In Vitro, in Vivo, and Spectroscopic Assessment of Lead Exposure Reduction via Ingestion and Inhalation Pathways Using Phosphate and Iron Amendments.

This study compared lead (Pb) immobilization efficacies in mining/smelting impacted soil using phosphate and iron amendments via ingestion and inhalation pathways using in vitro and in vivo assays, in conjunction with investigating the dynamics of dust particles in the lungs and gastro-intestinal tract via X-ray fluorescence (XRF) microscopy. Phosphate amendments [phosphoric acid (PA), hydroxyapatite, monoammonium phosphate (MAP), triple super phosphate (TSP), and bone meal biochar] and hematite were applied at a molar ratio of Pb:Fe/P = 1:5. Pb phosphate formation was investigated in the soil/post-in vitro bioaccessibility (IVBA) residuals and in mouse lung via extended X-ray absorption fine structure (EXAFS) and X-ray absorption near edge structures (XANES) spectroscopy, respectively. EXAFS analysis revealed that anglesite was the dominant phase in the ingestible (<250 µm) and inhalable (<10 µm) particle fractions. Pb IVBA was significantly reduced (p < 0.05) by phosphate amendments in the <250 µm fraction (solubility bioaccessibility research consortium assay) and by PA, MAP, and TSP in the <10 µm fraction (inhalation-ingestion bioaccessibility assay). A 21.1% reduction in Pb RBA (<250 µm fraction) and 56.4% reduction in blood Pb concentration (<10 µm fraction) were observed via the ingestion and inhalation pathways, respectively. XRF microscopy detected Pb in the stomach within 4 h, presumably via mucociliary clearance.

Toxicity assessment of fresh and weathered petroleum hydrocarbons in contaminated soil- a review.

Soil contamination with total petroleum hydrocarbons (TPH) is widespread throughout the globe due to the massive production of TPH anthropogenically and its occurrence in the soil. TPH is toxic to beneficial soil organisms and humans and thus has become a serious concern among the public. Traditionally TPH toxicity in the soil is estimated based on chemical fractions and a range of bioassays including plants, invertebrates and microorganisms. There is a large inconsistency among ecotoxicology data using these assays due to the nature of TPH and their weathering. Therefore, in this article, we critically reviewed the weathered conditions of TPH, the potential fate in soil and the bioindicators for the assessment of the ecotoxicity. Based on the current research and the state-of-the-art problem, we also highlighted key recommendations for future research scope for the real-world solution of the ecotoxicological studies of hydrocarbons.

Can in vitro assays account for interactions between inorganic co-contaminants observed during in vivo relative bioavailability assessment?

In vitro assays act as surrogate measurements of relative bioavailability (RBA) for inorganic contaminants. The values derived from these assays are routinely used to refine human health risk assessments (HHRA). Extensive in vitro research has been performed on three major inorganic contaminants; As, Cd and Pb. However, the majority of these studies have evaluated the contaminants individually, even in cases when they are found as co-contaminants. Recently, in vivo studies (animal model) have determined that when the three aforementioned contaminants are present in the same soil matrix, they have the ability to influence each other's individual bioavailability. Since in vitro assays are used to inform HHRA, this study investigated whether bioaccessibility methods including the Solubility/Bioavailability Research Consortium (SBRC) assay, and physiologically based extraction test (PBET), have the ability to detect interactions between As, Cd and Pb. Using a similar dosing methodology to recently published in vivo studies, spiked aged (12 years) soil was assessed by evaluating contaminant bioaccessibility individually, in addition to tertiary combinations. In two spiked aged soils (grey and brown chromosols), there was no influence on contaminant bioaccessibility when As, Cd and Pb we present as co-contaminants. However, in a red ferrosol, the presence of As and Pb significantly decreased (p < 0.05) the bioaccessibility of Cd when assessed using gastric and intestinal phases of the SBRC assay and the PBET. Conceivable, differences in key physico-chemical properties (TOC, Fe, Al, P) between the study soils influenced contaminant interactions and bioaccessibility outcomes. Although bioaccessibility methods may not account for interactions between elements as demonstrated in in vivo models, in vitro assessment provides a conservative prediction of contaminant RBA under co-contaminant scenarios.

Assessment of arsenic speciation and bioaccessibility in mine-impacted materials.

