Preparation And Biological Activity Of Deer Antler Collagenase Hydrolysates

In this paper, we extracted collagen from sika deer antler with water at the optimal conditions and hydrolyzed by protease, which enhances the solubility of collagen.The methods such as thin-layer chromatography (TLC),high performance liquid chromatography (HPLC), ultraviolet spectrum (UV), infra-red spectrum(IR), circular dichroism spectrum (CD) and so on was used in studying on the physical-chemical properties of CSDV. We did the active test to the collagen. Osteoporosis rat models were established by ovariectomy and retinoid acid (RA).One kind of health food products was developed from sika deer antler collagen. And we lay down quality standard and operating procedure.Here are the details:1 Study on the optimum conditions of extraction of CSDV.The collagen protein quantity was measured with folin-phenol reagent, and as a target, we want to know the influence of different liquid ratio (1:1,1:2,1:3,1:4,1:5), different extraction time (1 h , 2 h, 3 h, 4 h, 5 h), different extraction times (1,2,3,4,5 times) three factors on the extraction yield of collagen. That was compared by single factor and orthogonal experiment. It was found that the greatest impact on the extraction rate was extraction times, followed by extraction time, solid to liquid ratio of no significant effect on the extraction rate. The optimum extraction conditions namely: liquid ratio 1:3, extracted 5 times, 4 h per times.By single factor experiments, we screen optimum hydrolysis conditions suchas the of the hydrolysis time (0 h, 0.5 h, 1 h, 2 h) and its and the role of the ratio of substrate (1:3000,1:5000,1:8000,1:10000) with the target molecular weight of collagen.Results were that optimal trypsin was enzyme and substrate is the ratio of 1:5000, hydrolysis time to 2 h. Validation tests confirmed that under the above conditions, the extraction ratio was 12.52 percent on average, and the average protein content was 56.89%.2 Preliminary study on physical and chemical properties.By SDS-polyacrylamide gel electrophoresis (SDS-PAGE), molecular weight of CSDV were measured .The result showed it under 31 kDa, consistent with the result of HPLC. Acid hydrolysis was applicated in amino acid composition analysis on CSDV.Result show that the highest content is glycin, nearly 1/3 in the CSDV, and there is 15.50% proline and 13.87% alanine of it. Unusual amino acids - ydroxyproline content was very high too, nearly 6.71%. This result is consistent with characteristics of collagen amino acids. TLC seized at the same position with the standard collagen spots appear. Ninhydrin color reaction was blue-violet. Water content of the test results showed 8.24% moisture content. The effect of viscosity with temperature changes was researched; the viscosity decreased rapidly when collagen was enzymolysised.The UVspectra, IR, CD spectra was used in researching the characters of sika deer antler collagen. UV absorption spectra showed its maximum absorption at 260 nm,α-helix,β-fold and the number of random coil were 3.6%, 42.7% and 53.7% analyzed by CD.3 Acute toxicity of CSDV was studied.The acute oral toxicity test for this disinfectant indicated that it belonged to practically non-toxic grade.4 Growth and development effects of CSDV on gonadal related organs.Serum glucose and testosterone levels detected CSDV can contribute to weight gain in mice, And the weight of relevant organs of mouse prostate glands - the seminal vesicles, testes, levator ani muscles - sponge ball muscle increased, blood testosterone increased, Meanwhile, the thymus weight increased, blood glucose decreased.5 Anti-inflammatory, immune and anti-stress effects of CSDV.The anti-inflammatory, immune and anti-stress effect was obvious. Experimental results show that foot swelling caused by carrageenin in mice inhibition by CSDV in 0.2 g?kg-1, P<0.05; dinitrochlorobenzene induced delayed-type hypersensitivity was inhibited when the dose of 0.5 g?kg-1 and 0.25 g?kg-1, and may to some extent to promote macrophage phagocytosis. Anti-stress test results show that when the dose on 0.5 g?kg-1 survival time lengthen; and on 1.0 g?kg-1, 0.5 g?kg-1 and 0.25 g?kg-1 can extend the swimming time, but only 1.0 g?kg-1 and 0.5 g?kg-1 significantly different.6 Prevention and therapeutic effects of CSDV on osteoporosis in rats.To evaluate the preventive and therapeutic effects of the collagen extracted from Sika deer velvet (CSDV) on osteoporosis in ovariectomized rats. Compared with the ovariectomized group, CSDV markedly improve bone mineral density (BMD) and bone weight in ovariectomized rats, regulate the ALP and BGP in serum. There was marked difference in biomechanical properties and static histomorphometric indexes, such as Maximum load, Maximum deflection, bone stress, bone strain, TBV, MTPS, and MBCT between the collagen-treated groups and without.CSDV was found to play a role in the prevention and treatment of osteoporosis in ovariectomized rats.The results of the study on prevention and therapeutic effects of CSDV on osteoporosis in rats by retinoic acid was similary with that in ovarectomized rats.7 Anti radiation effects of CSDV was researched.Results show that CSDV can only reduce the number of micronuclei bone marrow cells caused byγ-ray irradiation.8 Study on promoting cell adhesion in vitro effects of CSDV.The results of the experiments in vitro showed that there no effect on the cell adhesion in vitro.9 One kind of health food was produced basis on the sika deer antler.The chewable of CSDV was produced, and the quality standard was researching.