Separation and detection method for cis-trans isomers of 4-tert-butylcyclohexanol and application of separation and detection method

Abstract

The invention relates to the technical field of organic matter detection, in particular to a 4-tert-butyl cyclohexanol cis-trans isomer separation and detection method and application thereof. Aiming at the separation problem caused by similar physicochemical properties of cis-isomers and trans-isomers, efficient separation of the 4-tert-butyl cyclohexanol cis-isomers and trans-isomers is successfully realized by adopting an RB-5 type chromatographic column under specific chromatographic conditions, and quantitative analysis is realized by combining an internal standard method. Experimental results show that the linear fitting degree of a standard working curve reaches 99.9% (R2), the adding standard recovery rate is 97.6%-102.3%, and the relative standard deviation (RSD) of repeated measurement (n = 5) is less than or equal to 1.8%, which indicates that the method has good repeatability and accuracy. The detection method not only solves the problem of separation of cis-trans isomers of 4-tert-butylcyclohexanol, but also has the advantages of simplicity and convenience in operation and short time consumption, and has higher practical application value.

Description

Technical Field

[0001] This application relates to the field of organic matter detection technology, and in particular to a method for separating and detecting the cis-trans isomers of 4-tert-butylcyclohexanol and its application. Background Technology

[0002] 4-tert-butylcyclohexanol, an important organic compound, exhibits significant differences in physicochemical properties and biological activities between its cis and trans isomers due to their spatial configurations. The trans isomer, with its antagonistic effect on TRPV1 receptors, is widely used in cosmetics and pharmaceuticals to soothe sensitive skin, while the cis isomer can be used as an intermediate in antibacterial drugs or as a fragrance ingredient. Both have important applications in different fields.

[0003] However, due to the extremely similar molecular structures of cis-trans isomers, their efficient separation and accurate quantification have always been prominent challenges in analytical chemistry. In traditional detection methods, gas chromatography (GC) often employs medium-polarity columns, but under conventional chromatographic conditions, cis-trans isomers are prone to peak overlap or insufficient resolution, directly leading to significant quantitative errors and failing to meet the requirements for precise analysis. Although some studies have attempted to improve detection stability through derivatization reactions, such methods require the introduction of additional chemical reagents, which may not only alter the original properties of the sample but also increase operational complexity, limiting their practicality.

[0004] In addition to the challenges of separation and quantification, existing detection methods require high-temperature pretreatment to remove impurities during operation. This process not only increases energy consumption but also raises safety risks during the experiment.

[0005] Therefore, there is an urgent need to develop an efficient, safe and accurate method for separating and detecting the cis-trans isomers of 4-tert-butylcyclohexanol in order to overcome the current technical bottlenecks and meet the needs of practical applications. Summary of the Invention

[0006] The technical problem to be solved by the present invention is to overcome the defects and shortcomings of the prior art in which the separation of cis-trans isomers of 4-tert-butylcyclohexanol is difficult, the operation of existing detection methods is complicated and there are safety risks, and to provide a method for separating and detecting cis-trans isomers of 4-tert-butylcyclohexanol.

[0007] Another object of the present invention is to provide the application of the above-described detection method in determining the content of 4-tert-butylcyclohexanol cis-trans isomers in a product.

[0008] The above-mentioned objective of this invention is achieved through the following technical solution: This invention protects a method for separating and detecting the cis-trans isomers of 4-tert-butylcyclohexanol, comprising the following steps: adding an internal standard to the sample solution to be tested, mixing to obtain a test solution, detecting the obtained test solution by gas chromatography, and quantifying by the internal standard method; The conditions for the gas chromatography method include: Chromatographic column: RB-5 gas chromatographic column; Temperature program: 45~55 ℃, hold for 1~5 min, then increase the temperature to 200~230 ℃ at a rate of 5~15 ℃ / min and hold for 1~5 min; The internal standard is benzyl alcohol.

[0009] To address the separation challenge of cis and trans isomers due to their similar physicochemical properties, this invention achieves efficient separation of the cis and trans isomers of 4-tert-butylcyclohexanol using a specific chromatographic condition and an RB-5 column. Quantitative analysis is achieved by combining this method with an internal standard, avoiding errors caused by variations in injection volume and process variables in the external standard method. This detection method not only solves the separation problem of the cis and trans isomers of 4-tert-butylcyclohexanol but also offers advantages such as simple operation and short processing time, making it highly valuable for practical applications.

