Acidic Microclimate pH Distribution in PLGA Microspheres Monitored by Confocal Laser Scanning Microscopy

Purpose. The acidic microclimate pH (mu pH) distribution inside poly(lactic-co-glycolic acid) (PLGA) microspheres was monitored quantitatively as a function of several formulation variables. Methods. A ratiometric method by confocal laser scanning microscopy with Lysosensor yellow/blue (R) dextran was adapted from those previously reported, and mu pH distribution kinetics inside microspheres was examined during incubation under physiologic conditions for 4 weeks. Effects of PLGA molecular weight (MW) and lactic/glycolic acid ratio, microspheres size and preparation method, and polymer blending with poly(ethylene glycol) (PEG) were evaluated. Results. mu pH kinetics was accurately sensed over a broadly acidic range (2.8 <mu pH < 5.8) and was more acidic and variable inside PLGA with lower MW and lactic/glycolic acid ratio. Lower mu pH was found in larger microspheres of lower MW polymers, but size effects for lactic-rich polymers were insignificant during 4 weeks. Microspheres prepared by the oil-in-oil emulsion method were less acidic than those prepared by double emulsion, and blending PLGA 50/50 with 20% PEG increased mu pH significantly (mu pH > 5 throughout incubation). Conclusions. Coupling this method with that previously developed (SNARF-1 (R) dextran for mu pH 5.8-8.0) should provide microclimate pH mapping over the entire useful pH range (2.8-8.0) for optimization of PLGA delivery of pH-sensitive bioactive substances.