Synergistic, additive and antagonistic effects of Potentillafruticosa combined with EGb761 on antioxidant capacities and the possible mechanism

• Potentilla fruticosa leaves extracts combined with EGb761 showed synergism.
• The possible mechanism responsible for the synergistic effect was explored.
• Interaction of existing phytochemicals is likely responsible for the synergism.
• Caffeic acid combined with isorhamnetin played an important role for the synergism.

Potentillafruticosa and EGb 761 (Ginkgo biloba extracts) have been applied in traditional medicine and exhibit antioxidant properties. However, there is very little scientific evidence to support their use in combination. In this study, a bio-guided chromatographic fractionation strategy was used to obtain two effective fractions (C-3 and D-2) from the acetone crude extracts (A) of P. fruticosa leaves. Isobolographic analysis indicated that the combination of A + EGb had largely demonstrated synergism and 5:1 (A:EGb) displayed the best synergistic effect. In addition, C-3 + EGb exhibited a similar result as A + EGb, synergism in DPPH and ABTS assays, additive in FRAP assay. While D-2 + EGb showed only additive effect. The possible mechanism behind the synergism was explored by RP-HPLC. The changes in the total phenolic and flavonoids content were also investigated. While chemical composition did not seem to change in the mixture, no new peaks appeared in the chromatogram of C-3 + EGb mixture as compared to the individual chromatograms, no existing peaks disappeared either. Further studies were observed which may prove the possible mechanism responsible for the synergistic effect. Combinations of C-3 and 4 phenolic compounds in EGb ((+)-catechin, kaempferide, quercetin and isorhamnetin) were evaluated for their antioxidant capacities. Isorhamnetin showed synergism with C-3, and 5:1 exhibited the strongest synergism in DPPH (γ = 0.669) and ABTS (γ = 1.249), 1:1 in FRAP assays (γ = 1.385). Isorhamnetin was added to the other 6 phenolics in C-3 ((+)-catechin, caffeic acid, hyperoside, rutin, ellagic acid and quercetin), and the antioxidant capacity of mixtures was analyzed. For all combinations studied, caffeic acid + isorhamnetin displayed the greatest synergistic effect in various ratios by three antioxidant assays, the observed values were significantly higher than the expected values (P < 0.05) and 1:1 exhibited the strongest synergism in DPPH (γ = 0.711), ABTS (γ = 1.693) and FRAP (γ = 1.474) assays. Hyperoside + isorhamnetin also showed a slight synergism, but (+)-catechin + isorhamnetin exhibited an antagonism in DPPH assays. Thus, interaction of phytochemicals exist in extracts is likely responsible for the synergistic effects present in A + EGb, and caffeic acid + isorhamnetin may have played an important role. This study lends some credence to the concomitant use of P. fruticosa blended with EGb, which could be useful for developing new medicines and functional products from P. fruticosa and EGb.