Single Extraction Procedure Research Articles

Effects of Leaching Agents on Pb and Cd Immobilization in Battery Waste Contaminated Soils Amended with Bare and Stabilized Zero-Valent Iron Nanoparticles

This study compared the effectiveness of bare zero-valent iron nanoparticles (B-nZVI) and starch-stabilized zero-valent iron nanoparticles (S-nZVI) in immobilizing Pb and Cd from lead-acid battery waste soils. Both B-nZVI and S-nZVI were prepared in almost identical manner using the technique of reducing ferric chloride with sodium borohydride. X-ray diffraction (XRD) and dynamic light scattering (DLS) analyses confirmed that polydisperse B-nZVI and S-nZVI were synthesized. XRD and DLS analyses showed that B-nZVI and S-nZVI had different surface properties. To assess the immobilization capability of B-nZVI and S-nZVI, a composite soil sample was collected from an automobile lead-acid battery waste dumpsite. The soil sample had a pH of 3.85 and Pb and Cd levels of 16,674 mg/kg and 41 mg/kg, respectively. Single extraction procedures using 0.01M CaCl2, 0.1 M HCl, and 0.05 M EDTA were used to simulate phytoavailable Pb and Cd in the soil studied. Batch immobilization analysis showed that Cd was mobile in the control but immobile in B-nZVI and S-nZVI treated soils. Pb was however not immobile in either the control or treated soils. The mobility of Pb however decreased with increasing doses of S-nZVI and 0.003 g of S-nZVI was needed to make Pb completely immobile in soil. Batch immobilization also showed that S-nZVI was 1.8-2.49 times more efficient in immobilizing Pb than B-nZVI. Simulated phytoavailability of Pb was in the order of EDTA > HCl > CaCl2 > H2O while simulated photoavailable Cd was in the order of HCl > EDTA > H2O > CaCl2.

Synthesis and characterization of Restricted Access Magnetic Nanotubes (M-RACNTs) for the extraction of secondary metabolites from Lasiodiplodia sp. fermentation broth

This work proposed the synthesis and the characterization of Restricted Access Magnetic Nanotubes (M-RACNTs) to be used as sorbent in the magnetic dispersive solid-phase extraction (MDSPE) of organic compounds from the fermentation broth of the endophyte Lasiodiplodia sp. Synthesis was performed by functionalizing commercial multiple-walled carbon nanotubes with magnetic nanoparticles (M-CNTs) and by coating the M-CNTs with a layer of bovine serum albumin. The materials were characterized by infrared spectroscopy with Fourier Transform, thermogravimetry, zeta potential, and scanning electron microscopy. Different crude extracts were obtained, depending on the sample pH and the elution solvent tested. Chloroform and ethyl acetate extracts showed activity against a Gram-positive pathogen (Staphylococcus aureus). The crude extract's constitution was investigated. By GC/MS analysis were found hydrocarbons, aldehydes, alcohols, among others, while by LC-MS/MS analysis were found flavonoids, macrocyclic lactones, oleic acids, jasmonic acid derivatives and amino acids. A multivariate optimization, through fractional factorial planning, for four independent variables, was performed for a hydrocarbon (hexadecane) and one possible jasmonic acid derivative, both with antimicrobial activity related in the literature. The results showed that it was possible to obtain crude extracts with high concentrations of these analytes in a single extraction procedure. The increase in the analytical signal was 3.7 times for hexadecane and 1.4 times for the possible jasmonic acid derivative. The M-RACNTs also demonstrated their ability to prevent the adsorption of macromolecules, such as lysozyme, which was confirmed through SDS-PAGE analysis.

Liquid chromatography-tandem mass spectrometry based simultaneous quantification of tryptophan, serotonin and kynurenine pathway metabolites in tissues and cell culture systems

