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In vitro synergistic antioxidant activity and identification of antioxidant components from Astragalus membranaceus and Paeonia lactiflora.

Xu X, Li F, Zhang X, Li P, Zhang X, Wu Z, Li D - PLoS ONE (2014)

Bottom Line: Many traditionally used herbs demonstrate significantly better pharmacological effects when used in combination than when used alone.Among them, E1 exhibited the strongest synergistic effect in scavenging DPPH radicals and reducing ferric ions (P<0.05).A strong correlation between the increment of total phenolic/flavonoid and synergistic antioxidant activity, especially between the increment of total flavonoid and the increase in ferric reducing power was observed.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science, Shandong Agricultural University, Taian, China.

ABSTRACT
Many traditionally used herbs demonstrate significantly better pharmacological effects when used in combination than when used alone. However, the mechanism underlying this synergism is still poorly understood. This study aimed to investigate the synergistic antioxidant activity of Astragalus membranaceus (AME) and Paeonia Lactiflora (PL), and identify the potential antioxidant components by 1,1-diphenyl-2-picrylhydrazine (DPPH) radical spiking test followed by a high performance liquid chromatography separation combined with diode array detection and tandem mass spectrometry analysis (DPPH-HPLC-DAD-MS/MS). Eight AME-PL combined extracts (E1-E8) were prepared based on bioactivity-guided fractionation. Among them, E1 exhibited the strongest synergistic effect in scavenging DPPH radicals and reducing ferric ions (P<0.05). Moreover, E1 presented strong cytoprotection against H2O2-induced oxidative damage in MRC-5 cells by suppressing the decrease of the superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and catalase (CAT) activities. A strong correlation between the increment of total phenolic/flavonoid and synergistic antioxidant activity, especially between the increment of total flavonoid and the increase in ferric reducing power was observed. Finally, seven antioxidant substances were identified in E1 as oxypaeoniflora, catechin, calycosin-7-O-β-D-glucopyranoside, fomononetin-7-O-β-D-glucopyranoside, 9,10-dimethoxy-pterocarpan-3-O-β-D-glucopyranoside, quercetin and 2'-dihydroxy-3',4'-dimethyl-isoflavan-7-O-β-D-glucopyranoside.

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DPPH radical-scavenging activity (A) and CI values (B) of different combinations of AME and PL solvent-extracted fractions.Data are expressed as means ± SD (n = 3), and histograms marked with different letters are significantly different at P<0.05.
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pone-0096780-g001: DPPH radical-scavenging activity (A) and CI values (B) of different combinations of AME and PL solvent-extracted fractions.Data are expressed as means ± SD (n = 3), and histograms marked with different letters are significantly different at P<0.05.

Mentions: The DPPH radical-scavenging activity of different combined fractions from AME and PL are shown in Fig.1. Sixteen combined fractions exhibited a wide range of differences in scavenging DPPH free radicals. Among them, the ethyl acetate fraction of Paeonia lactiflora (EA-PL) presented the highest potency in scavenging DPPH radical when used in combination with four solvent-extracted fractions from AME (P<0.05), suggesting that EA-PL was rich in DPPH radical-scavenging activity (Fig.1A). Our results were in agreement with the previous report [19]. Herein, CIs were used to determine the possible interactive actions between the extracts or fractions. In order to calculate the CIs, dose-effect curves for the single extract or fraction were analyzed (data not shown). As shown in Fig.1B, the CIs for EA-PL+CF-AME and EA-PL+NB-AME were <1.0, indicating these combined extracts had a synergistic effect in scavenging DPPH radical. By contrast, the CIs for several combined fractions (i.e. CF-PL+EA-AME, CF-PL+NB-AME, and OH-PL+EA-AME) were >1.0, suggesting these combinations had antagonistic effects in scavenging DPPH radicals. As is well known, the phenolic and flavonoid compounds are the most commonly studied substances that greatly contributed to antioxidant activity of plant foods. Therefore, we measured the total phenolic and flavonoid contents in EA-PL+CF-AME combination, which exhibited the strongest activity in scavenging DPPH radicals. Results showed that the total phenolic and flavonoid contents were 603.339±22.894 mg GAE/g and 121.785±1.264 mg RE/g, respectively, which were significantly higher than those of other combinations (P<0.05, data not shown). These results suggested that the EA-PL fraction had the highest potency in scavenging DPPH radicals to warrant further fractionation. Thus, EA-PL was further chromatographed on a silica gel column (5.5×60 cm) using a stepwise elution of methanol/chloroform (methanol/chloroform  = 1∶19, 1.5∶18.5, 2∶18, 2.5∶17.5, 3∶17, 4∶16, 5∶15, 20∶0, v/v) to afford 8 fractions (A1-A8). Then, each fraction was combined with CF-AME to yield eight AME-PL combined extracts, i.e., E1 (A1+CF-AME), E2 (A2+CF-AME), E3 (A3+CF-AME), E4 (A4+CF-AME), E5 (A5+CF-AME), E6 (A6+CF-AME), E7 (A7+CF-AME) and E8 (A8+CF-AME). These samples were examined for in vitro antioxidant activity using DPPH free radical scavenging assay and FRAP test.


