The Effects Of Nutrition Elements On The CPT Accumulation In Camptotheca Acuminata Seedling And Extraction And Purification Technology Of CPT

Camptothecin (CPT),an alkaloid extracted from Camptotheca acuminata has high anti-tumor activity in spectrum range. CPT has become one of the most promising anticancer drug in the 21st century. To obtain higher output of CPT, one-year old seedlings of Camptotheca acuminata as the object, we investigated the effects of nutrition elements on the CPT content and dynamic changes.The new technology of extraction and purification of CPT from Camptotheca acuminata seedlings had been studied in this dissertation.The main results of our research as follows:RP-HPLC method of determining CPT content.1.The RP-HPLC method of determining CPT content was built up. phenomenex Luna C18(2)(4.6 mm×250 mm,5μm) column was used for this study and acetonitrile-water(30:70,v/v) was chosen as the mobile phase at the flow-rate of 1.0mL/min. The detection wavelength was 254nm. Column temperature was maintained at 40℃and The sample injection volume used was 20μl. The retention time of CPT was 10.4 min under this method, and the CPT was well separated and the apex was perfect.2.Sample solution was prepared as follows.1.0g powders of Camptotheca acuminata was weight accurately and ultrasonic extracted by 50 ml menthol at 50℃for one hour. The extracts was filtrated by filter membrane of 0.45μm.This method could extract CPT from Camptotheca acuminata sample sufficiently. Determination result was accurate.We investigated the alteration of dry weight and accumulation of CPT in Camptotheca acuminata seedlings.The following are the main results of our research.1.Camptotheca acuminata seedlings grow rapidly from July to October. CPT content of leaves exhibited a single apex curve from May to November, it reached the highest point at August. CPT content of roots and trunks exhibited a sinusoidal curve from May to November,it reached the maximum value in August and the minimal value at October.2.The CPT content in root, trunk and the whoole seedling of Camptotheca acuminata from gradually increased from May to November. The CPT content of leaves exhibited a single apex curve from May to November, it reached the highest amount at October.we investigated the effects of exogenous nutritient on the CPT accumulation in Camptotheca acuminata seedling.The main results of our research as follows.1.The effects of nitrogen on the growth of seedling and CPT accumulation in Camptotheca acuminata seedling.1) Nutrition was conducive to growth of Camptotheca acuminata from seedling. But this effect could be unremarkable when nutrition was greater than 0.95g per seedling.Under different level of nutrition, CPT content exhibited a single apex curve in every organ,it reached the highest level when the nutrition content was 1.90g per seedling.2) The CPT amount in whoole seedling of Camptotheca acuminata increased gradually with the increasing of nitrogen. But when the nutrition content was greater than 1.90g per seedling, the increasing rate declined.2.The effects of phosphorus on the growth of seedling and CPT accumulation.1) Phosphorus was very important to growth of Camptotheca acuminata from seedling. Under different level of phosphorus, CPT content exhibited a single apex curve in every organ,it reached the highest level when the amount of phosphorus was 0.14g per seedling.2) The CPT amount in every organ and whoole seedling gradually increased when the amount of phosphorus increased from 0 to 0.56g per seedling.3.The effects of potassium on the growth of seedling and CPT accumulation.1) Potassium at a fewer level was positive to the growth of Camptotheca acuminata seedling. But the growth of seedling and the content of CPT in in every organ would be restrained when the amount of potassium was greater than 1.06g per seedling.2) The CPT amount in every organ and whoole seedling reached the highest level when the content of potassium was 1.06g per seedling.We also investigated the extraction and purification technology of CPT from Camptotheca acuminata seedling.The main results of our research as follows.1. The best extraction method using NaOH as the solution was as follows:the concentration of NaOH aq. is 1%.1 g raw material was extracted by 10ml NaOH solvent at room temperature for 2 hours. This treatment was repeated for 3 times and all the extracts were collected.2.The purification technology of CPT was as follows:The extracts solvent was neutralized to PH 7.0 by HC1 and then filtrated after laying for 10 hours. The filtrate was adsorbed by AB-8 resin.The CPT in resin was eluted by chloroform. Chloroform solvent was carried out through silica gel column analysis.Then the detergent of methanol-chloroform (2:98,v/v) was used to elute.TLC method was used for identification of CPT.The CPT was obtained by collecting eligible products, which was measured by TLC, and concentrated at 70℃. The products after concentration were recrystallizated by methanol-chloroform (1:l,v/v).The content of CPT was about 99% by HPLC detection.