BackgroundKynurenine and respective metabolites exhibit bioactivity as well as tryptophan, an essential amino acid, and the neurotransmitter serotonin. Dysregulations in the kynurenine pathway are involved in neurodegenerative/neuropsychiatric disorders and diabetes mellitus type 2 but also in cancer. Therefore, measurements of kynurenine-related metabolites will improve the general understanding for kynurenine pathway relevance in disease pathogenesis. MethodsTryptophan, serotonin, picolinic acid, quinolinic acid, 3–OH-kynurenine, kynurenine, 3-OH-anthranilic acid, kynurenic acid, anthranilic acid as well as nicotinic acid and the redox cofactor NAD+ were analyzed in heterogeneous matrices by ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). After validation, the described method was applied for measurements of native metabolite concentrations in murine tissues and cellular systems including pathway-shift monitoring after treatment with the tryptophan-2,3-dioxygenase-inhibitor 680C91. In addition, the method was evaluated for its ability for integration into multi-omics approaches using a single sample metabolite extraction procedure. ResultsA simple and sensitive UPLC-MS/MS method for simultaneous quantification of up to 10 kynurenine-related metabolites in four biological matrices was developed. Within a run time of 6.5 min, chromatographic separation of kynurenine-related metabolites, including the isomers nicotinic acid and picolinic acid, was achieved without derivatization. Validation parameters, including interday precision (<14.8%), mean accuracy (102.4% ± 12.9%) and linear detection ranges of more than three orders of magnitude, indicate method reliability. Depending the investigated sample matrix, the majority of metabolites were successfully detected and quantified in native murine and cell culture derived sample materials. Furthermore, the method allowed to monitor the impact of a tryptophan-2,3-dioxygenase-inhibitor on kynurenine pathway in a cellular system and is suitable for multi-assay analyses using aliquots from the same cell extract. ConclusionThe described UPLC-MS/MS method provides a simple tool for the simultaneous quantification of kynurenine pathway metabolites. Due to its suitability for many physiological matrices, the method provides wide application for disease-related experimental settings.

A double-edged sword: Reductive soil disinfestation changes the fates of trace metal elements in soil

Although the effects of reductive soil disinfestation (RSD) in soil sterilization have been proven in several countries, the potential risks of trace metal elements (TMEs) caused by RSD require further assessment. Here, freshly Cd-spiked soil and historically contaminated greenhouse soil were exposed to RSD and the fates of TMEs, Cd, Co, Cu, Ni, Pb, and Zn, were investigated. All RSD treatments lasted for 21 days and subsamples were collected at different time intervals. Samples were open-air incubated for another 7 days until day 28 to simulate the situation after drainage. The bioavailability and geochemical fractionation of TMEs were investigated based on single and sequential extraction procedures and the environmental risks were assessed. The results showed that RSD increased the relative abundance of Firmicutes and Bacteroidetes, and the content of functional groups, including Fe, Mn, and S compounds respirations increased after RSD, highlighting the possible reductive dissolution of FeMn oxides and precipitation of TMEs. The dissolution decreased the reducible fractions of TMEs and increased the acid-soluble fractions of Co, Ni, Pb, and Zn, in the European Community Bureau of Reference results, reflecting the activation of TMEs in soils. However, the precipitation of sulfate resulted in the stabilization of Cd and Cu in two types of soils, increased their residual fractions, and decreased their acid-soluble fractions and bioavailabilities. After drainage, because the influence caused by precipitation rapidly disappeared and the impact of FeMn oxides dissolution remained, the acid-solubility of TMEs was greater than their initial status in the two soils. Furthermore, as a highly toxic metal, the activation of Cd at 28 days caused the rapid increase of ecological risks, which is particularly concerning. The results suggest that RSD temporarily increases the potential risks of TMEs and that certain measures must be taken.

Effect of incubation time of three single extraction procedures on trace element extraction from sediment and soil.

The use of standard single-extraction procedures to evaluate the mobility and availability of trace elements is a common practice in most laboratories dealing with soil or sediment analysis. Most standard single-extraction procedures describe incubations last for 2h. However, these were tested and validated for soil analysis. Applying them for sediment analysis without further investigation might be misleading and should be reviewed with care. This paper investigates the effect of incubation time on the extraction efficiency of three standard single-extracting reagents (0.01M CaCl2, 1M NH4NO3, and 0.05M EDTA). Incubation experiments with sediment and soil samples lasting for 2h, 10h, and 10 d were performed. The results indicated that 2h appears sufficient to reach equilibrium using CaCl2 or NH4NO3 for soil analysis; but when analyzing sediments, incubation for 10 d resulted in higher concentrations. Incubation experiments with 0.05M EDTA showed that incubation for 2h was enough to extract Cd from the soil sample, Mn and to a lesser extent Cd from the sediment samples; while for the other elements, incubation for 10 d yielded higher concentrations for both sample types compared to that obtained after 2h and 10h separately. Relative to the pseudo-total metal contents, more than 55% of all studied elements were extracted by using 0.05M EDTA, indicating high bioavailable metal fraction.

