Characterization of aggregate-stabilized dissolved organic matter release - A novel approach to determine soil health advances of conservation farming systems

Since principles of conservation agriculture mimic the soil conditions of undisturbed natural soils, linking aggregation and dissolved organic matter (DOM) occlusion would therefore provide a targeted descriptor for soil health advances of innovative farming systems. This study aimed to assess structure-related DOM patterns of conservation farming systems and underlying bio-chemical drivers by using a novel method for the combined analysis of aggregate breakdown and DOM release. Soil samples were collected from conventional farming, conservation farming and natural reference soil systems over a wide range of soil types. Ultrasonication aggregate breakdown combined with continuous UV–Vis measurement was used to characterize DOM release from soil. Measures of breakdown dynamics were related to soil physical and chemical properties to determine the strongest predictors of DOM release. The quantity of DOM released and aggregate stabilization showed a steady continuum starting from standard farming through conservation agriculture towards reference soil systems. DOM released from reference soils however was less complex and occluded in more stable soil aggregates than arable soils. The overall DOM release dynamics are shaped by agricultural management with site-specific modifiers driving aggregation and mineral-organic interactions in soils. The simultaneous quantification of aggregate breakdown and DOM release captures key biophysical effects in structure-related DOM stabilization and revealed significant differences between land-use and agricultural management systems. The linkage of physical with functional soil organic matter descriptors provides an improved approach to monitor soil health advances in arable cropping systems.

Methods Soil samples were collected from conventional farming, conservation farming and natural reference soil systems over a wide range of soil types. Ultrasonication aggregate breakdown combined with continuous UV-Vis measurement was used to characterize DOM release from soil. Measures of breakdown dynamics were related to soil physical and chemical properties to determine the strongest predictors of DOM release. Results The quantity of DOM released and aggregate stabilization showed a steady continuum starting from standard farming through conservation agriculture towards reference soil systems. DOM released from reference soils however was less complex and occluded in more stable soil aggregates than arable soils. The overall DOM release dynamics are shaped by agricultural management with site-specific modifiers driving aggregation and mineral-organic interactions in soils. Conclusions The simultaneous quantification of aggregate breakdown and DOM release captures key biophysical effects in structure-related DOM stabilization and revealed significant differences between land-use and agricultural management systems. The linkage of physical with functional soil organic matter descriptors provides an improved approach to monitor soil health advances in arable cropping systems.

Abstract
Purpose Since principles of conservation agriculture mimic the soil conditions of undisturbed natural soils, linking aggregation and dissolved organic matter (DOM) occlusion would therefore provide a targeted descriptor for soil health advances of innovative farming systems. This study aimed to assess structure-related DOM patterns of conservation farming systems and underlying bio-chemical drivers by using a novel method for the combined analysis of aggregate breakdown and DOM release.

