*** Global organic soils – organic matter quality, carbon and nitrogen stocks and fluxes *** Authors: Jaan Pärn, Ülo Mander Department of Geography, Institute of Ecology and Earth Sciences, University of Tartu Corresponding Author Jaan Pärn, john@ut.ee Department of Geography, University of Tartu Vanemuise 56 Tartu Estonia *** General Introduction *** We conducted a global survey of soil organic matter formation and transformation processes in organic soils, during the dry season (i.e. the annual water table minimum time of year including temperate and boreal summers) at each site between 2011 and 2022. We used 156 organic soil samples from the top 10 cm in 61 sites in our global organic soil database throughout the rainy tropical, temperate, and boreal climate zones. *** Purpose of campaign *** The main purpose of this campaign is to investigate water-extractable organic matter (WEOM) of soil to determine which ecosystem properties (e.g. temperature, water content, land use, plant carbon inputs, nutrients, respiration or methanogenesis rates) control the persistence of organic matter. We applied ultrahigh-resolution mass spectrometry to analyse molecular species and nominal oxidation state of carbon (NOSC) of WEOM. *** Methods of soil organic matter quality analysis *** The samples (n=156) were shipped to the Max-Planck-Institute for Biogeochemistry in Jena, Germany, to generate molecular-level information on WEOM. We employed Orbitrap ultra-high resolution mass spectrometry (UHR-MS) to analyse WEOM in a global collection of organic soil samples. UHR-MS provides insight into the molecular composition of SOM. Extracted fractions of WEOM highly depend on the extraction method. For instance, water-based, alkaline, organic solvent extraction, and mineral dissolution have been reported to extract a certain fraction of SOM selectively, thus influencing the molecular composition measure and complicating comparisons between WEOM fractions; however, trends of derived indices such as aromaticity have been deemed more robust across datasets. In addition, UHR-MS features ionization techniques that maximize or reduce specific peak intensities, depending on the nature of the analyte composition extracted, and potential interfering species. The chemical characteristics of extracted SOM, such as its acid-base properties, solubility (i.e., hydrophobicity), functional groups, molecular size, and extent of electron delocalization, can all impact the ionization efficiency. Electrospray ionization (ESI) in negative mode coupled with an ultrahigh resolution mass spectrometer (such as an Orbitrap) is commonly used to characterize the molecular composition of organic matter. Electrospray ionization is used in two modes: negative mode, which favor the deprotonation of acidic functional groups such as carboxylic acids and phenolic groups to form negative ions; positive mode, preferable to protonate the functional groups such as amines, alcohols, and carbonyls to form cations. Here, negative mode is employed because we study WEOM that is soluble by definition. Dissolution in water requires acidic, i.e., oxidized functional groups that are key drivers of solubility, hence explaining WEOM concentration, yield, and characteristics. The soil samples were shipped on dry ice to Jena, Germany on 18 February 2020 and stored at –20°C since. The samples were thawed at 4°C for several days, and kept at 4°C if not processed. To avoid unnecessary freeze-thaw cycles, samples were thawed sequentially depending on the time required for further processing steps. Before weighing in, samples were homogenized by manual shaking. Otherwise, a representative aliquot of sample was used (sampled with clean spatula). 7g (± 0.1 g) of material were transferred to an empty, cleaned 50 mL Falcon tube. After weighing in all samples of an extraction run (maximum 19 + Blank), 30 mL of ultrapure water were added to each Falcon tube and then shaken for 30 minutes on a shaker (ratio V/m = 4.3). Afterwards, samples were centrifuged for 5 minutes. Falcon tubes were carefully taken out of the centrifuge to avoid stirring up the sedimented material. Supernatant was carefully poured over an ultrapure-water-rinsed paper filter to filter out particles and isolate WEOM. Filter holders and extract bottles were cleaned before use (rinsed with ultrapure water and ultrapure water acidified to pH 2 by adding 1 mL of 32% HCl per L ultrapure water). WEOM extracts and remaining samples were then stored at –20°C. The WEOM samples were measured for dissolved organic carbon and total nitrogen at MPI-BGC according to established protocols​. The collected aqueous extracts were then filtered with a syringe filter (PTFE, 0.2 µm) to exclude colloidal and clay particles and adjusted to a concentration of 20 mg-C/L (20 ppm C) in 50% MeOH. Samples were then directly injected via a liquid chromatograph (100 µl volume) and measured according to established protocols. The dataset was first checked for general consistency. Numbers and fraction (number and intensity) of different formula classes were checked and showed an expectable decline in distribution of CHO, CHNO, CHOS and CHOP formulae, in line with general experience of DOM data, and organic soils. More “exotic” combinations of heteroatoms were in an acceptable range, indicating no bias in molecular formula assignment. Orbitrap identified 14,890 molecular formulae with masses ranging from 100 to 950 Daltons. *** Description of data *** MAT Mean annual temperature soil_temp Soil temperature at 10cm depth measure during the field campaign, centigrade LogMAP Logarithm of mean annual precipitation, mm SWC Volumetric soil water content, m3 water per m3 soil sum_i Sum of spectral intensity values n_forms Number of molecular formulae bp_i Base peak intensity value av_i Average spectral intensity Properties of individual formulae in the dataset. Samples are characterised in terms of these indices to yield proxies of organic matter composition. nC  Number of C atoms    nH  Number of H atoms    nO  Number of O atoms    nN  Number of N atoms    nS  Number of S atoms    nP  Number of P atoms  wa Weighted average   HC  Hydrogen to Carbon ratio  (Kim et al., 2003)?  OC  Oxygen to Carbon ratio  (Kim et al., 2003)?  DBE  Double bond equivalents (Koch & Dittmar, 2006, 2016)?  AImod  Modified aromaticity index (Koch & Dittmar, 2006, 2016)?  DBE-O  DBE minus O atoms  (Herzsprung et al., 2014)?  DBE-O/2  DBE minus half number of O atoms  (Gonsior et al., 2016)?  NOSC  Nominal oxidation state of Carbon (Boye et al., 2017)?  dG0_Cox  Gibbs free energy of oxidation half reaction  (LaRowe & Van Cappellen, 2011)?  CA  Condensed aromatics (Hawkes et al., 2020)?  AR  Aromatics***  HO  Highly saturated, Oxygen-rich LO  Highly saturated, Oxygen-poor AL  Aliphatics  LIP  Lipid-like (Minor et al., 2014)?  PROT  Protein-like AMIN  Aminosugar-like SUG  Carbohydrate-like  COND  Condensed hydrocarbon-like LIG  Lignin-like TAN  Tannin-like NA  Other formulae  GPP Gross primary production calculated from MOD17A2H 8-day 500 m grid V006 data developed from the MODIS sensor data onboard the Terra and Aqua satellites, mg C per square meter per h δVPDB (13C/12C) / ‰ Soil stable carbon isotope ratio compared to Vienna Pee Dee Belemnite (VPDB) standard, ‰ ER Field-measured ecosystem respiration, mg C per square meter per h CH4 Field-measured methane flux, µg C per square meter per h N2O Field-measured methane flux, µg C per square meter per h N2 Denitrification potential, measured from laboratory incubations of intact soil cores, µg N per square meter per h δVAIR (15N/14N) / ‰ Soil stable nitrogen isotope ratio compared to air standard OrgM Soil organic matter content, g organic matter per g soil CtoN Soil carbon-to-nitrogen ratio *** Licence *** - License: [CC-BY-NC-ND 4.0]