Mode of action of 6-oxophenolic triterpenoids against Bacillus subtilis

Four 6-oxophenolic triterpenoids were isolated from Maytenus blepharodes Lundell, with activity against Gram positive bacteria, especially on spore forming bacteria, and the yeast Candida albicans [1]. Chemically, these triterpenoids differ in the functional group in ring A and E (Figure 1). The MIC values against B. subtilis suggest that the increase of the electronegativity produces a decrease of the antibacterial potency (1 zeylasterone 3µg/ml < 2 demethylzeylasterone 13µg/ml and 3 zeylasteral 10µg/ml < 4 demethylzeylasteral 13µg/ml).

Fig.1: Structure of 6-oxophenolic triterpenoids

	R1	R2	MIC (µg/ml)
1.	COOH	COOCH3	3
2.	COOH	COOH	13
3.	COH	COOCH3	10
4.	COH	COOH	13

The activity of the compounds (between 2 and 3×MIC) on B. subtilis cultures varies according to the bacteria concentrations, showing bactericidal activity (≥3 log₁₀ in CFU/ml reduction) at lowest inocula size (10⁵-10⁴ CFU/ml), except for demethylzeylasterone and demethylzeylasteral. At highest inocula (10⁷-10⁶ CFU/ml), the phenols showed bacteriostatic activity (<3 log₁₀ in CFU/ml reduction), except demethylzeylasteral.

Treatments of B. subtilis cultures with the phenolic compounds revealed a blockage of biosynthetic pathways as DNA, RNA, protein and cell wall synthesis. However, inhibition of all processes did not occur simultaneously. The macromolecular synthesis of peptidoglycan was stopped 2min after addition of the phenols, and later DNA, RNA and protein synthesis. These results suggest that the compounds could act on multiple targets on B. subtilis cells.

Acknowledgements: This study was supported by the Spanish Grant CTQ2006–13376/BQU.

References:

1. De León, L., Beltrán, B., Moujir, L. (2005) Plant. Med. 71: 313–319.