Introduction
Land management and climate change play a crucial role in the sequestration and loss of soil organic carbon (SOC) stocks (Sanderman et al. 2017). Appropriate soil management such as conservation tillage, cover cropping, and crop diversification can enhance SOC sequestration by either increasing soil organic matter (SOM) inputs to soil and/or decreasing its losses (Lal 2004(Lal , 2013Conceição et al. 2013;Poeplau and Don 2015;Minasny et al. 2017;Chenu et al. 2019).
The occlusion of SOC in soil aggregates is currently considered as a major mechanism for sequestering organic C in soil and responsive to soil management (Grandy and Neff 2008;Lehmann and Kleber 2015;Wiesmeier et al. 2019). In light of this, SOC stabilization in soil aggregates via physical protection is a vital early indicator of SOC changes and hence constitutes an opportunity to identify the response of agricultural practices to SOC sequestration (Chenu et al. 2019).
Among the various organic fractions in soil, dissolved organic matter (DOM) is one of the most active fractions in soil despite representing only approximately 2% of the total SOM pool (Bolan et al. 2011;Lavallee et al. 2019;Basile-Doelsch et al. 2020).This fraction plays a vital role in biogeochemical processes and mediates some of the most prominent environmental nutrient pathways (Van Hees et al. 2005;Kaiser and Kalbitz 2012;Lehmann and Kleber 2015;Basile-Doelsch et al. 2020;Gmach et al. 2020). Since approximately 67% of DOM is constituted by C and due to its high bioavailability (Neff and Asner 2001;Bolan et al. 2011;Malik and Gleixner 2013;Li et al. 2021), DOM is considered as a primary source of C for soil microorganisms. It has a significant role in SOC build up via microbial C assimilation and the subsequent accumulation of microbial necromass (Sae-Tun et al. 2022), which is a major contributor to SOC in agricultural soils in temperate regions (Liang et al. 2019). Even though DOM represents only a small proportion of SOM in soil, bioavailable DOM can stimulate soil microbial activities and subsequently promote soil aggregation and stabilization; in turn, this mechanism protects DOM from microbial mineralization and prevents losses from the soil system (Marschner and Kalbitz 2003;Six et al. 2006). This poses an important link between DOM occluded in soil aggregates and its role in aggregate stabilization. There is a large variety of sources for DOM in soil, ranging from plant root and microbial exudates to decomposed organic matter (Neff and Asner 2001;Bolan et al. 2011;Malik and Gleixner 2013;Schomakers et al. 2014). The organic compounds include, for example, simple carbohydrates and amino acids with low molecular weight, organic acids and proteins, amino sugars, or more complex polysaccharides, which show a wide range of degradability (Bolan et al. 2011). Some of DOM are extracellular polymeric substances (EPS) which directly contribute to soil aggregation as binding agents (Costa et al. 2018). In soil systems, the quality of DOM controls physical (e.g., leaching), chemical (e.g., sorption, complexation), and biological (e.g., decomposition) processes (Kalbitz et al. 2005;Van Hees et al. 2005;Grandy and Neff 2008;Kaiser and Kalbitz 2012;Karavanova 2013;Roth et al. 2019). In recent years, studies on soil organic C fraction have been increasingly conducted, with the dissolved fraction receiving higher attention (Gmach et al. 2020). However, we still lack a comprehensive biochemical understanding about this vital C fraction in soil.
Recently, several studies reported DOM quantity and quality changes with soil management on arable land. Conservation agriculture practices have shown to increase DOM contents in soil (i.e., Steenwerth and Belina 2008;Guo et al. 2016;Sharma et al. 2017;Martínez-García et al. 2018;Schmidt and Martínez 2019;Li et al. 2021;Jakab et al. 2022). This increase could in turn positively feedback on SOC sequestration via the soil microbial C pump (Liang et al. 2017). Also, DOM quality (as indicated from its C quality) was proposed to be a more useful indicator than its quantity for monitoring soil heath (Jones et al. 2014). However, alteration in DOM quality due to conservation agriculture practices remains controversial (Cawthray 2003;Cuss and Guéguen 2013;Sharma et al. 2017;Martínez-García et al. 2018;Schmidt and Martínez 2019;Jakab et al. 2022). Under field conditions, factors such as soil type (Li et al. 2019) may have greater influence on DOM dynamics than biotic controls.
DOM quantity and quality in soil studies are commonly represent a total fraction. This however hardly approximates DOM availability (and thus a relevant part of SOM stabilization mechanisms) in real soil where it is tightly bound to soil aggregates and the related pore spaces (Karavanova 2013). The aggregate hierarchy theory suggests that smaller units constitute larger units of soil aggregates (Tisdall and Oades 1982). These subunits are more stable and more persistent than the larger ones due to differences in binding agents and sorption mechanisms. Accordingly, the determination of DOM release from soil aggregates could provide mechanistic insight into stabilization processes of this active fraction in soil and thus better represent potential SOC effects of different soil managements where aggregate-bound C is a key sensitive indicator (Six et al. 2000;Kiem et al. 2002;Wiesmeier et al. 2019). The application of ultrasonic energy effectively disperses soil aggregates, which leads to a continuous release of occluded DOM (Schomakers et al. 2011(Schomakers et al. , 2014. A level of ultrasonic energy application during a dispersion process can be used to infer mechanical stability of soil aggregates and subsequently occluded SOC (Kaiser and Berhe 2014). An ultrasonic energy of 60 J ml −1 was suggested to completely disperse soil macroaggregate (250 -2000 μm), while ≥ 440 J ml −1 are required for microaggregate separation (20 -250 μm). A study by Schomakers et al. (2014) reported that 6.7 J ml −1 was sufficient to release dissolved organic fractions obtained from plant and microbial origins in the soils.
In the present study, we established a novel approach combining low energy ultrasonication and the simultaneous monitoring and characterization of the quality and quantity of DOM released during soil aggregate breakdown using ultraviolet-visible (UV-Vis) spectroscopy. UV-Vis spectroscopic measurement has been widely used in environmental research and monitoring. In general, absorption at short wavelengths refers to low molecular weight molecules (LMW) and less aromaticity molecules, while high molecular weight molecules (HMW) and high aromaticity (or complex) molecules are indicated by measurement at higher wavelengths (Helms et al. 2008). We expect that combining aggregate breakdown with DOM release can improve soil health assessment of agricultural management systems by integrating physical (aggregate stability) with organic C (DOM release) descriptors. In this study, we aim to investigate the effect of agricultural management and pedological properties on the release of DOM during gradual aggregate breakdown by using the novel established method. We compare conventional and conservation agricultural systems and relate those to permanently covered reference soils. We hypothesize that (1) the quantity of released DOM and aggregate stabilization show a steady continuum starting from conventional farming through conservation agriculture towards reference soil systems; moreover, the quality of DOM released differs between management systems; (2) the extent of agricultural management effects on DOM characteristics and release mechanisms is dependent on site-specific soil characteristics.

Study sites and site selection
We chose an on-farm approach on 21 study sites across North-Eastern Austria, where the majority of agricultural land is cropland (Statistik Austria 2021). These sites cover a wide range of relevant arable soil types. Mean annual precipitation ranges between 540-700 mm. For more details, we refer to Rosinger et al. (2022). Each site comprised of a conventional farming system (from now on referred to as "standard farming"), a conservation farming system (from now on referred to as "pioneer farming") and a permanently vegetated reference soil system. The standard farming systems are characterized by regular ploughing at 20-25 cm soil depth, no cover or inter cropping, a rather low diversity crop rotation, predominantly mineral fertilization, and the use of pesticides. Pioneer farming systems have set themselves the operational target of increasing SOC and improving soil health through the implementation of conservation management techniques (i.e., conservation tillage, intensive use of cover/inter cropping and/or a highly diverse crop rotation, organic fertilization). Conservation agriculture practices have been implemented for a minimum duration of 9 years (mean 26 years). Reference systems are shaped by cumulative ecological legacy from undisturbed permanent vegetation cover under given climatic and soil type conditions . Common grass species found in the reference systems are Lolium perenne, Poa trivialis, Dactylis glomerata, Alopecurus pratensis, Festuca rubra or Agropyron repens. Soil samples were collected from adjacent fields ensuring identical soil type (texture) at every study site. To analyze the site-specificity of management effects, farms were selected over a soil type gradient covering all common arable soil types in the area (BFW 2020).

