Method for determining short chain fatty acid content in intestinal contents or excrement

A technology for short-chain fatty acids and content, which is used in measurement devices, instruments, scientific instruments, etc., can solve the problems of damage to the detector, low extraction rate of short-chain fatty acids, and low efficiency of ultra-pure water extraction of short-chain fatty acids, etc. effect of loss

Inactive Publication Date: 2019-07-23
JIANGNAN UNIV
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

First of all, the extraction efficiency of short-chain fatty acids in ultrapure water is not high, which will cause the detection value of the sample to be lower than the actual value; secondly, the effect of metaphosphoric acid in this method is to reduce the loss of short-chain fatty acids, but the treatment time of metaphosphoric acid is long (more than 2h) , which greatly reduces the experimental efficiency; finally, the sample contains a large amount of water, and direct injection will not only lead to a serious reduction in the life of the chromatographic column, but also seriously damage the detector.
Zhang Mengjie and others also used gas chromatography to quantify the content of short-chain fatty acids in feces. They also used dilute acid solution (2-ethylbutyric acid + hydrochloric acid) to extract short-chain fatty acids, and directly detected them in the gas phase through the membrane. This method also has shortcomings. The extraction rate of chain fatty acids is low (the recovery rate of standard addition is 83.75%~94.95%), the chromatographic analysis takes a long time (about 40min), and it is easy to damage the chromatographic column and instruments, etc.

Method used

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  • Method for determining short chain fatty acid content in intestinal contents or excrement
  • Method for determining short chain fatty acid content in intestinal contents or excrement
  • Method for determining short chain fatty acid content in intestinal contents or excrement

Examples

Experimental program
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Effect test

Embodiment 1

[0029] Embodiment 1: Determination of short-chain fatty acid mixed standard standard curve

[0030]The sample is a mixed standard solution of short-chain fatty acids, and the concentration range of each short-chain fatty acid standard product in the standard curve is set as: 0.015~0.200 μL / mL (short-chain fatty acid standard product: solvent; V / V); The mass volume concentration range of each short-chain fatty acid in the corresponding standard product is: acetic acid 15.738-209.840 μg / mL, propionic acid 14.850-198.000 μg / mL, isobutyric acid 14.235-189.800 μg / mL, butyric acid 14.460-192.800 μg / mL mL, isovaleric acid 13.962-186.160 μg / mL, valeric acid 14.085-187.800 μg / mL. Set 7 concentrations of each standard analyte (for example, 0.015, 0.025, 0.050, 0.075, 0.100, 0.150, 0.200 μL / mL); each concentration of standard analyte was determined in parallel three times. Take 3 times of the standard deviation of the peak area as the lower limit of detection (LOD) of each short-chain f...

Embodiment 2

[0041] Embodiment 2: Determination of the content of short-chain fatty acids in the colon contents of SD rats

[0042] (1) Sample dissolution: Mix about 50 mg of sample with 500 μL of saturated sodium chloride solution, add 0.1 times the volume of saturated sodium chloride in 10% sulfuric acid solution to acidify, and oscillate to fully dissolve the sample;

[0043] (2) Extraction of short-chain fatty acids: add 800 μL of ether, shake for 30 s, and fully extract the short-chain fatty acids in the sample;

[0044] (3) Sample preparation: centrifuge the extract at 12 000r / min, 4°C for 10min, take the supernatant, add 0.25g of anhydrous sodium sulfate solid, shake for 30s, centrifuge at 4500r / min, 4°C for 3min, remove traces Measure the water, pass through a 0.22μm organic microporous membrane, and enter the chromatograph for analysis;

[0045] (4) Determination by gas chromatography: Chromatographic column: Agilent DB-FFAP strong polar capillary column. The experimental data w...

Embodiment 3

[0052] Embodiment 3: Measuring the content of short-chain fatty acids in the fresh feces of SD rats

[0053] (1) Sample dissolution: Mix about 50 mg of sample with 500 μL of saturated sodium chloride solution, add 0.05 times the volume of saturated sodium chloride in 10% sulfuric acid solution to acidify, and oscillate to fully dissolve the sample;

[0054] (2) Extraction of short-chain fatty acids: add 800 μL of ether, shake for 30 s, and fully extract the short-chain fatty acids in the sample;

[0055] (3) Sample preparation: centrifuge the extract at 12 000r / min, 4°C for 10min, take the supernatant, add 0.25g of anhydrous sodium sulfate solid, shake for 30s, centrifuge at 4500r / min, 4°C for 3min, remove traces Measure the water, pass through a 0.22μm organic microporous membrane, and enter the chromatograph for analysis;

[0056] (4) Determination by gas chromatography: Chromatographic column: Agilent DB-FFAP strong polar capillary column (column parameters: 30m×0.25μm×0.2...

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Abstract

The invention discloses a method for determining short chain fatty acid content in intestinal contents or excrement. A sample is pretreated by diethyl ether, and the short chain fatty acid content inthe sample is determined through a gas chromatographic method; and pretreatment temperature is 2-8 DEG C. In the method provided by the invention, other derivatization pretreatment steps are not needed, pretreatment time is less than 15min, length of determining time is less than 20min; low temperature can effectively reduce volatilization loss of the short chain fatty acid, and recovery rate is up to 98.10 +/-0.29%-101.30 +/-0.27%; with the method provided by the invention, limit of detection can be as low as 0.0036 <mu>L/mL, limit of quantitation can be as low as 0.0041 <mu>L/mL, so the method is more accurate and efficient for detection of the short chain fatty acid.

Description

technical field [0001] The invention relates to a method for measuring the content of short-chain fatty acids in intestinal contents or feces, belonging to the field of biological sample detection. Background technique [0002] The intestine is not only an important place for human digestion and absorption, but also the largest immune organ, which plays an extremely important role in maintaining normal immune defense functions. The human intestinal tract provides a good habitat for microorganisms. The microbial population living in the human gastrointestinal tract can reach tens of trillions, which is 10 times the number of human cells. More than 1000 species of bacteria have been identified and about 160 species of microorganisms are present in all individuals. The genome of intestinal microorganisms includes about 3 million genes, which is 150 times that of the human genome, and has metabolic functions that the human body does not possess. Intestinal microecology refers ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06G01N2030/025
Inventor 刘元法叶展曹晨李进伟
Owner JIANGNAN UNIV
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