Research On Preparation And Characterization Of Collagen From Skin Of Grass Carp

In this paper, we study grass carp skin pretreatment conditions, using the method of sodium hydroxide adding hydrogen peroxide can bleaching and removing non-collagen and defatted to grass carp skin, the role of its collagen solubility effects. Effects of the key parameter extraction—temperature to the collagen extraction rate and characterization, Characterization different between Acid-soluble collagen and enzyme-soluble collagen; Purifying typeâ… and typeâ…¢collagens from Grass carp skin and pig skin and electrophoresis identification.1. Different concentrations of hydrogen peroxide and sodium hydroxide treatment grass carp skin, can also play a role on bleaching, removing non-collagen and defatted, the higher the concentration of sodium hydroxide, the faster shedding pigment. Concentration of hydrogen peroxide had no significant effect on bleaching. In 0.01 mol/L the sodium hydroxide solution containing 1% hydrogen peroxide, skin floating in surface of the solution, pigment gradual withdrawal, unable to see melanin turn to yellow; 0.01 mol/L-0.2 mol/L sodium hydroxide can effectively remove the skin of non-collagenous proteins, However, the higher concentration of sodium hydroxide, may lead to the degradation of collagen protein in the pretreatment solution; At 20℃conditions, 0.01 mol/L NaOH solution containing 1% H₂O₂ can effectively remove fat, defatted rate about 70%.2. The treatment of different concentrations of hydrogen peroxide and sodium hydroxide, have an impact on the Characterizations of collagen. At 20℃, 1% hydrogen peroxide, sodium hydroxide concentration solution, leaching treatment 24 hours later, have great impact on the Characterizations of collagen, even in the acid+enzyme xtracted 48 hours, Extraction rate only 8%. In the conditions of 0.01 mol/L NaOH or 4℃for dealing with collagen, have less impact on the collagen extraction rate, so it can only choose lower concentrations of sodium hydroxide or low temperature skin.3. Key parameter temperature of extraction of collagen and pepsin have great impact on the extraction rate. At 4℃in the acetic acid+pepsin solution extraction rate more than double extraction rate in acetic acid, At 20℃and 30℃, the extraction rate of solution containing pepsin is far higher than that of acetic acid. The higher the temperature, the higher collagen extraction rate. However, SDS-PAGE showed that peptide chain of collagen extracted at 30℃degrade, indicating collagen denaturation. Collagen extracted at 20℃and below 20℃,triple helix structure keep intact with a complete biological activity.4. Acid-soluble and pepsin-soluble collagens (ASC and PSC) were extracted from the skin of grass carp and partially characterized. the UV spectra of ASC and PSC from grass carp showed that the distinct absorption was obtained near 223 nm; SDS-PAGE showed that acid-soluble collagen protein contain moreβ,γand polymer. Molecular weight of acid-soluble collagen higher than it of pepsin-soluble collagen, The denaturation temperature of ASC and PSC from grass carp was 33.8℃and 34.5℃, respectively, about 3℃lower than that of the porcine skin collagen(37℃). These results suggest that grass carp skin has potential application in the functional food, healthcare, cosmetic, and pharmaceutical industries.5. Under the condition of pH 7.4, different salt concentrations fraction precipitate collagens from grass carp skin and pig skin.Typeâ…¢collagen were completely precipitated at 1.8 mol/L salt concentration,at this time the majority of typeâ… collagen dissolved in the solution, in the final were precipitated at 2.5 mol/L salt concentration. the samples purifying from varying salt concentration, analyzed by SDS-PAGE containing urea. It can make distinction between the electrophoresis mobility of a1(â… ) and a1(â…¢) which used be the same. SDS-PAGE showed that grass carp skin samples without a1(â…¢), however the pigskin contains a1(â…¢). but purity of typeâ…¢collagen protein is not high, only about 20%. Typeâ… collagen reached electrophoresis purity. Conclusion: grass carp skin contains only typeâ… collagen without proteinâ…¢collagen, Pigskin main contain typeâ… collagen protein also contain a small amount of typeâ…¢collagen.