Maria Elisa Soares

Effect of surface coating on the biodistribution profile of gold nanoparticles in the rat

Successful application of gold nanoparticles (AuNPs) in biomedicine requires extensive safety assessment for which biokinetic studies are crucial. We evaluated the biodistribution of AuNPs (∼20 nm) with different surface coatings: citrate, 11-MUA and 3 pentapeptides, CALNN, CALND and CALNS, after i.v. administration to rats (0.6-1 mg Au/kg). Biodistribution was evaluated based on Au tissue content measured by GFAAS. Citrate-AuNPs were rapidly removed from circulation with 60% of the injected dose depositing in the liver. Thirty minutes post-injection, the lungs presented about 6% of the injected dose with levels decreasing to 0.7% at 24 h. Gold levels in the spleen were of 2.6%. After 24 h, liver presented the highest Au level, followed by spleen and blood. A similar biodistribution profile was observed for MUA-coated AuNPs compared to Cit-AuNPs at 24h post-injection, while significantly higher levels of peptide-capped AuNPs were found in the liver (74-86%) accompanied by a corresponding decrease in blood levels. TEM analysis of liver slices showed AuNPs in Kupffer cells and hepatocytes, trapped inside endosomes. Our data demonstrate that AuNPs are rapidly distributed and that the liver is the preferential accumulation organ. Peptide capping significantly increased hepatic uptake, showing the influence of AuNPs functionalization in biodistribution.

Influence of the surface coating on the cytotoxicity, genotoxicity and uptake of gold nanoparticles in human HepG2 cells

The toxicological profile of gold nanoparticles (AuNPs) remains controversial. Significant efforts to develop surface coatings to improve biocompatibility have been carried out. In vivo biodistribution studies have shown that the liver is a target for AuNPs accumulation. Therefore, we investigated the effects induced by ~20 nm spherical AuNPs (0–200 μM Au) with two surface coatings, citrate (Cit) compared with 11‐mercaptoundecanoic acid (11‐MUA), in human liver HepG2 cells. Cytotoxicity was evaluated using the 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) reduction and lactate dehydrogenase (LDH) release assays after 24 to 72 h of incubation. DNA damage was assessed by the comet assay, 24 h after incubation with the capped AuNPs. Uptake and subcellular distribution of the tested AuNPs was evaluated by quantifying the gold intracellular content by graphite furnace atomic absorption spectrometry (GFAAS) and transmission electron microscopy (TEM), respectively. The obtained results indicate that both differently coated AuNPs did not induce significant cytotoxicity. An inverse concentration‐dependent increase in comet tail intensity and tail moment was observed in Cit‐AuNPs‐ but not in MUA‐AuNPs‐exposed cells. Both AuNPs were internalized in a concentration‐dependent manner. However, no differences were found in the extent of the internalization between the two types of NPs. Electron‐dense deposits of agglomerates of Cit‐ and MUA‐AuNPs were observed either inside endosomes or in the intercellular spaces. In spite of the absence of cytotoxicity, DNA damage was observed after exposure to the lower concentrations of Cit‐ but not to MUA‐AuNPs. Thus, our data supports the importance of the surface properties to increase the biocompatibility and safety of AuNPs. Copyright

Short- and long-term distribution and toxicity of gold nanoparticles in the rat after a single-dose intravenous administration

UNLABELLED Surface chemistry plays an important role in gold nanoparticles (AuNPs) stability and biocompatibility, which are crucial for their implementation into the clinical setting. We evaluated short- (30 min) and long-term (28 days) biodistribution and toxicity of ~20 nm citrate- and pentapeptide CALNN-coated AuNPs after a single intravenous injection in rats. The pattern of AuNPs distribution in Cit- and CALNN-AuNPs-injected rats was very similar in the assessed time-points. Both AuNPs were quickly removed from the bloodstream and preferentially accumulated in the liver. At 28 days liver remained the main accumulation site but at significantly lower levels compared to those found at 30 min. Spleen atrophy and hematological findings compatible with mild anemia were observed in CALNN-AuNPs-administered rats. Under our experimental conditions, surface coating had more impact on toxicity rather than on biodistribution of the AuNPs. Improvements in the design of capping peptides need to be done to increase biomedical applicability of peptide-coated AuNPs. FROM THE CLINICAL EDITOR The biodistribution and toxicity of ~ 20 nm citrate- and pentapeptide CALNN-coated gold nanoparticles was investigated after a single intravenous injection in rats. Rapid clearance and hepatic accumulation was found at 30-minutes, whereas mild anemia and spleen atrophy was seen 28 days post injection. The authors also concluded that the toxicity was related to the capping proteins as opposed to the biodistribution of the particles, providing important suggestion for future design of gold nanoparticles.

