All laboratory analyses carried out in this study are summarized in Fig. S1.3. Fresh leaf litter subsamples were subjected to the measurement of microbial extracellular enzyme activities. Samples were weighed corresponding to 50–100 mg dry weight and cut into small pieces before they were homogenized with a tissue-tearer homogenizer (Dremel 395 MultiPro) in 10 mL of 0.10 M Na–K phosphate buffer on ice. Enzyme extraction was performed at (24 ± 2)  °C for 1 h in a rotary mixer (Elmi Intelli-Mixer RM-2L) at 90 rpm. The homogenates were centrifuged for 5 min, with 4000 g at 4 °C. The obtained supernatants were directly analysed or kept refrigerated (4 °C) until analysis within a few days. The enzymatic activity assays were analysed by UV–Vis spectrophotometer (Shimadzu UV-1800).

Phenol oxidase activity was determined through the spectrophotometric determination of 4-(N-proline)-o-benzoquinone resulting from enzymatic oxidation of catechol as the substrate in the presence of L-proline (adopted from Zimmer 2005b; Perucci et al. 2000). Upon mixing of 200 µL sample supernatant, 900 µL 0.05 M L-proline and 900 µL 0.05 M pyrocatechol, absorbance changes (ΔA) at 520 nm was recorded at 1-min intervals over 10 min. Relative catechol oxidation was determined as the slope of ΔA produced by linear regression analysis. Phenol oxidase activity was expressed as relative phenol oxidation capacity (RAU g−1 h−1).

Cellulase activity was quantified according to Zimmer (2005a), Skambracks and Zimmer (1998) and Linkins et al. (1990) with some modifications. A mix of 90 mg α-cellulose, 900 µL supernatant solution and 900 µL citrate–phosphate buffer (with 0.05% NaN3) was incubated in a rotary mixer for 18 to 24 h and then centrifuged for 15 min at 1000 g at room temperature. Blank solutions were prepared by mixing 700 µL supernatant with 300 µL citrate–phosphate buffer and measured at 340 nm to obtain absorbance at time zero (A0). For samples, 700 µL supernatant was mixed with 300 µL glucose (HK) assay kit solution (Sigma-Aldrich), incubated for 15 min at room temperature, and then the absorbance was measured (A15) at 340 nm. The final absorbance was calculated by subtracting A0 from A15 (ΔA).

Protease activity was determined through enzymatic decay of azocasein as a chromogenic protein substrate (after de Menezes et al. 2014; Charney and Tomarelli 1947). A mix of 500 µL supernatant solution and 500 µL 1% azocasein solution was incubated at room temperature for 1 h in a rotary mixer at 90 rpm. The proteolytic reaction was stopped by adding 500 µL 20% TCA to precipitate the remaining substrate. Then, the mixture was kept on ice for 10 min and centrifuged at 1000 g for 15 min at room temperature. For the blank solution, 20% TCA was added before the addition of sample supernatant. Both sample and blank solutions were alkalinized with an equal volume of 2 M NaOH solution prior to the photometric measurement at 440 nm. As the proteolysis of azocasein results in a mix of peptides and amino acids, the final result was expressed as relative protease activity (RAU g−1 h−1).

For leaf chemistry analyses –total carbon content, total nitrogen content, total phenolic content, protein-precipitation capacity and phenolic fingerprint– samples were shipped to the Leibniz Centre for Tropical Marine Research (ZMT), Bremen, Germany.

Approximately 1 to 2 g sub samples of fine-ground leaf-litter were weighed and encapsulated in 10 × 10 mm tin capsules for the measurement of carbon and nitrogen contents (Eurovector EA3000 Elemental Analyzer). Birch leaf standard (BLS) was used as standard. Another 50 mg of the samples were extracted with 2 mL of 70% ethanol at room temperature for 1 h with a rotary mixer at 90 rpm, and then the supernatants were separated from the precipitates by centrifugation at 10,000 g, 4 °C for 10 min.

