A method for analyzing thiol compounds

Abstract

The present invention provides an analytical method for thiol compounds, comprising: 1) dissolving standard substances of various thiol compounds in an aqueous solution with a pH of 10-12, and derivatizing them with 2,4,6-trifluorobenzylamine and o-phthalaldehyde to obtain standard substances of various derivatized thiol compounds. 19 FNMR signal spectrum; 2) dissolving the thiol compound to be tested in an aqueous solution with a pH of 10-12, and derivatizing it with 2,4,6-trifluorobenzylamine and o-phthalaldehyde to obtain the nuclear magnetic resonance of each derivatized thiol compound to be tested 19 FNMR signal spectrum; 3) superimposing and comparing the two spectra to identify the thiol compound to be tested; at the same time, the product after derivatization can be 19 The relationship between the ortho or para peak area and the concentration of FNMR is quantified and mutually verified; the method of the invention is simple and easy to implement, can prepare and measure samples immediately, has high sensitivity and selectivity, can effectively avoid interference from other biological molecules, and can be used for qualitative and quantitative analysis of thiol compounds in biological systems.

Description

Technical Field

[0001] The invention belongs to the technical field of chemical analysis, and in particular relates to an analysis method for thiol compounds. Background Art

[0002] Thiol is the most active group in cells. Common thiol compounds include Figure 1 As shown, among them, cysteine ​​(Cys), homocysteine ​​(Hcys) and N-acetylcysteine ​​(NAC) are abundant biological small molecule thiol compounds in the human body. These biological molecules have strong reducing and nucleophilic properties, participate in various physiological processes of the organism, and maintain normal physiological functions of the body, such as protein synthesis, cell redox homeostasis, etc. It has been reported that the metabolic abnormalities of thiol compounds in organisms are also related to various diseases, such as diabetes, leukemia, acute liver failure, nervous system diseases, etc. Based on these findings, more and more studies have begun to focus on finding biomarkers of diseases. For example, Hcys has been considered to be related to cardiovascular diseases, and excessive Cys in urine can also be used to diagnose cystinuria; in addition, some thiol compounds, such as NAC, can also be used to treat hypertension and liver diseases, etc. Therefore, a method for detecting thiol compounds with high sensitivity and selectivity is of great significance for biochemical research and disease diagnosis.

[0003] At present, the traditional analysis methods for biothiol compounds mainly include high performance liquid chromatography (HPLC), mass spectrometry (MS), liquid chromatography-mass spectrometry (HPLC-MS), capillary electrophoresis and fluorescence methods, but most of the above methods require complex sample pretreatment, long detection time, large background signal interference, and low selectivity for biothiol compounds. Among them, fluorescence method has been widely used in biological sample analysis, such as the specific detection of Cys / Hcy in living human fibrosarcoma cells in the presence of glutathione (GSH), but its low resolution is still the most critical problem limiting its detection application. Compared with fluorescence detection, nuclear magnetic resonance (NMR) is an important analytical technique that can provide a large amount of material structure information while analyzing the mixture, including the composition, configuration and conformation of unknown compounds, and has low background interference. Due to 1 The natural abundance of H is 99.985%, which has a high detection sensitivity and is often used in metabolomics analysis; however, the spectral width of hydrogen spectrum is relatively narrow, and for samples with more complex structures, the spectrum is prone to peak overlap, which brings certain difficulties to the detection of thiol compounds. Therefore, there are few reports on the use of highly sensitive nuclear magnetic resonance in the analysis of thiol compounds.

[0004] In summary, there is an urgent need to develop a method that is rapid and simple, has high sensitivity, good selectivity, no background interference, and can simultaneously detect a variety of small molecule thiol compounds. Summary of the invention

[0005] The technical problem to be solved by the present invention is to provide a method which is simple and easy to use, can prepare samples and test samples immediately, has high sensitivity and selectivity for the detection of thiol compounds, can effectively avoid interference from other biological molecules, can detect a variety of small molecule thiol compounds at the same time, and is suitable for qualitative and quantitative analysis of thiol compounds in biological systems.

