Method for determining 10 antioxidants in medical device leachables

Abstract

The application discloses a kind of 10 kinds of antioxidants in medical device leachable determination method.The present application, by ultra-high performance liquid chromatography time-of-flight mass spectrometry technology realizes the simultaneous determination of ten kinds of antioxidants in medical device leachable, covers different types of antioxidants such as phenolic and phosphite.The pretreatment process only needs to extract the measured product, and the extracted liquid is blown dry by nitrogen, methanol is added, vortexed and filtered through microporous filter to complete the preparation of test solution, and the operation steps are simple, without complex derivatization or purification process, greatly reducing the difficulty and time cost of sample processing.In the analysis process, the ultra-high performance liquid chromatography system is used in conjunction with specific chromatographic column and mobile phase gradient elution program, which can quickly realize the baseline separation of ten kinds of antioxidants, combined with the high sensitivity detection of time-of-flight mass spectrometry, significantly improves the analysis efficiency, and is suitable for rapid screening and quantitative analysis of large quantities of samples.

Description

Technical Field

[0001] This invention belongs to the field of medical device testing technology, specifically a method for determining 10 antioxidants in extractable materials of medical devices. Background Technology

[0002] In the manufacturing process of medical devices, polymeric materials (such as polyethylene, polypropylene, and polyvinyl chloride) are widely used due to their excellent physicochemical properties. However, these materials are susceptible to oxidative degradation under the influence of heat, light, and oxygen during processing, sterilization (such as gamma irradiation and high-temperature steam), and long-term use, leading to a decline in material properties (such as brittleness, discoloration, and reduced mechanical strength) or the release of harmful substances. Therefore, antioxidants are often added to polymeric materials used in medical devices to slow down oxidation reactions and extend product lifespan.

[0003] Phenolic antioxidants, such as Antioxidant 1010 (pentaerythritol tetrakis[β-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate]), are primarily high molecular weight and resistant to extraction, making them suitable for long-term implantable devices. Antioxidant 1076 (octadecyl β-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate) has good compatibility with polyolefins and is widely used in medical catheters and packaging materials. Antioxidant 330 (1,3,5-trimethyl-2,4,6-tris(3,5-di-tert-butyl-4-hydroxybenzyl)benzene) is structurally stable and radiation-resistant, making it suitable for high-requirement medical devices. Antioxidant 2246 (2,2'-methylenebis(4-methyl-6-tert-butylphenol)) has the main advantage of low cost, but it has high migration properties and is mostly used for short-term contact devices. Phosphites are primarily used as auxiliary antioxidants. For example, antioxidant 168 (tris(2,4-di-tert-butylphenyl)phosphite) is often compounded with phenolic antioxidants to synergistically improve thermal stability. Propyl gallate (PG) is a natural derivative used in some biodegradable materials, but it is prone to migration. Antioxidant 1098 (N,N'-bis[3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionyl]hydrazine) is heat-resistant and suitable for engineering plastics such as nylon. Antioxidant TH-1790 is specifically designed for resistance to gamma radiation and is used in disposable sterilization devices. Antioxidant 565 is mainly suitable for post-processing stabilization of unsaturated rubbers and is very effective for elastomers, protecting materials from thermal oxidative degradation during production, processing, and end-use. Antioxidant 3114, due to its high molecular weight and melting point, has extremely low volatility, low migration, and good water extraction resistance, imparting excellent heat oxidation resistance and light oxidation resistance to plastics.

[0004] The safety evaluation of antioxidants uses the "Exposure Limit Ratio (ELR)" as the core assessment indicator, which quantifies the risk level by calculating the ratio of Actual Exposure Dose (AED) to Tolerable Intake (TI). Tolerable Intake (TI), a key toxicological parameter, is defined as the maximum dose threshold at which continuous exposure to a compound via a predetermined route of administration within a specific exposure period can occur without producing a No Observed Adverse Effect Level (NOAEL). When ELR ≥ 1, it indicates that the systemic exposure level of the antioxidant has exceeded the safety threshold, requiring the initiation of a risk assessment procedure. Therefore, quantitative investigation of antioxidants migrating from medical devices to the human body is necessary. Currently, the detection of various antioxidants in extractables from medical devices mainly relies on HPLC-UV, GC-MS, or triple quadrupole LC-MS / MS. However, Time-of-Flight Mass Spectrometry (TOF-MS) still suffers from problems in this field, such as incomplete antioxidant coverage, severe matrix interference, and insufficient quantitative accuracy.

