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Method for extracting, purifying and detecting acyl-homoserine lactones in soil and application thereof

A technology of homoserine lactone and determination method, which is applied in the field of extraction, purification and determination of acyl homoserine lactone in soil, can solve the problems of weak specificity, low sensitivity and difficult quantification, and achieves strong specificity, high sensitivity, short cycle effect

Inactive Publication Date: 2017-01-04
INST OF SOIL SCI CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In view of the fact that the existing HSLs detection methods are more restricted by environmental conditions, and the detection in complex media (such as soil) is rarely reported, the present invention provides a method for the extraction and purification of acyl homoserine lactones in soil and its The application overcomes the shortcomings of the existing HSLs biosensor detection methods in soil, such as long detection time, low sensitivity, low specificity, and difficulty in quantification.

Method used

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  • Method for extracting, purifying and detecting acyl-homoserine lactones in soil and application thereof
  • Method for extracting, purifying and detecting acyl-homoserine lactones in soil and application thereof
  • Method for extracting, purifying and detecting acyl-homoserine lactones in soil and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] This example focuses on the comparison of different eluents for the purification of acyl homoserine lactones (HSLs) in soil extracts.

[0038] Step 1: Take 20g of Jiangsu yellow brown soil in a beaker, add 3mL of 100mg L -1 HgCl 2 The solution is sterilized, stirred evenly and set aside.

[0039] Step 2: Weigh 2g of the prepared yellow-brown soil, add 1g of diatomaceous earth, mix well and pour into a 15mL ASE extraction pool, mix well and transfer to a 15mL ASE extraction pool, and fill the remaining space of the extraction pool with quartz sand. Ethyl acetate / acetone (2:1, v / v) was selected as the extraction agent, and the conditions for accelerated solvent extraction were: extraction temperature 100°C, pressure 1500psi, preheating for 5 minutes, static extraction for 5 minutes, cycle once, nitrogen purge for 40s to collect all extract. The extract was transferred to a 50mL pear-shaped bottle, and concentrated by rotation at 50°C to nearly 1mL.

[0040] Step 3: Ta...

Embodiment 2

[0044] This embodiment focuses on comparing the extraction efficiency of acyl homoserine lactones (HSLs) in soil by accelerated solvent extraction compared with conventional extraction methods (vibration extraction and ultrasonic extraction). The steps are:

[0045] Step 1: Take 50g of Hainan brick red soil and Jiangsu yellow brown soil in different beakers, add 5mL 100mg L -1 HgCl 2 The solution is sterilized and stirred evenly;

[0046] Step 2: Weigh 2g of sterilized brick red soil and yellow brown soil into different 50mL beakers, add 2μL and 5μL of 100mg L -1 HSLs standard stock solution to a spiked concentration of 0.1mg kg -1 and 0.5mg kg -1 ;

[0047] Step 3: Carry out accelerated solvent extraction according to step 2 described in Example 1. The extract was concentrated to 1 mL by a rotary evaporator, ethyl acetate was used for solid-phase extraction column purification, the eluate was concentrated by a rotary evaporator, and the ethyl acetate was fixed to 1 mL be...

Embodiment 3

[0053] This embodiment focuses on comparing the extraction recovery rates of acyl homoserine lactones (HSLs) in soil extracted by accelerated solvent extraction with different solvents.

[0054] Step 1: Take 2 g of the sterilized brick red soil and yellow brown soil described in Step 1 of Example 2 in different 50 mL beakers, add 2 μL of 100 mg L -1 HSLs standard stock solution to a spiked concentration of 0.1mg kg -1 .

[0055] Step 2: Carry out accelerated solvent extraction according to Step 2 described in Example 1. The extraction solvents were ethyl acetate / acetone (4:1, v / v) and ethyl acetate solution, and the extract was concentrated to 1mL by a rotary evaporator at 50°C, and ethyl acetate was used for SPE purification, and the eluent was rotated to Concentrate with an evaporator at 50°C, dilute to 1 mL with ethyl acetate, and measure by GC / MS;

[0056] Step 3: Take 2 g each of the sterilized and unspiked brick red soil and yellow brown soil, and repeat the operation...

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Abstract

The invention discloses a method for extracting, purifying and detecting acyl-homoserine lactones (HSL) in soil and application thereof, and belongs to the technical field of pretreatment analysis of compounds in soil. An analysis method capable of efficiently and quickly extracting and purifying HSL in the soil is established by utilizing ASE and SPE technologies, an ASE extracting agent is ethyl acetate / acetone (4:1, v / v), the extraction is carried out every 5 minutes of preheating and 5 minutes of static extraction at a temperature of 100 DEG C and a pressure of 1500psi, and all extracting liquid is collected by sweeping nitrogen for 40 seconds; SPE column packing is prepared from anhydrous sodium sulfate and silica gel, and 8mL of ethyl acetate is used as an eluting agent. The method has the advantages of simple operation and high detection accuracy, can fill up the blank of the field of simultaneously qualitatively and quantitatively analyzing multiple HSLs signal molecular compounds in soil and other complicated mediums, and establishes foundation for intensively understanding the quorum sensing behavior and mechanism of microorganism in a complicated soil environment.

Description

technical field [0001] The invention belongs to the technical field of pretreatment and analysis of compounds in soil, and more specifically relates to an extraction, purification and determination method of acyl homoserine lactone in soil and its application. Background technique [0002] In recent years, bacterial quorum sensing (QS) is closely related to many physiological activities of microorganisms, and has become a hot spot in microbiology research. Quorum sensing is information exchange between microorganisms. Microorganisms sense the changes in the number of themselves and other flora in the surrounding environment and control the expression of genes by releasing specific signaling molecules. N-acyl homoserine lactones (Acyl-homoserine lactones, HSLs), also known as autoinducers, are regulated and synthesized by the LuxI / R type information system. Gram-negative bacteria communicate and coordinate group behavior by secreting HSLs. When the concentration of signal mo...

Claims

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Application Information

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IPC IPC(8): G01N30/08G01N30/88
CPCG01N30/08G01N30/88
Inventor 王芳生弘杰宋洋卞永荣谷成刚杨兴伦叶茂蒋新
Owner INST OF SOIL SCI CHINESE ACAD OF SCI
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