Studies On Extraction, Separation, Determination And Hypoglycemic Activity Of Effective Components In Fresh Rehmannia Root

Radix Rehmanniae is the tuberous root of Rehmannia glutinosa Libosch. of the family Scrophulariaceae, which can be processed to fresh rehmannia root, rehmanniae praeparatum and Radix Rehmanniae preparata. It backs to Shennong bencao jing and is recorded by Chinese Pharmacopoeia (2005 Edition) now and it is mainly distributed in Henan, Hebei, Shandong, Shanxi, Inner mongolia, and now cultivated in most parts of China. It is widely used in clinic and diabetes with a long history. Documents reported that the hypoglycemic activity is saccharide, catalpol and rehmannioside D, which of the last two are the relative higher contents in iridoid glycosides.Research of the pharmacological effect of Radix Rehmanniae reported that the active component of hypoglycemic is more than one content, but did not report the difference of them. In order to identify the difference of them, need to get the departed components. Up to now, the main components of lowing blood sugar in Radix rehmanniae is oligosaccharide and iridoid glycosides, which are similar in polarity and solubility, so it is difficult to be separated. First of all, we should separate and enrich them with meeting the requirements of the total content of effective components is more than 50% respectively.In order to guarantee the quality of finished products and semi-manufactured goods and provide reasons of dosage, the content of the effective compositions needed to know in the whole process of discovery and development. The determination method of iridoid glycosides is not easy to establish because its chemical characters, which obstructs the development of Radix rehmanniae.To solve the above problems and develop Chinese new drug, we investigated the extraction and isolation technology, the determination method and studied the pharmacological effect of Rehmannia glutinosa oligosaccharide (ROS) and iridoid glycosides.1. Studies on the extraction and separation technology of ROS and iridoid glycosidesDetection of indexes: catalpol and saccharide which was monitored by the means of HPLC and Molish teast respectively. We studied the technology of extraction and separation of saccharide and iridod glycosides, then separated them successfully.The objective products were extracted effectively by solvent distillation witn using suitable solvent, quantity of solvent, extraction times and extraction process. Then, the efficiency of 4 types of macroreticular resins were compared and draw the conclusion that H103 was the most suitable resin to separate ROS and iridoid glycosides. And the content of ROS and iridoid glycosides was more than 50% respectively, which meets the requirements of the V class of new Chinese drugs.2. Studies on the determination method and content determinationAfter getting ROS and iridoid glycosides, studied the determination method by UV spectrophotometry and established a method to determine the content of iridoid glycosides of Radix Rehmanniae finally, by which we determined the total content of iridoid glycosdies , at one time, we determined the content of ROS in effective compositions by phenol-sulfuric acid method. The iridoid glycosides content of medical material, water extract-alcohol precipitated and effective parts is 2.7%, 16.2% and 87.4% respectively, the ROS content in the effective compositions of polysaccharides is 76%.3. Studies on hypoglycemic effect of components in Radix RehmanniaeIn alloxan-induced diabetic mice, we compared the hypoglycemic effect of chemical compositions which including iridoid glycosides and its chief constituent catalpol, ROS and its chief constituent stachyose and the mixtrue of iridoid glycosides and ROS that comed from Radix Rehmanniae. We observed these indexes:body weight, random blood sugar, fasting plasma glucose, two-hour postprandial plasma glucose, high density lipoprotein cholesterol(HDL-C), total cholesterol(TC), triglycerides(TG), alanine aminotransferase(ALT), aspartate aminotransferase(AST) and weight of viscera, which indicated that those components could lower two-hour postprandial plasma glucose, decrease total cholesterol (TC) and triglycerides (TG) level , but the effect is difference from each other. It was observed that the drugs could increase high density lipoprotein cholesterol (HDL-C) content in alloxan-induced diabetic mice, suggesting that the drugs had some preventive effects on diabetic complications. Hyperglycemia-resulted injury of liver, spleen was also ameliorated. In conclusion, the mechanisms of hypoglycemic activity of objective drugs might relate to modulating carbohydrate and lipid metabolism, protecting liver and spleen against impairment by hyperglycemia.The preliminary research indicated that Radix Rehmanniae could be used as assistant drugs of chemical medicine, which would take effect in complication in diabetes.4. One of the important part of natural products is the extraction–separation- purification of monomers, which is could not be replaced by anything else. We obtained several monomers in simple way and identified the structure by TLC, determination of melting point, analysis of spectrum. These monomers could meet the requirement of the lab, and it is also available for industrial production.