The Isotopic Composition of Selected Phosphate Sources (δ18O-PO4) from the Area of the Vistula and Bug Interfluve (Poland)

Abstract

Phosphorus belongs to the crucial bioelements that cause eutrophication, and phosphates, easily assimilated by organisms, are widespread in the environment. Phosphates can be of natural or anthropogenic origin and can derive from various point or non-point sources. Knowledge about the origin of nutrients is necessary to effectively manage, protect, and revitalize water resources. To recognize various phosphate sources in the study area of our research, i.e., the Vistula and Bug interfluve (SE Poland), we used the oxygen isotopic signature of phosphate ions (δ¹⁸O-PO₄), which has been successfully used in recent decades as a tracer of phosphorus cycling in water studies. We measured the δ¹⁸O-PO₄ of dissolved inorganic phosphates (DIPs) extracted from various phosphate sources. The obtained results are as follows: For springs, the δ¹⁸O-PO₄ value varied from +14.8‰ to +18.5‰; for riverine samples, from +10.3‰ to +18.6‰, which were significantly location-dependent; while waste water treatment plant effluents ranged from +12.4‰ to +15.6‰. Two tested drainage water samples had similar isotopic compositions (+16.7‰ and +17.3‰). In the case of two analyzed bedrock samples, the δ¹⁸O-PO₄ values, which were similar (+20.5‰ and +21.7‰), are close to the existing data on sedimentary bedrocks derived from similar geological periods. The obtained results can be helpful in future research aimed at identifying phosphate sources and P cycling in the studied area.

1. Introduction

Phosphorus plays a major role in many processes necessary for life, like DNA and RNA formation, and is crucial in the transmission of chemical energy through ATP molecules. P is also a structural ingredient in many components in cell membranes, teeth, and bones. The phosphorus content of the Earth’s crust is 0.10–0.12% on a weight basis [1], making it the 11th most abundant element [2]. Minor P reservoirs are oceans, with about 88 ppb of P content [3]. Phosphorus, whose bioreactivity increases from mineral to occluded to organic form [4], occurs in a +5-oxidation state in most mineral and organic compounds, mainly in the form of phosphate, PO₄³⁻. It is taken up by plants in the form of phosphate ions dissolved in water, especially H₂PO₄⁻ in an acidic environment and HPO₄²⁻ in an alkaline environment.

Phosphorus, which is necessary for the growth and proper functioning of organisms, in excessive quantities leads to the eutrophication of surficial waters, resulting in the deterioration of water quality and a disruption of the ecological balance. The total phosphorus content is one of the indicators for assessing water trophic levels, originating from the system proposed by Vollenweider in the 1960s [5]. Currently, in Poland, the Regulation of the Minister of Infrastructure from 2021 [6], regarding the limit values of the indicators of surface water quality that relate to river water bodies, determines the permissible limits of individual indicators, depending on the category of the considered surface water. In accordance with the above regulation, the permissible concentrations of P-PO₄ are within the range of 0.005–0.065 mg/dm³, and the total phosphorus content should not exceed 0.05–0.2 mg/dm³, depending on the character of the surface water.

Poland also faces the problem of bad-quality surface water, whereby the state of approximately 90% of river surface waters is bad (more information can be found in Rączka et al. [7]), according to the Water Framework Directive, which states that the condition of surface waters is assessed as good, if both their ecological and chemical status are good. To assess the ecological status of surface waters, both biological and physicochemical elements are taken into account, while in the assessment of the chemical status, the concentrations of particularly harmful substances are determined. While the content of nitrogen and phosphorus compounds in this case rarely exceeds the permissible limits for drinking water, their presence favors water eutrophication. For example, in the area of our study, the results of daily measurements of the orthophosphate concentration in the water from the Wieprz River, conducted at the research station in Guciów in 2021–2023, showed that the monthly averages of PO₄³⁻ concentration varied from 0.4 to 0.6 mg/dm³. To effectively protect our water resources and effectively counteract unfavorable changes, we need the tools to identify the source of the problem. In this case, we are primarily interested in whether the phosphates in the studied waters come from natural or anthropogenic sources and what the impact of individual sources in the total phosphorus pool is.

In order to be able to follow the P cycle in the studied ecosystem and to be able to determine what is the main source of phosphates, we can use an isotope analysis of the oxygen in the phosphate ions. Phosphorus has only one stable isotope, ³¹P, but in most compounds, phosphorus is strongly bound to oxygen, with three stable isotopes and with atomic masses of 16, 17, and 18. Without biological activity, the O–P bond is not broken, and the isotopic fractionation associated with abiotic processes is not greater than 1‰ [8,9,10], so we can measure δ¹⁸O-PO₄ as an indicator of the P source and P cycling. The δ¹⁸O-PO₄ analysis is widely used in the recent literature for identifying different phosphate sources and to understand P cycling both in marine and freshwater environments [11,12,13,14,15,16,17].

Our study is focused on selected areas of the Vistula and Bug interfluve (south-eastern Poland), in which the main groundwater reservoir feeding the rivers is developed by Maastrichtian and Tertiary formations, developed in the form of opokas, marls, chalk, gaizes, limestones, and sands [18]. The studied opokas are sedimentary rocks, composed mainly of silica in the form of a mixture of quartz, opal, and other phases, with the participation of carbonate grains and with an admixture of clay minerals, glauconite aggregates, iron oxides, and hydroxides. The marls occurring next to them are mainly composed of carbonate grains and also a significant admixture of clay minerals. Gaizes are rocks composed of siliceous (mainly opal) fragments of bioclasts, together with an admixture of clay minerals, iron oxides, hydroxides, and carbonates. Decalcified gaizes not containing carbonates were also found in this area. Limestones constitute a smaller share of the discussed rocks, composed mainly of carbonate grains and a small admixture of clay minerals, glauconite, iron oxides and hydroxides. Additionally, quartz grains and muscovite flakes may occur in these rocks. Chalk rocks, composed mainly of carbonate microplankton, remains were also encountered. Sands were loose rocks, composed mainly of quartz grains and an admixture of plagioclases (crystalloclasts), carbonaceous fragments (lithoclasts), and also crushed fragments of shells (bioclasts). Apatite in these rocks occurs in the vicinity of carbonates and silica, and in sands, the source of phosphate is monazite minerals, constituting a small admixture below 5% by volume.

In this area, in the region of Rachów–Annopol, the presence of phosphate deposits has been documented. Although extraction in the Annopol mine was terminated in the 1970s for economic reasons, the abundance of phosphate concretions in the Annopol deposit was estimated to total 568 kg/m² in the 2020s [19], and the P₂O₅ content in phosphate concretions in the deposit profile was estimated to be at 13.5 (%), with the phosphate concretions having diameters of > 10 mm; phosphate deposits that are slightly less abundant are located in the area of Gościeradów (496 kg/m², 15.2% of P₂O₅, and phosphate concretions of > 2 mm in diameter). Other deposits in this area are less abundant and only have local significance. However, the average orthophosphate concentration in water extracts from collected bedrocks, selected for a preliminary examination, was determined to be at the level of 0.5 mg/dm³ (unpublished data), whereas for the nearest spring waters, the concentration levels were similar (from about 0.3 to 0.7 mg/dm³), which may suggest a high content of phosphates derived from the leaching of natural phosphorus minerals from sedimentary rocks in rivers fed by these springs. In order to broaden our knowledge on this subject, multi-proxy research was undertaken, including physicochemical, geochemical, mineralogical, and isotopic analyses.

The first part of our project was to identify potential phosphate sources, which can be introduced to surface waters in a studied area, that is, to determine the range of variability of the oxygen isotopic composition for various local phosphate sources. Research on this topic has not been conducted in Poland so far, apart from the preliminary studies conducted by the authors of [20]. Therefore, the first point of our project was to estimate the isotopic composition of the oxygen isotopic composition of phosphates from different sources (e.g., spring waters, sewage treatment plant effluents or bedrock samples). The δ¹⁸O-PO₄ variability ranges in phosphates from various sources may have similar and quite narrow ranges and may strongly depend on local conditions; therefore, their recognition was necessary before undertaking further research. These results, supplemented by further seasonal studies of chemical parameters and a water δ¹⁸O analysis, was later used to identify the main phosphate sources in studied waters.

The results of the oxygen isotopic composition analyses in DIPs presented here constitute the beginning of creating a database of potential phosphate sources in the Lublin Upland and Roztocze and ultimately also in other areas of Poland. Knowledge of the δ¹⁸O-PO₄ of individual local phosphate sources is necessary to better understand the processes occurring in the P cycle and to determine the origin of the main P inflows to waters, which may help in their protection and revitalization. Here, we present and briefly discuss the results of the spatial recognition of the δ¹⁸O-PO₄ of some phosphate sources studied in selected areas of the Lublin Upland and Roztocze (SE Poland).

