Identification and separation of water tracing dyes using pH response characteristics

Problems encountered in field applications of water tracing dyes include background fluorescence and interference between dyes, where the complexity of hydrological systems, particularly in karst areas, makes multiple dye tracing experiments desirable. The fluorescence of water tracing dyes may vary dramatically according to the pH of the water, and this property can be used to address these problems. The response of a number of commonly available water tracing dyes to pH is investigated; the distinctive curves provide a means of distinguishing between dyes, and between dyes and background fluorescence caused by algae. It is not necessary to reproduce the full pH response curve to utilize these properties. Recommendations are made in the use of pH buffering to improve sensitivity in detecting dyes, and for quantitative laboratory analysis. Buffering to a small number of predetermined pH values can be used effectively to distinguish both between dyes which fluoresce at the same wavelength, and between natural and induced fluorescence. It is particularly useful in the case of the green dyes, both for detection when there are high natural backgrounds and because it allows more than one green dye to be used simultaneously. If Rhodamine is used together with, for example, Fluorescein and Pyranine, triple dye tracing experiments can be executed.