[0006] The technical solution adopted by the present invention to solve the above technical problems is:

[0007] A method for analyzing thiol compounds comprises the following steps:

[0008] 1) The standard substances of each thiol compound are respectively dissolved in an aqueous solution with a pH of 10-12, and after being fully mixed with 2,4,6-trifluorobenzylamine, o-phthalaldehyde and a deuterated reagent are added to perform a nuclear magnetic resonance test to obtain the nuclear magnetic resonance of the standard substances of each derivatized thiol compound. 19 F NMR signal spectrum;

[0009] 2) dissolving the thiol compound to be tested in an aqueous solution with a pH of 10-12, and fully mixing with the 2,4,6-trifluorobenzylamine, adding the o-phthalaldehyde and the deuterated reagent, and performing a nuclear magnetic resonance test to obtain a nuclear magnetic resonance of the derivatized thiol compound to be tested. 19 F NMR signal spectrum;

[0010] 3) performing nuclear magnetic resonance (NMR) of the derivatized thiol compound to be tested in step 2) 19 F NMR signal spectrum and the NMR of the standard substance of the thiol compound after each derivatization in step 1) 19 The F NMR signal spectra are superimposed and compared to identify the chemical properties of the thiol compound to be tested.

[0011] A method for analyzing thiol compounds comprises the following steps:

[0012] S1: Select different concentrations of the standard substance of each thiol compound and dissolve them in an aqueous solution with a pH of 10-12. After fully mixing with 2,4,6-trifluorobenzylamine, add o-phthalaldehyde and a deuterated reagent to prepare a series of standard solutions of the thiol compounds with different concentrations. Perform nuclear magnetic resonance tests to obtain the nuclear magnetic resonance images of the standard solutions of the thiol compounds with different concentrations. 19 F NMR spectrum;

[0013] S2: the standard solution of the thiol compound with different concentrations in step S1 19 The ortho or para peak area of ​​the F NMR spectrum is integrated to obtain the standard solution of the thiol compound with different concentrations. 19 The ortho or para peak area integral value of F NMR is used to draw the standard solution of the thiol compound. 19 The linear relationship curve between the ortho- or para-position peak area integral value of F NMR and the concentration is obtained to obtain the ortho- or para-position linear equation;

[0014] S3: dissolving the thiol compound to be tested in an aqueous solution with a pH of 10-12, and fully mixing with the 2,4,6-trifluorobenzylamine, adding the o-phthalaldehyde and the deuterated reagent, and performing a nuclear magnetic resonance test to obtain a nuclear magnetic resonance image of the derivatized thiol compound to be tested. 19 F NMR signal spectrum, for the thiol compound to be tested 19 The ortho or para peak area of ​​the FNMR spectrum is integrated to obtain the 19 The ortho- or para-peak area integral value of F NMR is substituted into the ortho- or para-linear equation to calculate the content of the thiol compound to be measured.

[0015] Furthermore, the molar ratio of the thiol compound to the 2,4,6-trifluorobenzylamine is 1:1.

[0016] Furthermore, the molar ratio of the 2,4,6-trifluorobenzylamine to the o-phthalaldehyde is greater than 1.

[0017] Furthermore, the molar ratio of the thiol compound, the 2,4,6-trifluorobenzylamine and the o-phthalaldehyde is 2:2:1.

[0018] Furthermore, the aqueous solution with pH=10-12 is a borax-sodium hydroxide buffer solution.

[0019] Furthermore, the deuterated reagent is selected from deuterated water, deuterated methanol, deuterated acetonitrile, or deuterated dimethyl sulfoxide.

[0020] Furthermore, the thiol compound is any one or more of 3-mercaptopropionic acid, furfuryl mercaptan, 1-propanethiol, 2-naphthalenethiol, β-mercaptoethanol, butyl thioglycolate, 4-methoxybenzyl mercaptan, cyclohexyl mercaptan, 2,3-butanedithiol, 1,2-ethanedithiol, 1,5-pentanedithiol, L-cysteine, N-acetyl-L-cysteine ​​or 1,4-benzenedithiol.

[0021] Compared with the prior art, the advantages of the present invention are:

[0022] The present invention utilizes commercial reagents 2,4,6-trifluorobenzylamine and o-phthalaldehyde (OPA) to carry out derivatization reaction with different thiol compounds to introduce two environments of ortho and para positions. 19 F, based on NMR of the derivatized product 19 FNMR is sensitive to the chemical environment. The structure of thiol compounds will affect the ortho- and para-position NMR of the product. 19 FNMR chemical shift signals, that is, the products generated by reactions with different thiol compounds have different nuclear magnetic resonance 19 FNMR chemical shifts, thereby drawing characteristic "pseudo-2D" NMR of various thiol compounds 19 F NMR spectrum is used to identify thiol compounds; the detection method of the invention is simple and convenient, can realize rapid analysis and identification of various thiol compounds, and has low cost.

