Method for detecting content of effective components in gibberellic acid soluble concentrate

Abstract

The invention relates to the technical field of chemical detection, and particularly discloses a method for detecting the content of effective components of a gibberellic acid soluble concentrate. The detection method comprises the following steps: weighing 20 [mu] L of a solution and 20 [mu] L of a reference substance solution, respectively injecting the solutions into a liquid chromatograph, carrying out determination by taking a composite filler as a filler and a potassium dihydrogen phosphate-methanol mixed solution as a mobile phase under the conditions that the detection wavelength is 284 nm, the column temperature is 30 DEG C and the flow velocity is 1.0 ml per minute, recording a chromatogram, and carrying out gradient elution. And calculating the content of 5-hydroxymethylfurfural by peak area according to an external standard method. The detection method provided by the invention is good in detection accuracy and stability.

Description

Technical Field

[0001] The present application relates to the technical field of chemical detection, and more specifically, to a method for detecting the content of effective ingredients in a soluble solution of epi-rutin gibberellic acid. Background Art

[0002] Gibberellic acid has many functions, such as promoting cell elongation, preserving flowers and fruits, breaking seed dormancy, inducing parthenocarpy, increasing fruit setting rate, enlarging fruits, and increasing yield. Brassinolide is known as the sixth plant growth regulator, which has the functions of promoting germination, root growth, plant growth, preserving flowers and fruits, and increasing yield. When the two are mixed in a certain proportion, the synergistic effect is very significant, which can significantly improve the drought and cold resistance of the plant, and the yield-increasing effect is very significant. The epi-brassic gibberellic acid soluble solution mixed with gibberellins and brassinolide can promote the growth of vegetables, melons, fruits and other crops, improve quality, increase yield, make the color brighter, and make the leaves thicker. It can also advance the time of tea leaf picking, and make melons and fruits have higher sugar content, larger individuals, higher yield, and more storage resistance. However, the simultaneous detection method for 28-epibrassinolide and gibberellic acid A4+A7 is to perform component analysis only through liquid chromatography, but the existing method has problems such as complex operation, poor accuracy of analysis results and error control, and cannot meet the company's needs for precise control of product quality. Summary of the invention

[0003] In order to solve the problems of low detection accuracy and poor stability in the related art, the present application provides a method for detecting the content of effective ingredients in a soluble solution of gibberellic acid.

[0004] The present application provides a method for detecting the content of active ingredients in a soluble solution of epirutin gibberellic acid, which adopts the following technical scheme: A method for detecting the content of active ingredients in a soluble solution of epirutin gibberellic acid, comprising the following steps: Step 1: Weigh 20 μL of epirutin gibberellic acid soluble solution and control solution, concentrate under reduced pressure, dissolve in methanol, add to the boric acid methanol solution and stir for 20 minutes to obtain the test solution and control solution; Step 2: inject the test solution and the reference solution into a liquid chromatograph respectively, use the first composite filler as the filler, use a potassium dihydrogen phosphate-methanol mixed solution as the mobile phase, and perform the determination under the conditions of a detection wavelength of 284 nm, a column temperature of 30° C., and a flow rate of 1.0 ml per minute, record the chromatogram, and calculate the content of 28-epi-brassinolide by the peak area according to the external standard method; Step 3: Inject the test solution and the control solution into a liquid chromatograph respectively, use the second composite filler as the filler, potassium dihydrogen phosphate-methanol mixed solution as the mobile phase, the detection wavelength is 284nm, the column temperature is 30°C, and the flow rate is 2.0ml per minute. The chromatogram is recorded, and the content of gibberellic acid A4+A7 is calculated by the peak area according to the external standard method.

[0005] The detection method provided in the present application can effectively improve the detection accuracy and stability of the active ingredients of epirutin gibberellic acid soluble solution.

[0006] Preferably, the reference solution contains 10 μg of 28-epibrassinolide solution and 10 μg of gibberellic acid A4+A7 solution per 1 ml.

