RESUMEN
INTRODUCTION: Rhazya stricta Decne. (Apocynaceae) is a medicinal plant rich in terpenoid indole alkaloids (TIAs), some of which possess important pharmacological properties. The study material including transgenic hairy root cultures have been developed and their potential for alkaloid production are being investigated. OBJECTIVE: In this study, a comprehensive GC-MS method for qualitative and quantitative analysis of alkaloids from Rhazya hairy roots was developed. METHODS: The composition of alkaloids was determined by using GC-MS. In quantification, the ratio between alkaloid and internal standard was based on extracted ion from total ion current (TIC) analyses. RESULTS: The developed method was validated. An acceptable precision with RSD ≤ 8% over a linear range of 1 to 100 µg/mL was achieved. The accuracy of the method was within 94-107%. Analysis of hairy root extracts indicated the occurrence of a total of 20 TIAs. Six of them, pleiocarpamine, fluorocarpamine, vincamine, ajmalicine and two yohimbine isomers are reported here for the first time in Rhazya. Trimethylsilyl (TMS) derivatisation of the extracts resulted in the separation of two isomers for yohimbine and also for vallesiachotamine. Clearly improved chromatographic profiles of TMS-derivatives were observed for vincanine and for minor compounds vincamine and rhazine. CONCLUSION: The results show that the present GC-MS method is reliable and well applicable for studying the variation of indole alkaloids in Rhazya samples.
Asunto(s)
Apocynaceae/química , Cromatografía de Gases y Espectrometría de Masas/métodos , Raíces de Plantas/química , Alcaloides de Triptamina Secologanina/análisis , Alcaloides/análisis , Alcaloides/química , Alcaloides/aislamiento & purificación , Apocynaceae/genética , Isomerismo , Estructura Molecular , Raíces de Plantas/genética , Plantas Modificadas Genéticamente , Reproducibilidad de los Resultados , Alcaloides de Triptamina Secologanina/química , Alcaloides de Triptamina Secologanina/aislamiento & purificación , Técnicas de Cultivo de Tejidos/métodos , Compuestos de Trimetilsililo/análisis , Compuestos de Trimetilsililo/química , Compuestos de Trimetilsililo/aislamiento & purificación , Vincamina/análisis , Vincamina/química , Vincamina/aislamiento & purificación , Yohimbina/análisis , Yohimbina/química , Yohimbina/aislamiento & purificaciónRESUMEN
OBJECTIVE: To study tissue culture of Vinca minor and determine the content of vincamine. METHOD: Leaf blades, stalks, root segment of V. minor were used as explants to study the effect of 2, 4-D,6-BA,NAA on its callus induction and vincamine contents in the orthogonal design experiment. In the peak period of callus formation, vincamine content in callus of V. minor and sterile plants was determined by HPLC. The experimental data was statistically analyzed. RESULT: The content of 6-BA and NAA had no significant effect on its callus induction. But the content of 2, 4-D had significant effect on its callus induction. Within 20,40,60 d, the content of vincamine in sterile plant was (0.015 +/- 0.003)%, (0.097 +/- 0.001)% , (0.113 +/- 0.06)%, respectively. In the peak period of callus formation, vincamine content in callus of leaf blades, stalks, root segment was (0.024 +/- 0.0025)%, (0.016 +/- 0.0015)%, (0.010 +/- 0.0015)%, respectively. To 30 days of subculture, vincamine content in callus of leaf blades, stalks, root segment was (0.041 +/- 0.002)%, (0.019 +/- 0001)%, (0.016 +/- 0.002)%, respectively. CONCLUSION: The optimal hormone combination for callus initiation was MS +2, 4-D 1.0 mg x L(-1) +6-BA 0.5 mg x L(-1) + NAA 0.5 mg x L(-1). In different growth periods, vincamine content in sterile plants is significantly different. From different explants in callus vincamine content is different, in which leaves callus is significantly higher than that of stems, roots produced callus organization.