Method for detecting content of myristoyl propyl dimethylamine in corneal contact lens care product by HPLC (High Performance Liquid Chromatography) method

Abstract

The invention provides a method for detecting the content of myristoyl propyl dimethylamine in a corneal contact lens care product by an HPLC (High Performance Liquid Chromatography) method, which is a novel method for accurately detecting the content of myristoyl propyl dimethylamine in the corneal contact lens care product. In the prior art, MPA is widely used as a preservative in cosmetics and corneal contact lens care products, but MPA is insoluble in water due to good solubility in an organic solvent and is easily interfered by other polar components in a care solution, so that the existing analysis method has limitation. According to the method, the MPA characteristic peak is effectively separated by optimizing the mobile phase ratio and selecting the chromatographic column, the interference of other polar components is overcome, and the accuracy and the sensitivity of a detection result are ensured. A methodology verification result shows that the method has a good linear relation, a high recovery rate and good repeatability. The HPLC method provides a reliable technical means for the quality control of the corneal contact lens care product, and is beneficial to ensuring the quality safety of the product and protecting the eye health of a user.

Description

Technical Field

[0001] The invention relates to the field of medical detection, and in particular to a method and a detection system for detecting the content of myristoylpropyl dimethylamine in corneal contact lens care products by HPLC. Background Art

[0002] As a preservative, myristoylpropyl dimethylamine is widely used in cosmetics and contact lens care products. Since it is a fat-soluble substance and insoluble in water, but has good solubility in organic solvents (such as ethanol, acetonitrile, etc.), and may irritate the eyes and skin at a certain concentration, it is particularly important to monitor the content of myristoylpropyl dimethylamine in contact lens care products. However, the existing quality control methods for myristoylpropyl dimethylamine are mostly aimed at the analysis of pure substances, while the care solution products contain a large amount of highly polar surfactants, moisturizing lubricants and other ingredients, which will seriously interfere with the analysis of myristoylpropyl dimethylamine.

[0003] In order to solve the above problems, the applicant proposed a method for detecting the content of myristoylpropyl dimethylamine in corneal contact lens care products by HPLC. Summary of the invention

[0004] The object of the present invention is to provide a method for detecting the content of myristoylpropyl dimethylamine in a corneal contact lens care product by HPLC, so as to solve the problems in the prior art.

[0005] To achieve the above object, the present invention provides the following technical solution: a method for detecting the content of myristoylpropyl dimethylamine in a corneal contact lens care product by HPLC, comprising the following steps:

[0006] S1. Prepare a mobile phase, which is a mixture of 0.025 mol / L sodium dihydrogen phosphate solution and acetonitrile in a volume ratio of 65:35, wherein the 0.025 mol / L sodium dihydrogen phosphate solution is prepared by accurately weighing 3.9 g of sodium dihydrogen phosphate (NaH2PO4·2H2O), adding it to 1000 mL of water to dissolve it, and adjusting the pH value of the solution to 3.80 with phosphoric acid, and then filtering it through a 0.45 micron water system membrane to obtain a clear mobile phase solution;

[0007] S2. Prepare the contact lens care product to be tested as the test solution, and prepare a blank solution as a control to ensure that there is no external interference during the test;

[0008] S3. High performance liquid chromatography (HPLC) was used for detection. Agilent ZORBAX SB-CN chromatographic column was selected as the separation medium and equipped with a UV detector as the detection means. The chromatographic operation parameters were set to include a flow rate of 1.0 mL / min, a UV detection wavelength of 218 nm, a column temperature of 30 °C, a total operation time of 15 minutes, and an injection volume of 50 μL each time.

[0009] S4. Under the above-set chromatographic conditions, the blank solution, the control sample solution with a known concentration, and the test solution to be tested are analyzed by high performance liquid chromatography in turn, and the chromatogram of each solution is recorded and obtained, wherein the reference retention time of myristoylpropyl dimethylamine is about 11 minutes;

[0010] S5. Based on the characteristic peak area of ​​myristoylpropyl dimethylamine in the chromatogram, the standard curve (y=2.2377x-0.0841, R 2 =0.99996) to calculate the exact content of myristoylpropyl dimethylamine in the test solution.

