Investigation of metal-binding metallothioneins in the tissues of rats after oral intake of cinnabar

The multi-metal-binding MT fractions in rat tissues after oral intake of cinnabar were characterized by hyphenated size-exclusion chromatography (SEC) and inductively coupled plasma mass spectrometry (ICP–MS). With the increase of both the feeding time and dosage of cinnabar the amounts of Hg-binding MT fractions in rat kidney of groups fed cinnabar increased significantly compared with the control group. Meanwhile, more Cu-binding MT were synthesized in the rat kidney, which confirmed the manipulating effect of MT in the homeostasis of Cu for detoxification of nephritic mercury. Although the Hg-binding MT fractions in rat liver of all cinnabar groups were almost independent of cinnabar dosage (2.5–5.0 g kg^–1 bw) and feeding time (2–4 weeks), the amount was much higher than that of the control group. The amounts of Hg-MT in other rat tissues of the cinnabar groups and the control group were compared and their significance is discussed.     

Investigation of mercury metallothionein complexes in tissues of rat after oral intake of HgCl2

Mercury, a highly toxic metal found widely throughout the environment, is a potent inducer of metallothionein (MT) expression. The role of MTs in the detoxification of mercury after its oral intake in mammals is studied. After feeding rats with mercuric chloride by gastric gavage, the distribution of heavy metals in rat tissues was investigated by inductively coupled plasma mass spectrometry (ICP‐MS). Extensive accumulation of mercury, copper and zinc in kidney and liver is observed. A homemade preparative size‐exclusion chromatography (SEC) column (30 cm × 1.9 cm) packed with Sephadex G‐75 (40–120 µm particle size) gel (Pharmacia) was used for the purification of MT fractions in rat tissues. Preliminary results from SEC indicate that the mercury‐binding MT levels in liver were much lower than in kidney. The MT fractions were collected, desalted, and then separated by reversed‐phase high‐performance liquid chromatography (HPLC) with UV–Vis spectrometry, ICP‐MS and electrospray ionization MS detection. One major and several minor peaks were observed in the HPLC chromatograms of the MT fraction for the kidney sample. UV absorption spectra indicate that MTs were found to bind with mercury. There were no significant mercury‐binding MTs detected in the liver sample using UV detection. ICP‐MS detection showed that mercury‐binding MTs in kidney contained large amounts of mercury and copper but little zinc. Further characterization with ESI‐MS showed that the major peak found in kidney contained Hg₆Cu, Hg₅Cu₂‐MT‐2c and Hg₆‐MT‐2β, Hg₆Cu‐MT‐1γ, Hg₇‐MT‐2α. However, distinction between copper and zinc could not be made based on current mass spectrometric analysis because of instrumental resolution limitations. Copyright © 2005 John Wiley & Sons, Ltd.     

Effects of aqueous extracts of Acacia albida stem bark on Wistar albino rats infected with Trypanosoma evansi

The effect of aqueous extract of Acacia albida stem bark was investigated in Wistar albino rats infected with Trypanosoma evansi. The extract showed highest reduction in parasitemia at the dose of 600 mg/kg body weight (bw). A dose of 300 mg/kg bw improved packed cell volume the most by 14.35%. The group treated with 150 and 600 mg/kg bw of the extract showed significant decrease (P < 0.05) in alanine transaminase and aspartate transaminase levels which were lower than those of the group treated with diminazene aceturate. The group treated with 150 mg/kg bw of the extract showed the least urea, albumin and protein level and lowest relative organ weight. There was a significant difference (P < 0.05) in the levels of catalase and Thiobarbituric acid reactive substances in liver and kidney of the animals in the infected-untreated group and the extracts-treated groups. The results of this study show that the extracts of A. albida have antitrypanosomal activity against T. evansi infection.     

