| Rapeseed is one of the important oilseed crops in China. The main by-product of the oil extraction process is rapeseed meal, which has relatively high protein content of 35-45%, making it an attractive and promising source of vegetable proteins. Annually, there is a rich production of 6.0×10~9 kg rapeseed meal in China. However, the presence of anti-nutritional compounds is a major drawback in the use of this bioresource as human food. Hence, this protein rich meal is usually only used for animal feed and fertilizer at present. Thus, efforts are being made to develop efficient methods to recovery proteins from cold-pressing double-low defatted rapeseed meal and to prepare acceptable products for human consumption. The purpose of this study was to provide some useful academic information and novel effective approach for deep processing of defatted rapeseed meal and developed some high value-added products for human consumption.In the present study, the preparation of rapeseed protein concentrate (RPC) with the technology of ethanol leaching and novel methods of utilizing ultrasonic and microwave to modify alcohol leached rapeseed protein concentrate were studied; the possibility of using low concentration of alkali extraction combining with ultra-filtration to prepared rapeseed protein from defatted rapeseed meal were also studied; the Alcalase was selected to hydrolysis rapeseed protein for obtaining the rapeseed protein hydrolysates (RPH); several kinds of experimental systems in vitro and the animal experiments were used to evaluate the antioxidative activities of rapeseed protein hydrolysates. The results are as follows:The effects of ethanol concentration, extraction temperature, extraction times and the ratio of solid to liquid on the protein content were investigated. The parameters of extraction process were optimized through orthogonal test and average. The best extraction condition is ethanol concentration 70%, 25 min at 55℃, 5 times of extraction, extraction time 25 min, the ratio of solid to liquid 1:6. Under this condition the content and the yield of protein were 60.5% and 75.7% respectively, the removal rate of the anti-nutritional compounds such as tannin, phytic acid and glucosinolate were 90.5%, 66.7%, 99.3% respectively. The rapeseed protein concentrate could reach the national standard (GB NY/T 126-2005) of first grade rapeseed meal for feedstuffs. The orthogonal test results showed that the optimum ultrasonic modification parameters were as follows: RPC concentration 3%, modification time 5 min, ultrasonic power 300 W and pH 11. Under the above-mentioned modification parameters, the modified alcohol leached rapeseed protein concentrate with Nitrogen Solubility Index (NSI) 86.94% can be obtained. The orthogonal test results showed that the optimum microwave modification parameters were as follows: RPC concentration 5%, microwave power 300 W, modification time 1 minute and pH value 10, under the above-mentioned modification parameters, the modified alcohol leached rapeseed protein concentrate with NSI 73.38% and foaming stability 91.25% can be obtained.On the basis of low concentration of alkali extraction combining with ultra-filtration, the preparation of rapeseed protein from defatted meal was studied and the respective reaction conditions were optimized. With the extraction rate of protein as the index and through the optimization of single factor and orthogonal experiments, the optimum conditions of the protein extraction were: colloid mill clearance 10μm, pH 9.5, 50℃, 3 times extraction, the ratio of solid to liquid 1:15, and the time of extraction was 1.2 h, precipitation of protein at pH4.5. The extraction rate of protein was 88.1%. The optimum technical conditions of ultra-filtration process were: pH 9.0, operating temperature 40℃, operating pressure 0.20 MPa, 5 h. The volume flow of solution through membrane recovers 90% of original by washing. The intake and exit temperature of the spray dryer were 180℃and 80℃respectively. Through centrifugal spray drying to the homogenized protein solution, the flaxen rapeseed protein was obtained, the content and the yield of the rapeseed protein were 66.5% and 70% respectively, the content of the anti-nutritional compounds were not detected. The physicochemical, functional and nutritional properties of the rapeseed protein were comprehensive studied. Results indicated that rapeseed protein has relatively high quality protein due to their well-balanced amino acid composition and high nutritive value.Four proteases of food grade, including Alcalase, Protamex, Neutrase and Flavorzyme, were selected to hydrolysis rapeseed protein, and the degree of hydrolysis (DH) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activities of the resulting hydrolysates were determined. There results showed that the hydrolysates of Alcalase possess strong antioxidant potential than others, therefore, the Alcalase was determined to be used to hydrolysis rapeseed protein for acquire rapeseed protein hydrolysates with high antioxidative activities. The optimal conditions for the enzyme hydrolysis are as follows: 55℃, pH 8.5, concentration of substrate 2.5%, enzyme/substrate 2.0%, hydrolysis time 4h. Under these conditions, the DPPH radical scavenging activity of the resulting rapeseed protein hydrolysates was 76.16% and the degree of hydrolysis is 24.45%. The antioxidative activity of the resulting hydrolysate is the strongest and the Molecular mass of the main composition with high antioxidative activity of the hydrolysates seperated by ultra-filtration was lower than 3 kDa.Rapeseed protein hydrolysates prepared as per the protocol in this study exhibited antioxidant function in five different test models in vitro. The free radical scavenging activities of RPH were concentration dependent. Antioxidative activity of Rapeseed protein hydrolysates was also observed in senile animal model induced by injection of D-galactose. The results showed that the rapeseed protein hydrolysates can effectively prevent the elevation of MDA level in serum and liver and prevent the decrease of SOD and GSH-Px activities in serum, liver and kidney induced by D-galactose. These results showed that rapeseed protein hydrolysates have obviously antioxidative activity in vivo. Rapeseed protein hydrolysates containing a high content of essential amino acids is suggestive of its high nutritive value. The results obtained in this study clearly suggest that rapeseed protein hydrolysates is a natural source for antioxidants, which could serve as functional food ingredient with potential applications in food products and related health benefits.
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