[0023] The present invention uses o-phthalaldehyde, 2,4,6-trifluorobenzylamine and different thiol compounds to react, the reaction conditions are mild and rapid, and the nuclear magnetic resonance of the corresponding product can be detected when the nuclear magnetic resonance sample is prepared. 19 F NMR signal can realize instant sample preparation and instant sample measurement.

[0024] The present invention uses nuclear magnetic resonance 19 The chemical shift of F NMR is highly sensitive to the local environment, the spectrum peak is widely distributed, there is no background signal interference, and the peak signal overlap is small. It can be quantified and mutually verified according to the relationship between the peak area integral value of the ortho or para position of the product and the concentration, which improves the accuracy of the present invention and can perform qualitative and quantitative analysis of thiol compounds in biological systems. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 It is a common thiol compound;

[0026] Figure 2 It is the reaction principle diagram of the present invention;

[0027] Figure 3 The nuclear magnetic resonance of the thiol compound labeled with 2,4,6-trifluorobenzylamine under the condition of pH=10 in Example 1 of the present invention is 19 F NMR spectrum overlay;

[0028] Figure 4 The nuclear magnetic resonance of the thiol compound labeled with 2,4,6-trifluorobenzylamine under the condition of pH=10 in Example 1 of the present invention is 19 F NMR chemical shift value "pseudo-two-dimensional" map;

[0029] Figure 5 1-Propanethiol labeled with 2,4,6-trifluorobenzylamine at pH = 10 19Linear relationship curve between F-NMR peak area integral value and 1-propanethiol concentration. DETAILED DESCRIPTION

[0030] The present invention is further described in detail below with reference to the accompanying drawings.

[0031] In order to further understand the present invention, preferred embodiments of the present invention are described below in conjunction with examples. However, it should be understood that these descriptions are only for further illustrating the features and advantages of the present invention, rather than limiting the claims of the present invention.

[0032] Nuclear Magnetic Resonance 19 F NMR is characterized by high natural abundance, which can reach 100%, high sensitivity, and easy to obtain high-resolution spectra. Its importance cannot be underestimated. 19 The high sensitivity of F NMR chemical shift to the local environment results in a broad distribution of spectral peaks, which reduces the overlap of peak signals and is not present in commonly used matrices. 19 F background signal, thus avoiding interference with the analyte signal, are advantages for analyzing thiol compounds in biological samples.

[0033] The invention discloses a qualitative analysis method for thiol compounds, comprising the following steps:

[0034] 1) The standard substances of each thiol compound were dissolved in an aqueous solution with a pH of 10-12, and mixed thoroughly with 2,4,6-trifluorobenzylamine. Then, o-phthalaldehyde and a deuterated reagent were added to obtain the NMR spectra of the standard substances of each derivatized thiol compound. 19 F NMR signal spectrum; 2) the thiol compounds to be tested are dissolved in an aqueous solution of pH = 10-12, and after being fully mixed with 2,4,6-trifluorobenzylamine, o-phthalaldehyde and a deuterated reagent are added to obtain the NMR signal of each derivatized thiol compound to be tested. 19 F NMR signal spectrum; 3) superimposing and comparing the two spectra to identify the chemical properties of the thiol compound to be tested.

[0035] The present invention also discloses a quantitative analysis method of thiol compounds, comprising the following steps:

[0036] S1: Select different concentrations of standard substances of various thiol compounds and dissolve them in aqueous solution with pH=10-12. After being fully mixed with 2,4,6-trifluorobenzylamine, o-phthalaldehyde and deuterated reagent are added to prepare a series of standard solutions of thiol compounds with different concentrations. The NMR spectra of standard solutions of thiol compounds with different concentrations are obtained respectively. 19 F NMR spectrum;