In vitro investigation on cinnabar dissolution

To study the effects of different chemical factors in the gastrointestinal tract, i.e. pH, proteins, amino acids, ionic strength and Na₂S, on the dissolution of cinnabar. The content of the total mercury in various dissolutions of cinnabar was analyzed by UV/VIS Spectrophotometer. Laser Particle Size Analyzer measured the particle distributions in the dissolution of cinnabar. The chemical species of dissolved substance of cinnabar in the presence of Na₂S were determined using ESI-MS. The results indicate that the solubility of cinnabar could be increased significantly in the presence of Na₂S/S⁰, and strong acidic pH, respectively. While the influence of thiol amino acid on promoting dissolution remains relatively low. Cinnabar did not dissolve in the form of nanoparticle. It is postulated that cinnabar could be dissolved in the gastrointestinal tract in various forms of sulfur-containing mercury complexes. __________ Translated from China Journal of Chinese Materia Medica, 2007, 32(3): 231–234 (in Chinese) [译自: 中国中药杂志]     

Sulfur isotope analysis of cinnabar from Roman wall paintings by elemental analysis/isotope ratio mass spectrometry – tracking the origin of archaeological red pigments and their authenticity

The most valuable pigment of the Roman wall paintings was the red color obtained from powdered cinnabar (Minium Cinnabaris pigment), the red mercury sulfide (HgS), which was brought from mercury (Hg) deposits in the Roman Empire. To address the question of whether sulfur isotope signatures can serve as a rapid method to establish the provenance of the red pigment in Roman frescoes, we have measured the sulfur isotope composition (δ³⁴S value in ‰ VCDT) in samples of wall painting from the Roman city Aventicum (Avenches, Vaud, Switzerland) and compared them with values from cinnabar from European mercury deposits (Almadén in Spain, Idria in Slovenia, Monte Amiata in Italy, Moschellandsberg in Germany, and Genepy in France). Our study shows that the δ³⁴S values of cinnabar from the studied Roman wall paintings fall within or near to the composition of Almadén cinnabar; thus, the provenance of the raw material may be deduced. This approach may provide information on provenance and authenticity in archaeological, restoration and forensic studies of Roman and Greek frescoes. Copyright © 2010 John Wiley & Sons, Ltd.     

LC Tissue Distribution Study of Paeonol in Rats after Oral Administration

Paeonol, an important constituent of the traditional Chinese medicine Cortex Moutan, has a variety of bioactivity. A simple and sensitive HPLC?CUV method has been developed for analysis of paeonol in different rat tissue (heart, liver, spleen, lung, kidney, and brain). Bio-samples were prepared by simple protein precipitation, and separation of paeonol was achieved on a C₁₈ column with methanol?C2% glacial acetic acid solution 70:30 (v/v) as mobile phase at a flow rate of 1.0 mL min^?1. UV detection was at 274 nm and the column temperature was 30 °C. Linearity was good between 0.025 and 5.0 ??g mL^?1 (r ² ?? 0.9990) for tissue samples. Inter-day and intra-day accuracy (as relative error, RE) and precision (as relative standard deviation, RSD) were <5.94 and 6.05%, respectively. The limit of detection was 0.025 ??g mL^?1 and extraction recovery for all samples was ??85.86%. The method was successfully applied to a tissue-distribution study after oral administration of 40 mg kg^?1 paeonol to healthy Sprague?CDawley rats. The study showed that paeonol was quickly distributed and eliminated after oral administration; liver and kidney were the major distribution tissues of paeonol in rats, and paeonol quickly passed through the blood?Cbrain barrier. It was also found there was no long-term accumulation of paeonol in rat tissues.     

剌梨利康饮对汞中毒大鼠汞铜锌含量的影响

用高汞水喂养大鼠８周,复制出慢性汞中毒模型,再分别自由饮用利康饮饮料和腹腔注射二巯基丙磺酸钠３周,探讨剌梨利康饮和二巯基丙磺酸钠对慢性汞中毒大鼠体内汞、铜、锌含量的影响。结果显示：慢性汞中毒引起血清、肝、脑和肾中汞含量升高的同时,引起血清、脑和肾中铜、锌含量及肝中锌含量降低;利康饮可降低血清和肾中汞含量,并可提高血清、脑和肾中铜、锌含量及肝含量;二巯基丙磺酸钠虽可降低血清、肝和肾中汞含量,升高血清     

Methylmercury in marine fish from Malaysian waters and its relationship to total mercury content

