Investigation on the interaction between Ag+ and bovine hemoglobin using spectroscopic methods.

Silver ions (Ag+) can be released by silver nanoparticles (AgNPs) which are widely used in diverse fields. Ag+ can exist inside cells to produce cytotoxicity. This report uses spectroscopic methods to reveal the interactions between Ag+ and bovine hemoglobin (BHb). The results of the quenching rate constant (Kq) and the fluorescence lifetime detection showed that the quenching mechanism of BHb by Ag+ was static. Thermodynamic investigations indicated that Ag+ can interact with BHb with one binding site to form complex mainly through van der Waals interactions and hydrogen bonds. The UV-vis absorption and synchronous fluorescence spectra showed that Ag+ changed the conformation of BHb, which may affect protein functions. This research is favorable for understanding the molecular toxic mechanism of Ag+ in vivo.

Comparative study on the toxic mechanisms of medical nanosilver and silver ions on the antioxidant system of erythrocytes: from the aspects of antioxidant enzyme activities and molecular interaction mechanisms.

BACKGROUND: The wide application of silver nanoparticles (AgNPs) in medicals and daily utensils increases the risk of human exposure. The study on cell and protein changes induced by medical AgNPs (20 nm) and Ag+ gave insights into the toxicity mechanisms of them. RESULTS: AgNPs and Ag+ affected the enzymatic and non-enzymatic antioxidant systems of red blood cells (RBCs). When RBCs were exposed to AgNPs or Ag+ (0-0.24 µg/mL), catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPX) were more sensitive to Ag+, whereas the RBCs had slightly higher glutathione (GSH) contents treated by AgNPs. Both AgNPs and Ag+ increased the malondialdehyde (MDA) content of RBCs, but the difference was not significant. The difference in the change of the enzyme activity indicated that AgNPs and Ag+ have different influencing mechanisms on CAT and GPX. And SOD has stronger resistance to both of AgNPs and Ag+. When AgNPs or Ag+ (0-10 µg/mL) was directly applied on enzymatic proteins, although AgNPs or Ag+ at a high concentration was toxic, at the concentration below 0.4 µg/mL could promote the activities of CAT/SOD/GPX. The spectroscopic results (fluorescence, synchronous fluorescence, resonance light scattering and ultraviolet absorption), including the changes in amino acid microenvironment, peptide chain conformation, and aggregation state, indicated that the interaction mechanism and conformational changes were also the important factors for the changes in the activities of SOD/CAT when SOD/CAT were directly exposed to AgNPs or Ag+. CONCLUSIONS: Low concentration (< 0.4 µg/mL) of AgNPs is relatively safe and the direct effects of AgNPs and Ag+ on enzymes are important reasons for the change in antioxidant capacity of RBCs.

Study on the mechanism of action between dimethyl phthalate and herring sperm DNA at molecular level.

Dimethyl phthalate (DMP), a typical phthalic acid ester, is widespread in the environment and causes extensive concern due to its adverse effects on human health. To understand the genotoxicity of DMP at molecular level, the toxic interaction of DMP with herring sperm (hs) deoxyribonucleic acid (DNA; hs-DNA) was investigated in vitro under simulated physiological conditions using multi-spectroscopic techniques and a molecular modeling method. The results of Ultraviolet-Visible absorption spectroscopy, fluorescence emission spectroscopy, and circular dichroism spectra indicated that DMP interacts with hs-DNA in a groove-binding mode that changes the double helical structure of DNA. The binding constant and the number of binding sites calculated from the fluorescence quenching data were 565.718 L mol(-1) and 0.7872, respectively. A molecular modeling study revealed that DMP tends to bind with DNA in the A-T-rich regions of minor groove and that hydrogen bonding and van der Waals forces play main roles in the interaction. This research can help to elucidate the mechanism of DMP toxicity in vivo.

Binding of the veterinary drug tetracycline to bovine hemoglobin and toxicological implications.

