Electron-energy-loss spectroscopic imaging of calcium and nitrogen in the cell walls of apple fruits

Changes in texture are an integral part of ripening in most fleshy fruits a nd these changes are thought to be determined, primarily, by alterations in cell wall structure. Electron energy loss spectroscopy (EELS) imaging was used to obtain quantitative information on the levels of calcium and nitrog en in the cell walls of apple (Malus domestica Borkh. cv. Cox's Orange Pipp in) fruits. Samples of fruit cortex were prepared for EELS by high-pressure freezing and molecular distillation drying to minimize loss and redistribu tion of soluble cell wall components such as calcium. The EELS imaging succ essfully resolved calcium and nitrogen levels in the middle lamella and pri mary cell wall. When the elemental compositions of the cell walls of Cox's apples from two sites in the UK were compared at harvest or after 6 months storage, the orchard which always produced consistently firmer fruit had si gnificantly lower levels of cell wall calcium and higher levels of cell wal l nitrogen. This result was unexpected since firm texture in apples and oth er fruits has been commonly associated with elevated levels of fruit calciu m. The nitrogen-rich material in the sections used for EELS was insoluble i n acidified methanol? indicating that it represented a high-molecular-weigh t component in the cell wall. Furthermore, total tissue hydroxyproline leve ls were greatest in material with elevated cell wall nitrogen, suggesting e nhanced levels of wall structural proteins in the tissue. These data indica te a correlation between increased amounts of cell wall nitrogen and firm f ruit texture. The possible role of cell wall proteins in determining the te xtural properties of fruit tissue is discussed.