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Step-step Enzymatic Hydrolysis And Characterization Of Collagen Peptide From Fish Scales

Posted on:2012-01-31Degree:MasterType:Thesis
Country:ChinaCandidate:L L YangFull Text:PDF
GTID:2181330344453226Subject:Aquatic Products Processing and Storage Engineering
Abstract/Summary:PDF Full Text Request
Fish scales are the by-product of scaly freshwater fish in processing.There are 19.98 million tons freshwater fish in 2008, including 13.1 million tons scaly freshwater fish which could produce approximately 0.65 million tons fish scales every year in China, and most of fish scales has not been utilized except that some were treated as raw materials exported to Japan. Currently much work has been done on the preparation of collagen peptide from fish scales and the evaluation of its characterizations, but most of them were to get higher nitrogen recovery and hydrolysis degree by increasing the dosage of enzyme, extending the time of enzymatic hydrolysis and maintaining pH vaule of the reaction system, and these technology could make the product have overweight bitterness and salty which need be taken off in the industrial production of collagen peptide from fish scales. Used freshwater fish scales as raw materials, the pretreatment and hydrolysis conditions of fish scales were studied after analying the composition of scales of different fish species and different stored time, and then the functional properties of enzymatic hydrolysates from grass carp scales by three proteases during the enzymatic hydrolysis were evaluated, and the aim of this work was to build the preparation of antioxidant peptide from fish scales with step-step enzymatic hydrolysis, providing the foundation of theory and applications for the development and utilization of fish scales. The main results are as follows:(1) The fish scales were determined. The basic composition of scales of Grass carp and Silver carp stored for 0.5 year to 4 year was measured, as all as the composition of scales of different fish species such as grass carp, crucian carp, carp, silver carp and bream fish. There was no significant effect of storage time on collagen content of fish scales. Scales of grass carp, silver carp and bream fish has higher collagen content, but grass carp scales has larger flake and was more convenient for gather and application. And thus the grass carp scales stored 0.5 year was used as raw material for enzymatic hydrolyzate.(2) The effects of hydrolyzing conditions on the enzymatic hydrolysates from grass carp scales by three proteases were studied. There was a significant effect of heat treatment temperature, heat treatment time, initial pH value, substrate concentration, protease dosage on the hydrolyzing degree and nitrogen recovery of the enzymatic hydrolysates (p<0.01). The optimum conditions of Alcalase hydrolysates of grass carp scales were heat treatment temperature 85℃, heat treatment time 1.5h, initial pH value 8.0~8.5, protease dosage 420U/gPro and substrate concentration 1:17(w/w), respectively, and then hydrolyzing temperature and hydrolysis time were 60℃and 4.5h. The optimum conditions of Neutrase and Protamex hydrolysates were heat treatment temperature 85℃, heat treatment time 1.5h, initial pH value 7.0-7.5, protease dosage 450U/g pro and substrate concentrationwere 1:17 (w/w), and hydrolyzing temperature and hydrolysis time were 50℃and 4.5h.(3) The methods of pretreatment of grass carp scales were studied. There was a significant effect of pretreatment (heating-crushing, heating-no crushing, no crushing-no heating) of grass carp scales on nitrogen recovery, hydrolysis degree and free amino acids of enzymatic hydrolysates (p< 0.01), and hydrolysates of fish scales crushed after heating treatment has the highest nitrogen recovery, hydrolysis degree and free amino acids. There was no significant effect of pretreatment of scales on antioxidant activity of enzymic hydrolysates for Neutrase and Protamex hydrolysis, but the trend of antioxidant activity was diametrical for Alcalase hydrolysates. For Alcalase hydrolysates, fish scales which received no treatments of enzyme hydrolysates has the strongest antioxidant activity (IC50 value of DPPH clearance was minimum), but the nitrogen recovery was only 71%, far below that of hydrolysates of fish scales with crushing and heating treatment (about 99%). There was no significant effect of three methods of pretreatment on the molecular weight distribution of enzyme hydrolysis.(4) The characteristics of enzymatic hydrolysates during the enzymatic hydrolysis were studied. The trend rule of nitrogen recovery, hydrolysis degree, free amino acid, emulsification (ESI) and antioxidant activity of three kinds of protease hydrolysates were similar with hydrolysis time extended. When hydrolysis time was between 0-4h, its nitrogen recovery, hydrolysis degree, free amino acid and the DPPH radical clearance were increasing, but ESI and molecular weight was exhibiting a reduced tendency. When hydrolysis time was more than 4h, nitrogen recovery of Alcalase, Protamex and Neutrase hydrolysis maintained at 97%,97% and 90%, and the scavenge of DPPH free radical was declined with time, and there was no significantly effect of hydrolysis time on molecular weight distribution.(5)The preparation of antioxidant peptides with step-step enzymatic hydrolysis was determined. The hydrolysis degree and total free amino acids of the enzymatic hydrolysates of combination of Alcalase and Flavourzyme protease are the highest, when the total enzyme activity are 840u/gpro, the value of which was 15% and 130mg/g fish scales-DPPH radical clearance increased with the increment of total enzyme activity. When total enzyme activity was more than 525 u/g pro, enzymatic hydrolysates has highest ability of clearance of DPPH free radical when the protease of the second step enzyme hydrolysis was Protamex; when total enzyme activity was 840 u/gpro, the enzymatic hydrolysis of combination of Alcalase and Protamex has smaller average molecular weight, and only 14% of them are above 9952Da while 50% of them are under 5946Da.
Keywords/Search Tags:Grass carp scales, Collagen peptide, Scavenge of DPPH free radical, The molecular weight distribution, Antioxidant peptides
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