Trace elements and antioxidant enzymes in extremely low birthweight infants.

Oxygen radicals are believed to contribute to typical diseases of prematurity, such as bronchopulmonary dysplasia (BPD), intraventricular haemorrhage (IVH), retinopathy of prematurity (ROP) and necrotising enterocolitis (NEC). Our aim was to investigate whether these disorders are associated with disturbances in antioxidant enzyme activities and with low trace elements, which are co-factors of antioxidant enzymes. 209 infants with birthweight less than 1000g were enrolled into a European multicentre randomised erythropoietin (rhEPO) trial; 155 developed one or more of the above mentioned diseases. We analysed Zn, Cu, Fe, Se in plasma and red blood cells (RBCs), superoxide dismutase (CuZn-SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) in RBCs on the 3rd and 68th day of life. Zn, Fe, Se in plasma, and Se in RBCs decreased (p<0.01), and Zn in RBC (p<0.001), CuZn-SOD (p<0.01) and CAT increased (p<0.05), whereas GSH-Px remained unchanged. No differences were observed between the rhEPO and control groups. Antioxidant enzyme activities did not correlate with gestational age. In infants with BPD, IVH, ROP, or NEC, CuZn-SOD and CAT (p<0.05) were higher at day 68 than in infants without these diseases. CuZn-SOD and GSH-Px at 3 days and CuZn-SOD at 68 days correlated positively (p<0.05) with the duration of oxygen treatment. In conclusion, in ELBW infants, trace element concentrations decreased over the first 10 weeks of life. Lower trace element concentrations, did not affect the activities of CuZn-SOD, GSH-Px, and CAT. Typical diseases of prematurity were not associated with decreased antioxidant enzyme activities.

Determination of normal human intrathyroidal iodine in Caracas population.

This study focuses on the determination of iodine content in healthy thyroid samples on male population from Caracas, Venezuela. Contribution to establish a baseline of iodine content in thyroid glands and hence to compare the iodine thyroid concentration of the Venezuelan population with other countries is also our objective. Male post-mortem individual samples were analyzed using a spectrophotometric flow injection method, based on the Sandell-Kolthoff reaction. The median intrathyroidal iodine concentration was 1443+/-677 microg/g (wet weight), ranging from 419 to 3430 microg/g, which corresponds to a median of total iodine content of 15+/-8 mg (ranging from 4 to 37). These results were higher than those values reported in the literature. No correlation of iodine content with age or weight of the healthy gland was found.

Nutrition of very low birth weight infants fed human milk with or without supplemental trace elements: a randomized controlled trial.

BACKGROUND: Very low birth weight infants (<1500 g) have high nutritional needs. Deficiencies of minerals, trace elements (especially zinc) may develop as a result of rapid growth, low body stores and low content of these substances in human milk We hypothesized that fortification of human milk might prevent deficiencies. METHODS: Prospective, randomized trial to evaluate mineral, trace element, thyroid status and growth of infants fed human milk fortified with different amounts of calcium, phosphorus and protein, with (BMF) or without (FM 85) trace elements. Sixty-two infants, 1000 to 1499 g birth weight, were randomized. Minerals and trace elements in serum, red blood cells and human milk and alkaline phosphatase activity, TSH, T4 and FT4 in serum were measured once until the fifth day and at 3 and 6 weeks of life. Clinical course and anthropometric measurements were recorded. RESULTS: Intake of zinc, copper, manganese, calcium, phosphorus and magnesium was higher in the BMF group (P < 0.001). Serum zinc concentrations <0.49 mg/L occurred in 12% of the FM 85 group and 7% of the BMF group at 6 weeks (not significant). Median alkaline phosphatase activity was 436/379 IU/L in the FM 85/BMF group at 6 weeks (P < 0.01). The FM 85 group showed a higher weight gain (P < 0.05), possibly because of higher caloric (P < 0.01) and protein intake (P < 0.05) at 3 weeks. CONCLUSIONS: Zinc deficiency was rare. Elevated intake of calcium, phosphorus and zinc was associated with lower serum alkaline phosphatase activity but did not influence serum zinc concentration.

CZE-ICP-MS separation of metallothioneins in human brain cytosols: comparability of electropherograms obtained from different sample matrices.

