Introduction

Naturally occurring radionuclides, such as uranium-238 (238U) and thorium-232 (232Th), are extensively dispersed in the environment and building materials, including concrete [1,2,3]. Given the increasing utilization of uranium and thorium in rare earth and nuclear industries, workers and the general public are increasingly exposed to both radionuclides, which create severe toxicological and radiobiological health issues.

The world’s average 238U and 232Th contents in building materials is 50 Bq kg−1 (approximately 238U: 4.05 mg kg−1 and 232Th: 12.32 mg kg−1) [4], although greater 238U and 232Th contents are observed in concrete and other building materials made with fly ash and phosphogypsum (238U: 8.42–12.63 mg kg−1 and 232Th: 34.24–49.51 mg kg−1) [5,6,7]. The naturally occurring 238U and 232Th contents in concrete and steel used in nuclear installations (power plants, fuel production or reprocessing plants, and research reactors) can be activated or radioactively contaminated by various radionuclides throughout their service life [8, 9]. Kato et al. [10] discovered that uranium and thorium concentrations in activated concrete resulting from nuclear facility activities were relatively low, ranging between 0.76 and 0.96 mg kg−1 and 4.32 and 5.28 mg kg−1, respectively. Meanwhile, contamination of radionuclides to the sub-surface can be carried through micropores or fissures in the cement binder [11]. Uranium, strontium (Sr), cesium (Cs), plutonium (Pu), technetium-99 (99Tc), cobalt-60 (60Co), carbon-14 (14C), americium (Am), and other heavy metals can be linked with concrete contamination at the 300 Area of the Hanford Site due to spills on floors and other concrete surfaces [12]. However, the concentrations of uranium and thorium in the contaminated concrete from 300 Area Hanford site, the Fernald Environmental Management Project (FEMP), Oak Ridge Reservation (ORR) K-25, Paducah Gaseous Diffusion Plant (PGDP), and Portsmouth Gaseous Diffusion Plant (PORTS) have not been recorded [12]. In addition, the Building 21 site of the US Department of Energy Mound facility’s concrete floor in Miamisburg exhibited 136.5 mg kg−1 of thorium species [13].

A significant amount of radioactive concrete waste can be produced globally in the upcoming years due to the decommissioning of rare earth and nuclear facilities, including nuclear power plants (NPPs), and hence, even minor amounts of 238U and 232Th should be considered for potential radiobiological risks. Furthermore, catastrophic nuclear incidents such as the Chernobyl and Fukushima Daiichi tragedies can be extremely hazardous to nearby residents as well as the environment [14,15,16]. Despite this, there is limited information on the acute and chronic effects of incidental ingestions of 238U- and 232Th-contaminated concrete dust, particularly in workers or the public living near uranium and thorium mining sites, decommissioned rare earth, and nuclear facilities. Although workers in nuclear facilities are the most susceptible to uranium or/and thorium exposures, emerging data suggest that children and adults are equally vulnerable [17,18,19].

The estimated incidental daily soil ingestion rate from hand-to-mouth activity is 0.023 g for children [20], but as high as 0.2 g has been used [21, 22]. Unlike soil, incidental intakes of cement or concrete dust are uncommon. Their occurrence is often attributed to particular behaviors and linked to suicidal symptoms or psychiatric disorders [23,24,25]. Internal exposure to radioactive materials could damage biological tissues as they deteriorate over time. 238U and 232Th are released from the concrete matrix into the gastrointestinal (GI) fluids upon ingestion and transferred into the bloodstream, where blood is the primary carrier of soluble 238U and 232Th to target organs and tissues. However, when ingested, not all concrete-borne 238U and 232Th are absorbed into the systemic circulation. Previous studies on metals bioavailability ingestion have demonstrated that exposure risks based on the total concentrations of the contaminant’s calculations tend to overestimate the actual magnitude [26,27,28,29]. In addition, few findings indicated that thorium could result in hematotoxicity [30,31,32,33,34], while the effects of ingesting uranium are still unclear [19, 32, 35,36,37].

Previously, we addressed the absolute bioavailability of 238U and 232Th in IAEA-312-certified reference material, which was used as a surrogate for co-contaminated soil [22]. Post-48 h acute ingestion of IAEA-312 demonstrated noticeable histopathological and hematological alterations, indicating that intake of 238U in co-contaminated soil may lead to erythrocytes and proximal tubules damage, whereas 232Th intake may harm erythrocytes. However, studies are yet to link the relative bioavailability (RBA) and toxicity of 238U and 232Th following exposure to co-contaminated concrete dust. Therefore, this study aims to investigate the RBA and toxicity of 238U and 232Th in the female Sprague Dawley (SD) rats’ blood following oral ingestion exposure to co-contaminated concrete dust. The SD rats’ diet pellet was amended with co-contaminated concrete dust at three different dose levels (~ 50, 100, and 200 mg kg−1). In addition, blood samples collected at 48 h were prioritized to monitor the short-term or acute impacts of 238U and 232Th exposure on histopathological and hematological markers. This information and the applied techniques were envisaged to benefit research and exposure assessment processes for environmental and contaminated building materials.