Tetracycline (TC) is a widely used veterinary drug in animal breeding and fishery. Because of its low bioavailability, the TC residue extensively exists in the environment (e.g. soils, lakes and rivers), which can enter the human body, being potentially harmful. Hemoglobin (Hb) is a protein responsible for oxygen carrying in the vascular system of animals. The aim of this study was to investigate the interaction of bovine hemoglobin (BHb) with TC through spectroscopic and molecular modeling methods. The experimental results revealed that TC can interact with BHb with one binding site to form a TC-BHb complex, mainly through van der Waals interactions and hydrogen bonds. The UV-visible absorption, synchronous fluorescence, and circular dichroism (CD) results revealed that the binding of TC can cause conformational and some microenvironmental changes of BHb, which may affect BHb physiological functions. The synchronous fluorescence experiment disclosed that TC binds into BHb central cavity, which was verified by molecular modeling study. The work contributes to clarify the molecular mechanism of TC toxicity in vivo.

Study on the combined toxicity of DEHP and lead on the blood system of rats.

Di(2-ethylhexyl) phthalate (DEHP) is a commonly used phthalate ester compound, while lead is a persistent and bioaccumulative heavy metal. Both can be exposed to the body through a variety of ways, which may have an impact on the blood system. In this study, we examined the impact of co-exposure to DEHP (0, 10, 100 mg/kg) and Pb (0, 5, 50 mg/kg) on the blood system of male SD rats. The study revealed that continuous exposure to DEHP and Pb for 20 days resulted in a decrease in leukocytes and lymphocytes, while an increase in neutrophils and monocytes. Co-exposure led to a significant decrease in the spleen coefficients. Furthermore, the combined exposure could increase the ratio of bone marrow cells in G1 phase, and decrease the ratio of cells in S phase and G2 phase. Cytokine testing showed that combined exposure affects the secretion of hematopoietic factors and may cause bone marrow cell apoptosis. Single or combined exposure to DEHP and Pb can cause oxidative stress in serum and bone marrow. Overall, these results indicate that the co-exposure of DEHP and Pb adversely affected the blood system of rats, mainly due to the induction of oxidative stress and ultimately affects the secretion of cytokines. The combined effect of the two substances is primarily antagonistic. These results have important implications for the risk assessment of combined pollution and provide valuable theoretical guidance.

Dimethyl phthalate induces blood immunotoxicity through oxidative damage and caspase-dependent apoptosis.

Dimethyl phthalate (DMP), a low-molecular-weight phthalate ester, exists in ectoparasiticides, plastics, and insect repellants, and has been linked to neurotoxic, reproductive, and endocrine disruptive responses. However, its blood immunotoxic effects and mechanism are still poorly understood. In this study, rats were exposed to gradient concentrations of DMP through intragastric administration to assess the blood immunotoxic effects in the combined assay of biomarker, cytometry, and transcriptomics. DMP treatment altered the redox status of rats, thus causing oxidative damage. Significantly decreased blood cell counts and disordered antibody and cytokine secretion were observed in treated rats, suggesting the suppressed immune defense and destructed inflammatory regulation. Flow cytometry showed that in lymphocytes, especially CD3+CD4+ T cells, the occurrence of apoptosis/necrosis was positively related to DMP exposure level. Transcriptomics revealed an oxidative stress-related mechanism. The overexpression of the Bcl-2 family genes and the activation of the Fas/FasL pathway triggered downstream caspase cascade and caused reactive oxygen species signaling-mediated apoptosis/necrosis. To the best of our knowledge, it was the first report that the exposure to low-molecular-weight phthalate esters potentially triggered blood immunotoxicity. The result and underlying mechanisms can provide an essential basis for understanding phthalate ester toxicity and usage regulation.

Phenotypic characterization of the binding of tetracycline to human serum albumin.