Capillary zone electrophoresis, with its high resolution capability in the separation of different compounds, is well suited for the investigation of metal-containing proteins, especially when elemental detection is conducted using hyphenated inductively coupled plasma-mass spectrometry. A major problem in the separation of proteins in body fluids is caused by the effects of different sample matrix composition. The migration time of proteins varies significantly, depending on the nature of the matrix. Electropherograms are consequently difficult to compare and the peak identification is uncertain. Pre-analytical steps for the reduction of matrix compounds enhance the quality of the data, but the results are still unsatisfactory. This paper describes a technique for obtaining electropherograms that can be used for comparison purposes by correction of the data with the aid of time markers. A mixture of five substances (caesium chloride, arsenocholine, arsenobetaine, dimethylarsinic acid and monomethylarsonic acid) was added in a separate injection step. Ionic caesium eluted at the start of the separation and the other four markers appeared throughout and at the end of the electropherogram. All electropherograms were normalized to a reference run by recalculation of the time axis using the time markers. The method was applied to the analysis of human brain cytosols. Samples were separated after different pre-treatment steps and were compared, with special emphasis on the detection of the isoform metallothionein-3.

Determination of protein-bound trace elements in human cell cytosols of different organs and different pathological states.

In the human body, there exists over 200 different cell types, which differ in size and structure and have specialised functions in the organism. Therefore it can be assumed that these different cells also contain different proteins necessary to carry out the respective specialised functions. This supposed different metalloprotein composition in different human organs cannot be demonstrated by determination of total element concentrations. Therefore investigations of the different protein-bound forms of the elements were achieved by speciation analysis: The biomolecules were separated by size exclusion chromatography and the elements detected on-line in the eluate by a hyphenated inductively coupled plasma mass spectrometer (ICP-MS). For the interpretation of the obtained element profiles, an identification of the signals and their assignment to different metalloproteins was necessary. This identification was carried out by means of specific protein assays, i.e. enzymatic assays or immunochemical reactions, in collected fractions of the chromatographic separations. A comparison of the element binding pattern in cytosols of different human organs was then possible. The optimised method was applied to tissue cytosols of different human organs. As expected, the element patterns varied for different organs of the same patient and for the same organ of patients with different diseases. Metalloproteins and their bound metals could consequently be considered as biological markers for physiological differences or pathological changes in human tissues.

Environmental exposure to trace elements and risk of amyotrophic lateral sclerosis: a population-based case-control study.

We analyzed the association between the environmental exposure to trace elements and the risk of sporadic amyotrophic lateral sclerosis (ALS) in a population-based case-control study in the Emilia-Romagna region in northern Italy. We evaluated exposure to selected trace elements by measuring toenail concentrations of the same by means of inductively coupled plasma optical spectrometry and instrumental neutron activation analysis. The final number enrolled in the study was 22 patients and 40 controls. Disease progression, assessed through a clinical score, was generally unassociated with toenail trace element levels, with the exception of an inverse relation with zinc and selenium content and a direct correlation with copper concentration. In logistic regression analysis, we found no evidence of an association between ALS risk and toenail content of cadmium, lead, copper, zinc, manganese, selenium, chromium, cobalt, iron, and aluminum. This investigation does not suggest a major role in sporadic ALS etiology of environmental exposure to these trace elements, though results for zinc, selenium, and copper should be evaluated with caution due to the potential limitations of toenails as biomarkers of chronic exposure in patients.

Characterization and quantification of metallothionein isoforms by capillary electrophoresis-inductively coupled plasma-isotope-dilution mass spectrometry.

In a new approach to the characterization and quantification of metallothionein isoforms an on-line isotope-dilution method in combination with the coupling of capillary electrophoresis (CE) to an inductively coupled plasma-sector field mass spectrometer (ICP-SFMS) is reported. Metallothionein (MT) isoforms are separated by CE and the elements Cu, Zn, Cd, and S are detected simultaneously by use of ICP-SFMS in the medium resolution mode. On-line isotope dilution is performed by continuous introduction of an isotopically enriched, species-unspecific spike solution after the separation step. MT from rabbit liver and a further purified MT-1 isoform were quantified by determination of sulfur, and the stoichiometric compositions of the metalloprotein complexes are characterized by determination of their sulfur-to-metal ratios.

Speciation analysis of trace elements in the brains of individuals with Alzheimer's disease with special emphasis on metallothioneins.

