Estudo dos alcaloides de amaryllidaceae e sua correlação com a atividade anti-acetilcolinesterase

Abstract

The Amaryllidaceae family is characterized by the high occurrence of isoquinoline alkaloids so-called Amarylidaceae alkaloids (AA), which have chemotaxonomic and biological importance. AA stands out for the inhibitory action on the enzyme acetylcholinesterase (AchE), which is an important therapeutic target for the treatment of Alzheimer's disease. Galantamine (GAL), a clinical drug used for this purpose, is the main representative of this chemical class, being obtained mainly from botanical sources that occur or are cultivated in temperate regions. Such alkaloid was previously reported in Hippeastrum elegans, an Amarylidaceae species native to the Caatinga, where it is known as açucena and used for ornamental purposes. The present work aimed to investigate potential tropical sources of galantamine and other bioactive AA, by evaluating the chemical compositions of bulbs from the Embrapa Collection of Amaryllidaceae (ECA) and cultivated H. elegans, then correlating them with their antiAchE activity. Our study consisted of the following experiments: (I) development and validation of an ultra-efficient liquid chromatography method coupled to mass spectrometry (UPLC-ESI-QTOF and UPLC-QDA) and Hydrogen Nuclear Magnetic Resonance (1H NMR) analysis for identification AA and quantification of the alkaloids GAL, pseudochlorine (PSE), narciclasin (NAR) and sanguinine (SAN) in 67 extracts from ECA (Habranthus, Hippeastrum, Hymenocallis, Zephyranthes and Griffinia species) as well as six extracts from H. elegans bulbs harvested in six harvest times (150 , 210, 270, 330, 390 and 450 days of cultivation); (II) evaluation in vitro and ex vivo AchE inhibitory activity assays for the aforementioned extracts. In addition, the UPLC-ESI-QTOF and NMR data were analyzed by chemometrics analysis based on partial least squares regression (PLS) to identify biomarkers. Thus, the alkaloids were extracted from dry and ground bulbs (100 mg) using liquid-liquid microextraction (hexane/CH3OH-H2O) followed by solid phase extraction in cation exchange cartridges. The UPLC-QDA quantification method showed good linearity (R ≥ 0.9968), selectivity, sensitivity (LOD: 5-100 ng / mL and LOQ: 20-350 ng / mL), repeatability (CV = 1.3- 8.4% for intraday and interday precision) and acceptable recovery limits (87.5-96.2%), except for narciclasin. In the ECA evaluation, the Hippeastrum extracts showed the highest levels of SAN, PSE, NAR, while those of Griffinia showed the highest levels of GAL. The greatest anti-AChE activities were observed for the Habranthus, Hippeastrum and Griffinia extracts. Regarding the investigation of harvest time, H. elegans was found to be rich in PSE and NAR, reaching higher levels at 270 and 390 days of cultivation, respectively. However, the greatest anti-AChE activities were observed for 450-day bulbs. PLS analyses indicated PSE as a discriminating compound for this extract, therefore it was pointed out as responsible for its greater anti-AChE activity along with the known AChE inhibitors GAL and SAN. Accordingly, our study indicates that Griffinia species and H. elegans are promising sources of bioactive AA. However, agronomic studies for genetic breeding and production system development are necessary to make them commercially competitive materials with their congeners grown in temperate countries.