| Summary: | 博士 === 國立臺灣大學 === 藥學研究所 === 99 === Part 1 Characterization of acetylcholinesterase inhibitory constituents from Annona glabra assisted by HPLC micro-fractionation
Acetylcholinesterase (AChE) inhibitors are used as an effective approach to treat Alzheimer’s disease (AD). Up to now, only a few AChE inhibitors have been approved for such purpose due to the side effects. Thus, the search for safer and effective AChE inhibitors is still a demand for the treatment of AD. Since natural products are versatile and often serve as drug lead for further development, a simple assay system was applied to explore the potential AChE inhibitors from higher plants. The EtOH extract of the stem of Annona glabra L. (Annonaceae) was found to be active against AChE. Thus the characterization of the anti-AChE constituents from this plant extract was the aim of this study.
The active fraction was analyzed by combining HPLC micro-fractionation with a bioassay. The analytical-scale sample was fractionated by HPLC-DAD into 96-well microplates, which, after condentration using centrifugal vacuum evaporator, were assayed against AChE. The active subfractions were separated over semi-preparative HPLC to give twenty compounds. Four of these, (7S,14S)-10-O-demethylxylopinine N-metho salt (3), S-(-)-7,8-didehydro-10-O-demethylxylopininium salt (10), S-(-)-7,8-didehydrocorydalminium salt (11), and 5-methoxymarcanine A (17), were assigned as new natural products. In addition, compounds 10 and 11 represent the first natural occurrence of 7,8-didehydroprotoberberines. Compound 3, pseudocolumbamine (12), palmatine (15), and pseudopalmatine (16) showed anti-AChE IC50 values of 8.4, 5.0, 0.4 and 1.8 μM, respectively.
Part 2 Flavonoid composition in the leaves of 14 Neolitsea and Litsea plants
This study was aimed to study the chemodiversity of flavonoids in the Formosan Litsea and Neolitsea plants. Applications of LC-SPE-NMR and LC/MS hyphenated techniques in analyzing polar constituents from the leaves of L. acuminata, L. akoensis var. Chitouchiaoensis, L. krukovii, N. acuminatissima, and N. konishi led to the identification of 13 known flavonoids and one new flavonol dioside, quercetin 3-O-(2-O-β-D-apiofuranosyl)-α-L-rhamnopyranoside. The quantity and variety of flavonoid composition in the leaves of 14 Litsea and Neolitsea plants were examined to enable more effective utilization of such bioactive ingredients. Of these, N. acuminatissima was found to contain the most quantity of flavonoids (ca. 0.24%, w/w, leaves).
Part 3 ALZ-1 attenuates scopolamine-induced amnesia in rats
The deficiency in cholinergic neurotransmitter is an important feature of Alzheimer''s disease (AD). Acetylcholinesterase (AChE) inhibitor is one of the rational therapeutic targets to improve AD. From in vitro study, IC50 of ALZ-1 for AChE inhibitory activity is 1.56 uM. The effects of ALZ-1 on the scopolamine-induced memory deficits in rats were examined in Morris water maze and active shuttle avoidance. Scopolamine (1 mg/kg, i.p.), the antagonist of cholinergic receptors, caused significant impairments on a wide range of learning and memory tasks. We investigated the effects of ALZ-1 on memory disturbances induced by scopolamine, since ALZ-1 (1 and 10 mg/kg, p.o.) had greater efficacy than scopolamine group on the improvement of scopolamine-induced learning and memory errors, respectively. These in vivo findings by two behavioural paradigms suggest ALZ-1, as a therapeutic potential of orally active AChE inhibitor, for improving learning and memory in AD.
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