[0005] Therefore, there is an urgent need for a new detection method that can achieve high sensitivity and high specificity in the detection of 10 antioxidants in medical devices, in order to meet the actual needs of medical device production and regulation. Summary of the Invention

[0006] The purpose of this invention is to provide a method for determining 10 antioxidants in extractable materials of medical devices in order to solve the problems mentioned above.

[0007] The technical solution adopted in this invention is as follows: a method for determining 10 antioxidants in medical device extracts, the method comprising the following steps:

[0008] S1: Instrument preparation: Ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometer, with a delay column added between the pump and the autosampler; adjust all parameters to operating conditions, use nitrogen as the collision gas; use a C18 column; column temperature 40℃.

[0009] S2: To prepare antioxidant stock solutions, accurately weigh 15.20 mg of antioxidant 565 standard, 12.02 mg of antioxidant 3114 standard, 1.33 mg of antioxidant 330 standard, 12.11 mg of antioxidant 168 standard, 10.28 mg of antioxidant 1076 standard, 14.25 mg of antioxidant 1010 standard, 12.48 mg of antioxidant TH-1790 standard, 10.59 mg of propyl gallate standard, 10.76 mg of antioxidant 1098 standard, and 15.22 mg of antioxidant 2246 standard. Dilute each standard stock solution to 10 mL with methanol or ethyl acetate to obtain 10 antioxidant standard stock solutions. Mix appropriate amounts of each standard stock solution and dilute to 10 mL with methanol to obtain a mixed standard stock solution with each antioxidant concentration of 10 μg / mL.

[0010] S3: Prepare linear solutions. Using methanol as the matrix, quantitatively dilute the mixed standard stock solution in S2 into seven standard working solutions with increasing concentrations. Test them using ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry. With the concentration of the mixed standard curve solution as the ordinate and the corresponding peak area as the abscissa, standard curves for ten antioxidants can be plotted separately.

[0011] S4: Preparation of detection limit solution: Prepare a 0.025 μg / mL mixed standard solution of ten antioxidants as the detection limit solution. The signal-to-noise ratio of the response value of this solution shall not be lower than 3:1; prepare a 0.05 μg / mL mixed standard solution of ten antioxidants as the quantitation limit solution. The signal-to-noise ratio of the response value of this solution shall not be lower than 10:1.

[0012] S5: Accuracy solutions are prepared by adding an appropriate amount of standard solution to the test sample solution to prepare accuracy solutions at three concentration levels: low, medium, and high. Three copies of each concentration are prepared in parallel.

[0013] S7: Repeatability solution: Take 9 portions of the accuracy solution to obtain the solution;

[0014] S8: Prepare the test solution. After extracting the test sample with water or ethanol, the extract is obtained. Take 10 ml of the extract and blow it dry with nitrogen. Add 1 mL of methanol, vortex for 1 min, filter through a 0.22 μm microporous membrane, and take the filtrate to obtain the test solution.

[0015] S9: The contents of ten antioxidants (antioxidant 565, antioxidant 3114, antioxidant 330, antioxidant 168, antioxidant 1076, antioxidant 1010, antioxidant TH-1790, propyl gallate, antioxidant 1098, and antioxidant 2246) in the test solution were determined by ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry and standard curve analysis for each antioxidant.

[0016] In a preferred embodiment, in step S1, the liquid chromatography conditions of the ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometer are as follows: the chromatographic column is C10 ... 18 Column: 3.0 mm × 100 mm, 1.7 μm; injection volume: 0.5 μL; flow rate: 0.4 mL / min.

[0017] In a preferred embodiment, in step S1, mobile phase A solution comprises: 4.5 mmol / L ammonium formate, 0.5 mmol / L ammonium fluoride, 0.1% formic acid, and purified water in solution A; mobile phase B solution comprises: 80% methanol, 20% isopropanol, 4.5 mmol / L ammonium formate, 0.5 mmol / L ammonium fluoride, 0.1% formic acid in positive mode, and 0.1% formic acid in negative mode in solution B.