Materials and Methods

Concrete Spiking and Characterization

The present research applied analytical grade chemicals without further purification and used ultrapure water (Milli-Q, Millipore, 18.2 MΩ·cm) as the general solvent. The mass ratio of concrete samples was based on the NPPs reports: 24.6 g of dry sand, 14.0 g of ordinary Portland cement (OPC, Type I), and 3.4 g of fly ash were mixed for the homogeneity of the concrete composition [38,39,40]. After mixing thoroughly, 8.0 mL of ultrapure water was added. Next, the uranyl- and thorium-nitrate solutions (1000 mg L−1 in 2% HNO3, Pure Plus, Perkin Elmer) were spiked uniformly into concrete samples to simulate various contamination levels of 238U and 232Th (Table 1) [10, 41,42,43]. After 28 days of curing, the unspiked- (UC) and spiked-concretes (SC) were crushed, ground, sieved (≤ 75 µm), and uniformly mixed for physicochemical characterization, bioavailability, and toxicity evaluations. Concrete particles of ≤ 75 μm were selected under the assumption of ease of finger adherence and a high probability of being ingested incidentally [44]. Subsequently, the UC and SC samples’ total concentration, moisture and organic contents, pH, mineral composition, and microstructure were measured and described in the Supplementary Information (SI) file.

Table 1 Physicochemical properties of the concrete, UC, and SC samples
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Assessing the 238U and 232Th Bioavailability via an SD Model

Throughout in vivo bioassays, the present study complied well with the animal care and experimental protocols outlined by the National University of Malaysia Animal Ethics Committee. Each of 174 female SD rats (aged 7–9 weeks) with 190–220 g body weight (bw) was separately housed in metabolic cages under standard conditions: 25 °C, 50% humidity, and 12/12 h light and dark cycle with free access to ultrapure water (Milli-Q, Millipore, 18.2 MΩ·cm).

The pellets were prepared by mixing the UC- or SC-amended diet in a mill, homogenized with ultrapure water, and kneaded into a paste. The diet was supplemented with UC or SC at a 1:10 concrete-to-diet mass ratio. Since the actual weight of concrete ingestion in humans is uncertain, this study used ~ 0.2 g of UC and SC based on a prior study [22] that utilized this quantity in soil ingestion in SD rats. In addition, uranyl nitrate and thorium nitrate were employed as reference doses (R1−R3) to compute the 238U- and 232Th-RBA in the SC and were separately added into the diet to obtain similar concentrations while control SD rats consumed unmodified diet pellets.

An in vivo SD rat bioassay was utilized to assess the influence of co-exposure on 238U- and 232Th-RBA in contaminated concrete dust. After a week of acclimation, each rat was placed in a metabolic cage. The rats were not fed for 24 h before being exposed acutely via modified pellets intake. Rats were randomly assigned into eight groups and the final concentrations of 238U and 232Th in unmodified or modified diet pellets are shown in Table 2. The unmodified diet pellets were 0.20 ± 0.01 mg of 238U⋅kg−1 and ≤ 0.01 mg of 232Th⋅kg−1.

Table 2 Each female SD rat in a specific group was exposed to a diet pellet (either unmodified or modified) containing total concentrations of 238U and 232Th
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Six SD rats were sacrificed by carbon dioxide anesthesia at each time interval (6, 12, 24, and 48 h) following UC, SC, and R exposure. In this study, the blood samples collected within 48 h were based on Rashid et al. [22], which suggested that the concentrations of 238U and 232Th in the SD rat’s blood increased within 48 h following acute ingestion of IAEA-312 and then gradually decreased. In addition, blood samples of control and treated SD rats were collected into a pre-calibrated tube containing dipotassium ethylenediaminetetraacetic acid (K2EDTA) and quantified via instrumental neutron activation analysis (INAA) after 24 h of freeze-drying.

