A kind of method for accurately detecting phytosterol ester content by using microplate reader

The technology of phytosterol ester and microplate reader is applied in the field of detection of phytosterol ester content, which can solve the problem of not being suitable for the detection of phytosterol ester, and achieve the effects of low detection limit, high detection throughput and good effect.

Active Publication Date: 2021-08-03
ZHEJIANG UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, for the detection of phytosterol esters, the sample pretreatment method used when the conventional UV-visible spectrophotometer detects phytosterols is not suitable for the detection of phytosterol esters on a multifunctional microplate reader.

Method used

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  • A kind of method for accurately detecting phytosterol ester content by using microplate reader
  • A kind of method for accurately detecting phytosterol ester content by using microplate reader
  • A kind of method for accurately detecting phytosterol ester content by using microplate reader

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Experimental program
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Effect test

Embodiment 1

[0045] The present embodiment provides a kind of method that utilizes microplate reader to accurately detect phytosterol ester content, and this method is to obtain after a large number of experiment groping (test groping process is as shown in embodiment 2), concrete steps are as follows:

[0046] (1) Take 1 mL of the sample to be tested containing phytosterol esters, add it to about 40 mL of solvent ethyl acetate, and treat it with ultrasonic (100W) for 5 minutes, then dilute to 50 mL with ethyl acetate, and the phytosterol esters are completely dissolved in ethyl acetate In, obtain phytosterol ester test solution (the phytosterol ester equivalent to 3.75mg / mL);

[0047] (2) Mix and shake 200 μL of the phytosterol ester test solution and 100 μL of the phosphorus-sulfur iron color developer with a mass concentration of 0.0015%, and develop the color for 25 minutes at room temperature to obtain the color-developing solution;

[0048] (3) Take 200 μL of the chromogenic solution...

Embodiment 2

[0057] This embodiment explores, optimizes and verifies the conditions of each step in the detection method. In order to facilitate the exploration, optimization and verification of the detection method, this embodiment uses phytosterol esters directly for testing. The phytosterol esters are conventional commercially available Sales product, and in the process of groping for the following conditions, except that the mentioned steps are changed, all the other contents are the same as in Example 1.

[0058] 1. Solvent selection and ultrasonic time

[0059] Take 100 mg of phytosterol ester and put it in a beaker, add 80 mL of organic solvent, ultrasonically treat until it is completely dissolved, transfer it to a 100 mL volumetric flask, and use the same organic solvent to make up the volume to obtain the test solution of phytosterol ester.

[0060] When in use, draw 10 mL of the above-mentioned solution to be tested, and dilute to 100 mL with an organic solvent. It is proposed ...

Embodiment 3

[0086] Embodiment 3 precision and rate of recovery

[0087] 1) Draw the standard curve of ΔA and different concentrations of phytosterol esters: add 200 μL of ethyl acetate solutions of different concentrations of phytosterol esters to the microwell microplate plate, 100 μL of phosphorous sulfur iron chromogen, and put in the multifunctional enzyme standard Oscillate in the instrument, develop color at room temperature for 25 minutes, and measure the absorbance value at the optimal wavelength of 492nm, where the reagent blank is marked as A 0 , the ethyl acetate solution containing phytosterol esters is denoted as A, and calculates ΔA=A-A 0 , to establish a standard curve of ΔA and phytosterol ester concentration.

[0088] 2) Preparation of phosphorus sulfur ferric color reagent: weigh 1gFeCl 3 ﹒ 6H 2 Dissolve O (AR grade) in 85% concentrated phosphoric acid, dilute to 100mL with phosphoric acid, store in a brown bottle, keep refrigerated, and can be stored for one year to o...

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Abstract

The invention discloses a method for accurately detecting the content of phytosterol esters by using a microplate reader. The method comprises the following steps: taking a sample to be tested, adding it to ethyl acetate, and ultrasonically treating it until it is completely dissolved to obtain a phytosterol ester test solution; After mixing and shaking the liquid to be tested and the phosphorus-sulfur-iron color developer with a mass concentration of 0.0015-0.015%, develop color at room temperature to obtain a color-developing solution; take 50-300 μL of the color-developing solution and put it in a microplate reader for detection. Obtain the absorbance value of the sample to be tested; draw a standard curve to obtain the content of phytosterol ester in the sample to be tested. The method of the present invention adopts ethyl acetate as the solvent of the phytosterol ester, adopts the phosphorus-sulfur ferric solution with a mass concentration of 0.0015% to 0.015% as the color developing agent, cooperates with the microplate reader detection, and not only realizes detection under trace conditions , to achieve the purpose of high-throughput detection, and high detection accuracy, high precision, low detection limit, and stable color development.

Description

technical field [0001] The invention relates to the technical field of detection of phytosterol ester content, in particular to a method for accurately detecting the content of phytosterol ester by using a microplate reader. Background technique [0002] Phytosterols have the effect of lowering plasma total cholesterol and low-density lipoprotein cholesterol. Since the water solubility and oil solubility of phytosterols are not ideal, and the solubility in edible oil and fat can be significantly improved after esterification, sterol esters can be converted into sterols and fatty acids in the human body, and the phytostanol esters in the intestinal tract The hydrolysis rate reaches 90%, so it has two physiological functions of phytosterol and fatty acid. Phytostanol esters mainly include β-sitosterol esters, phytosterol esters, and campesterol esters. At present, phytosterol esters are mainly used in food spreads and salad dressings. Phytosterol ester products have been app...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/78G01N21/31
CPCG01N21/31G01N21/78
Inventor 冯思敏孙培龙王章铁王丹邵平
Owner ZHEJIANG UNIV OF TECH
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