Analysis method for establishing typical steroid female excitation in water environment based on solid-phase extraction

Abstract

The invention discloses an analysis method for establishing typical steroid female excitation in a water environment based on solid-phase extraction, which comprises the following steps: S1, optimizing a sample pretreatment condition, and reducing or eliminating the influence of a highly complex matrix in a water body on extraction; s2, respectively selecting different solid-phase extraction columns and different organic solvents as leacheate, and finally optimizing the pretreatment conditions through the recovery effect of the standard substance; s3, enabling the standard substance to pass through different types of analytical columns; and S4, optimizing by using a mass spectrum, exploring optimal mass spectrum conditions of parent ions, characteristic ions, cracking voltage and collision energy of typical SEs, and establishing an analysis method. The pretreatment method disclosed by the invention is simple, does not need a derivatization step, is high in automation degree, has the advantages of accurate quantitative analysis of SEs components, high sensitivity and the like, and realizes automatic and high-throughput analysis of different types of SEs.

Description

Technical Field

[0001] The invention relates to the technical field of environmental engineering, and in particular to an analytical method for typical steroid estrogens in water environment based on solid phase extraction. Background Art

[0002] Environmental endocrine disruptors (EDCs) refer to exogenous substances that interfere with the synthesis, secretion, transport, combination, reaction and metabolism of hormones in organisms, thus affecting the normal maintenance of organisms and endangering reproduction, development or behavior. They are also called environmental hormones. Many chemicals in the environment have endocrine disrupting effects. The properties of these compounds vary greatly. There are persistent organic pollutants (POPs) (such as polychlorinated biphenyls, polycyclic aromatic hydrocarbons and organochlorine pesticides), hormones (steroids) excreted by animals and humans, birth control drugs, detergent degradation products (alkylphenols), natural plant hormones, polar herbicides, etc. Among the many endocrine disruptors, steroid estrogens (SEs) are the most concerned type of substances. It is a class of fat-soluble biologically active substances. The basic core of its structure is a cyclopentane polyhydrophenanthrene formed by the fusion of three six-membered rings and one five-membered ring, that is, a steroid ring. This type of substance has stable chemical properties and is easily accumulated in organisms after entering the environment. Compared with other EDCs, it has a stronger endocrine disrupting effect and its impact on the ecological environment is particularly significant.

[0003] Common SEs components mainly include naturally occurring estrone (Estrone, E1), 17α-estradiol (17α-Estradiol, 17α-E2), 17β-estradiol (17β-Estradiol, 17β-E2), estriol (E3) and synthetic 17α-ethynylestradiol (17α-ethynylestradoil, EE2).

[0004] Due to the extremely low concentration of SEs in the environment and the interference of complex matrix in the environment, the detection of this type of substance is difficult. At present, domestic research is still in its early stages and there is no standard analysis method.

[0005] The analysis of SEs in complex environmental media generally includes two processes: sample pretreatment and instrument detection. Sample pretreatment includes extraction, purification, concentration and derivatization, which can enrich trace components, eliminate matrix interference and improve method sensitivity. The analysis of trace SEs requires highly sensitive analytical instruments. With the advancement of modern science and technology, various advanced analytical instruments and methods have been greatly improved in terms of applicability and sensitivity. In particular, the popularity of gas chromatography and liquid chromatography with mass spectrometry detectors has greatly promoted the accurate analysis of SEs.

[0006] Common problems with existing analytical techniques:

[0007] Due to the extremely low concentration of steroid estrogens in the environment and the interference of complex matrices in the environment, the detection of this type of substance is difficult. Currently, research is still in its early stages and there is no standard analysis method.

[0008] At present, the methods used to determine endocrine disruptors in environmental media at home and abroad are mostly liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry. Gas chromatography-mass spectrometry technology has low cost and high sensitivity, but its pretreatment requires a derivatization step, which is cumbersome. Liquid chromatography-mass spectrometry technology has relatively simple pretreatment, good selectivity and no derivatization. Therefore, high-performance liquid chromatography-mass spectrometry is currently used to determine endocrine disruptors in the environment.

[0009] Some domestic scholars have used ultra-high performance liquid chromatography-mass spectrometry to analyze the estrogen level in sewage, but the types of substances analyzed are relatively single. Different types of hormones have different ionization modes in the mass spectrometry source and usually need to be measured separately, resulting in long analysis time and the need to improve the analysis efficiency. In actual research, multiple types of endocrine disruptors often coexist in environmental water bodies, and there are relatively few efficient analytical methods that can simultaneously measure multiple types of endocrine disruptors. Summary of the invention

[0010] The purpose of the present invention is to solve the above technical problems and to propose an analytical method for typical steroid estrogens in water environment based on solid phase extraction.

