Inhibition of Reactive Oxygen Species Production in Rat Peritoneal Macrophages by Ethanolic Extract Of MOR US ALBA

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Rapeeporn Phromgate
Bunkerd Kongyingyoes
Patchareewan Pannangpetch
Veerapol Kukongviriyapan
Yupa Kukongviriyapan


Free radicals and reactive oxygen species are known to implicate in the

pathogenesis of various human diseases such as atherosclerosis, diabetes and

ischaemia-reperfusion injury. Numerous plant constituents have been shown to have

free radical scavenging or antioxidant activity. Mulberry (Marus alba) leaves have

been used to feed the silk worms and now they are also widely prepared as a beverage

for healthy, mulberry tea. It has been reported that the extract of mulberry leaves had

many pharmacological effects including antioxidant activity. However, most of the

information of antioxidant activity of mulberry leaves was evaluated in cell free

system. In order to obtain an additional important information for implementing the

mulberry leaves in therapeutic intervention, therefore, we examined the intracellular

antioxidative activity of mulberry leaves. An ethanolic extract of dried leaves of

M.alba var.Nakhonrajchasima 60 was used in all investigations. The effect of the extract

on superoxide produced within the rat peritoneal macrophages was tested by using

H2DCFDA (2',7'-dichlorodihydro fluorescein diacetate) probe. The production of

H2O2 was stimulated by phorbol-12-myristate-13- acetate. The fluorescence intensity

is proportional to the amount of H202 produced by cells. The extract of M. alba of

100 μg/ml significantly inhibited the production of peroxide (n=5, P<0.05). The free

radical scavenging and the reductive activities of the extract of M. alba were also

investigated. The free radical scavenging activity was determined by a method based

on the reduction of coloured stable free radical DPPH (1, 1-diphenyl-2-picrylhydrazyl).

The M. alba extract (1-300 μg/ml) scavenged the DPPH in the dose-dependent

manner with the IC50 of 20.1 μg/ml. The reductive activity was examined

by using the ferric reducing/antioxidant power (FRAP) assay. The extract was able to

reduce ferric complex to ferrous form in a dose-dependent mode. The extract at 10

μg/ml had the ferric reducing activity equivalent to vitamin C 1.2 μg/ml. It can be

concluded that the ethanolic extract of M. alba var.Nakhonrajchasima 60 has the free

radical scavenging and the reducing activities in cell free system together with the

intracellular antioxidant activity.


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2002 Annual Meeting Abstracts/Lectures