Estimating remobilization of potentially toxic elements in soil and road dust of an industrialized urban environment.

The mobility of potentially toxic elements (PTEs) is of paramount concern in urban settings, particularly those affected by industrial activities. Here, contaminated soils and road dusts of the medium-size, industrialized city of Volos, Central Greece, were subjected to single-step extractions (0.43M HNO3 and 0.5M HCl) and the modified BCR sequential extraction procedure. This approach will allow for a better understanding of the geochemical phase partitioning of PTEs and associated risks in urban environmental matrices. Based on single extraction procedures, Pb and Zn exhibited the highest remobilization potential. Of the non-residual phases, the reducible was the most important for Pb, and the oxidizable for Cu and Zn in both media. On the other hand, mobility of Ni, Cr, and Fe was low, as inferred by their dominance into the residual fraction. Interestingly, we found a significant increase of the residual fraction in the road dust samples compared to soils. Carbonate content and organic matter controlled the extractabilities of PTEs in the soil samples. By contrast, for the road dust, magnetic susceptibility exerted the main control on the geochemical partitioning of PTEs. We suggest that anthropogenic particles emitted by heavy industries reside in the residual fraction of the SEP, raising concerns about the assessment of this fraction in terms of origin of PTEs and potential environmental risks. Conclusively, the application of sequential extraction procedures should be complemented with source identification of PTEs with the aim to better estimate the remobilization of PHEs in soil and road dust influenced by industrial emissions.

Optimization of a Multiresidue Analysis of 65 Pesticides in Surface Water Using Solid-Phase Extraction by LC-MS/MS.

An analytical method was developed and validated for simultaneous quantitation of 65 pesticides, including one single solid-phase extraction (SPE) procedure in surface water by liquid chromatography coupled to tandem mass spectroscopy. Different parameters that have an influence on extraction efficiency were evaluated in this research. Different types of cartridges, elution solvents, and sorbent drying time were investigated, and the most appropriate one was selected. Moreover, various pretreatment techniques were applied to remove sediments from water without the loss of pesticides. Centrifugation was introduced as the best option at the beginning of sample preparation to resolve the clogging of the sorbent cartridges. The recoveries of all pesticides ranged from 70% to 120%, with a relative standard deviation of less than 13.7%. The feasibility of the method was evaluated on 10 surface water samples with different concentrations of sand, sediment, and particles.

Effects on metal availability of the application of tree biochar and municipal waste biosolid in a metalliferous mine tailings substrate.

The phytostabilization of mine tailings requires a previous assessment of the effects of soil amendments on metal mobility. The goal of this work was to evaluate the response of metal availability (both labile and potentially available pools) to the addition of two organic amendments (a municipal waste biosolid and a tree biochar), separately and in combination, in a mine tailings substrate. For this purpose, a comprehensive comparison among several single extraction procedures and a sequential extraction procedure was performed. The effects on metals phytotoxicity were assessed through a germination test using seeds of Zygophyllum fabago. When evaluating the effect of the amendments in the labile metal pool, the biochar resulted effective in decreasing metal-extractable concentrations, especially for Cd, Mn and Zn. The treatment with biochar also showed better germination parameters (percentage of germinated seeds and sooner germination) than the rest of the unamended and amended treatments. The use of the municipal organic biosolid increased labile metal concentrations and potentially available metal pools assessed with EDTA and did not contribute to achieve better results of seed germination. Compared to the single biosolid treatment, the combination of biochar/biosolid modulated some labile metal concentrations and showed similar germination parameters to those obtained for the treatment amended only with biochar. This positive effect of biochar in modulating the soluble metal concentrations associated with certain urban/agricultural organic materials supported the suitability of using these combinations in field applications, although a higher rate of biochar application would be recommended to obtain a more beneficial effect.