The study evaluated methylmercury concentrations, the methylmercury to total mercury ratio (%MeHg) and their correlations in ten fish species from different trophic levels. Methylmercury levels in fish studied were in the range of 0.007 to 0.914 µg g^?1 wet wt. Muscle tissue of predatory fish contained significantly (p < 0.05) higher content of methylmercury than non-predatory fish. The methylmercury to total mercury ratio ranged from 49.1% to 87.5%, with the highest ratio in predatory fish. This ratio was always higher in muscle tissue compared to the liver tissues, indicating tissue-specific binding and accumulation of methylmercury in the muscle. All the fish species showed strong positive correlation between methylmercury and total mercury levels (R ²> 0.86). Except for long tail tuna and short-bodied mackerel, all fish species showed lower methylmercury levels and estimated weekly intake as compared to the maximum values established by US FDA (of 0.5 µg g^?1) and by FAO/WHO (1.5 µg kg^?1 bodyweight), respectively. This study showed that the percentage of methylmercury is rather high in fish and fish represents the major source of this toxic mercury form to the local population.     

Determination of 3-chloro-1,2-propanediol in biological samples by quick-easy-cheap-effective-rugged-and-safe extraction coupled with gas chromatography-mass spectrometry and its biodistribution in rats

3-Chloro-1,2-propanediol is a common food contaminant, but reports on its determination in biological tissues are lacking. In the present study, a method was developed to detect 3-chloro-1,2-propanediol contents in rat tissues by quick-easy-cheap-effective-rugged-and-safe extraction and gas chromatography-mass spectrometry analysis. Biological samples were extracted with ethyl acetate and purified with adsorbents. The optimized adsorbent for each sample was selected from 4–5 combinations of N-propylethylenediamine, octadecylsilane, graphitized carbon black, strong anion exchange, and florisil. Extracted 3-chloro-1,2-propanediol was derivatized with heptafluorobutyric anhydride and subjected to gas chromatography-mass spectrometry. This method had good linearity (correlation coefficients >0.99) in the range of 2–2000 ng/g for blood, kidney, liver, testis, and brain samples. The limits of detection were under 0.8 ng/g; the limits of quantification were 2 ng/g; the recovery rates were 85%–102%; and the matrix effects were 1.98%–7.67%. This method also had good precision. The dynamic changes in 3-chloro-1,2-propanediol in rats gavaged with 20 mg/kg b.w. for 24 h were detected using this method. The 3-chloro-1,2-propanediol content in each tissue sharply increased to a peak, rapidly decreased within 2 h, and stabilized at 12 h. 3-Chloro-1,2-propanediol persisted in the kidney, testis, and liver 24 h after gavage.     

Bioaccumulation of metals and induction of metallo‐ thioneins in selected tissues of common carp (Cyprinus carpio L.) co‐exposed to cadmium,mercury and lead

The concentrations of mercury (Hg), cadmium (Cd) and lead (Pb) at various exposure periods were determined in the gill, kidney, liver and muscle of common carp (Cyprinus carpio L.) co‐exposed to 1.0 µg ml^?1 each of Cd²⁺, Hg²⁺ and Pb²⁺ for up to 10 days. Metallothionein fractions (MTs) in these organs were characterized using the hyphenated technique of size‐exclusion chromatography (SEC) and inductively coupled plasma mass spectrometry (ICP‐MS). After 10 days of exposure, maximum toxic metal concentrations of Hg, Cd and Pb were 10.7 (gill), 0.145 (kidney) and 0.112 µg g^?1_dryweight (gill), respectively. The pattern of accumulation of Hg and Pb was in the order gill > kidney > liver > muscle. In the case of Cd, accumulation was in the order kidney > gill > liver > muscle. Cd and Hg binding MTs were significantly induced in the gill, kidney and liver of all the exposure groups in comparison with the control group (p < 0.05), and the amounts of them increased with the longer exposure time. Despite the higher intracellular Hg concentration and the stronger Hg? SH binding affinity, the amount of Cd‐binding MTs was much higher than that of Hg‐binding MTs. The results indicate that MT synthesis in these organs was clearly metal‐specific. MTs in gill may be used as a bio‐marker to detect the metal pollution caused by Hg and Cd. Zinc and copper binding MTs in the organs of the exposed fish were also increased. This may be due to the MTs' important role in the homeostatic regulation of essential metals and their protective role against the acute toxicity of non‐essential metals. Even though there was considerable accumulation of lead in the organs of the exposed fish, Pb‐binding MT synthesis was non‐significant. Copyright © 2006 John Wiley & Sons, Ltd.     

