Mechanical and enzymatic separation of ripening rice (Oryza sativa L.) caryopsis tissues

Abstract

The ripening rice (Oryza sativa L.) caryopsis contains several maternal and embryonic tissues that transport assimilates along the partitioning pathway. Experimental access to transport steps in the pathway is limited by the separability of the tissues. Hence, the extent to which tissues can be mechanically and enzymatically separated was assessed. The caryopsis coat was isolated during mid-ripening, dissected and microscopically characterized. In centripetal order, the pericarp (epidermis, parenchyma-cell layer, cross-cell layer, tube-cell layer), inner integument, nucellar epidermis, aleurone layer and subaleurone layer adhered to each other. There was mechanical separation at the tube-cell layer, but not at the maternal/embryonic interface. Aleurone- and subaleurone-layer cells exposed on the inner surface of isolated caryopsis coats were macerated with Pectolyase Y-23 and manually sheared, which freed the endosperm and exposed the nucellar epidermis. Yield of endosperm cells increased linearly with the number of coats and reached a maximum after 1 h. The proportion of cells that were viable was approximately 10%. Subaleurone-layer cells contributed less than 25% to the yield. These results suggest that pectin polymers are necessary for adherence between the nucellar epidermis and the aleurone layer, and that enzymatic maceration is useful for separating the tissues at the maternal/embryonic interface.