Sample pre-treatment method for determining endogenesis hormone content of towel gourd seeds by adopting HPLC (High Performance Liquid Chromatography)

A sample pretreatment and endogenous hormone technology is applied in the field of sample pretreatment for measuring the endogenous hormone content of loofah seeds by HPLC. The effect of saving time and reducing loss

Inactive Publication Date: 2012-12-19
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This pretreatment method is not only expensive, but also complicated and time-consuming.
At present, there is no public report on the determination of hormone content in loofah seeds, and the effect of using the above pretreatment method to treat loofah seeds is not ideal

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Processing steps include:

[0023] Accurately weigh 1.000 g of the loofah seeds with the outer testa removed, grind them into powder under liquid nitrogen, add 6 ml of cold acetonitrile, seal them and put them in a refrigerator at 4 °C for extraction for 24 h, then centrifuge at low temperature for 10 min (10000 r / m ) to take the supernatant, and then add 4 ml of cold acetonitrile to the residue for low-temperature ultrasonic oscillation for 1 h, centrifuge for 10 min (10,000 r / m) to get the supernatant, repeat the extraction twice, and combine the supernatant three times. The use of acetonitrile extraction can not only reduce the dissolution of impurities, but also acetonitrile is easy to volatilize, which is beneficial to speed up the efficiency of nitrogen blowing concentration and shorten the experimental time.

[0024] Blow dry with nitrogen at 35°C, and add 3 ml of phosphate buffer solution (0.1 mol / L) with pH=8.0. The nitrogen blowing instrument can process mult...

Embodiment 2

[0029] This method was used to determine the content of endogenous hormone ABA in developing loofah seeds.

[0030] Accurately weigh 1.000 g of the loofah seeds with the outer testa removed, grind them into powder under liquid nitrogen, add 6 ml of cold acetonitrile, seal them and put them in a refrigerator at 4 °C for extraction for 24 h, then centrifuge at low temperature for 10 min (10000 r / m ) to take the supernatant, then add 4 ml of cold acetonitrile to the residue, ultrasonically shake at low temperature for 1 h, centrifuge for 10 min (10,000 r / m) to get the supernatant, repeat the extraction twice, and combine the supernatant three times.

[0031] Blow dry with nitrogen at 35 °C, add 3 ml of phosphate buffer solution (0.1 mol / L) with pH=8.0, freeze in -80 °C ultra-low temperature refrigerator for 30 min, thaw at 4 °C, and centrifuge at low temperature for 15 min (10000 r / m), filter to discard impurities, adjust to pH = 3.0~3.5 with hydrochloric acid (0.5 mol / L), then e...

Embodiment 3

[0035] Determination of endogenous hormone GA in developing loofah seeds by this method 3 content.

[0036] Accurately weigh 1.000 g of the loofah seeds with the outer testa removed, grind them into powder under liquid nitrogen, add 6 ml of cold acetonitrile, seal them and put them in a refrigerator at 4 °C for extraction for 24 h, then centrifuge at low temperature for 10 min (10000 r / m ) to take the supernatant, then add 4 ml of cold acetonitrile to the residue, ultrasonically shake at low temperature for 1 h, centrifuge for 10 min (10,000 r / m) to get the supernatant, repeat the extraction twice, and combine the supernatant three times.

[0037] Blow dry with nitrogen at 35 °C, add 3 ml of phosphate buffer solution (0.1 mol / L) with pH=8.0, freeze in -80 °C ultra-low temperature refrigerator for 30 min, thaw at 4 °C, and centrifuge at low temperature for 15 min (10000 r / m), filter to discard impurities, adjust to pH = 3.0~3.5 with hydrochloric acid (0.5 mol / L), then extract ...

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PUM

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Abstract

The invention provides a sample pre-treatment method for determining an endogenesis hormone content of towel gourd seeds by adopting an HPLC (High Performance Liquid Chromatography), comprising the following treating steps of: preparing a towel gourd seed plant hormone rough extraction solution to be detected; concentrating the rough extraction solution and adding 3 ml of phosphoric acid buffering solution with the pH (Potential of Hydrogen) of 8.0 and the concentration of 0.1 mol/L; freezing and degreasing; filtering and removing impurities; adjusting the pH value to 3.0-3.5 by using diluted hydrochloric acid with the concentration of 0.5 mol/L and utilizing ethyl acetate with the same volume to extract for three times; mixing ester phrases; concentrating the ester phases and utilizing a flow phase to adjust the volume to 1 ml; and filtering with a micro-pore filtering film with the size of 0.45 microns and utilizing the HPLC to analyze. The method disclosed by the invention has the advantages of simplicity in operation and low cost; the method can treat a plurality of samples one time so that the time can be saved, column treatment is not carried out in a purification process, and loss of objects to be detected in an extraction process is reduced. The method disclosed by the invention can be used for pre-treatment before extracting a plurality of types of plant hormones and can meet the requirements of treating samples by adopting the high performance liquid chromatography.

Description

[0001] technical field [0002] The invention belongs to the field of analytical chemistry, and more specifically relates to a sample pretreatment method for HPLC determination of endogenous hormone content in luffa seeds. [0003] Background technique [0004] Loofah is a vegetable crop that can be cultivated in both north and south of my country. In addition to its rich nutritional value, loofah also has medical and health care functions, such as clearing away heat and resolving phlegm. In-depth study of loofah seeds can provide a theoretical basis for the breeding of loofah seeds. [0005] Phytohormones are important regulators of plant life activities. The content of phytohormone itself in plants is very small, and its composition is complex. In addition, there are many interfering substances in seed materials. Therefore, it is difficult to measure phytohormones in seeds. [0006] High performance liquid chromatography (high performance liquid chromatography, also kn...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06
Inventor 胡晋秦国臣王洋朱岩芳祝水金
Owner ZHEJIANG UNIV
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