Acetylcholinesterase inhibitory-active components of Rhodiola rosea L. Hua Wang, Gaochao Zhou, Xiangdong Gao, Yidan Wang, Wenbing Yao * School of Life Science and Technology, China Pharmaceutical University, 24 Tongjia xiang, Nanjing 210009, PR China 授授授授 授授授 授授 : 授授授 授授授授 授授授 :、
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Acetylcholinesterase inhibitory-active components of Rhodiola rosea L.
Acetylcholinesterase inhibitory-active components of Rhodiola rosea L. Hua Wang, Gaochao Zhou, Xiangdong Gao, Yidan Wang, Wenbing Yao * School of Life Science and Technology, China Pharmaceutical University, 24 Tongjia xiang, Nanjing 210009, PR China 授課老師:詹于誼 教授 報告者:田仲源、莫博欽. Introduction. - PowerPoint PPT Presentation
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Acetylcholinesterase inhibitory-active components of
School of Life Science and Technology, China Pharmaceutical University, 24 Tongjia xiang, Nanjing 210009, PR China
授課老師:詹于誼 教授報告者:田仲源、莫博欽
IntroductionRhodiola rosea ("golden root" or "rose root") : 景天科
(Crassulaceae) 紅景天屬 (Rhodiola sp.) is a popular plant in traditional medical systems in Eastern Europe and Asia.
西藏稱“索羅瑪寶 ”
Rhodiola rosea: in a variety of diverse areas of physiological function, including neurotransmitter levels, central nervous system activity, and cardiovascular function.
紅景天的多樣性
生理功能的主要有效成份為 : 紅景天苷 (salidroside ;C14H20O7) and 酪醇 (p-tyrosol ; C6H10O2)
Application :decreasing depression, enhancing work performance, eliminating fatigue, and preventing high altitude sickness…
Acetylcholinesterase (AChE) inhibitors have been widely used in the treatment of Alzheimer’s disease (AD)
The medicinal effect of RR in the treatment of AD is probably mediated by the acetylcholinesterase inhibitory capability of this plant (Wu, Yao, Gao, & Wang, 2003). However, the specific compounds responsible for the anti-cholinesterase activity of RR remain unknown.
BPSD, which includes symptoms such as depression, anxiety, delusions, and agitation, is currently treated with psychotropic medications including antipsychotics, antidepressants, and hypnotics.
Cholinesterase inhibitors may be useful in the management of behavioral and psychological symptoms of dementia (BPSD).
Materials and methodsChemicals
Acetylthiocholine iodide (ATCI) as substrate
AChE as enzyme
5,5’-dithio-bis-(2-nitrobenzoic acid) (DTNB)
Plant materials
the dried rhizome of Rhodiola rosea L.
HPLC mobile phases :Solvent A (water) and solvent B (acetonitrile)
detector : 280 nm
UV–visible spectrophotometric analysis The sample solution was scanned from 200 to 700 nm
Molecular weight estimation ionization energy was 70 eV; ion source temperature was 230
℃. The scan range of m/z was 40–800.
NMR spectroscopy
Extraction and isolation proceduresplant material (200 g)
boiled ethanol (1 l) for 1 h
filtered through a filter paper
boiled ethanol (1 l) for 1 h
filtered through a filter paper
Filtered extracts evaporated to dryness (46.4 g)
Filtered extracts evaporated to dryness (46.4 g)
Extract (40 g) in 400 ml of deionized water
centrifugation (3000 rpm, 5 min)
Supernatant (360 ml) was precipited with gelatine
centrifuged ( 3000 rpm ,10 min )to remove tannins
The supernatant was reprecipitatedwith ethanol
centrifuged ( 3000 rpm , 10 min ) remove the remnant gelatine
The supernatant was evaporated to dryness
supernatant(dryness)
H2Ochloroform
EtOAcn-butanol
column chromatography
chloroform/ethyl acetate (1:1)
100 fractions (A)
Fractions A36–A59
Sephadex LH-20 column
methanol/water (1:1)
100 fractions (B)
FractionsB29–B39compound 1purifier HPLC
system
Bioassay for anti-cholinesterase activitythe colorimetric method of Ellman
ATCIthiocholin
e
Compare the method of Ellman and Rhee Ellman’ s method Rhee’ s method(false-positive
effect)
First step substrate (1 ml) sodium phosphate buffer (0.1 mM, pH 7.4) 1 ml 0.1 ml of plant extract incubated at 37 for 5 min.℃
First step substrates (1 ml) sodium phosphate buffer (1 ml) enzyme (0.1 ml) Incubated at 37 for 5 min. ℃
Second step 0.1 ml of enzyme was added and the mixture was incubated at 37 for a ℃ further 8 min.
Second step Plant extract (0.1 ml) was added and the mixture was incubated at 37 for ℃ another 8 min.
Third step The reaction was terminated by adding 3% SDS (1 ml), then 0.1 ml of 0.2% DTNB was added to produce the yellow anion of 5-thio-2-nitrobenzoic acid.