| Chinese herbal medicine Euryale ferox is the only species in Euryale genus of the family Nymphaeaceae, it has been recorded in Shen Nong’s Herbal earliest. Euryale ferox is considered to have the function of tonifying spleen and kidney, arresting seminal emission or leucorrhoea, appetizing and antidiarrheal. Euryale ferox is also a kind of natural invigorant with comestible and medicinal value. In this paper, we choose Euryale ferox seeds to study its chemical composition and inhibition effect of proteinuria induced by diabetic nephropathy.This study includes the following parts:1. Optimization extract solventsUsing different concentrations of ethanol or acetone as extraction solvents, 80% ethanol has been chosen as the large scale extraction solvent, based on the extraction yield and the content of phenolic. Then after extraction by ethanol, water extraction is made.2. Determination of anti-oxidant abilityDetermination of the antioxidant ability in vitro of Euryale ferox extract, the result shows that when the concentrations of ethanol extract reaching 0.6,0.8, 1.Omg/ml, it can clear O2-, OH-, and DPPH free radical obviously.3. Acute toxicity testing18-22g KM mice are chosen for acute toxicity test and each mouse is administrated at dose of 20g/kg Euryale ferox extract. The result shows that 20g/kg is the safe dose and no mice deaths.4. Decrease urine protein testSprague Dawley male rats, weighting 150±20g, are divided into normal group, operation group and non-operation group. The operation group undergoes left uninephrectomy first. Then all rats except normal group are given a single intraperitoneal injection with STZ at a dose of 60mg/kg. After that rats are administrated with different dose of Euryale ferox ethanol extract (75mg/kg, 150mg/kg,300mg/kg, and 600mg/kg) for 12 weeks, collecting 24 hours urine for detecting urinary protein and urinary microalbumin per four weeks. And blood serums are collected for glucose, Scr, BUN, TC and TG after rats sacrificed. Results show that Euryale ferox ethanol extract may decrease 24h urinary protein and urinary microalbumin significantly at dose of 300mg/kg and 600mg/kg. These dosages also can ameliorate kidney damage process.5. Determination contentsFolin-Phenol method, vanillic aldehyde-glacial acetic acid-perchloric acid method, phenol-sulfuric acid method and m-hydroxybiphenyl method are combined to determine the phenolic content, terpenoid content and glycoside content. The results show that the content of these three compositions are 27.3%, 22.4% and 26.0% in ethanol extract.6. UPLC-MS analysisEuryale ferox ethanol extract is detected by UPLC-MS. Ten possible compositions are identified as gallic acid, ethyl gallate,5,7,3’,4’,5’-pentahydroxy-flavanone,5,7,4’-trihydroxy-flavanone, epicatechin gallate and isolariciresinol- 9- O-β- D- glucopyranoside, galloyl glucosidase, proanthocyanidins, polyhydroxy xanthone, tannic acid.7. Monosaccharide composition and molecular weight analysisDetecting the monosaccharide composition and the molecular weight distribution of ethanol extract, aqueous extract supernatant and sediment, results show that the main monosaccharide composition is glucose. The molecular weight is focused on less than 2500 in ethanol extract. The molecular weight of supernatant and sediment are more than 10000. |
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