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Method for quantitative detection of endogenous brassinosteroids in plant sample

A quantitative detection method, brassinosterol technology, applied in the field of analytical chemistry, can solve the problems of precious samples that cannot be realized, solvent consumption and high cost, etc., and achieve the effect of small sample size and effective detection method

Inactive Publication Date: 2013-03-20
WUHAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods all put forward very high requirements on the sample volume, most of which require more than 40g fresh weight. Obviously, such a large sample volume cannot be realized for some precious samples. At the same time, it also brings problems such as large solvent consumption and high cost to the sample pretreatment process.

Method used

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  • Method for quantitative detection of endogenous brassinosteroids in plant sample
  • Method for quantitative detection of endogenous brassinosteroids in plant sample
  • Method for quantitative detection of endogenous brassinosteroids in plant sample

Examples

Experimental program
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Effect test

Embodiment 1

[0020] Determination of brassinosterols in rice leaves

[0021] Accurately weigh 1g of rice leaf and place it in a mortar, freeze and grind it into powder with liquid nitrogen, transfer it to a 10mL centrifuge tube, and add the isotope internal standard [ 2 h 3 ] brassinolide and [ 2 h 3 ] Brassinosterone (castasterone) each 4ng, at the same time 5mL acetonitrile placed in the refrigerator at -18 ℃ for 12h extraction. The sample was centrifuged at 10,000 rpm for 10 min, and the supernatant was taken. Add 250 mg of sodium chloride to the supernatant, shake vigorously with a vortex mixer for 1 min to separate the aqueous phase and the organic phase, and discard the aqueous phase. Add 1 g of anhydrous magnesium sulfate to the centrifuge tube to further remove water. Centrifuge at 10,000 rpm for 10 min, and take the supernatant. Then the resulting supernatant was passed through a double-layer solid-phase extraction cartridge (previously activated by 5 mL of acetonitrile) thr...

Embodiment 2

[0030] Determination of Brassinosterols in Rice Roots

[0031] Accurately weigh 1g of rice root and place it in a mortar, freeze and grind it into powder with liquid nitrogen, transfer it to a 10mL centrifuge tube, and add the isotope internal standard [ 2 h 3 ]brassinolide and [ 2 h 3 ] each 4ng of castasterone, and extract with 5mL of acetonitrile in a refrigerator at -20°C for 12h. The sample was centrifuged at 10,000 rpm for 10 min, and the supernatant was taken. Add 250 mg of sodium chloride to the supernatant, shake vigorously with a vortex mixer for 1 min to separate the aqueous phase and the organic phase, and discard the aqueous phase. Add 1 g of anhydrous magnesium sulfate to the centrifuge tube to further remove water. Centrifuge at 10,000 rpm for 10 min, and take the supernatant. Then the resulting supernatant was passed through a double-layer solid-phase extraction cartridge (previously activated by 5 mL of acetonitrile) through the solid-phase extraction ca...

Embodiment 3

[0033] Determination of Brassinosterols in Rice Flower Spikes

[0034] Accurately weigh 1g of rice flower spikes and place them in a mortar, freeze and grind them in liquid nitrogen until they are powdery, transfer them to a 10mL centrifuge tube, and add an isotope internal standard [ 2 h 3 ]brassinolide and [2 h 3 ] each 4ng of castasterone, and extract with 5mL of acetonitrile in a refrigerator at -20°C for 12h. The sample was centrifuged at 10,000 rpm for 10 min, and the supernatant was taken. Add 250 mg of sodium chloride to the supernatant, shake vigorously with a vortex mixer for 1 min to separate the aqueous phase and the organic phase, and discard the aqueous phase. Add 1 g of anhydrous magnesium sulfate to the centrifuge tube to further remove water. Centrifuge at 10,000 rpm for 10 min, and take the supernatant. Then the resulting supernatant was passed through a double-layer solid-phase extraction cartridge (previously activated by 5 mL of acetonitrile) through ...

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Abstract

The invention discloses a method for quantitative detection of endogenous brassinosteroids in a plant sample. According to the method, firstly the endogenous brassinosteroids are extracted by using a solvent, solid-phase extraction impurity removal is carried out by double-layer solid-phase extraction small columns, liquid-liquid extraction impurity removal is further carried out, and then, a high-performance liquid-electrospray-tandem mass spectrometry is adopted, so that the quantitative detection of the endogenous brassinosteroids is realized. The method disclosed by the invention has the advantages that the operation is simple and fast, and the sample consumption is little; and meanwhile, most of plant matrixes are effectively removed, thus the quantitative detection of low-content endogenous brassinosteroids can be realized.

Description

technical field [0001] The invention relates to a quantitative detection method for endogenous brassinosterol in plant samples, belonging to the field of analytical chemistry. Background technique [0002] Brassinosteroids (BRs) are polyhydroxylated sterol compounds, which are the sixth class of plant hormones found after auxin, gibberellin, cytokinin, abscisic acid and ethylene. So far, it has been found to include 28-norbrassinolide, 28-norcastasterone, brassinolide, 24-epibrassinolide, castasterone, 24-epibrassinolide (24-epicastasterone), homobrassinolide (homobrassinolide) and dozens of BRs compounds. The content of BRs in plants is extremely low, usually 0.01-1ng / g. Brassinosterol regulates a series of physiological and metabolic processes in plants, including seed germination, seedling growth to maturity, and seed development. At the same time, BRs also play a role in the elongation, division and differentiation of plant cells, the increase of crop yield, reproduct...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/06
Inventor 冯钰锜丁俊
Owner WUHAN UNIV
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