RESUMEN

The determination of ginkgolic acids in Ginkgo products is one of the principal components of quality control. However, a number of ginkgolic acids with different side chains may be present and this makes their analysis by conventional chromatographic methods more complex. In this study, 1H-NMR spectrometry was applied to the analysis of the total content of ginkgolic acids in leaves of Ginkgo biloba and in six types of commercial Ginkgo products in the absence of chromatographic purification. For this analysis, protons H-3, H-4, and H-5, which are well separated in the range 8 (ppm) 6.5-7.5 in the 1H-NMR spectrum, were utilised. For further confirmation, the correlations of H-3, H-4 and H-5 were examined by 1H-1H COSY spectra in all extracts. The quantity of the compounds was calculated from the relative ratio of the integral of each peak to the integral of the peaks of a known amount (100 microg) of anthracene used as an internal standard. The quantitative results obtained by 1H-NMR analysis were compared with those obtained by GC, which showed that the 1H-NMR method allows a simple quantification of total ginkgolic acids in Ginkgo extracts without any pre-purification steps.

Asunto(s)

Ginkgo biloba/química , Salicilatos/análisis , Cromatografía de Gases , Resonancia Magnética Nuclear Biomolecular/métodos , Hojas de la Planta/química , Plantas Medicinales/química

RESUMEN

Geraniol 10-hydroxylase (G10H) is a cytochrome P450 monooxygenase involved in the biosynthesis of iridoid monoterpenoids and several classes of monoterpenoid alkaloids found in a diverse range of plant species. Catharanthus roseus (Madagascar periwinkle) contains monoterpenoid indole alkaloids, several of which are pharmaceutically important. Vinblastine and vincristine, for example, find widespread use as anti-cancer drugs. G10H is thought to play a key regulatory role in terpenoid indole alkaloid biosynthesis. We purified G10H from C. roseus cells. Using degenerate PCR primers based on amino acid sequence information we cloned the corresponding cDNA. The encoded CYP76B6 protein has G10H activity when expressed in C. roseus and yeast cells. The stress hormone methyljasmonate strongly induced G10h gene expression coordinately with other terpenoid indole alkaloid biosynthesis genes in a C. roseus cell culture.

Asunto(s)

Alcaloides/biosíntesis , Sistema Enzimático del Citocromo P-450/genética , Sistema Enzimático del Citocromo P-450/metabolismo , Magnoliopsida/enzimología , Magnoliopsida/genética , Oxigenasas de Función Mixta/genética , Oxigenasas de Función Mixta/metabolismo , Monoterpenos Acíclicos , Secuencia de Aminoácidos , Secuencia de Bases , Línea Celular , Clonación Molecular , Ciclopentanos/farmacología , Sistema Enzimático del Citocromo P-450/química , Sistema Enzimático del Citocromo P-450/aislamiento & purificación , Cromatografía de Gases y Espectrometría de Masas , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Hidroxilación , Oxigenasas de Función Mixta/química , Oxigenasas de Función Mixta/aislamiento & purificación , Datos de Secuencia Molecular , Oxilipinas , Plantas Modificadas Genéticamente , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN de Planta/genética , ARN de Planta/metabolismo , Especificidad por Sustrato , Terpenos/metabolismo , Levaduras