[0011] Optionally, the method for preparing the control sample solution of known concentration specifically includes: accurately weighing 0.01 g of myristoylpropyl dimethylamine of known content, placing it in a 100 mL volumetric flask, adding the prepared mobile phase solution, fully dissolving and diluting to the scale line, shaking well, then accurately pipetting 0.5 mL of the above solution into another 10 mL volumetric flask, adding the mobile phase again to dilute to the scale line, shaking well for later use.

[0012] Optionally, the linear relationship of the method is verified, and its linear equation is y=2.2377x-0.0841, and the correlation coefficient R2 is as high as 0.99996, indicating that the method has a good linear response within different concentration ranges.

[0013] Optionally, the accuracy, precision and repeatability of the method are all verified by strict methodology, and the recovery rate ranges from 93% to 101%, which meets the relevant requirements of the 9101 analytical method validation guidelines of Part IV of the Chinese Pharmacopoeia.

[0014] A detection system for the detection method according to any one of claims 1 to 4, the system comprising a liquid chromatograph, an ultraviolet detector, an Agilent ZORBAX SB-CN chromatographic column or an equivalent chromatographic column, and various equipment and reagents required for accurately preparing a mobile phase.

[0015] Optionally, the liquid chromatograph has a programmable control function, which can automatically set and adjust chromatographic parameters such as flow rate, column temperature, detection wavelength, etc., to improve detection efficiency and accuracy.

[0016] Optionally, data processing and analysis software is also included for automatically collecting chromatogram data, calculating peak areas, and automatically calculating the content of myristoylpropyl dimethylamine based on a standard curve, thereby reducing human errors and improving the reliability of analysis results.

[0017] Optionally, the method and system are not only applicable to corneal contact lens care products, but also extended to the quality inspection of other cosmetics or daily chemical products containing myristoylpropyl dimethylamine.

[0018] Beneficial effects: Efficient separation: By optimizing the mobile phase composition and chromatographic detection conditions, the present invention can effectively separate surfactants such as polyoxyethylenediamine and amine salt preservatives with similar polarity from the substance to be analyzed, myristoylpropyl dimethylamine, and the resulting chromatogram has a stable baseline.

[0019] High accuracy: This method has been methodologically validated, and its linearity, repeatability, and recovery rate are in line with the relevant statements in the guidelines for validation of analytical methods in Part IV of the Chinese Pharmacopoeia 9101. The sensitivity reaches 0.8 μg / mL, and it can accurately determine the content of myristoylpropyl dimethylamine in corneal contact lens care products.

[0020] Wide application: This method is not only suitable for the quality control of corneal contact lens care products, but can also be extended to the quality inspection of other cosmetics or daily chemical products containing myristoylpropyl dimethylamine, and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 : shows the chromatogram of the blank solution according to this protocol.

[0022] Figure 2 : shows the chromatogram of the reference solution according to this protocol.

[0023] Figure 3 : A chromatogram showing the contact lens care solution according to this protocol.

[0024] Figure 4 : Shows the chromatogram of adding a reference substance to the contact lens care solution according to this protocol.

[0025] Figure 5 : shows the chromatogram of the reference solution according to Comparative Example 1.

[0026] Figure 6 : shows the chromatogram of the contact lens care solution according to Comparative Example 1.

[0027] Figure 7 : shows the chromatogram of the reference solution according to Comparative Example 2.

[0028] Figure 8: shows the chromatogram of the contact lens care solution according to Comparative Example 2.

[0029] Fig. 9 : shows the chromatogram of the reference solution according to Comparative Example 3.

[0030] Fig.10 : shows the chromatogram of the contact lens care solution according to Comparative Example 3.

[0031] Fig.11 : shows the chromatogram of the reference solution according to Comparative Example 4.

[0032] Fig.12 : shows the chromatogram of the contact lens care solution according to Comparative Example 4.

[0033] Fig.13 : shows the chromatogram of the reference solution according to Comparative Example 5.

