Visualization of enzyme activity in germinating cereal seeds using a lipase sensitive fluorochrome

Abstract

The development patterns of hydrolytic enzymes during germination of barley, wheat, rye and sorghum were examined by a fluorescence staining technique. The method was based upon conversion of non-fluorescent fluorescein dibutyrate to highly fluorescent fluorescein, a reaction known to be preferentially catalyzed by lipase. The formation of hydrolases was compared with cell wall breakdown in germinating barley. Fluorescence microscopy of longitudional sections of germinated cereal seeds revealed that a considerable amount of hydrolytic enzymes reacting with fluorescein dibutyrate was produced as germination proceeded. The initial formation site of the enzymes was found to be in the scutellum. Later in germination the hydrolases gradually diffused into the entire endosperm. The diffusion pattern of hydrolases in barley was found to coincide with cell wall breakdown. It is concluded that the fluorescein dibutyrate method has a potential as an indicator of the transport of de novo synthetized enzymes in cereal seeds. Practical applications of the fluorescein dibutyrate method are discussed. © 1982 Carlsberg Laboratory.