Quantitative assessment of the metabolic products of iron oxide nanoparticles to be used as iron supplements in cell cultures.

Iron nanoparticles (NPs) metabolism is directly associated to human health due to their use as anemia treatment and should be studied in detail in cells. Here we present a speciation strategy for the determination of the metabolic products of iron oxide nanoparticles coated by tartaric and adipic acids in enterocytes-like cell models (Caco-2 and HT-29). Such methodology is based on the use of SDS-modified reversed phase high performance liquid chromatography (HPLC) separation using inductively coupled plasma-mass spectrometry (ICP-MS) as Fe selective detector. Post-column isotope dilution analysis is used as quantification tool by adding 57Fe as isotopically enriched standard. To assess the separation capability of the method, two different iron nanostructures: iron sucrose nanoparticles -Venofer®- used as model suspension and iron tartrate/adipate-modified nanoparticles, both of about 4 nm (core size) were evaluated. The two nanostructures were injected into the system showing good peak profiles and quantitative elution recoveries (>80%) in both cases. In addition, both nanoparticulate fractions could be based-line separated from ionic iron species, which needed to be complexed with 1 mM citrate to elute from the column. Exposed cells up to 0.5 mM of iron tartrate/adipate-modified nanoparticles were specifically treated to extract the internalized NPs and the extracts examined using the proposed strategy. The obtained results revealed the presence of three different fractions corresponding to nanoparticle aggregates, dispersed nanoparticles and soluble iron respectively in a single chromatographic run. Quantitative experiments (column recoveries ranging from 60 to 80%) revealed the presence of the majority of the Fe in the nanoparticulated form (>75%) by summing up the dispersed and aggregate particles. Such experiments point out the high uptake and low solubilization rate of the tartrate/adipate NPs making these structures highly suitable as Fe supplements in oral anemia treatments.

Single-cell analysis by ICP-MS/MS as a fast tool for cellular bioavailability studies of arsenite.

Single-cell inductively coupled plasma mass spectrometry (SC-ICP-MS) has become a powerful and fast tool to evaluate the elemental composition at a single-cell level. In this study, the cellular bioavailability of arsenite (incubation of 25 and 50 µM for 0-48 h) has been successfully assessed by SC-ICP-MS/MS for the first time directly after re-suspending the cells in water. This procedure avoids the normally arising cell membrane permeabilization caused by cell fixation methods (e.g. methanol fixation). The reliability and feasibility of this SC-ICP-MS/MS approach with a limit of detection of 0.35 fg per cell was validated by conventional bulk ICP-MS/MS analysis after cell digestion and parallel measurement of sulfur and phosphorus.

An approach for quantification of platinum distribution in tissues by LA-ICP-MS imaging using isotope dilution analysis.

Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) has been revealed as a convenient technique for trace elemental imaging in tissue sections, providing elemental 2D distribution at a quantitative level. For quantification purposes, in the last years several approaches have been proposed in the literature such as the use of CRMs or matrix matched standards. The use of Isotope Dilution (ID) for quantification by LA-ICP-MS has been also described, being mainly useful for bulk analysis but not feasible for spatial measurements so far. In this work, a quantification method based on ID analysis was developed by printing isotope-enriched inks onto kidney slices from rats treated with antitumoral Pt-based drugs using a commercial ink-jet device, in order to perform an elemental quantification in different areas from bio-images. For the ID experiments 194Pt enriched platinum was used. The methodology was validated by deposition of natural Pt standard droplets with a known amount of Pt onto the surface of a control tissue, where could be quantified even 50pg of Pt, with recoveries higher than 90%. The amount of Pt present in the whole kidney slices was quantified for cisplatin, carboplatin and oxaliplatin-treated rats. The results obtained were in accordance with those previously reported. The amount of Pt distributed between the medullar and cortical areas was also quantified, observing different behavior for the three drugs.

Properties of in situ generated gold nanoparticles in the cellular context.

