| Our job aimed chiefly at developing and utilizing a new resource of natural medicines, Isodon excisa(Maxin.)Hara, and elucidating its main component--diterpenoids, and contents. The author quantified Kamebakaurin and Kamebanin depended on HPLC in the I. excia and determinated the optimum harvest days and the best ultrasonic extraction process. Meanwhile, we obtained Kamebakaurin and Kamebanin, which are two purified compounds, from the herbs, using new technology for extracting. Besides, we measured the contents of flavonoids and polysaccharides in different collecting time and parts of I. excia which might supply the reference data for the future researching works.RP-HPLC was used to determinate the content of kamebakaurin and kamebanin in different harvest period of I. excia. Test result showed:1. The HPLC separation was performed on a C18 analytical column gradient eluted with a mixture consisting of acetonitrile and water (30: 70) at a flow rate of 1.0 mL/min with UV detector at 230nm. The temperature of column was 25℃. The method was accurate, sensitive and reproducible.2. It indicated that the contents of Kamebakaurin in I. excia in later July is the highest, up to 0.5246％, the Kamebanin in I. excia in early and mid-August is the highest, up to 0.1349％, and the contents of them in later July is the highest, and then when the I. excia comes into the flower season and underage fruit period, the contents of them decrease slowly. So the optimum collecting period is in the later July when branches and leaves are luxuriance before blooming.An optimum method for getting diterpenoids of I. excia was found, which could be described as follows: Kamebakaurin and Kamebanin were extracted by EtOH(70％) with treatment of ultrasonic, radiating 3 times, 30 minutes each time. This method has the advantages of high efficiency of soaking and economization of solvent, environmental protection.Diterpenoids in I. excia were extracted and separated by using new method. The dried stems and leaves of I. excia were extracted with 70％EtOH in ultrasonic at 50℃, 100Hz, radiating 3 times, 30 minutes each time. The combined extracts were concentrated in vacuo and got the residue, which degreased with ligarine. Then we pulled twice as much water into it. Filtered and then put the liquid into the refrigerator for one week. Took out the liquid then made the filtering went through the D-101 and used 10％, 40％, 60％, 80％, 90％EtOH to smash the column separately, one by one collected. The third part of the fluid was concentrated and evaporated in vacuo to give yellow powder 4g. The second part was concentrated and evaporated in vacuo and then supernatant fluid extracted with ethyl acetate. We evaporated ethyl acetate, got yellow white powder 10g then made the powder go through the JTY-1 reversed-phase colophony and used 15％, 20％, 25％, 30％, 35％EtOH to smash the column separately. The pan of 20％was concentrated and evaporated in vacuo to give crystallization. After purified further, we got compounds 1. The pans of 25％and 30％combined and the residue was separated on a silica gel column with CHCl3: Me2CO: EtAc (3: 1: 1) as eluent. Followed by prep. TLC, Ft. Nos. 8-9 and Fr. Nos. 12-15 was combined and recrystallized from the MeOH gave the compounds 1 and 2. Two compounds isolated in the experiments identified by chemical reaction, phytochemical properties and spectral analysis(MS), structures of them was elucidated as diterpenoids. They are Kamebakaurin and Kamebanin, which structure belongs to the ent-kaurane, might had the action of anti-tumor etc.At the same time, the tube dilution test was used for determing in the antibacterial activity of ethanolic extract of I. excia. The experiment told us: the ethanolic extract of I. excia has preferable antibacterial action in vitro.We quantified the contents of total flavonoids and polysaccharides in different collecting period and parts of I. excia by colorimetry technique. The experimental results indicate that: the contents of total flavonoids and polysaccharides in different parts of I. excia were different significantly. |
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