N. Violante

The Assessment of Reference Values for Elements in Human Biological Tissues and Fluids: A Systematic Review

Abstract A survey is presented of the rationale of and the operative procedures for the assessment of reference values of minor and trace elements in human blood (whether entire, plasma, or serum), urine, milk, hair, kidneys, liver, and lungs. Literature sources available on this subject matter have been scanned over the past 30 years and reference values have been critically and selectively reported. The need for further harmonization of strategies and better comparability of experimental data is emphasized.

Exosome Release and Low pH Belong to a Framework of Resistance of Human Melanoma Cells to Cisplatin

Intrinsic resistance to cytotoxic drugs has been a main issue in cancer therapy for decades. Microenvironmental acidity is a simple while highly efficient mechanism of chemoresistance, exploited through impairment of drug delivery. The latter is achieved by extracellular protonation and/or sequestration into acidic vesicles. This study investigates the importance of extracellular acidosis and nanovesicle (exosome) release in the resistance of human tumour cell to cisplatin (CisPt); in parallel to proton pump inhibitors (PPI) ability of interfering with these tumour cell features. The results showed that CisPt uptake by human tumour cells was markedly impaired by low pH conditions. Moreover, exosomes purified from supernatants of these cell cultures contained various amounts of CisPt, which correlated to the pH conditions of the culture medium. HPLC-Q-ICP-MS analysis revealed that exosome purified from tumour cell culture supernatants contained CisPt in its native form. PPI pre-treatment increased cellular uptake of CisPt, as compared to untreated cells, in an acidic-depend manner. Furthermore, it induced a clear inhibition of exosome release by tumour cells. Human tumours obtained from xenografts pretreated with PPI contained more CisPt as compared to tumours from xenografts treated with CisPt alone. Further analysis showed that in vivo PPI treatment induced a clear reduction in the plasmatic levels of tumour-derived exosomes which also contained lower level of CisPt. Altogether, these findings point to the identification of a double mechanism that human malignant melanoma use in resisting to a dreadful cellular poison such as cisplatin. This framework of resistance includes both low pH-dependent extracellular sequestration and an exosome-mediated elimination. Both mechanisms are markedly impaired by proton pump inhibition, leading to an increased CisPt-dependent cytotoxicity.

Assessment of reference values for elements in human hair of urban schoolboys

Hair samples of youngsters (3-15 years of age) from several urban areas of Rome were analyzed to determine the content of 19 minor and trace elements with the aim of assessing Reference Values (RVs). Thirteen essential elements were taken into account, Ca, Co, Cr, Cu, Fe, Mg, Mn, Mo, Ni, P, Se, V and Zn. On the other hand, Al, As, Cd, Pb, Sr and Ti were also evaluated on the basis of their potential toxicity. Procedures were developed for the collection, storage and pre-analytical treatment of samples. Measurements were performed by inductively coupled plasma atomic emission spectrometry. Subgroups were formed according to age and sex. Significant differences were found for certain elements depending on age and sex. This was the case, e.g., for Ca which showed a mean value of 336 mg/kg for males and of 537 mg/kg for females. The sex-dependent pattern for this element was also apparent when the three age subgroups of 3-6, 7-10 and 11-15 years were compared. The overall RVs obtained (mg/kg) are as follows Al, 10.2; As, 0.09; Ca, 450; Cd, 0.23; Co, 0.67; Cr, 0.99; Cu, 22.1; Fe, 19.0; Mg, 28.0; Mn, 0.35; Mo, 0.43; Ni, 1.49; P, 195; Pb, 7.11; Se, 0.77; Sr, 1.20; Ti, 0.79; V, 1.22; and Zn 150.

Trace and major elements in whole blood, serum, cerebrospinal fluid and urine of patients with Parkinson's disease.

Summary.Quantifications of Al, Ca, Cu, Fe, Mg, Mn, Si and Zn were performed in urine, serum, blood and cerebrospinal fluid (CSF) of 26 patients affected by Parkinson’s disease (PD) and 13 age-matched controls to ascertain the potential role of biological fluids as markers for this pathology. Analyses were performed by Inductively Coupled Plasma Atomic Emission Spectrometry and Sector Field Inductively Coupled Plasma Mass Spectrometry. The serum oxidant status (SOS) and anti-oxidant capacity (SAC) were also determined. Results showed a decreasing trend for Al in all the fluids of PD patients, with the strongest evidence in serum. Calcium levels in urine, serum and blood of PD patients were significantly higher than in controls. Copper and Mg concentrations were significantly lower in serum of PD patients. Levels of Fe in urine, blood and CSF of patients and controls were dissimilar, with an increase in the first two matrices and a decrease in CSF. No significant difference was found in levels of Mn between patients and controls. Urinary excretion of Si was significantly higher in PD subjects than in controls. No clear difference between Zn levels in the two groups was found for serum, urine or CSF, but an increase in Zn levels in the blood of PD patients was observed. The SOS level in PD was significantly higher while the corresponding SAC was found to be lower in patients than in controls, in line with the hypothesis that oxidative damage is a key factor in the pathogenesis of PD. The results on the whole indicate the involvement of Fe and Zn (increased concentration in blood) as well as of Cu (decreased serum level) in PD. The augmented levels of Ca and Mg in the fluids and of Si in urine of patients may suggest an involuntary intake of these elements during therapy.