Gold nanostructures that serve as probes for nanospectroscopic analysis of eukaryotic cell cultures can be obtained by the in situ reduction of tetrachloroauric acid (HAuCl4). To understand the formation process of such intracellularly grown particles depending on the incubation medium, the reaction was carried out with 3T3 fibroblast cells in three different incubation media, phosphate buffer, Dulbecco's Modified Eagle Medium (DMEM), and standard cell culture medium (DMEM with fetal calf serum). The size, the optical properties, the biomolecular corona, and the localization of the gold nanoparticles formed in situ vary for the different conditions. The combination of surface-enhanced Raman scattering (SERS) and laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) microscopic mapping and transmission electron microscopy (TEM) provides complementary perspectives on plasmonic nanoparticles and non-plasmonic gold compounds inside the cells. While for the incubation with HAuCl4 in PBS, gold particles provide optical signals from the nucleus, the incubation in standard cell culture medium leads to scavenging of the toxic molecules and the formation of spots of high gold concentration in the cytoplasm without formation of SERS-active particles inside the cells. The biomolecular corona of nanoparticles formed in situ after incubation in buffer and DMEM differs, suggesting that different intracellular molecular species serve for reduction and stabilization. Comparison with data obtained from ready-made gold nanoparticles suggests complementary application of in situ and ex situ generated nanostructures for optical probing.

A simple metal staining procedure for identification and visualization of single cells by LA-ICP-MS.

High lateral resolution of metal detection in single cells by use of laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) demands powerful staining methods. In this work different staining procedures for the single cell analysis with LA-ICP-MS were optimized. An iridium intercalator was utilized to stain the cell nuclei whereas the whole cell was stained by the use of maleimido-mono-amide-DOTA (mDOTA) complexing lanthanide(iii) ions. The content of the artificially introduced metals per cell was quantified using a matrix matched calibration approach based on cellulose membranes onto which standards were spotted by a microarray spotter. Absolute metal stain amounts in the range of 2.34 to 9.81 femtomole per cell were determined. The metal staining procedures allow direct identification and visualization of single cells and their cell compartments by element microscopy without the use of bright field images of the sample.

A survey of lead pollution in Chhattisgarh State, central India.

Lead (Pb) is of major environmental concern due to its toxicological importance. The anthropogenic emission of Pb is at least 100 times higher than natural emissions. Soil and dust are significant sources of Pb exposure. Lead is generally immobile in soil and accumulates in the upper layers. Lead particles may enter homes via shoes, clothes, pets, and windows. Central India is rich in deposits of natural resource materials such as coal, pyrite, dolomite, and alumina that contain Pb and other heavy metals at the trace levels, and the substantial exploitation of these materials has tended to increased contamination of water and geological formations. Here we present data on Pb concentrations in the water, soil and sediment samples (n=158) collected from 70 locations in Chhattisgarh state, Raipur region. Lead concentrations in the surface water (n=44), groundwater (n=44), soils (n=60) and sediments (n=10) ranged from 6 to 1410, 3 to 52, 12.8 to 545, and 31 to 423 microg g(-1), with mean values of 305, 16, 102 and 190 microg g(-1), respectively. Most of the Pb fractions of >80% can be leached out with the chemical extractants EDTA, acetic acid, and hydroxylamine hydrochloride. Lead has accumulated in the soil clay fraction due to its relatively large surface area and decreases with increasing depth in the soil profile.

Arsenic contamination in water, soil, sediment and rice of central India.

Arsenic contamination in the environment (i.e. surface, well and tube-well water, soil, sediment and rice samples) of central India (i.e. Ambagarh Chauki, Chhattisgarh) is reported. The concentration of the total arsenic in the samples i.e. water (n = 64), soil (n = 30), sediment (n = 27) and rice grain (n = 10) were ranged from 15 to 825 microg L(-1), 9 to 390 mg kg(-1), 19 to 489 mg kg(-1) and 0.018 to 0.446 mg kg(-1), respectively. In all type of waters, the arsenic levels exceeded the permissible limit, 10 microg L(-1). The most toxic and mobile inorganic species i.e. As(III) and As(V) are predominantly present in water of this region. The soils have relatively higher contents of arsenic and other elements i.e. Mg, Al, Si, K, Ca, Ti, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, As, Ga, Zr, Sn, Sb, Pb and U. The mean arsenic contents in soil of this region are much higher than in arsenic soil of West Bengal and Bangladesh. The lowest level of arsenic in the soil of this region is 3.7 mg kg(-1) with median value of 9.5 mg kg(-1). The arsenic contents in the sediments are at least 2-folds higher than in the soil. The sources of arsenic contamination in the soil of this region are expected from the rock weathering as well as the atmospheric deposition. The environmental samples i.e. water, soil dust, food, etc. are expected the major exposure for the arsenic contamination. The most of people living in this region are suffering with arsenic borne diseases (i.e. melanosis, keratosis, skin cancer, etc.).

Size fractionation of non-volatile dissolved organic compounds and metal species in German white wines by combining on-line tangential-flow multistage ultrafiltration, a home-built carbon analyser, and inductively coupled plasma mass spectrometry.

