Liquid chromatographic analysis method for detecting content of pyruvic acid
Through the liquid chromatography analysis method, using aQ column and phosphoric acid aqueous solution, the problems of insufficient retention time and peak shape asymmetry of pyruvate content analysis methods in the prior art were solved, and accurate and efficient detection of pyruvate was achieved.
Patent Information
- Application Number
- CN202510354395.3
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-03-25
- Publication Date
- 2025-04-25
AI Technical Summary
The existing pyruvate content analysis methods have problems such as insufficient retention time and interference of solvent peaks, and the peak shape of ion-to-chromatography is asymmetric, which affects the accuracy of the analysis.
Using liquid chromatography analysis method, using aQ column and phosphoric acid aqueous solution as mobile phases, the peak area of the sample to be tested was detected by preparing the standard solution and determining the standard curve of pyruvate, ensuring that the retention time of pyruvate is close to 4 minutes, and the peak shape is sharp and symmetrical.
Accurate pyruvate content determination is achieved, retention time is extended, peak shape is optimized, solvent peak interference is reduced, and analysis accuracy and efficiency is improved.
Smart Images
Figure CN119881189A_ABST
Abstract
Description
Technical Field
[0001] The present application relates to the technical field of chromatography analysis, and in particular to a liquid chromatography analysis method for detecting pyruvic acid content. Background Art
[0002] Methacrylic acid undergoes ozonation reaction with ozone, followed by reduction with the addition of hexyl sulfide, and finally purified by molecular distillation to obtain pyruvic acid. In order to optimize the reaction temperature, reaction time, reducing agent equivalent, reduction time and other parameters of the two-step synthesis of pyruvic acid, it is necessary to perform real-time analysis of the pyruvic acid content.
[0003] At present, the liquid phase content analysis method of pyruvic acid has almost no retention time and will be interfered by the solvent peak. The use of ion pair chromatography can greatly extend the retention time of pyruvic acid, but the peak shape is asymmetric and the ion pair reagent is more harmful to the chromatographic column. Summary of the invention
[0004] The purpose of the present application is to provide a liquid chromatography analysis method for detecting pyruvic acid content, aiming to solve the problems existing in the existing pyruvic acid content analysis method.
[0005] To achieve the above objectives, the present application provides a liquid chromatography analysis method for detecting the content of pyruvic acid, comprising: Prepare a standard solution and determine a standard curve of pyruvic acid, in which Y is the peak area corresponding to 3.883 min-3.992 min; The sample to be tested was tested by liquid chromatography analysis. In the obtained chromatogram, the peak area corresponding to the target object was the peak area corresponding to 3.883min-3.992min. The liquid chromatography detection parameters included: The chromatographic column was an aQ column and the mobile phase was aqueous phosphoric acid.
[0006] In some embodiments, the phosphoric acid aqueous solution is a 0.1% by volume phosphoric acid aqueous solution.
[0007] In some embodiments, the aQ column is a Thermo Hypersil GOLD™ aQ.
[0008] In some embodiments, the flow rate of the mobile phase is 0.9-1.1 mL / min.
[0009] In some embodiments, the flow rate of the mobile phase is 1 mL / min.
[0010] In some embodiments, the detection wavelength is 200 nm.
[0011] In some embodiments, the column temperature is 20-30°C; preferably 25°C.
[0012] In some embodiments, the standard solution is a sodium pyruvate solution.
[0013] In some embodiments, the injection volume of the sample to be tested is 0.01 mg.
[0014] In some embodiments, the detection time is more than 15 minutes.
[0015] Compared with the prior art, the beneficial effects of this application include: The liquid chromatography analysis method for detecting the pyruvate content provided in the present application adopts an aqueous phosphoric acid solution as the mobile phase and an aQ column that can withstand 100% aqueous phase as the chromatographic column. Under the liquid chromatography detection parameter conditions, the retention time of pyruvate is close to 4 minutes, and the blank solution has no interference here, the peak shape is sharp and symmetrical, and it can be used as an accurate method for determining the pyruvate content. BRIEF DESCRIPTION OF THE DRAWINGS
[0016] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings required for use in the embodiments will be briefly introduced below. It should be understood that the following drawings only show certain embodiments of the present application and therefore should not be regarded as limiting the scope of the present application.