Dynamic reaction cell ICP-MS for determination of total As, Cr, Se and V in complex matrices: Still a challenge? A review

Mass interferences, caused by atomic or polyatomic species and having the same mass/charge ratio of the analyte, can be a severe limit for a reliable assay of trace and ultratrace elements by ICP-MS. The DRC™ technology uses a reaction gas to overcome these interferences. Reactions of charge exchange, atom transfer, adduct formation, condensation and analyte association/condensation are the main mechanisms. Interfering ions tend to react with the gas exothermally, while, the analyte reacts endothermally. Selecting the most appropriate reaction gas in DRC-ICP-MS is the very critical point for the determination of strongly interfered elements. A careful evaluation of the reaction mechanisms and the chemistry involved are required. The DRC allows the use of different gases, among them, ammonia (NH(3)), methane (CH(4)), hydrogen (H(2)) and oxygen (O(2)) are the most known, but there are other potentially useful gases like nitrous oxide (N(2)O), nitrogen oxide (NO), carbon dioxide (CO(2)), fluoromethane (CH(3)F), sulphur hexafluoride (SF(6)) and carbon disulfide (CS(2)). This paper provides a review on the analytical challenges for a reliable assay of As, Cr, Se and V by DRC-ICP-MS and illustrates different approaches and mechanisms involved in the analysis of polymers, biological fluids (serum, urine and whole blood), rock, soil and particulate matter.

Assessment of reference values for elements in hair of urban normal subjects

Hair is quite attractive as a biological indicator because of the simplicity of sampling and storage. The analysis of trace and minor elements in this tissue has been used in a diversity of applications, including nutritional, environmental, medical, and occupational studies. Within this context the availability of reliable reference values plays a key role. Scalp hair samples taken from 225 urban subjects not older than 15 years were analyzed for 18 elements. The method chosen for the quantification of elements in hair has been ICP-AES in all cases because of its multielemental capability, wide dynamic range, adequate detection limits, and relative freedom from matrix interferences. The following mean values were obtained (in μg g−1): Al, 11.6; As, 0.10; Ca, 416; Cd, 0.20; Co, 0.86; Cr, 0.67; Cu, 22.4; Fe, 17.9; Mg, 25.9; Mn, 0.41; Mo, 0.44; Ni, 0.97; P, 185; Pb, 8.10; Se, 0.84; Ti, 1.05; V, 1.44; and Zn 144. These data were analyzed statistically to ascertain possible significant differences according to sex and age.

Method validation for determination of arsenic, cadmium, chromium and lead in milk by means of dynamic reaction cell inductively coupled plasma mass spectrometry.

With Regulation No. 1881/2006 the European Union fixed a maximum level for lead in milk. Consequently, there is the need to determine very low concentration of elements that may be present in milk in trace and ultratrace levels. Quadrupole inductively coupled plasma mass spectrometry (Q-ICP-MS) combined with dynamic reaction cell (DRC) has been widely employed in order to reach very low concentration, requested for this product. Furthermore, the DRC technology can help in removing polyatomic and argon-based interferences. In the present study, a method for the determination of arsenic, cadmium, chromium and lead in bovine milk was validated according to the EU common standards by means of DRC-ICP-MS. The main parameters evaluated in the validation were: recovery, repeatability and within-laboratory reproducibility, detection and quantification limits, linearity range and measurement uncertainty. Additionally, stability studies of the analyte in solution and ruggedness studies were carried out. The results obtained for limit of detection (LoD) and limit of quantification (LoQ) in microg kg(-1) were respectively: As, 3.1 and 9.5; Cd, 0.08 and 0.24; Cr, 0.229 and 0.693; Pb, 0.5 and 1.5. While for the recovery: As, 91%; Cd 96%; Cr 99%; Pb, 95%. As for the repeatability: As, 7%; Cd, 3%; Cr, 6%; Pb, 4%.