1.1. Global Phosphorus Cycle

Phosphorus belongs to the class of the most important elements, limiting nutrients for terrestrial biological productivity. Unlike other most important environmental elements, such as a carbon, nitrogen, or sulfur, P does not typically undergo oxidation-reduction reactions and does not exist in gaseous forms; therefore, the phosphorus cycle is not as complex as the C, N, or S cycles. The availability of P in ecosystems is limited by the rate of release of this element during the weathering of exposed continental rocks and soils [2,21,22]. Phosphorus occurs mainly in the form of inorganic phosphate minerals and phosphorus-containing organic compounds.

The most abundant group of phosphate minerals is apatite, comprising hydroxyapatite, Ca₅(PO₄)₃OH; fluorapatite, Ca₅(PO₄)₃F; and chlorapatite, Ca₅(PO₄)₃Cl. However, the chemical composition of apatites is usually much more complex than their simple formulas, which is caused by isomorphic substitutions between Ca²⁺, Al ³⁺, Fe²⁺, and Fe³⁺, as well as trace-element inclusions [3,23,24,25,26,27]. Phosphate minerals are generally formed in the environment of magmatic processes or through precipitation from a solution (which may be microbially mediated), and the chemical composition of the formed minerals depends on the kinds of ions present in the solution at the time of precipitation [1]. These natural deposits of phosphate minerals are usually diverse and are collectively called ‘phosphorites’ to reflect differences in their chemical compositions.

The phosphorus cycle includes various living and nonliving environmental reservoirs and different transport pathways (see Figure 1). In the P cycle, micro-organisms play a very important role, being involved in the transformation of this element in various P pools, such as soil or aquatic environments. Briefly, we can summarize, after Ruttenberg [28], that the global P cycle begins from a tectonic uplift, followed by the physical erosion and chemical weathering of exposed rocks, resulting in soils producing and transporting dissolved and particulate phosphorus to rivers, whence P’s transportation to lakes and the ocean. The sedimentation of phosphorus in organic and mineral matter and burial in sediments constitute the last stage of the global P cycle.

Photosynthetic organisms utilize dissolved phosphorus (and other important nutrients) to build their tissues, and biological productivity depends on the P availability to these organisms, which form the basis of the food chain in terrestrial and aquatic systems. However, the phosphorus present in the lithosphere is not directly available to organisms; more easily assimilated forms, such as orthophosphates, occur through both geochemical and biochemical reactions at various stages of the phosphorus cycle. The biomass production results in the deposition of organic matter in soils and sediments in the next stage, where it becomes a source of fuel and nutrients to microbial communities [1,21,28]. The land cycle, in which P is transferred from soil to the biosphere, then back to soil, lasts an average of one year. The water cycle, in which organic phosphorus circulates among organisms living in aquatic reservoirs, lasts only a few weeks [29]. The principal phosphorus sinks of dissolved phosphorus formed from surface waters are downwelling and biological uptake. The primary sink for P in a marine environment is lost to sediments, whereas the main part of the particulate flux from rivers is lost to sediments on continental shelves, and a smaller portion is lost to deep-sea sediments, as was reported by, for example, Mackey and Paytan [1] and Paytan and McLaughlin [8].

The small amounts of phosphorus in the atmosphere may derive from aeolian dust, from the combustion of fossil energy resources, or from modern biomasses. This element, present in the atmosphere, does not affect air quality and is even essential for terrestrial ecosystems in areas poor in P, but, leading to the eutrophication of reservoirs, it may deteriorate the quality of standing waters.

1.2. The Human Impact on the Global Phosphorus Cycle

Human activity has significantly disturbed the natural P cycle. The most important processes, related to human activity and disrupting the natural cycle of phosphorus [4,28,30] are (1) deforestation and related losses of phosphorus due to soil erosion, (2) the production of P-containing fertilizers, and (3) municipal and industrial wastewater. A schematic view of the human impact on the global phosphorus cycle is presented in Figure 2.

According to Schlesinger [31], deforestation, usually by burning after the selective harvesting of trees, converts phosphorus from plant matter to ash, which is then rapidly dissolved, leached from the ash, and transported in rivers over timescales of a year or two.

Soil erosion is caused by various mechanical factors, mainly by water or wind, while human activity, including the destruction of vegetation, improper land cultivation, and the drainage of swamps, accelerates the degradation of soils. When fine particles of soil, such as clay or sand, are transported by water through runoffs, they can transfer the phosphorus bound to the particles, causing an export of particulate phosphorus from lands to water bodies. The total amount of phosphorus available in a soil profile decreases over time, as soil P is lost through surface and subsurface runoffs, whereas an increase in the P content of a water body can cause its eutrophication.

Another problem is related to the long-term use of agricultural lands. As a consequence of this anthropogenic activity, soils have a depleted phosphorus content, and humans try to compensate for the deficiency of this element in soils by adding P-containing fertilizers. According to the IFA report [32], at the beginning of the 21st century, the annual production of phosphate was around 53 million tones (P₂O₅), from which around 80% is used as fertilizers and another 5% in mineral feeds. A huge problem, both for the environment and the industry, is losses generated from the production and distribution of fertilizers (about 35%) and losses related to the production, distribution, and consumption of food (up to 60% [33]). Phosphorus utilization in animal production varies among various groups of livestock, which is why the dominant part of phosphorous from feeds goes to manure. Human waste, with waste from foodstuff processing, and the industrial use of detergents are additional, but minor, anthropogenic P sources [3,4].

Figure 2

The human impact on the global phosphorus cycle (based on Turner & Raboy [34]).

[Figure omitted. See PDF]

Nowadays, we can say that the global phosphorus cycle is an anthropogenic cycle; according to Fillipelli [4], the net input of dissolved P from lands to oceans has doubled compared to prehuman periods. Also, approximately 30% of atmospheric P transfers is caused by human activity, which plays a more significant role than was previously thought [35]. Regardless of the accuracy of estimating the anthropogenic P pool, it is obvious that the additional flow of phosphorus introduced into the P cycle by humans, on the one hand, will lead to the anthropogenic eutrophication of water reservoirs, and, on the other hand, the excessive exploitation of P-containing bedrocks will result in the rapid depletion of non-renewable P resources. Another problem related to the excess of phosphates in drinking water, reported by Penuelas and Sardans [36], is higher the risk of lung and skin tumors for domestic animals.

1.3. How to Use δ¹⁸O-PO₄ as a Tracer for Sources and the Cycling of Phosphates

Information about the origin of nutrients and their circulation in the environment is important for understanding the functioning of the ecosystem and its management. One of the nutrients necessary for the proper functioning of aquatic ecosystems is phosphorus. In opposition to other environmentally important light elements, phosphorus has only one stable isotope, ³¹P, and its double-marked stable isotope analysis, as in the case of carbonates, nitrates, or sulphates is not available. However, in most compounds, phosphorus is strongly bound to oxygen, with three stable isotopes (¹⁶O, ¹⁷O, and ¹⁸O), which allows for a measurement of the oxygen isotopic composition in phosphate ions (δ¹⁸O-PO₄).

The isotopic composition of oxygen, expressed in delta notation, is calculated as follows: ${\mathrm{\delta }}^{18 }\mathrm{O} \left[‰\right]=\left({\displaystyle \frac{R_{sample}}{{R }_{standard}}}-1\right)\times 1000$, where R denotes the ¹⁸O/¹⁶O ratio in the sample and standard, respectively. The oxygen isotopic composition is expressed relative to the international standard, the Vienna Standard Mean Ocean Water (VSMOW). Also, in isotopic studies, the isotopic fractionation factor (α), given as ${\alpha }_{A-B}={\displaystyle \frac{R_A}{R_B}}$, which expresses enrichment or depletion in a heavy ¹⁸O isotope between two substances A and B, where R_A and R_B denote the ¹⁸O/¹⁶O ratio in samples A and B, respectively, can be useful. The fractionation factor is close to 1; therefore, in isotopic studies, it is more often expressed by ε [in ‰], which is given as $\mathrm{\epsilon }=\left(\alpha -1\right)\ast 1000$. Here, ε represents the enrichment (ε > 0) or the depletion (ε < 0) of the rare isotope in B with respect to A.

Phosphorus can be derived by aquatic reservoirs, both from natural or anthropogenic sources, like phosphate rocks, guano, or excreta, or from anthropogenic sources, like fertilizers or detergents, and its sources can be separated into point and non-point sources. Among the first, we can include, for example, sewage discharge sites, and for the second, soil leaching and agricultural runoffs [9]. Identifying the sources of pollution is critical for the proper resource management and caring of the state of aquatic ecosystems. In recent years, the isotopic study of oxygen in P compounds is increasingly being used by researchers, and our dataset is constantly expanding, but still, the amount of available data is much lower than in the case of other nutrients. The δ¹⁸O-PO₄ values, measured for potential natural and anthropogenic sources of P, which could be delivered to water bodies, are shown in Figure 3.

The information about isotope fractionations associated with several important reactions and transformations occurring in the phosphorus cycle provides a basis for the interpretation of isotope data (δ¹⁸O-PO₄) obtained from phosphate analyses of the natural environment. Without biological activity, the isotope exchange between phosphate oxygen and water (or other solutions) is very slow and is negligible for most environmental applications. Other abiotic processes, such as transport by water or air, the precipitation/dissolution of phosphate minerals (apatite), or the adsorption/desorption of P to/from mineral surfaces, are related to very small isotopic effects and are usually omitted in environmental studies.