Because of the widely usage of the veterinary drug tetracycline (TC), its residue exist extensively in the environment (e.g., animal food, soils, surface water, and groundwater) and can enter human body, being potential harmful. Human serum albumin (HSA) is a major transporter for endogenous and exogenous compounds in vivo. The aim of this study was to examine the interaction of HSA with TC through spectroscopic and molecular modeling methods. The inner filter effect was eliminated to get accurate binding parameters. The site marker competition experiments revealed that TC binds to site II (subdomain IIIA) of HSA mainly through electrostatic interaction, illustrated by the calculated negative ΔH° and ΔS°. Furthermore, molecular docking was applied to define the specific binding sites, the results of which show that TC mainly interacts with the positively charged amino acid residues Arg 410 and Lys 414 predominately through electrostatic force, in accordance with the conclusion of thermodynamic analysis. The binding of TC can cause conformational and some microenvironmental changes of HSA, revealed by UV-visible absorption, synchronous fluorescence, and circular dichroism (CD) results. The accurate and full basic data in the work is beneficial to clarifying the binding mechanism of TC with HSA in vivo and understanding its effect on protein function during the blood transportation process.

Study on the binding of cerium to bovine serum albumin.

The interaction of Ce(3+) to bovine serum albumin (BSA) has been investigated mainly by fluorescence spectra, UV-vis absorption spectra, and circular dichroism (CD) under simulative physiological conditions. Fluorescence data revealed that the quenching mechanism of BSA by Ce(3+) was a static quenching process, the binding constant is 6.70 × 10(5) , and the number of binding site is 1. The thermodynamic parameters (ΔH = -29.94 kJ mol(-1) , ΔG = -32.38 kJ mol(-1) , and ΔS = 8.05 J mol(-1) K(-1) ) indicate that electrostatic effect between the protein and the Ce(3+) is the main binding force. In addition, UV-vis, CD, and synchronous fluorescence results showed that the addition of Ce(3+) changed the conformation of BSA.

Biodegradation performance and biofouling control of a halophilic biocarriers-MBR in saline pharmaceutical (ampicillin-containing) wastewater treatment.

This work develops a halophilic biocarriers-MBR for saline pharmaceutical wastewater treatment. The system has effectively treated the ampicillin-containing saline wastewater for 32 days, when the ampicillin concentration is lower than 20 mg/L. The system can tolerate the saline organic wastewater with a reasonable biodegradability (removals of COD over 75%) when the ampicillin concentration is 50 mg/L. The system has a bad performance in biodegradation (COD removals around 60-70%) and fouled within 16 days at a high ampicillin concentration of 100 mg/L. At high transmembrane pressures over 30 KPa, some ampicillin molecules may permeate through the membrane causing decreases in the ampicillin removal. The concentrations of protein and carbohydrate in EPS and SMP have increased over time and with increasing the ampicillin concentration. The method of biofouling control in MBR for the ampicillin situations has been proposed based on monitoring the concentrations of EPS and SMP. The drying-assisted monitoring of membrane biofoulants has showed a better efficiency than the monitoring of transmembrane pressure for membrane anti-biofouling in the treatment of pharmaceutical saline wastewaters where a spectroscopic detection can be hardly applied. This work may benefit relative research works for the control of biodegradation performance and membrane biofouling to better treat saline pharmaceutical wastewaters.

Revealing the toxicity of dimethyl phthalate (DMP) to the oxygen-carrying function of red blood cells (RBCs): The iron release mechanism.

