A gas phase fingerprint of trans-seed and a construction method and application thereof

Abstract

The application discloses a gas phase fingerprint of transverse and longitudinal seat, which comprises peaks No.1 to No.5, the retention time of the peaks No.1 to No.5 is 16.89±0.1 min, 17.36±0.1 min, 17.89±0.1 min, 19.37±0.1 min and 19.6±0.1 min respectively; the construction method of the gas phase fingerprint of the transverse and longitudinal seat is as follows: the transverse and longitudinal seat is crushed to obtain transverse and longitudinal seat powder, then water is added into the transverse and longitudinal seat powder, and then ethyl acetate is added, and then micro-boiling is carried out for 4 hours, then natural cooling is carried out to room temperature, and then layering is carried out after standing, and the ethyl acetate layer is collected as a test solution; liquid chromatography detection is carried out on the test solution; more than 10 kinds of transverse and longitudinal seat medicinal materials are collected, and the detection operation is carried out according to the above method, the obtained detection results are compared, and the common characteristic peaks are selected to obtain the gas phase fingerprint of the transverse and longitudinal seat. The gas phase fingerprint constructed by the application can be used for quickly and accurately determining and identifying the transverse and longitudinal seat medicinal material.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of detection of Calophyllum inophyllum L., and particularly relates to a gas phase fingerprint of Calophyllum inophyllum L. and a construction method and application thereof. BACKGROUND

[0002] Calophyllum inophyllum L., the scientific name of which is Calophyllum membranaceum Gardn. et Champ, is a plant belonging to the family Guttiferae and the genus Calophyllum, and is commonly known as thin-leaf tallow tree. The plant mainly grows in Guangdong and Guangxi and is a common Chinese herbal medicine. The roots and leaves of the plant can be used as medicine and have the functions of dispelling wind and dampness, strengthening bones and muscles, and activating blood and relieving pain.

[0003] Calophyllum inophyllum L. has certain similarities in appearance with other plant varieties belonging to the same genus, such as Jatropha gossypiifolia and Jatropha curcas. These plant varieties have similarities in leaf shape, surface characteristics, and powder microscopic characteristics, which makes it difficult to accurately identify Calophyllum inophyllum L. In order to effectively identify Calophyllum inophyllum L., researchers usually adopt the method of thin layer chromatography determination. However, the fixed phase length of the thin layer chromatography plate is limited, which limits its separation capacity. At the same time, since the thin layer chromatography is an open system, it is easily affected by external factors such as humidity and temperature, which in turn affects the accuracy of the determination, so there is still a need to develop a simple, rapid, reproducible, and accurate detection method suitable for Calophyllum inophyllum L. SUMMARY

[0004] In view of the above problems, the application discloses a gas phase fingerprint of Calophyllum inophyllum L. and a construction method thereof, and the gas phase fingerprint can be used to quickly and accurately determine and identify Calophyllum inophyllum L.

[0005] The application is implemented by adopting the following technical scheme:

[0006] The gas phase fingerprint of Calophyllum inophyllum L. comprises peaks 1-5, and the retention times of the peaks 1-5 are 16.89±0.1 min, 17.36±0.1 min, 17.89±0.1 min, 19.37±0.1 min, and 19.6±0.1 min, respectively.

[0007] The construction method of the gas phase fingerprint of Calophyllum inophyllum L. comprises the following steps:

[0008] (1) Calophyllum inophyllum L. is crushed and passed through a 50-mesh sieve to obtain Calophyllum inophyllum L. powder, then water is added to the Calophyllum inophyllum L. powder, followed by the addition of ethyl acetate, and then micro-boiling is performed for 4 h, followed by natural cooling to room temperature and static stratification, and the ethyl acetate layer is collected as a test solution;

[0009] (2) Take the test solution obtained in step (1) to carry out liquid chromatography detection, and the conditions of the liquid chromatography detection are as follows: a ZB-264 chromatographic column is selected, the carrier gas flow is 1 mL / min, the split ratio is 25:1, the injection amount is 1 μL, the temperature of the injection port is 250 DEG C, and the temperature of the FID detector is 250 DEG C; the temperature program of the chromatographic column is as follows: the initial temperature is 80 DEG C, is kept for 2 min, is raised to 250 DEG C at a rate of 10 DEG C / min, and then is kept for 6 min;

[0010] (3) More than 10 kinds of horizontal seat medicine are collected, and the detection operation is carried out according to the method described in steps (1)-(2), the detection results obtained are compared, and the common characteristic peaks are selected to obtain the gas chromatography fingerprint of horizontal seat.

[0011] Further, in step (1), the mass of horizontal seat powder and the volume of water are 20g:800mL, and the mass of horizontal seat powder and the volume of ethyl acetate are 20g:2mL.

[0012] Further, in step (1), the temperature in the micro-boiling process is controlled at 80-100 DEG C.

