A GC quantitative detection method for polyoxyethylene (35) castor oil in microemulsion extract and its application

Abstract

The present invention proposes a GC quantitative detection method for polyoxyethylene (35) castor oil in a microemulsion extract, obtains the ratio between CrEL and ricinoleic acid methyl ester through research, then determines the content of ricinoleic acid methyl ester in the microemulsion extract to be tested, and then calculates the content of CrEL in the microemulsion extract to be tested according to the ratio between the two, thereby realizing the quantitative determination of CrEL in the microemulsion extract. The method has good repeatability and high accuracy, can quantitatively determine the CrEL in the microemulsion extract of Zexie decoction, and has important significance for the determination of the residual amount of CrEL in the effective components of Zexie decoction separated and purified in the later stage.

Description

Technical Field

[0001] The invention belongs to the technical field of chemical testing, and specifically relates to a GC quantitative detection method for polyoxyethylene (35) castor oil in a microemulsion extract and an application thereof. Background Art

[0002] With the development of synthetic chemistry, surfactants with different properties have been developed and widely used in the pharmaceutical industry. Surfactants are composed of two groups with different properties, hydrophilic and lipophilic, and are often used in formulation molding as emulsifiers, solubilizers, wetting agents, cosolvents and penetrants. There are many types of surfactants, among which non-ionic surfactants are widely used in oral pharmaceutical preparations, injections and topical preparations due to their excellent properties such as high stability, good compatibility and low toxicity. Commonly used non-ionic surfactants in the pharmaceutical industry include alkyl polyoxyethylene ethers, sorbitan fatty acid esters and alkyl glucosides.

[0003] Polyoxyethylene (35) castor oil (hereinafter referred to as CrEL) is a commonly used alkyl polyoxyethylene ether nonionic surfactant. It is a complex mixture of ethylene oxide and glycerol castor oil in a molar ratio of 35:1. Its main chemical component is polyoxyethylene triricinoleate (structural formula see Figure 1a ) and polyethylene glycol ricinoleate (structural formula see Figure 1b ). CrEL is generally considered to be relatively non-toxic, but studies have shown that CrEL is the root cause of severe hypersensitivity reactions to the anti-tumor drug paclitaxel. In addition, this type of surfactant can promote the release of histamine in the human body during metabolism, causing adverse reactions such as neurotoxicity, cardiotoxicity and toxic kidney damage. At present, research on CrEL is mostly focused on component analysis and application, and there is a lack of suitable quantitative analysis methods for quality control. Although the pre-column derivatization method and liquid chromatography / tandem mass spectrometry reported in the literature have high sensitivity, they can only quantify the ricinoleic acid and polyethylene glycol compounds present in CrEL, respectively, and cannot achieve the overall quantification of the CrEL mixture. Although the colorimetric method and titration method can roughly quantify the mixture as a whole, the sensitivity and feasibility of the overall experiment are low.

[0004] Therefore, it is urgent to establish a new method with high sensitivity and quantitative detection of CrEL, in order to provide a stable and feasible detection method for the removal of surfactants in microemulsion extracts. Summary of the invention

[0005] In order to solve the above problems in the prior art, the present invention provides a GC quantitative detection method and application of polyoxyethylene (35) castor oil in microemulsion extract.

[0006] In the first aspect, the present invention proposes a GC quantitative detection method for polyoxyethylene (35) castor oil in a microemulsion extract. The ratio relationship between CrEL and ricinoleic acid methyl ester is obtained by research, and then the content of ricinoleic acid methyl ester in the microemulsion extract to be tested is determined. Then, the content of CrEL in the microemulsion extract to be tested is calculated based on the ratio relationship between the two, thereby achieving the quantification of CrEL in the microemulsion extract.

[0007] According to the present invention, the chemical structure of the compound of CrEL contains a ricinoleic acid ester-like structure, and ricinoleic acid ester can undergo alcoholysis with methanol under the catalysis of a base to generate methyl ricinoleate (structural formula: Figure 1c ). Methyl ricinoleate is the content determination index of castor oil in the 2020 edition of the Chinese Pharmacopoeia. Firstly, CrEL was taken as the research object to obtain the proportional relationship between CrEL and methyl ricinoleate. The drug preparation containing CrEL was treated with the same sample preparation method to determine the content of methyl ricinoleate. According to the proportional relationship between the two, the content of CrEL in the sample was calculated to achieve the quantification of CrEL.