Phthalic acid esters (PAEs), as typical hormone pollutants, do harms to human health after enrichment over a long term exposure, causing the loss of oxygen-carrying function of red blood cells (RBCs). This study has investigated the mechanism for the toxicity of dimethyl phthalate (DMP) on the oxygen-carrying function of RBCs by measuring the iron release content of hemoglobin (Hb) in vivo and in vitro. The hematologic examination showed that the high dose of DMP at 1000 mg/kg significantly reduced the Hb content and increased the granulocyte content, whereas such toxicity was not relatively observed at a low (50 mg/kg) or a medium (250 mg/kg) dose of DMP. The in vitro experiments showed that DMP, incubated with RBCs, increased the iron release content as a function of DMP concentration. Interestingly, such a phenomenon was not observed when DMP was incubated with Hb alone, indicating that the release of hemoglobin iron could not directly caused by the combination of DMP and hemoglobin. The in vivo experiments indicated that DMP induced iron release and oxidative stress for rat RBCs. Moreover, vitamin C and E was found to reduce the level of iron release by recovering erythrocytes from the oxidative stress induced by DMP. This work has revealed that the oxidative stress induced by DMP, causing the release of Hb iron from RBCs, is the reason for the toxicity of DMP to the oxygen-carrying function.

Noncovalent interaction of oxytetracycline with the enzyme trypsin.

Oxytetracycline (OTC) is a kind of widely used veterinary drugs. The residue of OTC in the environment (e.g., animal food, soils, surface, and groundwater) is potentially harmful. In this article, the binding mode of OTC with trypsin was investigated using spectroscopic and molecular docking methods. OTC can interact with trypsin with one binding site to form OTC-trypsin complex, resulting in inhibition of trypsin activity and change of the secondary structure and the microenvironment of the tryptophan residues of trypsin. After elimination of the inner filter effect, the association constant, K, was calculated to be K(290K) = 1.36 × 10(5) L mol(-1), K(298K) = 7.30 × 10(4) L mol(-1), and K(307K) = 3.58 × 10(4) L mol(-1) at three different temperatures. The calculated thermodynamic parameters (negative values of ΔH(○) and ΔS(○)) indicated that van der Waals interactions and hydrogen bonds play a major role during the interaction. The molecular docking study revealed that OTC bound into the S1 binding pocket, which illustrates that the trypsin activity was competitively inhibited by OTC, in accordance with the conclusion of the trypsin activity experiment. This work establishes a new strategy to probe the toxicity of OTC and contributes to clarify its molecular mechanism of toxicity in vivo. The combination of spectroscopic and molecular docking methods in this work can be applied to investigate the potential enzyme toxicity of other small organic pollutants and drugs.

Spectroscopic investigation on the toxic interaction of melamine with herring sperm DNA.

The toxic interaction of melamine with herring sperm DNA (hs-DNA) was investigated by using fluorescence and UV-vis absorption spectra techniques. The experimental results showed that the toxic interaction between melamine and hs-DNA occurred. Fluorescence quenching experiments indicated the existence of electrostatic binding between melamine and hs-DNA. The binding constants K(A) and the binding site numbers were calculated by means of the Stern-Volmer equation and were 9.8 × 10(4) L mol(-1) and 1.3, respectively. Both the results of fluorescence spectra and UV-vis absorption spectra verified that there are electrostatic binding between melamine and hs-DNA. The possibility in the presence of a classical intercalation binding mode could be ruled out by using DNA unwinding experiments.

A novel mitochondrial-targeted two-photon fluorescent probe for ultrafast monitoring of SO2 derivatives and its applications.

SO2 derivatives maintain a certain balance and play an important role in many metabolic processes. However, excessive ingestion of them can lead to serious complications of various diseases. Therefore, a rapid and precision detection of SO2 derivatives with high selectivity and sensitivity would be an advance for their bio-analytic studies. Accordingly, a novel deep red and two-photon fluorescent SO2 derivatives probe (DRQ) was developed here for the first time. The probe showed fast ratio response rate (within 5 s), excellent sensitivity (the detection limit is 103 nM for red channel and 17 nM for green channel), and outstanding selectivity toward SO2 derivatives in 10 mM PBS buffer. Moreover, the ratiometric probe DRQ displays a 99 nm blue-shift in emission upon addition of SO2 derivatives. Intriguingly, the probe can be made into test papers for real-time monitoring of SO2 derivatives. Moreover, it can be also applied for visual ratio imaging of SO2 derivatives in cellular mitochondria and zebrafish under two-photon absorption (green channel) and one-photon absorption (red channel).