Evaluation of Element Mobility in River Sediment Using Different Single Extraction Procedures and Assessment of Probabilistic Ecological Risk

In this manuscript, samples of Kupa River sediments were examined using three different extraction agents. The aim of this study was to evaluate the applicability of single extraction procedures to investigate the bioavailability and mobility of major and trace elements (Al, As, Ba, Cd, Co, Cr, Cu, Fe, K, Li, Mg, Mn, Na, Ni, P, Pb, S, Si, Sr, Ti, V, and Zn) from river sediment. Two forms of studied elements were evaluated: mobile, the most toxic element form (extraction with 1 M CH3COONH4 and 0.01 M CaCl2) and potentially mobilized form (2 M HNO3 extraction). The estimation of the ecological risk, with the application of the probability distribution of RI (potential ecological risk index) values, is yielded with the help of the Monte Carlo simulation (MCS). Ammonium acetate is proved to be a better extraction agent than calcium chloride. A positive correlation between the content of all extracted elements with nitric acid and the total element content indicates that 2 M HNO3 efficiently extracts all studied elements. Results showed anthropogenic sources of cadmium and copper and high barium mobility. The MCS suggests that risk of Cr, Cu, Ni, Pb, and Zn was low; As and Cd posed a lower and median ecological risk in the studied areas.

A simple and easy-to-perform liquid chromatography–mass spectrometry method for the quantification of tacrolimus and its metabolites in human whole blood. Application to the determination of metabolic ratios in kidney transplant patients

Tacrolimus is the cornerstone of immunosuppressive therapy in solid organ transplantation and its blood concentrations are routinely monitored. Tacrolimus is extensively metabolized into metabolites that are supposed to be nephrotoxic. Yet, few analytical methods have been described to simultaneously quantify tacrolimus and its main metabolites. We developed and validated a simple liquid chromatography-mass spectrometry method for the quantification of tacrolimus and its three desmethylated metabolites, 13-O, 15-O, and 31-O-desmethylated tacrolimus (M−I, M−III, and M−II respectively) in human whole blood. Protein precipitation of 50 µL of whole blood with 100 µL methanol and zinc sulfate was used as a single-extraction procedure. Tacrolimus and its metabolites were quantified using electrospray ionization-triple quadrupole mass spectrometry in combination with selected reaction monitoring detection in the positive ionization mode. The method was validated following FDA recommendations. This method was precise (intra- and inter-assay coefficients of variation: 2.88–7.81% and 3.96–12.10% for low and high levels of internal quality controls, respectively) and accurate (intra- and inter-assay biases: −1.67–10.30%, and −0.77–-9.36%, respectively).In adult kidney transplant patients who were treated with tacrolimus prolonged release formulation, the median (10th-90th percentiles) trough concentrations (n = 16) of tacrolimus, M−I, and M−III were 5.85 (3.37–7.09), 0.100 (0.037–0.168), 0.051 (0.03–0.104), respectively. M−II was measured in only 2 trough samples. The metabolic ratios M−I/tacrolimus and M−III/tacrolimus were 0.017 (0.009–0.027) and 0.009 (0.006–0.015) when measured on trough concentration and 0.022 (0.011–0.037) and 0.008 (0.006–0.015) when measured on area under the curves 0–24 h.This method is a suitable and easy-to-perform tool for future pharmacokinetic-pharmacodynamics studies investigating the importance of tacrolimus and its metabolites blood exposure for solid organ graft survival.

Selecting efficient methodologies for estimation of As and Hg availability in a brownfield.

The determination of soil metal(loid) availability presents controversy and there is no consensus or uniformity on used analytical methods. In this study nine single extraction methods (H2O, CaCl2, NaNO3, NH4NO3, DTPA, EDTA, HCl, LMWOA, TCLP) and four sequential extraction procedures (Tessier, BCR, Wenzel and Fernández-Martínez) have been compared to estimate the availability of As and Hg in two soils from a highly polluted brownfield, especially with As. The metal(loid) concentrations were also determined in three native plant species (Lotus corniculatus, Betula celtiberica and Dactylis glomerata) collected in the habitat under study. Each single extractant showed a particular capacity of As/Hg extraction because they do not extract the same forms of each element. The availability of As and Hg depended on the element characteristics, soil properties, type of extractant and degree of pollution, thus the use of a single extraction procedure provides limited information of metal(loid) availability and to reach general conclusions is difficult. Regarding the sequential extractions, each procedure showed a specific pattern for As and Hg regardless of the soil. Thus, the choice of one or other method depends on the environmental conditions, metal(loid) and soil properties. In risk assessment studies it would be recommendable to select one of the more aggressive extractants, so as not to underestimate the environmental risk. In this regard, the sequential extraction procedures render more detailed information about metal(loid) potential availability in relation to soil properties. The analysis of native plant species showed higher metal(loid) concentrations in roots than in aerial parts and differences were observed depending on the metal(loid) and the species. In general, plants showed a higher BCFs for Hg than As even though the total and available As concentrations were higher than those found for Hg, which highlights the influence of plant species on the metal(loid) uptake.

