Mid Sweden University

In this work the distribution of free and weekly adsorbed aromatic acids (phthalic acid and ten phenolicacids; gallic, p-hydroxybenzoic, salicylic, vanillic, protocatechuic, p-coumaric, syringic, sinapic, ferulic and caffeicacid), which could participate in weathering and soil formation processes, were studied for O, E and Bhorizons in a podzol soil in central Sweden. For the analysis a simple and rapid quantitative and qualitative liquidchromatography–tandem mass spectrometry method (using gradient elution) was developed with LODs rangingfrom 5 to 25 nM. Different soil solution sampling techniques (tension-lysimeter and soil centrifugation) and soilextraction with either 10 mM phosphate buffer (pH 7.2) or 50:50 (v/v) 10 mM phosphate buffer:MeOHwere compared. All eleven acids were found in detectable or trace concentrations. The most abundant aromaticacids were vanillic and phthalic acid with concentrations around 1 μM for O and E horizon respectively.Lysimeter samples resulted in the lowest concentrations followed by centrifugation samples. Ingeneral, buffer:MeOH extraction resulted in the highest concentrations for the O horizon, likely due toMeOH's ability to compete for hydrophobic sites on soil organic matter (SOM). Then again, pure bufferwith its higher ion strength, interfering with the acids electrostatic interactions with clay particles, leads tohigher extracted concentrations for the E and B horizons. Since the efficiency of the extraction solutions, to alarge extent, depends on the sample properties, a general approach is hard to appoint. However, the extractionof substituted cinnamic acids is in general facilitated by adding MeOH to the extraction solution. The use of statisticalmethods for the evaluation of the results showed a large and significant difference in aromatic acid concentrationsreceived using different sampling techniques and sample preparations. In fact, sampling methodsresulted in higher variations in aromatic acid concentrations than sampled horizon.

Samples of apatite, biotite and oligoclase were incubated in different soil horizons of a boreal forest podzol in central Sweden. After four years, soil from the bulk and the surfaces of the minerals were extracted and analyzed in regard to fungal biomass, enzymatic activity, low molecular mass organic acids and hydroxamate siderophores.  The nutrient status of the study area was evaluated using historical data from foliar and soil solution analysis. Soil solution indicated a possible shortage of P in the mineral soil even though foliar content was above target levels. The foliar analysis instead revealed possible shortage of Fe. Apatite is the primary source of inorganic P in soil and biotite is rich in Fe compared to the soil at the site, and it was hypothesized that both minerals could be attractive sources of weatherable mineral nutrients, hence associated with higher microbial activity. Parameters measured as indicators of microbial activity revealed few significant (p<0.05) differences between the minerals and bulk soil in the O horizon, probably due to large variations among replicates, although an overall higher microbial activity was found in the bulk soil. In the E horizon a distinct trend appeared indicating higher levels of low molecular mass organic acids, enzymatic activity and fungal biomass associated with the biotite mineral, although this observation was only found to be significant for some of the parameters when analyzed individually. No clear trends were observed in the B horizon.

This method employs liquid chromatography-tandem mass spectrometry to rapidly quantify chitin-derived glucosamine for estimating fungal biomass. Analyte retention was achieved using hydrophilic interaction liquid chromatography, with a zwitter-ionic stationary phase (ZIC-HILIC), and isocratic elution using 60 % 5 mM ammonium formate buffer (pH 3.0) and 40 % ACN. Inclusion of muramic acid, and its chromatographic separation from glucosamine, enabled calculation of the bacterial contribution to the latter. Galactosamine, an isobaric isomer to glucosamine, found in significant amounts in soil samples, was also investigated. The two isomers form the same precursor and product ions, and could not be chromatographically separated using this rapid method. Instead, glucosamine and galactosamine were distinguished mathematically, using the linear relationships describing the differences in product ion intensities for the two analytes. The m/z transitions of 180→72 and 180→84 were applied for the detection of glucosamine and galactosamine and that of 252→126 for muramic acid. Limits of detection were in the pico-molar range for all included analytes. The total analysis time was 6 min, providing a high sample through-put method.

A liquid chromatography-electrospray ionization-mass spectrometry method was developed to quantitatively and qualitatively analyze thirteen hydroxamate siderophores (ferrichrome, ferrirubin, ferrirhodin, ferrichrysin, ferricrocin, ferrioxamine B, D₁, E and G, neocoprogen I and II, coprogen and triacetylfusarinine C). Samples were pre-concentrated on-line via switch-valve setup prior to analyte separation on a Kinetex C18 column. Gradient elution was performed using a mixture of an ammonium formate buffer and acetonitrile. Total analysis time including column conditioning was 20.5 minutes. Analyte were fragmented by applying collision induced dissociation, enabling structural identification via tandem mass spectrometry. Limit of detection values for the selected ion monitoring method ranged from 71 pM to 1.5 nM with corresponding values of two to nine times higher for the multiple reaction monitoring method. The liquid chromatography-mass spectrometry method resulted in a robust and sensitive quantification of hydroxamate siderophores as indicated by retention time stability, linearity, sensitivity, precision and recovery. The analytical error of the methods, assessed trough random-order, duplicate analysis of soil samples extracted with a mixture of 10 mM phosphate buffer and methanol, appears negligible in relation to between-sample variations.