[0034] Fig.14 : shows the chromatogram of the contact lens care solution according to Comparative Example 5. DETAILED DESCRIPTION

[0035] The following describes the preferred embodiments of the present invention with reference to the drawings in the specification, so that the technical content is clearer and easier to understand. The present invention can be embodied in many different forms of embodiments, and the protection scope of the present invention is not limited to the embodiments mentioned in the text.

[0036] Example 1

[0037] The present invention provides a method for detecting the content of myristamidopropyl dimethylamine (MPDMA) in a contact lens care product by using high performance liquid chromatography (HPLC). The method aims to solve the problem that it is difficult to accurately detect the content of MPDMA in a complex matrix in the prior art, especially in a complex system such as a contact lens care solution containing a variety of polar substances such as surfactants and moisturizing lubricants. By optimizing the mobile phase composition, chromatographic column selection and detection conditions, the present invention successfully achieves efficient separation and accurate detection of MPDMA, and has the advantages of good repeatability and high recovery rate.

[0038] Summary of the technical solution: The present invention adopts HPLC method to accurately determine the content of MPDMA in corneal contact lens care solution by precisely controlling the composition of mobile phase, selecting a suitable chromatographic column and optimizing detection conditions. The key to this method is the selection of mobile phase and the optimization of chromatographic conditions to ensure that the characteristic peak of MPDMA can be effectively separated from other components, while obtaining good peak shape and baseline stability.

[0039] Specific implementation steps

[0040] Instrument conditions

[0041] Liquid chromatograph: Agilent 1260HPLC system or other equivalent HPLC system equipped with UV detector (VWD).

[0042] Chromatographic column: Agilent ZORBAX SB-CN chromatographic column (150 mm × 4.6 mm, 5 μm) or other equivalent chromatographic columns are used. This chromatographic column is suitable for analyzing polar compounds and has a high sample loading capacity and good selectivity.

[0043] Other auxiliary equipment: including electronic balance, ultrasonic cleaner, pipette, volumetric flask, etc.

[0044] Reagents: Myristoylpropyl dimethylamine standard: purity not less than 98%.

[0045] Mobile phase: 0.025 mol / L sodium dihydrogen phosphate solution and acetonitrile in a volume ratio of 65:35. Sodium dihydrogen phosphate solution is prepared by weighing 3.9 g of sodium dihydrogen phosphate (NaH2PO4·2H2O) and dissolving it in 1000 mL of water, adjusting the pH to 3.80 with phosphoric acid, and filtering through a 0.45 μm water membrane before use. Acetonitrile must be HPLC grade.

[0046] Corneal contact lens care products: commercially available or homemade, used as samples to be tested.

[0047] Other reagents: including purified water (in compliance with relevant standards of the Chinese Pharmacopoeia), methanol (HPLC grade, used for comparative tests), etc.

[0048] The blank solution is the mobile phase and no additional preparation is required.

[0049] Accurately weigh 0.01 g of MPDMA standard with known content, place it in a 100 mL volumetric flask, dissolve and dilute to the scale with mobile phase, shake well, and obtain a stock solution with a concentration of 100 μg / mL.

[0050] Accurately pipette an appropriate amount of the above stock solution and dilute it with mobile phase to the required concentration (e.g. 5 μg / mL) as the reference solution.

[0051] Directly take an appropriate amount of corneal contact lens care product as the test solution, and dilute it appropriately if necessary to meet the test requirements.

[0052] Flow rate: 1.0mL / min.

[0053] Column temperature: 30℃.

[0054] Detection wavelength: 218 nm (selected according to the ultraviolet absorption characteristics of MPDMA).

[0055] Injection volume: 50 μL.

[0056] Running time: 15 min (determined according to the retention time of MPDMA).

[0057] For corneal contact lens care products, usually no complicated pre-treatment is required, and an appropriate amount can be directly taken as the test solution for analysis. If the sample matrix is ​​complex or the MPDMA content is low, appropriate dilution or purification can be considered.

[0058] Chromatographic analysis: Inject the blank solution, reference solution and test solution into the HPLC system for analysis.

