Derivatives of oxazine dyes were synthesized on mulitigram scales via efficient synthetic strategies. One practical route was selected to prepare compounds 6, 9 and 10, especially water-soluble compound 6 was obtained in better yield than reported, and compound 10 was insoluble in aqueous media in absence of phenolic-OH. Compounds 3 and 9 were found to be clear pH-dependent between pH = 4.0 and 10.0, and could be used as acid-base indicators to measure intracellular pH. Compounds 6, 9, 10 all have carboxylic acid functionalities, which could be activated and used to conjugate the dyes to biomolecules. In addition, compounds 6 and 9 with good solubility in aqueous media were used to develop a simple, quick, safe, highly sensitive staining method to detect PHAs-producing bacteria on heat-fixed smears, which was confirmed by fluorescence images of PHAs granules of bacteria.