Biogeochemistry of marine dissolved organic matter: molecular composition, reactivity and new methods

Dissolved organic matter (DOM) is an ultimate chemical product of all life on earth. It integrates energy, carbon dioxide and nutrients into a vast compositional and structural variety of molecules further modified by biological, chemical, and physical processes. In the ocean, organic matter production depends mainly on the photosynthetic activity of autotrophs and most of it is immediately consumed and respired by heterotrophs. Some of this fresh organic matter, however, escapes immediate turnover and accumulates in dissolved form in the entire water column. During isopycnal transport and seasonal convective overturn, microbial, photochemical, and physical processes remove most of the fresh DOM. The remaining organic matter is an old, chemically poorly characterized heterogeneous mixture of small, partially oxidized and unsaturated molecules: refractory DOM. The main topic of this thesis is the chemical characterization of DOM: elemental composition and reactivity with regard to environmental boundary conditions as well as causalities of persistence. All studies involved substantial chemical and physical gradients of temperature, pressure, salinity, irradiation, biological communities, and nutrients. These gradients allowed for testing the main research hypotheses with different end members to obtain functional relationships between the physico-chemical variables and the observed properties of DOM. Different methods were applied to achieve these aims. High resolution inorganic and organic mass spectrometry, chromatography, statistical analysis and modeling were performed on samples obtained from oceanic research cruises. Additional seasonal surveys in an estuarine system and experimental setups addressed the influence of the various physico-chemical boundary conditions on the chemical composition and phase distribution of DOM. The most comprehensive study of this work included more than 200 samples from the tropical to the polar open ocean and from the surface to the seafloor and represents the so far largest consolidated dataset for ultrahigh resolution organic mass spectrometry in the ocean. A method was established that enabled for the first time separation and quantification of organic phosphorus and sulfur in marine DOM in a coupled chromatography mass spectrometry system. It was shown that the compositional diversity of DOM, i.e., the contributions from the heteroatoms phosphorus and sulfur, was reflected in the chemical properties of the molecules as revealed by polarity separation. Further, the method was shown to be applicable for determining metal ions that are also part of the chemical entity of DOM. However, not all investigated metal ions showed a strong and selective affinity for organic matter, e.g., uranium. A rare isotope of uranium, 236U, determined for the first time in an oceanic depth profile, was demonstrated to be a suitable transient tracer in oceanographic studies, reflecting an anthropogenic marker for water mass circulation. Very different compounds, surface active sulfonic acids, were identified as part of the total DOM pool in a sea surface microlayer study. Although sulfonic acids are widely known as potential contaminants in surface waters, this study demonstrated the analytical capability of ultrahigh resolution organic mass spectrometry and fragmentation to study thousands of DOM molecules and their responses to changing physico-chemical conditions, e.g., the ionic strength of the aqueous phase. An even deeper insight into the composition and long-term transformation of DOM was achieved by comparing the molecular signatures of DOM samples from the East Atlantic and Southern Ocean. Using statistical tools, it was demonstrated that distinct patterns of mass peak magnitude changes could be related to the consecutive ageing of this mixture of molecules. A modeling of the degradation rates of individual DOM molecules demonstrated that the chemical composition of the bulk DOM changes with age towards a proposed island of stability . The broad distribution of these degradation rates is proposed as an extension of the contemporary perception of marine DOM cycling and reworking. Bringing together inorganic and organic biogeochemistry as well as (molecular) microbiology to study the complex biogeochemical interactions in the ocean will be an important future research direction in marine sciences. The combined efforts from multidisciplinary research groups are a prerequisite to resolve the unanswered questions on the response of the microbial communities, the fate of anthropogenic carbon dioxide, the chemical processes and equilibria in the ocean, and its crucial feedback mechanisms in a changing climate

Impacts of litter decay on organic leachate composition and reactivity

Litter decomposition produces labile and recalcitrant forms of dissolved organic matter (DOM) that significantly affect soil carbon (C) sequestration. Chemical analysis of this DOM can provide important knowledge for understanding soil DOM dynamics, but detailed molecular analyses on litter derived DOM are scarce. Here we use ultrahigh resolution mass spectrometry (FT-ICR MS) to characterize the molecular composition of DOM from fresh and progressively decomposed litter samples. We compared high reactive (HR) and low reactive (LR) litter sources with regard to changes in the chemistry and bioavailability of leachates throughout the early phase of litter decay. We show that litter reactivity is a driver of chemical changes in the leached DOM of litter species. Birch, alder and Vaccinium (i.e. HR) litter initially produced more DOM with a higher lability than that of spruce, pine and wood (i.e. LR) litter. Labile oxidized phenolic compounds were abundant in leachates produced during the initial HR litter decay stages, indicating litter lignin degradation. However, the similarity in chemistry between HR and LR leachates increased during the litter decay process as highly leachable structures in HR litter were depleted. In contrast, chemistry of leachates from LR litter changed little during the litter decay process. The oxygenated phenolic compounds from HR litter were driving the lability of HR leachates and the changes in relative abundance of molecules during DOM incubation. This appeared to result in the creation of stable aliphatic secondary microbial compounds. In LR leachates, lability was driven by labile aliphatic compounds, while more resistant phenolic compounds were associated with recalcitrance. These results show how DOM dynamics follow different paths depending on litter reactivity, which has important implications for soil biogeochemistry and C sequestration

