Study Of Antiosteoporotic Activity Of Collagen Peptid And Psoralea Corylifolia Extract

Fructus Psoraleae is the dried fruits of the leguminous plant Psoralea Corylifolia L. It is recorded in the china pharmacopoeia(2005 edition).Fructus Psoraleae is a common tonic chinese medicine which is pungent and bitter in flavor,warm in nature,and attributive to the kidney and spleen meridians.It invigorates the kidney and strengthens yang,warm the spleen and stop diarrhea.The doctors in past dynasties used Fructus Psoraleae for invigorating the kidney and strengthening the bones.The circs of clinical applications and modern study of pharmacology suggest that Fructus Psoraleae can assuredly strengthen the bones and cure osteoporosis.Collagen is the main component of extra-cellular matrix (ECM). It is very important to maintain the flexibility of skin and blood vessel, keep hair and nail soft and lustrous, improve the lubricative property of cartilage. Collagen peptid have anti-osteoporotic, antioxi dative, ACE inhibitory activity,have received increasing attention for their unique physiochemical property and excellent physiological active function.The collagen peptid from Tilapia fish scale has been extracted by using pepsin method. The effects of six factors including enzymatic species,enzyme concentration,substrate concentration,hydrolyzing time,temperature and pH. The best enzymatic hydrolysis condition was determined based on extraction ratio and average molecular weight of peptid. The optimum condition of enzymatic species,enzyme concentration, substrate concentration,hydrolyzing time,temperature and pH for the collagen peptid as 1%MG+1%HHM,150g/L,12h,50℃,pH6.WST-8 assay and ELISA were used to detect the proliferation and the secretion of type I collagen of cultured osteoblast MC3T3-E1 cell. The results of the experiment showed that the collagen peptid can stimulate the mouse osteoblasts proliferation,and can increase type I collagen secretion.Draw a comparison between the composition and content of amino acids in some aquatic animals and mammal by HITACHI L-8800 Amino Acid Automatic Analyzer.18 kinds of amino acids were assayed and 8 of all were essential or conditionally essential amino acids.There was a significant difference in the content of methionine, threonine and hydroxyproline between aquatic animals and mammal. The content of essential amino acid methionine, threonine in aquatic animals, it was significantly higher than mammal. The difference of amino acids content can be used as reference of rational development.Fructus psoraleae preparation was extracted with distilled water,30% alcohol,60% alcohol,95% alcohol. A mouse osteoblast cell line MC3T3-E1 was used as a cell model for screening potency. The content of ALP and type I collagen was measured. The results of the experiment showed that 95% alcoholic extracts could stimulate the activity of ALP,and could increase type I collagen secretion. The expression of ALP,type I collagen and osteocalcin mRNA in MC3T3-E1 cell was detected by real-time PCR.The 95% alcoholic extracts enhanced the expression of ALP,type I collagen and osteocalcin mRNA in MC3T3-E1 cell. The expression of ALP mRNA was enhanced for 7-14d,and the expression of type I collagen and osteocalcin mRNA was enhanced for 14-21d.To investigate the effect of fructus psoraleae and fish collagen peptid on the differentiation of cultured osteoblast MC3T3-E1 cell. A mouse osteoblast cell line MC3T3-E1 was used as a cell model for screening potency. It was cultured and exposed to different concentrations and proportion of fructus psoraleae extract and fish collagen peptid. The expression of ALP,type I collagen mRNA in MC3T3-E1 cell was detected by real-time PCR. The results showed that type I collagen mRNA was gradually increased. ALP activity reached the maximum level on the 12th day. The extract of fructus psoralea and fish collagen peptid enhanced the expression of ALP,type I collagen mRNA in MC3T3-E1 cell, It was significantly higher than fructus psoralea group,the optimum proportion was 1:1,and the concentration was 0.2mg/mL. The results showed that there was a synergic action of fructus psoraleae and fish collagen peptid to stimulate the differentiation of osteoblasts in vitro.The anti-osteoporotic effectiveness and safety of fructus psoraleae and fish collagen peptid were ssessed on bone formation in rat in vivo. Growthing rat was used as model, the rat bone mineral density(BMD) was measured by dual-energy X-ray absorptiometry(DXA).The results showed that there was no significant change of rat bone mineral density in femur and tibia. The results of chronic toxicity experiment showed that there was no evident toxic effect. Ovariectomized rat was used as model, ssessed the anti-osteoporotic effectiveness of fructus psoraleae and fish collagen peptid, The results showed that fructus psoraleae and fish collagen peptid could significantly increase the BMD in ovariectomized rats, and could decline the excretion of urine calcium.The study suggested that fructus psoraleae and fish collagen peptid could regulate the osteoporosis of female rat without ovary.The research combined the nanomaterials collagen peptid with traditional Chinese medicine Fructus Psoraleae, proved there was a synergic action of fructus psoraleae and fish collagen peptid to prevent or alleviate osteoporosis.As a new attempt, it was provided a new research idea for further work on discovery and development of new bone-building drug.