[0037] S2: Standard solutions of thiol compounds with different concentrations in step S1 19 The ortho or para peak area of ​​the F NMR spectrum was integrated to obtain the standard solutions of thiol compounds with different concentrations. 19 The integrated value of the ortho or para peak area of ​​F NMR is used to plot the standard solution of the thiol compound. 19 The linear relationship curve between the ortho- or para-position peak area integral value of F NMR and the concentration is obtained to obtain the ortho- or para-position linear equation;

[0038] S3: dissolving the thiol compound to be tested in an aqueous solution with a pH of 10-12, and fully mixing with 2,4,6-trifluorobenzylamine, adding o-phthalaldehyde and the deuterated reagent, and performing a nuclear magnetic resonance test to obtain a nuclear magnetic resonance image of the derivatized thiol compound to be tested. 19 F NMR signal spectrum of the thiol compound to be tested 19 The ortho or para peak area of ​​the F NMR spectrum was integrated to obtain the peak area of ​​the thiol compound to be tested. 19 The integral value of the ortho or para peak area of ​​F NMR is substituted into the ortho or para linear equation to calculate the content of the thiol compound to be measured.

[0039] In the present invention, 2,4,6-trifluorobenzylamine (a), o-phthalaldehyde (c) and different thiol compounds (b) react under pH=10-12 to generate corresponding products (d), as shown in the attached Figure 2 As shown; Due to the different structures of different thiol compounds, the ortho and para positions of the product (d) 19 The different chemical environments induced by F produce different chemical shifts, thereby achieving the purpose of analyzing different thiol compounds. 19 The F signal peak area integration ratio of 1:2 can effectively carry out the quantitative analysis of thiol compounds and verify each other.

[0040] In the present invention, the molar ratio of the thiol compound (b) to 2,4,6-trifluorobenzylamine (a) is 1:1; the molar ratio of 2,4,6-trifluorobenzylamine (a) to o-phthalaldehyde (c) is greater than 1; the molar ratio of the thiol compound (b), 2,4,6-trifluorobenzylamine (a) to o-phthalaldehyde (c) can be 2:2:1 or 3:3:1, etc., and the preferred molar ratio is 2:2:1.

[0041] In the present invention, the deuterated reagent only needs to be miscible with the water or ethanol in the reaction system. Preferably, the deuterated reagent can be deuterated water, deuterated methanol, deuterated acetonitrile, or deuterated dimethyl sulfoxide.

[0042] In the present invention, a buffer solution can be used to adjust the pH of the mixed solution to 10-12. The buffer solution is mainly used to keep the pH and ion concentration values ​​within a certain range and reduce the influence of the solution environment on the product nuclear magnetic resonance spectrum peak signal. Preferably, the buffer solution can be a borax-sodium hydroxide buffer solution.

[0043] In the present invention, the thiol compound can be any one or more of 3-mercaptopropionic acid, furfuryl mercaptan, 1-propanethiol, 2-naphthalenethiol, β-mercaptoethanol, butyl thioglycolate, 4-methoxybenzyl mercaptan, cyclohexyl mercaptan, 2,3-butanedithiol, 1,2-ethanedithiol, 1,5-pentanedithiol, L-cysteine ​​(Cys), N-acetyl-L-cysteine ​​(NAC), and 1,4-benzenedithiol.

[0044] In order to further illustrate the scheme of the present invention, a method for identifying amino acids provided by the present invention is described in detail below in conjunction with the accompanying embodiments. However, it should be understood that the protection scope of the present invention is not limited to the following embodiments.

[0045] Main reagents: 2,4,6-trifluorobenzylamine, CAS No. 214759-21-4, β-mercaptoethanol, CAS No. 60-24-2, 1,4-benzenedithiol, CAS No. 624-39-5, 4-methoxybenzylmercaptan, CAS No. 6258-60-2, 1,2-ethanedithiol, CAS No. 540-63-6, cyclohexylmercaptan, CAS No. 1569-69-3, 3-mercaptopropionic acid, CAS No. 107-96-0, 2,3-butanedithiol, CAS No. 4532-64-3, 1,5-pentanedithiol, CAS No. 928-98-3 were purchased from San Chemical Technology (Shanghai) Co., Ltd.; butyl thioglycolate, CAS No. 10 047-28-6, purchased from TCI (Shanghai) Chemical Industry Development Co., Ltd.; furfurylthiol, CAS No. 10047-28-6, N-acetyl-L-cysteine ​​(NAC), CAS No. 1679-07-8, 1-propanethiol, CAS No. 107-03-9, o-phthalaldehyde, CAS No. 643-79-8, purchased from Shanghai Aladdin Biochemical Technology Co., Ltd.; dithiothreitol (DTT), CAS No. 3483-12-3, purchased from Sangon Biotech (Shanghai) Co., Ltd.; L-cysteine, CAS No. 52-90-4, 2-naphthalenethiol, CAS No. 91-60-1, purchased from Shanghai MacLean Biochemical Technology Co., Ltd.