Voltammetric Studies of Cadmium‐ and Zinc‐Containing Metallothioneins at Nafion‐Coated Mercury Thin Film Electrodes

Voltammetric studies of rabbit liver metallothioneins (MTs, containing both Zn and Cd ions) and Zn₇‐MT were carried out at Nafion‐coated mercury film electrodes (NCMFEs). The accumulation of MT molecules into the NCMFEs enhances the voltammetric signals and the electrostatic interaction between the Nafion membrane and MT facilitates facile electron transfer reactions. Two well‐defined redox waves, with reduction potential (E_pc) values at ?0.740 and ?1.173 V, respectively, were observed. The peak at E_pc =?0.740 V is attributable to the reduction of the Cd‐MT complex, whereas that at E_pc=?1.173 V was assigned to the reduction of the Zn‐MT complex. Zn₇‐MT exhibits only one redox wave with E_pc=?1.198 V. The NCMFE was found to be more advantageous than thin mercury film electrode (MFE), because the pristine metal ions in MTs (e.g., Cd²⁺ and/or Zn²⁺) are not significantly replaced by Hg²⁺. The NCMFE is also complementary to Nafion‐coated bismuth film electrode in that it has a greater hydrogen overpotential, which allows the reduction of the Zn‐MT complex to be clearly observed. Moreover, intermetallic compound formation between Cd and Zn appears to be less serious at NCMFEs. Consequently, the amounts of Cd and Zn deposited into the electrode upon the reduction reactions can be quantified more accurately.     

Pharmacokinetics,tissue distribution and excretion of a recombinant fusion protein <Superscript>125</Superscript>I-rhTNT-IL2

This study examined the tissues distribution and pharmacokinetics of rhTNT-IL2, a fusion protein, in rats and macaques after iv. injection. The rhTNT-IL2 was labeled with ¹²⁵I. The ¹²⁵I-rhTNT-IL2 was administered into rats and macaques at a dose of 250 μg·kg⁻¹ and 125 μg·kg⁻¹, respectively. Upon administration, rhTNT-IL2 declined in tri-exponentially with the half-lives of 1.78 h (T _1/2α), 25.06 h (T _1/2β), 114.19 h (T _1/2γ) for rats and 1.87 h (T _1/2α), 9.82 h (T _1/2β), 43.17 h (T _1/2γ) for macaques. The fraction of ¹²⁵I-rhTNT-IL2 excreted in feces was low (about 1%), while the majority of ¹²⁵I-rhTN-IL2 was excreted in urine. The tissue distributions showed that the liver, spleen and heart were major organs for deposition of the fusion protein in rats and macaques in 48-hour post-injection, and the fusion protein could not penetrate through blood brain barrier.     

Direct analysis of mercury in Traditional Chinese Medicines using thermolysis coupled with on-line atomic absorption spectrometry

The purpose of this study is to develop and apply a mercury analyzer system capable of quantitative analysis of mercury in Traditional Chinese Medicines (TCM) drugs in the concentrations range from ng g⁻¹ to mg g⁻¹. No sample pre-treatment was needed and this greatly simplifies the analytical procedure and minimizes potential sources of contamination. The precisions of analyzing solid mercury standard sample and real TCM materials were 2.1% and 2.5-8.2%, respectively; and the recovery based on the analysis of standard reference materials ranged from 95.2% to 105%. The performance of the method has been compared with inductively coupled plasma-mass spectrometry (ICP-MS) technique and excellent agreements were observed between the two methods. The method has been applied to the investigation of Hg content in several TCM drugs containing or not containing cinnabar. Mercury concentration in the same TCM products differs widely with different manufacturers, suggesting that external contamination and the Hg presence in raw herbal materials are the main sources of Hg. In addition, comparison of mercury thermal releasing profiles between TCM drug and cinnabar suggests that mercury conversion from cinnabar to biological matrices-bound Hg could occur because of the aid of other ingredients in the formulated drug.     