Soil sampling
Soils were sampled to a depth of 0-5 cm using a soil core auger (Ø 7 cm) with 4 field replicates (sample points) (Supplementary Fig. 1) during the growing season (March -July) in 2020. The soil samples from the standard and the pioneer farming systems were collected between crop rows from 5 × 5 m plots with a 4 m buffer strip between them. For reference systems, soils were sampled from the most nearby permanent grass strip or hedge row along approximately 14 m line transects according to its dimension and shape. The soil samples were immediately sieved to 2 mm to obtain a minimum of 200 g from each field replicate and subsequently air-dried. Prior to laboratory analysis, the field replicate soil samples were pooled corresponding to their treatments and study sites with equal volume. Thereby, a total of 63 composite soil samples were obtained.
Dissolved organic matter (DOM) extraction DOM was extracted from a wide soil:water ratio solution to avoid filter blocking and maintain constant flow rate during UV-Vis spectroscopic online measurement. 2 g air-dried soil was added to 200 ml ultrapure water (resistivity 18.2 MΩ⋅cm at 25 °C; total organic C ≤ 2 ppb) immediately before the ultrasonication and stirred with a magnetic device to homogenize the soil-water suspension (Schomakers et al. 2011). An in-house developed ultrasonic equipment with a cylindrical probe of diam. 30 mm was used for the experiments to allow application of precise lowenergy ultrasonic energy. The probe was inserted in a cylindrical beaker (Ø 80 mm) at a depth of 10 mm with a distance of 30 mm to the bottom (Fig. 1a). The probe was vibrating at an amplitude of 1.2 µm at a frequency of 19.1 Hz. Using a probe with a large diameter that vibrates at small amplitude close to the cavitation threshold was found to be most appropriate to differentiate the influence of soil management on soil aggregate stability (Schomakers et al. 2011). At the selected amplitude, the ultrasonic equipment produced a power of 4.66 W, which was determined by caloric calibration procedure following Schomakers et al. (2011). The ultrasonication was performed for 500 s until the DOM release was steady, thereby yielding a total energy of 2,330 J which was equal to 11.7 J ml −1 . The soil solutions were collected after the ultrasonication and stored at -20 °C for DOM quantity and quality determination as illustrated in the analysis scheme (Fig. 1b).