We observe a different situation in the case of processes involving micro-organisms. In the case of biological processes, both intracellular and extracellular processes, significant isotopic fractionation can be observed, from several to even several dozen per mill. More details can be found, for example, in the literature from the last few decades, including Paytan and McLaughlin [8], McLaughlin et al. [9], Tamburini et al. [10], and von Sperber et al. [37].

The most important enzymatic process in the aquatic P cycle, controlling δ¹⁸O in the environment, is the intracellular activity of pyrophosphatase, which involves an equilibrated isotopic exchange [8,9,10]. In this case the pyrophosphatase enzyme catalyzes the hydrolysis of pyrophosphate (P₂O₇); during this reaction, one atom of oxygen from the surrounding water is incorporated into the P₂O₇ molecule, and, as a consequence, two inorganic orthophosphate molecules are formed and subsequently released. The exchange of O isotopes, in this case, is subject to a thermodynamic isotopic fractionation, leading to a temperature-dependent equilibrium between the water and phosphate, as described by the empirical equation of Longinelli and Nuti [38], $T=111.4-4.3·\left({\mathrm{\delta }}^{18}\mathrm{O}\cdots \mathrm{P}{\mathrm{O}}_4-{\mathrm{\delta }}^{18}\mathrm{O}\cdots {\mathrm{H}}_2\mathrm{O}\right)$, where T denotes the temperature in °C, and δ¹⁸O$\cdots$PO₄ and δ¹⁸O$\cdots$H₂O denote the oxygen isotopic composition in the phosphate and in water, respectively, expressed in ‰, with respect to the VSMOW. Therefore, due to measurements of the water temperature and oxygen isotopic composition, the calculation of the equilibrium value of δ¹⁸O in phosphate ions is possible.

Usually, phosphates studied in water are not in an isotope equilibrium with the surrounding H₂O (the measured δ¹⁸O-PO₄ differs from the estimated equilibrium δ¹⁸O-PO₄), which indicates that the complete intracellular biological cycle of the orthophosphate does not take place, and the “isotope record” of the source can be preserved. Deviations from the equilibrium values are associated with the mixing of waters with phosphates from natural or anthropogenic sources [8,9]. The values of δ¹⁸O-PO₄ for various local potential phosphate sources, which can supply phosphates to the examined waters, should be also determined during research. A more detailed discussion can be found, for example, in the review article by Gebus-Czupyt and Wach [39].

2. Study Area

Our study focuses on selected areas of the Vistula and Bug interfluve of south-eastern Poland (see Figure 4), where Cretaceous or Quaternary carbonate rocks develop the main useful water-bearing level [40]. The main groundwater reservoir feeding the rivers in this region is developed by cracked formations of the Maastricht and Tertiary developed in the form of opokas, marls, chalk, gaizes, limestones, and sands [18]. In the north-western part of this region, the water-bearing horizon occurs in Paleocene gaizes and marly limestones, and in the southern part, in Neogene limestones and sandstones. In the Lublin Upland and Roztocze, for about 37% of the springs in this area, the water flows from Quaternary sediments and 63% from fissure rocks, including 56% from Upper Cretaceous carbonate formations and 7% from Tertiary rocks [41]. The occurrence of particular rock types is visibly differentiated both in their vertical profile and on a regional basis, and the existing rock layers decelerate the infiltration and formation of upper levels (see, e.g., Michalczyk et al. [40] and the references cited therein). As has been studied for this area, almost 75% of river runoffs come from underground resources, whereas during dry periods, even 100% of the river water comes from groundwater resources [42].

The Lublin Upland and Roztocze cover an area of approximately 12,200 km², and this region is one of the richest in springs (it is second only to the Tatras) [43]. About 1550 springs were recorded in this area, and nearly 80% of them are still functioning. In this area, springs with different discharge levels are present, from springs with a small yield, <1 dm³/s (the most common, about 57% of the studied springs), to springs a with discharge of above 10 dm³/s (13% of springs), with 11 springs permanently exceeding 100 dm³/s, most of which are located in Roztocze. The most abundant springs with a high discharge (continuously or periodically exceeding 100 dm³/s, with a maximum discharge level of 400 dm³/s) are supplied from the Cretaceous aquifer [42].

A total of 35 samples were taken for our reconnaissance research: 16 water samples from springs, 9 from rivers, and 6 from WWTP effluents, as well as 2 samples of drainage water and 2 phosphate rocks. All collection points are marked in Figure 4.

Figure 4

The main elements of the geological structure of our study area, Lublin Upland and Roztocze (based on Dobrowolski et al. [44] and the references therein), with marked sampling points (yellow = spring water, blue = riverine water, brown = WWTP effluent, violet = bedrock, green = drainage water).

[Figure omitted. See PDF]

3. Materials and Methods

3.1. Sample Collection

This research provides a preliminary spatial recognition of the variability in phosphate isotopic composition in the study area, which was carried out mainly in the autumn of 2021. The drainage waters were additionally collected in the spring of 2023. Water samples (from springs, rivers, and waste water treatment plant effluents (WWTPEs)) were collected in HDPE bottles and canisters for chemical and isotopic analyses. In the case of spring and riverine samples, we collected 10 dm³ of water for a WWTPE analysis—1 dm³ of the samples was used for an isotopic analysis, and 0.25 dm³ of the samples was used for a chemical analysis. The sampled points are marked in Figure 4.

In order to remove any organic impurities, before sampling the bottles and canisters, samples were rinsed with 3% HCl and then with deionized water. Parameters such as pH, temperature, conductivity, and dissolved oxygen were measured in the field. The chemical analysis was performed in a laboratory in the Department of Hydrology and Climatology at the Institute of Earth and Environmental Sciences at Maria Curie-Sklodowska University in Lublin. The isotopic analysis of δ¹⁸O-PO₄ was performed in the Stable Isotope Laboratory of the Institute of Geological Sciences at the Polish Academy of Sciences in Warsaw.

The studied bedrock samples were collected for preliminary recognition from two different areas with documented high contents of P-bearing minerals. The first sample came from the Annopol area, where numerous concretions occur in Upper Cretaceous glauconitic sandstones. The phosphate rock deposits were discovered in the 1920s. Further studies showed that these deposits contained about 19% of P₂O₅, making them the richest phosphate deposits in Poland [45]. In the discussed rocks, phosphate concretions are composed of quartz crystals and redeposited glauconite, cemented with apatite with an admixture of chalcedony and single-calcite crystals. These concretions are usually found in the vicinity of fossil accumulations [46]. It is assumed that they were formed as a result of the redistribution of dissolved phosphorus due to increased deep-water circulations [47]. The second sample derived from Brzeziny (near Lubartów), where gravel, sand, clay, and kaolin are currently being mined (as reported by, for example, Cywicka [48]). In the second location, the discussed sandstone formations are probably related to glacial processes that served to redeposit abraded minerals and rocks occurring in Pleistocene formations [49]. The geochemical and mineralogical analyses were performed in a laboratory at the Department of Geology, Soil Science and Geoinformation of the Institute of Earth and Environmental Sciences at Maria Curie-Sklodowska University in Lublin.

3.2. Methods

Physicochemical Analysis

The pH, temperature, conductivity, and dissolved oxygen were measured in the field by using a portable meter (YSI Professional). For the determination the concentrations of NO₂⁻, NO₃⁻, SO₄²⁻, and NH₄⁺ ions, the MIC 3 ion chromatograph (Metrohm) was used. The anions were separated on a Metrosep A SUPP5 250 column, and the cations, on a Metrosep C2 150 column. A photometer by HACH DR 900 was used for the determination of the concentration of orthophosphate ions (PO₄³⁻) (using the Hach 8048 method). Bicarbonates were determined by titration, with 0.05 M of HCl and bromocresol used as dye indicators. The analysis of performance correctness was verified by a certified reference material (Environment Canada MISSIPPI-14).

3.3. Phosphate Extraction from Water and Rock Samples

3.3.1. Water Samples

Dissolved inorganic phosphates were extracted from water samples using the method described by McLaughlin et al. [12]. As an internal reference material for checking reproducibility of the extraction procedure, a pure chemical reagent, KH₂PO₄ (ROTH), was used, whose average values of measured δ¹⁸O-PO₄ values are equal to 11.7 ± 0.3‰ and 11.5 ± 0.3‰ for the raw and extracted KH₂PO₄ reagent, respectively.