[0017] Figure 1 is the chromatogram of the blank solution; Figure 2 is the chromatogram of the system suitability solution; Figure 3 is the chromatogram of reference solution 1; Figure 4 It is the chromatogram of the acid-destroyed solution of the test sample; Figure 5 is the chromatogram of the test sample's alkaline-destroyed solution; Figure 6 It is the chromatogram of the oxidative destruction solution of the test sample; Figure 7 This is the chromatogram of the high temperature destruction solution of the test sample; Figure 8 It is the chromatogram of the light-destroyed solution of the test sample; Fig. 9 is a linear graph of sodium pyruvate. DETAILED DESCRIPTION
[0018] As used herein: "Prepared from" is synonymous with "comprising." As used herein, the terms "comprising," "including," "having," "containing," or any other variation thereof, are intended to cover a non-exclusive inclusion. For example, a composition, process, method, article, or apparatus that comprises the listed elements is not necessarily limited to only those elements but may include other elements not expressly listed or inherent to such composition, process, method, article, or apparatus.
[0019] The conjunction "consisting of excludes any unspecified element, step, or component. If used in a claim, this phrase renders the claim closed-ended so that it does not include materials other than those described, except for conventional impurities associated therewith. When the phrase "consisting of" appears in a clause of the body of a claim rather than immediately following the subject matter, it limits only the elements described in that clause; other elements are not excluded from the claim as a whole.
[0020] When an amount, concentration, or other value or parameter is expressed as a range, a preferred range, or a range defined by a series of upper preferred values and lower preferred values, this should be understood as specifically disclosing all ranges formed by any pairing of any range upper limit or preferred value with any range lower limit or preferred value, regardless of whether the range is disclosed separately. For example, when a range of "1 to 5" is disclosed, the described range should be interpreted as including ranges "1 to 4", "1 to 3", "1 to 2", "1 to 2 and 4 to 5", "1 to 3 and 5", etc. When a numerical range is described in this article, unless otherwise stated, the range is intended to include its end values and all integers and fractions within the range.
[0021] In these examples, parts and percentages are by mass unless otherwise indicated.
[0022] "Mass parts" refers to the basic unit of measurement for expressing the mass ratio of multiple components. 1 part can represent any unit mass, such as 1g or 2.689g. If we say that the mass parts of component A are a parts and the mass parts of component B are b parts, it means the ratio of the mass of component A to the mass of component B is a:b. Alternatively, it means that the mass of component A is aK and the mass of component B is bK (K is an arbitrary number, indicating a multiple factor). It should not be misunderstood that, unlike the mass parts, the sum of the mass parts of all components is not limited to 100 parts.
[0023] "And / or" is used to indicate that one or both of the stated situations may occur, for example, A and / or B includes (A and B) and (A or B).
[0024] The present application provides a liquid chromatography analysis method for detecting the content of pyruvic acid, comprising: Prepare a standard solution and determine a standard curve of pyruvic acid, in which Y is the peak area corresponding to 3.883 min-3.992 min; The sample to be tested was tested by liquid chromatography analysis. In the obtained chromatogram, the peak area corresponding to the target object was the peak area corresponding to 3.883min-3.992min. The liquid chromatography detection parameters included: The chromatographic column was an aQ column and the mobile phase was aqueous phosphoric acid.
[0025] The liquid chromatography analysis method for detecting the pyruvate content provided in the present application adopts an aqueous phosphoric acid solution as the mobile phase and an aQ column that can withstand 100% aqueous phase as the chromatographic column. Under the liquid chromatography detection parameter conditions, the retention time of pyruvate is close to 4 minutes, and the blank solution has no interference here, the peak shape is sharp and symmetrical, and it can be used as an accurate method for determining the pyruvate content.
[0026] In some embodiments, the phosphoric acid aqueous solution is a 0.1% by volume phosphoric acid aqueous solution.
[0027] In some embodiments, the aQ column is a Thermo Hypersil GOLD™ aQ.
[0028] In some embodiments, the flow rate of the mobile phase is 0.9-1.1 mL / min.
[0029] In some embodiments, the flow rate of the mobile phase is 1 mL / min.
[0030] In some embodiments, the detection wavelength is 200 nm.
[0031] In some embodiments, the column temperature is 20-30°C, for example, it can be 20°C, 21°C, 22°C, 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C, 30°C or any value between 20-30°C; preferably 25°C.
[0032] In some embodiments, the standard solution is a sodium pyruvate solution.
[0033] In some embodiments, the injection volume of the sample to be tested is 0.01 mg.