UV-Visible spectroscopic measurement of DOM release
Absorbance at different UV-Vis wavelengths is commonly used for (semi-) quantitative and qualitative (by means of absorbance ratios) determination of DOM in environmental studies (Minor et al. 2014;Li and Hur 2017). In general, the absorbance at 254 nm is specific for DOC determination and can be used for calculating its concentration (UV-DOC) (Brandstetter et al. 1996;Kasper et al. 2009;Schomakers et al. 2011). However, DOM and DOC terminologies are sometime interchangeable due to high C content in DOM. In the current study, we refer DOM to dissolved organic fraction and DOC exclusively to constituent C of DOM. Hence, during the entire DOM extraction process by ultrasonication, UV-Vis absorbance of the released DOM at 210, 254 and 400 nm were simultaneously measured online to determine DOM release characteristics.
The DOM extraction and the measurement units were connected by a silicone and polytetrafluoroethylene (PTFE) tubing circuit. The flow rate of 4 ± 0.2 ml min −1 was controlled by a peristaltic pump. Two filtration systems comprising a 5 ml glass woolfilled syringe filter and a double 0.45 µm PTFE membrane filter (VWR International, LLC) were installed to eliminate the interference from coarse and fine particles, respectively. The UV-Vis absorbance of the filtrated soil solution was continuously measured by UV-Vis spectrophotometry (Agilent 8453 UV-Vis spectroscopy system, Agilent Technologies, Inc.) in a 10 mm quartz flow-through cuvette. Total measurement time was 500 s at a 5 s interval and a 71 s delay time due to the flow rate and the tube distance to the Fig. 1 Schematic presentation of (a) our device setup. The ultrasonication device was connected to the UV-Vis spectrophotometer for the online determination of dissolved organic matter (DOM) release characteristics. Immediately after adding a soil sample into ultrapure water and throughout the measurement, the soil solution is homogenized by the magnetic stirrer. During ultrasonication, the soil solution simultaneously circulates through the whole system. It passed through two filtering steps prior to UV-Vis absorbance measurement by the spectrophotometer. The glass wool filter and the double 0.45 μm PTFE membrane filter were installed to remove coarse and fine soil particles, respectively. The flow rate of soil solution was controlled by a peristaltic pump. (b) Analysis scheme demonstrates simultaneous UV-Vis spectroscopic measurement at wavelength 210, 254 and 400 nm at every 5 s along 500 s of aggregate dispersion by ultrasonication to assess DOM release characteristics. Subsequently, soil solution sample is collected for non-purgeable organic C (NPOC) measurement to quantify total dissolved organic C (DOC) and UV-Vis spectroscopic measurement at wavelength ranging from 200 to 650 nm with 1 nm interval to assess the quality of extracted DOM flow-through cuvette. Thus, DOM release characteristics display the accumulation of gradually released DOM from dispersion of soil aggregates within a 0 -10 J ml −1 energy range. Ultrapure water was used for routine blank calibration for every sample.
DOM quantity and quality determination The collected soil solution sample was passed through a 0.45 µm PTFE membrane filter prior to UV-Vis absorbance and non-purgeable organic C (NPOC) measurements. The absorbance of UV-Vis spectra was measured at wavelengths ranging from 200 to 650 nm with 1 nm interval using an UV-Vis spectrophotometer. The absorbance of ultrapure water was also recorded and served as a blank and as a correction value to minimize absorbance interferences. Prior to quantity measurement, the filtrate soil solution samples were treated with 1.5% sulfuric acid to remove inorganic C and purgeable organic C following the DIN EN 1484 (2019). NPOC contents were then measured using a Shimadzu total organic carbon analyzer (TOC-L series). This measurement represents the total DOC concentration in the samples. It was also required for the aromaticity determination of DOC indicated by specific UV absorbances as ratios between UV absorbance at 254 to NPOC (SUVA 254 ) (Weishaar et al. 2003;Gao et al. 2018;Zhang et al. 2022).
Physical and chemical soil property analyses The soil particle size distribution was analyzed following ÖNORM L 1061-2 (2019). Soil particle sizes were re-classified according to Nemes et al. (1999) procedure and then used to categorize soil texture into three main classes (coarse, medium, and fine texture) following the USDA classification (USDA-NRCS 2017). Soil aggregate stability was determined by wet sieving (ÖNORM L 1072(ÖNORM L 2004. Briefly, soil aggregates with diam. 1000-2000 µm were dipped on a 250 µm sieve. Here, 4 g of soil (EW) were used. The mass of stable aggregates after dipping (m K ) and the mass of sand after chemical dispersion of the remaining aggregates (m A ) is determined. These parameters are used to calculate the relative amount of stabile aggregates (%SAS, Eq. 1). (1) Soil pH and electric conductivity (EC) were measured in 1:10 soil:water (w/w) (ÖNORM L 1083(ÖNORM L 2006. Cation exchange capacity (CEC) were analyzed according to ÖNORM L 1086ÖNORM L (2001. Total C (TC) and total nitrogen (TN) contents were quantified by dry combustion. SOC was calculated from the difference between TC and inorganic C content obtained from the Scheibler method (ÖNORM L 1084(ÖNORM L 2006. Concentrations of aluminium (Al), iron (Fe) and manganese (Mn) oxides were assessed from dithionite (Al D , Fe D and Mn D , respectively) and oxalate extraction (Al O , Fe O and Mn O , respectively) (Candra et al. 2021). Ratios of SOC to TN (C:N) as well as SOC to clay content (SOC:clay), and between Fe D and Fe O (Fe D :Fe O ) were calculated. The SOC:clay ratio is indicative of soil structure quality (Johannes et al., 2017) while the Fe D :Fe O ratio refers to the degree of Fe oxide crystallinity (McKeague and Day 1966;Candra et al. 2021). For more details on the methods, we refer to Rosinger et al. (2022).

DOM release characteristics
Pre-processing of DOM release data was necessary to eliminate noise and other inconsistencies. The data were smoothed by a simple moving average method over 5 absorbance measurements. UV-Vis absorbance baselines were corrected. Spline interpolation was subsequently applied due to its smoother and better fit. Ultrasonic energy in J ml −1 was calculated from measurement time (with delay time exclusion) multiplied by ultrasonic power. As the DOM release followed a sigmoidal shape (Fig. 2), we inferred the following points to characterize the curve pattern: a starting point of release (A int ), a starting point of rapid release (A inc ), a maximum slope (A smax ) where a maximum DOM release rate occurred, a starting point of steady release (A dec ), a point where the absorbance was a half of the maximum absorbance (A half = A max /2), and a maximum UV-Vis absorbance (A max ) ( Fig. 2 and Table 1).
The UV-Vis absorbance and the applied energy indicate the abundance of released DOM and the stability of soil to release DOM at the given points, respectively. The quality of the released DOM was assessed by computing ratios of the absorbance at 210, 254, and 400 nm at every point. The ratio of UV absorbance at 210:254 nm refers to the UV absorbance ratio index (URI) -an indication of the relative proportions of unsaturated compounds to UV-absorbing functional groups (Her et al. 2004). A rate of the rapid release (R inc ) was calculated from the difference of absorbance between A inc and A dec divided by the difference of energy used at the respective points (Fig. 2) as an indication of the most unstable soil condition in releasing DOM within the range of energy applied.

DOM quantity and quality
DOC concentrations in mg kg −1 were assessed from NPOC while DOM quality assessment was based on UV-Vis spectrophotometric data. UV-Vis absorbance spectra were preprocessed by subtracting the absorbances with the blank prior to baseline correction. The spectral baseline was corrected by the absorbance spectra at 600 -620 nm (due to their steadiness and close to zero) using staRdom package (Pucher et al. 2019). The corrected absorbances were subsequently used to assess DOM spectral parameters such as the absorbance at 254, 260 and 280 nm (A 254 , A 260 and A 280 respectively), and SUVA 254 (details in Table 1).

