Use of total reflection X-ray fluorescence (TXRF) for the evaluation of heavy metal poisoning due to the improper use of a traditional ayurvedic drug

An Indian patient referred to Clinica del Lavoro ‘L.Devoto’ of Milano showed clinical signs of heavy metal poisoning, possibly related to a sustained 6-month use of approx. 3 g/day of a traditional preparation (a whitish powder with a ‘mineral’ appearance) to treat urological problems. To confirm the causal relationship between the disease and the use of such product, metal testing was performed on the patient's hair and the ayurvedic remedy samples by total reflection X-ray fluorescence (TXRF). For TXRF analysis 1-cm cut of the patient's hair was directly deposited onto the quartz glass sample carrier, then 10 μl of nitric acid 65% were added and dried in air. TXRF showed high versatility, rapid and simultaneous element detection, and short analysis time, thus supporting a wider use in emergency medicine and in forensic analyses.     

Quantitative histochemistry for macrophage biodistribution on mice liver and spleen after the administration of a pharmacological-relevant dose of polyacrylic acid-coated iron oxide nanoparticles

Understanding in vivo biodistribution of iron oxide nanoparticles (IONs), and the involvement of the phagocyte system in this process, is crucial for the assessment of their potential health risk. In the present study, the histochemical expression of iron in liver and spleen sections of CD-1 mice (aged 8 weeks) was quantified, 24?h after intravenous administration of polyacrylic acid-coated IONs (PAA-coated IONs) (8, 20, 50?mg/kg). Organ sections were stained with Perls’ Prussian blue for iron detection, followed by the quantification of iron deposition with ImageJ software. Our study revealed the existence of a linear dose-dependent increase of iron deposition in macrophages of both organs. Exposed animals showed hepatic iron deposition in all zones, although most marked in periportal region. In the spleen, no iron was detected in the white splenic pulp of both control and treated animals. When compared with control mice, a positive correlation between histochemical detection of iron and PAA-coated ION doses was observed in splenic red pulp of animals. The results confirmed our assumption that liver and spleen are involved in the clearance pathways of PAA-coated IONs from the blood. Excess iron was cytotoxic at the highest dose of PAA-coated IONs tested, but no significant morphologic alterations were observed for the lower doses. Clusters of early necrotic hepatocytes were observed in the hepatic periportal region of mice injected with the higher dose (50?mg/kg) of PAA-coated IONs. Further studies are necessary to determine if liver and spleen macrophages will degrade these IONs, or eventually exocytose both the degraded and intact ones.     

Lipidomic changes in rat liver after long-term exposure to ethanol

Alcoholic liver disease (ALD) is a serious health problem with significant morbidity and mortality. In this study we examined the progression of ALD along with lipidomic changes in rats fed ethanol for 2 and 3 months to understand the mechanism, and identify possible biomarkers. Male Fischer 344 rats were fed 5% ethanol or caloric equivalent of maltose-dextrin in a Lieber-DeCarli diet. Animals were killed at the end of 2 and 3 months and plasma and livers were collected. Portions of the liver were fixed for histological and immunohistological studies. Plasma and the liver lipids were extracted and analyzed by nuclear magnetic resonance (NMR) spectroscopy. A time dependent fatty infiltration was observed in the livers of ethanol-fed rats. Mild inflammation and oxidative stress were observed in some ethanol-fed rats at 3 months. The multivariate and principal component analysis of proton and phosphorus NMR spectroscopy data of extracted lipids from the plasma and livers showed segregation of ethanol-fed groups from the pair-fed controls. Significant hepatic lipids that were increased by ethanol exposure included fatty acids and triglycerides, whereas phosphatidylcholine (PC) decreased. However, both free fatty acids and PC decreased in the plasma. In liver lipids unsaturation of fatty acyl chains increased, contrary to plasma, where it decreased. Our studies confirm that over-accumulation of lipids in ethanol-induced liver steatosis accompanied by mild inflammation on long duration of ethanol exposure. Identified metabolic profile using NMR lipidomics could be further explored to establish biomarker signatures representing the etiopathogenesis, progression and/or severity of ALD.     