[0018] In a preferred embodiment, in step S1, the mass spectrometry conditions of the ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometer are as follows: ionization mode is electrospray ionization positive ion mode ESI+, sheath gas temperature: 325℃, sheath gas flow rate: 8L / min, nebulizer temperature: 350℃, nebulizer gas flow rate: 11L / min, capillary voltage: 4000V for positive mode and 3500V for negative mode.

[0019] In a preferred embodiment, in step S1, the other mass spectrometry conditions for the ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometer are: antioxidant 565 molecular ion peak (M+H). + The m / Z is 589.4048, and the antioxidant 3114 molecular ion peak (M+NH4) is also present. + The m / Z is 801.5548, and the antioxidant has a molecular ion peak at 330 (M+NH4). + The m / Z is 792.6318, and the antioxidant 168 molecular ion peak (M+H) is also present. + The m / Z is 647.4612, and the antioxidant molecular ion peak (M+NH4) is at 1076. + The m / Z is 548.5075, and the peak of the antioxidant 1010 molecular ion (M+NH4) is also observed. + The m / Z is 1194.822, and the molecular ion peak (M+NH4) of the antioxidant TH-1790 is also observed. + The m / Z of the propyl gallate molecular ion peak (MH) is 717.4586, the m / Z of the antioxidant 1098 molecular ion peak (MH) is 211.0620, the m / Z of the antioxidant 2246 molecular ion peak (MH) is 635.4814, and the m / Z of the antioxidant 2246 molecular ion peak (MH) is 339.2371.

[0020] In a preferred embodiment, in step S3, the mixed standard stock solution is quantitatively diluted with methanol to form seven standard working solutions with increasing concentrations, each antioxidant having a concentration range of 0.050 μg / mL, 0.10 μg / mL, 0.20 μg / mL, 0.40 μg / mL, 0.60 μg / mL, 0.80 μg / mL, and 1.0 μg / mL.

[0021] In a preferred embodiment, in step S4, a mixed standard solution of ten antioxidants is prepared as the detection limit solution with a concentration of 0.020 μg / mL. This solution is measured using ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry (UHPLC-MS / MS) to ensure that the signal-to-noise ratio (SNR) of each antioxidant response value is not less than 3:1. A mixed standard solution of ten antioxidants with a concentration of 0.05 μg / mL is prepared as the quantitation limit solution, and this solution is also measured using the same instrument, requiring a SNR of not less than 10:1. The same liquid chromatography and mass spectrometry conditions as for the linear solution are used during the determination, including column specifications, mobile phase composition, flow rate, column temperature, ionization mode, and mass spectrometry parameters, to verify the correspondence between solution concentration and SNR, ensuring that the method detection limit and quantitation limit meet the detection requirements.

[0022] In a preferred embodiment, in step S5, an accuracy solution is prepared by spiking the sample. An appropriate amount of standard solution is added to the test sample solution to prepare accuracy solutions at low, medium, and high concentration levels. Three replicates are prepared for each concentration level. The test sample solution is prepared by extracting the test sample with water or ethanol, drying 10 mL of the extract under nitrogen, adding 1 mL of methanol, vortexing for 1 min, and filtering through a 0.22 μm microporous membrane to obtain the filtrate. During the spiking process, the amount of spiked at each level needs to be determined based on the linear range of the antioxidant and the expected detection concentration to ensure that the low, medium, and high concentrations cover different intervals from the limit of quantitation to the upper limit of the linear range. Strict consistency in operation is maintained during the parallel sample preparation process to evaluate the accuracy of the method.

[0023] In a preferred embodiment, in step S6, an appropriate amount of standard solution is added to the test sample solution to prepare accuracy solutions at three concentration levels: low, medium, and high. Three copies of each concentration are prepared in parallel.

[0024] In a preferred embodiment, in step S7, nine portions of the accuracy solution are taken to obtain the repeatability solution.