Impact Assessment of heavy metal cations to the characteristics of photosynthetic phycocyanin.

This work has assessed the impact of typical heavy metal cations on C-phycocyanin in vitro and in silico. At low concentrations (<2×10-6 mol/L), the influence of Pb2+ is the highest on the light absorption of C-phycocyanin trimer. At higher concentrations, however, a new order of influence on the light absorption has been observed with Cd2+ < Cu2+ < Pb2+ < Zn2+. The fluorescence polarization has changed from the order of Cd2+ < Pb2+≈Cu2+ < Zn2+ to Cd2+ < Cu2+ < Pb2+ < Zn2+, when the metal concentrations reaches 2×10-6 mol/L. The mechanisms for these findings have been studied using FTIR, hydrophobic probe, isothermal titration calorimetry and molecular docking for the analysis of structure disorder of C-phycocyanin. It has been suggested that the secondary structure of C-phycocyanin affects more to the light absorbance while the fluorescence characteristics relies more on the tertiary structure. The interaction between Pb2+ and C-phycocyanin is both enthalpically and entropically favoured, whereas the interactions for Cd2+, Cu2+ and Zn2+ are entropically driven. The ion-molecular docking suggests that the structure disorder of C-phycocyanin relies on the molecular interactions with metal ions. The in silico study also showed that the binding cites of Zn2+ are closer to chromophores.

A new strategy to probe the genotoxicity of silver nanoparticles combined with cetylpyridine bromide.

The environmental genotoxic behavior of silver nanoparticles (nanoAg) combined with the detergent cetylpyridine bromide (CPB) was examined in vitro. The experimental results showed that the genotoxicity of nanoAg itself is weak, but nanoAg shows obvious genotoxicity after combined with CPB. The combined materials have a strong coeffect on calf thymus DNA (ctDNA) at a concentration of 3.3 x 10(-6 )gmL(-1) nanoAg and 6.0 x 10(-6) molL(-1) CPB. After the addition of ctDNA to the nanoAg-CPB system, the particles are scattered and the diameter decreases, which indirectly reveal that nanoAg-CPB has genotoxicity.

In vitro toxicity of dimethyl phthalate to human erythrocytes: From the aspects of antioxidant and immune functions.

In the study, the effects of dimethyl phthalate (DMP) on the antioxidant defense capacity and immune functions of human erythrocytes were experimentally explored. DMP affected the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) and the contents of glutathione (GSH) and malondialdehyde (MDA) in erythrocytes, thus impairing the function of antioxidant defense system of erythrocytes. When DMP concentration increased from 0 to 28 µmol L-1, the SOD and GPX activities were increased firstly and then gradually decreased. When DMP concentration was below 20 µmol L-1, the relative activity of SOD was enhanced by DMP and the effect was known as hormesis. The relative activity of GPX was also increased when the concentration of DMP was below 12 µmol L-1. The CAT activity was more significantly inhibited by DMP than the activities of SOD and GPX, whereas the relative GSH content was increased by DMP. MDA levels were significantly changed after the exposure to DMP (0-24 µmol L-1). The experimental results of the activity of SOD and CAT, and the content of MDA also suggested that DMP could inhibit the immune functions of red blood cells (RBCs), which were further proved by the decrease of two indicators (RBC-C3b and RBC-IC) due to the destruction of C3b receptor with immune adherence function on erythrocyte membrane. The study provides a deep understanding of the toxicity of DMP on erythrocytes.

The toxicity of cadmium ion (Cd2+) to phycocyanin: an in vitro spectroscopic study.