[0010] Furthermore, the detector used in the gas chromatography is a flame ionization detector.

[0011] Furthermore, the injection mode of the gas chromatography is splitless.

[0012] Furthermore, the carrier gas in the gas chromatography is nitrogen.

[0013] Furthermore, the flow rate of the carrier gas is 3.5~4.5 mL / min.

[0014] Furthermore, the injection volume for the gas chromatography method is 0.1~1 μL.

[0015] Preferably, the RB-5 chromatographic column is a Fuli RB-5 chromatographic column.

[0016] Preferably, the RB-5 chromatographic column has dimensions of 30 m × 0.32 mm × 0.50 μm.

[0017] Furthermore, the stationary phase of the RB-5 column is 5% phenyl-95% dimethylpolysiloxane.

[0018] In a preferred embodiment, the test solution is prepared by mixing the test sample solution with the internal standard solution.

[0019] Furthermore, the test sample solution is obtained by fully dissolving the analyte in a solvent.

[0020] In a preferred embodiment, the preparation of the test sample solution includes the following steps: dissolving and diluting the analyte with a solvent, and making up to a final volume to obtain the test sample solution.

[0021] Furthermore, the internal standard solution is obtained by fully dissolving the internal standard in a solvent.

[0022] In a preferred embodiment, the preparation of the internal standard solution includes the following steps: dissolving and diluting the internal standard with a solvent, and making up to a final volume to obtain the internal standard solution.

[0023] Furthermore, the quantification using the internal standard method includes the following steps: S1. Prepare a mixed isomer standard working solution containing an internal standard; S2. Prepare the test solution; S3. Detect the mixed isomer standard working solution containing internal standard obtained in step S1 by gas chromatography and plot the standard working curve; S4. The test solution obtained in step S2 is detected by gas chromatography to obtain the peak area. Based on the standard working curve obtained in step S3, the contents of cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol are calculated. In step S2, the definition of the test solution is the same as described above.

[0024] Furthermore, the solvents of the test solution or the mixed isomer standard working solution containing the internal standard are each independently selected from one or more of ethanol, ethyl acetate, and tetrahydrofuran.

[0025] Further, in step S1, the preparation of the mixed isomer standard working solution containing the internal standard includes the following steps: Dissolve and dilute the analyte standard sample with a solvent, make up to volume, and prepare a series of analyte standard solutions with different concentration gradients. Add 0.5 to 2 mL of internal standard solution to each solution to obtain a mixed isomer standard working solution containing internal standard.

[0026] Furthermore, the concentration of the internal standard in the test sample solution or the mixed isomer standard working solution containing the internal standard is 5~6.8 g / L.

[0027] Furthermore, the horizontal axis of the standard working curve represents the ratio of the concentration of any isomer to the concentration of the internal standard, and the vertical axis represents the ratio of the peak area of ​​any isomer to the peak area of ​​the internal standard.

[0028] This invention protects the application of the above-described separation and detection method in determining the content of 4-tert-butylcyclohexanol cis-trans isomers in products.

[0029] Furthermore, the products include cosmetics, pharmaceuticals, or food.

[0030] Compared with the prior art, the present invention has the following beneficial effects: To address the separation challenge of cis and trans isomers due to their similar physicochemical properties, this invention successfully achieved efficient separation of the cis and trans isomers of 4-tert-butylcyclohexanol using an RB-5 column under specific chromatographic conditions, and further facilitated quantitative analysis using an internal standard method. Experimental results show that the linearity of the standard working curve reaches 99.9% (R²). 2 The spiked recoveries ranged from 97.6% to 102.3%, and the relative standard deviation (RSD) for repeated measurements (n=5) was ≤1.8%, indicating good repeatability and accuracy. This detection method not only solves the problem of separating the cis-trans isomers of 4-tert-butylcyclohexanol, but also has the advantages of simple operation and short time consumption, making it highly valuable for practical applications. Attached Figure Description

[0031] Figure 1 This is the standard working curve of cis-4-tert-butylcyclohexanol using the internal standard method in Example 1.