[0059] Record the chromatogram and observe the retention time, peak shape and separation degree of the MPDMA characteristic peak.

[0060] The linear relationship between the peak area and concentration of MPDMA was determined based on the chromatogram of the reference solution, and a standard curve was established.

[0061] Calculate the MPDMA content based on the chromatogram of the test solution.

[0062] To ensure the reliability of the detection method, a series of methodological validations are required, including but not limited to linear range, precision, accuracy, specificity and detection limit.

[0063] Prepare a series of MPDMA reference solutions of different concentrations, inject and analyze them separately, and record the peak area. Plot a standard curve with concentration (C) as the horizontal axis and peak area (A) as the vertical axis, and calculate the regression equation and correlation coefficient (R2). The linear range of this method can generally cover the expected concentration range of MPDMA in the sample to be tested.

[0064] Select MPDMA reference solutions of low, medium and high concentrations, inject them 6 times in a row, and calculate the relative standard deviation (RSD) of the peak area to evaluate the intra-day precision. Take another batch of reference solutions, repeat the above operation on different days, and calculate the inter-day precision. This method should have good intra-day and inter-day precision.

[0065] The accuracy is evaluated by the spike recovery test. Take an appropriate amount of corneal contact lens care product with a known MPDMA content, add different amounts of MPDMA standards, treat it according to the test solution preparation method, and then analyze it. Calculate the recovery rate and RSD value to evaluate the accuracy of the method. The recovery rate of this method should be within the specified range (usually 80% to 120%), and the RSD value should be small.

[0066] The specificity of the method was evaluated by comparing the chromatograms of the blank solution, the reference solution, and the test solution. It should be ensured that the characteristic peak of MPDMA is completely separated from the peaks of other components and that there are no interfering peaks in the blank solution.

[0067] The limit of detection (LOD) and limit of quantification (LOQ) of the method were determined by gradually diluting the reference solution to concentration points where the signal-to-noise ratio (S / N) was approximately 3 and 10. The LOD and LOQ of this method should meet the actual detection requirements.

[0068] Under the optimized chromatographic conditions, the characteristic peak of MPDMA was effectively separated from other component peaks (see Figure 1 The retention time of MPDMA was stable, the peak shape was symmetrical, and the baseline was smooth, indicating that the method had good separation and selectivity.

[0069] Linear range: This method has a good linear relationship in the concentration range of 0.1-10 μg / mL, and the regression equation is y=2.2377x-0.0841, R 2 =0.99996.

[0070] Precision: The intra-day and inter-day precision RSD values ​​were both less than 2%, indicating that the instrument had good precision.

[0071] Accuracy: The spiked recoveries were between 93% and 101%, and the RSD values ​​were less than 5%, indicating that the method had high accuracy.

[0072] Specificity: No interfering peaks appeared in the blank solution, and the characteristic peak of MPDMA was completely separated from the peaks of other components (see Figure 1 ).

[0073] Detection limit and quantification limit: LOD is 0.02μg / mL, LOQ is 0.08μg / mL, which meets the actual detection needs.

[0074] Here are 5 comparative examples:

[0075] The difference between Comparative Example 1 and Example 1 is that the mobile phase is methanol: acetonitrile: 0.01 mol / L potassium dihydrogen phosphate aqueous solution (pH = 4.00) = 20:60:20 (v / v / v), flow rate: 0.8 mL / min, injection volume: 80 μL, and other chromatographic conditions are the same as those in Example 1. The sample solution to be tested is tested under the chromatographic conditions of this comparative example, and the obtained HPLC chromatogram is as shown in FIG. Figure 5 , Figure 6 shown.

[0076] By comparison Figure 5 , Figure 6 It can be seen that myristoylpropyl dimethylamine eluted early, the baseline of the sample solution to be tested was too high, and the characteristic peaks were difficult to separate.

[0077] Comparative Example 2

[0078] The difference between Comparative Example 2 and Example 1 is that the mobile phase is methanol: 0.025 mol / L sodium dihydrogen phosphate aqueous solution (pH = 3.80) = 60:40 (v / v), and the other chromatographic conditions are the same as those of Example 1. The sample solution to be tested is tested under the chromatographic conditions of this comparative example, and the obtained HPLC chromatogram is as follows Figure 7 , Figure 8 shown.