Acute toxicity of zinc oxide nanoparticles to the rat olfactory system after intranasal instillation

With the increased applications of zinc oxide (ZnO) nanoparticles (NPs), the toxicity of ZnO NPs arouses great concerns from the nano community and the general public. In this study, we report the toxicity of ZnO NPs (30 nm) to the rat olfactory system after intranasal instillation revealed by non‐invasive magnetic resonance imaging (MRI). MRI scans were performed on a 4.7‐T scanner at 1, 2, 3 and 7 days post‐exposure, and the histological changes of the rat olfactory epithelium were evaluated. The influences of chemical component and dispersant of the NPs were also investigated. In addition, an olfactory behavior test was performed. The MRI and histological results indicated that ZnO NPs induced significant damages to the olfactory epithelium, including disruption of the olfactory epithelial structures and inflammation. The destruction of mitochondria in epithelial cells was observed under transmission electron microscopy (TEM), suggesting that the possible toxicological mechanism might involve cellular energy metabolic dysfunction. Further, the lesion of the olfactory epithelium disturbed sniffing behaviors of the treated animals. The results suggest that MRI is potentially useful as a screening tool to assess the consequence of occupational exposure of ZnO NPs. Caution should therefore be taken during the use and disposal of ZnO NPs to prevent the unintended public health impacts. Copyright © 2012 John Wiley & Sons, Ltd.     

Effect of surface coating on the biocompatibility and in vivo MRI detection of iron oxide nanoparticles after intrapulmonary administration

Abstract

Superparamagnetic iron oxide nanoparticles (SPIONs) have attracted special attention as novel nanoprobes capable of improving both the therapy and diagnosis of lung diseases. For safe prospective clinical applications, their biocompatibility has to be assessed after intrapulmonary administration. This study was therefore conducted to understand the biological impact of SPIONs and their further surface-functionalization with polyethylene glycol (PEG) having either negative (i.e. carboxyl) or positive (i.e. amine) terminal in a 1-month longitudinal study following acute and sub-acute exposures. Noninvasive free-breathing MR imaging protocols were first optimized to validate SPIONs detection in the lung and investigate possible subsequent systemic translocation to abdominal organs. Pulmonary Magnetic Resonance Imaging (MRI) allowed successful in vivo detection of SPIONs in the lung using ultra-short echo time sequence. Following high-dose lung administration, MR imaging performed on abdominal organs detected transient accumulation of SPIONs in the liver. Iron quantification using Inductive coupled plasma – Mass mass spectroscopy (ICP-MS) confirmed MRI readouts. Oxidative stress induction and genotoxicity were then conducted to evaluate the biocompatibility of SPIONs with their different formulations in a mouse model. A significant increase in lipid peroxidation was observed in both acute and sub-acute sets and found to regress in a time-dependent manner. PEG functionalized SPIONs revealed a lower effect with no difference between both terminal modifications. Genotoxicity assessments revealed an increase in DNA damage and gene expression of CCL-17 and IL-10 biomarkers following SPIONs administration, which was significantly higher than surface-modified nanoparticles and decreased in a time-dependent manner. However, SPIONs with carboxyl terminal showed a slightly prominent effect compared to amine modification.     

Aluminium oxide nanoparticles induced morphological changes,cytotoxicity and oxidative stress in Chinook salmon (CHSE‐214) cells

Aluminium oxide nanoparticles (Al₂O₃ NPs) are increasingly used in diverse applications that has raised concern about their safety. Recent studies suggested that Al₂O₃ NPs induced oxidative stress may be the cause of toxicity in algae, Ceriodaphnia dubia, Caenorhabditis elegans and Danio rerio. However, there is paucity on the toxicity of Al₂O₃ NPs on fish cell lines. The current study was aimed to investigate Al₂O₃ NPs induced cytotoxicity, oxidative stress and morphological abnormality of Chinnok salmon cells (CHSE‐214). A dose‐dependent decline in cell viability was observed in CHSE‐214 cells exposed to Al₂O₃ NPs. Oxidative stress induced by Al₂O₃ NPs in CHSE‐214 cells has resulted in the significant reduction of superoxide dismutase, catalase and glutathione in a dose‐dependent manner. However, a significant increase in glutathione sulfo‐transferase and lipid peroxidation was observed in CHSE‐214 cells exposed to Al₂O₃ NPs in a dose‐dependent manner. Significant morphological changes in CHSE‐214 cells were observed when exposed to Al₂O₃ NPs at 6, 12 and 24 h. The cells started to detach and appear spherical at 6 h followed by loss of cellular contents resulting in the shrinking of the cells. At 24 h, the cells started to disintegrate and resulted in cell death. Our data demonstrate that Al₂O₃ NPs induce cytotoxicity and oxidative stress in a dose‐dependent manner in CHSE‐214 cells. Thus, our current work may serve as a base‐line study for future evaluation of toxicity studies using CHSE‐214 cells. Copyright © 2015 John Wiley & Sons, Ltd.     

