Diferenciação e caracterização química de espécies de Phyllanthus utilizando técnicas hifenadas
Resumen
Several species of Phyllanthus genus are widely used in popular
medicine, even though very few phytochemical reports on species such as: P.
caroliniensis, P. stipulatus and P. tenellus are found in the literature. Furthermore, the
official reference quality control methods, especially those related with authenticity
assessment, are nonspecific for the species under analysis. Thereby, this work
reports on the online chemical characterization of the ethanolic extract of six
Phyllanthus species by liquid chromatography hyphenated to mass spectrometry
(LC-MS) and liquid chromatography hyphenated to nuclear magnetic spectroscopy
using automated solid phase extraction system, as interface (LC-SPE-NMR). The
chapter 2 discusses the classification of commercial Phyllanthus samples acquired
from different establishments in several Brazilian municipalities. The identification of
various chemical markers from authentic samples of each species allowed the
classification of commercial samples. The obtained results showed that they were
misused, in spite of their labels nominating them as P. niruri, the majority of the tested
samples were P. tenellus. None was classified as P. niruri. Chapter 3 discusses the
use of statistical models for the chromatographic method development and also for
the handling of MS data. Development of the method was carried out by
experimental design furnishing chromatographic runs with short analysis time and
high number of chromatographic bands. Statistical analysis using principal
component analysis (PCA) allowed the unmistakable differentiation of species and
pointed out to several of compounds responsible for this. Through the interpretation
of the MS/MS data, using the mass analyzers QqToF and IT-MS, 22 compounds had
their chemical structure elucidated, some of which have never been reported in the
literature. Chapter 4 describes the results obtained by LC-SPE-NMR for the
unequivocal structural elucidation of the secondary metabolites of the ethanolic
extract of P. tenellus. The use of automated SPE interface facilitated the isolation of
the compounds of interest, eliminating the need of using semi-preparative
chromatography. The interpretation of NMR spectra enabled the identification of 9
compounds confirming the structures inferred by LC-MS.