Hard clam polypeptide, and preparation method and application thereof
A technology of clams and amino acids, which is applied in the field of marine biology, can solve many problems, and achieve a wide range of applications, good economic and social effects, and good anti-oxidation effects
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Embodiment 1
[0041] Embodiment 1, preparation of clam polypeptide MmP13
[0042] Fresh clams are shelled, washed with water for 3-4 times to remove the sediment, and the soft clam is fully homogenized. The homogenate is centrifuged at 5000g for 10 minutes, the supernatant is vacuum freeze-dried, and extracted with 60-80% ethanol for 24 hours , centrifuged at 10,000g for 30min, and the supernatant was intercepted by an ultrafiltration membrane with a molecular weight cut-off of 3kDa to obtain an ultrafiltration fraction with a MW<3kDa.
[0043] Sephadex G-25 gel chromatography column (1.5cm×1.2m, Bio-Rad Company) was used to separate the polypeptide components with MWfigure 1
[0044] Depend on figure 1 The results showed that the peptide fractions with MW<3kDa were separated by gel chromatography, and five fractions were mainly obtained, namely F1, F2, F3, F4 and F5.
[0045] Take the F1 component and dissolve it in an aqueous solution containing 0.1v / v% formic acid and 2v / v% acetonitril...
Embodiment 2
[0048] Example 2, Anti-paraquat-induced oxidative stress experiment
[0049] The clam polypeptide MmP13 prepared in Example 1 was added to the Caenorhabditis elegans culture medium prepared by 70mM paraquat according to the concentrations of 0.5mM, 1mM, 2mM and 4mM respectively, and 2mM reduced glutathione (GSH) was added to the positive control group as a positive control group. Positive test substances, and the blank control group plus an equal volume of S Medium were incubated at 20°C, and the survival rate of Caenorhabditis elegans was counted every 12 hours until all the worms died. Survival rate is represented by survival curve, and Kaplan-meier method is used to analyze and compare the difference between the clam polypeptide MmP13 and its analogue group and the control group. The result is as Figure 4 shown.
[0050] The results showed that the clam polypeptide MmP13 prepared in Example 1 had significant antioxidant effect when the concentration was 0.5mM, 1mM, 2mM a...
Embodiment 3
[0051] Embodiment 3, life experiment
[0052] The clam polypeptide MmP13 prepared in Example 1 was prepared according to the concentration of 1mM, 2mM and 4mM, and the analogs MmP13-1, MmP13-2, MmP13-3, MmP13-4, MmP13-5 of the clam polypeptide MmP13 prepared in Example 1 At a concentration of 4mM, add it to wild-type C. elegans in the early adult stage, and add an equal volume of S Medium to the control group, place it at 20°C for continuous culture, and count the survival of C. elegans in each group every other day under the microscope. , until all C. elegans died. The survival rate of nematodes in each group was statistically processed, and the results were expressed as survival curves. The Kaplan-meier method was used to analyze and compare the differences between clam polypeptide MmP13 and its analogs and the control group. The result is as Figure 5-10 shown.
[0053] The results show that the clam polypeptide MmP13 prepared in Example 1 can prolong the average surviva...
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