[0025] In summary, due to the adoption of the above technical solution, the beneficial effects of the present invention are:

[0026] This invention exhibits excellent precision and accuracy. Through optimized mass spectrometry conditions and standard curve plotting methods, the stability and reliability of the antioxidant determination results are ensured. The linear range covers multiple levels from low to high concentrations, meeting the detection needs of antioxidants with varying concentrations. Furthermore, the reasonable limits of detection and quantitation allow for the accurate determination of trace antioxidant components in extractable materials of medical devices, providing reliable technical support for assessing the migration risk of antioxidants in medical devices. This efficient and accurate detection method helps strengthen the quality control of medical devices and ensure the safety of clinical use. Attached Figure Description

[0027] Figure 1 This is a positive mode chromatogram of ten antioxidants in this invention.

[0028] Figure 2 The image shows the negative mode chromatograms of ten antioxidants in this invention.

[0029] Figure 3 This is the positive mode EIC diagram of ten antioxidants in this invention.

[0030] Figure 4 This is the EIC diagram of the negative modes of ten antioxidants in this invention.

[0031] Figure 5 This is a blank chromatogram of the sample in this invention.

[0032] Figure 6 This is a chromatogram of the sample test solution in this invention.

[0033] Figure 7 This is a schematic diagram of the process principle in this invention. Detailed Implementation

[0034] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention.

[0035] Reference Figure 1-7 A method for determining 10 antioxidants in a medical device extract, the method comprising the following steps:

[0036] S1: Instrument preparation: The ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometer is an Agilent 1290Ⅱ-6450, equipped with an electrospray ionization source ESI (Agilent Technologies (China) Co., Ltd.), and a delay column is added between the pump and the autosampler;

[0037] S2: Preparation of reference solutions: Accurately weigh 15.20 mg of antioxidant 565 standard, 12.02 mg of antioxidant 3114 standard, 1.33 mg of antioxidant 330 standard, 12.11 mg of antioxidant 168 standard, 10.28 mg of antioxidant 1076 standard, 14.25 mg of antioxidant 1010 standard, 12.48 mg of antioxidant TH-1790 standard, 10.59 mg of propyl gallate standard, 10.76 mg of antioxidant 1098 standard, and 15.22 mg of antioxidant 2246 standard. Dilute each standard to 10 mL with methanol or ethyl acetate to prepare ten standard stock solutions. Take an appropriate amount of each standard stock solution, mix them, and dilute to 10 mL with methanol to obtain a mixed standard stock solution with a concentration of 10 μg / mL.

[0038] S3: Preparation of standard working solutions. Using methanol as the base, the mixed standard stock solution in S2 was quantitatively diluted to prepare seven standard working solutions with increasing concentrations. The mass concentrations of the standard working solutions are shown in Table 1.

[0039] Table 1. Concentration of Standard Working Solution:

[0040]

[0041] The standard working solutions of each concentration were analyzed by ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry.

[0042] The liquid chromatography conditions for the determination and analysis were as follows: the chromatographic column was C100000. 18 Chromatographic column: 3.0 mm × 100 mm, 1.7 μm; injection volume: 0.5 μL; flow rate: 0.4 mL / min; mobile phase A solution: includes 4.5 mmol / L ammonium formate, 0.5 mmol / L ammonium fluoride, 0.1% formic acid, and purified water in solution A; mobile phase B solution: includes 80% methanol, 20% isopropanol, 4.5 mmol / L ammonium formate, 0.5 mmol / L ammonium fluoride, 0.1% formic acid in positive mode, and 0.1% formic acid in negative mode in solution B; the specific elution program is shown in Table 2.

[0043] Table 2. Mobile phase gradient elution program:

[0044]

[0045] The mass spectrometry conditions for the analysis were as follows: ionization mode: electrospray ionization (ESI+) positive ion mode and ESI- negative ion mode; sheath gas temperature: 325℃; sheath gas flow rate: 8 L / min; nebulizer temperature: 350℃; nebulizer gas flow rate: 11 L / min; capillary voltage: 4000 V for positive mode and 3500 V for negative mode; other mass spectrometry conditions are shown in Table 3.

[0046] Table 3 Qualitative and quantitative ions

[0047]

[0048] The tests were performed using ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry. With the concentration of the mixed standard working solution as the ordinate and the corresponding peak area as the abscissa, standard curves for the six antioxidants could be plotted.