[0032] Figure 2 This is the standard curve of trans-4-tert-butylcyclohexanol using the internal standard method in Example 1.

[0033] Figure 3 This is a gas chromatogram of the cis-trans isomer of 4-tert-butylcyclohexanol in Example 1.

[0034] Figure 4 This is a gas chromatogram of the cis-trans isomer of 4-tert-butylcyclohexanol in Example 2.

[0035] Figure 5 The gas chromatogram is shown for the concentration of 4-tert-butylcyclohexanol cis-trans isomer in Example 4, which is 0.25 g / L. Detailed Implementation

[0036] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, but the embodiments do not limit the present invention in any way. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in this technical field.

[0037] Unless otherwise specified, all reagents and materials used in the following examples are commercially available.

[0038] cis-4-tert-butylcyclohexanol (98%) was purchased from Shanghai Maclean Biochemical Technology Co., Ltd. trans-4-tert-butylcyclohexanol (99.5%) was purchased from Shanghai Maclean Biochemical Technology Co., Ltd. 4-tert-butylcyclohexanol cis-trans isomers (i.e., mixtures containing cis-trans isomers) were purchased from Dongying Kehong Chemical Co., Ltd.; among them, the detected mass content of cis-4-tert-butylcyclohexanol was 29.31%, and the detected mass content of trans-4-tert-butylcyclohexanol was 70.13%.

[0039] Example 1: A method for separating and detecting the cis-trans isomers of 4-tert-butylcyclohexanol S1. Draw the standard working curve. S1-1 Preparation of internal standard solution: Weigh 0.2 g of internal standard benzyl alcohol into a 20 mL volumetric flask using an analytical balance, dilute to the mark with ethanol, mix well, seal and shake to obtain a 10 g / L benzyl alcohol-ethanol solution, which is the internal standard solution. S1-2. Preparation of mixed isomer standard working solution containing internal standard: Weigh 0.0513 g of cis- and trans-4-tert-butylcyclohexanol standards into 10 mL volumetric flasks using an analytical balance. Dilute to the mark with ethanol, mix well, seal, and shake to obtain a standard working solution with a total concentration of 10.26 g / L. Dilute this solution at different ratios to obtain a series of standard working solutions with actual concentrations of 1.14 g / L, 2.16 g / L, 3.96 g / L, 6.02 g / L, and 10.26 g / L, respectively. Mix 1 mL of each of the above standard working solutions with 1 mL of the internal standard solution prepared in step S1-1 to obtain a mixed isomer standard working solution containing internal standard (internal standard concentration of 5 g / L), wherein the concentrations of cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol are 0.57 g / L, 1.08 g / L, and 1.98 g / L, respectively. g / L, 3.01 g / L, 5.13 g / L; S1-3. Construction of Standard Working Curve: The mixed isomer standard working solutions containing different concentrations of internal standard obtained in step S1-2 were mixed thoroughly, centrifuged, and 1.0 μL of the supernatant was taken as the sample solution to be tested. Gas chromatography was performed using a Fuli GC9790II gas chromatograph. A Fuli RB-5 gas chromatograph column (30 m × 0.32 mm × 0.50 μm, stationary phase: 5% phenyl-95% dimethyl polysiloxane) was used in splitless injection mode to achieve chromatographic separation of cis- and trans-4-tert-butylcyclohexanol. The gas chromatographic parameters were as follows: temperature program: initial temperature 50 ℃, hold for 1 min, then increase to 220 ℃ at 10 ℃ / min, hold for 2 min; carrier gas: nitrogen, carrier gas flow rate: 4.0 mL / min; injection port temperature: 200 ℃; detector: detection temperature: 220 ℃. Using a flame ionization detector (FID) at ℃, standard working curves for cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol were plotted with the concentration ratio (concentration of any isomer / concentration of internal standard) as the x-axis and the peak area ratio (peak area of ​​any isomer / peak area of ​​internal standard) as the y-axis.