[0034] In some embodiments, the detection time is more than 15 minutes, for example, it can be 16 minutes, 17 minutes, 18 minutes, 19 minutes, 20 minutes, 21 minutes, 22 minutes, 23 minutes, 24 minutes, 25 minutes, preferably 20 minutes-25 minutes.
[0035] The embodiments of the present application will be described in detail below in conjunction with specific examples, but it will be appreciated by those skilled in the art that the following examples are only used to illustrate the present application and should not be considered as limiting the scope of the present application. If specific conditions are not specified in the examples, they are carried out according to normal conditions or the conditions recommended by the manufacturer. If the manufacturer is not specified for the reagents or instruments used, they are all conventional products that can be purchased commercially.
[0036] Example 1 Example 1 provides a liquid chromatography analysis method for detecting the content of pyruvic acid. The specific experimental method is as follows: Preparation of mobile phase: Measure 1L of ultrapure water into a clean beaker, accurately pipette 1mL of chromatographic pure phosphoric acid and add it, remove bubbles by ultrasonication before use.
[0037] Preparation of reference and test products: Accurately weigh about 31.25 mg of sodium pyruvate reference substance (Tanmo quality inspection, 99.1%), place it in a 50 mL volumetric flask, add mobile phase to dilute to the scale, mix well, and the reference substance is used as a standard solution. On the one hand, it is used for qualitative analysis, and the peak time of the test sample should be consistent with the peak time of the reference substance; on the other hand, it is used for quantitative analysis, and the content of pyruvic acid or sodium pyruvate in the test sample is calculated by the external standard method.
[0038] Accurately weigh about 25 mg of pyruvic acid test sample, place it in a 50 mL volumetric flask, add mobile phase to dilute to the scale, mix well, and obtain.
[0039] Liquid chromatography detection conditions: Instrument: Agilent 1260 Chromatographic column: Thermo Hypersil GOLDTM aQ (250mm*4.6mm, 5um) Detector: DAD Detection wavelength: 200nm Mobile phase: 0.1% phosphoric acid in water Flow rate: 1mL / min Column temperature: 25°C Injection volume: 20uL Detection time: 20 minutes.
[0040] Example 2 System suitability test Solution preparation: Diluent: mobile phase of Example 1.
[0041] Blank solution: mobile phase of Example 1.
[0042] Linear stock solution: Accurately weigh 124.96 mg of sodium pyruvate reference substance (Tanmo quality inspection, 99.1%), place in a 20 mL volumetric flask, add diluent to dissolve and dilute to the scale, mix well, and obtain.
[0043] Stock solution for specificity study of test sample: Accurately weigh 97.60 mg of the test sample from batch 20240913-02, place in a 20 mL volumetric flask, add diluent to dissolve and dilute to the scale, mix well, and obtain.
[0044] Reference solution 1: Accurately pipette 1 mL of the linear stock solution into a 10 mL volumetric flask, add diluent to the mark, mix well, and the solution is ready.
[0045] Reference solution 2: Accurately weigh 31.49 mg of sodium pyruvate reference substance, place in a 50 mL volumetric flask, add diluent to dissolve and dilute to the scale, mix well, and obtain.
[0046] System suitability solution: Accurately weigh 11.45 mg of methacrylic acid reference substance (Tanmo quality inspection, 99.9%) and 31.56 mg of sodium pyruvate reference substance, place in a 50 mL volumetric flask, add diluent to dissolve and dilute to the scale, mix well, and obtain.
[0047] In the liquid chromatography detection system of Example 1, the blank solution was injected twice, the system suitability solution was injected once, the reference solution 1 was injected five times, the reference solution 2 was injected twice, and the reference solution 1 was injected once to detect the suitability of the liquid chromatography detection system of Example 1. The results are shown in Table 1. The chromatogram of the blank solution is shown in Figure 1 The chromatogram of the system suitability solution is shown in Figure 2 As shown, the chromatogram of reference solution 1 is as follows Figure 3 shown.
[0048] Table 1 System suitability results
[0049] In Table 1, the blank solution had no interference with the main peak determination, the separation degree of sodium pyruvate and methacrylic acid in the system applicability solution was ≥1.5, the RSD of the main peak area of the reference solution 1 after five consecutive injections was ≤2.0%, the recovery rates of the reference solution 2 and the reference solution 1 were between 98% and 102%, and the recovery rate of the reference solution 1 after repeated measurement was between 98% and 102%, which proved that the liquid chromatography detection method of Example 1 had good system applicability.