In the first stage, filtration of the samples was performed: for the WWTPE and riverine samples, GF glass filters were used for pre-filtration, and 0.45 µm of MCE membrane filters were used. Subsequently, the DIP was stripped via MagIC (magnesium-induced coprecipitation). Deposited on the bottom of the vessel, the precipitated brucite, Mg(OH)₂, with adsorbed DIP was then transferred to a 250 mL bottle for centrifugation, followed by a removal of the supernatant. The obtained precipitate was dissolved by adding acetic acid and 10 M of nitric acid, and this solution was then buffered at pH 5.5 with 1 M of potassium acetate. Subsequently, a cerium nitrate solution was added to the precipitate cerium phosphate overnight, separated by centrifugation, and rinsed with 0.5 M of potassium acetate to remove the chloride ions. The retained precipitate was dissolved in a nitric acid solution, and next, the solution was mixed with 4 mL of a BioRad AG-50W×8 cation exchange resin and was shaken overnight to remove cerium ions from the solution. The resin was subsequently separated from the solution, which was collected in 50 m centrifuge tubes. In the next step of extraction, a bromothymol blue indicator was added for each sample (to easily check pH levels), as well as 1 mL of ammonium hydroxide and 1 mL 3 M of ammonium nitrate. The pH value was controlled by adding 3 M of nitric acid (pH should be neutral). Finally, the silver phosphate was precipitated by adding a freshly prepared AgNO₃ solution. The precipitated Ag₃PO₄ was rinsed several times and separated by centrifugation and was then dried at 50 °C overnight. The purified, dried silver phosphate was used for the analysis of δ¹⁸O-PO₄.

3.3.2. Bedrock Samples

The bedrock samples were first dried in the air and grounded in an agate mortar and pestle. To perform an isotopic analysis of the δ¹⁸O of phosphates in the samples, a procedure used to extract PO₄³⁻ ions from mammalian enamel/bone samples was used [50]. To check the accuracy and reproducibility of this extraction procedure, the Florida Phosphate Rock standard NIST 120c was used, for which the estimated δ¹⁸O-PO₄ value ranged from −21.5 ± 0.2‰ to −22.9 ± 0.2‰ during numerous laboratory intercomparisons in recent years (see, e.g., Lécuyer et al. [51] and the references cited therein); in our study, the average δ¹⁸O-PO₄ value of the Florida Phosphate Rock standard was −22.3 ± 0.2‰. Due to the similar type of materials analyzed, this was also applied to the rock samples collected from the vicinities of Annopol and Brzeziny.

In the first stage, approximately 2 g of the rock samples was placed in a plastic vial, a solution of 30% perhydrol was added for 24 h to remove any organic impurities. After centrifugation and the removal of the supernatant, the solution was rinsed several times with ultrapure water until neutral pH was achieved. To remove the carbonate fraction, 0.1 M of acetic acid was added, and after 72 h, as before, the solution was rinsed several times with ultrapure water, until neutral pH was achieved, and finally, the supernatant was discarded. Then, 2 M of HF was added to the sample for 24 h, after which the solid fraction was separated by centrifugation, and the supernatant was used in the next step of the analytical protocol. Then, NH₄OH was added to the solution to obtain neutral pH, and phosphate ions precipitated in the form of Ag₃PO₄ by adding a silver nitrate solution. The precipitated silver phosphate was then separated by centrifugation and rinsed several times by ultrapure water. The purified Ag₃PO₄ precipitate was then dried at a temperature of 50 °C in an oven.

3.3.3. Isotopic Analysis of δ¹⁸O-PO₄ in Phosphates

The analysis of the oxygen isotopic composition in phosphates (δ¹⁸O-PO₄) were performed using the Flash EA 1112 HT elemental analyzer coupled with the ConFlo IV peripheral and Delta V Advantage IRMS (Thermo Scientific, Waltham, MA, USA) in a continuous flow mode. The silver capsule with the phosphate sample (about 0.5 mg of Ag₃PO₄) was decomposed at a temperature of 1400 °C in the presence of C. The obtained separated and purified CO gas was then introduced into a mass spectrometer, in which its isotopic composition was measured. The measurement results were then normalized to three international standards, USGS 80 (δ¹⁸O = 13.1‰), B 2207 (δ¹⁸O = 21.7‰), and USGS 81 (δ¹⁸O = 35.4‰), which are reported here relative to the VSMOW. The internal standard was measured several times during each measurement cycle to control the quality of the analysis. The measurement precision and reproducibility of the results were generally better than ± 0.3‰.

4. Results and Discussion

In this study, we analyzed different types of water samples, including spring, river, and drainage waters and sewage treatment plant effluents, as well as bedrock samples. The results of the measured basic physical and chemical parameters of the collected samples are presented in Table 1. The content of the phosphates and the average values of the δ¹⁸O-PO₄ of the collected samples are presented in Table 2. The measured δ¹⁸O-PO₄ values for the different types of water samples in this study, compared with literature data, are presented in Figure 5.

The studied springs belong to moderately mineralized waters, with a slightly alkaline pH and a predominance of HCO₃–Ca ions, which are the result of the dissolution of carbonate rocks. The discharge of the springs varied in a wide range, from 7.5–8.2 dm³/s (springs in Bezek and Nowosiółki) to 93 dm³/s (the spring in Wąwolnica). The HCO₃⁻ content was in the range of 230–461mg/dm³, while the concentration of Ca²⁺ and Mg²⁺ ions varied from 92 to 140 mg/dm³ and from 1 to 24 mg/dm³, respectively. The content of substances such as nitrates, sulphates, or chlorides, usually considered indicators of environmental pollution, was relatively low, from 6 to 28 mg/dm³ in the case of NO₃⁻ ions, from 13 to 56 mg/dm³ for SO₄²⁻ ions, and from 5 to 27 mg/dm³ in the case of Cl⁻ ions. The content of Na⁺ and K⁺ ions did not exceed 9 mg/dm³ and 5 mg/dm³, respectively, while the average content of fluoride ions was 0.1 mg/dm³ and did not exceed 0.3 mg/dm³. Only in two springs were trace amounts of NO₂⁻ ions detected (0.1 mg/dm³ in the Bezek and Sulów springs), which proves the relatively high quality of the examined spring waters. All spring samples were characterized by a relatively high content of phosphate ions, ranging from 0.2 to 1.1 mg/dm³, while the average concentration of PO₄³⁻ ions was estimated to be 0.5 mg/dm³. The highest orthophosphate concentrations were detected in springs, whose bedrocks are opokas (see Table 2). The δ¹⁸O-PO₄ values varied from +14.8 to +18.5‰ and depended on the sampling site and type of bedrock. In the Husiny spring, with the highest content of orthophosphates (1.1 mg/dm³), the lowest value of δ¹⁸O-PO₄ was measured, while for the remaining springs, whose bedrocks were mainly opokas, a δ¹⁸O-PO₄ value in the range from +16 to +17‰ predominated.

The studied rivers included rivers that are right-bank tributaries of Vistula, Wyżnica, and Wieprz (including its tributaries, the Łabuńka and Wolica rivers). Two small rivers flow into Bystrzyca (approximately 22.75 km and approximately 32 km long, respectively), as well as a left-bank tributary of the Bug and Huczwa rivers. The discharge of these rivers varied in a wide range, from 153 dm³/s (Tuszów) to 12,600 dm³/s (Wieprz, in the town of Krasnystaw), depending on the sampling sites and collection period (between July and December 2021). All rivers had a slightly alkaline pH and a conductivity that ranged from 381 µS/cm (Wieprz in the town of Guciów) to 713 µS/cm (Łabuńka, in the town of Krzak). The average orthophosphate concentration in these rivers was estimated to be 0.4 mg/dm³, while the highest content of PO₄³⁻ ions was detected in July 2021 in the Czerniejówka river (0.7 mg/dm³). The δ¹⁸O-PO₄ values varied from +10.3 to +18.6‰, while the higher values from this range predominated.

For the identification of δ¹⁸O in dissolved inorganic phosphates extracted from sewage treatment plant effluents, a few nearby WWTPs were selected: those in Celejów, Krasnobród, Krasnystaw, Piaski, Piotrowice, and Urzędów (marked as numbers 26 to 31 in Figure 4). All samples were characterized by high conductivity and contents of dissolved components. The highest phosphate content was determined for the WWTP effluents collected in September 2021 (sample no. 26 and no. 29, 26.5 mg/dm³ and 16.90 mg/dm³, respectively, see Table 1 and Table 2). The concentration of PO₄³⁻ ions in other WWTP samples varied from 3 mg/dm³ to 10 mg/dm³ (samples collected between October and December 2021). Higher orthophosphate concentrations were recorded in September compared to subsequent months, which is typical for urbanized areas, where an increase in sewage production during the summer period can be observed. The δ¹⁸O-PO₄ values varied from +12.4 to +15.6‰, while the lower values from this range predominated.

The analyzed two drainage water samples, collected in May 2023, were significantly different. The first one, collected from a drainage hole near a small airport in Świdnik, with a conductivity equal to 1665 µS/cm, contained a large amount of dissolved components, including ions, usually indicating anthropogenic pollution from nitrates, chlorides, sodium, and potassium, whose concentrations were approximately 203 mg/dm³, 487 mg/dm³, 269 mg/dm³, and 98 mg/dm³. Also, the nitrite concentration was very high (1.2 mg/dm³), while the measured orthophosphate concentration was 1.9 mg/dm³. The second sample, collected in Wilkołaz, with a conductivity of 578 µS/cm, contained much less of all tested compounds compared to the first sample (see Table 1), and its orthophosphate content was, ca., 0.2 mg/dm³. Despite the different natures of both samples, the δ¹⁸O-PO₄ values were similar (+16.7 and +17.3‰, respectively).