Direct Imaging of Plant Metabolites in the Rhizosphere Using Laser Desorption Ionization Ultra-High Resolution Mass Spectrometry

The interplay of rhizosphere components such as root exudates, microbes, and minerals results in small-scale gradients of organic molecules in the soil around roots. The current methods for the direct chemical imaging of plant metabolites in the rhizosphere often lack molecular information or require labeling with fluorescent tags or isotopes. Here, we present a novel workflow using laser desorption ionization (LDI) combined with mass spectrometric imaging (MSI) to directly analyze plant metabolites in a complex soil matrix. Undisturbed samples of the roots and the surrounding soil of Zea mays L. plants from either field- or laboratory-scale experiments were embedded and cryosectioned to 100 mm thin sections. The target metabolites were detected with a spatial resolution of 25 mm in the root and the surrounding soil based on accurate masses using ultra-high mass resolution laser desorption ionization Fourier-transform ion cyclotron resonance mass spectrometry (LDI-FT-ICR-MS). Using this workflow, we could determine the rhizosphere gradients of a dihexose (e.g., sucrose) and other plant metabolites (e.g., coumaric acid, vanillic acid). The molecular gradients for the dihexose showed a high abundance of this metabolite in the root and a strong depletion of the signal intensity within 150 mm from the root surface. Analyzing several sections from the same undisturbed soil sample allowed us to follow molecular gradients along the root axis. Benefiting from the ultra-high mass resolution, isotopologues of the dihexose could be readily resolved to enable the detection of stable isotope labels on the compound level. Overall, the direct molecular imaging via LDI-FT-ICR-MS allows for the first time a nontargeted or targeted analysis of plant metabolites in undisturbed soil samples, paving the way to study the turnover of root-derived organic carbon in the rhizosphere with high chemical and spatial resolution

Iron Exports From Catchments Are Constrained by Redox Status and Topography

Fe(III) hydroxides stabilize organic carbon (OC) and P in soils. Observations of rising stream Fe concentrations are controversially posited to result from a flushing of iron-rich deeper soil layers or a decrease of competing electron acceptors inhibiting Fe reduction (NO3- and SO42-). Here, we argue that catchment topography constrains the release of Fe, OC, and P to streams. We therefore incubated organic topsoil and mineral subsoil and modified the availability of NO3-. We found that Fe leaching was highest in topsoil. Fe, OC, and P released at quantities proportional to their ratios in the source soil. Supply of NO3- reduced Fe leaching to 18% and increased pore water OC:Fe and P:Fe ratios. Subsoil, however, was an insignificant Fe source (<0.5%). Here, the leached quantities of Fe, OC and P were highly disproportionate to the soil source with an excess of released OC and P. We tested if experimental findings scale up using data from 88 German catchments representing gradients in NO3- concentration and topography. Average stream Fe concentrations increased with decreasing NO3- and were high in catchments with shallow topography where high groundwater levels support reductive processes and topsoils are hydrologically connected to streams; but Fe concentrations were low in catchments with steep topography where flow occurs primarily through subsoils. OC:Fe and P:Fe ratios in the streams similarly varied by NO3- and topography. This corroborates the findings from the laboratory experiment and suggests that catchment topography and competing electron acceptors constrain the formation of Fe-reducing conditions and control the release of Fe, OC, and P to streams. © 2022. The Authors

New Insights into the Seasonal Variation of DOM Quality of a Humic-Rich Drinking-Water Reservoir—Coupling 2D-Fluorescence and FTICR MS Measurements

Long-term changes in dissolved organic matter (DOM) quality, especially in humic-rich raw waters, may lead to intensive adaptions in drinking-water processing. However, seasonal DOM quality changes in standing waters are poorly understood. To fill this gap, the DOM quality of a German drinking water reservoir was investigated on a monthly basis by Fourier-transform ion cyclotron resonance mass spectrometry (FTICR MS) measurements and 2D fluorescence for 18 months. FTICR MS results showed seasonal changes of molecular formula (MF) intensities, indicating photochemical transformation of DOM as a significant process for DOM quality variation. For an assessment of the two humic-like components, identified by parallel factor analysis (PARAFAC) of excitation–emission matrices (EEM), their loadings were Spearman’s rank-correlated with the intensities of the FTICR MS-derived MF. One of the two PARAFAC components correlated to oxygenrich and relatively unsaturated MF identified as easily photo-degradable, also known as coagulants in flocculation processes. The other PARAFAC component showed opposite seasonal fluctuations and correlated with more saturated MF identified as photo-products with some of them being potential precursors of disinfection byproducts. Our study indicated the importance of elucidating both the chemical background and seasonal behavior of DOM if raw water-quality control is implemented by bulk optical parameters

Multi-Class Cancer Subtyping in Salivary Gland Carcinomas with MALDI Imaging and Deep Learning