Metal changes in CSF and peripheral compartments of parkinsonian patients

BACKGROUND Involvement of metals in the risk of developing Parkinsons disease (PD) has been suggested. In the present study, concentration of metals in cerebrospinal fluid (CSF), blood, serum, urine and hair of 91 PD patients and 18 controls were compared. METHODS Blood and hair were microwave digested, while CSF, serum and urine were water-diluted. Elements quantification was achieved by Inductively Coupled Plasma Atomic Emission Spectrometry and Sector Field Inductively Coupled Plasma Mass Spectrometry. RESULTS Some metal imbalances in PD were observed: i), in CSF, lower Fe and Si; ii), in blood, higher Ca, Cu, Fe, Mg and Zn; iii), in serum, lower Al and Cu; iv), in urine, lower Al and Mn, higher Ca and Fe; and v), in hair, lower Fe. The ROC analysis suggested that blood Ca, Fe, Mg and Zn were the best discriminators between PD and controls. In addition, hair Ca and Mg were at least 1.5 times higher in females than in males of patients and controls. A decrement with age of patients in hair and urine Ca and, with less extent, in urine Si was observed. Magnesium concentration in CSF decreased with the duration and severity of the disease. Elements were not influenced by the type of antiparkinsonian therapy. CONCLUSIONS Variation in elements with the disease do not exclude their involvement in the neurodegeneration of PD.

Human hair as a marker of pollution by chemical elements emitted by a thermoelectric power plant

Abstract A preliminary study was undertaken to assess the risk posed to the environment and health of residents by trace elements released by fuel combustion in the thermoelectric power plant of Pian de’ Gangani (Montalto di Castro, Latium, Italy). In this context, analysis of human hair can be considered as an advantageous approach to monitor the exposure of population to the adverse effects of the power plant, which is now fully operative. To this purpose 92 healthy children (aged 9–10 years) were selected from two primary schools of Montalto di Castro and Pescia Romana. The young age of the donors guarantees that the effects of cosmetic treatment and occupational exposure on the representativeness of data are minimized. Hair samples were collected, washed and digested as prescribed by already well-established procedures. Inductively coupled plasma mass spectrometry (ICP-MS) and inductively coupled plasma atomic emission spectrometry (ICP-AES) were employed for the determination of the trace and minor elements of interest (As, Ca, Cd, Co, Cr, Cu, Fe, Mg, Mn, Ni, P, Pb, Sb, Se, Tl, V and Zn). The entire analytical procedure was checked for both measurement precision and accuracy by using the Certified Reference Material BCR No. 397 Human Hair. The mean values obtained were (in μg g−1): As, 0.159±0.044; Ca, 393±187; Cd, 0.047±0.055; Co, 0.040±0.038; Cr, 0.704±0.208; Cu, 12.0±10.3; Fe, 12.1±10.8; Mg, 40.6±27.7; Mn, 0.383±0.296; Ni, 0.868±1.331; P, 141±138; Pb, 1.64±1.63; Sb, 0.041±0.058; Se, 0.430±0.135; Tl, 0.001±0.001; V, 0.185±0.158; and Zn, 177±173. These experimental figures can be assumed to be the baseline values prior to the full operation of the power plant. Within the next 4–5 years it is planned to conduct a similar study with a new group of subjects, comparable for age, sex, style of life and exposure mode, to the present ones so as to detect possible trends in the bioaccumulation of the above elements.

Occupational exposure of goldsmith workers of the area of Rome to potentially toxic metals as monitored through hair analysis

In continuation of an investigation recently carried out to monitor through hair analysis the occupational exposure of goldsmith workers to potentially toxic elements, another study was performed to extend the same methodological approach to the goldsmiths of Rome. This research was part of the project P.R.O.Art. undertaken by the Italian National Research Council in cooperation with the Ministry of Industry and the National Craftsmens Federation with the purpose of supporting goldsmith activities and trade. Sampling of hair, washing and sample digestion followed well-established procedures. Silver, Au, Cd, Co, Cr, In, Ni, Pb and Pt were determined by means of inductively coupled plasma mass spectrometry (ICP-MS), whereas Hg was analyzed using the flow injection mercury system (FIMS). On the other hand, the expected relatively high concentrations of Cu and Zn in hair allowed for the use of inductively coupled plasma atomic emission spectrometry (ICP-AES). Data obtained were statistically treated by applying the non-parametric Kruskal-Wallis test. A significant difference, at the level of P < 0.05, between exposed and unexposed subjects in the Rome area was observed only for Au.