An X-ray analysis confirmed the presence of phosphate minerals (fluorapatite and apatite) in both collected bedrock samples. Depending on the analyzed phase, the content of P ranged from 2% (quartz and glauconite) to 13% (francolite) in the concretion collected from the Annopol area, while the average δ¹⁸O-PO₄ value was found to be at +21.2 ± 0.3‰. The second sample, collected from Brzeziny, contained significant amounts of apatite (about 9.3% of P was detected), while the average δ¹⁸O-PO₄ value was +20.5 ± 0.5‰. An isotopic signature is dependent on the formation mechanisms of bedrocks. The various ranges of δ¹⁸O-PO₄ variabilities of phosphates minerals derived from bedrocks of marine sedimentary origin, from bedrocks associated with guano and igneous deposits, have been collected, and these are further discussed in Smith et al. [53]. The results obtained in this study (+21.2 ± 0.3‰ and +20.5 ± 0.5‰) are in the range of the existing data on sedimentary bedrocks (from about +8‰ to, ca., +25‰), and they are close to the values of samples derived from similar geological periods (younger sedimentary bedrock samples have a higher δ¹⁸O-PO₄ than those of older ones [53]).

The confirmed presence of phosphate minerals in the examined bedrock samples and the high content of orthophosphates in the spring waters from this area, with the simultaneous absence/very small visibility of an anthropogenic factor, confirmed by the results of chemical analyses, indicate that the leaching of phosphorus from bedrocks is the main source of phosphate ions in the spring waters of the Lublin Upland and Roztocze. At the same time, it should be noted that the leaching of phosphates from a bedrock depends on several factors, such as its chemical composition, the presence of Fe or similar oxides and hydroxides, or the pH of the environment; these relationships are the subject of our future investigations.

During spatial reconnaissance, no additional analyses of the oxygen isotopic composition of the water samples were performed, apart from several test samples that were collected from the rivers. The estimated δ¹⁸O-PO₄ values from the Longinelli and Nuti equation equilibrium were, in all cases, higher than the measured δ¹⁸O-PO₄, which indicates that a full intracellular cycle did not take place, and the studied waters were mixed with phosphates derived from other P sources with lower δ¹⁸O values. These were probably both phosphates introduced from local spring waters and effluents from nearby waste water treatment plants.

Information about the isotopic composition of phosphate oxygen has led to more and more applications of it, both for tracking processes in the P cycle in the environment and for determining the origin of phosphates in water bodies. Research regarding the δ¹⁸O-PO₄ value in P-bearing bedrocks is also becoming increasingly important, especially in soil studies, where this isotopic signature is used as a proxy for the biological phosphorus cycle. These data can be very useful, especially when it is impossible to collect a sample of source bedrocks or applied mineral fertilizers for isotopic analyses. Phosphates derived from the leaching of phosphate minerals can also be an important source of PO₄³⁻ for surface waters, and this subject is a topic of our future research.

As can be seen, the oxygen isotopic composition of inorganic phosphates has rather narrow ranges of variability (see Figure 3 and Figure 5), and these ranges may partially overlap for some sources. Therefore, special attention must be paid to identifying local hydrogeological conditions and potential sources of phosphates for this study area, both of natural and anthropogenic origin. It is also worth noting that the global database on δ¹⁸O of inorganic phosphates, despite the constant increase in researchers’ interests in this topic, is still relatively poor. Most publications in this field concern research in the USA, although in recent years, there has been an increase in the research conducted in Europe, which relates to the topic of eutrophication (e.g., Gruau et al. [13] and Gooddy et al. [54]), drinking water supplies in England and Wales [55], and case studies of rivers in the United Kingdom [56]. Both an expansion of the global database to include information on the isotopic composition of P_i oxygen, e.g., phosphorus fertilizers derived from various P-bedrock reserves, as well as advanced studies of seasonal and daily variabilities of the δ¹⁸O-PO₄ values of sewage waters are still needed, as was suggested by, e.g., Davies et al. [52]. In this paper, we add new data from Poland to the global database.

5. Conclusions

Here, we presented novel data of the δ¹⁸O of dissolved inorganic phosphates extracted from different types of waters from the Lublin Upland and Roztocze (Poland), with additional results for P-bearing bedrock samples from the same study area. The preliminary results of our study show slightly different ranges of δ¹⁸O-PO₄ values for various phosphate sources: spring waters, rivers, drainage waters, WWTP effluents, and bedrocks, indicating that the oxygen isotopic signature could be useful tool to recognize phosphate sources and P cycling in the studied area. However, in contrast to other most important environmental elements, such as C, N, or S, in the case of phosphates, researchers only have one marker (oxygen isotopic composition), and local potential sources of P should be particularly well-recognized.

The results of δ¹⁸O-PO₄ measurements of the examined springs showed a rather narrow range of δ¹⁸O variability, ranging from +14.8 to +18.5‰, with dominant values from the middle of this range for samples with opokas as bedrocks. For the riverine samples, the obtained results are in a wider range, from +10.3‰ to +18.6‰, while the higher values from this range predominated. In the case of the WWTP effluents, the lowest values of δ¹⁸O-PO₄ were observed, from +12.4 to +15.6‰, with a predominance of lower values in this range. In the case of the two analyzed bedrock samples, the δ¹⁸O-PO₄ values were similar (+20.5‰ and +21.7‰) and close to the existing data on sedimentary bedrocks derived from similar geological periods.

The undertaken research provides a preliminary insight into the spatial oxygen isotopic composition of dissolved inorganic phosphates in the studied area, covering the Vistula and Bug interfluve. Ongoing studies are being conducted, with a particular emphasis on assessing the qualitative and quantitative role of phosphates leached from sedimentary bedrocks on the total pool of available P in the groundwater and surface waters of this study area.

Author Contributions

B.G.-C.—conceptualization, methodology, chemical preparation and isotopic analysis, validation, investigation, resources, visualization, and writing—original draft preparation; S.C.—conceptualization, methodology, validation, investigation, data curation, and writing—review and editing; M.K., M.H. and M.R.—laboratory analysis, data curation, and writing—review and editing; J.S.—visualization, laboratory analysis, data curation, and writing—review and editing; K.S., M.P. and B.Z.—field measurements and writing—review and editing. All authors have read and agreed to the published version of the manuscript.

Data Availability Statement

The original contributions presented in the study are included in the article, further inquiries can be directed to the corresponding author.

Conflicts of Interest

The authors declare no conflicts of interest.

Footnotes

Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Figures and Tables

Figure 1. Schematic diagram of the phosphorus cycle, showing phosphorus reservoirs (living—green boxes; nonliving—gray boxes); physical transport pathways (blue arrows); and microbially mediated transformations (green arrows). Based on Mackey and Paytan [1].

Figure 3. Ranges of δ18O-PO4 values for some different P sources (simplified from Tamburini et al. [10]). Animal feces also include guano, detergents also include toothpaste, and high T rocks represent δ18O-PO4 from lithogenic material formed at a high temperature (e.g., igneous and volcanic rocks).

Figure 5. The range of δ18O-PO4 values within aquatic systems as adapted from Davies et al. [52], with results from this study.

Table 1

Basic physical and chemical parameters of the collected water samples.