[0008] According to the present invention, the microemulsion extract is prepared from ethyl oleate, CrEL, and 50% ethanol as raw materials to prepare a stable O / W type microemulsion, and used as an extraction solvent for extracting Zexie decoction to obtain a microemulsion extract. Using microemulsion as an extraction solvent can improve the extraction efficiency of effective components in Zexie decoction to a certain extent, especially the most lipophilic Zexie alcohol B and 23-acetyl Zexie alcohol B, which are increased by 6.4 times and 5.5 times respectively, but a certain amount of CrEL exists in the Zexie decoction liquid extracted by microemulsion, so the present invention establishes a quantitative detection method of CrEL, which can provide a method for removing CrEL in the microemulsion extract, and provide a reference for the quality control of other pharmaceutical preparations containing CrEL.

[0009] As a specific embodiment of the present invention, the following steps are included:

[0010] S1: Mix the quantitative CrEL with potassium hydroxide-methanol solution and heat it under reflux, then add boron trifluoride ether-methanol solution and heat it under reflux, extract, and obtain the research sample solution;

[0011] S2: Dehydrate and filter the sample solution to test the peak area of ​​ricinoleic acid methyl ester;

[0012] S3: repeat steps S1 and S2 to obtain the average content ratio of ricinoleic acid methyl ester in CrEL;

[0013] S4: mixing the microemulsion extract to be tested with a potassium hydroxide-methanol solution and heating under reflux, then adding a boron trifluoride ether-methanol solution and heating under reflux for extraction to obtain a sample solution to be tested;

[0014] S5: dehydrating and filtering the sample solution to be tested to test the peak area of ​​ricinoleic acid methyl ester to obtain the content of ricinoleic acid methyl ester;

[0015] S6: The content of CrEL in the microemulsion extract to be tested is calculated by using the mean value of the content ratio of ricinoleic acid methyl ester in CrEL obtained in step S3.

[0016] As a specific embodiment of the present invention, in the step S1 and step S4, independently, the concentration of potassium hydroxide in the methanol solution in the potassium hydroxide-methanol solution is 0.25-1.5 mol / L, preferably 1 mol / L; the methanol solution is pure methanol;

[0017] The mass volume ratio of the CrEL to the potassium hydroxide-methanol solution is 1 g:(50-150) ml, preferably 1 g:100 ml.

[0018] As a specific embodiment of the present invention, in the step S1 and the step S4, independently, the mass ratio of boron trifluoride etherate to methanol in the boron trifluoride etherate-methanol solution is 1:(2-4), preferably 1:3;

[0019] The mass volume ratio of the CrEL to the boron trifluoride ether-methanol solution is 1 g:(50-250) ml, preferably 1 g:100 ml.

[0020] As a specific embodiment of the present invention, in the step S1 and step S4, independently, the heating temperature is 50-70° C., preferably 60° C., and the heating method is preferably water bath heating;

[0021] As a specific embodiment of the present invention, in step S1 and step S4, independently, the reflux time is 15-60 min, preferably 15 min.

[0022] As a specific embodiment of the present invention, in the step S1 and the step S4, independently, the extraction comprises extracting with n-hexane for 1-4 times, preferably 2 times;

[0023] The mass volume ratio of CrEL to n-hexane is 1 g:(50-150) ml, preferably 1 g:100 ml;

[0024] Preferably, saturated sodium chloride is added during the first extraction, and the amount added is 2-5 times, preferably 3.5 times, of n-hexane.

[0025] As a specific embodiment of the present invention, in the step S2 and the step S5, independently, the dehydration includes dehydration using anhydrous sodium sulfate; the filtration includes filtration using a filter membrane, and preferably the pore size of the filter membrane is 0.22 μm.

[0026] As a specific implementation of the present invention, in the step S3, the average content ratio of methyl ricinoleate in CrEL is 0.2084.

[0027] As a specific embodiment of the present invention, in the step S4, in actual implementation, after the microemulsion extract to be tested is evaporated and dried, the residue is mixed with potassium hydroxide-methanol solution and heated to reflux.

[0028] As a specific implementation of the present invention, in step S6, the CrEL content = the ricinoleic acid methyl ester content obtained in step S5 / 0.2084.