The dose–response of 238U or 232Th accumulation in the blood was established for SD rats receiving reference materials (R: uranyl or thorium nitrates) following zero correction. 238U- or 232Th-RBA in spiked concrete (SC) was then determined using individual endpoints as the ratio of the dose-normalized 238U or 232Th accumulation in the blood following spiked concrete of 238U or 232Th exposure compared to reference materials (R: uranyl or thorium nitrates) exposure (Eq. (1)) [45]. The dose–response was determined using linear regressions for 238U and 232Th accumulation in the blood.

$$^{238}{\rm{U}}{\text{-}}\, {\rm{or}} ^{232}{\text{Th-RBA}} (\% ) = \left[ \frac{ {\rm{Blood}}_{[{\rm{spiked\, concrete}}]}} {{\rm{Blood}}_{[{\rm{reference \, material}}]}} \right] \, \times \left[\frac{ {\rm{Dose}}_{[{\rm{reference \ material}}]}} {{\rm{Blood}}_{[{\rm{spiked \ concrete}}]} }\right] \, \times 100 \%$$
(1)

where Blood [spiked concrete] = Concentrations of 238U or 232Th in the SD rat’s blood (mg kg−1) following exposure to modified pellets containing spiked concrete (SC). Blood [reference material] = Concentrations of 238U or 232Th in the SD rat’s blood (mg kg−1) following exposure to modified pellets containing (R: uranyl or thorium nitrates). Dose [spiked concrete] = Dosing level of 238U or 232Th (mg kg−1 bw h−1) in the modified pellets containing spiked concrete (SC). Dose [reference material] = Dosing level of 238U or 232Th (mg kg−1 bw h−1) in the modified pellets containing reference material (R: uranyl or thorium nitrates).

Histopathological and Hematological Analyses

The 238U and 232Th exposure toxicity were assessed at a cellular level by integrating histopathological and hematological analyses. Morphological changes in the SD rat blood cells were detected with high-resolution transmission electron microscopy equipped with electron-dispersive X-ray (HRTEM-EDX) spectrometer post-acute 238U and 232Th ingestion. Red blood cell (RBC), hemoglobin (Hb), erythrocytic indices [mean corpuscular volume (MCV) and mean corpuscular hemoglobin concentration (MCHC)], white blood cells (WBC), and leukocytes differential [including monocytes, lymphocytes, neutrophils, and platelets (PLT)] count, and packed cell volume (PCV) were assessed during hematological evaluations. Samples were prepared for both analyses as the methods reported by Almhanawi et al. [46] and Rahman et al. [47].

Quality Control and Assurance

A certified reference material (IAEA-312) was analyzed for quality control and assurance [48]. The IAEA-312 was irradiated alongside concrete and blood samples and three standard solutions designated for U/Th, RR1/5, and RR2/5 to assess the reliability of the INAA [49, 50]. During the measurement, each sample was assessed in triplicates.

The 238U and 232Th recoveries from the SC samples (n = 40) were 104.7 ± 0.3–109.2 ± 0.1% and 103.9 ± 0.4–113.8 ± 0.4%, respectively (Table 1). These results confirm the presence of 238U and 232Th in the SC. Therefore, the samples are practical for further investigations regarding their bioavailability and toxicity. Furthermore, the spiked process showed acceptable precision among replicates and good recoveries compared to the theoretical values. All data are presented as geometric mean (GM) ± geometric standard deviation (GSD), while statistical analyses were performed at a 95% confidence level with OriginPro software 9.8 [OriginLab Inc., Massachusetts, United States of America (USA)].

Results and Discussion

In Vivo Bioavailability of U and Th Co-exposures

No mortality or physical stress on SD rats was observed during the trials. The 238U and 232Th concentrations in the control SD rat’s blood were 5.01 ± 0.01 and 0.033 ± 0.007 μg kg−1, respectively, whereas the 238U and 232Th concentrations in SD rat’s blood following oral exposure to UC-amended diets were 13.8 ± 3.03 and 0.057 ± 0.004 μg kg−1, respectively. The low 238U and 232Th levels in the blood resulted from their low concentrations in the SD diet pellet and UC-amended diet. The 238U and 232Th blood concentrations of the SC- and R-amended diet groups were 4–120 times greater than the control and UC-amended diet groups, indicating low background values of 238U and 232Th in the blood.

Initially, a dose–response relationship was established based on the SD rats’ blood assay following 48 h exposure to different dose levels of R-amended diet. Figure 1 illustrates a strong linear dose–response relationship between 238U or 232Th accumulation in the blood with correlation coefficient (R2) values of 0.93 (Fig. 1A) and 0.96 (Fig. 1B) after zero correction, demonstrating the suitability of employing blood as the biological endpoint to quantify 238U- and 232Th-RBA in co-contaminated concrete dust. Moreover, determining bioavailability via time-course blood data is often more reliable than single tissue measurements [51].

