Camptothecin is a strong anti-tumor compound isolated from Camptotheca acuminata. One of the most important way for the production of Camptothecin is by cell cultures of Camptotheca acuminata. The effect of Cu2+ on camptothecin accumulation in Camptotheca acuminata cell line was described in this paper. The results showed that the optimum CuCl2 concentration in B5 medium was 0.008 mg/mL, which increased camptothecin production for 30 times compare to the control while has no inhibitive effects on cell growth, at the same time, the peroxidase activity was increased and the anthocyanidin accumulation was inhibited. The promotive effects of Cu2+ on camptothecin accumulation in light was higher than that in dark.
Anthocyanins ; biosynthesis ; Antineoplastic Agents, Phytogenic ; biosynthesis ; Camptotheca ; growth & development ; metabolism ; Camptothecin ; biosynthesis ; Copper ; pharmacology ; Light

OBJECTIVETo investigate the effects of nitrogen concentration on the camptothecin (CPT) content in Camptotheca acuminata seedlings:

METHODThe seedlings of C. acuminata with 6 pair of leaves were subjected to five nitrogen concentrations treatments by sand culture in a greenhouse. The CPT content in the seedlings was determined by HPLC on the 20th, 35th, 50th, 65th and 80th day respectively.

RESULTThe CPT content in the young leaves of C. acuminata seedlings supplied with different nitrogen concentration was significantly higher than that in other organs (P < 0.01), and it showed a single peak curve with the time course, the highest CPT content was observed on the 50th day after treatment. The CPT content in the young leaves obviously declined with increasing nitrogen concentration, and it reached the highest (6.72%) when nitrogen concentration was 4 mmol x L(-1), equal to 1.1 times that of 16 mmol x L(-1) nitrogen.

CONCLUSIONThe results demonstrate that proper deficient nitrogen stress can significantly enhance CPT accumulation in young leaves of C. acuminata seedlings.

Camptotheca ; drug effects ; metabolism ; Camptothecin ; metabolism ; Chromatography, High Pressure Liquid ; Nitrogen ; pharmacology ; Seedlings ; drug effects ; metabolism

Three-dimensional (3D) fluorescence spectra and thin layer fluorescence chromatogram of common Camptotheca fruit (CCF) crude drug, camptothecin (CPT) and 10-hydroxycamptothecin (HCPT) have been studied, and a novel fluorimetric method for determination of CPT in CCF crude drug has been established. In 3D fluorescence spectra, CPT presented 3 fluorescence peaks with excitation wavelengths lambdaex of 215, 255 and 365 nm, separately, and all peaks with emission wavelength lambdaem of 430 nm. HCPT presented 4 fluorescence peaks with lambdaex of 220, 265, 325 and 375 nm, separately, and all peaks with lambdaem of 555 nm. The fluorescence of CPT is much stronger than that of HCPT. Comparison of 3D fluorescence spectra and analysis of thin layer fluorescence chromatogram revealed that the main fluorescent component of CCF is CPT. HCPT and other components basically do not interfere with the fluorescence of CPT. Under the condition of pH 3.0-6.7, CCF aqueous solution can produce strong and steady fluorescence. Using methanol as solvent, the extracting solution of CCF was prepared, and diluted properly with water, then measured fluorescence intensity at lambdaex/lambdaem = 365/430 nm to determine the content of CPT. A linear calibration curve covered the concentration range 0.002 35-0.235 microg x mL(-1). The regression equation was IF = 9 + 30,844 c, with correlation coefficient r = 0.999 (n=9). The method has been applied to the analysis of CPT in a CCF sample, with a result of 0.127% and a spiked recovery rate of 102%. The reliability of the method has been verified by a thin layer chromatography-fluorescence scanning method. Experimental results demonstrated that this method can be used for quality evaluation of CCF crude drug.
Antineoplastic Agents, Phytogenic ; analysis ; Camptotheca ; chemistry ; Camptothecin ; analogs & derivatives ; analysis ; Chromatography, Thin Layer ; Fruit ; chemistry ; Plants, Medicinal ; chemistry ; Quality Control ; Reproducibility of Results ; Spectrometry, Fluorescence