[0029] As a specific embodiment of the present invention, the test of ricinoleic acid methyl ester is preferably carried out by gas chromatography, and the test conditions include: using -WAX chromatographic column (30m×0.25mm×0.25μm); detector FID, column temperature programmed to 190-240℃, starting temperature 190℃, rising to 240℃ at 10℃ / min, maintaining for 20min, carrier gas is high-purity nitrogen; carrier gas flow rate 1ml / min; detector temperature 250℃; injection port temperature 220℃; injection volume 1μl; split ratio 5:1.

[0030] The above raw materials in the present invention can be prepared in-house or commercially available, and the present invention is not particularly limited thereto.

[0031] In a second aspect, the present invention provides application of the detection method described in the first aspect in the field of CrEL removal.

[0032] Compared with the prior art, the present invention has the following beneficial effects:

[0033] The GC quantitative detection method of polyoxyethylene (35) castor oil in a microemulsion extract of the present invention obtains the ratio relationship between CrEL and ricinoleic acid methyl ester through research, then determines the content of ricinoleic acid methyl ester in the microemulsion extract to be tested, and then calculates the content of CrEL in the microemulsion extract to be tested according to the ratio relationship between the two, thereby realizing the quantification of CrEL in the microemulsion extract. The method is stable and feasible.

[0034] The GC quantitative detection method of polyoxyethylene (35) castor oil in the microemulsion extract of the present invention has good repeatability and high accuracy, and can quantify CrEL in the microemulsion extract of Zexie decoction, which is of great significance for the determination of the residual amount of CrEL in the effective components of Zexie decoction separated and purified in the later stage. CrEL is often used as a solubilizer and emulsifier for poorly soluble drugs, and is used in large amounts, which is an important cause of adverse reactions in preparations. The CrEL detection method established in this study can quantify CrEL in preparations, improve the safety of its clinical application from the perspective of product quality control, and provide a detection method for feasibility analysis of microemulsion as an extraction solvent. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1a It is the structural formula of polyoxyethylene glyceryl triricinoleate;

[0036] Figure 1b It is the structural formula of polyethylene glycol ricinoleate;

[0037] Figure 1c is the structural formula of methyl ricinoleate;

[0038] Figure 2 This is an experimental flow chart of the production of methyl ricinoleate by CrEL in Example 1 of the present invention;

[0039] Figure 3A is the gas chromatogram of the reference substance in Example 1 of the present invention;

[0040] Figure 3B is the gas chromatogram of CrEL in Example 1 of the present invention;

[0041] Figure 3C It is a gas chromatogram of the tested Zexie decoction microemulsion extract sample in Example 1 of the present invention.

[0042] Among them, 1-ricinoleic acid methyl ester. DETAILED DESCRIPTION

[0043] The present invention will be further described below in conjunction with specific embodiments, but they do not constitute any limitation to the present invention.

[0044] The specific information of the experimental equipment used in each embodiment of the present invention is as follows:

[0045] 7890A gas chromatograph, Agilent, USA;

[0046] BD125 1 / 100,000 electronic analytical balance, Sartorius Scientific Instruments (Beijing) Co., Ltd.;

[0047] 1011015 constant temperature water bath, Gongyi Yuhua Instrument Co., Ltd.;

[0048] JY2002 electronic balance, Shanghai Sunyu Hengping Scientific Instrument Co., Ltd.;

[0049] 98-1-B electronic temperature control heating jacket, Tianjin Test Instrument Co., Ltd.;

[0050] WB2000-D agitator, WIGGENS, Germany.

[0051] The specific information of the reagents used in the embodiments of the present invention is as follows:

[0052] Ricinoleic acid methyl ester, purity ≥99%, batch number H2112152, purchased from Aladdin Reagent Co., Ltd.;

[0053] n-Hexane, analytical grade, Beijing Mairuida Technology Co., Ltd.;

[0054] Boron trifluoride ethyl etherate, analytical grade, Beijing Mairuida Technology Co., Ltd.;

[0055] Potassium hydroxide, analytical grade, from Tianjin Damao Reagent Factory;

[0056] Sodium chloride, analytical grade, Beijing Mairuida Technology Co., Ltd.;

[0057] Methanol, analytical grade, Sinopharm Chemical Reagent Co., Ltd.;

[0058] Ethyl oleate, Beijing Fengli Jingqiu Trading Co., Ltd., batch number 0000881168;

[0059] Polyoxyethylene (35) castor oil CrEL, Beijing Fengli Jingqiu Trading Co., Ltd., batch number 14026224UO;

[0060] The medicinal materials of Alisma and Atractylodes were purchased from Sichuan Xinhehua Chinese Medicine Pieces Co., Ltd. and were identified by He Xirong from the Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences as the dried tubers of Alisma plantago-aquatica Linn., a plant of the Alismataceae family, and the dried rhizomes of Atractylodes macrocephala Koidz., a plant of the Compositae family.