[0007] The present application can ensure the accuracy and stability of detection in the subsequent comparative detection process by limiting the content of the reference solution.

[0008] Preferably, the potassium dihydrogen phosphate-methanol mixed solution is a mixed solution of potassium dihydrogen phosphate solution and methanol solution, and the volume ratio of the potassium dihydrogen phosphate solution to the methanol solution is 3-5:1.

[0009] The present application can ensure that the mobile phase has good performance by limiting the volume ratio of the potassium dihydrogen phosphate solution and the methanol solution in the potassium dihydrogen phosphate-methanol mixed solution, thereby improving the accuracy and stability of the detection.

[0010] Preferably, the concentration of the potassium dihydrogen phosphate solution is 0.04-0.06 mol / L.

[0011] The present application can ensure the stability of the performance of the mobile phase by limiting the concentration of the potassium dihydrogen phosphate solution, thereby avoiding the influence of the mobile phase on the overall detection effect.

[0012] Preferably, the composite filler is an octadecylsilane bonded silica filler and a naloxone bonded silica filler.

[0013] The present application uses octadecylsilane bonded silica gel filler and naloxone bonded silica gel filler as composite filler, which can effectively separate polar and non-polar components without tailing, with high column efficiency and sensitive analysis.

[0014] Preferably, the mass ratio of octadecylsilane bonded silica filler to naloxone bonded silica filler in the first filler is 1:2-3, and the mass ratio of octadecylsilane bonded silica filler to naloxone bonded silica filler in the second filler is 1:3-4.

[0015] The present application can effectively ensure high column separation efficiency and detection accuracy by controlling the mass ratio of octadecylsilane bonded silica gel filler and naloxone bonded silica gel filler.

[0016] Preferably, the particle size of the octadecylsilane bonded silica filler and the naloxone bonded silica filler in the first filler and the second filler is 3-5 μm.

[0017] The present application can ensure high column efficiency and separation degree by limiting the particle sizes of the octadecylsilane bonded silica gel filler and the naloxone bonded silica gel filler in the composite filler.

[0018] Preferably, the preparation method of the naloxone bonded silica filler comprises the following steps: (1) adding silicon spheres to hydrochloric acid, reacting for 2-3 hours, and drying at 110-130° C. to obtain activated silicon spheres for later use; (2) mixing the activated silica spheres with dioxane, hydrochloric acid and triethoxysilane, heating and refluxing for 12-15 hours, washing with a 30% acetone aqueous solution and dioxane, and drying at 110-130° C. to obtain modified silica gel; (3) Add naloxone to the modified silica gel, add chloroform and isopropanol solution of chloroplatinic acid, stir, heat under reflux for 8-10 hours, wash, and dry to obtain.

[0019] The present application can ensure the performance of the composite filler and thus ensure the detection accuracy by defining the preparation method of the naloxone bonded silica gel filler.

[0020] Preferably, the mass molar ratio of the activated silica spheres to triethoxysilane is 1:0.005-0.006 g / mol; the mass volume ratio of the activated silica gel to dioxane is 1:23-25 ​​g / ml.

[0021] The present application can effectively ensure the performance of the naloxone bonded silica filler by limiting the ratio of the active silica spheres, triethoxysilane and dioxane.

[0022] Preferably, the mass molar ratio of the modified silica gel to naloxone is 1:5-8 g / mol, the organic solvent is chloroform, and the molar ratio of chloroplatinic acid to naloxone is 10: -6 -10 -5 :1; the mass volume ratio of the modified silica gel to chloroform is 1:25-45g / mL.

[0023] The present application limits the contents of modified silica gel, naloxone, chloroplatinic acid and chloroform, thereby ensuring that the finally prepared naloxone bonded silica gel filler does not contain by-products, thereby avoiding affecting the final detection.

