 0.05 mol / L potassium dihydrogen phosphate solution - acetonitrile (volume ratio 60:40).

[0066] Test solution: Accurately measure an appropriate amount of calcium gluconate zinc oral solution and dilute it with blank solvent to prepare a solution containing approximately 1 mg of lysine hydrochloride per 1 ml.

[0067] Preparation of blank excipients: Prepare a mixed solution of lysine hydrochloride-free according to the prescription ratio. Accurately measure 1 ml of the blank excipient solution, place it in a 10 ml volumetric flask, dilute to the mark with blank solvent, and shake well.

[0068] Reference solution: Weigh an appropriate amount of lysine hydrochloride reference standard accurately, dissolve and quantitatively dilute it with a blank solvent to prepare a reference solution containing approximately 1.0 mg per ml.

[0069] For the assay, accurately measure blank solvent, blank excipient, test solution, and reference solution, and inject them separately into the liquid chromatograph, recording the chromatograms. Results are shown below. Figure 2 .

[0070] 3. Results

[0071] Figure 2 The blank solvent and blank excipient do not produce a peak at the retention time of lysine hydrochloride, meaning they do not interfere with the detection of lysine hydrochloride content, which meets the requirements of the Chinese Pharmacopoeia. However, after adjusting the proportion of acetonitrile solution in the mobile phase to 72%, the peak time of lysine hydrochloride shifts later, and the sensitivity of the analytical method decreases slightly. The result of the lysine hydrochloride content determination is 98.5%, and the running time needs to be adjusted to 50 min.

[0072] Example 3

[0073] 1. Instruments and chromatographic conditions

[0074] Shimadzu LC-20AT high performance liquid chromatograph, quaternary pump, UV detector, autosampler.

[0075] Amino-bonded silica gel was used as the filler (Kromasil amino column 4.6mm×250mm, 5μm).

[0076] The mobile phase consisted of 0.05 mol / L potassium dihydrogen phosphate solution-acetonitrile solution (acetonitrile-water volume ratio in acetonitrile solution was 9:1) or (potassium dihydrogen phosphate solution-acetonitrile solution volume ratio was 30:70).

[0077] The mobile phase flow rate was 1.0 ml per minute, the detection wavelength was 203 nm, the column temperature was 30 °C, the injection volume was 20 μl, and the run time was 40 min.

[0078] 2. Experimental Procedure

[0079] Blank solvent: 0.05 mol / L potassium dihydrogen phosphate solution - acetonitrile (volume ratio 60:40).

[0080] Test solution: Accurately measure an appropriate amount of calcium gluconate zinc oral solution and dilute it with blank solvent to prepare a solution containing approximately 1 mg of lysine hydrochloride per 1 ml.

[0081] Preparation of blank excipients: Prepare a mixed solution of lysine hydrochloride-free according to the prescription ratio. Accurately measure 1 ml of the blank excipient solution, place it in a 10 ml volumetric flask, dilute to the mark with blank solvent, and shake well.

[0082] Reference solution: Weigh an appropriate amount of lysine hydrochloride reference standard accurately, dissolve and quantitatively dilute it with a blank solvent to prepare a reference solution containing approximately 1.0 mg per ml.

[0083] For the assay, accurately measure blank solvent, blank excipient, test solution, and reference solution, and inject them separately into the liquid chromatograph, recording the chromatograms. Results are shown below. Figure 3 .

[0084] 3. Results

[0085] Figure 3 The blank solvent and blank excipient do not produce a peak at the retention time of lysine hydrochloride, meaning they do not interfere with the detection of lysine hydrochloride content, which meets the requirements of the Chinese Pharmacopoeia. However, after adjusting the acetonitrile solution ratio to 9:1, the peak time of lysine hydrochloride shifts later, and the lysine hydrochloride content determination result is 98.9%. The running time needs to be adjusted to 40 minutes.

[0086] Comparative Example 1

[0087] 1. Instruments and chromatographic conditions

[0088] Same as Example 1, except that the mobile phase is changed to 0.05 mol / L potassium dihydrogen phosphate solution-acetonitrile (potassium dihydrogen phosphate solution-acetonitrile volume ratio 36.4:63.6).

[0089] 2. Experimental Procedure

[0090] Blank solvent: 0.05 mol / L potassium dihydrogen phosphate solution - acetonitrile (volume ratio 60:40).

[0091] Test solution: Accurately measure an appropriate amount of calcium gluconate zinc oral solution and dilute it with blank solvent to prepare a solution containing approximately 1 mg of lysine hydrochloride per 1 ml.

[0092] Assay: Accurately measure the test solution and inject it into the liquid chromatograph, then record the chromatogram. Results are shown below. Figure 4 .