[0079] By comparison Figure 7 , Figure 8 It can be seen that myristoylpropyl dimethylamine eluted earlier, the baseline of the sample solution to be tested was too high, and the characteristic peaks were difficult to separate.

[0080] Comparative Example 3

[0081] The difference between Comparative Example 3 and Example 1 is that the mobile phase is methanol: 0.025 mol / L sodium dihydrogen phosphate aqueous solution (pH = 3.80) = 0min ~ 10min (45:55), 20min (70:30), 30min ~ 37min (85:15) (v / v), and the other chromatographic conditions are the same as those in Example 1. The sample solution to be tested is tested under the chromatographic conditions of this comparative example, and the obtained HPLC chromatogram is as shown Fig. 9 , Fig.10 shown.

[0082] from Fig. 9 , Fig.10 It can be seen that myristoylpropyl dimethylamine has a peak in the gradient change, and the peak shape is not good. The peak of the sample solution to be tested is flattened and cannot be accurately identified.

[0083] Comparative Example 4

[0084] The difference between Comparative Example 4 and Example 1 is that the mobile phase is methanol: 0.025 mol / L sodium dihydrogen phosphate aqueous solution (pH = 3.80) = 0 min (45:55), 10 min to 20 min (60:40), 21 min to 25 min (45:55) (v / v), and the other chromatographic conditions are the same as those in Example 1. The sample solution to be tested is tested under the chromatographic conditions of this comparative example, and the obtained HPLC chromatogram is as shown Fig.11 , Fig.12 shown.

[0085] from Fig.11 , Fig.12 It can be seen that the two chromatographic peaks are close to each other, the two peaks have not achieved complete baseline separation, and the separation degree is poor.

[0086] Comparative Example 5

[0087] The difference between Comparative Example 5 and Example 1 is that the mobile phase is acetonitrile: 0.025 mol / L sodium dihydrogen phosphate aqueous solution (pH=3.80)=0 min (45:55), 10 min to 20 min (60:40), 21 min to 25 min (45:55) (v / v), and the other chromatographic conditions are the same as those in Example 1. The sample solution to be tested is tested under the chromatographic conditions of this comparative example, and the obtained HPLC chromatogram is as shown in FIG. Fig.13 , Fig.14 shown.

[0088] from Fig.13 , Fig.14 It can be seen that the two chromatographic peaks are close to each other, the two peaks have not achieved complete baseline separation, and the separation degree is poor.

[0089] In summary, this method uses a mobile phase with a ratio of organic phase to buffer salt of 35:65 and a buffer salt pH of 3.8. This method can separate surfactants and amine salt preservatives with similar polarity such as polyoxyethylenediamine from the substance to be analyzed, myristoylpropyl dimethylamine, and the baseline of the resulting chromatogram is stable.

[0090] The HPLC method provided by the present invention for detecting the content of MPDMA in a corneal contact lens care product has the advantages of good separation effect, high accuracy, good repeatability, etc. By optimizing the mobile phase composition and chromatographic conditions, efficient separation and accurate detection of MPDMA in a complex matrix are successfully achieved. The method can be used for daily quality control of corneal contact lens care products to ensure that the content of MPDMA in the product meets the standard requirements and ensures the safety of consumers.

[0091] The above shows and describes the basic principles and main features of the present invention and the advantages of the present invention. It is obvious to those skilled in the art that the present invention is not limited to the details of the above exemplary embodiments, and the present invention can be implemented in other specific forms without departing from the spirit or basic features of the present invention. Therefore, no matter from which point of view, the embodiments should be regarded as exemplary and non-restrictive. The scope of the present invention is defined by the attached claims rather than the above description, and it is intended that all changes falling within the meaning and scope of the equivalent elements of the claims are included in the present invention. Any figure mark in the claims should not be regarded as limiting the claims involved.