Correlation analysis
The obtained data did not meet the criteria for normal distribution and variance homogeneity. Hence, a robust one-way analysis of variances (ANOVA) was applied to determine significant effects of management systems on DOM release characteristics (i.e., every point on the curve) at p < 0.05 using WRS2 package Wilcox 2017, 2020) in R software. Subsequently, linear contrast was applied as a post hoc test on the 20% trimmed mean.
Categorical regression analysis was used to determine significant soil physical (sand and clay content, aggregate stability), chemical (soil pH, EC, SOC, TN, SOC:clay ratio, SOC:TN ratio, sum of dithionite-extractable Fe-and Al-sesquioxides) and management predictors of the obtained DOM release parameters. This approach allows the sophisticated use of ordinal and nominal variables, since scale values are assigned to each category of every variable such that these values are optimal with respect to the regression. The solution of a categorical regression maximizes the squared correlation between the transformed response and the weighted combination of transformed predictors. In this regard, "management" was used as a nominal variable, with the values 1, 2 and 3 assigned to the standard, pioneer and  The UV-Vis absorbance at 254 nm Aromaticity of the DOM and proxy for DOM concentration estimation (Brandstetter et al. 1996;Weishaar et al. 2003 The aromaticity of the DOM (Weishaar et al. 2003) reference system, respectively, to evaluate parameters inherent to the different management systems but not yet accounted for with the above-mentioned variables (e.g., biochemical parameters). Silt content and other oxides-related variables were excluded due to multicollinearity. The analysis was conducted under optimal scaling and with cross validation in SPSS 26. Significant models and relationships were determined at p < 0.05.

Wavelengths dependent DOM release characteristics
Across all management systems, UV-Vis absorbances at 210 nm (A 210 ) were highest among measured wavelengths for all points followed by 254 (A 254 ) and 400 (A 400 ) nm, respectively (p < 0.05) (Fig. 3). Energy application to extract 210 nm UV-absorbing DOM was lowest before reaching A half (p < 0.01) compared to the absorbance at 254 (A 254 ) and 400 nm (A 400 ). After A half , the energy required to extract 210 nm UVabsorbing DOM turned to be significantly higher than that required to extract 254 and 400 nm UV-absorbing DOM (p < 0.05).

Management system dependent DOM release characteristics
Significant differentiation of the UV-Vis absorbances and the energy applied to release DOM among management systems started to be apparent at A smax (p < 0.05) for all detected wavelengths (Fig. 4). The reference systems showed the highest absorbance at all measured wavelengths (p < 0.05) from A half to A max ; absorbances in standard farming systems were lowest (p < 0.05). DOM characteristics at A 254 differed significantly (p < 0.05) across management systems, with the following order: standard farming systems < pioneer farming systems < reference soil systems. DOM release from the soils detected at 210 nm required different amounts of energy depending on management systems from A smax to A half (p < 0.05). In contrast, the amount of energy needed to release 254 and 400 nm absorbing DOM only differed significantly (p < 0.05) at A max among the management systems. The energy for DOM release from reference systems was significantly (p < 0.05) higher than the standard and the pioneer farming systems.
Rates of DOM release (R inc ) were not significantly different among agricultural management systems (Table 2) but with soil texture classes (p < 0.001). Coarse-textured soils had the lowest R inc , while finetextured soils showed the highest R inc . Medium-and fine-textured soils released DOM at similar rates at all detected wavelengths.
Absorbance ratios of A 254 to A 400 (A 254 :A 400 ) had significant differences among management systems between A half and A max (p < 0.001). The reference systems showed significantly higher A 254 :A 400 ratios of 7.0 -7.2 on average compared to the standard Fig. 3 (a) UV-Vis absorbance and energy application during DOM release and (b) energy applied at observed points measured at different wavelengths averaged over all three management systems. Given is the mean ± SE (n = 63) and the pioneer farming (approximately 6.0 and 6.3, respectively) (Fig. 5).

Management induced differentiation of cumulative DOM quantity and quality
The concentrations of DOC as determined via NPOC were different among the management systems (p < 0.001), ranging from the highest (602 mg kg −1 ) in the reference to the lowest (390 mg kg −1 ) in the standard farming systems (Fig. 6a). The UV-Vis absorbance of extracted DOM at every measured wavelength from the standard farming systems was significantly lower compared to the reference systems (p < 0.05), while absorbance of pioneer farming was more comparable to the reference (Fig. 6b). However, SUVA 254 was not significantly different among management systems and ranged between 2.98 and 3.18 L mg -1 C m -1 on average.

Correlation analysis
For the absorbance and the absorbance ratios, significant relationships between DOM release characteristics and soil properties, as indicated by significant models (p < 0.05), were most strongly related from A inc onwards (Figs. 7 and 8). Among soil properties, soil pH and EC were the most significant positive and negative predictors, respectively, for A 210 as well as its derived absorbance ratios (A 210 :A 254 and A 210 :A 400 , p < 0.05). On the other hand, the variable management was the strongest positive predictor (p < 0.001) for A 254 and A 400 (Fig. 7a). Apart from that, CEC and management significantly (p < 0.05) affected A 210 at A inc , while clay content and aggregate stability significantly predicted A 400 at A dec (Fig. 7a). Interestingly, the C:N ratio had a significant effect on the ratios A 210 :A 254 and A 210 :A 400 (Fig. 8). There were no clear patterns in the prediction of the energy applied to release DOM through management and physico-chemical predictors (Fig. 7b). R inc of A 210 was significantly (p < 0.05) predicted by both soil pH and EC. Soil texture together with management significantly (p < 0.05) regulated R inc of A 254 and A 400 (Fig. 8). Furthermore, DOC concentrations after ultrasonic extraction (as determined by NPOC) were significantly (p < 0.05) predicted by soil aggregate stability and TN as well as management (Fig. 9). Soil pH had a significant (p < 0.05) negative effect on the absorbance at 254 and 280 nm.