[0013] Further, in step (2), the size of the ZB-264 chromatographic column is 30m*0.25mm*1.40 μm. The ZB-264 chromatographic column is a chromatographic column produced by the American Fenome Phenomenex Company.

[0014] The application of the gas chromatography fingerprint of horizontal seat is used for the identification of horizontal seat medicine, specifically, the medicine to be identified is detected according to the method described in steps (1)-(2), and then the gas chromatography results obtained are compared with the gas chromatography fingerprint of horizontal seat, if the peaks 1-5 are present, it can be preliminarily determined that the medicine to be tested is horizontal seat.

[0015] The technical scheme has the following beneficial effects compared with the prior art:

[0016] The gas chromatography fingerprint of horizontal seat medicine is constructed by the gas chromatography method, then the medicine sample is detected according to the method of the application, the results obtained are compared with the constructed gas chromatography fingerprint, the authenticity and quality of the horizontal seat medicine can be quickly and accurately identified, the medicine raw materials that do not meet the quality requirements can be screened out, the quality control of horizontal seat medicine is realized, and the safety and stability of the use of horizontal seat medicine are improved. BRIEF DESCRIPTION OF DRAWINGS

[0017] Figure 1is the gas chromatogram of No. 1 Hengjingxi medicinal material sample detected in Example 1.

[0018] Figure 2 is the gas chromatogram of No. 2 Hengjingxi medicinal material sample detected in Example 1.

[0019] Figure 3 is the gas chromatogram of No. 3 Hengjingxi medicinal material sample detected in Example 1.

[0020] Figure 4 is the gas chromatogram of No. 4 Hengjingxi medicinal material sample detected in Example 1.

[0021] Figure 5 is the gas chromatogram of No. 5 Hengjingxi medicinal material sample detected in Example 1.

[0022] Figure 6 is the gas chromatogram of No. 6 Hengjingxi medicinal material sample detected in Example 1.

[0023] Figure 7 is the gas chromatogram of No. 7 Hengjingxi medicinal material sample detected in Example 1.

[0024] Figure 8 is the gas chromatogram of No. 8 Hengjingxi medicinal material sample detected in Example 1.

[0025] Figure 9 is the gas chromatogram of No. 9 Hengjingxi medicinal material sample detected in Example 1.

[0026] Figure 10 is the gas chromatogram of No. 10 Hengjingxi medicinal material sample detected in Example 1.

[0027] Figure 11 is the gas chromatogram of No. 11 Hengjingxi medicinal material sample detected in Example 1.

[0028] Figure 12 is the gas chromatogram obtained when using anhydrous ethanol solvent for detection in Experimental Example 1.

[0029] Figure 13 is the gas chromatogram obtained when using cyclohexane solvent for detection in Experimental Example 1.

[0030] Figure 14 is the gas chromatogram obtained when using petroleum ether solvent for detection in Experimental Example 1.

[0031] Figure 15 is the gas chromatogram obtained when using ethyl acetate solvent for detection in Experimental Example 1.

[0032] Figure 16 is the gas chromatogram obtained when using HP-1 chromatographic column for detection in Experimental Example 2.

[0033] Figure 17 is a gas chromatogram obtained when the ZB-624 column was used for detection in Experimental Example 2.

[0034] Figure 18 is a gas chromatogram obtained when the ZB-WAXplus column was used for detection in Experimental Example 2.

[0035] Figure 19 is a gas chromatogram obtained when the blank control was detected in Experimental Example 3.

[0036] Figure 20 is a gas chromatogram obtained when Parallel Sample 1 was detected in Experimental Example 3.

[0037] Figure 21 is a gas chromatogram obtained when Parallel Sample 2 was detected in Experimental Example 3.

[0038] Figure 22 is a gas chromatogram obtained when Parallel Sample 3 was detected in Experimental Example 3.

[0039] Figure 23 is a gas chromatogram obtained when Parallel Sample 4 was detected in Experimental Example 3.

[0040] Figure 24 is a gas chromatogram obtained when Parallel Sample 5 was detected in Experimental Example 3.

[0041] Figure 25 is a gas chromatogram obtained when Parallel Sample 6 was detected in Experimental Example 3.

[0042] Figure 26 is a gas chromatogram obtained when Parallel Sample 7 was detected in Experimental Example 3.

[0043] Figure 27 is a gas chromatogram obtained when Experimentalist 1 was detected in Experimental Example 4.

[0044] Figure 28 is a gas chromatogram obtained when Experimentalist 2 was detected in Experimental Example 4.

[0045] Figure 29 is a gas chromatogram obtained when the same Hengxishi medicinal material sample was detected on the first day in Experimental Example 5.

[0046] Figure 30 is a gas chromatogram obtained when the same Hengxishi medicinal material sample was detected on the second day in Experimental Example 5.