The pollution of heavy metals is a severer problem for the ecosystems in waters. The toxicity of Cd2+ on phycocyanin (PC) is studied in molecular level in this work. The fluorescence quenching of PC is observed by the adding Cd2+ from 0 to 500 × 10-7 mol L-1. From the theoretical calculation and the time-resolved fluorescence decay profiles, the fluorescence quenching of PC by Cd2+ is found to be static. The synchronous fluorescence spectra are used to study the change in amino acid residues of PC molecules, indicating that the effect of Cd2+ on the Trp of PC is more significant than the Tyr. The UV-Vis absorbance of tetrapyrrole decreases from 0.26 to 0.23 cps with increasing Cd2+ concentration, suggesting that Cd2+ affects the light adsorption and the photosynthesis function of PC. The circular dichroism spectra reveal that adding Cd2+ also changes the secondary structure (α-helix) of PC.

In vitro assessment of the toxicity of small silver nanoparticles and silver ions to the red blood cells.

This work reports the toxicity of small silver nanoparticles (nanoAg, 20 nm) and silver ions (Ag+) to the red blood cells with the silver concentration level of 10-6 g/mL. Results show that red blood cells (RBCs) start hemolysis when treated by nanoAg of 1.5 × 10-5 g/mL or Ag+ of 2.9 × 10-7 g/mL. A low ATPase activity of 30% has been observed after RBCs being treated with Ag+ of 2.6 × 10-7 g/mL, while the nanoAg does not obviously affect the ATPase activity. In molecular level, Ag+ is more toxic to the amino acid residues than nanoAg according to the change of fluorescence characteristics of hemoglobin (Hb). However, the nanoAg has been found to be more toxic than Ag+ to the secondary structure of Hb in terms of the loss of α-helix content.

In vitro assessment of the toxicity of lead (Pb2+) to phycocyanin.

This work reports the influence of lead (Pb2+) on fluorescence characteristics and protein structure of phycocyanin molecules experimentally in vitro. The fluorescence intensity decreases with the increasing concentration of Pb2+ from 0 to 5 × 10-5 mol L-1, showing the fluorescence quenching of phycocyanin by Pb2+. The quenching process is suggested to be static regarding the calculation results and the experimental results of time-resolved fluorescence decay profiles. The synchronous fluorescence spectra show that the effect of Pb2+ on the Tyr residues of phycocyanin is more significant than the Trp residues. The forming of aggregation by the interaction of Pb2+ with phycocyanin molecules is suggested from the results of resonance light scattering spectra. The UV-Vis spectra of the protein skeleton of phycocyanin have a red-shift of about 10 nm with increasing the Pb2+ concentration from 0 to 5 × 10-5 mol L-1, indicating a change in the protein skeleton and its secondary structure. With the increasing Pb2+ concentration, the two negative peaks (209 nm and 218 nm) on circular dichroism spectra become smaller, showing a decrease of the α-helix structure. These results may give people a deeper understanding of that how the heavy metal (Pb2+) can affect the chemo-physical properties of phycocyanin.

New and clean strategy for the determination of Cu2+ in electroless copper plating baths.

This article concerns a new and clean strategy for the determination of Cu(2+) in electroless copper plating baths with differential spectrophotometry. With this strategy, the problem of too high absorbance of Cu(2+) under plating conditions has been solved. We investigated the influence of copper sulfate, sodium hypophosphite, nickel sulfate, sodium citrate, polyglycol, temperature and pH on the absorption spectrum of Cu(2+) in electroless copper plating baths. Five grams per litre of CuSO(4).5H(2)O solution was selected as the reference solution. Experimental results prove that, this strategy has the merits of fast and high accuracy compared to the traditional techniques. Linearly dependent coefficient of the working curve is 0.9999 and the components in the formula have no obvious effect on the detection of Cu(2+) under experimental conditions. Therefore, we can directly move solutions from the EC plating baths for detection, after that the sample can still go back to the baths without any pollution from the plating process to the environment.