4 methoxycinnamaldehyde

Trans-cinnamaldehyde (99%) and 10 of its derivatives: cinnamaldehyde oxime (95%), α-methylcinnamaldehyde (95%), 2-methoxycinnamaldehyde (95%), 2-nitrocinnamaldehyde (98%), 4-bromocinnamaldehyde (95%), 4-methoxycinnamaldehyde (95%), 4-nitrocinnamaldehyde (95%), 4-dimethylaminocinnamaldehyde (98%), 4-fluorocinnamaldehyde (97%), and 4-chlorocinnamaldehyde (95%) were purchased from Sigma-Aldrich (St. Louis, MO, USA) or Combi-Blocks (San Diego, CA, USA). Their molecular weights and chemical structures are given in Table 1. Stocks of 100 mg/ml were prepared in dimethyl sulfoxide (DMSO) and kept at −20°C until required. A total of 0.1% (v/v) DMSO was used as the control for antibacterial and biofilm experiments; at this concentration DMSO did not effect on bacterial growth or biofilm formation.

Agonist (cinnamaldehyde, natural, ≥95%, FG-grade) and the blocker (HC-030031, ≥98%, HPLC-grade) were obtained from Sigma-Aldrich, Overijse, Belgium. Derivates (2-methoxycinamaldehyde, >96%, FG-grade; 4-methoxycinnamaldehyde, >96%, FG-grade; hydrocinnamaldehyde, >90%, FCC-grade; cinnamic acid >99%) were obtained from Sigma-Aldrich (Overijse, Belgium). Dimethyl sulfoxide (DMSO) and phenylmethanesulfonylfluoride (PMSF) were also purchased from Sigma-Aldrich (Overijse, Belgium) as well as. Chemicals to make Krebs-Ringer buffer (KRB), as described in the previous section, were purchased from the same company. Cytotoxicity detection kit (PLUS) LDH (Roche Diagnostics, Indianapolis, IN, USA) was also purchased from Sigma-Aldrich.