[0049] S4: Prepare the test solution. After extracting the test sample with water or ethanol, the extract is obtained. Take 10 ml of the extract and blow it dry with nitrogen. Add 1 mL of methanol, vortex for 1 min, filter through a 0.22 μm microporous membrane, and take the filtrate to obtain the test solution.

[0050] S5: The contents of ten antioxidants (antioxidant 565, antioxidant 3114, antioxidant 330, antioxidant 168, antioxidant 1076, antioxidant 1010, antioxidant TH-1790, propyl gallate, antioxidant 1098, and antioxidant 2246) in the test sample solution were determined by ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry (UHPLC-Q-TOF) and concentration standard curves of each standard stock solution.

[0051] The contents of various antioxidant components in the test sample are calculated as follows:

[0052] W=C×V / 10

[0053] In the formula:

[0054] W represents the content of each antioxidant component in the test sample, expressed in micrograms per device (μg / instrument).

[0055] C represents the content of each antioxidant component in the test solution obtained from the concentration standard curve, expressed in micrograms per milliliter (μg / mL).

[0056] V represents the volume of the extract, expressed in milliliters (mL).

[0057] The method for determining ten antioxidants in the extractable material of the medical device described in this invention was validated:

[0058] Limits of detection, limits of quantitation, and linear range: Using the method described in this invention and according to the method in 5.4.2.2 of GB / T27417-2017, the measurement signals of a known low-concentration analyte sample and a blank sample were compared. The concentration with a signal-to-noise ratio greater than or equal to 3:1 was used as the limit of detection, and the concentration with a signal-to-noise ratio greater than or equal to 10:1 was used as the limit of quantitation. The results are shown in Table 4.

[0059] Table 4 Results of Limit of Detection and Limit of Quantification

[0060]

[0061] Using the method described in this invention, with methanol as the base, the standard working solution curve equations for ten antioxidants are shown in Table 5:

[0062] Table 5. Standard working solution curve equations for six antioxidants

[0063]

[0064] As shown in Table 5, the 10 antioxidants exhibited good linearity in the range of 0.05 μg / mL to 1.0 μg / mL (using methanol as the matrix and standard preparation curves), with correlation coefficients R all greater than 0.99.

[0065] Spiking recovery test: Blank spiking tests were conducted using standard substances. Three groups of samples were taken, with three parallel samples in each group. The amount of standard substance added to each sample is shown in Tables 6-8. The spiking recovery rate (%) and RSD were calculated. The results are shown in Tables 6-8.

[0066] Table 6. Spike recoveries of ten antioxidants (1)

[0067]

[0068] Table 7. Spike recoveries of ten antioxidants (2)

[0069]

[0070] Table 8. Spike recoveries of ten antioxidants (3)

[0071]

[0072] As shown in Tables 6-8, the spiked recoveries were all between 88.55% and 110.25%, indicating that the method and instrument have good accuracy and can meet the testing requirements.

[0073] From the above, we can conclude that:

[0074] This invention utilizes ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry (UHPLC-TSF-MS / MS) to simultaneously determine ten antioxidants in extractable materials from medical devices, covering different types of antioxidants such as phenols and phosphites. The pretreatment process is simple: extract the sample, dry the extract with nitrogen, add methanol, vortex, and filter through a microporous membrane to prepare the test solution. The operation is straightforward, eliminating the need for complex derivatization or purification processes, significantly reducing the difficulty and time cost of sample processing. The UHPLC system used in the analysis, combined with a specific column and gradient elution program, enables rapid baseline separation of the ten antioxidants. Combined with the high sensitivity of time-of-flight mass spectrometry, this significantly improves analytical efficiency and is suitable for rapid screening and quantitative analysis of large batches of samples.

[0075] This invention exhibits excellent precision and accuracy. Through optimized mass spectrometry conditions and standard curve plotting methods, the stability and reliability of the antioxidant determination results are ensured. The linear range covers multiple levels from low to high concentrations, meeting the detection needs of antioxidants with varying concentrations. Furthermore, the reasonable limits of detection and quantitation allow for the accurate determination of trace antioxidant components in extractable materials of medical devices, providing reliable technical support for assessing the migration risk of antioxidants in medical devices. This efficient and accurate detection method helps strengthen the quality control of medical devices and ensure the safety of clinical use.