[0046] Main instrument: Bruker AVANCE 500MHZ superconducting Fourier transform nuclear magnetic resonance instrument.

[0047] Instrument parameters:

[0048] 19The sampling parameters of F NMR spectrum were as follows: using Bruker instrument pulse program zgig (F19CPD), detection temperature 298K, spectral width 113636.367Hz, center frequency -47068.65Hz, and scanning times 16 times; adjusting instrument parameters, tuning, temperature control, shimming, sampling, and Fourier transform to obtain 19 F NMR spectrum.

[0049] Experimental Example 1

[0050] Step 1): Preparation of 0.05M borax-sodium hydroxide solution: Take 50mL 0.05M borax and 43mL 0.2M NaOH and dilute to 200mL with water; Prepare samples: Weigh thiol compounds (3-mercaptopropionic acid, furfuryl mercaptan, 1-propanethiol, 2-naphthalenethiol, β-mercaptoethanol, butyl thioglycolate, 4-methoxybenzyl mercaptan, cyclohexyl mercaptan, 2,3-butanedithiol, 1,2-ethanedithiol, 1,5-pentanedithiol, L-cysteine ​​(Cys), N-acetyl-L-cysteine ​​(NAC), 1,4-benzenedithiol) and dissolve in borax-sodium hydroxide buffer at pH = 10 to prepare a 0.1M solution; Weigh o-phthalaldehyde and 2,4,6-trifluorobenzylamine and dissolve in ethanol to prepare 0.05M and 0.1M solutions respectively.

[0051] Step 2): Select a 1.5mL epoxide tube, add 50μL 0.05M borax-sodium hydroxide solution and 50μL 0.1M 2,4,6-trifluorobenzylamine, mix well, then add 50μL DMSO-d6, and add ethanol to 500μL, where the final concentration of 2,4,6-trifluorobenzylamine (c) is 10mM. Transfer the mixed solution to a 5mm NMR tube for NMR. 19 F NMR signal acquisition, respectively, to obtain the nuclear magnetic resonance of 2,4,6-trifluorobenzylamine (c) 19 F NMR spectrum.

[0052] Step 3): Select 14 1.5mL ep tubes, add 50μL 0.1M target thiol compound and 50μL 0.1M 2,4,6-trifluorobenzylamine, mix thoroughly, add 50μL 0.05M o-phthalaldehyde (a), then add 50μL DMSO-d6, and add ethanol to 500μL, wherein the final concentration of each thiol compound and 2,4,6-trifluorobenzylamine (c) is 10mM, and the final concentration of o-phthalaldehyde (a) is 5mM. Transfer each mixed solution to a 5mm NMR tube for 19 F NMR signal acquisition, 14 corresponding 2,4,6-trifluorobenzylamine (c) labeled target thiol compounds were obtained. 19 F NMR spectrum, and overlay analysis, as shown in the attached Figure 3As shown, 19 The F NMR chemical shift table is shown in Table 1.

[0053] Table 1 NMR of 2,4,6-trifluorobenzylamine labeled thiol compounds at pH = 10 19 Chemical shift table of F NMR spectral signals

[0054]

[0055] Experimental results:

[0056] From the attached Figure 3 From Table 1, we can see that due to 19 F- 19 F coupling, ortho position 19 The F splitting was divided into a doublet, and the average value of the doublet was set as δpF; the para F splitting was divided into a triplet, and the average value of the triplet was set as δoF. The values ​​of δpF were used as the abscissa and δoF as the ordinate to obtain the attached Figure 4 , under the condition of pH=10, 14 thiol compounds including 3-mercaptopropionic acid, furfuryl mercaptan, 1-propanethiol, 2-naphthalenethiol, β-mercaptoethanol, butyl thioglycolate, 4-methoxybenzyl mercaptan, cyclohexyl mercaptan, 2,3-butanedithiol, 1,2-ethanedithiol, 1,5-pentanedithiol, L-cysteine ​​(Cys), N-acetyl-L-cysteine ​​(NAC), and 1,4-benzenedithiol can be identified with good recognition effect. Among them, 1-propanethiol and 4-methoxybenzyl mercaptan can be identified under this condition. 19 The F NMR chemical shifts are very close, with δpF being 107.647 and 107.644, and δoF being -111.920 and -111.924, respectively, but they can still be distinguished.