[0011] In order to achieve the above object, the present invention adopts the following technical solutions:

[0012] The analytical method for typical steroid estrogens in water environment is established based on solid phase extraction, including the following steps:

[0013] S1, optimize sample pretreatment conditions to reduce or eliminate the impact of highly complex matrix in water on extraction;

[0014] S2, select different solid phase extraction columns and different organic solvents as eluents, and finally optimize the pretreatment conditions through the recovery effect of standard substances;

[0015] S3, passing the standard substance through different types of analytical columns, analyzing the peak shape of the target substance in the sample, the separation of the chromatographic target peak and the interference peak to determine the best analytical column;

[0016] S4, using mass spectrometry to optimize, explore the optimal mass spectrometry conditions for parent ions, characteristic ions, fragmentation voltage and collision energy of typical SEs, and establish an analytical method.

[0017] Preferably, the organic solvent in S2 is methanol or acetonitrile.

[0018] Preferably, the mass spectrometry optimization is automatic optimization or manual optimization.

[0019] Compared with the prior art, the present invention has the following beneficial effects:

[0020] The pretreatment method of the present invention is simple, does not require a derivatization step, has a high degree of automation, has the advantages of accurate quantitative analysis of SEs components, and has high sensitivity, thereby realizing automated and high-throughput analysis of different types of SEs. DETAILED DESCRIPTION

[0021] In the description of the present application, it should be noted that the terms used herein are only for describing specific embodiments and are not intended to limit the exemplary embodiments according to the present application. The techniques, methods and devices known to those of ordinary skill in the relevant art may not be discussed in detail, but in appropriate cases, the techniques, methods and devices should be considered as part of the authorization specification. In all examples shown and discussed here, any specific value should be interpreted as being merely exemplary and not as a limitation. Therefore, other examples of the exemplary embodiments may have different values.

[0022] The technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all the embodiments.

[0023] The analytical method for typical steroid estrogens in water environment is established based on solid phase extraction, including the following steps:

[0024] S1, optimize sample pretreatment conditions to reduce or eliminate the effects of highly complex matrix in water on extraction; select the most suitable extraction technology based on the properties of the target analyte (such as polarity, volatility, stability, etc.) and the characteristics of the complex matrix;

[0025] S2, select different solid phase extraction columns and different organic solvents as eluents, and finally optimize the pretreatment conditions through the recovery effect of standard substances; when selecting solid phase extraction columns, first of all, we must have a deep understanding of the materials, particle sizes, specific surface areas, functional groups and other characteristics of various extraction columns, which determine their adsorption capacity and selectivity for different analytes; according to the chemical properties of the target analytes (such as polarity, functional groups, etc.) and the characteristics of the complex matrix, select the most suitable extraction column type;

[0026] S3, passing the standard substance through different types of analytical columns, analyzing the peak shape of the target substance in the sample, the separation of the chromatographic target peak and the interference peak to determine the best analytical column;

[0027] Select standard substances with known concentration and high purity to ensure their accuracy and reliability during the analysis; select appropriate solvents for dissolution and dilution according to the properties of the target analytes to ensure their effective separation and detection on the analytical column; prepare analytical columns of various materials, particle sizes, specific surface areas and functional groups to cover target analytes of different polarities and molecular weights; ensure that each analytical column is properly pretreated and activated before use to improve its separation effect and stability; observe the peak shape of the target analyte through the chromatogram, including the peak height, width, symmetry and other characteristics. The ideal peak shape should be sharp, symmetrical and without obvious tailing, which helps to accurately measure the peak area and concentration; carefully analyze the chromatogram to determine the target peak position of the target analyte; observe whether there are other interfering peaks near the target peak and evaluate the separation between them.