Mine-impacted materials were collected from Victoria, Australia and categorized into three source materials; tailings (n=35), calcinated (n=10) and grey slimes (n=5). Arsenic (As) concentrations in these materials varied over several orders of magnitude (30-47,000mgkg(-1)), with median concentrations of 500, 10,800 and 1500mgkg(-1), respectively. When As bioaccessibility was assessed using the Solubility Bioaccessibility Research Consortium (SBRC) assay, As bioaccessibility ranged between 4 and 90%, with mean gastric phase values of 30%, 49% and 82% for tailings, calcinated and grey slimes, respectively. An analysis of variance (ANOVA) determined that As bioaccessibility was significantly different (P<0.05) between source materials. This was due to differences in As mineralogy, soil particle size as well as the concentration and nature of Fe present. X-ray Absorption Near Edge Structure (XANES) analysis identified arseniosiderite, yukonite, realgar, loellingite and mineral sorbed arsenate species in mine-impacted materials. Despite differences in physicochemical properties, 'mine wastes' are often reported under a generic descriptor. Outcomes from this research highlight that variability in As bioaccessibility can be prescribed to As mineralogy and matrix physicochemical properties, while categorizing samples into sub-groups can provide some notional indication of potential exposure.

Assessment of DDT relative bioavailability and bioaccessibility in historically contaminated soils using an in vivo mouse model and fed and unfed batch in vitro assays.

In this study, DDTr (DDTr = DDT + DDD + DDE) relative bioavailability in historically contaminated soils (n = 7) was assessed using an in vivo mouse model. Soils or reference materials were administered to mice daily over a 7 day exposure period with bioavailability determined using DDTr accumulation in adipose, kidney, or liver tissues. Depending on the target tissue used for its calculation, some variability in DDTr relative bioavailability was observed; however, it did not exceed 25% (range 2-25%). When DDTr bioaccessibility was determined using organic physiologically based extraction test (Org-PBET), unified BARGE method (UBM), and fed organic estimation human simulation test (FOREhST) in vitro assays, bioaccessibility was less than 4% irrespective of the assay utilized and the concentration of DDTr in the contaminated soil. Pearson correlations demonstrate a poor relationship between DDTr relative bioavailability and DDTr bioaccessibility (0.47, 0.38, and 0.28, respectively), illustrating the limitations of the static in vitro methods for predicting the dynamic processes of the mammalian digestive system for this hydrophobic organic contaminant.

Assessment of lead bioaccessibility in peri-urban contaminated soils.

Lead (Pb) bioaccessibility was assessed in a range of peri-urban soils (n=31) with differing sources of Pb contamination, including shooting range soils, and soils affected by incinerator, historical fill, mining/smelting, and gasworks activities. A gossan soil sample was also included. Lead bioaccessibility was determined using both gastric and intestinal phases of the SBRC in vitro assay and in vitro data was then incorporated into in vivo-in vitro regression equations to calculate Pb relative bioavailability. Lead bioaccessibility ranged from 26.8-105.2% to 5.5-102.6% for gastric and intestinal phase extractions respectively. Generally, Pb bioaccessibility was highest in the shooting range soils and lowest in the gossan soil. Predictions of relative Pb bioavailability derived from in vitro data were comparable for shooting ranges soils, but highly variable for the other soils examined. For incinerator, historical fill, gasworks and gossan soils, incorporating in vitro gastric data into the in vivo-in vitro regression equation resulting in more conservative Pb relative bioavailability values than those derived using the intestinal in vitro data.

Environmental metabolites of fluoroquinolones: synthesis, fractionation and toxicological assessment of some biologically active metabolites of ciprofloxacin.