Assessment of bioavailability and mobility of major and trace elements in agricultural soils collected in Port St Johns, Eastern Cape, South Africa using single extraction procedures and pseudo-total digestion.

The aim of this study was to evaluate the applicability of ultrasound assisted single extraction and pseudo-total digestion procedures to investigate the bioavailability and mobility of major and trace elements collected from agricultural soil collected in Mzimvubu farmstead area located in Port St Johns, Eastern Cape Province (South Africa). The potential metal availability was assessed using complexing agent (ethylenediaminetetraacetic acid (EDTA)), mild (cacium chloride (CaCl2) and ammonium nitrate (NH4NO3)), deionized water and acidic (CH3COOH) extractants with the assistance of ultrasound to evaluate mobility and bioavailability of metals. The ultrasound radiation was used to assist the single extraction of major and trace metals from agricultural soils as well shortening the extraction time. The pseudo-total metal content in agricultural soils was obtained using pseudo-total digestion employing aqua-regia. The results obtained using different solvents for single extraction procedures, revealed that higher extraction of Al (256-681mgkg-1), Fe (172-430mgkg-1) and Mn (35-136mgkg-1), was observed compared to other metals. Among the investigated solvents, deionized water, acetic acid and EDTA proved to be the most aggressive extractants. The geo-accumulation index (2-6) and contamination factor (3-6) demonstrated that Fe, Cr, Mn, Co, Cu, Ni, and Zn could pose significant environmental contamination risk. Based on the results obtained, it can be concluded that the high levels of some studied metals was due to anthropogenic activities. The online version contains supplementary material available at 10.1007/s40201-020-00581-x.

An Optimization of Liquid-Liquid Extraction of Urinary Volatile and Semi-Volatile Compounds and Its Application for Gas Chromatography-Mass Spectrometry and Proton Nuclear Magnetic Resonance Spectroscopy.

Urinary volatile compounds (VCs) have been recently assessed for disease diagnoses. They belong to very diverse chemical classes, and they are characterized by different volatilities, polarities and concentrations, complicating their analysis via a single analytical procedure. There remains a need for better, lower-cost methods for VC biomarker discovery. Thus, there is a strong need for alternative methods, enabling the detection of a broader range of VCs. Therefore, the main aim of this study was to optimize a simple and reliable liquid–liquid extraction (LLE) procedure for the analysis of VCs in urine using gas chromatography-mass spectrometry (GC-MS), in order to obtain the maximum number of responses. Extraction parameters such as pH, type of solvent and ionic strength were optimized. Moreover, the same extracts were analyzed using Proton Nuclear Magnetic Resonance Spectroscopy (1H-NMR), to evaluate the applicability of a single urine extraction for multiplatform purposes. After the evaluation of experimental conditions, an LLE protocol using 2 mL of urine in the presence of 2 mL of 1 M sulfuric acid and sodium sulphate extracted with dichloromethane was found to be optimal. The optimized method was validated with the external standards and was found to be precise and linear, and allowed for detection of >400 peaks in a single run present in at least 50% of six samples—considerably more than the number of peaks detected by solid-phase microextracton fiber pre-concentration-GC-MS (328 ± 6 vs. 234 ± 4). 1H-NMR spectroscopy of the polar and non-polar extracts extended the range to >40 more (mainly low volatility compounds) metabolites (non-destructively), the majority of which were different from GC-MS. The more peaks detectable, the greater the opportunity of assessing a fingerprint of several compounds to aid biomarker discovery. In summary, we have successfully demonstrated the potential of LLE as a cheap and simple alternative for the analysis of VCs in urine, and for the first time the applicability of a single urine solvent extraction procedure for detecting a wide range of analytes using both GC-MS and 1H-NMR analysis to enhance putative biomarker detection. The proposed method will simplify the transport between laboratories and storage of samples, as compared to intact urine samples.

