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Visible tracking and detecting method for protease in leather treatment process

A technology for leather processing and tracking detection, applied in measuring devices, instruments, scientific instruments, etc., can solve problems such as difficulty in detecting permeability

Active Publication Date: 2013-12-11
SHAANXI UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At the earliest, someone transferred the protease of different levels in the enzymatic dehaired pigskin to casein, and the enzyme reacted with the casein to produce white spots, and the penetration depth and quantity of the enzyme could be known through the spots (Yang Y.Z.Test of enzyme in dehaired pelt.China Leather, 2002,31:20–22), however, it is difficult to detect the permeability of proteases that have been inactivated by enzymatic treatment

Method used

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  • Visible tracking and detecting method for protease in leather treatment process

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] 1) Prepare 2 mg / mL Genecor TanG protease solution and 1 mg / mL FITC solution with phosphate buffer solution with a pH value of 8.0 as solvent;

[0025] 2) Slowly add 0.2mL of FITC solution to 10mL of Genecor TanG protease solution and shake well, so that the mass ratio of Genecor TanG protease to FITC in the resulting reaction system is 100:1, and then react at 4°C in the dark for 10h , to obtain the reaction solution;

[0026] 3) Put the reaction solution into a dialysis bag with a molecular cut-off range smaller than the molecular weight of Genecor TanG protease, and then put it into a phosphate buffer with a pH value of 8.0 for oscillating dialysis. During the oscillating dialysis process, change the pH value once every 2 hours. 8.0 phosphate buffer solution until the eluate obtained by dialysis has an absorbance of 0 at 480nm, indicating that there is no free FITC in the eluate obtained by dialysis, and the dialysis is stopped. At this time, the remainder in the dial...

Embodiment 2

[0029] 1) Prepare 2 mg / mL Genecor TanG protease solution and 1 mg / mL FITC solution with phosphate buffer solution with a pH value of 8.0 as solvent;

[0030] 2) Slowly add 0.2mL of FITC solution to 10mL of Genecor TanG protease solution and shake well, so that the mass ratio of Genecor TanG protease to FITC in the resulting reaction system is 100:1, and then react at 4°C in the dark for 10h , to obtain the reaction solution;

[0031] 3) Put the reaction solution into a dialysis bag with a molecular cut-off range smaller than the molecular weight of Genecor TanG protease, and then put it into a phosphate buffer with a pH value of 8.0 for oscillating dialysis. During the oscillating dialysis process, change the pH value once every 2 hours. 8.0 phosphate buffer solution until the eluate obtained by dialysis has an absorbance of 0 at 480nm, indicating that there is no free FITC in the eluate obtained by dialysis, and the dialysis is stopped. At this time, the remainder in the dial...

Embodiment 3

[0035] 1) Prepare 2 mg / mL Genecor LimeG protease solution and 1 mg / mL FITC solution with glycine-sodium hydroxide buffer solution with a pH value of 9.0 as solvent;

[0036] 2) Slowly add 0.5mL of FITC solution to 10mL of Genecor LimeG protease solution and shake well, so that the mass ratio of Genecor LimeG protease to FITC in the resulting reaction system is 100:2.5, and then react at 4°C in the dark for 10h , to obtain the reaction solution;

[0037] 3) Put the reaction solution into a dialysis bag with a molecular cut-off range smaller than the molecular weight of Genecor LimeG protease, and then put it into a glycine-sodium hydroxide buffer solution with a pH value of 9.0 for oscillating dialysis. During the oscillating dialysis process, change 1 Glycine-sodium hydroxide buffer solution with a secondary pH value of 9.0, until the eluate obtained by dialysis has an absorbance of 0 at 480nm, indicating that there is no free FITC in the eluate obtained by dialysis, stop dial...

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Abstract

The invention discloses a visible tracking and detecting method for protease in a leather treatment process. The visible tracking and detecting method comprises the following steps of firstly, under an alkaline condition, marking protease through fluorescein isothiocyanate (FITC) to obtain FITC marker enzyme, then treating a skin block without furs with the marker enzyme under room temperature and shading conditions, and preparing a slice from the skin block treated by the marker enzyme to observe a distribution state of the enzyme in the skin under a fluorescence microscope. According to the visible tracking and detecting method, the permeability of the enzyme in the skin and the acting degree of the enzyme to the skin can be detected according to the strong fluorescence characteristic of the FITC, and a permeation mode of the enzyme in the skin can be obtained according to the difference of treatment time. The enzyme firstly permeates into the skin through pores and the peripheries of hair follicles and then gradually diffuses into a skin substrate to act on fibrous stroma proteins. According to the visible tracking and detecting method, a theoretical basis is supplied to the application of the enzyme to manufacturing of leather; enzyme with proper molecular weight is selected for treating collagen according to fiber structures of the collagen in different treatment stages of manufacturing of leather, and the controllable treatment of enzyme on the leather protein and the collagen is realized by analyzing the permeability and the acting degree of the enzyme in the collagens.

Description

technical field [0001] The invention relates to a method for detecting enzymes in leather collagen fibers, in particular to a method for visually tracking and detecting proteases in leather processing. Background technique [0002] With the seriousness of pollution in the tanning industry, the research on environmentally friendly materials for tanning has aroused widespread concern from tanning researchers. Among them, enzyme preparations, as an efficient and environmentally friendly material, have been used in tanning for a long time. Hydrolytic enzymes are mainly used to remove non-collagen components such as protein and fat in the hide, so as to prepare for the penetration and combination of materials in the later process. (Swarna V.Kanth; R.Venba; B.Madhan; N.K.Chandrababu; S.Sadulla. Cleaner tanning practices for tannery pollution abatement: Role of enzymes in eco-friendly vegetable tanning.J.Clean Prod.,2009,17:507 –515; Puvanakrishnan R. Ecofriendly lime and Sulfide ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/573
Inventor 马建中侯雪艳高党鸽吕斌
Owner SHAANXI UNIV OF SCI & TECH
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