Fig. 4

Synergistic antiviral activity of caffeic acid, perilla aldehyde, and perillyl alcohol. A Pooled icELISA data of 3 independent experiments of SARS-CoV-2-infected Vero E6 and Caco-2 cells after treatment with components of herbal infusions at 1 h p.i or 1.5 h p.i. for 1.5 h. Cells were fixed at 20 to 24 h p.i. and stained with α-S (Vero E6) or α-N mAb (Caco-2). Each condition was analyzed in triplicate. The treated conditions were compared to the untreated control by one-way ANOVA. *, p < 0.05. **, p < 0.01. ***, p < 0.001. IC50 values (compound concentration sufficient to reduce the virus-specific icELISA signal to 50%) were calculated by nonlinear regression for Vero E6 and Caco-2. Caffeic acid, 82.95 and 34.64 µg/ml. Perilla aldehyde, 94.99 and 77.04 µg/ml. Perillyl alcohol, 252.7 and 66.62 µg/ml. B icELISA data of SARS-CoV-2-infected Vero E6 cells after treatment with indicated compounds (caffeic acid, 25 µg/ml; perilla aldehyde and perillyl alcohol, 125 µg/ml) at 1 h p.i. for 1.5 h. Cells were fixed at 20 h p.i. and stained with α-S mAb. Each condition was analyzed in triplicate. See Additional file 1 for individual data values. C Combination of different concentrations of caffeic acid and perilla aldehyde or perillyl alcohol were tested for their antiviral activity. The inhibition (%) compared to the untreated control was calculated and the mean values of 3 replicates were included to test for synergistic effects. The R package SynergyFinder was used to calculate the Loewe synergy score [33]. The analysis was performed without impute method and baseline correction. D Pooled icELISA data of SARS-CoV-2-infected cells (Vero E6, 4 independent experiments; Caco-2, 3 independent experiments) after treatment with perilla or thyme infusion. Data are expressed as relative change in optical density compared to the untreated control. All treated conditions were compared to the untreated control by one-way ANOVA. *, p < 0.05. **, p < 0.01. ***, p < 0.001. E, F Dose-response curves of SARS-CoV-2-infected Vero E6 cells (2000 PFU per well) after treatment for 1 h or 30 min with aqueous infusions of mint (E, fresh mint; F, 1 h treatment with aqueous infusion of 2 commercially available mint tea bags in 100 ml water). Each condition was analyzed in triplicate. See Additional file 1 for individual data values. The mint-treated conditions were compared to the untreated control by one-way ANOVA. *, p < 0.05. **, p < 0.01. ***, p < 0.001