Pharmacokinetics,protein binding and metabolism of a quinoxaline urea analog as an NF‐κB inhibitor in mice and rats by LC‐MS/MS

13–197 is a novel NF‐κB inhibitor that shows promising in vitro efficacy data against pancreatic cancer. In this study, we characterized the pharmacokinetics, tissue distribution, protein binding and metabolism of 13–197 in mice and rats. A valid, sensitive and selective LC‐MS/MS method was developed. This method was validated for the quantification of 13–197, in the range of 0.1 or 0.2‐500 ng/mL in mouse plasma, liver, kidney, lung, heart, spleen, brain, urine and feces. 13–197 has low bioavailability of 3 and 16% in mice and rats, respectively. It has faster absorption in mice with 12‐fold shorter T_max than in rats. Tissue concentrations were 1.3–69.2‐fold higher in mice than in rats at 72 h after intravenous administration. 13–197 is well distributed to the peripheral tissues and has relatively high tissue–plasma concentration ratios, ranging from 1.8 to 3634, in both mice and rats. It also demonstrated more than 99% binding to plasma proteins in both mice and rats. Finally, <1% of 13–197 is excreted unchanged in urine and feces, and metabolite profiling studies detected more than 20 metabolites in mouse and rat plasma, urine and feces, which indicates that 13–197 is extensively metabolized and primarily eliminated by metabolism rather than by excretion. Copyright © 2013 John Wiley & Sons, Ltd.     

Determination of d‐limonene in mice plasma and tissues by a new GC–MS/MS method: Comparison of the pharmacokinetics and tissue distribution by oral and inhalation administration in mice

The aim of the present study was to develop a method based on gas chromatography–tandem mass spectrometry (GC–MS/MS) to determine and quantify the d ‐limonene in mouse plasma and tissue samples. This new method was validated for the quantification of d ‐limonene with the linearity ranges 1.0–1000.0 ng/mL (r² > 0.9952) for plasma samples and 5.0–5000.0 ng/g (r² > 0.9940) for tissue samples. The intra‐ and inter‐day assay of precisions in plasma and tissues were <13.4% and the accuracies were within 91.1–105.8%. In the oral/inhalation administration pharmacokinetics and tissue distribution studies, the main pharmacokinetic parameters were the peak concentration = (97.150 ± 34.450)/(4336.415 ± 1142.418) ng/mL, the area under the curve = (162.828± 27.447)/(2085.721 ± 547.787) h ng/mL and the half‐life = (3.196 ± 0.825)/(0.989 ± 0.095) h. The tissue distribution of d ‐limonene in mice after oral/inhalation administration demonstrated a decreasing tendency in different tissues (liver > kidney > heart > lung > spleen).     

Simple mercury fractionation in biological samples by CV AAS following microwave-assisted acid digestion or TMAH pre-treatment

A simple and reliable method to determine total and inorganic mercury in biological certified reference material (CRM) by cold vapor atomic absorption spectrometry (CV AAS) is proposed. After the CRM treatment at room temperature with tetramethylammonium hydroxide (TMAH), inorganic mercury is determined by CV AAS. Total mercury is measured by the same technique, after sample acid digestion in a microwave oven. Organic mercury, basically methylmercury, is obtained by difference. In both procedures, the quartz tube is kept at room temperature. By means of analysis of the following reference materials: pig kidney, lobster hepatopancreas, dogfish liver and mussel tissue, it was clear that the difference between the total and inorganic mercury concentrations agrees with the methylmercury concentration. Only one calibration curve against aqueous standards in acidic medium was carried out for both procedures. The concentrations obtained by both procedures are in agreement with the certified values according to the t-test at a 95% confidence level. The relative standard deviations were lower than 3.0% for digested CRM and 6.0% for CRM treated with TMAH for most of the samples. The limits of detection in the samples were 0.02 µg g^{− 1} and 0.04 µg g^{− 1} for inorganic and total Hg, respectively, since the sample mass for total mercury was half of that for inorganic mercury determination. Simplicity and high efficiency without using chromatographic techniques are some of the qualities of the proposed method, being adequate for fractionation analysis of mercury in biological samples.     