Simple Summary The correct diagnosis of different salivary gland carcinomas is important for a prognosis. This diagnosis is imprecise if it is based only on clinical symptoms and histological methods. Mass spectrometry imaging can provide information about the molecular composition of sample tissues. Using a deep-learning method, we analyzed the mass spectrometry imaging data of 25 patients. Using this workflow we could accurately predict the tumor type in each patient sample. Abstract Salivary gland carcinomas (SGC) are a heterogeneous group of tumors. The prognosis varies strongly according to its type, and even the distinction between benign and malign tumor is challenging. Adenoid cystic carcinoma (AdCy) is one subgroup of SGCs that is prone to late metastasis. This makes accurate tumor subtyping an important task. Matrix-assisted laser desorption/ionization (MALDI) imaging is a label-free technique capable of providing spatially resolved information about the abundance of biomolecules according to their mass-to-charge ratio. We analyzed tissue micro arrays (TMAs) of 25 patients (including six different SGC subtypes and a healthy control group of six patients) with high mass resolution MALDI imaging using a 12-Tesla magnetic resonance mass spectrometer. The high mass resolution allowed us to accurately detect single masses, with strong contributions to each class prediction. To address the added complexity created by the high mass resolution and multiple classes, we propose a deep-learning model. We showed that our deep-learning model provides a per-class classification accuracy of greater than 80% with little preprocessing. Based on this classification, we employed methods of explainable artificial intelligence (AI) to gain further insights into the spectrometric features of AdCys

Dissolved organic matter in continental hydro-geothermal systems: insights from two hot springs of the East African Rift valley

Little is known about the quantity and quality of dissolved organic matter (DOM) in waters from continental geothermal systems, with only a few reports available from the Yellowstone US National Park. In this study, we explored the chemodiversity of DOM in water samples collected from two geothermal hot springs from the Kenyan East African Rift Valley, a region extremely rich in fumaroles, geysers, and spouting springs, located in close proximity to volcanic lakes. The DOM characterization included in-depth assessments performed by negative electrospray ionization Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR-MS). Reduced, saturated and little aromatic DOM compounds were dominant in the hot spring waters collected from either the Ol Njorowa gorge (ON) or the south shore of the soda-saline Lake Elementaita (ELM). Oxygen-poor and sulfur-bearing DOM molecules prevailed in ON, probably reflecting abiotic sulfurization from sulfide-rich geofluids. Nitrogen-bearing aliphatic and protein-like molecules were abundant in ELM, possibly perfusing through the organic-rich sediments of the adjacent Lake Elementaita. Notably, the heat-altered DOM of ancient autochthonous derivation could represent an overlooked source of aliphatic organic carbon for connected lentic environments, with a potential direct impact on nutrient cycling in lakes that receive geothermal water inputs

Molecular transformation and degradation of refractory dissolved organic matter in the Atlantic and Southern Ocean

More than 90% of the global ocean dissolved organic carbon (DOC) is refractory, has an average age of 4,000–6,000 years and a lifespan from months to millennia. The fraction of dissolved organic matter (DOM) that is resistant to degradation is a long-term buffer in the global carbon cycle but its chemical composition, structure, and biochemical formation and degradation mechanisms are still unresolved. We have compiled the most comprehensive molecular data set of 197 Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) analyses from solid-phase extracted marine DOM covering two major oceans, the Atlantic sector of the Southern Ocean and the East Atlantic Ocean (ranging from 50° N to 70° S). Molecular trends and radiocarbon dating of 34 DOM samples (comprising Δ14C values from -229 to -495‰) were combined to model an integrated degradation rate for bulk DOC resulting in a predicted age of >24 ka for the most persistent DOM fraction. First order kinetic degradation rates for 1,557 mass peaks indicate that numerous DOM molecules cycle on timescales much longer than the turnover of the bulk DOC pool (estimated residence times of >100 ka) and the range of validity of radiocarbon dating. Changes in elemental composition were determined by assigning molecular formulae to the detected mass peaks. The combination of residence times with molecular information enabled modelling of the average elemental composition of the slowest degrading fraction of the DOM pool. In our dataset, a group of 361 molecular formulae represented the most stable composition in the oceanic environment (“island of stability”). These most persistent compounds encompass only a narrow range of the elemental ratios H/C (average of 1.17 ± 0.13), and O/C (average of 0.52 ± 0.10) and molecular masses (360 ± 28 and 497 ± 51 Da). In the Weddell Sea DOC concentrations in the surface waters were low (46.3 ± 3.3 μM) while the organic radiocarbon was significantly more depleted than that of the East Atlantic, indicating average surface water DOM ages of 4,920 ± 180 a. These results are in accordance with a highly degraded DOM in the Weddell Sea surface water as also shown by the molecular degradation index IDEG obtained from FT-ICR MS data. Further, we identified 339 molecular formulae which probably contribute to an increased DOC concentration in the Southern Ocean and potentially reflect an accumulation or enhanced sequestration of refractory DOC in the Weddell Sea. These results will contribute to a better understanding of the persistent nature of marine DOM and its role as an oceanic carbon buffer in a changing climate