The cortical epithelium of the rat thymus after in vivo exposure to bis(tri-n-butyltin)oxide (TBTO)

Bis(tri-n-butyltin)oxide (TBTO) induces cortical atrophy in the rat thymus. We studied the potential involvement of the cortical epithelium in TBTO-induced thymotoxicity by (immuno) histology and electron microscopy. Juvenile male Wistar rats were orally intubated once with either 30 or 90 mg/kg TBTO and sacrificed 4 or 10 days later. A dose-dependent thymic atrophy occurred. Anti-keratin labelling showed epithelial cell aggregation in some animals at 10 days after exposure to 90 mg/kg TBTO, when recovery of the thymus was apparent. At the ultrastructural level, a relative shift was observed from “pale” to darker epithelial cell subtypes at the 30 mg/kg dose level, both at day 4 and day 10 after intubation. This phenomenon was not observed after exposure to 90 mg/kg TBTO. Both the altered keratin distribution and the increased electron density of the epithelium probably represent non-specific phenomena. The present morphological observations support the concept that TBTO affects the thymus via its action on lymphoid cells rather than on the epithelial compartment.     

Nitric oxide induced sinusoidal relaxation after a propranolol priming dose in the perfused rat liver reduces propranolol availability on subsequent dosing

1. The present study sought to explain the mechanism leading to reduced availability of propranolol when given after a priming dose in the single‐pass perfused rat liver. 2. Extracellular sucrose space (as a measure of sinusoidal relaxation) in perfused rat liver before and after propranolol or propranolol and N^G‐nitro‐l ‐arginine methyl ester (l ‐NAME; nitric oxide (NO) synthase inhibitor) treatment were examined. The results showed that propranolol induces sinusoidal relaxation in the perfused liver and this effect could be abolished by NO synthase inhibitor l ‐NAME. 3. Two bolus injections of propranolol were given to the isolated perfused rat liver and outflow concentration‐time profiles of intact propranolol were determined. A two‐phase physiologically based organ pharmacokinetic model was applied to estimate hepatocellular influx, efflux, binding, ion‐trapping and metabolic elimination pharmacokinetic parameters for propranolol. The recovery of propranolol in the second injection was approximately 54% of that in the first injection. The permeability‐surface area product, the binding and the intrinsic clearance all increased significantly after prior exposure of the rat liver to the first bolus of propranolol (P < 0.05). 4. Based on the findings of the present study, we propose that the most likely explanation for the reduced availability of a second propranolol dose (after administration of a priming dose) in the perfused liver is a consequence of the NO‐mediated sinusoidal relaxation effect of propranolol, arising from the priming dose. This observation supports the view that the pharmacokinetics of some drugs might be altered by the pharmacodynamic effects of the same drug given earlier in the perfused liver.     

The toxicity and distribution of iron oxide–zinc oxide core‐shell nanoparticles in C57BL/6 mice after repeated subcutaneous administration