DOM release characteristics upon ultrasonication-induced aggregate decay
In the present study, we evaluated the gradual release of DOM from the dispersion of soil aggregates by a novel approach combining continuous aggregate breakdown by application of ultrasonic energy along with the simultaneous measurement of UV-Vis absorbance. The ultrasonic energy used in this study (≤ 10 J ml −1 ) is sufficient to break macroaggregates into sub-macroaggregates (250 -2,000 μm) without the interference from mineral associated DOM and fragmentation of organic particles (Mentler et al. 2004;Kaiser and Berhe 2014;Schomakers et al. 2014). Thus, the obtained measures provide appropriate descriptors for the physically uncomplexed DOM (such as free DOM) and occluded DOM in macroaggregates, being the primary sources of available DOM in soil upon disturbance (e.g., rainfall and/or irrigation impact).

Wavelength-dependent DOM release characteristics
The release of DOM during ultrasonication as detected by UV-Vis absorbance at a wavelength of 210, 254 and 400 nm is characterized by a sigmoidal shape Table 2 DOM release rates (R inc ) measured at different wavelengths (λ) under different management systems (standard farming, pioneer farming, and reference soil systems) and soil texture (coarse, medium, and fine). Given is the mean ± SE ( 1) n = 21, 2) n = 17, 3) n = 24, 4) n = 22), and different letters indicate significant differences between texture classes as determined by robust one-way ANOVA followed by linear contrast post hoc test on trimmed means (p < 0.05) R inc (× 10 -3 J −1 ⋅ml)

Management system
Soil texture Standard farming 1) Pioneer farming 1) Reference 1) Coarse 2) Medium 3) Fine 4) λ 210 56.1 ± 6.2 63.1 ± 5.7 74.7 ± 7.5 40.5 ± 4.5 a 72.0 ± 5.9 b 75.2 ± 6.4 b λ 254 18.5 ± 2.6 18.2 ± 1.4 19.1 ± 1.3 12.5 ± 1.2 a 18.3 ± 0.9 b 23.7 ± 2.2 b λ 400 3.54 ± 0.54 3.28 ± 0.26 3.16 ± 0.25 2.14 ± 0.21 a 3.32 ± 0.20 b 4.26 ± 0.47 b Fig. 5 Absorbance ratios of A 254 :A 400 at A half , A dec , and A max for the standard farming, the pioneer farming and the reference soil systems. Given is the mean ± SE (n = 21). Different letters indicate significant differences between management systems as revealed by robust one-way ANOVA followed by linear contrast post hoc test on trimmed means (p < 0.05) Fig. 6 (a) DOC concentrations extracted from soils at 0-5 cm depth in the standard farming, the pioneer farming and the reference soil systems using 11.7 J ml −1 ultrasonic energy and (b) its UV-Vis absorbance at selected wavelengths. Given is the mean ± SE (n = 21). Different letters indicate significant differences between management systems as revealed by robust one-way ANOVA followed by linear contrast post hoc test on trimmed means (p < 0.05) (Fig. 3a). On average, 0.8 -1.3 J ml −1 of energy were required for the initial release of DOM (A int ). This is comparable with the energy of rainfall. The kinetic energy of rain depends on the intensity (van Dijk et al. 2002); however, it is typically in the regime between 20 and 35 J m −2 mm −1 (van Dijk et al. 2002;Shin et al. 2016). The ultrasonic probe had a diameter of 30 mm (area 7.1 cm 2 ) while the beaker's diameter was 80 mm (area 50.3 cm 2 ), i.e., the ultrasonic energy was introduced at 14% of the surface. Thus, the emitted energy for the initial release of DOM was between 4.5 and 7.3 kJ m −2 . The same amount of energy would be transported to the surface in a rainfall scenario with 150 -240 mm of rain. The strongest change in DOM release -the turning point of the sigmoidal curve (A smax ) -was approximately at 3 J ml −1 ultrasonic energy, which corresponds to the typical kinetic energy of 560 mm rain and resulted in DOM release at the maximum rate. Relatively stable absorbance starting from A dec (> 5 J ml −1 ) until 10 J ml −1 suggests that tremendously higher energy levels are required to disperse and release DOM from the more stable aggregates Fig. 7 Results of a categorical regression analyses showing the prediction of (a) the UV-Vis absorbance and (b) the energy applied during DOM release determined at wavelength 210, 254, and 400 nm through soil properties and management (n = 63). The colour gradient represents regression coefficients when significant, while displayed numbers are F-values. F-value of the regression coefficients are only displayed when the overall regression model (in grey column) is significant (p < 0.05). x , *, **, and *** indicate a significant prediction at p < 0.1, p < 0.05, p < 0.01, and p < 0.001, respectively (i.e., free and occluded smaller aggregates named subaggregates). Kaiser and Berhe (2014) suggested that 60 J ml −1 of ultrasonic energy are required to disperse macroaggregates (≥ 250 µm), at least 440 J ml −1 to break microaggregates (< 250 µm), and even more to release mineral-complexed organic compounds. In general, absorption at short wavelengths refers to LMW and molecules with less aromaticity Fig. 8 Categorical regression analyses showing the prediction of UV-Vis absorbance ratios among selected wavelengths and rate of rapid DOM release (R inc ) determined at 210, 254 and 400 nm (λ 210 , λ 254 and λ 400 respectively) through soil properties and management (n = 63). The colour gradient represents regression coefficients when significant, while displayed numbers are F-values. F-value of the regression coefficients are only displayed when the overall regression model (in grey column) is significant (p < 0.05). x , *, **, and *** indicate a significant prediction at p < 0.1, p < 0.05, p < 0.01, and p < 0.001, respectively Fig. 9 Categorical regression analyses showing the prediction of DOC concentration and UV-Vis spectral parameters of DOM extracted using 11.7 J ml −1 ultrasonic energy through soil properties and management (n = 63). The colour gradient represents regression coefficients when significant, while dis-played numbers are F-values. F-value of the regression coefficients are only displayed when the overall regression model (in grey column) is significant (p < 0.05). x , *, **, and *** indicate a significant prediction at p < 0.1, p < 0.05, p < 0.01, and p < 0.001, respectively (hereafter referred to as "easily degradable compounds"), while HMW and high aromaticity (or complex) molecules (hereafter refers to as "complex compounds") are indicated by increased absorbance at longer wavelengths (Helms et al. 2008). Hence, easily degradable compounds as detected at 210 nm would be initially released and dissolved in the soil solution. Differentiation of the energy used for releasing 210 nm absorbing compounds at A dec and A max demonstrates that > 5 J ml −1 can apparently liberate easily degradable compounds from more stable aggregate structures, while this is not obvious for complex compounds.