[0028] S4, use mass spectrometry for optimization, explore the optimal mass spectrometry conditions for the parent ion, characteristic ion, fragmentation voltage and collision energy of typical SEs, and establish an analytical method; prepare standard solutions or actual sample solutions containing typical SEs-related substances, and ensure that their concentrations are moderate for mass spectrometry detection; if possible, use isotope-labeled standard substances as internal standards to improve the accuracy and reliability of the analysis; according to experimental requirements, select the appropriate type of mass spectrometry instrument, such as liquid chromatography-mass spectrometry (LC-MS), gas chromatography-mass spectrometry (GC-MS) or direct infusion mass spectrometry (DI-MS); perform necessary preheating and calibration of the mass spectrometer to ensure that the instrument is in good condition and meets the experimental requirements.

[0029] In the present invention, the organic solvent in S2 is methanol or acetonitrile.

[0030] In the present invention, the mass spectrometry optimization is automatic optimization or manual optimization.

[0031] Specifically:

[0032] ① Sample pretreatment optimization

[0033] Optimize sample pretreatment conditions to reduce or eliminate the effects of highly complex matrices in water on extraction, efficiently extract the analyte, eliminate interfering components, reduce matrix effects on subsequent mass spectrometry, and improve analytical sensitivity and accuracy. Select different solid phase extraction columns and different organic solvents (methanol, acetonitrile) as eluents, and finally optimize pretreatment conditions through the recovery effect of standard substances.

[0034] ②Optimization of chromatography-mass spectrometry conditions

[0035] The standard substances were passed through different types of analytical columns, and the peak shape of the target substance in the sample, the separation of the chromatographic target peak and the interference peak were analyzed to determine the best analytical column. The optimal mass spectrometry conditions such as the parent ion, characteristic ion, fragmentation voltage and collision energy of typical SEs (five research targets were tentatively identified: E1, 17α-E2, 17β-E2, E3, and EE2) were explored by automatic or manual mass spectrometry optimization, and the analytical method was established. At present, the parent ion, characteristic ion, fragmentation voltage and collision energy of the first four substances (E1, 17α-E2, 17β-E2, and E3) have been basically found (Estrdiol-d4 is the mass spectrometry parameter of the internal standard).

[0036] Table 1 Mass spectrometry parameters of target SEs

[0037]

[0038]

[0039] Due to the extremely low concentration of steroid estrogens in the environment and the interference of complex matrices in the environment, the detection of such substances is difficult. The current research is still in its infancy and there is no standard analysis method. In actual research, multiple types of endocrine disruptors often coexist in environmental water bodies, and there are relatively few efficient analysis methods that can simultaneously determine multiple types of endocrine disruptors. The present invention establishes a reliable solid phase extraction-high performance liquid chromatography-triple quadrupole tandem mass spectrometry analysis method to achieve automation and high-throughput analysis of different types of SEs.

[0040] At present, the methods used to determine endocrine disruptors in environmental media at home and abroad are mostly liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry. Gas chromatography-mass spectrometry technology has low cost and high sensitivity, but its pretreatment requires a derivatization step, which is cumbersome to operate. The pretreatment of liquid chromatography-mass spectrometry technology is relatively simple, has good selectivity and does not require derivatization. Therefore, high-performance liquid chromatography-mass spectrometry is currently used to determine endocrine disruptors in the environment. The present invention selects a solid phase extraction-high-performance liquid chromatography-triple quadrupole tandem mass spectrometry analysis method with a simple pretreatment method, no derivatization step, a high degree of automation, and the advantages of accurate quantitative analysis of SEs components and high sensitivity.

[0041] The above description is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can make equivalent replacements or changes according to the technical scheme and inventive concept of the present invention within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.

Claims

1. A method for analyzing typical steroid estrogens in water environment is established based on solid phase extraction, characterized in that: The following steps are involved: S1, optimize sample pretreatment conditions to reduce or eliminate the impact of highly complex matrix in water on extraction; S2, select different solid phase extraction columns and different organic solvents as eluents, and finally optimize the pretreatment conditions through the recovery effect of standard substances; S3, passing the standard substance through different types of analytical columns, analyzing the peak shape of the target substance in the sample, the separation of the chromatographic target peak and the interference peak to determine the best analytical column; S4, using mass spectrometry to optimize, explore the optimal mass spectrometry conditions for parent ions, characteristic ions, fragmentation voltage and collision energy of typical SEs, and establish an analytical method.

2. The method for analyzing typical steroid estrogens in water environment based on solid phase extraction according to claim 1, characterized in that: The organic solvent in S2 is methanol or acetonitrile.

3. The method for analyzing typical steroid estrogens in water environment based on solid phase extraction according to claim 1, characterized in that: The mass spectrum optimization is automatic optimization or manual optimization.

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