Therapeutic cancer vaccines promote immune responses by delivering tumour‐specific antigens. Recently, we developed iron oxide (Fe₃O₄)–zinc oxide (ZnO) core‐shell nanoparticles (CSNPs) as carriers for antigen delivery into dendritic cells (DCs), and the CSNPs were injected subcutaneously into C57BL/6 mice to examine the systemic toxicity, tissue distribution and excretion of the CSNPs. The doses injected were 0, 4, 20 and 200 mg kg^–1 weekly for 4 weeks. No significant changes were observed after the CSNPs administration with respect to mortality, clinical observations, body weight, food intake, water consumption, urinalysis, haematology, serum biochemistry,and organ weights. A dose‐dependent increase in granulomatous inflammation was observed at the injection site of the CSNP‐treated animals, but no other histopathological lesions in other organs could be attributed to the CSNPs. The Zn concentration, which is an indicator for CSNPs, was not significantly higher in the sampled tissues, urine, or faeces after the CSNP injection. In contrast, the Zn concentration at the subcutaneous skin of the site injected with the CSNPs increased in a dose‐dependent manner, along with a macroscopic deposition of the CSNPs. The CSNP residue at the injection site resulted in a foreign body response with the appearance of macrophage infiltration, but otherwise did not show any systemic distribution or toxicity at up to 200 mg kg^–1 during this study. In conclusion, CSNPs could be used as good antigen carriers for DC‐based immunotherapy, although further study is needed to completely clear the residue of the CSNPs at the injection site. Copyright © 2015 John Wiley & Sons, Ltd.     

Investigating the toxic effects induced by iron oxide nanoparticles on neuroblastoma cell line: an integrative study combining cytotoxic,genotoxic and proteomic tools

Abstract

Nanomaterials have gained much attention for their use and benefit in several fields. Iron Oxide Nanoparticles (IONPs) have been used in Biomedicine as contrast agents for imaging cancer cells. However, several studies reported the potential toxicity of those nanoparticles in different models, especially in cells. Therefore, in our present study, we investigated the effects of IONPs on the SH-SY5Y neuroblastoma cell line. We carried out cytotoxic and genotoxic studies to evaluate the phenotypic effects, and proteomic investigation to evaluate the molecular effects and the mechanisms by which this kind of NPs could induce toxicity. Our results showed that the use of three different sizes of IONPs (14, 22 and 30?nm) induced cell detachment, cell morphological changes, size, and concentration-dependent IONP internalization and cell mortality. IONPs induced slight genotoxic damage assayed by modified comet assay without affecting cell cycle, mitochondrial function, membrane integrity, intracellular calcium level, and without inducing ROS generation. All the studies were performed to compare also the effects of IONPs to the ferric iron by incubating cells with equivalent concentration of FeCl₃. In all tests, the NPs exhibited more toxicity than the ferric iron. The proteomic analysis followed by gene ontology and pathway analysis evidenced the effects of IONPs on cytoskeleton, cell apoptosis, and cancer development. Our findings provided more information about IONP effects on human cells and especially on cancer cell line.     

The inhibitory effect of superparamagnetic iron oxide nanoparticle (Ferucarbotran) on osteogenic differentiation and its signaling mechanism in human mesenchymal stem cells

Superparamagnetic iron oxide (SPIO) nanoparticles are very useful for monitoring cell trafficking in vivo and distinguish whether cellular regeneration originated from an exogenous cell source, which is a key issue for developing successful stem cell therapies. However, the impact of SPIO labeling on stem cell behavior remains uncertain. Here, we show the inhibitory effect of Ferucarbotran, an ionic SPIO, on osteogenic differentiation and its signaling mechanism in human mesenchymal stem cells. Ferucarbotran caused a dose-dependent inhibition of osteogenic differentiation, abolished the differentiation at high concentration, promoted cell migration, and activated the signaling molecules, β-catenin, a cancer/testis antigen, SSX, and matrix metalloproteinase 2 (MMP2). An iron chelator, desferrioxamine, suppressed all the above Ferucarbotran-induced actions, demonstrating an important role of free iron in the inhibition of osteogenic differentiation that is mediated by the promotion of cell mobilization, involving the activation of a specific signaling pathway.     