[0040] The experimental results of the standard working curve are shown in Tables 1-2 and 2. Figures 1-2 As shown: Table 1 Standard curves for cis-4-tert-butylcyclohexanol

[0041] Table 2 Standard Curve for Trans-4-tert-Butylcyclohexanol

[0042] The linear equation for cis-4-tert-butylcyclohexanol is y = 0.9791x + 0.02, with a correlation coefficient R0. 2 The value is 0.9998. For specific data and standard working curves, please refer to Table 1 and [Table 2]. Figure 1 The linear equation for trans-4-tert-butylcyclohexanol is y = 1.0249x + 0.0198, with a correlation coefficient R0. 2 The value is 0.9998. For specific data and standard working curves, please refer to Table 2 and [Table 2]. Figure 2 .

[0043] S2, Sample Testing S2-1. Weigh 0.01 g of the cis-trans isomer of 4-tert-butylcyclohexanol and dissolve it in 1.0 mL of anhydrous ethanol (i.e., the concentration of the test sample solution is 10 g / L). Take 1 mL of the test sample solution and 1 mL of the internal standard solution (10 g / L) and mix them well to obtain the test solution. After centrifugation, take the supernatant and inject and detect it according to the conditions of step S1-3 above to obtain the peak area. S2-2. Substitute the peak area obtained in step S2-1 into the standard working curve obtained in step S1 to calculate the contents of cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol, respectively.

[0044] The detection results of Example 1 are as follows Figure 3 As shown, the retention time of cis-4-tert-butylcyclohexanol was 10.970 min, and the retention time of trans-4-tert-butylcyclohexanol was 11.210 min. By substituting the peak areas of the two isomers into the standard curve, the mass content of trans-4-tert-butylcyclohexanol in the sample was calculated to be 70.4%, and the mass content of cis-4-tert-butylcyclohexanol was 29.6%, which is almost consistent with the manufacturer's detection results (the detected mass content of cis-4-tert-butylcyclohexanol was 29.31%, and the detected mass content of trans-4-tert-butylcyclohexanol was 70.13%). This indicates that the separation and detection method of this application can achieve the separation of the cis and trans isomers of 4-tert-butylcyclohexanol, and the quantitative detection of the isomers.

[0045] Example 2: A method for separating and detecting the cis-trans isomers of 4-tert-butylcyclohexanol S1. Weigh 0.05 g each of cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol standards and dissolve them in 10.0 mL of anhydrous ethanol (i.e., the concentration of the test sample solution is 10 g / L). Take 1 mL of the test sample solution and 1 mL of the internal standard solution (10 g / L) and mix them well to obtain the test solution. After centrifugation, take the supernatant and inject and detect it according to the conditions of Example 1 to obtain the peak area. S2. Substitute the peak area obtained in step S1 into the standard working curve obtained in Example 1 to calculate the contents of cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol, respectively.

[0046] The detection results of Example 2 are as follows Figure 4 As shown: the retention time of solvent ethanol was 1.358 min, the retention time of internal standard benzyl alcohol was 8.088 min, the retention time of cis-4-tert-butylcyclohexanol was 11.078 min, and the retention time of trans-4-tert-butylcyclohexanol was 11.313 min. These results indicate that the separation and detection method of this application can effectively separate the cis and trans isomers of 4-tert-butylcyclohexanol. Specific peak area data are as follows: cis-4-tert-butylcyclohexanol peak area was 354453, trans-4-tert-butylcyclohexanol peak area was 338960.3, and the internal standard benzyl alcohol peak area was 1184389.7.

[0047] Example 3: A method for separating and detecting the cis-trans isomers of 4-tert-butylcyclohexanol The difference from Example 2 is that the heating program is different in the detection conditions. The specific heating program is changed to the following: initial temperature 45 ℃, hold for 2 min, then increase the temperature to 220 ℃ at 8 ℃ / min, and hold for 3 min.

[0048] The other steps and conditions are the same as in Example 2.

[0049] The detection results of Example 3 are basically the same as those of Example 2, both of which can effectively separate the cis-trans isomers of 4-tert-butylcyclohexanol, which will not be repeated here.