[0061] Preparation of blank microemulsion

[0062] Accurately weigh 5.17 g of ethyl oleate, 29.06 g of CrEL, and 10.76 g of 50% ethanol into a beaker, stir, add deionized water to dilute to 200 mL, and stir at 600 r / min. -1 The mixture was stirred for 30 min, and the stirred liquid was divided into three equal parts. Deionized water was added to dilute the mixture to 966, 773, and 773 mL, which were used as solvents for the first, second, and third heating reflux extractions, respectively.

[0063] Test sample: Preparation of Zexie decoction microemulsion extract

[0064] Weigh 69 g of Alisma orientalis and 27.6 g of Atractylodes macrocephala and place them in a 2000 mL round-bottom flask, add blank microemulsion solution, heat under reflux and extract for 1, 0.5, and 0.5 h respectively, combine the extracts and let cool to obtain the Alisma orientalis decoction microemulsion extract.

[0065] The GC quantitative detection method of polyoxyethylene (35) castor oil in the microemulsion extract of the present invention comprises the following steps:

[0066] S1: Mix the quantitative CrEL with potassium hydroxide-methanol solution and heat it under reflux, then add boron trifluoride ether-methanol solution and heat it under reflux, extract, and obtain the research sample solution;

[0067] S2: Dehydrate and filter the sample solution to test the peak area of ​​ricinoleic acid methyl ester;

[0068] S3: repeat steps S1 and S2 to obtain the average content ratio of ricinoleic acid methyl ester in CrEL;

[0069] S4: After the microemulsion extract to be tested is evaporated and dried, the residue is mixed with a potassium hydroxide-methanol solution and heated under reflux, and then a boron trifluoride ether-methanol solution is added and heated under reflux for extraction to obtain a sample solution to be tested;

[0070] S5: dehydrating and filtering the sample solution to be tested to test the peak area of ​​ricinoleic acid methyl ester to obtain the content of ricinoleic acid methyl ester;

[0071] S6: The content of CrEL in the microemulsion extract to be tested is calculated by using the mean value of the content ratio of ricinoleic acid methyl ester in CrEL obtained in step S3.

[0072] Among them, the gas chromatograph test was used, and the test conditions included: -WAX chromatographic column (30m×0.25mm×0.25μm); detector FID, column temperature programmed to 190-240°C, starting temperature 190°C, rising to 240°C at 10°C / min, maintaining for 20min, carrier gas is high-purity nitrogen; carrier gas flow rate 1mL / min; detector temperature 250°C; injection port temperature 220°C; injection volume 1μL; split ratio 5:1.

[0073] Example 1

[0074] This example provides a GC quantitative detection method for polyoxyethylene (35) castor oil in a microemulsion extract, wherein CrEL is used as a research object to obtain the ratio between CrEL and ricinoleic acid methyl ester. The specific details are as follows:

[0075] About 0.1 g of CrEL was accurately weighed in a conical flask, and the effects of the concentration, dosage, and 60°C water bath heating reflux time of potassium hydroxide-methanol solution, the dosage of boron trifluoride ether-methanol (1:3) solution, 60°C water bath heating reflux time, and the number of n-hexane extractions on the formation of ricinoleic acid methyl ester were investigated respectively as shown in Table 1. The experimental flow chart is shown in Figure 2 The extracted samples were combined into a 50 ml volumetric flask, and n-hexane was added to the scale, shaken, dehydrated with 2 g of anhydrous sodium sulfate, shaken, and filtered through a 0.22 μm filter membrane.

[0076] Accurately inject 1 μl into the gas chromatograph and record the peak area of ​​ricinoleic acid methyl ester.

