| Rationales: Aging is inevitable.With aging,the body's ability to resist external stress is gradually weakened and the incidence of diseases related to aging is exacerbated.Moreover,with the increasing economic and living standards,health and anti-aging has become an urgent need for people at the moment.Therefore,the selection of effective anti-aging ingredients from traditional Chinese medicine has become a hot topic of research.Scutellaria baicalensis is the dry root of Scutellaria baicalensis Georgi.Its active ingredients are mainly flavonoids,including baicalein,baicalin,wogonoside,wogonin,etc.Their pharmacological effects are extensive,including antioxidation,anti-inflammatory,neuroprotective,anti-viral and anti-tumor.The literatures reported that crude extracts of Scutellaria baicalensis such as alcohol extracts,aqueous extracts,flavonoids from stems and leaves,and polysaccharides have anti-aging activity.At present,the researches on antiaging active ingredients in Scutellaria baicalensis mainly focus on the baicalin and baicalein.The anti-aging study of other flavonoids in Scutellaria baicalensis has not been reported.Therefore,this project aims to screen out the anti-aging active parts of Scutellaria baicalensis and evaluate the antiaging activity and mechanism of the main flavonoids that have no reported anti-aging effects in various parts.Objective: 1.Confirm the effective fractions preparation process and verify their anti-aging activity;identify the anti-aging activity and mechanism of the main flavonoids in various parts.Methods: 1.Macroporous adsorption resin was used to separate the ethanol extract of Scutellaria baicalensis.Three fractions were obtained: water,50% ethanol and 95% ethanol,and their main chemical components were analyzed used high performance liquid chromatography(HPLC).Using Drosophila melanogaster as a model,three fractions were obtained to feed fruit flies at two different concentrations(0.15 mg/mL and 3 mg/mL),and the blank was used as control group to examine the effect on the lifespan of natural sensational Drosophila.2.The composition of the anti-senile parts(50% ethanol fraction)was analyzed by HPLC,and the anti-aging effect of the wogonoside which has not reported anti-aging activity was studied.The flies were fed with different concentrations of wogonoside(0 mg/mL,0.02 mg/mL and 0.5 mg/mL)to investigate the effect on the lifespan of fruit flies.The anti-oxidation activity in vivo was evaluated by detecting the activities of superoxide dismutase(SOD)and catalase(CAT),and the expression levels of the copper zinc superoxide dismutase(SOD1)gene,manganese superoxide dismutase(SOD2)gene,CAT gene,and methuselah(MTH)gene.A metabolomics study by ultra performance liquid chromatography tandem mass spectrometry(UPLCMS/MS)was used to investigate the mechanism of the effect of wogonoside on the lifespan of fruit flies.3.The composition of the 95% ethanol fraction was analyzed by HPLC,and the anti-aging effect of the wogonin which has not reported anti-aging activity was studied.The flies were fed with different concentrations of wogonin(0 mg/mL,0.02 mg/mL and 0.5 mg/mL)to investigate the effect on the lifespan of fruit flies.The anti-oxidation activity in vivo was evaluated by detecting the activities of SOD and CAT,and the expression levels of the gene SOD1,SOD2,CAT,and MTH.A metabolomics study by UPLCMS/MS was used to investigate the mechanism of the effect of wogonin on the lifespan of fruit flies.Results: 1.The HPLC results showed that the ultraviolet absorption of the water fraction was weak and should be mainly polysaccharides.The main components of 50% ethanol fraction are baicalin and wogonoside,and the main components of 95% ethanol fraction are baicalein and wogonin.The lifespan results showed that the water and 50% ethanol fractions could significantly prolong the mean lifespan(P<0.001),and 50% ethanol fraction could significantly prolong the maximum lifespan of Drosophila(P<0.05 or P<0.01).2.Wogonoside can significantly prolonged the mean(P<0.01)and maximum lifespan(P<0.05 or P<0.001)of Drosophila.The results of biochemical indicators showed that wogonoside significantly increased the activity of antioxidant enzymes SOD and CAT in Drosophila(P<0.05 or P<0.01).The RT-PCR results showed that wogonoside significantly upregulated the expression levels of SOD1,SOD2 and CAT(P<0.01 or P<0.001),and down-regulated the expression of MTH(P<0.001).The metabolic profile of different fruit fly groups was analyzed and a total of 27 differential metabolites related to aging were found.And there were 17 potential biomarkers were recalled by wogonoside,including: L-Arginine,Pyroglutamic Acid,L-Glutamine,L-Acetylcarnitine,UDP-Nacetylglucosamine,L-Isoleucine,3-Hydroxydecanoic acid,LysoPE(18:3),LysoPC(18:3),LysoPE(16:1),LysoPC(16:1),LysoPC(16:0),Linoleyl carnitine,LysoPE(18:1),LysoPC(18:1),L-Palmitoylcarnitine,Elaidiccarnitine.These potential biomarkers are mainly involved in amino acid metabolism(D-glutamine and D-glutamate metabolism;alanine,aspartate and glutamate metabolism;arginine and proline metabolism;valine,leucine and isoleucine biosynthesis)and energy metabolism(nitrogen metabolism).3.The lifespan results showed that wogonin could prolong the mean lifespan(P<0.05 or P<0.001)and maximum lifespan(P<0.01)of Drosophila.The results of biochemical indicators showed that wogonin significantly increased the activity of antioxidant enzymes SOD and CAT in Drosophila(P<0.05).The RT-PCR results showed that wogonin significantly upregulated the expression levels of SOD1 and SOD2(P<0.05),and downregulated the expression of MTH(P<0.05 or P<0.01).The metabonomics results showed that wogonin could regulate 16 differential metabolites related to aging in Drosophila,including: L-Arginine,Pyroglutamic Acid,Choline,L-Glutamine,UDP-N-acetylglucosamine,L-Isoleucine,3-Hydroxydecanoic acid,LysoPE(18:3),LysoPE(16:1),LysoPC(16:1),LysoPC(16:0),Linoleyl carnitine,LysoPE(18:1),LysoPC(18:1),L-Palmitoylcarnitine,Elaidic carnitine.These differential metabolites are mainly involved in amino acid metabolism(D-glutamine and D-glutamate metabolism;alanine,aspartate and glutamate metabolism;arginine and proline metabolism;valine,leucine and isoleucine biosynthesis),energy metabolism(nitrogen metabolism)and lipid metabolism(glycerophospholipid metabolism).Conclusion: The water and 50% ethanol fractions of Scutellaria baicalensis can prolong the lifespan of Drosophila.Both wogonoside and wogonin can delay the senescence of Drosophila melanogaster.Their mechanism is closely related to increasing antioxidant enzyme activity,upregulating antioxidant gene expression and regulating different metabolic pathways,thereby ameliorating oxidative stress and metabolic disorders in the body. |
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