[0076] It should be noted that, in this document, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitations, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0077] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit it. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for determining 10 antioxidants in a medical device extract, characterized in that: The method includes the following steps: S1: Instrument preparation: Ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometer, with a delay column added between the pump and the autosampler; adjust all parameters to operating conditions, use nitrogen as the collision gas; use a C18 column; column temperature 40℃. S2: To prepare antioxidant stock solutions, accurately weigh 15.20 mg of antioxidant 565 standard, 12.02 mg of antioxidant 3114 standard, 1.33 mg of antioxidant 330 standard, 12.11 mg of antioxidant 168 standard, 10.28 mg of antioxidant 1076 standard, 14.25 mg of antioxidant 1010 standard, 12.48 mg of antioxidant TH-1790 standard, 10.59 mg of propyl gallate standard, 10.76 mg of antioxidant 1098 standard, and 15.22 mg of antioxidant 2246 standard. Dilute each standard stock solution to 10 mL with methanol or ethyl acetate to obtain 10 antioxidant standard stock solutions. Mix appropriate amounts of each standard stock solution and dilute to 10 mL with methanol to obtain a mixed standard stock solution with each antioxidant concentration of 10 μg / mL. S3: Prepare linear solutions. Using methanol as the matrix, quantitatively dilute the mixed standard stock solution in S2 into seven standard working solutions with increasing concentrations. Test them using ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry. With the concentration of the mixed standard curve solution as the ordinate and the corresponding peak area as the abscissa, standard curves for ten antioxidants can be plotted separately. S4: Preparation of detection limit solution: Prepare a 0.020 μg / mL mixed standard solution of ten antioxidants as the detection limit solution. The signal-to-noise ratio of the response value of this solution shall not be lower than 3:1; Prepare a 0.050 μg / mL mixed standard solution of ten antioxidants as the quantitation limit solution. The signal-to-noise ratio of the response value of this solution shall not be lower than 10:

1. S5: Accuracy solutions are prepared by adding an appropriate amount of standard solution to the test sample solution to prepare accuracy solutions at three concentration levels: low, medium, and high. Three replicates of each concentration are prepared. S7: Repeatability solution: Take 9 portions of the accuracy solution to obtain the solution; S8: Prepare the test solution. After extracting the test sample with water or ethanol, the extract is obtained. Take 10 ml of the extract and blow it dry with nitrogen. Add 1 mL of methanol, vortex for 1 min, filter through a 0.22 μm microporous membrane, and take the filtrate to obtain the test solution. S9: The contents of ten antioxidants (antioxidant 565, antioxidant 3114, antioxidant 330, antioxidant 168, antioxidant 1076, antioxidant 1010, antioxidant TH-1790, propyl gallate, antioxidant 1098, and antioxidant 2246) in the test solution were determined by ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry and standard curve analysis for each antioxidant.

2. The method for determining 10 antioxidants in a medical device extract as described in claim 1, characterized in that: In step S1, the liquid chromatography conditions of the ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometer are as follows: the chromatographic column is C10 ... 18 Column: 3.0 mm × 100 mm, 1.7 μm; injection volume: 0.5 μL; flow rate: 0.4 mL / min.

3. The method for determining 10 antioxidants in a medical device extract as described in claim 1, characterized in that: In step S1, mobile phase A solution comprises 4.5 mmol / L ammonium formate, 0.5 mmol / L ammonium fluoride, 0.1% formic acid, and purified water in solution A; mobile phase B solution comprises 80% methanol, 20% isopropanol, 4.5 mmol / L ammonium formate, 0.5 mmol / L ammonium fluoride, 0.1% formic acid in positive mode, and 0.1% formic acid in negative mode in solution B.