In vitro synergistic antioxidant activity and identification of antioxidant components from Astragalus membranaceus and Paeonia lactiflora.

Xu X, Li F, Zhang X, Li P, Zhang X, Wu Z, Li D - PLoS ONE (2014)

DPPH radical-scavenging activity (A) and CI values (B) of different combinations of AME and PL solvent-extracted fractions.Data are expressed as means ± SD (n = 3), and histograms marked with different letters are significantly different at P<0.05.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4016014&req=5

pone-0096780-g001: DPPH radical-scavenging activity (A) and CI values (B) of different combinations of AME and PL solvent-extracted fractions.Data are expressed as means ± SD (n = 3), and histograms marked with different letters are significantly different at P<0.05.
Mentions: The DPPH radical-scavenging activity of different combined fractions from AME and PL are shown in Fig.1. Sixteen combined fractions exhibited a wide range of differences in scavenging DPPH free radicals. Among them, the ethyl acetate fraction of Paeonia lactiflora (EA-PL) presented the highest potency in scavenging DPPH radical when used in combination with four solvent-extracted fractions from AME (P<0.05), suggesting that EA-PL was rich in DPPH radical-scavenging activity (Fig.1A). Our results were in agreement with the previous report [19]. Herein, CIs were used to determine the possible interactive actions between the extracts or fractions. In order to calculate the CIs, dose-effect curves for the single extract or fraction were analyzed (data not shown). As shown in Fig.1B, the CIs for EA-PL+CF-AME and EA-PL+NB-AME were <1.0, indicating these combined extracts had a synergistic effect in scavenging DPPH radical. By contrast, the CIs for several combined fractions (i.e. CF-PL+EA-AME, CF-PL+NB-AME, and OH-PL+EA-AME) were >1.0, suggesting these combinations had antagonistic effects in scavenging DPPH radicals. As is well known, the phenolic and flavonoid compounds are the most commonly studied substances that greatly contributed to antioxidant activity of plant foods. Therefore, we measured the total phenolic and flavonoid contents in EA-PL+CF-AME combination, which exhibited the strongest activity in scavenging DPPH radicals. Results showed that the total phenolic and flavonoid contents were 603.339±22.894 mg GAE/g and 121.785±1.264 mg RE/g, respectively, which were significantly higher than those of other combinations (P<0.05, data not shown). These results suggested that the EA-PL fraction had the highest potency in scavenging DPPH radicals to warrant further fractionation. Thus, EA-PL was further chromatographed on a silica gel column (5.5×60 cm) using a stepwise elution of methanol/chloroform (methanol/chloroform  = 1∶19, 1.5∶18.5, 2∶18, 2.5∶17.5, 3∶17, 4∶16, 5∶15, 20∶0, v/v) to afford 8 fractions (A1-A8). Then, each fraction was combined with CF-AME to yield eight AME-PL combined extracts, i.e., E1 (A1+CF-AME), E2 (A2+CF-AME), E3 (A3+CF-AME), E4 (A4+CF-AME), E5 (A5+CF-AME), E6 (A6+CF-AME), E7 (A7+CF-AME) and E8 (A8+CF-AME). These samples were examined for in vitro antioxidant activity using DPPH free radical scavenging assay and FRAP test.

Bottom Line: Many traditionally used herbs demonstrate significantly better pharmacological effects when used in combination than when used alone.Among them, E1 exhibited the strongest synergistic effect in scavenging DPPH radicals and reducing ferric ions (P<0.05).A strong correlation between the increment of total phenolic/flavonoid and synergistic antioxidant activity, especially between the increment of total flavonoid and the increase in ferric reducing power was observed.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science, Shandong Agricultural University, Taian, China.

ABSTRACT
Many traditionally used herbs demonstrate significantly better pharmacological effects when used in combination than when used alone. However, the mechanism underlying this synergism is still poorly understood. This study aimed to investigate the synergistic antioxidant activity of Astragalus membranaceus (AME) and Paeonia Lactiflora (PL), and identify the potential antioxidant components by 1,1-diphenyl-2-picrylhydrazine (DPPH) radical spiking test followed by a high performance liquid chromatography separation combined with diode array detection and tandem mass spectrometry analysis (DPPH-HPLC-DAD-MS/MS). Eight AME-PL combined extracts (E1-E8) were prepared based on bioactivity-guided fractionation. Among them, E1 exhibited the strongest synergistic effect in scavenging DPPH radicals and reducing ferric ions (P<0.05). Moreover, E1 presented strong cytoprotection against H2O2-induced oxidative damage in MRC-5 cells by suppressing the decrease of the superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and catalase (CAT) activities. A strong correlation between the increment of total phenolic/flavonoid and synergistic antioxidant activity, especially between the increment of total flavonoid and the increase in ferric reducing power was observed. Finally, seven antioxidant substances were identified in E1 as oxypaeoniflora, catechin, calycosin-7-O-β-D-glucopyranoside, fomononetin-7-O-β-D-glucopyranoside, 9,10-dimethoxy-pterocarpan-3-O-β-D-glucopyranoside, quercetin and 2'-dihydroxy-3',4'-dimethyl-isoflavan-7-O-β-D-glucopyranoside.

Show MeSH
Related in: MedlinePlus