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Method for detecting content of L-hydroxyproline in dairy product

A technology of hydroxyproline and detection method, which is applied in the directions of measuring devices, instruments, scientific instruments, etc., can solve the problems of restricting wide application and restricting the wide configuration of instruments, and achieves simple and fast operation, simple and fast operation, and rich structural information. Effect

Inactive Publication Date: 2012-07-18
上海德诺产品检测有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The amino acid automatic analyzer method is faster, more accurate, more automated and more sensitive, but this method requires a special instrument, an amino acid analyzer, which can only be used to analyze amino acids, which limits the wide configuration of the instrument, thereby limiting the post-column derivatization method Wide Application of Amino Acid Analyzer Method in Analyzing L-Hydroxyproline in Dairy Products

Method used

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  • Method for detecting content of L-hydroxyproline in dairy product
  • Method for detecting content of L-hydroxyproline in dairy product
  • Method for detecting content of L-hydroxyproline in dairy product

Examples

Experimental program
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Effect test

Embodiment 1

[0047] Collagen powder was added to the pure milk sample (L-hydroxyproline was not detected) to prepare the simulated positive sample of liquid dairy product in this embodiment.

[0048] 1. Sample pretreatment.

[0049] Weigh 2.0 g of sample, add 6 mL of 10.6 mol / L hydrochloric acid solution, seal the tube in vacuum, and hydrolyze at 110 °C for 12 h. Evaporate the solvent, add water to dissolve, adjust the NaOH solution to neutral, add water to make it 25mL. Transfer to a 50mL centrifuge tube, centrifuge at 8000 rpm for 5min, pass through a 0.22μm microporous membrane, and wait for detection.

[0050] 2. Hydrophilic chromatography (HILIC) analytical column for high performance liquid chromatography-tandem mass spectrometry

[0051] In the present invention, the basic principle of carrying out high-performance liquid chromatography-tandem mass spectrometry through the hydrophilic chromatography (HILIC) analytical column is that the sample is hydrolyzed into various amino acid...

Embodiment 2

[0077] (1) Sample pretreatment:

[0078] Weigh 1.5g of commercially available yogurt, add 1.0mL of distilled water and mix well, add 6-9mL of 10.6mol / L hydrochloric acid solution, seal the tube in vacuum, hydrolyze at 110°C for 16h, evaporate the solvent, add water to dissolve, and adjust to Neutral, add water to 25mL, transfer to a 50mL centrifuge tube, centrifuge at 8000 rpm for 10min, dilute the supernatant 5 times with 10mmol / L ammonium acetate aqueous solution with a volume percentage of 20%, pass through a 0.22μm microporous membrane, Prepare a test solution.

[0079] (2) High-performance liquid chromatography-tandem mass spectrometry was performed on a hydrophilic chromatography (HILIC) analytical column, and the method was the same as in Example 1.

[0080] The same positive sample was analyzed 5 times by the above analysis method, and accurate spectra and quantitative information could be obtained for each analysis. Compared with the standard curve, the results are s...

Embodiment 3

[0085] (1) Sample pretreatment

[0086] Weigh 0.5g of commercially available milk powder, add 2.0mL of distilled water to reconstitute, add 6-9mL of 10.6mol / L hydrochloric acid solution, seal the tube in vacuum, hydrolyze at 110°C for 14h, evaporate the solvent, add water to dissolve, adjust to neutral with NaOH solution , add water to make up to 25mL, transfer to a 50mL centrifuge tube, centrifuge at 8000 rpm for 8min, and pass through a 0.22μm microporous membrane to prepare the test solution.

[0087] (2) High-performance liquid chromatography-tandem mass spectrometry was performed on a hydrophilic chromatography (HILIC) analytical column, and the method was the same as in Example 1.

[0088] The same positive sample was analyzed 5 times by the above analysis method, and accurate spectra and quantitative information could be obtained for each analysis. Compared with the standard curve, the results are shown in Table 4 below:

[0089] Table 4: Determination table of L-hydro...

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Abstract

The invention discloses a method for detecting the content of L-hydroxyproline in a dairy product. The method comprises the following steps of: 1) pre-treating a sample, namely adding acid into the diary product and hydrolyzing to obtain test liquid; 2) analyzing the content of L-hydroxyproline by liquid chromatogram-tandem mass spectrometry, wherein hydrophilic interaction chromatography (HILIC) is adopted, a flowing phase is a mixed solution a flowing phase A and a flowing phase B, and the flowing phase A is a 10mM ammonium acetate aqueous solution of which the pH is 5.6; and the flowing phase B is acetonitrile, and the mass spectrometry condition is that: an electrospray ion source is adopted, a scanning mode is a positive ion scanning mode, a detection mode is a multi-reaction detection mode, electrospray voltage is 5,500V, the temperature of the ion source is 600 DEG C, the pressure force of a gas curtain is 20Psi, the atomization pressure force is 50Psi, and auxiliary pressure force is 30Psi. By the method, the detection accuracy and sensitivity can be improved obviously, and the content of the L-hydroxyproline in the dairy product can be determined easily, conveniently and quickly.

Description

technical field [0001] The invention relates to a method for detecting L-hydroxyproline content in dairy products. Background technique [0002] L-Hydroxyproline, English name hydroxyproline, HYP for short, also known as trans-4-hydroxy-L-proline, molecular formula is C 5 h 9 NO 3 , molecular weight 131.13. [0003] L-Hydroxyproline is an imino acid, usually with a hydroxyl group in the 4th position of proline, but sometimes also in the 3rd position. L-Hydroxyproline is an amino acid unique to collagen, which accounts for about 10% of the total amino acids in collagen, and is very small in other proteins, which makes it almost a unique component of collagen, so it can be passed through The content of L-hydroxyproline is measured to calculate the content of collagen, so as to determine the overall level of collagen. [0004] Animal collagen hydrolyzed protein is a cheap protein raw material. It is a hydrolyzed protein made by rough processing of cowhide and its product w...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/88
Inventor 孟瑾郑小平何亚斌韩奕奕韩慧雯章寅
Owner 上海德诺产品检测有限公司
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