Histological alterations in gills and liver of rainbow trout (Oncorhynchus mykiss) after exposure to the antibiotic oxytetracycline

The aim of this study was to investigate the histopathological effects of oxytetracycline (OTC) on the gill and liver tissues of rainbow trouts (Oncorhynchus mykiss) following acute (96 h: 0.005–50 mg/L) and chronic (28 days: 0.3125–5 μg/L) exposures. Results suggest the existence of a cause-and-effect relationship between the exposure to OTC and tissue damage. Most predominant disorders observed in gills were progressive (e.g. hypertrophy of mucous cells and hyperplasia of epithelial cells) in acute exposure and regressive (e.g. lamellar fusion, epithelial lifting of lamellae and some changes in tissue architecture) in chronic exposure. However, only the acute exposure was responsible for a significant increase of the total gill pathological index. PAGE index, reflecting the extent of gill tissue available for gas exchanges in fish, remained unchanged for both exposures. In liver, circulatory (e.g. hemorrhage and increase of sinusoidal space), regressive (e.g. pyknotic nucleus, vacuolization and hepatocellular degenerations) and progressive (e.g. hypertrophy of hepatocytes) changes were observed, but just after acute exposure. After chronic exposure, only inflammatory changes (e.g. leucocytes infiltration) were observed. Following both exposures, a significant increase of the total liver pathological index was recorded. Despite the increase of the histological damage in individuals exposed to OTC, lesions observed were of minimal or moderate pathological importance, non-specific and reversible. The data gathered following acute and chronic exposures also suggest the onset of adaptive mechanisms of fish, namely for longer exposure periods. Furthermore the observed histological alterations appear to be result of several physio-metabolic disorders consequence of the biochemical and molecular modes of action of OTC.     

Pharmacokinetics and bio‐distribution of novel super paramagnetic iron oxide nanoparticles (SPIONs) in the anaesthetized pig

Manufactured nanomaterials have a variety of medical applications, including diagnosis and targeted treatment of cancer. A series of experiments were conducted to determine the pharmacokinetic, biodistribution and biocompatibility of two novel magnetic nanoparticles (MNPs) in the anaesthetized pig. Dimercaptosuccinic acid (DMSA) coated superparamagnetic iron oxide nanoparticles (MF66‐labelled 12 nm, core nominal diameter and OD15 15 nm); at 0.5, or 2.0 mg/kg) were injected intravenously. Particles induced a dose‐dependent decrease in blood pressure following administration which recovered to control levels several minutes after injection. Blood samples were collected for a 5‐h period and stored for determination of particle concentration using particle electron paramagnetic resonance (pEPR). Organs were harvested post‐mortem for magnetic resonance imaging (MRI at 1.5 T field strength) and histology. OD15 (2.0 mg/kg) MNP had a plasma half‐life of approximately 15 min. Both doses of the MF66 (0.5 and 2.0 mg/kg) MNP were below detection limits. MNP accumulation was observed primarily in the liver and spleen with MRI scans which was confirmed by histology. MRI also showed that both MNPs were present in the lungs. The results show that further modifications may be required to improve the biocompatibility of these particles for use as diagnostic and therapeutic agents.     

Effects of subchronic exposure to zinc nanoparticles on tissue accumulation,serum biochemistry,and histopathological changes in tilapia (Oreochromis niloticus)

Zinc nanoparticles (ZnNPs) are among the least investigated NPs and thus their toxicological effects are not known. In this study, tilapia (Oreochromis niloticus) were exposed to 1 and 10 mg/L suspensions of small size (SS, 40–60 nm) and large size (LS, 80–100 nm) ZnNPs for 14 days under semi‐static conditions. Total Zn levels in the intestine, liver, kidney, gill, muscle tissue, and brain were measured. Blood serum glucose (GLU), glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), and lactate dehydrogenase (LDH) were examined to elucidate the physiological disturbances induced by ZnNPs. Organ pathologies were examined for the gills, liver, and kidney to identify injuries associated with exposure. Significant accumulation was observed in the order of intestine, liver, kidney, and gills. Zn levels exhibited time‐ and concentration‐dependent increase in the organs. Accumulation in kidney was also dependent on particle size; NPs SS‐ZnNPs were trapped more effectively than LS‐ZnNPs. No significant accumulation occurred in the brain (p > 0.05) while Zn levels in muscle tissue increased only marginally (p ≥ 0.05). Significant disturbances were noted in serum GOT and LDH (p < 0.05). The GPT levels fluctuated and were not statistically different from those of controls (p > 0.05). Histopathological tubular deformations and mononuclear cell infiltrations were observed in kidney sections. In addition, an increase in melano‐macrophage aggregation intensity was identified on the 7th day in treatments exposed to LS‐ZnNPs. Mononuclear cell infiltrations were identified in liver sections for all treatments. Both ZnNPs caused basal hyperplasia in gill sections. Fusions appeared in the gills after the 7th day in fish treated with 10 mg/L suspensions of SS‐ZnNPs. In addition, separations in the secondary lamella epithelia were observed. The results indicated that exposure to ZnNPs could lead to disturbances in blood biochemistry and cause histopathological injuries in the tissues of O. niloticus. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1213–1225, 2017.     

