Primary Study On The Extraction And Purification Of Folium Eucommiae And The Aucubin Accumulation In Seeds Of Eucommiae

Cortex Eucommiae is the dry root of Eucommia ulmoides Oliv.,and Folium Eucommiae is the dry leaf of Eucommia ulmoides Oliv.,Folium Eucommiae is a well-known traditional Chinese medicine which can be used to reinforce liver and kidney and strengthen muscles and bones,and which is collected by the Chinese Pharmacopoeia 2005 (1).On base of the total review of chemical components, pharmacology and clinical researches of Cortex Eucommiae and Folium Eucommiae,the active constituents of Chlorogenic acid of Folium Eucommiae,and so on,based on the single factor and the orthogonal test guided by the content of Chlorogenic acid,the optimum process of extraction and purification was determined and verified,the optimal extraction conditions on Chlorogenic acid in Folium Eucommiae by cellulase enzymes , and Chlorogenic acid residue extraction with high effciency by ultrasonic extraction method,best extraction process to Chlorogenic acid in Folium Eucommiae were chosen finally:cellulase enzymes quantity was2mg/g Folium Eucommiae,16 times of water with pH4.0,and kept the temperature of water 45℃with the time 1.5h,once by extraction each time,and made Chlorogenic acid residue by ultrasonic extraction method with 16 times of water with pH4.0,the ultrasonic extraction conditions were 40min (180W,40kHz),once by extraction each time. verified the best optimal extraction. And studied on the condition on the purification of NKA-Ⅱmacroporous resin and decided on the best conditions: concentration of evaporated extraction 0.09g pharmacognostics/mL,the pH3～4,the upper sample liquid:dry macroporous resin 24:1, 1.0 mL/min by dynamic absorption,used the water with 10 times of dry macroporous resin quantity,and used 70% ethanol of dry macroporous resin by eluting, with vacuum concentration at 40℃,and got dry extract of Chlorogenic acid in Folium Eucommiae.The parameters of purification process of dry extract of Chlorogenic acid in Folium Eucommiae by the Silica gel were determined and verified,and made the best purification process:the dry extract of Chlorogenic acid: dry Silica gel quantity 1:40,Ethyl acetate- Formic acid-Water(12:1:1)400 mL by eluting, and used the identification method of TLC,combined withthe same flowing Portions,with vacuum concentration at 40℃,and got the refinement of Chlorogenic acid.A RP-HPLC method is used for establishing a way to determine the content of Chlorogenic acid in Folium Eucommiae,using Chlorogenic acid standard as index,the range of the content of Chlorogenic acid in Folium Eucommiae is 2.46～2.51%,the average of it is 2.48%; the range of the content of the dry extract of Chlorogenic acid in Folium Eucommiae is 27.63～27.89%,the average content is 27.52%.the average content of the refinement of Chlorogenic acid is 93.13%,the average recovery is 78.42%.A RP-HPLC method is used for establishing a way to determine the content of Aucubin of the seed of Eucommiae,the results show that the Carpodermis of Eucommiae has no aucubin,and the content of aucubin in the seed of Eucommiae from different strains has a greater difference,but the same variety not the different habitat has almost no difference on the content,and there are greater differences on the contents of aucubin in the seed of Eucommiae from the same collecting date but different habitat.And it is highest of the content of aucubin n the seed of Eucommiae from Huazhong No.6,Shangqiu,Henan,which can reach 10.93%;the content of aucubin in the seed of Eucommiae,from the same habitatof Shangqiu,Henan,has obvious difference,the content of aucubin in the seed of Eucommiae,from Sep 25th,2007 to Oct 25th,can keep the higher level,considering the extraction and utilization of aucubin,it is suitable for the date from Sep 25th to Oct 25th to collect the sample of the seed of Eucommiae.To establish a new RP-HPLC method for simultaneous determination of Geniposidic acid,Chlorogenic acid and Geniposide of Folium Eucommiae from different harvest seasons but the same habitat.The results show that, there are some differences on the contents of the three active constituents of Folium Eucommiae , and the contents of the three active constituents of Folium Eucommiae from May 5th,2009 to Jun 20th can keep the higher level,it is highest of the contents of the three active constituents of Folium Eucommiae from Jun 5th,the average of the contents of the three active constituents of it can reach 4.58%,2.23%,0.13%,but collecting leaves will have a severe impact on the Eucommia ulmoides Oliv. trees.then with the leaves growing older,the contents of the three active constituents of Folium Eucommiae will become lower.Considering these influences,it is suitable for the date from Jul.to the middle date of Aug.to collect Folium Eucommiae.