| As a special elastic tissue of translucent vertebrates,cartilage has the effect of reducing the friction of hard bones and maintaining the lubricity of the friction surface of the joints.This is due to the rich glycosaminoglycans and protein polypeptide molecules in the cell matrix.In recent years,research on the development and utilization of cartilage has risen sharply,and most of them have focused on the extraction of acidic polysaccharides such as chondroitin sulfate.At present,chondroitin sulfate has been proven to have many functions such as antiinflammatory and antioxidant,and has been widely used in health care drugs or clinical medicine.In addition,cartilage protein peptides have also been found to have obvious physiological activities,such as promoting the proliferation and differentiation of osteoblasts,resisting osteoarthritis,and protecting cartilage.The combination of active peptides and active polysaccharides can also significantly increase bone density and improve Anti-inflammatory and antioxidant effect.However,in the process of production and application of chondroitin sulfate,the protein components in cartilage are often removed,causing serious waste of resources.Based on the establishment of a high-performance liquid method for the accurate and rapid determination of chondroitin sulfate,this article uses chicken cartilage as raw material,and uses chondroitin sulfate content and peptide content as indicators,and uses enzymatic hydrolysis technology to prepare targeted preparations with high content of chondroitin sulfate and activity.The cartilage enzymatic hydrolysis products of peptides;and to explore the structural characteristics of chondroitin sulfate in chicken cartilage enzymatic hydrolysates;the rabbit joint damage model is further used to explore the joint protection effects of cartilage enzymatic hydrolysates,which is a high-value development and utilization of chicken cartilage and its enzyme The solution products provide new ideas for the application of joint health protection.The main research contents of this paper are as follows:According to the characteristic that chondroitin sulfate is composed of alternating glucuronic acid and N-acetylgalactosamine disaccharide units,the thesis uses galactosamine,the acid hydrolysis product of chondroitin sulfate,as the measurement target,and separates the high performance liquid chromatography column from the liquid phase.The determination conditions were optimized in order to establish an accurate and rapid method for determining the content of chondroitin sulfate in the enzymatic hydrolysate of chicken cartilage.The optimal determination parameters were as follows: ZORBAX C18 column(250 mm×4.6 mm,5 ?m)and acetonitrile were used,and ammonium acetate solution(0.025 mol/L)as the mobile phase for gradient elution,measured at 250 nm with UV detector.The results showed that the mass concentration of chondroitin sulfate at 100-500 mg/L has a good linear relationship with the peak area.The regression equation is y=3.3626x-9.8376,the repeatability deviation is 0.089%,the average recovery rate is 99.71%,and the relative deviation is 0.35%.Compared with the Elson-Morgan method and the carbazole method,the two traditional methods for the determination of chondroitin sulfate,the HPLC method determined in this paper has less interference from impurities,high accuracy,excellent repeatability,and both recovery and accuracy.Higher advantage.On the basis of establishing an accurate determination of the content of chondroitin sulfate in cartilage extract,using chondroitin sulfate content,protein recovery,peptide content,degree of hydrolysis and antioxidant capacity as screening indicators,a comparative analysis of papain,pancreatin,and complex protease(Protamex),neutral protease and 37071 five kinds of proteases affect the hydrolysis of chicken cartilage.It was found that pancreatin and compound protease(Protamex)showed better hydrolysis.And further explore to optimize the coordinated hydrolysis of the above two enzymes,to obtain the best enzymatic hydrolysis process: materialto-liquid ratio 1:2,amount of enzyme added 0.15%(enzyme/chicken cartilage,w/w),pancreatin:compound protease(Protamex)adding ratio 1 :0.5,p H 6.7,temperature 50 ?,enzymolysis time 6 h,the chondroitin sulfate content of the chicken cartilage enzymatic hydrolysis product prepared under these conditions is as high as 29.8%,the peptide content is 50.3%,and the antioxidant rate reaches 21.3%.Infrared spectroscopy and mass spectrometry are used to explore the structure and composition of chondroitin sulfate.In the infrared spectroscopy,the enzymatic hydrolysate has a characteristic absorption at 852 cm-1,and the hydrogen nuclear magnetic spectrum has a triplet near 2.0,all of which are characteristic signals of CS-4.The chondroitin sulfate in the enzymatic hydrolysate of cartilage is mainly chondroitin 4-sulfate.Through experimental rabbit bone joint modeling experiments,it is found that chicken cartilage enzymatic hydrolysate can significantly reduce rabbit chondrocyte apoptosis,increase cartilage matrix acid polysaccharide content,fight osteoarthritis,and increase cartilage polysaccharide content has significant effects.It is speculated that the sulfuric acid in chicken cartilage Chondroitin and peptides may have synergistic anti-inflammatory effects. |
PDF Full Text Request