Promoting neuroregeneration by applying dynamic magnetic fields to a novel nanomedicine: Superparamagnetic iron oxide (SPIO)-gold nanoparticles bounded with nerve growth factor (NGF)

Neuroregeneration imposes a significant challenge in neuroscience for treating neurodegenerative diseases. The objective of this study is to evaluate the hypothesis that the nerve growth factor (NGF) functionalized superparamagnetic iron oxide (SPIO)-gold (Au) nanomedicine can stimulate the neuron growth and differentiation under external magnetic fields (MFs), and dynamic MFs outperform their static counterparts. The SPIO-Au core-shell nanoparticles (NPs) (Diameter: 20.8 nm) possessed advantages such as uniform quasi-spherical shapes, narrow size distribution, excellent stabilities, and low toxicity (viability >96% for 5 days). NGF functionalization has enhanced the cellular uptake. The promotion of neuronal growth and orientation using NGF functionalized SPIO-Au NPs, driven by both the static and dynamic MFs, was revealed experimentally on PC-12 cells and theoretically on a cytoskeletal force model. More importantly, dynamic MFs via rotation performed better than the static ones, i.e., the cellular differentiation ratio increased 58%; the neurite length elongation increased 63%.     

Investigation into the pulmonary inflammopathology of exposure to nickel oxide nanoparticles in mice

We investigated the effects of nickel oxide nanoparticles (NiONPs) on the pulmonary inflammopathology. NiONPs were intratracheally installed into mice, and lung injury and inflammation were evaluated between 1 and 28 days. NiONPs caused significant increases in LDH, total protein, and IL-6 and a decrease in IL-10 in the BALF and increases in 8-OHdG and caspase-3 in lung tissues at 24 h. Airway inflammation was present in a dose-dependent manner from the upper to lower airways at 24 h of exposure as analyzed by SPECT. Lung parenchyma inflammation and small airway inflammation were observed by CT after NiONP exposure. 8-OHdG in lung tissues had increased with formation of fibrosis at 28 days. Focal adhesion was the most important pathways identified at 24 h as determined by protemics, whereas glutathione metabolism was the most important identified at 28 days. Our results demonstrated the pulmonary inflammopathology caused by NiONPs based on image-to-biochemical approaches.     

A comparative study on the in vitro cytotoxic responses of two mammalian cell types to fullerenes,carbon nanotubes and iron oxide nanoparticles

The present study was designed to evaluate and compare the time- and dose-dependent cellular response of human periodontal ligament fibroblasts (hPDLFs), and mouse dermal fibroblasts (mDFs) to three different types of nanoparticles (NPs); fullerenes (C₆₀), single walled carbon nanotubes (SWCNTs) and iron (II,III) oxide (Fe₃O₄) nanoparticles via in vitro toxicity methods, and impedance based biosensor system. NPs were characterized according to their morphology, structure, surface area, particle size distribution and zeta potential by using transmission electron microscopy, X-ray diffraction, Brunauer–Emmett–Teller, dynamic light scattering and zeta sizer analyses. The Mössbauer spectroscopy was used in order to magnetically characterize the Fe₃O₄ NPs. The hPDLFs and mDFs were exposed to different concentrations of the NPs (0.1, 1, 10, 50 and 100?μg/mL) for predetermined time intervals (6, 24 and 48?h) under controlled conditions. Subsequently, NP exposed cells were tested for viability, membrane leakage and generation of intracellular reactive oxygen species. Additional to in vitro cytotoxicity assays, the cellular responses to selected NPs were determined in real time using an impedance based biosensor system. Taken together, information obtained from all experiments suggests that toxicity of the selected NPs is cell type, concentration and time dependent.     