Organic metal species and their size fractions in three German white wines were characterized by combining multistage ultrafiltration (MST-UF), determination of non-volatile dissolved organic carbon (NV-DOC) by a home-built carbon analyser, and metal quantification by inductively coupled plasma mass spectrometry (ICP-MS). First, NV-DOC and metal species in selected "dry" German white wines were fractionated on-line using MST-UF in the size range of >100 kDa to <1 kDa. For this purpose a 20 mL sample of the wine under study diluted 1:10 with high-purity water was processed through a cascade system of hydrophilized polyethersulfone-based flat membranes of decreasing cut-off (100, 50, 10, 5, and 3 kDa). An aliquot of the fraction <3 kDa was additionally processed through a commercial UF tube (MidGee system, cut-off: 1 kDa) to obtain low-molecular size fractions also. A home-built carbon analyser was applied to determine NV-DOC in the wines and their size fractions. The NV-DOC found in a German reference wine and its size fractions was as follows: total NV-DOC: 8.97 mg mL(-1); F(1) (>100 kDa), 0.15%; F(2) (50-100 kDa), 0.44%; F(3) (10-50 kDa), 0.74%; F(4) (5-10 kDa), 0.76%; F(5) (5-3 kDa), 0.7%; F*(6) (3-1 kDa), 0.9%; F(7) (<1 kDa), 81.6% (related to total NV-DOC). The NV-DOC recovery was 85.2%. Accordingly, most of the NV-DOC in this wine consists of low-molecular mass organic compounds of <1 kDa, presumably carboxylic acids as typical in wine. Parallel metal determinations in these wines and their fractions were performed by ICP-MS. The measurements showed that the major part of the metals investigated, up to 25 elements, were dissolved in the size fraction of <1 kDa except Ba, Sr and Pb which appeared also in other fractions. In addition, conventional UV-VIS spectroscopy was applied to characterise the studied wines and their size fractions. According to this, the UV absorbance between 254 and 280 nm of these white wines shows a parallel trend to their NV-DOC.

Development of a procedure for the multi-element determination of trace elements in wine by ICP-MS.

An inductively coupled plasma mass spectrometric (ICP-MS) procedure has been developed for the determination of trace elements in wine. The procedure consists in simple 1+1 dilution of the wine and semi-quantitative analysis (without external calibration) using In as internal standard. Thirty-one elements at concentrations ranging from 0.1 mg mL(-1) to 0.5 ng mL(-1) can be determined by ICP-MS analysis with and without digestion. It was investigated whether a matrix effect observed for EtOH in the wine matrix can be overcome by application of a micro-concentric nebulizer with a membrane desolvator (MCN 6000). The results obtained for the MCN 6000 are compared with those obtained by use of a conventional Meinhard nebulizer. It is shown that the observed matrix effect can only be compensated by use of an internal standard for the Meinhard nebulizer, but not for the MCN 6000. Results for ICP-MS are compared with those obtained by total reflection X-ray fluorescence spectrometry (TXRF).

Analysis of protein phosphorylation by capillary liquid chromatography coupled to element mass spectrometry with 31P detection and to electrospray mass spectrometry.

A method for phosphopeptide identification by capillary liquid chromatography (muLC) interfaced alternatively to element mass spectrometry (inductively coupled plasma mass spectrometry, ICPMS) and to electrospray ionization mass spectrometry (ESI-MS) is described. ICPMS is used for 31P detection and ESI-MS provides the corresponding molecular weight information. Alignment of the two separate muLC runs is performed using the baseline distortion at the elution front, which shows up in both muLC-ICPMS and muLC-ESI-MS. Both a quadrupole and a magnetic sector field mass analyzer were used in combination with ICP. The detection limit achieved for the muLC-ICP-HRMS runs is approximately 0.1 pmol of phosphopeptide injected. Without any further precautions, contamination by phosphate-containing compounds at this level was found to be uncritical. The method is demonstrated for the analysis of a complex mixture of synthetic phosphopeptides and a set of tryptic digests of three phosphoproteins. These include beta-casein, activated human MAP kinase ERK1, and protein kinase A catalytic subunit. The tryptic phosphopeptides of these proteins could all be detected and identified by our new strategy. Analysis of three fractions of protein kinase A catalytic subunit with different phosphorylation status gives direct access to the order in which the phosphorylation of the four phosphorylation sites occurs. The two most important aspects of using muLC-ICPMS with 31P detection for phosphopeptide identification are (i) that a high selectivity is achieved and (ii) that the signal intensity is independent of the chemical form of phosphorus and directly proportional to the molar amount of 31P in the muLC eluate. Thus, muLC-ICPMS with 31P detection is introduced as a new, robust, and specific method in phosphoproteomics.

