Method for quantitatively detecting collagen by adopting fluorescence spectrum

A technique of quantitative detection and fluorescence spectroscopy, which is applied in the field of quantitative detection of collagen proteins by fluorescence spectroscopy, can solve the problems of easy errors and achieve good linearity and sensitive reproducibility

Inactive Publication Date: 2020-09-04
ANQING NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The technical problem to be solved by the present invention is that the quantitative detection method of collagen-like protein in the prior art is prone to errors, and a method for quantitatively detecting collagen-like protein by fluorescence spectrum is provided

Method used

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  • Method for quantitatively detecting collagen by adopting fluorescence spectrum
  • Method for quantitatively detecting collagen by adopting fluorescence spectrum
  • Method for quantitatively detecting collagen by adopting fluorescence spectrum

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Determination of Collagen Content in Bovine Hide Using Fixed Wavelength Fluorescence Scanning

[0036] (1) Take fresh cowhide, wash it with water after degreasing and dehairing, then cut into pieces and homogenate; weigh 10g of the homogenate, add 300mL of 0.5mol / L acetic acid and 0.1g of pepsin (activity value 1:10000), After acting at 4°C for 48h, centrifuge at 10000rpm for 15min. The supernatant was salted out with NaCl to a concentration of 3 mol / L. The precipitated sample was dissolved again in 0.5 mol / L acetic acid solution, and then salted out with 0.7 mol / L NaCl. The precipitate was redissolved in 0.5mol / L acetic acid, and dialyzed with 0.1mol / L acetic acid solution for 3 days. Finally, the sample was frozen for 48 hours with a freeze vacuum dryer (-55°C), and stored at 4°C for later use;

[0037] (2) Dissolving the collagen prepared in step (1) in 0.1mol / L acetic acid aqueous solution to prepare a 0.1-0.9mg / mL collagen solution, and standing at 4°C for 24h; ...

Embodiment 2

[0042] Determination of Collagen Content in Bovine Hide Using Simultaneous Fluorescence Spectroscopic Scanning

[0043](1) Take fresh cowhide, wash it with water after degreasing and dehairing, then cut into pieces and homogenate; weigh 10g of the homogenate, add 300mL of 0.5mol / L acetic acid and 0.1g of pepsin (activity value 1:10000), After acting at 4°C for 48h, centrifuge at 10000rpm for 15min. The supernatant was salted out with NaCl to a concentration of 3 mol / L. The precipitated sample was dissolved again in 0.5 mol / L acetic acid solution, and then salted out with 0.7 mol / L NaCl. The precipitate was redissolved in 0.5mol / L acetic acid, and dialyzed with 0.1mol / L acetic acid solution for 3 days. Finally, the sample was frozen for 48 hours with a freeze vacuum dryer (-55°C), and stored at 4°C for later use;

[0044] (2) Dissolving the collagen prepared in step (1) in 0.1 mol / L acetic acid aqueous solution to prepare a 0.2-0.6 mg / mL collagen solution, and standing at 4°...

Embodiment 3

[0049] Determination of Collagen Content in Bovine Hide Using Exogenous Fluorescent Probe Method

[0050] (1) Take fresh cowhide, wash it with water after degreasing and dehairing, then cut into pieces and homogenate; weigh 10g of the homogenate, add 300mL of 0.5mol / L acetic acid and 0.1g of pepsin (activity value 1:10000), After acting at 4°C for 48h, centrifuge at 10000rpm for 15min. The supernatant was salted out with NaCl to a concentration of 3 mol / L. The precipitated sample was dissolved again in 0.5 mol / L acetic acid solution, and then salted out with 0.7 mol / L NaCl. The precipitate was redissolved in 0.5mol / L acetic acid, and dialyzed with 0.1mol / L acetic acid solution for 3 days. Finally, the sample was frozen for 48 hours with a freeze vacuum dryer (-55°C), and stored at 4°C for later use;

[0051] (2) Dissolving the collagen prepared in step (1) in 0.1 mol / L acetic acid aqueous solution to prepare a 0.2-0.6 mg / mL collagen solution, and standing at 4°C for 24 hour...

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Abstract

The invention discloses a method for quantitatively detecting collagen by adopting a fluorescence spectrum. The method relates to the technical field of collagen detection, and comprises the followingsteps: (1) crushing a collagen-containing raw material, homogenizing, adding pepsase at 4 DEG C, reacting for 24 hours, carrying out centrifugation, salting-out and dialysis treatment, and carrying out freeze drying to obtain collagen; (2) dissolving collagen in a 0.1 mol/L acetic acid aqueous solution to prepare collagen solutions with different concentration gradients; (3) performing fluorescence scanning on the collagen solution, and drawing a standard curve; and (4) dissolving a to-be-detected sample containing collagen in 0.1 mol/L acetic acid, carrying out fluorescence scanning, and comparing the detected absorption peak intensity with the standard curve to obtain the content of collagen in the to-be-detected sample. The method has the beneficial effects of accuracy, rapidness, sensitivity and good reproducibility, and meets the requirement of accurate quantification.

Description

technical field [0001] The invention relates to the technical field of collagen detection, in particular to a method for quantitatively detecting collagen-like proteins using fluorescence spectroscopy. Background technique [0002] Collagen, as the main component of the extracellular matrix (ECM), is a natural macromolecular protein widely distributed in organs and tissues such as skin, tendon, cartilage and blood vessels of animal organisms, which supports and protects the body and organs of animals. Its content accounts for about one-third of the total protein in the animal body. Collagen has the characteristics of good biocompatibility, biodegradability and low immunogenicity, so it has important applications in the fields of tissue engineering, biomedicine and food. [0003] Currently, quantitative detection methods for proteins have been reported, such as TNBS method, Lorry method and Bergman method. However, the specific detection method for collagen protein content ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64
CPCG01N21/6486
Inventor 李从虎吴彦龙槿彦
Owner ANQING NORMAL UNIV
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