The Structure-Activity Relationship of Pterostilbene Against Candida albicans Biofilms

Abstract

Candida albicans biofilms contribute to invasive infections and dramatic drug resistance, and anti-biofilm agents are urgently needed in the clinic. Pterostilbene (PTE) is a natural plant product with potentials to be developed as an anti-biofilm agent. In this study, we evaluated the structure-activity relationship (SAR) of PTE analogues against C. albicans biofilms. XTT (Sodium 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide inner salt) reduction assay was used to evaluate the activity of the analogues against C. albicans biofilms. Knowing that hyphal formation is essential for C. albicans biofilms, anti-hyphal assay was further carried out. By comparing a series of compounds tested in this study, we found that compounds with para-hydroxy (-OH) in partition A exhibited better activity than those with other substituents in the para position, and the double bond in partition B and meta-dimethoxy (-OCH₃) in partition C both contributed to the best activity. Consistent results were obtained by anti-hyphal assay. Collectively, para-hydroxy (-OH), double bond and meta-dimethoxy (-OCH₃) are all needed for the best activity of PTE against C. albicans biofilms.

Keywords: Candida albicans; anti-biofilm; pterostilbene; structure-activity relationship.

Figure 1

Chemical structures of PTE and three partitions of PTE.

Figure 2

Effects of PTE and analogues C1–C5, B1, B2 on biofilm formation. (A) Chemical structures of PTE and analogues C1–C5, B1, and B2 are shown. The structures of analogue C1–C5, B1, and B2 are similar with PTE, and differences only exist in partition A; (B) Biofilm formation was evaluated by XTT reduction assay, and the results were presented as the percentage compared to the control biofilms formed without compound treatment. Biofilm formation results represent the mean ± standard deviation fromthree independent experiments. * p < 0.05 compared with the control biofilms, ** p < 0.01 compared with the control biofilms, *** p < 0.001 compared with the control biofilms. Statistical analysis was performed to compare the effects of various compounds, and significant difference was observed upon all the analogues compared with PTE. p = 0.035 (C4 vs. PTE), p < 0.001 (C1 vs. PTE), p < 0.001 (C2 vs. PTE), p = 0.0047 (C3 vs. PTE), p < 0.001 (B1 vs. PTE), p < 0.001 (B2 vs. PTE), and p < 0.001 (C5 vs. PTE).

Figure 3

Effects of different concentrations of PTE and analogues C1–C5, B1, and B2 on hyphal formation. Exponentially-growing C. albicans SC5314 cells were transferred to hypha-inducing liquid media, including Spider and RPMI1640. The cellular morphology was photographed after incubation at 37 °C for 3 h. Scale bar = 100 μm.

Figure 4

Effects of different concentrations of analogues C6, B3, C7, and B4 on biofilm formation and hyphal formation. (A) Chemical structures of analogues C6, B3, C7 and B4 are shown. C6, B3, C7, and B4 are both different from PTE in partition B; (B) Effects of different concentrations of analogues C6, B3, C7, and B4 on biofilm formation. Biofilm formation results represent the mean ± standard deviation for three independent experiments. * p < 0.05 compared with the control biofilms, ** p < 0.01 compared with the control biofilms, *** p < 0.001 compared with the control biofilms. Statistical analysis was performed to compare the effects of various compounds, and significant difference was observed upon C6, C7, and B4 compared with PTE. p = 0.037 (C6 vs. PTE), p = 0.175 (B3 vs. PTE), p = 0.0033 (C7 vs. PTE), and p < 0.001 (B4 vs. PTE); (C) Effects of different concentrations of analogues C6, B3, C7, and B4 on hyphal formation. Scale bar = 100 μm.

Figure 5

Effects of different concentrations of analogues B6, B5, C8. and C9 on biofilm formation and hyphal formation. (A) Chemical structures of analogues B6, B5, C8, and C9 are shown. B6, B5, C8, and C9 are different from PTE in partition C; (B) Effects of different concentrations of analogues B6, B5, C8, and C9 on biofilm formation. Biofilm formation results represent the mean ± standard deviation for three independent experiments. * p < 0.05 compared with the control biofilms, ** p < 0.01 compared with the control biofilms, *** p < 0.001 compared with the control biofilms. Statistical analysis was performed to compare the effects of various compounds, and significant difference was observed upon B5, C8, and C9 compared with PTE. p = 0.182 (B6 vs. PTE), p = 0.041 (B5 vs. PTE), p < 0.001 (C8 vs. PTE), and p < 0.001 (C9 vs. PTE); (C) Effects of different concentrations of analogues B6, B5, C8, and C9 on hyphal formation. Scale bar = 100 μm.