Double vs single primary tooth extraction in interceptive treatment of palatally displaced canines.

To compare the impact of primary canine and primary first molar extractions with extractions of only the primary canine regarding correction of palatally displaced canines (PDCs). Thirty-two children aged 9.5-13.5 years with 48 PDCs were randomly allocated to either the double-extraction group (DEG) or single-extraction group (SEG). Clinical and radiographic examinations were performed at baseline and at 6-month intervals until the canine emerged or orthodontic treatment was started. Outcome measures were: emergence of maxillary canine (yes/no), emergence of maxillary canine into a favorable position (yes/no), and maxillary canine positional change (angulation and sector). Factors influencing PDC emergence were analyzed using logistic regression. In the DEG, 64% (16/25) of canines emerged into the oral cavity vs 78% (18/23) in the SEG (P = .283). Favorable PDC position at trial end was seen in 64% (16/25) of the DEG vs 57% (13/23) of the SEG (P = .600). Significant distal movement of PDCs was recorded in the DEG and SEG, though no significant difference was observed between groups. Significant predictors of canine emergence were initial canine angulation (Angle A) (P = .008) and space conditions at T0 (P = .030). Double or single primary tooth extraction procedures are equivalent in supporting PDC eruption into the oral cavity and into a favorable position in the dental arch. Initial canine angulation and space assessments may be used as predictors of successful PDC eruption.

Validation and clinical application of a multiplex high performance liquid chromatography – tandem mass spectrometry assay for the monitoring of plasma concentrations of 12 antibiotics in patients with severe bacterial infections

ObjectiveUnpredictable pharmacokinetics of antibiotics in patients with life-threatening bacterial infections is associated with drug under- or overdosing. Therapeutic drug monitoring (TDM) may guide dosing adjustment aimed at maximizing antibacterial efficacy and minimizing toxicity. Rapid and accurate analytical methods are key for real-time TDM. Our objective was to develop a robust high-performance liquid chromatography-tandem mass spectrometry method (HPLC-MS/MS) for multiplex quantification of plasma concentrations of 12 antibiotics: imipenem/cilastatin, meropenem, ertapenem, cefepime, ceftazidime, ceftriaxone, piperacillin/tazobactam, amoxicillin, flucloxacillin, rifampicin, daptomycin. MethodsA single extraction procedure consisting in methanol plasma protein precipitation and H2O dilution was used for all analytes. After chromatographic separation on an Acquity UPLC HSS-T3 2.1 × 50 mm, 1.8 µm (Waters®) column, quantification was performed by electro-spray ionisation-triple quadrupole mass spectrometry with selected reaction monitoring detection. Antibiotics were divided in two pools of calibration according to the frequency of analyses requests in the hospital routine antibiotic TDM program. Stable isotopically-labelled analogues were used as internal standards. A single analytical run lasted less than 9 min. ResultsThe method was validated based on FDA recommendations, including assessment of extraction yield (96–113.8%), matrix effects, and analytical recovery (86.3–99.6%). The method was sensitive (lower limits of quantification 0.02–0.5 µg/mL), accurate (intra/inter-assay bias −11.3 to +12.7%) and precise (intra/inter-assay CVs 2.1–11.5%) over the clinically relevant plasma concentration ranges (upper limits of quantification 20–160 µg/mL). The application of the TDM assay was illustrated with clinical cases that highlight the impact on patients’ management of an analytical assay providing information with short turn-around time on antibiotic plasma concentration. ConclusionThis simple, robust high-throughput multiplex HPLC-MS/MS assay for simultaneous quantification of plasma concentrations of 12 daily used antibiotics is optimally suited for clinically efficient real-time TDM.