The total phenolic content was determined following the Folin-Ciocalteu assay (Bärlocher and Graça 2005; Ainsworth and Gillespie 2007). Aliquots of 10 to 200 µL of the supernatants were diluted to 500 µL with double-distilled water. Then, 250 µL 1 N Folin-Ciocalteu reagent and 1.25 mL 700 mM Na2CO3 were added sequentially and the final solutions were vortexed. Blanks contained 500 µL double-distilled water but no sample. The mixture was kept in the dark for 1–2 h, before the absorbance was read at 750 nm using a microplate reader (TECAN Infinite M200 Pro). Tannic acid served as standard, and results are expressed as mg tannic acid equivalents (TAE) g−1 litter.

The protein-precipitation capacity of the leaf litter phenolics was quantified through a radial diffusion assay, using bovine serum albumin (0.1% in ascorbic acid-containing agarose) as protein (Graça and Bärlocher 2005; Hagerman 1987). Four consecutive 9 μL aliquots from each supernatant were added to a single 2 mm well in the agarose gel plate. Each plate contained three tannic acid standards of 0.09, 0.18 and 0.27 mg mL−1. The gel plates were incubated for 3 to 4 days at room temperature. Then, the average of two perpendicular diameters of protein precipitation rings from both standards and samples was determined using a stereomicroscope. The ring area was calculated after subtracting the diameter of the wells. Protein-precipitation capacity is expressed in TAE g−1 sample dry weight.

As the Folin-Ciocalteu and the radial diffusion assays provide different proxies for the phenolic signature of a sample (Zimmer et al. 2015), a phenolic fingerprint was additionally depicted through HPLC analysis of litter sample extracts. The supernatants of these extracts were dried in a multi-evaporator system (Heidolph Synthesis 1) for approximately 1 h, 1000 rpm, 60 °C. The pressure was set to 175 mbar for 30 min to evaporate ethanol and changed to 60 mbar to evaporate water. The dried extracts were dissolved with 2 mL acetonitrile mix reagent (23% acetonitrile:76% water:1% acetic acid v/v) in an ultrasonic bath for 10 min and filtered through 0.45 µm PTFE filters. The sample solutions were placed in 1.0 mL glass vials and kept frozen at -24o C until analysis. Individual phenolic fingerprints of litter samples were obtained through reverse-phase high-performance liquid chromatography (Agilent 1260 Infinity), with a Luna 5 µm C18(2) non-polar column (100 Å, 250 × 4.6 mm) and detected at 280 nm with a Diode Array Detector. Flow-rate was kept at 1.25 mL/min, and temperature was set to 20 °C. From each sample, 20 µL was injected. The elution condition of the binary pump was: 0–11 min, 100% A (isocratic); 11–27.5 min, 75% A (linear gradient); 27.5–28.5 min, 0% A (linear gradient; 28.5–31 min, 0% A (isocratic); 31–32 min, 100% A (linear gradient); 32–35 min, 100% A (isocratic), with solvent A: 18% acetonitrile:0.5% acetic acid:81.5% water; solvent B: 100% acetonitrile. Gallic acid, tannic acid and sinapyl alcohol, a monomer of many lignins, served as standards for the identification and quantification of some of the resulting peaks.

Data and statistical analyses were conducted using the statistical package R version 3.4.4 (R Core Team 2018) and PAST software version 3.25 (Hammer et al. 2001). Normality of data was tested through Shapiro–Wilk’s tests and data were square root-transformed prior to the statistical analysis where necessary. The effects of ontogenetic and interspecific differences on leaf decay, leaf chemistry and extracellular enzyme activity over time were analysed through repeated measures analysis of variance (RM ANOVA) with fixed variations of species (4 levels), maturity stages (2 levels) and time intervals (4 levels). Bonferroni tests were used for post-hoc analyses (α = 0.05). Correlation coefficient matrices were created to determine the relationship of the leaf litter decay rate, represented as leaf half-life (t0.50), with initial leaf chemistry and extracellular enzymatic activity variables.

Differences in phenolic fingerprints (HPLC analysis) were visualized through non-metric multidimensional scaling (NMDS) with Morisita-Horn similarity index (Magurran 2004). Non-parametric permutational multivariate ANOVA (PERMANOVA) with 9999 permutations was utilized to test for significance of interspecies and ontogenetic differences, based on the selected distance measure (Anderson 2001). Similarity Percentage (SIMPER) analysis was used for assessing which HPLC peaks (= phenolic compounds) were primarily responsible for an observed difference among groups of samples (Clarke 1993).

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