Chronic exposure to ethanol exacerbates MDMA-induced hyperthermia and exposes liver to severe MDMA-induced toxicity in CD1 mice

3,4-Methylenedioxymethamphetamine (MDMA; ecstasy) is an amphetamine derivative drug with entactogenic, empathogenic and hallucinogenic properties, commonly consumed at rave parties in a polydrug abuse pattern, especially with cannabis, tobacco and ethanol. Since both MDMA and ethanol may cause deleterious effects to the liver, the evaluation of their putative hepatotoxic interaction is of great interest, especially considering that most of the MDMA users are regular ethanol consumers. Thus, the aim of the present study was to evaluate, in vivo, the acute hepatotoxic effects of MDMA (10mg/kg i.p.) in CD-1 mice previously exposed to 12% ethanol as drinking fluid (for 8 weeks). Body temperature was continuously measured for 12h after MDMA administration and, after 24h, hepatic damage was evaluated. The administration of MDMA to non pre-treated mice resulted in sustained hyperthermia, which was significantly increased in ethanol pre-exposed mice. A correspondent higher increase of hepatic heat shock transcription factor (HSF-1) activation was also observed in the latter group. Furthermore, MDMA administration resulted in liver damage as confirmed by histological analysis, slight decrease in liver weight and increased plasma transaminases levels. These hepatotoxic effects were also exacerbated when mice were pre-treated with ethanol. The activities of some antioxidant enzymes (such as SOD, GPx and Catalase) were modified by ethanol, MDMA and their joint action. The hepatotoxicity resulting from the simultaneous exposure to MDMA and ethanol was associated with a higher activation of NF-kappaB, indicating a pro-inflammatory effect in this organ. In conclusion, the obtained results strongly suggest that the consumption of ethanol increases the hyperthermic and hepatotoxic effects associated with MDMA abuse.

Chromium speciation analysis in bread samples.

Chromium is a controversial element with important essentiality and toxicity, depending on its different species; its speciation analysis in principal human foodstuffs, as in the case of bread, is of utmost importance. With this purpose, a method was validated, including a wet acid digestion procedure for total chromium dissolution, a selective alkaline extraction of hexavalent chromium, and ETAAS determination. The method was applied to the determination of total and hexavalent chromium in 152 bread samples. The total chromium contents were 47.3 +/- 20.0 and 50.9 +/- 22.2 microg/kg of dry weight for white and whole bread samples, respectively; those for hexavalent chromium were 5.65 +/- 5.44 and 6.82 +/- 4.88 microg/kg of dry weight. On the basis of a mean daily ingestion of three bread units, the calculated daily intake was up to 12.7 microg/day for total chromium and 1.98 microg/day for hexavalent chromium. Referring to total chromium, bread can contribute up to 10% of the Reference Daily Intake, 120 microg/day.

Comparative Study of Different Extractive Procedures to Quantify Morphine in Urine by HPLC-UV

Abstract A comparative study to extract and purify total morphine in urine is described using several procedure treatments of biological samples. Sep-Pak C18 cartridges, Extrelut columns and liquid-liquid conventional extraction with sequencial purification through Extrelut column were assayed in order to establish the best procedure to quantitatively extract morphine from urine free of endogenous interferents. Analysis of the drug was made by HPLC-UV with a C18 column and methanol: acetate buffer (pH6.9) (70+30) as eluent. Recovery of morphine from spiked urine was 88.0% and 79.8% for concentrations of 1μg/ml and 5μg/ml, respectively. The detection limit of the method was 0.2μg/ml and coeficient of variation was 1.8% (n+4).

Mercury fatal intoxication: two case reports.

We report two cases of fatal intoxications with mercury, one intentional and the other allegedly resulting from a drug formulation mistake. Both cases occurred in the year of 2004. The first case refers to a man who ingested a great portion of a mercuric chloride solution. He attended a hospital emergency, submitted to treatment, but died after 49 days. In the second case, a woman applied on the chest skin an ointment containing a great quantity of mercury bromide. After 7 days of treatment in the hospital, she died. In both cases, samples of tissues and organs were collected at autopsy for mercury analysis. Because methylation of mercury in humans after exposure to metallic or inorganic mercury is almost unknown, both total mercury and methylmercury were quantified in the post-mortem samples. The quantifications were carried out by Cold Vapour Generation Atomic Absorption Spectrometry for total mercury and by HPLC-UV for methylmercury. The total mercury contents found in the post-mortem fluid and tissue samples were consentaneous with mercury poisoning. For the first case, the concentrations found, expressed in microg/g wet weight, were in the liver 49.9, lung 3.27 and brain 0.33, and for blood 11.7 microg/mL. For the second case, the concentrations expressed in microg/g wet weight were in the liver 46.6, lung 14.6, brain 0.21, kidney 77.7, stomach 7.12, spleen 6.4 and heart 2.34, and for blood and urine 2.95 and 1.40 microg/mL, respectively. Only in the first case was methylmercury found and quantified in liver (1.70 microg/g wet weight) and in blood (0.15 microg/mL) samples.