Sampling Date	Sample No.	Sample Name	Q[dm3/s]	T [°C]	pH	Conductivity [μS/cm]	Dissolved O2		ORP [mV]	Na+[mg/dm3]	NH4+[mg/dm3]	K+[mg/dm3]	Ca2+ [mg/dm3]	Mg2+ [mg/dm3]	HCO3− [mg/dm3]	F−[mg/dm3]	Cl−[mg/dm3]	NO2−[mg/dm3]	NO3−[mg/dm3]	SO42− [mg/dm3]
							[mg/dm3]	[%]
Springs
14 October 2021	1	Bezek	7.5	9.6	7.5	596	7.5	67.2	104.0	1.1	<0.01	1.7	130.5	0.9	317.8	0.1	20.3	0.1	21.1	21.5
29 September 2021	2	Celejów	44.7	9.2	7.5	678	2.3	21.4	102.0	7.0	<0.01	1.9	129.6	24.2	461.2	0.3	14.4	<0.01	7.0	29.0
22 November 2021	3	Dębiny	16.9	9.1	7.7	446	6.1	55.5	131.0	1.3	<0.01	1.0	104.8	2.0	271.2	0.1	6.2	<0.01	10.8	33.9
30 September 2021	4	Gałęzów	34.0	9.1	7.6	563	10.1	88.5	107.7	1.7	<0.01	1.0	121.0	3.2	316.6	0.1	14.9	<0.01	20.2	13.2
23 November 2021	5	Husiny	42.0	8.8	7.5	487	3.1	27.0	94.6	1.6	<0.01	1.2	92.2	1.4	229.7	0.1	5.2	<0.01	18.7	28.9
22 November 2021	6	Krasnobród	39.7	8.8	7.7	431	5.9	52.6	135.0	2.2	<0.01	1.3	100.1	3.1	285.0	0.1	6.5	<0.01	14.5	20.4
22 November 2021	7	Łabunie	24.3	9.5	7.1	692	4.3	40.3	150.0	8.3	<0.01	5.0	136.4	16.2	405.1	0.2	27.3	<0.01	17.5	23.7
23 November 2021	8	Łosiniec	46.7	8.7	7.4	495	3.2	28.5	104.4	2.9	<0.01	1.6	96.5	3.1	262.3	0.1	5.4	<0.01	7.7	25.9
14 October 2021	9	Nowosiółki	8.2	9.5	7.5	606	8.0	71.4	109.0	2.1	<0.01	2.6	140.0	2.8	330.0	0.1	26.9	<0.01	27.6	27.3
29 September 2021	10	Pliszczyn	52.9	9.5	7.2	635	6.1	53.6	96.0	5.9	<0.01	1.4	129.8	22.1	420.9	0.0	22.5	<0.01	10.0	34.2
29 September 2021	11	Rogalów	62.3	9.6	7.5	684	6.1	53.6	67.3	8.3	<0.01	3.2	129.2	19.4	416.4	0.2	16.8	<0.01	5.9	37.7
13 October 2021	12	Słodków	41.6	9.5	7.2	532	4.7	42.4	112.4	5.4	<0.01	2.7	112.3	14.1	372.7	0.1	10.4	<0.01	16.2	15.6
30 September 2021	13	Stryjno Małe	78.0	9.4	7.4	599	7.2	63.4	100.3	4.7	<0.01	1.6	130.4	12.1	390.2	0.0	19.1	<0.01	23.2	24.5
13 October 2021	14	Sulów	57.8	9.3	7.4	546	4.9	43.4	105.2	3.8	<0.01	2.0	112.9	12.8	342.2	0.1	10.0	0.1	14.3	18.2
13 October 2021	15	Święta Otylia (Urzędów)	17.5	10.5	7.4	659	4.7	43.5	123.0	4.5	<0.01	2.5	129.7	2.4	338.5	0.1	13.5	<0.01	12.6	29.2
29 September 2021	16	Wąwolnica	93.0	9.5	7.7	692	4.0	35.6	53.7	4.5	<0.01	3.2	133.4	13.4	374.9	0.2	13.9	<0.01	9.2	56.3
Rivers
15 July 2021	1	Czerniejówka (Mętów)	309	14.1	7.6	614	9.4	98.6	143.0	5.2	<0.01	2.5	108.8	5.8	286.5	0.0	22.7	<0.01	12.6	26.0
06 December 2021	18	Huczwa (Werbkowice)	3064	10.4	8.3	747	12.1	88.9	116.5	12.2	0.3	4.7	151.2	18.1	535.7	0.2	22.5	<0.01	3.3	24.4
15 July 2021	19	Kosarzewka (Bychawka)	1201	14.9	7.8	560	11.1	110.0	172.0	4.6	<0.01	1.9	104.6	8.1	365.5	0.1	19.6	<0.01	10.7	19.9
23 November 2021	20	Łabuńka (Krzak)	2175	6.3	7.9	713	10.1	84.0	76.0	15.8	<0.01	5.4	133.6	12.4	409.1	0.0	37.5	0.2	12.8	29.7
23 November 2021	21	Wieprz (Guciów)	1030	7.0	7.9	381	10.1	88.0	98.3	5.2	0.1	2.4	89.8	5.4	264.1	0.1	8.1	<0.01	5.5	17.2
06 December 2021	22	Wolica	750	2.4	8.0	645	12.6	95.5	94.0	9.3	<0.01	3.0	146.3	22.2	525.7	0.0	11.5	<0.01	3.4	9.9
13 October 2021	23	Wyżnica	880	9.0	7.9	497	11.6	101.2	114.0	8.6	<0.01	3.7	105.7	6.9	296.7	0.1	18.1	<0.01	8.1	21.6
15 July 2021	24	Tuszów	153	11.4	7.5	514	10.9	110.2	104.3	3.5	<0.01	1.5	101.9	5.4	286.1	0.1	16.6	<0.01	18.4	22.0
06 December 2021	25	Wieprz (Krasnystaw)	12,600	2.7	7.8	627	12.2	92.2	117.0	8.3	<0.01	3.0	117.3	12.0	387.8	0.1	16.2	0.1	7.4	18.8
WWTP effluents
29 September 2021	26	Celejów	-	16.1	7.5	1450	4.8	49.0	13.5	133.8	25.4	42.4	193.2	28.5	698.6	0.2	174.8	6.5	52.3	58.5
23 November 2021	27	Krasnobród	-	11.0	7.4	1143	6.8	62.6	110.0	115.9	3.3	33.6	134.7	2.9	405.8	0.1	132.1	5.4	11.4	68.0
06 December 2021	28	Krasnystaw	-	11.3	7.6	1780	6.1	56.6	122.0	167.8	3.6	36.4	176.4	21.3	757.3	0.0	175.5	5.4	25.2	70.0
30 September 2021	29	Piaski	-	12.8	7.4	1540	1.8	18.3	86.0	95.4	72.5	29.3	145.7	12.6	749.3	0.1	117.7	0.1	1.0	30.0
13 October 2021	30	Piotrowice	-	17.9	7.1	1380	3.3	36.3	9.0	208.3	<0.01	42.7	142.6	28.3	547.9	0.3	272.5	5.2	31.1	90.0
13 November 2021	31	Urzędów	-	15.3	7.6	1440	6.2	63.5	85.0	104.9	86.8	29.7	146.6	4.4	789.0	0.1	110.6	<0.01	0.3	57.8
Drainage waters
31 May 2023	32	Świdnik (near airport)	-	-	-	1665	-	-	-	269.1	<0.01	97.7	198.0	26.9	430.7	0.0	486.7	1.2	203.0	13.8
31 May 2023	33	Wilkołaz	-	-	-	578	-	-	-	26.9	<0.01	2.5	114.6	7.7	218.4	0.1	66.6	0.2	41.0	41.5

Table 2

The results of the isotopic analysis of the δ¹⁸O- PO₄ of the collected samples.

Sampling Date	SampleNo.	Localization and Kind of Collected Sample	PO43− [mg/dm3]	δ18O-PO4 av [‰]	Bedrock Type
Springs
14 October 2021	1	Bezek	0.33	15.3 ± 0.3	Opokas
29 November 2021	2	Celejów	0.30	16.4 ± 0.6	Gaizes
22 November 2021	3	Dębiny	0.21	15.6 ± 0.5	Opokas
30 November 2021	4	Gałęzów	0.21	18.5 ± 0.4	Opokas
23 November 2021	5	Husiny	1.09	14.8 ± 0.4	Gaizes
22 November 2021	6	Krasnobród	0.76	16.0 ± 0.3	Opokas
22 November 2021	7	Łabunie	0.41	16.9 ± 0.4	Marls
23 November 2021	8	Łosiniec	0.59	17.8 ± 0.6	Opokas
14 October 2021	9	Nowosiółki	0.34	17.8 ± 0.5	Opokas
29 November 2021	10	Pliszczyn	0.54	16.4 ± 0.4	Gaizes
29 November 2021	11	Rogalów	0.21	16.3 ± 0.3	Gaizes
13 October 2021	12	Słodków	0.34	16.5 ± 0.2	Opokas
30 November 2021	13	Stryjno Małe	0.58	17.5 ± 0.3	Opokas
13 October 2021	14	Sulów	0.49	15.7 ± 0.3	Opokas
13 October 2021	15	Święta Otylia (Urzędów)	0.38	15.7 ± 0.5	Opokas
29 November 2021	16	Wąwolnica	0.54	16.7 ± 0.2	Gaizes
Rivers
15 July 2021	17	Czerniejówka (Mętów)	0.65	15.2 ± 0.5	Opokas
06 December 2021	18	Huczwa (Werbkowice)	0.23	14.9 ± 0.4	Marls/chalks
15 July 2021	19	Kosarzewka (Bychawka)	0.40	13.4 ± 0.2	Opokas
23 November 2021	20	Łabuńka (Krzak)	0.42	15.8 ± 0.5	Opokas/marls/chalks
23 November 2021	21	Wieprz (Guciów)	0.64	18.6 ± 0.4	Gaizes/opokas
06 December 2021	22	Wolica (Orłów Drewniany)	0.30	16.2 ± 0.3	Opokas/chalks
13 October 2021	23	Wyżnica (Bór)	0.30	10.3 ± 0.3	Opokas
15 July 2021	24	Tuszów (Tuszów)	0.45	12.9 ± 0.3	Gaizes/opokas
06 December 2021	25	Wieprz (Krasnystaw)	0.43	16.7 ± 0.3	Opokas/marls/chalks
WWTP effluents
29 November 2021	26	Celejów	26.50	12.4 ± 0.3	-
23 November 2021	27	Krasnobród	9.90	15.6 ± 0.3	-
06 December 2021	28	Krasnystaw	3.20	13.2 ± 0.3	-
30 November 2021	29	Piaski	16.90	12.4 ± 0.4	-
13 October 2021	30	Piotrowice	7.70	14.5 ± 0.1	-
13 November 2021	31	Urzędów	3.00	12.9 ± 0.2	-
Drainage waters
31 May 2023	32	Świdnik (near airport)	1.90	16.6 ± 0.3	-
31 May 2023	33	Wilkołaz	0.16	17.3 ± 0.4	-
Bedrocks
20 July 2022	34	Phosphorite concrection near Annopol	-	21.2 ± 0.3	Sandstone
19 July 2022	35	Phosphorite from Brzeziny (near Lubartów)	-	20.5 ± 0.5	Sands