[0093] 3. Results

[0094] Figure 4 The peak position of lysine hydrochloride in the test solution was interfered with, and the use of potassium dihydrogen phosphate solution-acetonitrile (35:65) as the mobile phase could not accurately quantify lysine hydrochloride.

[0095] Comparative Example 2

[0096] 1. Instruments and chromatographic conditions

[0097] Same as Example 1, except that:

[0098] Octadecylsilane-bonded silica gel is used as a filler;

[0099] The mobile phase was acetonitrile-0.5% sodium acetate solution (pH adjusted to 6.8 with dilute acetic acid) (the volume ratio of acetonitrile to 0.5% sodium acetate was 42:58).

[0100] 2. Experimental Procedure

[0101] Blank solvent: Acetonitrile-0.5% sodium acetate solution (pH adjusted to 6.8 with dilute acetic acid) (volume ratio of acetonitrile to 0.5% sodium acetate is 42:58) was used as blank solvent.

[0102] Preparation of blank excipients: Prepare a mixed solution of lysine hydrochloride-free according to the prescription ratio. Accurately measure 1 ml of the blank excipient solution, place it in a 10 ml volumetric flask, dilute to the mark with blank solvent, and shake well.

[0103] Reference solution: Weigh an appropriate amount of lysine hydrochloride reference standard accurately, dissolve and quantitatively dilute it with a blank solvent to prepare a reference solution containing approximately 1.0 mg per ml.

[0104] Test solution: Accurately measure an appropriate amount of calcium gluconate zinc oral solution and dilute it with blank solvent to prepare a solution containing approximately 1 mg of lysine hydrochloride per 1 ml.

[0105] Assay: Accurately measure blank solvent, blank excipient, reference solution, and test solution, inject them into the liquid chromatograph, and record the chromatogram. Results are shown below. Figure 5 .

[0106] 3. Results

[0107] Figure 5 The peak position of lysine hydrochloride in the test solution was interfered with. Acetonitrile-0.5% sodium acetate solution (pH adjusted to 6.8 with dilute acetic acid) (42:58) as the mobile phase could not accurately quantify lysine hydrochloride.

[0108] The embodiments described above are merely illustrative of several implementations of the present invention, designed to facilitate a detailed understanding of the technical solutions of the present invention. However, they should not be construed as limiting the scope of patent protection. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these modifications and improvements all fall within the scope of protection of the present invention.

Claims

1. A method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography, characterized in that, An amino column was used as the chromatographic column. The mobile phase consisted of a 0.04-0.06 mol / L potassium dihydrogen phosphate solution and an acetonitrile solution. The volume ratio of acetonitrile to water in the acetonitrile solution was 9-11:

1. The volume ratio of potassium dihydrogen phosphate solution to acetonitrile solution was 30:65-75. The detection wavelength was 200 nm to 206 nm. The calcium gluconate zinc oral solution was manufactured by Aono (China) Pharmaceutical Co., Ltd., batch number: A2203001.

2. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to claim 1, characterized in that, The concentration of the potassium dihydrogen phosphate solution is 0.045-0.055 mol / L; and / or, the volume ratio of acetonitrile to water in the acetonitrile solution is 9.5-10.5:

1.

3. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to claim 2, characterized in that, The concentration of the potassium dihydrogen phosphate solution is 0.05 mol / L; and / or, the volume ratio of acetonitrile to water in the acetonitrile solution is 10:

1.

4. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to claim 1, characterized in that, The volume ratio of the potassium dihydrogen phosphate solution to the acetonitrile solution is 30:

70.

5. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to claim 1, characterized in that, The chromatographic column used was a Kromasil amino column, 4.6 mm × 250 mm, 5 μm.

6. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to claim 5, characterized in that, The flow rate of the mobile phase is 0.9-1.1 ml per minute.

7. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to claim 6, characterized in that, The flow rate of the mobile phase is 1.0 ml per minute.

8. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to claim 6, characterized in that, The column temperature is 25-35℃; and / or the injection volume is 15-25 μl; and / or the run time is 30-50 min.

9. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to claim 8, characterized in that, The column temperature is 30℃; and / or the injection volume is 20 μl; and / or the run time is 35 min.

10. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to claim 1, characterized in that, The detection wavelength is 203nm.

11. The method for determining the lysine hydrochloride content in calcium gluconate zinc oral solution by high performance liquid chromatography according to any one of claims 1-10, characterized in that, The blank solvent consists of the potassium dihydrogen phosphate solution and the acetonitrile solution; the volume ratio of the potassium dihydrogen phosphate solution to the acetonitrile solution in the blank solvent is 50-70:40.

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