Determination of arsenic, cadmium and lead in porcine and bovine kidneys by electrothermal atomic absorption spectrometry

This paper describes an ETAAS method to quantify residues of arsenic, cadmium and lead in porcine and bovine kidneys. The results of a survey conducted during 1993 are tabulated. Analysis was performed by ETAAS with a stabilized-temperature platform furnace. The determinations were performed in the linear ranges 0.5-150, 0.04-2.0 and 1.1-75 micrograms l-1 for As, Cd and Pb, respectively, in the acid digests which were obtained under controlled conditions of temperature in a tetrafluoroethylene apparatus after appropriate dilution and addition of a suitable chemical modifier. Extensive quality assurance of the methods was performed by the standard additions method and by comparison with a certified reference material. The precision was better than 9.9, 8.5 and 9.2% for As, Cd and Pb, respectively. Low relative standard deviations of 7.4, 7.0, and 2.8% for As, Cd and Pb, respectively, were obtained by comparing the levels found in the kidney reference material and the certified values, showing this method to be satisfactory and suitable for routine analysis. The mean levels of As, Cd and Pb in porcine kidney were 2.11 (1.11-5.49), 0.81 (0.02-4.22) and 0.18 (0.02-0.42) microgram g-1 respectively. Those in the adult bovine kidney were 1.77 (0.42-3.42), 1.26 (0.01-6.16) and 0.73 (0.19-2.55) microgram g-1, respectively, and in the young bovine kidney they were 1.33 (0.61-2.56), 0.49 (0.15-1.82) and 0.20 (0.12-0.29) microgram g-1.

Inorganic mercury intoxication: A case report

Mercury is a heavy metal with unique physico-chemical properties, and it is well distributed throughout the environment, being present in soil, water and air. This non-essential element is considered by the World Health Organization (WHO) as one of the ten most troublesome chemical to public health. Its toxicity spectrum depends on the chemical form in which it presents: elemental (metallic), organic or inorganic. The known intoxications are mainly occupational (mining, agriculture, incineration) or related to the use of dental amalgams or the consumption of contaminated fish and shellfish. Nowadays, acute exposures to toxic amounts of mercury are increasingly rare, especially those involving inorganic mercury compounds. The rate is even lower if we refer to intentional poisonings. Although there is a growing understanding of the toxicokinetics of mercury, there is still a lack of studies that support the emerging theories about its bioavailability in humans. In this manuscript we describe a rare case of an individual who committed suicide by ingesting mercuric oxide. The aim is to offer a medical contribution to the better understanding of the kinetics of this metal, making a discussion based on published literature and analyzing its distribution, metabolism, internal doses, target and reservoir organs. The whole case - clinical course of the victim and her fatal destiny, the ante- and post-mortem sample concentrations and the necropsy findings - illustrates a situation that meets specific features of acute poisoning by ingestion of inorganic mercury, thus constituting an important support towards a more realistic and a based on evidence understanding of mercury biodistribution in humans.

Chromium speciation and biochemical changes vary in relation to plant ploidy

Uptake of trivalent chromium (Cr(III)-chloride), Cr speciation and consequences for the metabolism in chamomile plants with two ploidy levels have been studied. Depletion of fresh biomass, tissue water content and soluble proteins in response to high (120 μM) Cr(III) was ploidy-independent. Cr mainly accumulated in the roots (only negligibly in the shoots) and total root Cr amount was higher in tetraploid ones including the proof with specific fluorescent indicator (naphthalimide-rhodamine) of Cr(III). Quantification of Cr(VI) detected its higher content in tetraploid roots (up to 4.2% from total Cr), indicating partial oxidation of applied Cr(III). Higher H2O2 presence but lower activities of peroxidases were observed in tetraploid roots while nitric oxide, superoxide dismutase and glutathione reductase activities did not differ extensively. Soluble phenols, lignin, non-protein thiols, individual thiols (glutathione and phytochelatin 2) and ascorbic acid responded to high Cr(III) similarly in both cultivars while decrease of minerals was more pronounced in tetraploid ones. It seems that Cr(III)-induced oxidative stress arises from high root Cr uptake and Cr(VI) presence and is related to depletion of thiols. Assay of Krebs cycle acids confirmed rather depletion under 120 μM Cr(III) in both cultivars but increase in citric acid may indicate its involvement in root Cr chelation. Subsequent comparison of Cr(III)-chloride and Cr(III)-nitrate showed similar influence on Cr accumulation and majority of biochemical responses while different impact on phytochelatin 2 amount was the most distinct feature.