4. The method for determining 10 antioxidants in a medical device extract as described in claim 1, characterized in that: In step S1, the mass spectrometry conditions of the ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometer are as follows: ionization mode is electrospray ionization positive ion mode ESI+, sheath gas temperature: 325℃, sheath gas flow rate: 8L / min, nebulizer temperature: 350℃, nebulizer gas flow rate: 11L / min, capillary voltage: 4000V for positive mode and 3500V for negative mode.

5. The method for determining 10 antioxidants in a medical device extract as described in claim 1, characterized in that: In step S1, the other mass spectrometry conditions for the ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometer are: antioxidant 565 molecular ion peak (M+H). + The m / Z is 589.4048, and the antioxidant 3114 molecular ion peak (M+NH4) is also present. + The m / Z is 801.5548, and the antioxidant has a molecular ion peak at 330 (M+NH4). + The m / Z is 792.6318, and the antioxidant 168 molecular ion peak (M+H) is also present. + The m / Z is 647.4612, and the antioxidant molecular ion peak (M+NH4) is at 1076. + The m / Z is 548.5075, and the peak of the antioxidant 1010 molecular ion (M+NH4) is also observed. + The m / Z is 1194.822, and the molecular ion peak (M+NH4) of the antioxidant TH-1790 is also observed. + The m / Z of the propyl gallate molecular ion peak (MH) is 717.4586, the m / Z of the antioxidant 1098 molecular ion peak (MH) is 211.0620, the m / Z of the antioxidant 2246 molecular ion peak (MH) is 635.4814, and the m / Z of the antioxidant 2246 molecular ion peak (MH) is 339.2371.

6. The method for determining 10 antioxidants in a medical device extract as described in claim 1, characterized in that: In step S3, using methanol as a matrix, the mixed standard stock solution is quantitatively diluted into seven standard working solutions with increasing concentrations. The concentration ranges of each antioxidant are 0.050 μg / mL, 0.10 μg / mL, 0.20 μg / mL, 0.40 μg / mL, 0.60 μg / mL, 0.80 μg / mL, and 1.0 μg / mL.

7. The method for determining 10 antioxidants in a medical device extract as described in claim 1, characterized in that: In step S4, a mixed standard solution of ten antioxidants is prepared as the detection limit solution with a concentration of 0.020 μg / mL. This solution is measured using ultra-high performance liquid chromatography-tandem time-of-flight mass spectrometry (UHPLC-MS / MS) to ensure that the signal-to-noise ratio (SNR) of each antioxidant response is not less than 3:

1. A mixed standard solution of ten antioxidants with a concentration of 0.050 μg / mL is prepared as the quantitation limit solution, and this solution is also measured using the same instrument, requiring a SNR of not less than 10:

1. The same liquid chromatography and mass spectrometry conditions as for the linear solution are used during the measurement, including column specifications, mobile phase composition, flow rate, column temperature, ionization mode, and mass spectrometry parameters, to verify the correlation between solution concentration and SNR, ensuring that the method detection limit and quantitation limit meet the detection requirements.

8. The method for determining 10 antioxidants in a medical device extract as described in claim 1, characterized in that: In step S5, an accuracy solution is prepared using a blank spiking method. An appropriate amount of standard solution is added to the test sample solution to prepare accuracy solutions at low, medium, and high concentration levels. Three replicates are prepared for each concentration level. The test sample solution is prepared by extracting the test sample with water or ethanol, then drying 10 mL of the extract under nitrogen, adding 1 mL of methanol, vortexing for 1 min, and filtering through a 0.22 μm microporous membrane to obtain the filtrate. During the spiking process, the spiking amount for each level needs to be determined based on the linear range of the antioxidant and the expected detection concentration to ensure that the low, medium, and high concentrations cover different intervals from the limit of quantitation to the upper limit of the linear range. Strict consistency in operation is maintained during the parallel sample preparation process to evaluate the accuracy of the method.

9. The method for determining 10 antioxidants in a medical device extract as described in claim 1, characterized in that: In step S6, a blank spiking method is used to add an appropriate amount of standard solution to the test sample solution to prepare accuracy solutions at three concentration levels: low, medium, and high. Three copies of each concentration are prepared in parallel.

10. The method for determining 10 antioxidants in a medical device extract as described in claim 1, characterized in that: In step S7, nine portions of the accuracy solution are taken to obtain the repeatability solution.

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