| This study is a part of the project -Studies on immunoenhancement and mechanism of Chinese herbal medicinal ingredients and a new-type immunopotentor. The details are summarized as follows.â… Preparation of Chinese herbal medicinal ingredient (CHMI) Ten CHMIs, Astragalus polysaccharide (APS), Chinese angelica polysaccharide (CAPS), Isatis root polysaccharide (IRPS), Epimedium polysaccharide (EPS), Propolis polysaccharide (PPS), Astragalus flavone (AF), Epimedium flavone (EF), Propolis flavone (PF), Astragalosides (AS) and Ginsenosides (GS) , were choosed for the experiment. The five polysaccharides were extracted by Water-extracting-ethanol-precipitation method, AF and EF by Polyacrylamide chromatography, PF by Acid-alkali-precipitation, and AS by Colophony absorbent chromatography. The contents of five polysaccharides were assayed by Vitriol-anthracene ketone, three flavones by Rutin test, and AS by TLC spectrophotometry. The extractive rates (g Kg-1) of the nine ingredients were respectively 13.6, 14.5, 10.0, 4.1, 36.0,2.1,5.7, 11.0 and 1.0. The contents (%) were 82.48, 77.28, 82.94, 66.38, 51.41, 1.39, 38.37, 46.92 and 72.88 respectively. GS with the net content of 75% was purchased from Yawei Pharmaceutical Co., Jilin, China.â…¡ The determination of CHMI safety concentration for CEF Ten CHMIs were diluted with MEM medium into series concentrations and used for cultivating chick embryo fibroblasts (CEF) in two ways, at the beginning of culture and after CEF monolayer formation. Their maximal safty concentrations to CEF were judged according to OD value and morphological variety of the cells. The results showed that the maximal safety concentrations ( g-mL-1) of APS, CAPS, IRPS, AS, GS, PPS, EPS, EF, PF and AF were 1600, 1200, 600, 150, 120, 40, 40, 20, 20 and 2.5 respectively.â…¢ Effects of CHMI on CEF proliferating Ten CHMIs within safety concentrations were added into culture plates in two ways described previously. The cell OD values were assayed by MTT method respectively after 24, 36, 48, 60 and 72 hours cultivation for the groups adding ingredients initially and after 12, 24, 36, 48 and 60 hours for other groups to judge the effects of CHMI on cell proliferation. The results showed that all often ingredients at suitable concentrations from the former groups could stimulate cellproliferation. In other groups, some ingredients (AS, APS, IRPS and PF) promoted while EPS inhibited the cell proliferation and another five stimulated or inhibited cell proliferation, depending on concentrations and time points. It seems that there were dosage- and time-effect relationships.â…£ Effects of CHMI on cellular infectivity of virus Ten CHMIs at five concentrations and Newcastle disease virus (NDV) were respectively added into CEF in three different ways (synchronously, before and after inoculation of virus). After inoculating NDV for 72 hours, cell proliferation were tested by Neutral red assay to evaluate the effects often ingredients on infection of the virus to CEF. All of the ten, almost at every concentration, could inhibit infectivity of the virus to cells, in which AS was the stongest and then PPS>EF>IRPS>APS & AF>CAPS>EPS & PF & GS, depending on their concentrations. It suggested that they possibly behaved by the mechanism on inhibiting virus directly or promoting cell antivirus or both.â…¤ Effects of CHMI on antibody titer of ND vaccine In experiment 1, chicks were injected respectively with CAPS, APS, IRPS, EPS, PPS, EF, PF, AS and GS at two dosages once a day for three successive days after inoculation of Newcastle Disease virus â…£ strain (NDV-â…£) vaccine. Serum was collected on Day 7, 14, 21, 28 and 35 after inoculation. In experiment 2, the chicks were injected with those CHMIs once a day for three successive days then vaccinated with NDV-IV vaccine for two times with 14 days interval. Serum was collected on Day 7, 14, 21, 28, 35 and 49 after the first vaccination. The dynamic variation of serum HI antibody was determined by - micromethod. The results indicated that all of the nine ingred...
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