Evidence for Absence of Diurnal Variation of Chlorophyll Content in Mature Leaves of Soybean

Abstract

The fluctuation of the amount of chlorophyll about a mean daily value, i.e. diurnal variation of the chlorophyll content, in the leaves of higher plants has been studied by many investigators. However, it has become a matter of doubt because of recognition that the evidence is based on inadequate techniques. For example, Seybold and Falk (11) observed no diurnal change in chlorophyll content in mature leaves and suggested that the changes reported by other workers were reflections of the inherent variability in the methods of analysis employed. By applying analytical techniques similar to those of Seybold and Falk, Bauer (2) concluded that there was no daily change in the chlorophyll content of mature leaves of several species of plants, but that some variation, in addition to overall increase in chlorophyll content, occurred in young leaves. However, he restricted his sampling to a single leaf-or at most three or four leaves-in each experiment. Presumably aware of these arguments, other investigators (3, 8) have more recently asserted that diurnal variations in chlorophyll content do exist. We have confined our investigations to analyses of the chlorophyll contents in nonexpanding leaves of a relatively large population of soybean plants. The chlorophylls of leaf samples taken at 2-hour intervals were analyzed spectrophotometrically as their pheophytins in the extract solution, thus avoiding the analytical error introduced by transfer of the pigments into ether, a procedure which is utilized in methods employed by recent investigators (2, 11). Two preliminary experiments, involving six observations at each sampling time, indicated that there may indeed be a diurnal variation of the chlorophyll content in such leaves. However, the combined sampling and analytical error were of nearly the same magnitude as the apparent variation. Therefore, an experiment was designed whereby the number of observations was increased sixfold to decrease the level of the error arising from inherent variability in the plant material. Methods & Materials