[0077] Table 1 Single factor investigation results of the preparation method of ricinoleic acid methyl ester generated by CrEL

[0078]

[0079] The best preparation method finally determined in Example 1 is to prepare 10 mL of 1 mol / l potassium hydroxide-methanol solution and heat it in a water bath at 60°C for 15 min, prepare 10 mL of boron trifluoride ether-methanol (1:3) solution and heat it in a water bath at 60°C for 15 min, and extract with n-hexane twice.

[0080] The average percentage of methyl ricinoleate generated by CrEL in Example 1 was 20.84% ​​and the RSD value was 2.55%, indicating that the optimal preparation method had good reproducibility and was stable and feasible.

[0081] Example 2

[0082] This example provides a GC quantitative detection method for polyoxyethylene (35) castor oil in a microemulsion extract, and the specific details are as follows:

[0083] S1: Accurately pipette 10 ml of the test sample Zexie decoction microemulsion extract into an evaporating dish, evaporate to dryness, dissolve with 10 ml of 1 mol / l potassium hydroxide-methanol solution and transfer all of it into a conical flask, heat and reflux in a 60°C water bath for 15 min, accurately add 10 ml of boron trifluoride ether-methanol (1:3) along the wall of the condenser tube, heat and reflux in a 60°C water bath for 15 min; accurately add 10 ml of n-hexane into the conical flask, shake, cool to room temperature, add 10 ml of saturated sodium chloride solution into the conical flask, and reflux for 15 min. bottle, shake, and transfer to a separatory funnel, add 10, 10, and 5 ml of saturated sodium chloride solution to wash the conical flask in sequence, transfer all to a separatory funnel, shake, and let stand for 15 minutes; transfer all the supernatant to a 50 ml volumetric flask, add 10 ml of n-hexane to the lower layer for the second extraction, and transfer all the supernatant to a 50 ml volumetric flask; use n-hexane to make up the volume of the 50 ml volumetric flask to the scale, shake well, add 2 g of anhydrous sodium sulfate to dehydrate, shake, and pass through a 0.22 μm filter membrane to obtain the sample to be tested.

[0084] S2: Adoption -WAX chromatographic column (30m×0.25mm×0.25μm); detector FID, column temperature programmed to 190~240℃, starting temperature 190℃, at 10℃·min -1 Raise to 240℃, maintain for 20min, high purity nitrogen as carrier gas; carrier gas flow rate 1ml / ·min; detector temperature 250℃; injection port temperature 220℃; injection volume 1μl; split ratio 5:1; test the peak area of ​​methyl ricinoleate, such as Figure 3C As shown, the content of ricinoleic acid methyl ester is obtained;

[0085] S3: According to the average value of the percentage of ricinoleic acid methyl ester in CrEL obtained in Example 1, which is 20.84%, the CrEL content of the test sample Zexie decoction microemulsion extract is calculated to be 29.054g.

[0086] Embodiment 3-4

[0087] According to the steps of Example 2, the test sample was tested again. Example 3 measured the CrEL content of the test sample Zexie decoction microemulsion extract to be 30.495 g. Example 4 measured the CrEL content of the test sample Zexie decoction microemulsion extract to be 29.764 g.

[0088] The RSD value of the test result error obtained in Example 2-4 is 2.42%, indicating that the verification is qualified, indicating that the detection method of the present invention is stable and feasible.

[0089] In summary, the GC quantitative detection method of polyoxyethylene (35) castor oil in the microemulsion extract of the present invention obtains the proportional relationship between CrEL and ricinoleic acid methyl ester by studying, then determines the content of ricinoleic acid methyl ester in the microemulsion extract to be tested, and then calculates the content of CrEL in the microemulsion extract to be tested according to the proportional relationship between the two, thereby realizing the quantification of CrEL in the microemulsion extract. The method is stable, feasible, has good repeatability and high accuracy.

[0090] Any numerical value mentioned in the present invention, if there is only an interval of two units between any minimum value and any maximum value, includes all values ​​from the minimum value to the maximum value each time increasing by one unit. For example, if the amount of a component, or the value of a process variable such as temperature, pressure, time, etc. is declared to be 50-90, in this specification it means that 51-89, 52-88... and 69-71 and 70-71 are specifically listed. For non-integer values, 0.1, 0.01, 0.001 or 0.0001 can be appropriately considered as a unit. These are just some specially specified examples. In this application, in a similar manner, all possible combinations of numerical values ​​between the listed minimum and maximum values ​​are considered to have been disclosed.