References

1. Mackey, K.R.M.; Paytan, A. Phosphorus Cycle. Encyclopedia of Microbiology; 3rd ed. Schaechter, M. Academic Press: Cambridge, MA, USA, 2009; pp. 322-334. [DOI: https://dx.doi.org/10.1016/B978-012373944-5.00056-0]

2. Tiessen, H. Phosphorus in the global environment. The Ecophysiology of Plant-Phosphorus Interactions: Plant Ecophysiology; White, P.J.; Hammond, J.P. Springer: Dordrecht, The Netherlands, 2008; Volume 7, [DOI: https://dx.doi.org/10.1007/978-1-4020-8435-5_1]

3. O’Neill, P. Environmental Chemistry; 2nd ed. Chapman & Hall: London, UK, 1993.

4. Filippelli, G.M. The Global Phosphorus Cycle: Past, Present, and Future. Elements; 2008; 4, pp. 89-95. [DOI: https://dx.doi.org/10.2113/GSELEMENTS.4.2.89]

5. Vollenweider, R.A. The Scientific Basis for Lake and Stream Eutrophication with Particular Reference to Phosphorus and Nitrogen as Eutrophication Factors; Technical Report DAS/DS1/68.2 Organization for Economic Cooperation and Development: Paris, France, 1968.

6. Regulation on the Classification of Ecological Status, Ecological Potential, Chemical Status and the Method of Classifying the Status of Surface Water Bodies as Well as Environmental Quality Standards for Priority Substances. Available online: https://leap.unep.org/en/countries/pl/national-legislation/regulation-classification-ecological-status-ecological-1 (accessed on 25 September 2024).

7. Rączka, J.; Skąpski, K.; Tyc, T. Water Resources in Poland—Protection and Utilization; Fundacja Przyjazny Kraj: Warszawa, Russia, 2021; pp. 5-47. (In Polish)

8. Paytan, A.; McLaughlin, K. The Oceanic Phosphorus Cycle. Chem. Rev.; 2007; 107, pp. 563-576. [DOI: https://dx.doi.org/10.1021/cr0503613] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/17256993]

9. McLaughlin, K.; Young, M.B.; Paytan, A.; Kendall, C. The Oxygen Isotopic Composition of Phosphate: A Tracer for Phosphate Sources and Cycling; IAEA-TECDOC-1695 IAEA: Vienna, Austria, 2013; pp. 93-110.

10. Tamburini, F.; Pfahler, V.; von Sperber, C.; Frossard, E.; Bernasconi, S.M. Oxygen Isotopes for Unraveling Phosphorus Transformations in the Soil–Plant System: A Review. Soil Sci. Soc. Am. J.; 2014; 78, pp. 38-46. [DOI: https://dx.doi.org/10.2136/sssaj2013.05.0186dgs]

11. Blake, R.E.; O’Neil, J.R.; Surkov, A.V. Biogeochemical cycling of phosphorus: Insights from oxygen isotope effects of phosphoenzymes. Am. J. Sci.; 2005; 305, pp. 596-620. [DOI: https://dx.doi.org/10.2475/ajs.305.6-8.596]

12. McLaughlin, K.; Kendall, C.; Silva, S.; Stuart-Williams, H.; Paytan, A. A precise method for the analysis of δ¹⁸O of dissolved inorganic phosphate in seawater. Limnol. Oceanogr. Methods; 2004; 2, pp. 202-212. [DOI: https://dx.doi.org/10.4319/lom.2004.2.202]

13. Gruau, G.; Legeas, M.; Riou, C.; Gallacier, E.; Martineau, F.; Hénin, O. The oxygen isotope composition of dissolved anthropogenic phosphates: A new tool for eutrophication research?. Water Res.; 2005; 39, pp. 232-238. [DOI: https://dx.doi.org/10.1016/j.watres.2004.08.035]

14. Elsbury, K.E.; Paytan, A.; Ostrom, N.E.; Kendall, C.; Young, M.B.; McLaughlin, K.; Rollog, M.E.; Watson, S. Using Oxygen Isotopes of Phosphate to Trace Phosphorus Sources and Cycling in Lake Erie. Environ. Sci. Technol.; 2009; 43, pp. 3108-3114. [DOI: https://dx.doi.org/10.1021/es8034126]

15. Pistocchi, C.; Mészáros, É.; Frossard, E.; Bünemann, E.K.; Tamburini, F. In or Out of Equilibrium? How Microbial Activity Controls the Oxygen Isotopic Composition of Phosphate in Forest Organic Horizons with Low and High Phosphorus Availability. Front. Environ. Sci.; 2020; 8, 564778. [DOI: https://dx.doi.org/10.3389/fenvs.2020.564778]

16. Paytan, A.; Roberts, K.; Watson, S.; Peek, S.; Chuang, P.-C.; Defforey, D.; Kendall, C. Internal loading of phosphate in Lake Erie Central Basin. Sci. Total Environ.; 2017; 579, pp. 1356-1365. [DOI: https://dx.doi.org/10.1016/j.scitotenv.2016.11.133]

17. Tcaci, M.; Barbecot, F.; Hélie, J.-F.; Surridge, B.W.J.; Gooddy, D.C. A New Technique to Determine the Phosphate Oxygen Isotope Composition of Freshwater Samples at Low Ambient Phosphate Concentration. Environ. Sci. Technol.; 2019; 53, pp. 10288-10294. [DOI: https://dx.doi.org/10.1021/acs.est.9b00631]

18. Michalczyk, Z.; Wilgat, T. Waters. The Natural Environment of the Lublin Region; Uziak, S.; Turski, R. LTN EN: Brussels, Belgium, 2008; pp. 113-209. (In Polish)

19. Mazurek, S.; Roszkowska-Remin, J.; Bienko, T. New geological criteria for domestic phosphorite deposits—A discussion. Gospod. Surowcami Miner.—Miner. Resour. Manag.; 2024; 40, pp. 63-83. [DOI: https://dx.doi.org/10.24425/gsm.2024.149298]

20. Gebus-Czupyt, B.; Chmiel, S.; Trembaczowski, A.; Wach, B. The studies of nitrates (δ¹⁵N, δ¹⁸O) and phosphates (δ¹⁸O) isotopic composition as a tool for identifying eutrophic water pollution. Proceedings of the 9th National Conference Petrological and Mineralogical Research in Geology; Warsaw, Poland, 28–29 March 2019; (In Polish)

21. Mullen, M.D. Phosphorus in Soils—Biological Interactions. Reference Module in Earth Systems and Environmental Sciences; Elsevier: Amsterdam, The Netherlands, 2019; [DOI: https://dx.doi.org/10.1016/B978-0-12-409548-9.11992-X]

22. Fillipelli, G.M. Phosphorus and Life on a Water World. Geophys. Res. Lett.; 2022; 49, e2021GL097346. [DOI: https://dx.doi.org/10.1029/2021GL097346]

23. O’Sullivan, G.; Chew, D.; Kenny, G.; Henrichs, I.; Mulligan, D. The trace element composition of apatite and its application to detrital provenance studies. Earth-Sci. Rev.; 2020; 201, 103044. [DOI: https://dx.doi.org/10.1016/j.earscirev.2019.103044]

24. Yoder, C.H.; Stepien, K.R.; Dudrick, R.N. The distribution of carbonate in apatite: The environment model. Am. Miner.; 2023; 108, pp. 1072-1079. [DOI: https://dx.doi.org/10.2138/am-2022-8389]

25. Li, J.; Chen, S.-Y.; Zhao, Y.-H. Trace elements in apatite from Gejiu Sn polymetallic district: Implications for petrogenesis, metallogenesis and exploration. Ore Geol. Rev.; 2022; 145, 104880. [DOI: https://dx.doi.org/10.1016/j.oregeorev.2022.104880]

26. Bruand, E.; Fowler, M.; Storey, C.; Darling, J. Apatite trace element and isotope applications to petrogenesis and provenance. Am. Miner.; 2017; 102, pp. 75-84. [DOI: https://dx.doi.org/10.2138/am-2017-5744]

27. Qiu, K.-F.; Zhou, T.; Chew, D.; Hou, Z.-L.; Müller, A.; Yu, H.-C.; Lee, R.G.; Chen, H.; Deng, J. Apatite trace element composition as an indicator of ore deposit types: A machine learning approach. Am. Miner.; 2024; 109, pp. 303-314. [DOI: https://dx.doi.org/10.2138/am-2022-8805]

28. Ruttenberg, K.C. The Global Phosphorus Cycle. Treatise on Geochemistry; Holland, H.D.; Turekian, K.K. Pergamon: Oxford, UK, 2003; pp. 585-643. [DOI: https://dx.doi.org/10.1016/B0-08-043751-6/08153-6]

29. Liu, Y.; Villalba, G.; Ayres, R.U.; Schroder, H. Global Phosphorus Flows and environmental Impacts from a Consumption Perspective. J. Ind. Ecol.; 2008; 12, pp. 229-247. [DOI: https://dx.doi.org/10.1111/j.1530-9290.2008.00025.x]

30. Smil, V. Phosphorus in the environment: Natural flows and human interferences. Annu. Rev. Energy Environ; 2000; 25, pp. 53-88. [DOI: https://dx.doi.org/10.1146/annurev.energy.25.1.53]

31. Schlesinger, W.H. The Global Cycles of Nitrogen and Phosphorus: Biogeochemistry: An Analysis of Global Change; 2nd ed. Academic Press: San Diego, CA, USA, 1997; pp. 383-401.