Marginal Bone Resorption Around Dental Implants Placed in Alveolar Socket Preserved Sites: A 5 Years Follow-up Study

The present study evaluated the clinical and radiological stability of hard and soft tissues following alveolar socket preservation (ASP) procedure with a follow-up of 5 year from implant insertion. The initial sample consisted of seven patients who underwent single tooth extraction and ASP procedure by means of demineralized bovine bone mineral particles covered with a porcine-derived non-cross-linked collagen matrix (CM). Each patient received a submerged single implant in the healed site. Mesial and distal peri-implant marginal bone resorption (MBR) rates were assessed radiographically at 1 year (T1) and 5 years (T2) after implant placement (baseline value). No dropouts occurred up to 5 years. At T1, the MBR was 0.08 ± 0.16 mm at the mesial aspect and 0.1 ± 0.12 mm at the distal aspect. This difference was not statistically significant (P = 0.867). At T2, the mesial MBR was 0.15 ± 0.17 mm and the distal MBR was 0.11 ± 0.14 mm, with a non-statistically significant difference (P = 0.532). Therefore, no statistically significant differences were detected comparing mesial and distal MBR at any time point. With respect to the intra-group comparisons, no differences were observed comparing the different study periods within each variable. Indeed, the comparison between T0, T1 and T2 was non-statistically significant at both mesial (P = 0.06) and distal (P = 0.06) aspects. After 5 years, the volume of the soft tissues appeared clinically well maintained with a natural aspect around dental implants and adjacent teeth. ASP using demineralized bovine bone mineral in combination with CM proved to be an effective technique to maintain stable dimensional volumes of both hard and soft tissues.

Heavy metal availability assessment using portable X-ray fluorescence and single extraction procedures on former vineyard polluted soils

The periodic application of copper-based fungicides (Bourdeaux mixture) to vineyards of the Mediterranean region has generated an important pollution source that in some cases requires a quick intervention due to the high bioavailable copper content measured. Despite some vineyards were abandoned 40 years ago, noticeable amounts of Cu and other man-related metals are still nowadays detected in soils. In the present work, the development of a mobility test for the available heavy metal (Cu, Pb, Zn and As) content in soil has been performed using portable X-ray fluorescence (FP-XRF) combined with single leaching test, and was applied to a calcareous soil of a former vineyard area in Catalonia (NE Spain). The combined methodology has provided useful information for fast and detailed risk assessment, in terms of mobility and bioavailability of metals. The anthropogenic contribution was evaluated by means of the Concentration Enrichment Ratios (CER) in soil. The results reflect a clear anthropogenic contribution for Cu, a partial anthropogenic contribution for Pb proceeding from an external pollution source, and a non-significant external contribution for As and Zn. The topsoil concentration ranges for Cu (70–128 mg kg−1) were found to be above the background level and several samples above the regional governmental limits (Generic Reference Levels or GRL values) for soil ecosystem protection and for human health (90 mg kg−1). The present study reveals that the use of FP-XRF equipment constitute a highly valid option for quick decision making during the field location, characterization and quantitative elemental analysis of soil samples for screening of potential pollutants such as heavy metals.

Dynamic Risk Assessment of Lead Pollution of Shooting Range Soil by Applying the Delayed Geochemical Hazard Model – A Case Study in Botswana

ABSTRACTThe delayed geochemical hazard (DGH) model was used to show that environmental pollution risk from Pb is a dynamic process as opposed to static. The DGH model describes the potential transformation and release of Pb partitioned in the different fractions of the soil that may be inert but becoming reactive when the conditions of the soil such as pH, moisture, and organic matter become favorable or when the Pb deposited into the soil exceeds the soil-holding capacity. As opposed to single extraction procedures that investigate the total amount of Pb deposited into shooting range soils, irrespective of whether that Pb is from inert or labile Pb species, the DGH model takes into consideration the mineralogical state of the Pb species and the ever-changing physicochemical characteristics of the soil. The total concentration of Pb does not give details on the bioaccessibility, bioavailability, mobility, and quantitative environmental pollution risk of deposited Pb to biota. This model was applied on a case study in shooting range soils highly polluted with Pb at concentrations of up to 38 406.87 mg/kg. The findings indicated that TAB shooting range located inside a military airbase in the southern part of Botswana may be classified as high-risk area of Pb DGH with over 65% of Pb capable of being transformed into the mobile and bioavailable chemical forms.

