Method used for measuring content of lysinoalanine in whey protein

A determination method, whey protein technology, applied in the field of food processing, can solve the problems of no unified determination method of lysalanine content, etc., achieve the effect of reducing interference, improving accuracy and lowering detection limit

Inactive Publication Date: 2018-10-19
LANZHOU UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The purpose of the present invention is to provide a method for determining the content of lysalanine in whey pr...

Method used

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  • Method used for measuring content of lysinoalanine in whey protein
  • Method used for measuring content of lysinoalanine in whey protein
  • Method used for measuring content of lysinoalanine in whey protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] All reagents (unless otherwise specified) were from Shanghai Aladdin Reagent Company, China, and HPLC grade solvents were provided by Fisher Technology Chemicals, UK.

[0043] LL-LAL and LD-LAL Mixed Standards Swiss Bachem Company

[0044] Chloroformic acid-9-fluorenylmethyl ester (FMOC-Cl) Switzerland Bachem company

[0045] Amino acid solid phase extraction cartridge (3cc / 500mg) American waters company

[0046] The pH of 0.4mol / L, 0.2mol / L and 0.1mol / L borate buffer was adjusted to 12, 9.0 and 7.2 with 6mol / L NaOH, respectively.

[0047] FMOC-Cl derivative agent: Dissolve appropriate amount of FMOC-Cl in acetonitrile to make the concentration reach 5mg / mL.

[0048] SPE Activation Solution: Mix acetonitrile and 0.1mol / L borate buffer (pH=7.2) at a volume ratio of 1:4.

[0049] SPE eluent 1: Mix acetonitrile and water at a volume ratio of 1:4.

[0050] SPE eluent 2: Mix acetonitrile and water at a volume ratio of 1:1.

[0051] SPE eluent: acetonitrile and 0.2mol / L ...

Embodiment 2

[0065] 1. Drawing of standard curve

[0066] Concentrations of 0.5, 1, 2, 4, 8mg / Kg standards (LL-LAL and LD-LAL mixed standard) are prepared, and the pretreatment method is exactly the same as that of the sample to be tested, including hydrolysis, derivatization, and solid phase extraction. According to the method shown in the present invention, the response values ​​of different concentrations of standard derivatives are determined by reversed-phase high performance liquid chromatography, and the standard curve is drawn with the concentration of standard derivatives as the abscissa and the response value as the ordinate. Such as figure 1 shown.

[0067] The results show that the peak area response value of HPLC is linear when the LAL standard substance concentration is 0.5-8mg / Kg (r 2 =0.999), indicating that the measuring instrument has good repeatability.

Embodiment 3

[0069] 2. Measurement of precision, accuracy and reproducibility

[0070] Precision refers to the closeness between the results of the same sample to be tested after multiple sampling measurements, and is generally expressed by relative standard deviation (RSD). Add an equal volume of 4mg / Kg LAL standard substance to deionized water (blank control) without any LAL, after a series of pretreatment methods such as acid hydrolysis, derivatization, solid phase extraction, etc., repeat 9 times to measure its LAL The precision was expressed by calculating the RSD values ​​of retention time and peak area.

[0071] Accuracy refers to the closeness between the measured value of the sample measured by the above method and the true value, which can usually be expressed by the recovery rate. The accuracy is expressed by adding equal volumes of 0, 0.5, 2, and 8 mg / Kg LAL standard substances to the sample to be tested, and calculating the recovery rate and the RSD value of the peak area. ...

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Abstract

The invention relates to a method used for measuring content of lysinoalanine in whey protein. The method comprises following steps: 1, whey protein hydrolysis, 2, derivatization processing; 3, solidphase extraction (SPE), and 4, reversed-phase high-performance liquid chromatography (HPLC). The method is used for solving a problem in the prior art that no standard method used for measuring content of lysinoalanine in whey protein is disclosed. The operation flow is standard; detection precision is high; the detection limit reaches 0.5mg/Kg; and the precision, the accuracy, and the reproducibility are ideal.

Description

technical field [0001] The invention relates to the technical field of food processing, in particular to a method for determining the content of lysalanine in whey protein. Background technique [0002] Whey protein is a protein extracted from milk. It is known as the king of protein. It has the characteristics of high nutritional value, easy digestion and absorption, and contains a variety of active ingredients. It is recognized as one of the high-quality protein supplements for the human body. First, it is widely used in the food industry. [0003] In the application of whey protein in the food industry, it often undergoes two common processing methods, heat treatment and alkali treatment. However, heat treatment of whey protein, especially under alkaline conditions, can promote the production of cross-linked amino acids, including lysinoalanine (LAL). Lysalanine, that is, N(6)-(DL-2-amino-2-carboxyethyl)-DL-lysine, has a chemical structural formula as shown in Formula 1...

Claims

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Application Information

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IPC IPC(8): G01N30/02
CPCG01N30/02
Inventor 刘迪茹黄小丹罗斌张莹龙丹凤李星
Owner LANZHOU UNIVERSITY
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