Manufacture And Chelation Mechanism Of Peptide Chelating Calcium Prepared From Catfish By-products

The manufacturing process of freshwater fish surimi from catfish can produce 50-60%of by-products such as fish bone,head,skin,frittered meat and so on.Thus taking advantage of these by-products is important to improve additional value and is beneficial to practical production.The present study use sing enzymolysis-fermentation technology to prepare peptide chelating calcium from catfish by-products.In order to made peptide chelating calcium,firstly an efficient protease was chosen out of three,after enzymatic hydrolysis the by-products were further fermented by lactic acid bacteria,meanwhile both the process of enzymatic hydrolysis and fermentation were optimized.Then the structure of the peptide chelating calcium was characterized by using ultraviolet absorption spectrum,infrared absorption spectrum,scanning electron microscopy and x-ray photoelectron spectroscopy,the antioxidant and antibacterial properties of the peptide chelating calcium was analyzed too.Finally a new kind of calcium tablet was developed from the peptides chelating calcium.The main conclusions of the research are as follows:(1)Manufacture catfish flesh and bone paste from catfish by-products including fish head,skin,bones,fin and fish meat,and analyze its nutrient component.Treat the catfish flesh and bone paste with three kinds of protease(neutral protease,alkaline protease and pepsin),using calcium dissolution rate as the main index,combined with the yield of peptides,to determine the most efficient protease.The results showed that the total protein content was 15.50%,the total fat was 6.08% in catfish flesh and bone paste,and the total ash was 12.25%,in which the calcium content is high(42.60 g/kg),and the calcium/phosphorus ratio is 1.71 close to the best ratio for human.Pepsin was chosen as the protease to prepare peptide chelating calcium as it gave a calcium dissolution rate 57.04%,which was 50%higher than the other two enzymes,and a more peptide yield 44.40%.(2)Digest the catfish flesh and bone paste with pepsin,optimize three main factors as the amount of enzyme,enzymolysis time,and liquid-solid ratio,on the basis of single factor experiments,and further with response surface analysis,taking peptide yield and dissolution rate of calcium as the response.The result proved the optimum digestion conditions as:liquid-solid ratio(water/paste)6.73/1,pepsin 7000 U/g,enzymolysis time 6 h at 37 oC under pH 3.0.Under the optimum condition,the polypeptide conversion rate was 43.07%,the calcium dissolution rate was 44.62% in the catfish flesh and bone paste.(3)Collect the precipitation of the catfish flesh and bone paste after degestion by pepsin under the optimum digestion conditions,ferment the above precipitation using two kinds of lactic acid bacteria(Lyocarni SHI-59 and Lyocarni Pro-MIX5),to increase more peptide and dissolved calcium.The result of the response surface analysis showed the best condition for Lyocarni Pro-MIX5 fermentation was: the substrate content 20.12%,the inoculation quantity6.57 lg(cfu/g),fermentation time 24.12 h.Under this condition,the peptide yield was 11.8%,the calcium dissolution rate was 0.33%.And the best fermentation condition for Lyocarni SHI-59 was: substrate content 21.02%,the inoculation quantity 6.91 lg(cfu/g),fermentation time 35.28 h.Under this condition,the peptide yield was 14.3%,and calcium dissolution rate was 0.31%.Lyocarni SHI-59 was more suitable for catfish flesh and bone paste fermentation.(4)Calculate the calcium chelating rate by peptide in enzymatic hydrolysate and fermented supernate.Characterize the peptide chelating calcium by ultraviolet absorption spectrum,infrared absorption spectrum,scanning electron microscopy and X-ray photoelectron spectroscopy analysis,and evaluate its antioxidant and bacteria inhibition activity.The results showed that:(1)The maximum ultraviolet absorption peak of peptides chelating calcium had a red shift compared to polypeptide,evidenced a chelate formation that changed the light absorption performance of the ligand.(2)The N-H stretching vibration absorption peak of the peptides chelating calcium showed a blue shift,and the wave grew wider,meanwhile the N-H stretching vibration absorption peak for amide II disappeared,suggesting a integration of calcium ions with amino.There was a red shift in C=O stretching vibration absorption peak,and the Deformation vibration fingerprint region for O-H disappeared,evidenced a integration of calcium ions with carboxyl;(3)The scanning electron microscopy showed many calcium calcium crystals adhered to the smooth surface to the polypeptide;(4)X-ray photoelectron spectroscopy analysis proved that the elements in polypeptide surface were mainly O,C,P,Na and N,but the elements in the surface of peptides chelating calcium were mainly C,O,Ca,N,P,Mg and Na.The above mentioned analysis from four aspects,all proved that the mixture prepared by enzymolysis and fermentation from catfish flesh and bone paste was peptide chelating calcium.The calcium chelating rate in the enzymatic hydrolysate was 65.78%,and 58.94% in the fermented supernate.The peptide chelate calcium had a weaker antioxidant ability and lower reducing capacity than ascorbic acid.But it was more efficient in superoxide anion free radical and DPPH scavenging.The peptide chelating calcium had better antibacterial activity to gram positive bacteria than to gram-negative bacteria,but its efficiency was significantly lowerthan tea polyphenol.(5)Peptides chelating calcium tablets were made with 100 g peptides chelating calcium powder,adding 7.97% magnesium stearate,55.21% microcrystalline cellulose,and 5.94%CMC-Na.The calcium tablets had smooth surface,beautiful color and good taste,and the calcium content in the calcium tablet was detected to be 28.22 mg/g.