[0050] Example 3 Specificity Test Undamaged sample solution: Accurately pipette 1 mL of the sample specificity research stock solution of Example 2 into a 10 mL volumetric flask, add the diluent of Example 2 to dilute to the mark, and shake well to obtain the solution.
[0051] Acid destruction blank solution: Accurately pipette 1 mL of 1M hydrochloric acid solution into a 10 mL volumetric flask, accurately pipette 1 mL of 1M sodium hydroxide solution to neutralize, add diluent to dilute to the scale, shake well, and obtain.
[0052] Test sample acid destruction solution: accurately pipette 1 mL of the test sample specificity research stock solution of Example 2 into a 10 mL volumetric flask, accurately pipette 1 mL of 1 M hydrochloric acid solution, shake well, let stand at room temperature for 2 h, accurately pipette 1 mL of 1 M sodium hydroxide solution to neutralize, add diluent to dilute to the scale, shake well, and obtain.
[0053] Alkali destruction blank solution: Accurately pipette 1 mL of 1M sodium hydroxide solution into a 10 mL volumetric flask, accurately pipette 1 mL of 1M hydrochloric acid solution to neutralize, add diluent to dilute to the scale, shake well, and obtain.
[0054] Test sample alkaline destruction solution: accurately pipette 1 mL of the test sample specificity research stock solution of Example 2 into a 10 mL volumetric flask, accurately pipette 1 mL of 1 M sodium hydroxide solution, shake well, let stand at room temperature for 2 h, accurately pipette 1 mL of 1 M hydrochloric acid solution to neutralize, add diluent to dilute to the scale, shake well, and obtain.
[0055] Oxidative destruction of blank solution: Accurately pipette 1 mL of 1% hydrogen peroxide solution into a 10 mL volumetric flask, add diluent to dilute to the mark, shake well, and the solution is ready.
[0056] Oxidative destruction solution for the test sample: accurately pipette 1 mL of the test sample specificity research stock solution of Example 2 into a 10 mL volumetric flask, add 1 mL of 1% hydrogen peroxide solution, shake well, let stand at room temperature for 2 h, add diluent to dilute to the scale, shake well, and obtain.
[0057] High-temperature destruction solution of the test sample: accurately pipette 1 mL of the test sample specificity research stock solution of Example 2 into a 10 mL volumetric flask, then place it in a 60°C water bath for 2 h, take it out, add diluent to dilute to the scale, shake well, and obtain.
[0058] Test sample light-destruction solution: Accurately pipette 1 mL of the test sample specificity research stock solution of Example 2 into a 10 mL volumetric flask, then place it under light for 2 h, take it out, add diluent to dilute to the scale, and shake well to obtain the solution.
[0059] In the liquid chromatography detection system of Example 1, the blank solution of Example 1, the reference solution 1 and the above solution were injected once each. The test results are shown in Table 2. The chromatogram of the test acid destruction solution is shown in Figure 4 As shown, the chromatogram of the test sample alkali destruction solution is as follows Figure 5 As shown, the chromatogram of the oxidative destruction solution of the test sample is as follows Figure 6 As shown, the chromatogram of the high temperature destruction solution of the test sample is as follows Figure 7As shown, the chromatogram of the light-destroyed solution of the test sample is as follows Figure 8 shown.
[0060] Table 2 Specificity results
[0061] According to Table 2, the blank solution and each destroyed blank solution have no interference with the determination of the main peak. In each test sample destroyed solution, the degradation impurities have no interference with the determination of the pyruvic acid main peak. The separation degree between the methacrylic acid peak and the adjacent impurity peak is ≥1.5. In each test sample destroyed solution, the main peak retention time is consistent with the main peak retention time of the reference substance, and the retention time ratio is between 0.98 and 1.02, which proves that the method of this application has good specificity.
[0062] Example 4 Linearity and range test Prepare the linear solution according to Table 3, accurately pipette the corresponding volume of the linear stock solution of Example 2, place it in a 10 mL volumetric flask, add the diluent of Example 2 to dilute to the scale, and shake well.
[0063] In the liquid chromatography detection system of Example 1, the above solutions were taken and injected once each. The results are shown in Table 4 and Fig. 9 As shown, because the reference substance is sodium pyruvate, the molecular weight of sodium pyruvate is 110, and the molecular weight of pyruvic acid is 88, the concentration in the sodium pyruvate standard curve can be multiplied by 0.8 to calculate the content of pyruvic acid.