BACKGROUND, AIM, AND SCOPE: Biowastes produced by humans and animals are routinely disposed of on land, and concern is now growing that such practices provide a pathway for fluoroquinolone (FQs) antibacterial agents and their environmental metabolites (FQEMs) to contaminate the terrestrial environment. The focus of concern is that FQs and FQEMs may accumulate in amended soils to then adversely impact on the terrestrial environment. One postulated impact is the development of a selective environment in which FQ-resistant bacteria may grow. To find evidence in support of an accumulation of antibacterial-like activity, it was first necessary to establish whether any biologically active FQEMs could be synthesized by physicochemical factors that are normally present in the environment. However, many FQEMs are not commercially available to be used as standards in such studies. FQEMs were therefore synthesized using well-defined processes. They were subsequently analyzed using spectroscopy (UV-vis) and high performance liquid chromatography with mass spectral detection. The antibacterial-like activities of fractionated FQEMs were then assessed in novel bacterial growth inhibition bioassays, and results were compared to those obtained from instrumental analyses. MATERIALS AND METHODS: Parent FQs were either exposed to sunlight or were synthesized using defined aerobic microbial (Mycobacterium gilvum or a mixed culture derived from an agricultural soil) fermentation processes. Mixtures of FQEMs derived from photo- and (intracellular) microbial processes were isolated by preparative chromatography and centrifugation techniques, respectively. Mixtures were subsequently fractionated using analytical high-performance thin layer chromatography (HPTLC), and excised analytes were tested in bioautography assays for their antibacterial-like activities. Two bacteria, Escherichia coli (E. coli) and Azospirillum brasilense (A. brasilense) were used as reporter organisms in testing FQ standards and any subtle differences between biologically active FQEMs of ciprofloxacin (CF). RESULTS AND DISCUSSION: FQEMs produced in the photo-synthetic process had UV-vis profiles that were indistinguishable from the parent FQs, and yet mass spectral data revealed the presence of N-formylciprofloxacin (FCF). In contrast, the UV-vis profiles of FQEMs synthesized by M. gilvum and a mixed culture of microorganisms had UV-vis profiles that were similar to one another and markedly different to the parent fluoroquinolones. Mass spectral studies confirmed the presence of FCF and N-acetylciprofloxacin in both microbial ferments. In addition, a photo-FQEM (Cp 6), three M. gilvum FQEMs (Cm 5, Cm 8, and Cm 10) and a mixed culture FQEM (Cs 6) of CF and many other FQEMs of CF, norfloxacin (NF), and enrofloxacin (EF) were fractionated using HPTLC, although their identities have yet to be confirmed. Differences between bioautography results were obtained when E. coli or A. brasilense were used as reporter organisms. Parent FQs (CF and EF) and the FQEMs of CF (Cp 6, Cm 8, and Cs 6) displayed antibacterial-like activity when using E. coli as the reporter organism. In contrast, A. brasilense was insensitive to parent CF and sensitive to EF and all tested FQEMs of CF. Results are consistent with photo- and microbial processes modifying CF in different ways, with the latter changing the UV-vis chromophores. It can be inferred that a lack of detection of analytes (especially photo-FQEMs) when using UV-vis does not necessarily indicate an absence of analyte. Additionally, similarities between the UV-vis profiles of FQEMs extracted from the (monoculture) M. gilvum and the mixed culture microbial aerobic ferments are consistent with similar processes operating in both ferments. Results of HPTLC and bioautography studies revealed that mixtures of (photo- and microbial) FQEMs could be fractionated into individual components. CONCLUSIONS: Bioactive FQEMs of ciprofloxacin, as a representative FQ, can be synthesized by photo- and microbial processes, and their detection required the use of both instrumental and bioautography analytical techniques. It is likely that such FQEMs will also be present on agricultural land that has been repeatedly amended with FQ-contaminated biosolids. RECOMMENDATIONS AND PERSPECTIVES: The use of instrumental analytical techniques alone and especially photometric detection techniques will underestimate antibacterial-like activities of FQEMs. Moreover, the extraction technique(s) and the selected toxicological endpoint(s) require careful consideration when assessing bioactivity. It is therefore recommended that instrumental analytical techniques and several bioautography assays be performed concurrently, and bioautography assays should use a variety of reporter organisms. Two types of bacterial growth bioassays are recommended in any assessment of antibacterial-like activity derived from CF (and possibly from other FQs). A standardized E. coli bioassay should be used as a general screening procedure to facilitate intra- and inter-laboratory exchange of data. Additionally, soil-specific (region-specific) growth inhibition bioassays should be undertaken using several species of endemic soil bacteria. It is likely that the two sets of data will be useful in future risk assessment processes.

Assessment of four commonly employed in vitro arsenic bioaccessibility assays for predicting in vivo relative arsenic bioavailability in contaminated soils.

Currently, a number of in vitro methods are in use worldwide to assess arsenic (As) bioaccessibility in soils. However, a dearth of research has been undertaken to compare the efficacy of the in vitro methods for estimating in vivo relative As bioavailability. In this study, As bioaccessibility in contaminated soils (n = 12) was assessed using four in vitro assays (SBRC, IVG, PBET, DIN). In vitro results were compared to in vivo relative As bioavailability data (swine assay) to ascertain which methodologies best correlate with in vivo data. Arsenic bioaccessibility in contaminated soils varied depending on the in vitro method employed. For the SBRC and IVG methods, As bioaccessibility generally decreased when gastric-phase values were compared to the intestinal phase. In contrast, extending the PBET and DIN assays from the gastric to the intestinal phase resulted in an increase in As bioaccessibility for some soils tested. Comparison of in vitro and in vivo results demonstrated that the in vitro assay encompassing the SBRC gastric phase provided the best prediction of in vivo relative As bioavailability (R(2) = 0.75, Pearson correlation = 0.87). However, relative As bioavailability could also be predicted using gastric or intestinal phases of IVG, PBET, and DIN assays but with varying degrees of confidence (R(2) = 0.53-0.67, Pearson correlation = 0.73-0.82).