Gradient elution mode for the troubleshooting of matrix effect on the determination of G004 in different tissues by LC‐MS/MS

A steep gradient elution mode was applied to reduce the risk of matrix effect (ME) for the determination of G004, a novel sulfonylurea hypoglycemic drug, in a tissue distribution study by LC‐MS/MS. The mass spectra of the total‐ion‐current chromatograms combined with the post‐column infusion traces enabled the ‘unseen’ interfering species to be directly detected, and ensured that the chromatography conditions and sample preparation method were adequate to overcome the ME. According to this, a steep gradient elution mode was designed to overcome the intense ME from different tissues. The analysis was performed by monitoring the transitions m/z 558.1 → 419.0 for G004 and m/z 489.3 → 364.1 for glimepiride used as the internal standard. Calibration curves recovered over a range from 0.1 to 10000 ng/mL for seven different tissues. Sex‐related difference was found in the tissue distribution. The drug levels in the tissues of female rats were about two to three times higher than those in male counterparts. The highest level was observed in liver, then in kidney, heart, pancreas, lung and spleen, but no G004 was detected in brain. G004 was slowly eliminated from female rats compared with male rats. There was no long‐term accumulation of G004 in male or female rat tissues. Copyright © 2014 John Wiley & Sons, Ltd.     

Tissue distribution study of columbianadin and its active metabolite columbianetin in rats

Columbianadin, one of the main bioactive constituents of the roots of Angelica pubescens Maxim. f. biserrata Shan et Yuan, has been found to possess obvious pharmacological effects in previous studies. In this study, a valid and sensitive reverse‐phase high‐performance liquid chromatography (RP‐HPLC) method was established and validated for the determination of columbianadin (CBN) and its active metabolite columbianetin (CBT) in rat tissue samples. Sample separation was performed on an RP‐HPLC column using a mobile phase of MeOH–H₂O (75:25, v/v) at a flow rate of 1.0 mL/min. The UV absorbance of the samples was measured at the wavelength 325 nm. The calibration curves for CBN were linear over the ranges of 0.5–20 µg/g for brain, testes and muscle, 1.0–10.0 µg/g for stomach and intestine, and 0.2–20.0 µg/g for heart, liver, spleen, lung and kidney. The calibration curves for CBT were linear over the ranges of 0.5–25 µg/g for stomach and intestine, and 0.1–10.0 µg/g for heart, liver, spleen, lung and kidney. The analysis method was successfully applied to a tissue distribution study of CBN and CBT after intravenous administration of CBN to rats. The results of this study indicated that CBN could be detected in all of the selected tissues after i.v. administration. CBN was distributed to rat tissues rapidly and could be metabolized to CBT in most detected tissues. Of the detected tissues, heart had the highest uptake of CBN, which suggested that heart might be one of the main target tissues of CBN. Concentrations of CBT were obviously higher in the digestive system than in other assayed tissues. The information provided by this research is very useful for gaining knowledge of the capacities of CBN and CBT to access different tissues. Copyright © 2015 John Wiley & Sons, Ltd.     

Pathophysiology of Microwave Radiation: Effect on Rat Brain

The study aims to investigate the effect of 2.45 GHz microwave radiation on Wistar rats. Rats of 35 days old with 130 ± 10 g body weight were selected for this study. Animals were divided into two groups: sham exposed and experimental (six animals each). Animals were exposed for 2 h a day for 45 days at 2.45 GHz frequency (power density, 0.21 mW/cm²). The whole body specific absorption rate was estimated to be 0.14 W/kg. Exposure took place in a ventilated plexiglas cage and kept in an anechoic chamber under a horn antenna. After completion of the exposure period, rats were killed, and pineal gland and whole brain tissues were isolated for the estimation of melatonin, creatine kinase, caspase 3, and calcium ion concentration. Experiments were performed in a blind manner and repeated. A significant decrease (P < 0.05) was recorded in the level of pineal melatonin of exposed group as compared with sham exposed. A significant increase (P < 0.05) in creatine kinase, caspase 3, and calcium ion concentration was observed in whole brain of exposed group of animals as compared to sham exposed. One-way analysis of variance method was adopted for statistical analysis. The study concludes that a reduction in melatonin or an increase in caspase-3, creatine kinase, and calcium ion may cause significant damage in brain due to chronic exposure of these radiations. These biomarkers clearly indicate possible health implications of such exposures.