[0050] Example 4: Spiking recovery and detection limit test of a cis-trans isomer of 4-tert-butylcyclohexanol 1. Experimental Methods Internal standard solution: Weigh 0.2 g of benzyl alcohol into a 20 mL volumetric flask using an analytical balance, dilute to the mark with ethanol, mix well, seal and shake to obtain a 10 g / L benzyl alcohol-ethanol solution; Reference solutions: Accurately weigh cis / trans-4-tert-butylcyclohexanol reference standards, dissolve and dilute with ethanol to prepare standard solutions with concentration gradients of 0.57 g / L, 2.85 g / L, and 5.13 g / L; Test solution: Weigh 0.05 g of 4-tert-butylcyclohexanol cis-trans isomer sample and dissolve it in 10.0 mL of anhydrous ethanol (i.e., the concentration of the test sample solution is 5 g / L). Take 1 mL of the test sample solution and 1 mL of internal standard solution (10 g / L) and vortex mix for 1 min. Spiking design: Reference solution was added to the test sample solution to make the target isomer spiked concentration in the final solution 0.57 g / L, 2.85 g / L and 5.13 g / L. Three copies of each concentration were prepared in parallel and the chromatographic conditions of Example 1 were used for determination. The experimental results are shown in Table 3.

[0051] 2. Experimental Results Table 3 Recovery rates of cis / trans-4-tert-butylcyclohexanol

[0052] As shown in Table 3, within a concentration range of 0.57–5.13 g / L, the average recoveries of the cis-trans isomers of 4-tert-butylcyclohexanol were 97.6%–102.3%, with a relative standard deviation (RSD) ≤1.8%. These results indicate that the proposed detection method possesses high accuracy and good repeatability, meeting the requirements for practical quantitative detection.

[0053] Table 4. Detection limits for cis-trans isomers of 4-tert-butylcyclohexanol

[0054] In this application, the detection limit for the cis-trans isomer of 4-tert-butylcyclohexanol was calculated using the signal-to-noise ratio method. As shown in Table 4, the detection limit for the cis-trans isomer of 4-tert-butylcyclohexanol in Example 1 was 0.32 g / L; values ​​above this are suitable for qualitative analysis. The quantification limit was 0.57 g / L; values ​​above this exhibited typical Gaussian peak shapes, and both peak height and peak area were suitable for accurate quantification. Figures 3-4 In contrast, when the concentration of the cis-trans isomer of 4-tert-butylcyclohexanol was 0.25 g / L ( Figure 5 Within the expected retention time range of the target analyte (retention time 8.171 min for the internal standard benzyl alcohol, and retention time for the cis-trans isomer of 4-tert-butylcyclohexanol 11-12 min), only weak, diffuse signal fluctuations were observed, with no obvious chromatographic peaks. The instrument's standard integration parameters could not automatically identify effective chromatographic peaks. Even with manual baseline setting, the peak area varied greatly with repeated injections, and the linear relationship with concentration was extremely poor (R0). 2 If the value is less than 0.5, it indicates that the signal is unreliable noise or interference, and the existence of the target object cannot be confirmed, let alone quantitative analysis.

[0055] Comparative Example 1: A method for separating and detecting the cis-trans isomers of 4-tert-butylcyclohexanol. The difference from Example 2 is that, in the gas chromatography conditions, the Fuli RB-5 gas chromatographic column was replaced with a C-18 reversed-phase column (Agilent ZORBAX Eclipse XDB-C18, specifications: 4.6 m × 150 mm, 5 μm). The specific detection steps are as follows: Weigh 0.05 g each of cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol standards and dissolve them in 10.0 mL of anhydrous ethanol (i.e., the concentration of the test sample solution is 10 g / L). Take 1 mL of the test sample solution and 1 mL of the internal standard solution (10 g / L) and mix them well to obtain the test solution. After centrifugation, take the supernatant for injection and detection.

[0056] The detection results of Comparative Example 1 are as follows: When using the C-18 column, the target isomers (cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol) could not be effectively retained and separated, and no target peak was detected within a reasonable retention time. Based on this result, it can be determined that the physicochemical properties of the target substance are not suitable for analysis using reversed-phase liquid chromatography mode. Therefore, no further attempt was made to establish a standard working curve based on the C-18 reversed-phase column.

