In vivo physiological measurement is a major challenge in modern science and technology, as is environment conservation at the global scale. Proper toxicological testing of widely produced mixtures of chemicals is a necessary step in the development of new products, allowing us to minimize the human impact on aquatic ecosystems. However, currently available bioassay-based techniques utilizing small aquatic organisms such as fish embryos for toxicity testing do not allow assessing in time the changes in physiological parameters in the same individual. In this study, we introduce microencapsulated fluorescent probes as a promising tool for in vivo monitoring of internal pH variation in zebrafish embryos. The pH alteration identified under stress conditions demonstrates the applicability of the microencapsulated fluorescent probes for the repeated analysis of the embryo's physiological state. The proposed approach has strong potential to simultaneously measure a range of physiological characteristics using a set of specific fluorescent probes and to finally bring toxicological bioassays and related research fields to a new level of effectiveness and sensitivity.