[0057] Experimental Example 2

[0058] Quantitative experiments

[0059] (1) A series of 1-propanethiol with different concentrations were prepared. A series of standard solutions of 1-propanethiol with different concentrations were mixed with 2,4,6-trifluorobenzylamine (c) and o-phthalaldehyde (a) in 450 μL of 0.05 M borax-sodium hydroxide solution with a pH of 10, and then 50 μL of DMSO-D6 was added to obtain standard solutions with 1-propanethiol concentrations of 0.1, 0.2, 0.5, 1, 2, 5, 10, 25, and 50 mM, respectively. Nuclear magnetic resonance tests were performed to determine the 19 F NMR signals were collected to obtain the corresponding 1-propanethiol labeled with o-phthalaldehyde (a) at different concentrations. 19 F NMR spectrum; select the integral correction method and calculate the product obtained at each concentration of o-phthalaldehyde (a) labeled 1-propanethiol19 The peak area of ​​F NMR peaks was integrated to obtain the ortho- and para-positions of 1-propanethiol labeled with o-phthalaldehyde (a) at different concentrations. 19 The F NMR peak area integral value is based on the concentration of 1-propanethiol as the abscissa and the ortho position of the product as the 19 The F NMR peak area integral value is used as the ordinate, the concentration of 1-propanethiol is used as the abscissa, and the para-position of the product is used as the 19 The F NMR peak area integral value was used as the ordinate, and linear fitting was performed using OriginPro software. 19 The linear relationship curve between the FNMR peak area integral value and the 1-propanethiol concentration is shown in the attached figure. Figure 5 As shown, the linear equation of F(o) fitting corresponding to the adjacent position is: y(p) = 1.8 × 10 11 x-2.7×10 10 , correlation coefficient R 2 =99.8; the corresponding F(p) fitting linear equation is y(o)=3.9×10 11 x-1.5×10 11 , correlation coefficient R 2 =99.7.

[0060] (2) Weigh a certain amount of 1-propanethiol and dissolve it in 0.05 M borax-sodium hydroxide solution to obtain a test solution; mix the prepared test solution with 2,4,6-trifluorobenzylamine (c) and o-phthalaldehyde (a) in 450 μL of borax-sodium hydroxide buffer with a pH of 10, add 50 μL DMSO-D6, wherein the concentration of o-phthalaldehyde (a) is 50 mM, and transfer the test solution to a nuclear magnetic resonance tube for nuclear magnetic resonance. 19 F NMR signal acquisition, obtain the nuclear magnetic resonance of the solution to be tested 19 F NMR spectrum, found in the test solution o-phthalaldehyde (a) labeled 1-propanethiol NMR 19 The F NMR signal peaks were integrated to obtain the peak area integral values ​​y(p) and y(o) of the ortho-position F(p) and para-position F(o) of the test solution, which were 1.3×10 12 and 6.0×10 11 , respectively, into the linear regression equation y(p) = 1.8 × 10 11 x-2.7×10 10 , y(o)=3.9×10 11 x-1.5×10 11 , the concentration of 1-propanethiol in the test solution was 3.7 mM.

[0061] The quantitative experimental results show that the product can be 19The relationship between the F NMR peak area integral value and the concentration was quantitatively analyzed and verified with each other, which improved the accuracy of quantitative analysis.