[0064] Table 3 Linear solution preparation
[0065] Table 4 Linearity results
[0066] According to Table 4, the linear results can report the linear equation, and R 2 The absolute value of the ratio of the Y-axis intercept to the 100% concentration response value is not less than 0.999, and the linear range exists, which proves that the linearity of the method of this application is good.
[0067] Example 5 Repeatability Test Test solution (reproducible solution): Take about 25 mg of the test sample from batch 20240911, place it in a 25 mL volumetric flask, add the diluent of Example 2 to dissolve and dilute to the mark, and mix well. Accurately pipette 1 mL of the above solution, place it in a 10 mL volumetric flask, add the diluent to the mark, and mix well. Prepare 6 portions in parallel.
[0068] In the liquid chromatography detection system of Example 1, the above solutions were taken and injected once each. The results are shown in Table 5.
[0069] Table 5 Repeatability results
[0070] According to Table 5, the RSDs of the six solutions were all less than 2%, which proved that the precision (repeatability) of the method was good.
[0071] Example 6 Accuracy Test Test sample spiked solution (80% accuracy solution): Take about 25 mg of the test sample from batch 20240913-02 and 0.8 mL of the linear stock solution of Example 2, place in the same 50 mL volumetric flask, add diluent to dilute to the mark, mix well, and the solution is obtained. Prepare 3 portions in parallel.
[0072] Test sample spike solution (100% accuracy solution): Take about 25 mg of the test sample from batch 20240913-02 and 1.0 mL of the linear stock solution, place them in the same 50 mL volumetric flask, add diluent to dilute to the mark, mix well, and the solution is ready. Prepare three copies in parallel.
[0073] Test sample spike solution (120% accuracy solution): Take about 25 mg of the test sample from batch 20240913-02 and 1.2 mL of the linear stock solution, place them in the same 50 mL volumetric flask, add diluent to dilute to the mark, mix well, and the solution is ready. Prepare 3 copies in parallel.
[0074] In the liquid chromatography detection system of Example 1, the above solutions were taken and injected once each. The results are shown in Table 6.
[0075] Table 6 Accuracy results
[0076] According to Table 6, the average recovery rate of each concentration is between 80% and 120%, which proves that the accuracy of the method of this application is good.
[0077] Example 7 Stability Test The reference solution 1 of Example 2 and the undamaged solution of the test sample of Example 3 were injected once at different times to observe the ratio of the peak area to 0h. The results are shown in Table 7.
[0078] Table 7 Stability results
[0079] According to Table 7, the comparison of the subsequent test results of reference solution 1 and the test solution with those at 0 hours showed a ratio between 98% and 102%. The reference solution was stable within 15.7 hours at room temperature, and the test solution was stable within 12.5 hours at room temperature.
[0080] Example 8 Durability Test According to Table 8, the column temperature and flow rate were slightly changed. After the chromatographic system was balanced, two injections of the blank solution of Example 2, one injection of the system suitability solution, five injections of the reference solution 1, two injections of the reference solution 2, and one injection of the undamaged solution of the test sample of Example 3 were injected respectively.
[0081] Table 8 Chromatographic parameter adjustment
[0082] In all test results, the blank solution has no interference with the main peak determination. If there is interference, the blank response value does not exceed 0.05% of the reference solution response value; in the system suitability solution, the separation degree of sodium pyruvate and methacrylic acid is ≥1.5. The RSD of the main peak area of reference solution 1 after 5 consecutive injections is ≤2.0%; the recovery rate of reference solution 2 and reference solution 1 should be between 98% and 102%; the recovery rate of reference solution 1 after retest is between 98% and 102%; the ratio of the content determination result of the test solution to the determination result under the initial conditions is between 98% and 102%. All results meet the requirements, proving that the durability of this application method is good.
[0083] Comparative Example 1 Comparative Example 1 provides a liquid chromatography analysis method for detecting the content of pyruvic acid. The specific experimental method differs from that of Example 1 in that: The mobile phase was: 0.2 M potassium dihydrogen phosphate: acetonitrile = 95:5; The chromatographic column is: C18 Acclaim 120.
[0084] Other conditions were the same as those in Example 1. Under the conditions in Comparative Example 1, the elution time of pyruvic acid was 2.8 min, with no retention and interference by the solvent peak.
[0085] Comparative Example 2 Comparative Example 2 provides a liquid chromatography analysis method for detecting the content of pyruvic acid. The specific experimental method differs from that of Example 1 in that: Ion-pair chromatography with mobile phase: disodium hydrogen phosphate and tetrahexylammonium hydrogen sulfate; The chromatographic column is: TC-C18.