Management system dependent DOM release characteristics
The differentiation of DOM release characteristics among management systems started to emerge from A half (> 3 J ml −1 ) onwards, an energy equivalent to a heavy rainfall event (Schomakers et al. 2018). The greater release of DOM from reference soils was a result of higher DOM levels in these permanently vegetated systems. This subsequently enhanced DOM accumulation and stabilization in soil (Schmidt and Martínez 2019;Sokol et al. 2019;Sokol and Bradford 2019;Wiesmeier et al. 2019;Villarino et al. 2021).
Higher absorbance and energy applied at every point after A half (> 3 J ml −1 ) in the reference systems ( Fig. 4) indicated greater protection of aggregate-stabilized DOM as compared to both farming systems. The implementation of pioneer farming measures altered DOM release characteristics and tended to approximate characteristics inherent to reference soils. On the other hand, at energy below 2.5 J ml −1 -which is comparable to a typical rainfall regime (van Dijk et al. 2002) -similar amounts of DOM are prone to be lost from soils regardless of management systems if not being stabilized by soil microbial assimilation, occlusion in soil aggregates, soil mineral association, etc. Among the measured wavelengths, DOM release characteristics at 254 nm could best differentiate the management systems, supporting that A 254 is a highly valuable indicator for both DOM and DOC for changes in agricultural management (Zhou et al. 2000;Weishaar et al. 2003;Schreiber et al. 2005;Li and Hur 2017;Romero et al. 2017;Li et al. 2019).
Regarding DOM quality, the A 254 :A 400 ratio showed significant differences due to agriculture management from A half onwards (Fig. 5). In comparison, the highest A 254 :A 400 ratio in reference systems suggests a larger proportion of more easily degradable compounds occluded in more stable soil aggregates as compared to both farming systems. DOM quality varied with applied energy (Supplementary  Table 1), which refers to DOM components occluded in different aggregates. The increasing UV absorbance ratio index (URI) values starting from A inc in both farming systems implied a high proportion of complex compounds contained in unstable macroaggregates, while easily degradable compounds (i.e., amino acids and proteins) were occluded in more stable aggregates; this observation was not made in the reference systems.
The maximum DOM release (A max ) was reached with 20% lower energy in both farming system soils compared to the reference system soils. More than 50% of the DOM release curves obtained from farming systems showed a hump-backed shape after reaching A dec (see Supplementary Fig. 2a). This hump-back was particularly detected for clayey soil with low organic C content and high contents of Fe D . This is possibly caused by an interference of weakly bound and/or free ions, which likely occurred in higher concentrations in the farming system soils due to fertilizer application. The formation of DOM-ion complexes can alter DOM structure and thus UV-Vis absorbance (Li and Hur 2017). Interestingly, in more than 50% of the reference system soils, DOM release was characterized by a double-S shape (see Supplementary Fig. 2b). This indicates a further liberation of protected DOM from more stable aggregates in reference soils.
Management induced differentiation of cumulative DOM quantity and quality DOC concentrations significantly differed among management systems ranging from the highest under the reference systems to the lowest under the standard farming systems (Fig. 6a). The same trend was also evidenced for SOC contents , suggesting that DOC contributes to SOC sequestration through the microbial carbon pump (Liang et al. 2017). A positive relationship with TN ( Fig. 9) suggests that higher TN in the reference system soils ) enhanced plant and soil microbial growth and activities and thus increased DOC release in these soils. Furthermore, permanent soil cover in the reference systems and intensive cover cropping along with conservation tillage in the pioneer farming systems protect the soil and subsequently decreases DOC losses from soil erosion and leaching (Liebig et al. 2004;Morris et al. 2010;Scopel et al. 2013;Gmach et al. 2020). The positive relationship between aggregate stability and DOC contents further indicate that soil aggregation successfully decreases DOC losses from mineralization. Moreover, greater amounts of DOC would have in turn a positive feedback on aggregate stability through increased microbial activity (Sae-Tun et al. 2022). While the UV-Vis spectral indicators for DOM quality in the present study are in line with previously reported ones (Li and Hur 2017;Kholodov et al. 2020;Jakab et al. 2022), they were hardly affected by agricultural management measures (Fig. 6b). Instead, they were strongly controlled by soil pH (Fig. 9).

