Preparation of leaf litter leachates
We collected the leaf litter of 6 broadleaf and 5 coniferous species
commonly found in the plains and mountainous of northeastern Japan.
Although we did not check when they fell, we collected leaf litter on
top of the leaf litter layer on the
ground so that the litter did not have direct contact with the soil.
These fallen leaves were collected at Zao (N38.122, E140.451) on October
3, 2020, at Kawatabi (N38.745, E140.757) on October 15, 2020, and at
Aobayama (N38.259, E140.837) on November 19, 2020 in Miyagi Prefecture,
Japan (Table 1).
Before use, leaf litter was dried at room temperature (20 °C) and was
cut with scissors to mimic the early decomposition processes by
detritivores. For each tree species, we weighed the cut leaf litter and
placed it in two 1000 mL Nalgene bottles containing 800 ml of distilled
water. The weight of leaf litter in each bottle was adjusted to control
the final concentration of TDP to approximately 1-4 µM after 7 days of
the leaching process based on the release efficiency (Table 1) estimated
by Ho et al. (2023). The water in the two replicate bottles was aerated
in the dark for 7 days at 20 °C and mixed before being used as leaf
litter leachates.
We used Scenedesmus obliquusas the experimental algal species because it has been frequently used in
various studies . Prior to the incubation of Scenedesmus , the
leaf litter leachate was filtered through 40 µm mesh to remove litter
particles. DOC and total dissolved N (TDN) concentrations in the leaf
litter leachate were measured using a TOC/TNb analyzer (multi N/C 3100,
Analytik Jena GmbH, Jena, Germany). Total dissolved P (TDP)
concentrations were measured using the ascorbic acid-molybdenum blue
method after persulfate oxidation .