[0086] Other conditions were the same as those in Example 1. The retention time of pyruvic acid was extended to 5.6 min, but the peak shape was not good. The peak shape was not good when the pH was adjusted to 2 or 5.
[0087] The retention time of the liquid phase content determination method of pyruvic acid in the current literature is less than 3 minutes, there is no retention, and it is easily interfered by the solvent peak. Although ion pair chromatography can increase the retention time, the peak shape is asymmetric. This application uses an aQ column and a 0.1% phosphoric acid aqueous solution as the mobile phase to increase the retention time of pyruvic acid to nearly 4 minutes, and the peak shape is symmetrical, the mobile phase is simple to prepare, and the sample can be injected after dilution. It is an efficient, accurate and convenient method for determining the content of pyruvic acid.
[0088] The present application adopts sodium pyruvate as a standard to measure the content of pyruvic acid, because the content of the pyruvic acid standard purchased on the market is not high. For example, the pyruvic acid purchased from Tan Mo and the China Inspection Institute is used to calibrate the pyruvic acid content made by the applicant, and the content is more than 100%. The sodium pyruvate of Tan Mo or the China Inspection Institute is used to calibrate their own pyruvic acid, and the content is about 93%, while the sodium pyruvate of Tan Mo or the China Inspection Institute is used to calibrate the pyruvic acid made by the applicant, and the content is 99%. Therefore, the pyruvic acid made by the applicant is higher than the content of the pyruvic acid standard of Tan Mo and the China Inspection Institute. In addition, the process of the present application is also to make sodium pyruvate first, and then acidify and distill to obtain pyruvic acid. The content of the intermediate sodium pyruvate has a great influence on the content of the product pyruvic acid, and it is required to be above 98%. Therefore, the method of the present application can not only measure the content of the final product pyruvic acid, but also measure the content of the intermediate sodium pyruvate, which can be used for central control.
[0089] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present application, rather than to limit it. Although the present application has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present application.
[0090] In addition, those skilled in the art will appreciate that, although some embodiments herein include certain features included in other embodiments but not other features, the combination of features of different embodiments is meant to be within the scope of the present application and form different embodiments. For example, in the above claims, any one of the claimed embodiments may be used in any combination. The information disclosed in this background technology section is intended only to deepen the understanding of the overall background technology of the present application and should not be regarded as an admission or in any form of implication that the information constitutes prior art known to those skilled in the art.
Claims
1. A liquid chromatography analysis method for detecting pyruvic acid content, characterized in that: include: Prepare a standard solution and determine a standard curve of pyruvic acid, in which Y is the peak area corresponding to 3.883 min-3.992 min; The sample to be tested was tested by liquid chromatography analysis. In the obtained chromatogram, the peak area corresponding to the target object was the peak area corresponding to 3.883min-3.992min. The liquid chromatography detection parameters included: The chromatographic column was an aQ column and the mobile phase was aqueous phosphoric acid.
2. The liquid chromatography analysis method for detecting pyruvic acid content according to claim 1, characterized in that: The phosphoric acid aqueous solution is a phosphoric acid aqueous solution with a volume percentage of 0.1%.
3. The liquid chromatography analysis method for detecting pyruvic acid content according to claim 1, characterized in that: The aQ column is Thermo Hypersil GOLDTM aQ.
4. The liquid chromatography analysis method for detecting pyruvic acid content according to claim 1, characterized in that: The flow rate of the mobile phase is 0.9-1.1 mL / min.
5. The liquid chromatography analysis method for detecting pyruvic acid content according to claim 4, characterized in that: The flow rate of the mobile phase was 1 mL / min.
6. The liquid chromatography analysis method for detecting pyruvic acid content according to claim 1, characterized in that: The detection wavelength was 200 nm.
7. The liquid chromatography analysis method for detecting pyruvic acid content according to claim 1, characterized in that: The column temperature is 20-30°C; preferably 25°C.
8. The liquid chromatography analysis method for detecting pyruvic acid content according to claim 7, characterized in that: The standard solution is a sodium pyruvate solution.
9. The liquid chromatography analysis method for detecting pyruvic acid content according to claim 1, characterized in that: The injection volume of the sample to be tested is 0.01 mg.
10. The liquid chromatography analysis method for detecting pyruvic acid content according to claim 1, characterized in that: The detection time is more than 15 minutes.