To investigate the technological parameters of the isolation and purification of 10-hydroxycamptothecin and vincoside-lactam from Camptotheca acuminata seed by polyamide. The static arid dynamic adsorption characteristics of 10-hydroxycamptothecin and vincoside-lactam on polyamide were studied, and the contents were determined by HPLC. The optimum parameters for adsorption were as follows: the contents of 10-hydroxycamptothecin and vincoside-lactam in the extracts were 0.189 g x L(-1) and 0.334 g x L(-1), respectively, pH 6, flow rate was 1.0 mL x min(-1), processing volume was 3 BV; for desorption: ethanol-water (60:40), flow rate was 1.0 mL x min(-1), 5 BV as an eluent. After treated with polyamide, the contents of 10-hydroxycamptothecin and vincoside-lactam were 17.52% and 32.87%, respectively, the recovery yields were 66.05% and 75.86%, respectively. Results showed that polyamide revealed a good ability to separate 10-hydroxycamptothecin and vincoside-lactam. Therefore, we concluded that results in this study may provide scientific references for the large-scale production of 10-hydroxycamptothecin and vincoside-lactam extracted from C. acuminata seed.
Camptotheca ; chemistry ; Camptothecin ; analogs & derivatives ; chemistry ; isolation & purification ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Hydrogen-Ion Concentration ; Nylons ; chemistry ; Seeds ; chemistry

OBJECTIVETo study the callus induction from leaf, stem segments and stem segments with axillary's bud and the subculture conditions of callus in Camptotheca acuminata.

METHODThe explants were inoculated into the media of MS with different concentrations of 6-BA, NAA and 2, 4-D by orthogonal experiment.

RESULT AND CONCLUSIONThe optimal medium for callus induction was MS with 6-BA 2 mg x L(-1), NAA 2 mg x L(-1) and 2,4-D 2 mg x L(-1). It had better effects in leaf than in other explants. The induction ratio in leaf reached above 80%. The callus subcultured on the medium of MS with 6-BA 1 mg x L(-1) and 2, 4-D 1 mg x L(-1) grew vigorous and more quickly than that in other media. It was loose, friable in consistency and suitable for cell culture.

Camptotheca ; drug effects ; growth & development ; Culture Media ; pharmacology ; Plant Growth Regulators ; pharmacology ; Plant Leaves ; drug effects ; growth & development ; Plant Stems ; drug effects ; growth & development ; Plants, Medicinal ; drug effects ; growth & development ; Tissue Culture Techniques ; methods

OBJECTIVETo investigate the effects of nitrogen forms on the camptothecin (CPT) content, tryptophan synthase (TSB) and tryptophan decarboxylase (TDC) activities in Camptotheca acuminata seedlings.

METHODThe seedlings of C. acuminata with 6 pairs of leaves were subjected to 5 different NH4(+) -N/NO3(-) -N ratio (0 : 100, 75 : 25, 50 : 50, 25 : 75, 100 : 0) treatments by sand culture in a greenhouse. The CPT content, TSB activity in the young leaves and TDC in the stem barks of the seedlings were determined by HPLC on the 15th, 30th, 45th, 60th and 75th day, respectively.

RESULTThe obvious relationship between CPT content and nitrogen forms was observed. When NH4(+) - N /NO3(-) -N ratio was 25 : 75, CPT accumulation in young leaves displayed the best advantages (the highest value is 5.69 per thousand) and increased in the early 30 days of treatment and then declined. There was no obvious relationship between TSB activity in the young leaves and nitrogen forms. TDC activity in the stem bark was the highest when NH4(+) -N /NO3(-) -N ratio was 25 : 75, and the change of TDC activity paralleled to CPT content in the young leaves.

CONCLUSIONA short-term treatment that NH4(+) -N /NO3(-) -N ratio was 25:75 may gain high CPT content in the young leaves through enhancing the TDC activity in the stem bark of C. acuminata seedlings.

Aromatic-L-Amino-Acid Decarboxylases ; metabolism ; Camptotheca ; drug effects ; enzymology ; metabolism ; Camptothecin ; metabolism ; Drugs, Chinese Herbal ; metabolism ; Nitrogen ; chemistry ; pharmacology ; Plant Leaves ; drug effects ; enzymology ; metabolism ; Seedlings ; drug effects ; enzymology ; metabolism ; Tryptophan Synthase ; metabolism