[0062] The above is only a preferred embodiment of the present invention. It should be pointed out that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. A method for analyzing sulfhydryl compounds, characterized in that The following steps are involved: 1) The standard substances of each thiol compound are respectively dissolved in an aqueous solution with a pH of 10-12, and after being fully mixed with 2,4,6-trifluorobenzylamine, o-phthalaldehyde and a deuterated reagent are added to perform a nuclear magnetic resonance test, and the nuclear magnetic resonance of the standard substances of each derivatized thiol compound is obtained. 19 F NMR signal spectrum; 2) dissolving the thiol compound to be tested in an aqueous solution with a pH of 10-12, and fully mixing with the 2,4,6-trifluorobenzylamine, adding the o-phthalaldehyde and the deuterated reagent, and performing a nuclear magnetic resonance test to obtain the nuclear magnetic resonance of the derivatized thiol compound to be tested. 19 F NMR signal spectrum; 3) performing nuclear magnetic resonance (NMR) of the derivatized thiol compound to be tested in step 2) 19 F NMR signal spectrum and the NMR of the standard substance of the thiol compound after each derivatization in step 1) 19 The F NMR signal spectra are superimposed and compared to identify the chemical properties of the thiol compound to be tested; The thiol compound is any one or more of 3-mercaptopropionic acid, furfuryl mercaptan, 1-propanethiol, 2-naphthalenethiol, β-mercaptoethanol, butyl thioglycolate, 4-methoxybenzyl mercaptan, cyclohexyl mercaptan, 2,3-butanedithiol, 1,2-ethanedithiol, 1,5-pentanedithiol, L-cysteine, N-acetyl-L-cysteine ​​or 1,4-benzenedithiol.

2. A method for analyzing sulfhydryl compounds, characterized in that The steps include: S1: Select different concentrations of standard substances of various thiol compounds and dissolve them in an aqueous solution with a pH value of 10-12. After being fully mixed with 2,4,6-trifluorobenzylamine, o-phthalaldehyde and a deuterated reagent are added to prepare a series of standard solutions of the thiol compounds with different concentrations. The NMR test is performed to obtain the NMR of the standard solutions of the thiol compounds with different concentrations. 19 F NMR spectrum; S2: the standard solution of the thiol compound with different concentrations in step S1 19 The ortho or para peak area of ​​the F NMR spectrum is integrated to obtain the standard solution of the thiol compound with different concentrations. 19 The ortho or para peak area integral value of F NMR is used to draw the standard solution of the thiol compound. 19 The linear relationship curve between the ortho- or para-position peak area integral value of F NMR and the concentration is obtained to obtain the ortho- or para-position linear equation; S3: dissolving the thiol compound to be tested in an aqueous solution with a pH value of 10-12, and fully mixing with the 2,4,6-trifluorobenzylamine, adding the o-phthalaldehyde and the deuterated reagent, and performing a nuclear magnetic resonance test to obtain the nuclear magnetic resonance of the derivatized thiol compound to be tested. 19 F NMR signal spectrum, for the thiol compound to be tested 19 The ortho or para peak area of ​​the F NMR spectrum is integrated to obtain the peak area of ​​the thiol compound to be tested. 19 Substituting the ortho- or para-peak area integral value of F NMR into the ortho- or para-linear equation to calculate the content of the thiol compound to be measured; The thiol compound is any one or more of 3-mercaptopropionic acid, furfuryl mercaptan, 1-propanethiol, 2-naphthalenethiol, β-mercaptoethanol, butyl thioglycolate, 4-methoxybenzyl mercaptan, cyclohexyl mercaptan, 2,3-butanedithiol, 1,2-ethanedithiol, 1,5-pentanedithiol, L-cysteine, N-acetyl-L-cysteine ​​or 1,4-benzenedithiol.

3. according to the analytical method of a kind of sulfhydryl compound described in claim 1 or 2, it is characterized in that The molar ratio of the thiol compound to the 2,4,6-trifluorobenzylamine is 1:

1.

4. A method for analyzing sulfhydryl compounds according to claim 3, characterized in that The molar ratio of the 2,4,6-trifluorobenzylamine to the o-phthalaldehyde is greater than 1.

5. A method for analyzing sulfhydryl compounds according to claim 4, characterized in that The molar ratio of the mercapto compound, the 2,4,6-trifluorobenzylamine and the o-phthalaldehyde is 2:2:

1.

6. according to the analytical method of a kind of sulfhydryl compound described in claim 1 or 2, it is characterized in that The aqueous solution with pH=10-12 is borax-sodium hydroxide buffer solution.

7. The analytical method of a sulfhydryl compound according to claim 1 or 2, characterized in that The deuterated reagent is selected from deuterated water, deuterated methanol, deuterated acetonitrile, or deuterated dimethyl sulfoxide.

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