Relationship of DOM release with soil properties and management
The categorical regression analyses demonstrated that management-mediated and site-specific soil properties strongly regulate DOM release characteristics ( Fig. 7 and 8) as well as DOM quantity and quality (Fig. 9). However, the determinants for DOM release differ between easily degradable and complex compounds. The relationships between the absorbance at 210 nm and soil pH and EC (Fig. 7a) suggest that soil nutrient availability affects the abundance of easily degradable compounds being released. It should be noted that most of the studied soils had pH ≥ 7, thus slight decrease in soil pH would promote soil nutrient availability, and favor plant as well as fungal growth and activities (Rousk and Bååth 2011) in our study. On the other hand, management-related attributes, such as soil microbiological properties (Sae-Tun et al. 2022), significantly regulated the abundance of more complex compounds released from soil aggregates. In addition to the aforementioned soil properties, the observed negative relationship between C:N ratios and A 210 :A 254 as well as A 210 :A 400 ratios suggest that soil with a narrower C:N ratio, as observed in the pioneer farming and reference soil systems , promote the accumulation of more easily degradable compounds.
Significant relationships between energy used to release DOM and soil properties (Fig. 7b) emphasize an influence of site-specific conditions on DOM stabilization. The stabilization and accumulation of easily degradable compounds was found to be dependent on the amount of extractable Al-and Fe-sesquioxides. The release of more complex DOM compounds seems to be governed by soil texture, as indicated by its strong relationships with sand contents. This notion is also supported by significant relationships between R inc and soil properties (Fig. 8). We therefore hypothesize that easily degradable compounds are more likely stabilized by DOM-oxide complexes, while more complex compounds tend to be predominantly protected by either sorption on clay mineral surfaces or occlusion in soil aggregates (Wagner et al. 2007).

Conclusions
By using the combination of ultrasonic energy and continuous UV-Vis spectroscopic measurement, we could show that conservation agricultural practices, driven by the functional role of DOM in (agro)ecosystems, was clearly related to both higher DOM quantity and higher aggregate protection/stabilization as compared to soils from standard farming systems. Moreover, soil structural and organic matter (specifically the dissolved fraction) characteristics in the pioneer farming systems approximated those of the undisturbed soils under natural vegetation.
Compared to agriculturally managed soils, DOM released from the reference soil systems consisted of more easily available DOM compounds and was better protected in macroaggregates. The quality and quantity of DOM released was strongly shaped by soil physico-chemical properties and land management; while the release of easily degradable DOM compounds was strongly driven by soil chemistry, the release of more complex compounds was best predicted by management. Minimum mechanical soil disturbance leads to more stable aggregates resisting the simulated breakdown by ultrasonic energy and permanent (diverse) soil cover increasing the amount of functional C compounds (DOM).
Thus, the novel analytical approach implemented in this study is particularly suitable to simultaneously capture DOM quality and aggregate stability. The results of our studied soils prove that this is a successful method for monitoring soil health and the impact of the key principles of conservation agriculture. It provides relevant insights into the biophysical protection mechanisms for SOC sequestration. We thus encourage further studies on evaluating release patterns of this active and vital organic fraction in different land-use systems or along the soil profile in order to gain a better mechanistic understanding of SOC sequestration.
Acknowledgements The authors would like to thank Bernhard Scharf for his help in coordinating with farmers; Luca Bernardini for his support in data analysis; David Luger and Philipp Steiner for sample processing and their assistance in laboratory analyses; Astrid Hobel, Elisabeth Kopecky, and Karin Hackl for their support during the laboratory work as well as all support from fieldwork team for sample collection. Many thanks also for Boden.Leben and HUMUS Bewegung networks, the landowners, farmers, and other land managers for collaboration and permission to collect data and samples from their fields. The authors acknowledge Sabine Huber for third party reviewing.
Authors' contributions GB, KK, AM and OS contributed to the study conception and design as well as conducting fieldwork. OS and AM performed laboratory analyses. OS, KK and CR undertook data analysis. AM and HM supervised technical ultrasonic operation and validation. All authors contributed to interpretation of data, writing of the article and final approval of the version submitted.
Funding Open access funding provided by University of Natural Resources and Life Sciences Vienna (BOKU). Orracha Sae -Tun received the Ernst Mach Grant ASEA-UNINET from Austria´s Agency for Education and Internationalisation (OeAD). The study was carried out under research projects funded by the Gesellschaft für Forschungsförderung NÖ (GFF) as part of the RTI-strategy Lower Austria 2027 (Grant Nr. FTI19-002) and UMWELTFONDS Fonds zur Förderung einer nachhaltigen Entwicklung der Region rund um den Flughafen Wien (Environmental Fund for Sustainable Development of the Region around Vienna International Airport).

Data availability
The data supporting the findings of this study will be shared on reasonable request to the corresponding author.

Conflict of interest The authors declare no conflict of interest.
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