Speciation of nickel in airborne particulate matter by means of sequential extraction in a micro flow system and determination by graphite furnace atomic absorption spectrometry and inductively coupled plasma mass spectrometry.

A four-stage sequential extraction procedure for the speciation of nickel has been applied to ambient aerosol samples. The determination of the soluble, sulfidic, metallic and oxidic Ni fractions in particulate matter was carried out by graphite furnace (electrothermal) atomic absorption spectrometry (ETAAS) and inductively coupled plasma mass spectrometry (ICP-MS). An EDTA solution, a mixture of diammonium citrate and hydrogen peroxide, and a KCuCl3 solution were used as leaching agents for the determination of the soluble, sulfidic and metallic species, respectively, and nitric acid was used for the determination of oxidic compounds after microwave digestion of particulate matter sampled on filters. A new micro scale filter holder placed in a closed flow injection analysis (FIA) system for use in nickel speciation by means of sequential extraction, and the results of the optimisation of the extraction conditions are described. The temperature program for ETAAS was optimised for all extraction solutions with the aid of temperature curves. Pyrolysis temperatures of 900. 600 and 1,000 degrees C were found to be optimum for EDTA, hydrogen peroxide plus ammonium citrate and KCuCl3-containing solutions, respectively. Airborne dust was sampled on lilters at two locations near to a metallurgical plant in Dortmund, Germany. Concentrations in the low ng m(-3) range down to the detections limits (0.1-0.3 ng m(-3)) and various nickel species were found to be present in the collected dust. The mean fractions of total nickel (sampling period of one month) were found to contain 36+20% of soluble, 6 +/- 4% of sulfidic, 11 +/- 15% of metallic and 48 +/- 18% of oxidic nickel.

Determination of soluble and insoluble nickel compounds in airborne particulate matter by graphite furnace atomic absorption spectrometry and inductively coupled plasma mass spectrometry.

An extraction procedure for the determination of soluble and insoluble nickel and its compounds in ambient air dust was investigated employing a special device for the generation of test aerosols and using water-soluble NiCl2, partly water-soluble NiCO3 and water-insoluble NiO as model compounds. Additionally, results of the separation and determination of different nickel species down to some ng/m3 in ambient aerosols are discussed. The extraction was carried out with a solution containing 0.01 mol/L EDTA in order to determine partly water-soluble compounds such as NiCO3 and water-soluble, non-toxic nickel compounds in one step. Airborne dust was sampled on filters at locations close to two metallurgical plants in Northrhine-Westphalia (Germany), and first results on the nickel concentration (mean (median) values over a period of 4 months: 8.6+/-6.5 ng/m3 (6.7 ng/m3) and 27.7+/-36 ng/m3 (10.8 ng/m3), respectively) in the collected dust are presented. For EDTA-soluble nickel compounds the maximum and mean fractions of total nickel were found to be 77.1% and 18.6+/-12%, respectively.

Quantitative determination of DNA adducts using liquid chromatography/electrospray ionization mass spectrometry and liquid chromatography/high-resolution inductively coupled plasma mass spectrometry.

The quantitative determination of nucleotides from DNA modified by styrene oxide is described using a combination of inductively coupled plasma high-resolution mass spectrometry (ICP-HRMS) and electrospray ionization mass spectrometry (ESI-MS), both interfaced to reversed-phase high-performance liquid chromatography (HPLC). LC/ICP-MS (resolution > 1500 to discriminate against 15N16O+ and 14N16OH+) was employed to determine quantitatively the content of modified nucleotides in standard solutions based on the signal of phosphorus; phosphoric acid served as an internal standard. By means of the standard addition technique the sensitivity of the LC/ESI-MS approach was subsequently determined. Since a comparison of UV, ICP and ESI-MS data suggested that in ESI-MS the ionization efficiency of the adducts is identical within the error limits, quantitative determination of all adducts is possible. For LC/ESI-MS with single ion monitoring, the detection limit for styrene oxide adducts of nucleotides was determined to be 20 pg absolute or 14 modified in 10(8) unmodified nucleotides in a 5 micrograms DNA sample, which comes close to the best methods available for the detection of chemical modifications in DNA.