32. IFA. The Global “4R” Nutrient Stewardship Framework—Developing Fertilizer Best Management Practices for Delivering Economic, Social and Environmental Benefits; International Fertilizer Industry Association: Paris, France, 2009.

33. Sapek, A. Phosphorus in the human food chain and the environment in Poland. Inżynieria Ekol.; 2009; 21, pp. 62-72. (In Polish)

34. Turner, B.L.; Raboy, V. Phosphorus cycle. AccessScience; McGraw Hill: New York, NY, USA, 2019; [DOI: https://dx.doi.org/10.1036/1097-8542.508930]

35. Yuan, Z.; Jiang, S.; Sheng, H.; Liu, X.; Hua, H.; Liu, X.; Zhang, Y. Human Perturbation of the Global Phosphorus Cycle: Changes and Consequences. Environ. Sci. Technol.; 2018; 52, pp. 2438-2450. [DOI: https://dx.doi.org/10.1021/acs.est.7b03910] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/29402084]

36. Penuelas, J.; Sardans, J. Human-driven global nutrient imbalances increase risks to health. Eco-Environ. Health; 2023; 2, pp. 246-251. [DOI: https://dx.doi.org/10.1016/j.eehl.2023.08.003] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/38435356]

37. Von Sperber, C.; Tamburini, F.; Brunner, B.; Bernasconi, S.M.; Frossard, E. The oxygen isotope composition of phosphate released from phytic acid by the activity of wheat and Aspergillus niger phytase. Biogeosciences; 2015; 12, pp. 4175-4184. [DOI: https://dx.doi.org/10.5194/bg-12-4175-2015]

38. Longinelli, A.; Nuti, S. Revised phosphate-water isotopic temperature scale. Earth Planet. Sci. Lett.; 1973; 19, pp. 373-376. [DOI: https://dx.doi.org/10.1016/0012-821X(73)90088-5]

39. Gebus-Czupyt, B.; Wach, B. Application of δ¹⁸O-PO₄ analysis to recognize phosphate pollutions in eutrophic water. Ecohydrol. Hydrobiol.; 2022; 22, pp. 21-39. [DOI: https://dx.doi.org/10.1016/j.ecohyd.2021.05.005]

40. Michalczyk, Z.; Chmiel, S.; Głowacki, S.; Zielińska, B. Changes of springs’ yield of Lublin Upland and Roztocze Region in 1998–2008. J. Water Land Dev.; 2008; 12, pp. 113-125. [DOI: https://dx.doi.org/10.2478/v10025-009-0009-6]

41. Michalczyk, Z. Źródła Wyżyny Lubelskiej i Roztocza; Wydawnictwo UMCS: Lublin, Poland, 2001; pp. 1-298. (In Polish)

42. Michalczyk, Z.; Chmiel, S.; Głowacki, S.; Sposób, J.; Zielińska, B. Discharge of the springs of the Lublin Upland and Roztocze. Ecohydrol. Hydrobiol.; 2020; 20, pp. 599-609. [DOI: https://dx.doi.org/10.1016/j.ecohyd.2020.07.003]

43. Chełmicki, W.; Jokiel, P.; Michalczyk, Z.; Moniewski, P. Distribution, discharge and regional characteristics of springs in Poland. Episodes; 2011; 34, pp. 244-256. [DOI: https://dx.doi.org/10.18814/epiiugs/2011/v34i4/003]

44. Dobrowolski, R.; Harasimiuk, M.; Brzezińska-Wójcik, T. Structural control on the relief in the Lublin Upland and the Roztocze region. Przegląd Geol.; 2014; 62, pp. 51-56. (In Polish)

45. Available online: https://www.pgi.gov.pl/jubileusz/100-naj/wydarzen/11310-odkrycie-fosforytow-w-rejonie-annopola-rachowa.html (accessed on 24 March 2024).

46. Machalski, M.; Komorowski, A.; Harasimiuk, M. New perspectives of exploration of Cretaceous marine vertebrates in the abandoned phosphate mine in Annopol on the Vistula. Przegląd Geol.; 2009; 57, pp. 638-641. (In Polish)

47. Bardet, N.; Fischer, V.; Machalski, M. Large predatory marine reptiles from the Albian–Cenomanian of Annopol, Poland. Geol. Mag.; 2016; 153, pp. 1-16. [DOI: https://dx.doi.org/10.1017/S0016756815000254]

48. Cywicka, K. Geological Report on Reconnaissance Research for Clay Raw Material for the Production of Building Ceramics in the Eastern Part of the Lublin Voivodeship, in the Area of the Following Towns: Leszkowice, Brzeziny, Serniki, Czerniejów; Milejów Central Geological Archive of the Polish Geological Institute: Warsaw, Poland, 1980; (In Polish)

49. Krawczyk, M.; Ryzner, K.; Skurzyński, J.; Jary, Z. Lithological indicators of loess sedimentation of SW Poland. Contemp. Trends Geosci.; 2017; 6, pp. 94-111. [DOI: https://dx.doi.org/10.1515/ctg-2017-0008]

50. Dąbrowski, P.; Kulus, M.; Grzelak, J.; Radzikowska, M.; Oziembłowski, M.; Domagała, Z.; Krajcarz, M.T. Assessing weaning stress—Relations between enamel hypoplasia, δ¹⁸O and δ¹³C values in human teeth obtained from early modern cemeteries in Wroclaw, Poland. Ann. Anat.—Anat. Anz.; 2020; 232, 151546. [DOI: https://dx.doi.org/10.1016/j.aanat.2020.151546]

51. Lécuyer, C.; Fourel, F.; Seris, M.; Amiot, R.; Goedert, J.; Simon, L. Synthesis of In-House Produced Calibrated Silver Phosphate with a Large Range of Oxygen Isotope Compositions. Geostand. Geoanalytical Res.; 2019; 43, pp. 681-688. [DOI: https://dx.doi.org/10.1111/ggr.12285]

52. Davies, C.L.; Surridge, B.W.J.; Gooddy, D.C. Phosphate oxygen isotopes within aquatic ecosystems: Global data synthesis and future research priorities. Sci. Total Environ.; 2014; 496, pp. 563-575. [DOI: https://dx.doi.org/10.1016/j.scitotenv.2014.07.057]

53. Smith, A.C.; Pfahler, V.; Tamburini, F.; Blackwell, M.S.A.; Granger, S.J. A review of phosphate oxygen isotope values in global bedrocks: Characterising a critical endmember to the soil phosphorus system. J. Plant Nutr. Soil Sci.; 2021; 184, pp. 25-34. [DOI: https://dx.doi.org/10.1002/jpln.202000513]

54. Gooddy, D.C.; Lapworth, D.J.; Bennett, S.A.; Heaton, T.H.E.; Williams, P.J.; Surridge, B.W.J. A multi-stable isotope framework to understand eutrophication in aquatic ecosystems. Water Res.; 2016; 88, pp. 623-633. [DOI: https://dx.doi.org/10.1016/j.watres.2015.10.046]

55. Gooddy, D.C.; Lapworth, D.J.; Ascott, M.J.; Bennett, S.A.; Heaton, T.H.E.; Surridge, B.W.J. Isotopic Fingerprint for Phosphorus in Drinking Water Supplies. Environ. Sci. Technol.; 2015; 49, pp. 9020-9028. [DOI: https://dx.doi.org/10.1021/acs.est.5b01137]

56. Granger, S.J.; Heaton, T.H.E.; Pfahler, V.; Blackwell, M.S.A.; Yuan, H.; Collins, A.L. The oxygen isotopic composition of phosphate in river water and its potential sources in the Upper River Taw catchment, UK. Sci. Total Environ.; 2017; 574, pp. 680-690. [DOI: https://dx.doi.org/10.1016/j.scitotenv.2016.09.007]