Fig. 1
figure 1

The dose–response curves of 238U (A) and 232Th (B) accumulation in SD rats blood post-48 h feeding with the reference materials (R)-amended diet (uranyl and thorium nitrates)

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Figure 2 illustrates 238U and 232Th blood concentrations with time post-co-exposure. The maximum levels of 238U and 232Th were recorded 12 h after ingestion of the R-amended diet and decreased substantially between 24 and 48 h. In comparison, within 48 h, the concentrations of 238U and 232Th in the blood of rats fed with the SC-amended diet gradually increased. Similarly, Rashid et al. [22] reported that 238U and 232Th blood concentrations following the acute ingestion of IAEA-312 increased within 48 h and then continuously decreased. Although neither X-ray diffraction (XRD) nor field-emission scanning electron microscopy (FESEM)-EDX detected uranium- or thorium-bearing compounds in this study (Figs. S1, S2 and Table S1), the complex formation of uranium and thorium in the concrete influenced their lower absorptions in the SC- than R-amended diets.

Fig. 2
figure 2

Bioavailable 238U and 232Th concentrations in blood samples of SD rats fed spiked concrete (SC)- and reference materials (R)-amended diets

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Within 2 months, uranium in SC can form various phases, such as uranyl-oxyhydroxide, uranyl-silicate, and uranyl-phosphate [52]. Conversely, thorium forms the thorium-orthosilicate [Th(IV)SiO4] phase in cementitious materials [53]. The 238U and 232Th blood concentrations in rats fed with the R-amended diet were higher as uranyl and thorium nitrates are more soluble than other uranium and thorium compounds [17, 18]. Consequently, the uranium- and thorium-oxyhydroxides, silicates, and phosphates are less soluble with lower GI absorptions than uranyl or thorium nitrates. In contrast, following ingesting concrete dust contaminated with uranium and thorium, the soluble 238U and 232Th in the GI fluids will be absorbed into the bloodstream. Once in the blood, they are present such as bioavailable uranium and thorium species such as hexavalent (UO22+) and tetravalent (Th4+) states, respectively. According to Ansoborlo et al. [54], UO22+ moderately binds with ~ 50% of bicarbonate (HCO3−), 30% of transferrin (Tf), and 20% of hemoglobin (Hb), whereas Th4+ binds > 90% with Tf.

The RBA values of 238U and 232Th in the SD rat blood samples evaluated (in this study) were found to be in the range of 22.0% ± 0.86% to 30.8% ± 1.01% and 11.8% ± 0.14% to 13.7% ± 0.29%, respectively, post-exposure to co-contaminated concrete dust (Table 3). These findings revealed that the bioavailability of 238U and 232Th increased directly to their concentrations in the co-contaminated concrete dust. The RBA of 238U and 232Th demonstrated an ascending trend: SC1 < SC2 < SC3.

Table 3 Blood concentrations and relative bioavailability (RBA) of 238U and 232Th determined using a SD rat model
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Compared to 232Th, the 238U blood level of the SD rats fed with the SC-amended diets was ~ 100-fold higher, and its RBA values were approximately 2-fold greater in co-contaminated concrete dust. The variations in blood concentrations and bioavailability of 238U and 232Th suggested that compound solubility influences absorption in the GI tract. The GI pH value could also influence the absorption and bioavailability of 238U and 232Th in the blood since it affects the dissolution and solubility of radionuclides.

Höllriegl et al. [55] reported that 238U was more soluble in the GI tract than 232Th when the pH was increased (7.5), owing to the formation of highly soluble carbonate complexes with UO22+ ions. Conversely, 232Th solubility decreased, presumably due to precipitation at pH 7.5, which created insoluble or partially soluble 232Th compounds via the adsorption of soil minerals, organic material, and other suspended solids. These observations established that, despite exposure to 232Th, 238U absorption is higher and more soluble in the GI tract.

Biological Effects of U and Th on Blood Histopathology and Hematology

Notable erythrocyte anatomical changes were observed in controlled and treated rats during the end phase. Figure 3 depicts the well-structured cellular architecture of normal biconcave erythrocytes procured from the control SD rats. Similar changes were observed in erythrocytes exposed to a UC-amended diet and those exposed to a low dose of co-contaminated dust (SC1-amended diet), where only slight changes were observed compared to control erythrocytes. Meanwhile, significant changes were observed in erythrocytes exposed to modified diet pellets containing higher doses of co-contaminated concrete dust (SC2 and SC3) and reference materials (R1−R3). Erythrocytes were altered when subjected to a medium dose (SC2-amended diet), resulting in echinocytes, whereas a high dose (SC3-amended diet) resulted in echinocytes and aggregation. Similarly, following exposure to all R-amended diets, echinocytes and aggregation were observed.

