Comparison Between Fluorescent Probe and Ion-Selective Electrode Methods for Intracellular pH Determination in Leuconostoc mesenteroides

The intracellular pH (pH_in) of Leuconostoc mesenteroides subsp. mesenteroides 19D was evaluated by two different methods, fluorescent probe and ion-selective electrode. Two fluorescent probes 5 (and-6)-carboxyfluorescein diacetate succinimidyl ester (cFDASE) and 5 (and-6)-carboxy-2',7'-dichlorofluorescein diacetate succinimidyl ester (cDCFDASE) were tested to evaluate the intracellular pH (pH_in) of living cells at a medium pH (pH_ex) ranged from 5.0 to 6.5. Salicylic acid was used as a probe for the ion-selective electrode method. Cells kept 60-80% of cFDASE probe at all pH_ex values against 5-10% of cDCFDASE probe at pH_ex ≤ 6.0. The pH_in values measured by the ion-selective electrode were higher by 0.1-0.6 pH units at pH_ex ranged from 5.0 to 6.5 than those determinated by fluorescent probe method. The possibility to study the intracellular pH at a wide external pH range using a single probe, and the simplicity of the material and experimental protocol may make the ion-selective electrode method most useful and easy to measure the intracellular pH of lactic acid bacteria compared with the other techniques like fluorescent probes.