[0092] In addition, it should be understood that although the present specification is described according to implementation modes, not every implementation mode contains only one independent technical solution. This description of the specification is only for the sake of clarity. Those skilled in the art should regard the specification as a whole. The technical solutions in each embodiment may also be appropriately combined to form other implementation modes that can be understood by those skilled in the art.

Claims

1. A method for detecting the content of myristoylpropyl dimethylamine in a contact lens care product by HPLC, characterized in that: The following steps are involved: S1. Prepare a mobile phase, which is a mixture of 0.025 mol / L sodium dihydrogen phosphate solution and acetonitrile in a volume ratio of 65:35, wherein the 0.025 mol / L sodium dihydrogen phosphate solution is prepared by accurately weighing 3.9 g of sodium dihydrogen phosphate (NaH2PO4·2H2O), adding it to 1000 mL of water to dissolve it, and adjusting the pH value of the solution to 3.80 with phosphoric acid, and then filtering it through a 0.45 micron water system membrane to obtain a clear mobile phase solution; S2. Prepare the contact lens care product to be tested as the test solution, and prepare a blank solution as a control to ensure that there is no external interference during the test; S3. High performance liquid chromatography (HPLC) was used for detection. Agilent ZORBAX SB-CN chromatographic column was selected as the separation medium and equipped with a UV detector as the detection means. The chromatographic operation parameters were set to include a flow rate of 1.0 mL / min, a UV detection wavelength of 218 nm, a column temperature of 30 °C, a total operation time of 15 minutes, and an injection volume of 50 μL each time. S4. Under the above-set chromatographic conditions, the blank solution, the control sample solution with a known concentration, and the test solution to be tested are analyzed by high performance liquid chromatography in turn, and the chromatogram of each solution is recorded and obtained, wherein the reference retention time of myristoylpropyl dimethylamine is about 11 minutes; S5. Based on the characteristic peak area of ​​myristoylpropyl dimethylamine in the chromatogram, the standard curve (y=2.2377x-0.0841, R 2 =0.99996) to calculate the exact content of myristoylpropyl dimethylamine in the test solution.

2. The method according to claim 1, characterized in that The method for preparing the control sample solution of known concentration specifically comprises: accurately weighing 0.01 g of myristoylpropyl dimethylamine of known content, placing it in a 100 mL volumetric flask, adding the prepared mobile phase solution, fully dissolving and diluting to the scale line, shaking well, then accurately pipetting 0.5 mL of the above solution into another 10 mL volumetric flask, adding the mobile phase again to dilute to the scale line, shaking well for later use.

3. The method according to claim 1 or 2, characterized in that: The linear relationship of the method has been verified, and its linear equation is y=2.2377x-0.0841, and the correlation coefficient R 2 As high as 0.99996, it shows that the method has good linear response in different concentration ranges.

4. The method according to any one of claims 1 to 3, characterized in that The accuracy, precision and repeatability of the method were strictly verified by methodology, and the recovery rate ranged from 93% to 101%, which met the relevant requirements of the 9101 analytical method validation guidelines of Part IV of the Chinese Pharmacopoeia.

5. A detection system for the detection method according to any one of claims 1 to 4, characterized in that: The system includes a liquid chromatograph, a UV detector, an Agilent ZORBAX SB-CN column or equivalent, and various equipment and reagents required for accurately preparing the mobile phase.

6. The detection system according to claim 5, characterized in that: The liquid chromatograph has a programmable control function and can automatically set and adjust chromatographic parameters such as flow rate, column temperature, detection wavelength, etc., to improve detection efficiency and accuracy.

7. The detection system according to claim 5 or 6, characterized in that: It also includes data processing and analysis software for automatically collecting chromatogram data, calculating peak areas, and automatically calculating the content of myristoylpropyl dimethylamine based on a standard curve, thereby reducing human errors and improving the reliability of analysis results.

8. The method or system according to any one of claims 1 to 7, characterized in that: The method and system are not only applicable to corneal contact lens care products, but also can be extended to the quality inspection of other cosmetics or daily chemical products containing myristoylpropyl dimethylamine.

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