Abstract 243: Differential effects of combination treatment of biochanin A and statins on glioblastoma multiforme cell proliferation and cell metabolism

Abstract Glioblastoma Multiforme (GBM) is a deadly brain cancer and represents the most common central nervous system (CNS) tumors in adults. Although GBM does not metastasize, its aggressive growth and invasive nature are responsible for poor patient prognosis. The current standard therapy for newly diagnosed GBM patients involves surgical resection, followed by radiation and chemotherapy with Temozolomide. However, its rapid rate of infiltration into normal brain tissue ultimately renders the therapy ineffective. Epidemiological studies on dietary isoflavones (e.g., genistein, biochanin A) have shown their anti-cancer potential in different cancers. Though primarily used in management of hyperlipidemia and cardiovascular diseases, statins are known to exert anti-proliferative effects. We therefore hypothesized that a combination treatment of biochanin A &amp; statins (e.g., Atorvastatin, Lovastatin, Simvastatin, Fluvastatin and Pravastatin) exerts enhanced anticancer effects on GBM U-87 MG and T98 G cells. Our studies showed statins induced differential effects on viability of GBM cells in combination with biochanin A, with U-87 MG being more susceptible than T98 G cells. The combination treatment of biochanin A and atorvastatin also decreased invasion in U-87 MG cells. Additionally, cell metabolism studies using seahorse XFp analyzer showed a switch in their metabolic phenotype with an increase in mitochondrial respiration and a decrease in glycolytic activity with the combination treatment. We also performed metabolite extraction on GBM cells for a global unbiased profiling of metabolites using a single extraction procedure and dual separation analysis by LCMS. A differential analysis of alternative GBM treatment indicated statistically significant changes in carbohydrate and amino acid metabolism as well as alterations to various degradation pathway intermediates. Together, the combination treatment-induced effects on GBM cell lines are differential and our results may have potential implications in developing combination therapies with biochanin A in vivo and support the design of new and better therapies for the treatment of a lethal cancer like GBM. Citation Format: Vilas S. Desai, Eric Buchhalter, Max Cabanzo, Arushi Tiwari, Gagan Kaushal, James C. Lai, Alok Bhushan. Differential effects of combination treatment of biochanin A and statins on glioblastoma multiforme cell proliferation and cell metabolism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 243.

Development and Validation of a Fecal Extraction Procedure for the Assessment of Multiple Fecal Biomarkers of Intestinal Inflammation (P13-025-19)

ObjectivesFecal biomarkers have emerged as an important tool to assess intestinal inflammation and permeability. Commonly measured biomarkers include calprotectin (CP), myeloperoxidase (MPO), alpha-1-antitrypsin (AAT) and neopterin (NEO). We sought to develop a simple, fast and cost-effective single extraction procedure for use in determining all four biomarkers of interest. The applicability and sensitivity of this procedure for use in healthy adults was examined. MethodsSample extraction buffers and methods including sample weight, dilution, homogenization and centrifugation were all considered in the development of a single extraction procedure. An extraction buffer that included phosphate-buffered saline, phenylmethylsulfonyl fluoride, bovine serum albumin and Tween 20 was used to extract fecal samples. To assess the applicability and sensitivity of the single extraction procedure, concentrations of CP, MPO, AAT and NEO were measured using commercially available sandwich ELISA kits, according to manufacturer’s instructions. ResultsCP, MPO and AAT concentrations were measured in fecal samples of healthy adults (aged 50–80 years, n = 85) and found to be comparable to findings of previously published studies in healthy populations. Mean concentrations of CP and AAT were 3.6 ± 3.8 μg/g of wet weight (range 0.14–18.0 μg/g) and 2.3 ± 0.73 μg/g (range 0.76–5.2 μg/g), respectively. Mean fecal MPO concentrations were 135 ± 24 ng/g (range 3–1290 ng/g). NEO concentrations were examined in a subset of healthy adults (n = 10), with mean concentrations of 18 ± 1 ng/g (range 17–20 ng/g). ConclusionsWe demonstrated the efficacy of a single extraction procedure used to assess multiple fecal biomarkers of intestinal inflammation. This simple, fast and inexpensive extraction method will facilitate the determination of multiple fecal biomarkers which is critical in validating their use as clinical or predictive biomarkers of intestinal inflammation. Funding SourcesSupported by the U.S. Department of Agriculture – Agricultural Research Service (ARS), under Agreement No. 58–1950-4–003, the Bill & Melinda Gates Foundation and Canadian Institutes of Health Research Postdoctoral Fellowship.