Fig. 3
figure 3

High-resolution transmission electron microscopy (HRTEM) analysis of blood cells. Cross-section comparisons of the control and treated SD rat erythrocytes post-48 h feeding with unspiked concrete (UC)-, spiked concrete (SC)-, and reference material (R)-amended diets. The arrows with solid lines indicate normal erythrocytes (biconcave), while arrows with dashed lines denote echinocytes. Control erythrocytes were conventional biconcave shapes and echinocytes formed from low to medium doses of SC-amended diet. A high dose of the SC-amended diet and all levels of the R-amended diet resulted in echinocytes and aggregation at pH 7.3–7.5

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Generally, a higher 232Th dose induced severe damage to erythrocytes [30,31,32]. Thorium nitrate has been shown to trigger human erythrocytes to aggregate and hemolysis [30]. The 232Th bound to sialic acid in negatively charged membranes of glycophorin A (GpA) proteins reduces the surface charge and alters GpA while triggering erythrocyte aggregation. The EDX spectra confirmed the presence of 238U and 232Th in the erythrocyte samples of SD rats fed with the SC3 and all R-amended diets (Table S2). The observations proved that 238U and 232Th in co-contaminated concrete dust have the potential to harm erythrocytes.

In the present study, the hematological parameters of the control group of SD rats showed no significant changes compared to those exposed to the UC-amended diets (Table S3). Meanwhile, the hematological parameters of the SD rats exposed to the SC-amended diet increased progressively, concomitantly with the concentrations of 238U and 232Th in the blood of the SD rat’s post-exposure, as shown in Fig. 2. Concurrently, the results exhibited by the R-amended diet group revealed significant hematological effects at 12 h, with WBC, monocyte, lymphocyte, neutrophil, and PLT values corresponding with the maximum 238U and 232Th levels measured in the SD rat’s blood (Fig. 2). The data concurred with the results reported by prior studies [34, 35].

Hb and RBCs were observed to increase in female SD rats blood exposed to 24 mg L−1 of uranyl nitrate [35], while MCHC, WBC, and lymphocytes were elevated after exposure to 13.6 mg kg−1 of thorium nitrate [34]. The increments of WBCs and lymphocytes were related to the capacity of WBCs to combat inflammation due to 238U and 232Th ingestion [34]. In brief, steady increments in hematological parameters (RBC, Hb, PCV, MCV, MCHC, WBC, neutrophils, monocytes, lymphocytes, and PLT) and long-term exposure to 238U and 232Th of the SC- and R-amended diet groups would enhance polycythemia vera and thrombosis occurrences [56].

Conclusions

In this study, we presented the novel aspect of the bioavailability and hematotoxicity of 238U and 232Th following the acute ingestion of co-contaminated concrete dust containing these radioactive elements. The RBA values of 238U and 232Th in the SD rat blood samples ranged from 22.0% ± 0.86% to 30.8% ± 1.01% and from 11.8% ± 0.14% to 13.7% ± 0.29%, respectively. The RBA of 238U and 232Th demonstrated an ascending trend: SC1 < SC2 < SC3. The differences observed in blood concentrations and bioavailability of 238U and 232Th revealed that their absorption rates in the GI tract are affected by compound solubility. Moreover, the pH of the GI tract may influence the absorption and bioavailability of 238U and 232Th in the SD rats’ blood, which eventually alters their dissolution and solubility. Exposure to 238U and 232Th in co-contaminated concrete dust contributes to erythrocyte damage and elevated hematological attributes. The study observed significant changes in the erythrocytes exposed to modified diet pellets containing higher doses of co-contaminated concrete dust (SC2 and SC3) and reference materials (R1−R3). When subjected to a medium dose (SC2-amended diet), the erythrocytes were altered, resulting in echinocytes, whereas a high dose (SC3-amended diet) resulted in echinocytes and aggregation. Furthermore, exposure to all R-amended diets resulted in echinocytes and aggregation. Our study also indicates that there may be a correlation between the hematological parameters of SD rats and the concentrations of 238U and 232Th in their blood. Specifically, we observed a gradual increase within 48 h in the hematological parameters of rats exposed to a diet amended with SC. In contrast, rats fed an R-amended diet exhibited significant hematological effects after 12 h. Prolonged exposure to 238U and 232Th in co-contaminated concrete dust may result in blood cancer or even mortality if not effectively managed. These results would be highly significant in assessing the extremely hazardous impact of uranium and thorium exposure on humans due to incidental concrete dust ingestion.