[0057] Comparative Example 2: A method for separating and detecting the cis-trans isomers of 4-tert-butylcyclohexanol The difference from Example 2 is that, in the gas chromatography conditions, the Fuli RB-5 gas chromatographic column was replaced with a DB-624 column (stationary phase: 6% cyanopropylphenyl-94% dimethyl polysiloxane, dimensions: 30 m × 0.32 mm, 1.8 μm). The specific detection steps are as follows: Weigh 0.05 g each of cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol standards and dissolve them in 10.0 mL of anhydrous ethanol (i.e., the concentration of the test sample solution is 10 g / L). Take 1 mL of the test sample solution and 1 mL of the internal standard solution (10 g / L) and mix them well to obtain the test solution. After centrifugation, take the supernatant for injection and detection.

[0058] The detection results of Comparative Example 2 are as follows: Although the DB-624 column can produce peaks, the peaks are significantly broadened, have severe tailing, and have insufficient resolution. This indicates that the DB-624 column has insufficient selectivity for the target analyte and cannot meet the requirements of high resolution and sharp peak shape. Therefore, no further attempt was made to establish a standard working curve based on the DB-624 reversed-phase column.

[0059] In summary, this invention successfully achieved efficient separation of the cis-trans isomers of 4-tert-butylcyclohexanol using an RB-5 column under specific chromatographic conditions, and achieved quantitative analysis by combining the internal standard method. Experimental results show that the linearity of the standard working curve reached 99.9% (R²). 2 The spiked recoveries ranged from 97.6% to 102.3%, and the relative standard deviation (RSD) for repeated measurements (n=5) was ≤1.8%, indicating good repeatability and accuracy. When determining the content of cis-trans isomers of 4-tert-butylcyclohexanol in a product, effective separation and detection of the cis-trans isomers can be achieved as long as the dissolution of the target isomer is ensured and the peak times of impurities in the product do not overlap with those of the target isomer.

[0060] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. A method for separating and detecting the cis-trans isomers of 4-tert-butylcyclohexanol, characterized in that, The steps include: adding an internal standard to the sample solution to be tested, mixing well to obtain a test solution, detecting the obtained test solution by gas chromatography, and quantifying by the internal standard method; The conditions for the gas chromatography method include: Chromatographic column: RB-5 gas chromatographic column; Temperature program: 45~55 ℃, hold for 1~5 min, increase temperature at 5~15 ℃ / min to 200~230 ℃, hold for 1~5 min; The internal standard is benzyl alcohol.

2. The separation and detection method according to claim 1, characterized in that, The internal standard method for quantification includes the following steps: S1. Prepare a mixed isomer standard working solution containing an internal standard; S2. Prepare the test solution; S3. Detect the mixed isomer standard working solution containing internal standard obtained in step S1 by gas chromatography and plot the standard working curve; S4. The test solution obtained in step S2 is detected by gas chromatography to obtain the peak area. Based on the standard working curve obtained in step S3, the contents of cis-4-tert-butylcyclohexanol and trans-4-tert-butylcyclohexanol are calculated. In step S2, the definition of the test solution is consistent with that in claim 1.

3. The separation and detection method according to claim 2, characterized in that, The horizontal axis of the standard working curve represents the ratio of the concentration of any isomer to the concentration of the internal standard, and the vertical axis represents the ratio of the peak area of ​​any isomer to the peak area of ​​the internal standard.

4. The separation and detection method according to claim 1 or 2, characterized in that, The solvents of the test solution or the mixed isomer standard working solution containing the internal standard are each independently selected from one or more of ethanol, ethyl acetate, and tetrahydrofuran.

5. The separation and detection method according to claim 1 or 2, characterized in that, The concentration of the internal standard in the test sample solution or the mixed isomer standard working solution containing the internal standard is 5~6.8 g / L.

6. The separation and detection method according to claim 1, characterized in that, The stationary phase of the chromatographic column is 5% phenyl-95% dimethylpolysiloxane.

7. The separation and detection method according to claim 1, characterized in that, The sample solution to be tested is obtained by fully dissolving the analyte in a solvent.

8. The separation and detection method according to claim 1, characterized in that, The detector used in the gas chromatography is a flame ionization detector.

9. The application of the separation and detection method according to any one of claims 1 to 8 in determining the content of 4-tert-butylcyclohexanol cis-trans isomers in a product.

10. The application according to claim 9, characterized in that, The products mentioned include cosmetics, pharmaceuticals, or food.

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