Regenerated keratin has many excellent properties, can be applied to different areas, so keratin recycling has attracted more and more people's attention. Wool is a natural macromolecular material composed of keratin, which is one of the important sources of renewable proteins. Therefore, it is one of the current research focuses to dissolve keratin from wool. The key to dissolve the wool is to open the disulfide bond, destroying the horizontal relationship between wool macromolecules, but also to minimize the degradation of keratin macromolecules obtained, so the choice of reducing agent is very important. DTT and TCEP hydrochloride are disulfide reductant, the use of them in the wool keratin protein extraction will be compared.

Extraction of wool keratin with DTT: A certain amount of wool was added to the solution of metal salt ZnCl2 and surfactant SDS, adjust the pH value, microwave-assisted hydrolysis after putting at room temperature for 30min, take out from the microwave reactor into the three-necked flask, then gradually adding DTT solution (during which pure nitrogen is needed to cut off air), the temperature is adjusted and stirred evenly, the wool keratin solution is obtained after reacting for 2h, the undissolved wool is filtered and washed with water and dried; The hydrochloric acid solution of 10mol?L-1 was used to adjust the filtrate, and then removes the small molecule residual reagents such as metal salts, surfactants, and unreacted reducing agents in the crude solution through dialysis to obtain the wool keratin stock solution [1].

Extraction of wool keratin with TCEP: a certain amount of broken wool was dissolved in a beaker with TCEP solution, bath ratio was 1:15; then filled with nitrogen, water bath for 12h at 90℃; suction filtration for the wool solution, dialysis, centrifugation after 36h, freeze-dried to give keratin [2].

By contrast, both methods can extract wool keratin, but DTT often can not restore the disulfide bond embedded in the protein structure, so TCEP is better. Moreover, DTT is easily oxidized by air. After preparation, it needs to be cryopreserved, and inert gas treatment during the dissolution of wool to ensure the reduction effectiveness. TCEP is extremely selective for reducing disulfide bonds and has a great effect on the molecular weight of keratin, and almost no reaction with other amino acids other than cysteine, can be used in a wider pH range, including acidic conditions (pH=3), thereby effectively reducing the hydrolysis of the amide bond.

One of the key technologies of keratin recycle is how to get the keratin solution that meets the requirements, and the size of the molecular weight determines the use of keratin. TCEP is highly selective for disulfide bonds and can inhibit the hydrolysis of amide bond in wool to a great extent, which provides great convenience for the application of keratin solution. Meanwhile, TCEP has mild reactivity and little toxicity, easier to operate, and more stable in air and aqueous solutions, is a well-recognized alternative to DTT.

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