Visualisation of Freezing Behaviours in Flower Bud Tissues of Cold-hardy Rhododendron japonicum by Nuclear Magnetic Resonance Micro-Imaging

Abstract

¹H nuclear magnetic resonance (NMR) micro-imaging was used to study the freezing behaviour of wintering flower buds of Rhododendron japonicum (A. Gray) Suringer. Amulti-slice multi- echo pulse sequence was used to acquire images at different subfreezing temperatures. The images obtained predominantly reflected the density of mobile (i.e. non-ice) protons mainly from unfrozen water. By comparing these images taken at various subfreezing temperatures, we could determine which tissues produced high temperature exotherms and low temperature exotherms in differential thermal analyses. In flower buds of the cold-hardy R. japonicum, typical extra-organ freezing was successfully imaged. The scales readily froze at –7°C but some florets remained supercooled even at –21°C. The size of the supercooled florets was reduced with decreasing temperature which indicated a gradual decrease in floret water content. With decreasing temperature, there was a gradual decrease in the signal intensity of the flower bud axis including the peduncle and immature pith tissues, which implies either dehydration or partial freezing of these tissues. Deep supercooling in the entire mature pith tissues was also clearly visible in these images. Due to its non-invasive nature, NMR micro-imaging is a useful tool for studying freezing behaviours in various plant tissues, especially for imaging organised or harmonised freezing in complex organs as well as for clarifying the diversity and mechanisms involved in freezing behaviours.