Intrinsic biological property of colloidal fullerene nanoparticles (nC60): Lack of lethality after high dose exposure to human epidermal and bacterial cells

Colloidal fullerene nanoparticles (nC₆₀) were reported to be toxic to fish brains, human cells and microorganisms, while new observations suggest that the observed toxicity may be due to tetrahydrofuran (THF) solvent or its oxidative by-products in nC₆₀ preparations. Here, we report a novel method for preparing nC₆₀ nanoparticles that does not use THF solvent, but provides nC₆₀ with an average particle size of 43.8 nm and a yield approximately 100 times higher than the THF method. The prepared nC₆₀ showed a similar antioxidant capacity compared to a water-soluble vitamin E analog. No mortality to human epidermal keratinocytes was observed at a concentration 170 times higher than the reported LC₅₀ values for other human cell lines. No toxicity was observed to E. coli or B. subtilis at up to 342 μg/mL nC₆₀ for 16 h, which was hundred times higher than the reported minimum inhibitory concentrations of nC₆₀ prepared using THF method for these two bacteria. When E. coli was exposed to 85.5 μg/mL nC₆₀ with daily passage for 4 days, the stationary phase populations at different passages were not statistically different (p = 0.05) from the control without nC₆₀ nanoparticles. These results reveal that the intrinsic biological property of nC60 is non-toxic, confirming the prior non-toxic reports when using nC₆₀ prepared with non-THF methods.     

PCB methyl sulphones in rat liver after exposure to PCB (Clophen A50): analysis and radiosynthesis of selected methylsulphonyl-PCBs

1. The hepatic metabolism of polychlorinated biphenyls (PCBs) and formation of PCB methyl sulphone metabolites (MeSO₂-PCBs) was determined in the male Sprague-Dawley rat 1, 2, 4 or 8 weeks after dosage with Clophen A50 (a commercial PCB mixture). 2. The total concentration of the PCB congeners examined (ΣPCB) decreased during the experimental period, from 40 μg g^?1 lipid (l.w.) after 1 week to 4 μg g^?1 l.w. after 8 weeks. A 50% decrease of PCB in the liver were estimated to be 28, 13 and 11 days for 2,2′,4,4′,5,5′-hexachlorobiphenyl (CB153), 2,2′,3,3′,4,4′,5-heptaCB (CB170) and 2,2′3,4′,5,5′,6-heptaCB (CB187), respectively. 3. The total MeSO₂-PCB (ΣMeSO₂-PCB) concentration increased from 800 to 1020 ng g^?1 l.w. during the first 2 weeks of treatment and thereafter a decrease to 120 ng g^?1 l.w. after 8 weeks. The relative concentration of both 3′-MeSO₂-2,2′,4,4′,5-pentaCB (3′-MeSO₂-CB101) and 3′-MeSO₂-2,2′,3,4,5′-pentaCB (3′-MeSO₂-CB87) in rat liver showed significant increases during the 8 weeks. In contrast, the relative concentrations of 4-MeSO₂-2,4′,5,5-tetraCB (4-MeSO₂-CB64), 3-MeSO₂-2,3′,4′,5-tetraCB (3-MeSO₂- CB70) and 4-MeSO₂-2,2′,3,4,5′,6′-hexaCB (4′-MeSO₂-CB132) decreased significantly. 4. A route for the synthesis of radiolabelled MeSO₂-PCBs was developed employing the `Pummerer reaction’ to convert methylthio-PCBs (MeS-PCBs) to the corresponding mercapto-PCBs (SH-PCB). The SH-PCBs were methylated with radiolabelled methyl iodide and the resulting sulphides oxidized to yield the corresponding MeSO₂-PCBs. Using this approach, 4-[¹⁴C]-MeSO₂-2′,4′,5,5′,6-hexaCB (3-[¹⁴C]-MeSO₂-CB149), 4-[¹⁴C]-MeSO₂-2,2′,4′,5,5′,6-hexaCB (4-[¹⁴C]-MeSO₂-CB149), 3′-[¹⁴C]-MeSO₂-CB101,4′-[¹⁴C]-MeSO₂-2,2′,4,5,5′-pentaCB (4′-[¹⁴C]-MeSO₂-CB101) and 4′-[³C]-MeSO₂-CB101 were synthesized in quantitative radiochemical yields.