[0091] It should be noted that the embodiments described above are only used to explain the present invention and do not constitute any limitation to the present invention. The present invention has been described with reference to typical embodiments, but it should be understood that the words used therein are descriptive and explanatory words, rather than restrictive words. The present invention may be modified as specified within the scope of the claims of the present invention, and the present invention may be revised without departing from the scope and spirit of the present invention. Although the present invention described therein relates to specific methods, materials and embodiments, it does not mean that the present invention is limited to the specific examples disclosed therein, on the contrary, the present invention can be extended to all other methods and applications with the same functions.

Claims

1. A GC quantitative detection method for polyoxyethylene 35 castor oil CrEL in microemulsion extract, characterized in that: The ratio between CrEL and methyl ricinoleate is obtained by studying, and then the content of methyl ricinoleate in the microemulsion extract to be tested is determined, and then the content of CrEL in the microemulsion extract to be tested is calculated according to the ratio between the two, so as to achieve the quantification of CrEL in the microemulsion extract; The following steps are involved: S1: Mix the quantitative CrEL with potassium hydroxide-methanol solution, heat and reflux, then add boron trifluoride ether-methanol solution, heat and reflux, extract, and obtain the research sample solution; the concentration of potassium hydroxide in the methanol solution is 1 mol / L; the methanol solution is pure methanol; the mass volume ratio of the CrEL to the potassium hydroxide-methanol solution is 1g:100ml; the mass ratio of boron trifluoride ether to methanol in the boron trifluoride ether-methanol solution is 1:3; the mass volume ratio of the CrEL to the boron trifluoride ether-methanol solution is 1g:100ml; S2: Dehydrate and filter the sample solution to test the peak area of ​​ricinoleic acid methyl ester; S3: repeating steps S1 and S2 to obtain the average content ratio of ricinoleic acid methyl ester in CrEL; the average content ratio of ricinoleic acid methyl ester in CrEL is 0.2084; S4: Mix the microemulsion extract to be tested with potassium hydroxide-methanol solution, heat and reflux, then add boron trifluoride ether-methanol solution, heat and reflux, extract, and obtain the sample solution to be tested; the concentration of potassium hydroxide in the methanol solution is 1 mol / L; the methanol solution is pure methanol; the mass volume ratio of CrEL to potassium hydroxide-methanol solution is 1g:100ml; the mass ratio of boron trifluoride ether to methanol in the boron trifluoride ether-methanol solution is 1:3; the mass volume ratio of CrEL to boron trifluoride ether-methanol solution is 1g:100ml; S5: dehydrating and filtering the sample solution to be tested to test the peak area of ​​ricinoleic acid methyl ester to obtain the content of ricinoleic acid methyl ester; S6: The content of CrEL in the microemulsion extract to be tested is calculated by using the mean value of the content ratio of ricinoleic acid methyl ester in CrEL obtained in step S3; the CrEL content = the ricinoleic acid methyl ester content obtained in step S5 / 0.2084.

2. The detection method according to claim 1, characterized in that: In the step S1 and step S4, the heating temperature is independently 50-70°C. The reflux time is 15-60 min.

3. The detection method according to claim 2, characterized in that: The heating temperature is 60°C and the heating method is water bath heating; The reflux time is 15 min.

4. The detection method according to claim 1, characterized in that: In the step S1 and step S4, independently, the extraction comprises extracting with n-hexane 1-4 times; The mass volume ratio of CrEL to n-hexane is 1 g: (50-150) ml; During the first extraction, saturated sodium chloride is added in an amount 2-5 times that of n-hexane.

5. The detection method according to claim 4, characterized in that: The extraction comprises extracting twice with n-hexane; The mass volume ratio of the CrEL to n-hexane is 1 g:100 ml; During the first extraction, saturated sodium chloride was added in an amount 3.5 times that of n-hexane.

6. The detection method according to claim 1, characterized in that: In the step S2 and the step S5, independently, the dehydration includes dehydration using anhydrous sodium sulfate; and the filtration includes filtration using a filter membrane.

7. The detection method according to claim 6, characterized in that: The pore size of the filter membrane is 0.22 μm.

8. Application of the detection method according to any one of claims 1 to 7 in the field of CrEL removal.

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