Optimized Demineralization Technique for the Measurement of Stable Isotope Ratios of Nonexchangeable H in Soil Organic Matter

To make use of the isotope ratio of nonexchangeable hydrogen (δ²H_{n (nonexchangeable)}) of bulk soil organic matter (SOM), the mineral matrix (containing structural water of clay minerals) must be separated from SOM and samples need to be analyzed after H isotope equilibration. We present a novel technique for demineralization of soil samples with HF and dilute HCl and recovery of the SOM fraction solubilized in the HF demineralization solution via solid-phase extraction. Compared with existing techniques, organic C (C_org) and organic N (N_org) recovery of demineralized SOM concentrates was significantly increased (C_org recovery using existing techniques vs new demineralization method: 58% vs 78%; N_org recovery: 60% vs 78%). Chemicals used for the demineralization treatment did not affect δ²H_n values as revealed by spiking with deuterated water. The new demineralization method minimized organic matter losses and thus artificial H isotope fractionation, opening up the opportunity to use δ²H_n analyses of SOM as a new tool in paleoclimatology or geospatial forensics

Data on plant stoichiometry from Jena Trait-based experiment

The data set contains the lemental composition of plants in the Jena trait-based experiment, including derived variables. The columns are explained in the read me file

Observed C:N versus predicted C:N ratios.

<p>For every measured C:N ratio (2, 4, 8 and 16 species mixtures) the corresponding calculated C:N ratio is shown across years (2003–2006). The green line gives the fit of an orthogonal regression (intercept 1.03 (SD 0.423); estimated C:N 0.999 (SD 0.043). The blue line gives a linear regression (Intercept 8.1448 (SD 1.434); estimated C:N 0.756 (SD 0.046)). For comparison, identity is given by a black line.</p

Comparison of the coefficient of variance of observed C:N and predicted C:N ratios.

<p>The Coefficient of Variation (CV) of measured (oCN) and calculated C:N (pCN) ratios is shown for different diversity levels (2, 4, 8, and 16 species mixtures) separated by years (2003–2006).</p

P:K ratio versus plant species richness.

<p>GAMLSS (generalized additive model for location scale and shape) model of the molar P:K ratio versus species richness of the years 2003–2007. Black line stands for the mean. For better illustration of the variance, percentiles of the standard deviation are given as grey lines. Sown div. = sown diversity, leg = legume.</p

C:P ratio versus plant species richness.

<p>GAMLSS (generalized additive model for location scale and shape) model of the molar C:P ratio versus species richness of the years 2003–2007. Black line stands for the mean. For better illustration of the variance, percentiles of the standard deviation are given as grey lines. Sown div. = sown diversity, leg = legume.</p

C:N ratio versus plant species richness.

<p>GAMLSS (generalized additive model for location scale and shape) model of the molar C:N ratio versus species richness (natural logarithm) of the years 2003–2007. Black line stands for the mean. For better illustration of the variance, percentiles of the standard deviation are given as grey lines. Sown div. = sown diversity, leg = legume.</p

MANOVA results on bivariate elemental ratios for the years 2003 to 2007.

<p>For each factor, the Pillai Trace value and its significance level are given as well as all ratios for which the factor effect was significant at p<0.05. Significance levels: p<0.001 = ***, p<0.01 = **, p<0.05 = *, p<0.1 = .</p

Results of ANOVAs on the effect of the experimental variables on microbial community.

<p>Impact of plant diversity (plant species richness (PSR, log transformed) and number plant functional groups (FG)) and presence of distinct plant functional groups (legumes, grasses, small herbs, tall herbs) on total microbial biomass (MicBM), Gram-positive (Gram+), Gram-negative bacteria (Gram−), fungal biomass (Fungi) and the composition of the soil community, characterized by the fungal-to-bacterial biomass ration (F:B ratio). Numbers in bold display p values < 0.05 and numbers in italic display p values <0.1.</p

Impact of land use and plant diversity soil microbial community.

<p>Differences (<i>P</i><0.05) between experimental plots and control sites (arable land and semi-natural meadows) were analyzed with Tukey's HSD test and indicated by letters. Differences between bare ground vs. vegetated plots and significant effects of plant diversity were tested with ANOVA (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0096182#pone-0096182-t001" target="_blank">Table 1</a>) and indicated by asterisks (<i>P</i><0.05). Figures show effect of plant species richness on (a) total microbial biomass (b) fungal biomass and the fungal-to-bacterial biomass ratio and (c) number plant functional groups effect on fungal biomass and the fungal-to-bacterial biomass ratio.</p