[0024] In summary, this application has the following beneficial effects: The present application provides a method for detecting the content of effective ingredients in a soluble solution of epibrassic gibberellic acid. The method specifies octadecylsilane bonded silica gel filler and naloxone bonded silica gel filler as composite filler, potassium dihydrogen phosphate-methanol mixed solution as mobile phase, and specifies specific detection wavelength, column temperature and flow rate. The method can detect the content of 28-epibrassicin lactone and gibberellic acid A4+A7 more accurately and stably. DETAILED DESCRIPTION

[0025] The present application is further described in detail below with reference to the embodiments.

[0026] Preparation Example Preparation Example 1 The preparation method of naloxone bonded silica gel filler comprises the following steps: (1) adding 1 part of silicon spheres with an average particle size of 5 μm to 20 parts of 1 mol / L hydrochloric acid, reacting at 100° C. for 2 h, filtering and washing, and drying at 120° C. to obtain activated silicon spheres for later use; (2) Mixing activated silica balls with dioxane, 10 parts of 1 mol / L hydrochloric acid and triethoxysilane, heating and refluxing for 13 hours, washing with 30% acetone aqueous solution and dioxane, and drying at 120° C. to obtain modified silica gel; the mass molar ratio of activated silica balls to triethoxysilane is 1:0.005 g / mol; the mass volume ratio of activated silica gel to dioxane is 1:23 g / m; (3) Adding naloxone to the modified silica gel, and adding chloroform and an isopropanol solution of chloroplatinic acid, stirring, heating under reflux for 8-10 hours, washing, and drying to obtain a naloxone bonded silica gel filler; the mass molar ratio of the modified silica gel to naloxone is 1:7 g / mol, the organic solvent is chloroform, and the molar ratio of chloroplatinic acid to naloxone is 10:1. -6 :1; the mass volume ratio of the modified silica gel to chloroform is 1:35 g / mL.

[0027] Preparation Example 2 The preparation method of the first composite filler comprises the following steps: One part of octadecyl bonded silica gel and two parts of naloxone bonded silica gel prepared in Preparation Example 1 were ultrasonically treated at 200 W for 40 minutes in 150 parts of carbon tetrachloride solvent to prepare a first composite filler.

[0028] Preparation Example 3 The preparation method of the second composite filler comprises the following steps: One part of octadecyl bonded silica gel and four parts of naloxone bonded silica gel prepared in Preparation Example 1 were ultrasonically treated at 200 W for 40 minutes in 150 parts of carbon tetrachloride solvent to prepare a composite filler. Example

[0029] Example 1 The method for detecting the content of active ingredient in epirutin gibberellic acid soluble solution comprises the following steps: Step 1: Weigh 20 μL of epirutin gibberellic acid soluble solution and control solution, concentrate under reduced pressure, dissolve in methanol, add to the boric acid methanol solution and stir for 20 minutes to obtain the test solution and control solution; Step 2: inject the test solution and the reference solution into a liquid chromatograph respectively, use the first composite filler as the filler, use a potassium dihydrogen phosphate-methanol mixed solution as the mobile phase, and perform the determination under the conditions of a detection wavelength of 284 nm, a column temperature of 30° C., and a flow rate of 1.0 ml per minute, record the chromatogram, and calculate the content of 28-epi-brassinolide by the peak area according to the external standard method; Step 3: Inject the test solution and the control solution into a liquid chromatograph respectively, use the second composite filler as the filler, potassium dihydrogen phosphate-methanol mixed solution as the mobile phase, the detection wavelength is 284nm, the column temperature is 30°C, and the flow rate is 2.0ml per minute. The chromatogram is recorded, and the content of gibberellic acid A4+A7 is calculated by the peak area according to the external standard method.

[0030] Detection Methods The detection process of Example 1 was repeated using 4 groups of different test solutions. The test results are shown in Table 1.

[0031] Table 1 Test results of 4 groups of test solutions and reference solutions It can be seen from Example 1 and Table 1 that the method for detecting the content of the active ingredient in the soluble solution of gibberellic acid provided in the present application has good separation. It can also be seen that the detection method provided in the present application has high accuracy and good stability.

