Characterization of microcapsules by confocal laser scanning microscopy: structure, capsule wall composition and encapsulation rate.

The potential of confocal laser scanning microscope (CLSM) has been evaluated for characterizing microcapsules. The aim was to visualize the polymer distribution within the particle wall, and to localize and to quantify the encapsulated oil phase. Microcapsules were prepared by complex coacervation: the oil phase, gelatine, and arabic gum were labelled with fluorescent markers. For all compounds it was proved that fluorescence labelling did not alter physico-chemical properties critical to the encapsulation process. Labelling of the inner oil phase allowed us to identify and to localize, three-dimensionally, the encapsulated compound. A homogeneous distribution for both gelatine and arabic gum throughout the capsule wall was observed. The addition of fluorescently labelled casein as a macromolecular model compound to the coacervate resulted in an inhomogeneous distribution of casein within the wall material, the highest concentration of casein was found at the oil-wall interface. To determine the encapsulation rate, CLSM pictures of the microcapsule samples were acquired using different fluorescence labels for the microcapsule wall polymers and the incorporated oil phase, respectively. By applying computational image analysis, the volumes of the different phases were calculated. Comparing the results of non-destructive image analysis with those obtained by degradation, extraction and chemical analysis, a linear relation was found with correlation coefficients better than 0.980.