Screening For Acetylcholinesterase Inhibitors From Total Alkaloidal Extracts Of Medicinal Plants

Alzheimer's disease (AD) is a progressive degenerative neurologic disorder resulting in impaired memory and behavior. A promising treatment strategy for AD has been the use of acetylcholinesterase inhibitors (AChEIs). Several potent AChEIs such as huperzine and galanthamine were originally isolated from plants, indicating that medicinal plants are a good source for the search of potential cholinesterase inhibitors. In our study, to search for potential AChEIs, the total alkaloidal extracts of fifty Chinese herbal medicines were tested for their antiacetylcholinesterase activities by Ellman's colorimetric method in 96-well microplates in combination with modified thin layer chromatography (TLC) bioautography method. For the herbal extracts that were shown to exert a significant inhibition, the IC50 values for acetylcholinesterase inhibition were also determined. The results showed that alkaloidal extracts from Corydalis humosa, Bark of Phellodendron amurense, Uncaria rhynchophylla, Nelumbo nucifera, Zanthoxylum nitidum, Portulaca oleracea, Fritillaria pallidiflora, Fritillaria ussuriensis, Evodia rutaecarpa, Pinellia ternata, Ranunculus ternatus, Rhizome of Isatis indigotica, Cyperus rotundus and Stemona japonica exhibited remarkable antiacetylcholinesterase activities, their IC50 values were 1.93μg/mL,4.9μg/mL,10.8μg/mL,12.2μg/mL,17.4μg/mL,29.4μg/mL,31.2μg/mL,50.6μg/mL,50.7μg/mL,56.2μg/mL,59.5μg/mL,82.9μg/mL,88.8μg/mL and 102.6μg/mL. The rest of the alkaloidal extracts showed no or very weak AChE inhibitory activities.On the basis of screening results, We have further fractionated the alkaloidal extract of Zanthoxylum nitidum and obtained 10 compounds. Based on the modern spectroscopic methods (1HNMR,13CNMR,2D-NMR, EI-MS, etc.), these compounds were identified as: toddalolatone (1), skimmianine (2), dictamnine (3),γ- fagarine (4), magnolone (5), edulitine(6), haplopine (7), (-)-(S)-Edulinine (8),5,6,7-3-methyloxycoumarin (9), Zanthodioline(10). These compounds were tested for their antiacetylcholinesterase activities by Ellman's colorimetric method in 96-well microplates in combination with modified TLC bioautography method. The results showed that AChE inhibition was due to the presence of an quinoline alkaloid named skimmianine. The compound showed moderate acetylcholinesterase inhibitory activity with IC50 value of 33.3μmol/L. The rest of the compounds showed no or very weak AChE inhibitory activities.The results of this study indicate that the screening of traditional Chinese herbs for AChEIs may provide useful leads in the discovery of new drugs for AD. The screening strategies of acetylcholinesterase inhibitors also provide a useful reference for the screening of other types of natural enzyme Inhibitors.