[0047] Figure 31 is a gas chromatogram obtained when the same Hengxishi medicinal material sample was detected on the third day in Experimental Example 5.

[0048] Figure 32is the gas chromatogram obtained when the same sample of Hengjingxi medicinal material is detected on the fourth day in Experimental Example 5.

[0049] Figure 33 is the gas chromatogram obtained when the same sample of Hengjingxi medicinal material is detected on the fifth day in Experimental Example 5. DETAILED DESCRIPTION

[0050] The present application is further illustrated by the following examples, but not as a limitation to the present application. The specific experimental conditions and methods not mentioned in the following examples are generally the conventional methods well known to those skilled in the art.

[0051] Example 1: A gas phase fingerprint of Hengjingxi, which comprises peaks No. 1-5, the retention time of the peaks No. 1-5 is 16.89±0.1 min, 17.36±0.1 min, 17.89±0.1 min, 19.37±0.1 min, 19.6±0.1 min, respectively.

[0052] The method for constructing the gas phase fingerprint of Hengjingxi, which comprises the following steps:

[0053] (1) Hengjingxi is crushed and passed through a 50-mesh sieve to obtain Hengjingxi powder, then water is added to the Hengjingxi powder, followed by the addition of ethyl acetate, and then micro-boiling is carried out for 4 h, followed by natural cooling to room temperature and standing for layering, and the ethyl acetate layer is collected as the test solution; the temperature in the micro-boiling process is controlled at 90±2℃; the mass of the Hengjingxi powder and the volume of water are 20 g:800 mL, and the mass of the Hengjingxi powder and the volume of ethyl acetate are 20 g:2 mL;

[0054] (2) The test solution obtained in step (1) is subjected to liquid chromatography detection, and the conditions for the liquid chromatography detection are as follows: a ZB-264 chromatographic column is selected, the carrier gas flow rate is 1 mL / min, the split ratio is 25:1, the injection amount is 1 μL, the temperature of the injection port is 250℃, and the temperature of the FID detector is 250℃; the chromatographic column temperature program is that the initial temperature is 80℃, which is maintained for 2 min, then increased to 250℃ at a rate of 10℃ / min, and then maintained for 6 min; the size of the ZB-264 chromatographic column is 30 m x 0.25 mm x 1.40 μm; and the detection instrument model is Agilent 7890A;

[0055] (3) 11 kinds of Hengjingxi medicinal materials (see Table 1) are collected and subjected to detection operation according to the methods described in steps (1)-(2), and the obtained detection results (see Table 2 and Figures 1-11 ) are compared to select common characteristic peaks to obtain the gas phase fingerprint of Hengjingxi.

[0056] The application of the gas chromatography fingerprint of the Hengjingxi described in the embodiment is used for identification of Hengjingxi medicinal materials. Specifically, the method in steps (1)-(2) is used to detect the medicinal materials to be identified, and then the gas chromatography results obtained are compared with the gas chromatography fingerprint of the Hengjingxi. If the peaks 1-5 are present, it can be preliminarily determined that the medicinal materials to be detected are Hengjingxi.

[0057] Table 1 Collection table of different Hengjingxi medicinal materials

[0058]

[0059] Table 2 Retention time results (min) of detection of different Hengjingxi medicinal materials

[0060]

[0061] Experimental Example 1: The same Hengjingxi medicinal material sample was detected according to the method of Example 1 and the following three methods, and the detection results are shown in Table 3 and Figures 12-15 .

[0062] Method 1: Hengjingxi was crushed to obtain Hengjingxi powder, which was added with anhydrous ethanol, shaken for 30 minutes, ultrasonically treated for 30 minutes, and filtered to obtain the filtrate as a test sample solution. The mass of the Hengjingxi powder and the volume of the anhydrous ethanol were 1 g:5 mL.

[0063] Method 2: Hengjingxi was crushed to obtain Hengjingxi powder, which was added with cyclohexane, shaken for 30 minutes, ultrasonically treated for 30 minutes, and filtered to obtain the filtrate as a test sample solution. The mass of the Hengjingxi powder and the volume of the cyclohexane were 1 g:5 mL.

[0064] Method 3: Hengjingxi was crushed to obtain Hengjingxi powder, which was added with petroleum ether, shaken for 30 minutes, ultrasonically treated for 30 minutes, and filtered to obtain the filtrate as a test sample solution. The mass of the Hengjingxi powder and the volume of the petroleum ether were 1 g:5 mL.

[0065] Table 3 Retention time results (min) of detection by different methods

[0066]

[0067] According to the results in Table 3 and Figures 12-15 , it can be seen that only the method of the present application can obtain good peak effect, thereby realizing the detection and identification of Hengjingxi.

