Determination of MIC of combined γ-mangostin and penicillin G
Fifty microliters of two-fold serial dilution of penicillin G (final concentration ranging from 0.0125 to 6.25 μg/ml) was pipetted into well containing 50 μl ofγ-mangostin (final concentration ranging from 1.56 to 50 μg/ml). After mixing, 100 μl of leptospira inoculum (2×106 cells/ml) was added to each well. The plate was performed in the same conditions used to determine the MIC of the crude extracts and xanthones. The MIC of combination was deemed to be the lowest concentration of both γ-mangostin and penicillin G which inhibited the growth of leptospires in the same well.
Evaluation of the synergistic effect Synergy was evaluated by calculating the fractional inhibi- tory concentration (FIC) index as described previously [36]: FIC index = FICA + FIC B = [A]/MIC A + [B]/MIC B,
Table 1 Minimal inhibitory concentrations (MIC) of four crude extracts and five xanthones purified from G. mangostana against one non-pathogenic and four pathogenic leptospira Compounds MIC (μg/ml) Code MW Structure Type Patoc Bataviae Autumnalis Javanica Saigon SS-WS01 - ND Crude extract ≥ 800 ≥800 400 400 ≥800 SS-WS02 - ND Crude extract ≥800 400 200 400 ≥800 SS-WS03 - ND Crude extract ≥800 ≥800 400 200 ≥800 SS-WS04 - ND Crude extract ≥800 400 200 200 400 1 410 C24H26O6 α-Mangostin ≥800 400 ≥800 100 100 2 396 C23H24O6 γ-Mangostin 200 100 ≥800 100 100 3 414 C23H26O7 Garcinone C 200 100 100 100 100 4 428 C24H28O7 Garcinone D ≥800 ≥800 ≥800 200 200 5 380 C23H24O5 8-Desoxygartanin ≥800 ≥800 400 400 200 *Penicillin G 6.25 1.56