[0032] This specific embodiment is merely an explanation of the present application and is not a limitation of the present application. After reading this specification, those skilled in the art may make modifications to the present embodiment without any creative contribution as needed, but such modifications are protected by the patent law as long as they are within the scope of the claims of the present application.

Claims

1. A method for detecting the effective ingredient content of a soluble solution of epirutin gibberellic acid, characterized in that: The following steps are involved: Step 1: Weigh 20 μL of epirutin gibberellic acid soluble solution and control solution, concentrate under reduced pressure, dissolve in methanol, add to the boric acid methanol solution and stir for 20 minutes to obtain the test solution and control solution; Step 2: inject the test solution and the reference solution into a liquid chromatograph respectively, use the first composite filler as the filler, use a potassium dihydrogen phosphate-methanol mixed solution as the mobile phase, and perform the determination under the conditions of a detection wavelength of 284 nm, a column temperature of 30° C., and a flow rate of 1.0 ml per minute, record the chromatogram, and calculate the content of 28-epi-brassinolide by the peak area according to the external standard method; Step 3: Inject the test solution and the control solution into a liquid chromatograph respectively, use the second composite filler as the filler, potassium dihydrogen phosphate-methanol mixed solution as the mobile phase, the detection wavelength is 284nm, the column temperature is 30°C, and the flow rate is 2.0ml per minute. The chromatogram is recorded, and the content of gibberellic acid A4+A7 is calculated by the peak area according to the external standard method.

2. The detection method according to claim 1, characterized in that: The reference solution contains 10 μg of 28-epi-brassinolide solution and 10 μg of gibberellic acid A4+A7 solution per 1 ml.

3. The detection method according to claim 1, characterized in that: The potassium dihydrogen phosphate-methanol mixed solution is a mixed solution of potassium dihydrogen phosphate solution and methanol solution, and the volume ratio of the potassium dihydrogen phosphate solution to the methanol solution is 3-5:

1.

4. The detection method according to claim 3, characterized in that: The concentration of the potassium dihydrogen phosphate solution is 0.04-0.06 mol / L.

5. The detection method according to claim 1, characterized in that: The first composite filler and the second composite filler are octadecylsilane bonded silica filler and naloxone bonded silica filler.

6. The detection method according to claim 5, characterized in that: The mass ratio of the octadecylsilane bonded silica filler to the naloxone bonded silica filler in the first filler is 1:2-3, and the mass ratio of the octadecylsilane bonded silica filler to the naloxone bonded silica filler in the second filler is 1:3-4.

7. The detection method according to claim 5, characterized in that: The particle sizes of the octadecylsilane bonded silica filler and the naloxone bonded silica filler in the first filler and the second filler are 3-5 μm.

8. The detection method according to claim 5, characterized in that: The preparation method of the naloxone bonded silica gel filler comprises the following steps: (1) adding silicon spheres to hydrochloric acid, reacting for 2-3 hours, and drying at 110-130° C. to obtain activated silicon spheres for later use; (2) mixing the activated silica spheres with dioxane, hydrochloric acid and triethoxysilane, heating and refluxing for 12-15 hours, washing with a 30% acetone aqueous solution and dioxane, and drying at 110-130° C. to obtain modified silica gel; (3) Add naloxone to the modified silica gel, add chloroform and isopropanol solution of chloroplatinic acid, stir, heat under reflux for 8-10 hours, wash, and dry to obtain.

9. The detection method according to claim 8, characterized in that: The mass molar ratio of the activated silica spheres to triethoxysilane is 1:0.005-0.006 g / mol; the mass volume ratio of the activated silica gel to dioxane is 1:23-25 ​​g / ml.

10. The detection method according to claim 8, characterized in that: The mass molar ratio of the modified silica gel to naloxone is 1:5-8 g / mol, the organic solvent is chloroform, and the molar ratio of chloroplatinic acid to naloxone is 10 -6 -10 -5 :1; the mass volume ratio of the modified silica gel to chloroform is 1:25-45g / mL.

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