[0068] Experimental Example 2: The same *Platycladus orientalis* herb sample was tested according to steps (1) and (2) in Example 1, using different chromatographic columns: HP-1 (30m × 0.25mm × 0.25μm), ZB-624 (30m × 0.25mm × 1.40μm), and ZB-WAXplus (30m × 0.25mm × 0.25μm). The test results are shown in [reference needed]. Figures 16-18 .according to Figures 16-18 The results show that the ZB-624 column selected in this invention has better separation and no peak tailing.

[0069] Experimental Example 3: Following steps (1) and (2) of Example 1, samples of the herb *Phyllostachys edulis* were tested and divided into 7 equal portions for parallel testing. A test solution without *Phyllostachys edulis* samples was used as a blank control. The test results are shown in Table 4. Figures 19-26 .

[0070] Table 4. Results of Parallel Tests

[0071]

[0072] According to Table 4 and Figures 19-26 The results show that the method of the present invention has good specificity, and the method is stable and has good repeatability.

[0073] Experiment Example 4: Two researchers were assigned to use two instruments of the same model to test the same transverse-meridian herbal sample according to steps (1) and (2) in Example 1. The test results are shown in Table 5 and 6. Figures 27-28 .

[0074] Table 5. Retention time results (min) detected by different experimenters

[0075]

[0076] According to Table 5 and Figures 27-28 The results show that the repeatability and accuracy of the tests performed by different researchers using the method of this invention are good.

[0077] Experimental Example 5: Following the methods described in steps (1) and (2) of Example 1, the same transverse meridian herbal sample was tested for 5 consecutive days. The test results are shown in Table 6 and... Figures 29-33 .

[0078] Table 6 Results of testing over 5 consecutive days

[0079]

[0080] According to Table 6 and Figures 29-33 The results show that the present invention has good stability.

[0081] Furthermore, it should be understood that although the specification is described in terms of embodiments, not every embodiment includes only one independent technical solution, and the specification is described in this way only for the sake of clarity, and those skilled in the art should consider the specification as a whole, and the technical solutions in each embodiment can also be appropriately combined to form other embodiments that those skilled in the art can understand.

Claims

1. A method for constructing a vapor phase fingerprint spectrum of transverse latitude, characterized in that: The vapor phase fingerprint spectrum of the transverse meridian includes peaks 1 to 5, and the retention times of peaks 1 to 5 are 16.89±0.1 min, 17.36±0.1 min, 17.89±0.1 min, 19.37±0.1 min, and 19.6±0.1 min, respectively. The method for constructing the vapor phase fingerprint spectrum of the transverse meridian sheet includes the following steps: (1) Take the transverse warp mat, crush it and pass it through a 50-mesh sieve to obtain transverse warp mat powder. Then add water to the transverse warp mat powder, then add ethyl acetate, then boil gently for 4 hours, then cool naturally to room temperature and let it stand to separate into layers. Collect the ethyl acetate layer as the test solution. (2) Take the test solution obtained in step (1) and perform liquid chromatography detection. The liquid chromatography detection conditions are as follows: select ZB-624 column, carrier gas flow rate is 1 mL / min, split ratio is 25:1, injection volume is 1 μL, injection port temperature is 250℃, and FID detector temperature is 250℃; the column temperature program is as follows: initial temperature is 80℃, hold for 2 min, then increase to 250℃ at a rate of 10℃ / min, and then hold for 6 min. (3) Collect more than 10 kinds of transverse meridian herbs and perform detection operations according to the methods described in steps (1) to (2). Compare the obtained detection results and select the common characteristic peaks to obtain the gas phase fingerprint spectrum of transverse meridian herbs.

2. The method for constructing the vapor phase fingerprint spectrum of the transverse laminar flow pattern according to claim 1, characterized in that: In step (1), the mass ratio of the transverse warp powder to the volume ratio of water is 20g:800mL, and the mass ratio of the transverse warp powder to the volume ratio of ethyl acetate is 20g:2mL.

3. The method for constructing the vapor phase fingerprint spectrum of the transverse laminar flow pattern according to claim 1, characterized in that: The temperature during the micro-boiling process described in step (1) is controlled at 80-100℃.

4. The method for constructing a vapor phase fingerprint spectrum of a transverse diaphragm according to claim 1, characterized in that: The ZB-624 chromatographic column mentioned in step (2) has a size of 30m × 0.25mm × 1.40μm.

5. The application of the method for constructing a vapor phase fingerprint spectrum of a transverse laminar flow as described in claim 1, characterized in that: The gas chromatogram of the transverse meridian is used for the identification of transverse meridian Chinese medicinal materials. Specifically, the medicinal material to be identified is tested according to the method in steps (1) to (2). Then, the obtained gas chromatogram results are compared with the gas chromatogram of the transverse meridian. If peaks 1 to 5 are present, the medicinal material to be tested can be preliminarily determined to be transverse meridian.

Images