A speciation analysis of protein-bound elements in the cytosol of human brain was achieved by size exclusion chromatographical separation of the biomolecules and on-line detection of the metal profiles in the eluate by hyphenated inductively coupled plasma-mass spectrometry. Post-mortem samples from Alzheimer's disease brains and from brains of a control group were investigated to elucidate changes in the trace element distribution during the pathological process. Special attention was paid to the metallothioneins (MT) - cysteine-rich, metal-binding proteins of low molecular weight, existing in several isoforms. The isoform MT-3 is found especially in the brain and has a growth inhibition function on neurons. The MT peaks were identified in the element profiles. For this purpose, the metal binding capability and the heat stability of MT were taken into consideration. For verification, a comparison with pure MT-3 was carried out and further biochemical and analytical methods were applied to the fractions of the chromatographical run. A comparison between Alzheimer's disease and control brains showed a significant difference concerning the MT-1/-2 and MT-3 metal levels, leading to the assumption that there were oxidative processes having taken place in the Alzheimer's brain samples.

Sampling and processing of biopsy samples for speciation studies of cytosolic metalloproteins.

Studies to specify various metalloproteins in cell cytosol of tissues using chromatographic separation methods and plasma mass spectrometry for element detection require a careful step by step sample preparation. These steps involve (i) bioptic sample removal from tissue, (ii) extraction and preparation of the tissue supernatant, (iii) the chromatographic separation of the proteins, and finally (iv) the on-line transfer of the column eluate into the plasma mass spectrometer. Each of the analytical steps has to be carefully monitored to avoid undesired changes in the sample composition which could be caused by enzymatic and/or oxidative processes as well as by external element contamination. This presentation introduces a concept to ensure that environmental element contamination does not occur to bias the analytical results.

Brain Iron and Zinc Contents of German Patients with Alzheimer Disease.

Our first project aimed to determine the average values of Fe and Zn in normal German human brain (5 individuals, 10 brain parts). Determinations were carried out by instrumental neutron activation analysis in Berlin. Quality control measurements were performed using National Institute of Standard Technology standard reference materials. The present results show non-homogeneous distribution of Fe and Zn in normal human brain. Our second goal was to study the possible elemental concentration changes in German patients with Alzheimer disease (5 subjects, 10 brain regions). Fe and Zn values are found to be significantly changed in some AD brain regions compared to the controls. Another object of this work was to extend the method for the determination of elemental concentration not only in whole brain samples (high fat content) but - applying two types of solvent extraction - in lipid fraction and in brain tissue without lipid.

Niveles séricos de minerales y oligoelementos en recién nacidos sanos menores de un día / Seric levels of minerals and trace elements in healtly newborn of one day

Con el propósito de determinar valores séricos normales de algunos minerales y oligoelementos en recién nacidos durante el primer día de edad y de esta manera disponer de valores de referencia, se tomó muestra de sangre arterial a 55 niños menores de 25 horas de edad con examen. Después de separación inmediata del suero y su posterior congelación en hielo seco, las muestras fueron trasladadas a Alemania donde se determinaron Mg,Cu,S,P y Ca por espectroscopía de emisión atómica por plasma (ICP-AES); Cs,Co,Fe,Rb,Sc,Se, y Zn por análisis instrumental por activación de neutrones (INAA) y K,Na,Cl y proteínas totales por fotometría. Las concentraciones promedio (peso húmedo) fueron: Mg(19,9 ñ 2,6mg/L), Cu(0,41 ñ 0,14mg/L), S(901 ñ 76mg/L), P(110,3 ñ 12,5mg/L), Ca(96,9 ñ 7,6mg/L), Co(<0,56ug/L), FE(1,66 ñ 0,67mg/L), Sc(0,14 ñ 0,04ug/L), Se(0,13 ñ 0,03mg/L), ZN(0,79 ñ 0,18mg/L), K(5,1 ñ 0,5meq/L), Na(147,0 ñ5,9meq/L), y Cl(111,5 ñ 5,3meq/L). En el caso de Na,Mg y Rb se nota una significante variación dependiendo de las horas de edad del recién nacido. Encontramos en el suero correlaciones entre Fe y Cs; Cuy Ca; Rb y K; Se y S; Zn con Ca, K y S; Mg y Cl; Na y P; Ca con Cl, P y S. Por otra parte Ca,Cu,P y se correlacionaron con las proteínas totales del suero. El único oligoelemento que correlacionó con todos los parámetros hematológicos fue el Cu. El valor sérico del selenio en el recién nacido corresponde aprox. al 50 por ciento del valor de los adultos de la región.