| The adult of Martianus dermestoides Chevrolat has high economic value for its abundant functional ingredients, such as necessary amino acids, vitamins, microelements and unsaturated fatty acids et al. This paper attemped to make a research on extraction of protein, enzymed protein, it was focused on the antioxidative activity of Martianus dermestoides Chevrolat's beetles protein hydrolysates including the optimization of its hydrolysis process, isolation, and structural analysis of the hydrolysates with the highest functional activity. The results showed:The first, make a research on extraction of Martianus dermestoides Chevrolat's beetles protein. This experiment adopted three kind of protein extracting methods, they were extract method of alkali, the extract method of salt, the extract method of Tris-HCL buffer solution, respectively. This study aimed at filtering the best extract method of proteins and making sure of the technique guide lines of different measures respectively from fresh beetles Martianus dermestoides Chevrolat, the results showed that alkali extract method was the best of three, which obtained the highest proteins, the second was the Tris-HCl extract method, and the last was the salt extract method. The relevant condition of alkali method was that, concentration being 1.0%, temperature being 80℃, time being 60min, with the ratio of solid versus liquid being 1:15. The best condition of salt method was that, concentration being 1.0%, temperature being 20℃, time being 2h, with the ratio of solid versus liquid being 1:10. The best condition of Tris-HCl method was that, pH being 8.1, temperature being 10℃, time being 30min, with the ratio of solid versus liquid being 1:10.The defatted Martianus dermestoides Chevrolat's beetles protein was hydrolyzed with four different proteases (Papain, Neutral, Trypsin, and Pepsin), and the degree of hydrolysis, the stripping ratio of protein and the length of peptide chain were examined. Result show that the hydrolysates using Papain has the strongest advantage. So Papain was established as the first enzyme.The effects of pH, temperature, enzyme, time and substrate concentration on degree of hydrolysis were studied. With responsive surface methodology, the preparation conditions of the hydrolysates were optimized: the papain hydrolysates were studied. By choosing the amount of enzyme added as 2225U/g (the beetles protein), temperature as 42℃, and time as 4h, a relatively high degree of hydrolysis (22.67%) could be achieved, efficiently and energysavingly.The zymolytes were cut off into three parts by ultrafilt ration membranes with 10000 MWCO (Molecular Weight Cut Off), 6000 MWCO respectively. In the system of non-oil, the antioxidative activity of the different parts of peptides were studied respectively. The result showed that the different of the hydrolysate has a strong redical scavenging activity in eliminating DPPH·, OH·, the determination of power and superoxide anion radicals. Meanwhile the research found that the first part of the hydrolysates has the strongest activity. Zymolytes can clear superoxide negative ion (O2·), but the effect of clear was not far from Vc. when clear hydroxyl free radical (·OH), the effect was equivalence, when clear 1, 1-dipHenyl-2picrylh-yrazyl free radical (DPPH·) and the determination of power, the zymolyte was stronger than VcFinally, in the experiment of anti-aging, we choosed healthy animals. Each weight was 20±2g. 80 mice were randomly assigned to six groups, with 12 replications per group. The six groups were one control group, one aging model group, one VE control group, and three experiment groups. D-galactose was used to build aging model. Mice of experimental groups were perfused by three dose of zymolytes everyday. They were killed by extirpating eyeballs after 35 days. Serum, liver and brain were separated to determine biochemical parameters. Results showed that. The effect of zymolytes on activity of SOD in liver showed that, there was very significant difference observed between middle dose group, high dose group and aging model group(P＜0.01), and significant difference between high dose group and aging model group(P＜0.05). And the middle dose group was best and there was very significant difference observed (P＜0.01). The effect of AAP on activity of SOD in serum and in brain showed that, there was significant difference observed between middle dose group and aging mode group (P＜0.05), however, no significant difference observed among other groups. The effect of zymolytes on activity of GSH-PX in serum showed that, there was very significant difference observed between middle dose group and aging model group (P＜0.01), and significant difference between high, middle doze groups and aging model group (P＜0.05). The effect of zymolytes on activity of GSH-PX in liver showed that, there was very significant difference observed between high, middle doze groups and aging model group (P＜0.01), and significant difference between high, low dose groups and aging model group (P＜0.05). The effect of zymolytes on content of MDA in serum showed that, there was very significant difference observed between middle dose group, low dose group and aging model group (P＜0.01). The protein zymolytes of Martianus dermestoides Chevrola extracts could distinctly delay the mice's aging, zymolytes had a good anti-aging action. |
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