Comparison of in vivo and in vitro methodologies for the assessment of arsenic bioavailability in contaminated soils.

An in vivo swine assay was utilised for the determination of arsenic (As) bioavailability in contaminated soils. Arsenic bioavailability was assessed using pharmacokinetic analysis encompassing area under the blood plasma-As concentration time curve following zero correction and dose normalisation. In contaminated soil studies, As uptake into systemic circulation was compared to an arsenate oral dose and expressed as relative As bioavailability. Arsenic bioavailability ranged from 6.9+/-5.0% to 74.7+/-11.2% in 12 contaminated soils collected from former railway corridors, dip sites, mine sites and naturally elevated gossan soils. Arsenic bioavailability was generally low in the gossan soils and highest in the railway soils, ranging from 12.1+/-8.5% to 16.4+/-9.1% and 11.2+/-4.7% to 74.7+/-11.2%, respectively. Comparison of in vivo and in vitro (Simplified Bioaccessibility Extraction Test [SBET]) data from the 12 contaminated soils and bioavailability data collected from an As spiked soil study demonstrated that As bioavailability and As bioaccessibility were linearly correlated (in vivo As bioavailability (mgkg(-1))=14.19+0.93.SBET As bioaccessibility (mgkg(-1)); r(2)=0.92). The correlation between the two methods indicates that As bioavailability (in vivo) may be estimated using the less expensive, rapid in vitro chemical extraction method (SBET) to predict As exposure in human health risk assessment.

In vitro assessment of arsenic bioaccessibility in contaminated (anthropogenic and geogenic) soils.

Arsenic (As) bioaccessibility in contaminated soils (n=50) was assessed using the simplified bioaccessibility extraction test (SBET). Soils used in the study were collected from sites where As was used as an herbicide (railway corridor) or pesticide (cattle dip sites), from former gold mines and from highly mineralised locations containing geogenic As sources (gossans). In all but three soils, As bioaccessibility was less than 50% indicating that a significant proportion of the total As concentration may not be available for absorption in the gastrointestinal tract following incidental soil ingestion. When regression models were developed based on soil properties, the descriptive variables best able to describe As bioaccessibility in railway corridor, dip site and mine site soils were total As and total or dithionite-citrate extractable (free) iron (Fe). While As bioaccessibility could be predicted (r(2)=0.955, n=50) in these contaminated soils, As bioaccessibility for gossan soils was a poor fit using linear or multivariate regression analysis.

In vivo assessment of arsenic bioavailability in rice and its significance for human health risk assessment.

BACKGROUND: Millions of people worldwide consume arsenic-contaminated rice; however, little is known about the uptake and bioavailability of arsenic species after arsenic-contaminated rice ingestion. OBJECTIVES: In this study, we assessed arsenic speciation in greenhouse-grown and supermarket-bought rice, and determined arsenic bioavailability in cooked rice using an in vivo swine model. RESULTS: In supermarket-bought rice, arsenic was present entirely in the inorganic form compared to greenhouse-grown rice (using irrigation water contaminated with sodium arsenate), where most (approximately 86%) arsenic was present as dimethylarsinic acid (organic arsenic). Because of the low absolute bioavailability of dimethylarsinic acid and the high proportion of dimethylarsinic acid in greenhouse-grown rice, only 33 +/- 3% (mean +/- SD) of the total rice-bound arsenic was bioavailable. Conversely, in supermarket-bought rice cooked in water contaminated with sodium arsenate, arsenic was present entirely in the inorganic form, and bioavailability was high (89 +/- 9%). CONCLUSIONS: These results indicate that arsenic bioavailability in rice is highly dependent on arsenic speciation, which in turn can vary depending on rice cultivar, arsenic in irrigation water, and the presence and nature of arsenic speciation in cooking water. Arsenic speciation and bioavailability are therefore critical parameters for reducing uncertainties when estimating exposure from the consumption of rice grown and cooked using arsenic-contaminated water.