BioMetals (v.26, #1)

Mithramycin (Mith) forms a drug-metal complex with a 2:1 stoichiometry by chelation with a Ni(II) ion, which was determined using circular dichroism spectroscopy. Mith exhibits an increased affinity (~55 fold) for Ni(II) in the presence of DNA compared to the absence of DNA, suggesting that DNA acts as an effective template to facilitate chelation. Also, we characterized the DNA-acting properties of a Ni(II) derivative of Mith. Kinetic analysis using surface plasmon resonance and UV melting studies revealed that NiII(Mith)2 binds to duplex DNA with a higher affinity compared to MgII(Mith)2. The thermodynamic parameters revealed a higher free energy of formation for duplex DNA in the presence of NiII(Mith)2 compared to duplex DNA in the presence of MgII(Mith)2. The results of a DNA-break assay indicated that NiII(Mith)2 is capable of promoting one-strand cleavage of plasmid DNA in the presence of hydrogen peroxide; the DNA cleavage rate of NiII(Mith)2 was calculated to be 4.1 × 10−4 s−1. In cell-based experiments, NiII(Mith)2 exhibited a more efficient reduction of c-myc and increased cytotoxicity compared to Mith alone because of its increased DNA-binding and cleavage activity. The evidence obtained in this study suggests that the biological effects of NiII(Mith)2 require further investigation in the future.
Keywords: Mithramycin; Dimer; Nickel(II); Divalent metal; DNA-binding; DNA cleavage; c-Myc expression; Cell viability

Nickel induces apoptosis in human neutrophils by Marisa Freitas; Pedro Barcellos-de-Souza; Christina Barja-Fidalgo; Eduarda Fernandes (13-21).
Nickel is an ubiquitous transition metal that is industrially applied in many forms, which inevitably leads to a high degree of occupational and environmental exposure. Over-exposure to nickel can produce a variety of adverse effects on human health, including allergy and lung and nasal cancers. In the present study, it is demonstrated, for the first time, that nickel [(Ni(II)] (as a nickel nitrate salt) at concentrations that may be attained in vivo, induces neutrophils’ apoptosis by the intrinsic pathway. The use of diphenyleneiodonium, a NADPH oxidase inhibitor, delayed Ni(II)-induced apoptosis, suggesting that NADPH oxidase-derived reactive oxygen species and subsequent signaling could contribute to this event. This is an important finding since increased apoptosis mediated by nickel may disrupt the physiological activities of neutrophils, with potential impact in its immunological and antimicrobial role.
Keywords: Human neutrophils; Nickel; Apoptosis; NADPH oxidase

White light augments chemotherapeutic potential of cyclophosphamide: an in vitro study by Sandesh Chibber; Iftekhar Hassan; Mohd Farhan; Maria Salman; Imrana Naseem (23-31).
Cyclophosphamide (CYC) is a known chemotherapeutic drug used widely for the treatment of leukemias, lymphomas and some solid tumors. Copper is an essential constituent of chromatin and its level is usually elevated in various malignancies. Combined modality chemotherapy involves the use of drug with other components for cancer treatment, such as radiation therapy or surgery. Photosensitizer anticancer drugs can be used in combination with light and may have synergistic effect on cancer. The present study is an attempt to show that CYC acts as prooxidant when used in combination with Cu(II) and white light. We hypothesize that CYC when given as a chemotherapeutic agent possibly interact with endogenous copper associated with chromatin of the cancer cells and generate ROS besides acting as DNA alkylating agent. Thus, during chemotherapy the oxidative stress is possibly generated by the drug through mobilizing endogenous Cu(II) which may attribute to the cytotoxic death of cancer cell.
Keywords: Cyclophosphamide; White light; Copper; Pro-oxidant; Chemo-phototherapy

Effects of cadmium on the sub-cellular localization of β-catenin and β-catenin-regulated gene expression in NRK-52E cells by Joshua R. Edwards; Kevin Kolman; Peter C. Lamar; Nalini Chandar; Michael J. Fay; Walter C. Prozialeck (33-42).
The E-cadherin/β-catenin complex is a structural component of adherens-type junctions in epithelial cells. Moreover, β-catenin acts as an intracellular signaling molecule that can influence the expression of a variety of genes that regulate apoptosis and cell cycle control. Cadmium (Cd) is an environmental toxicant that causes renal dysfunction and disrupts cadherin-dependent cell–cell adhesion in various types of epithelial cells. In this study, we examined the effects of Cd on the subcellular localization of β-catenin, the cadherin/β-catenin complex and β-catenin-mediated gene transcription in rat proximal tubule NRK-52E cells. Exposure to 5–10 μM Cd for 4 h caused the NRK cells to separate from each other without killing the cells or causing them to detach from the growing surface. This effect was associated with the loss of β-catenin and E-cadherin from the cell–cell contacts and apparent changes in the accumulation of β-catenin in the nuclear cell subfraction. The expression of the β-catenin-sensitive gene, c-jun was significantly increased in cells exposed to 5 μM Cd. However, there was no change in the expression of several other β-catenin-regulated genes including: c-myc, cyclin D1 and matrilysin. Additional studies utilizing the TOPFLASH β-catenin reporter gene construct showed that Cd caused a 2–3 fold increase in the expression of the luciferase reporter gene. Overall, these results indicate that Cd disrupts the cadherin/β-catenin complex in NRK-52E cells, but this effect leads to only partial activation of β-catenin-mediated gene transcription.
Keywords: E-cadherin; N-cadherin; β-catenin; Cadmium; NRK-52E cells

Lanthanum chloride (LaCl3) can affect neurobehavioral development and impair cognitive abilities. The mechanism underlying LaCl3-induced neurotoxic effects is still unknown. The purpose of this research was to investigate the neuronal impairment induced by LaCl3 and discuss the possible mechanism from the aspects of the alteration of glutamate level, intracellular calcium concentration ([Ca2+]i), Bax, Bcl-2 and caspases expression in the hippocampus. Lactational rats were exposed to 0, 0.25, 0.50 and 1.0 % LaCl3 in drinking water, respectively. Their offspring were exposed to LaCl3 by parental lactation and then administrated with 0, 0.25, 0.50 and 1.0 % LaCl3 in drinking water for 1 month. The results showed that 0.25, 0.50 and 1.0 % LaCl3 exposure induced neuronal impairment in the hippocampus of young rat. Hippocampal glutamate level, [Ca2+]i and ratio of Bax and Bcl-2 expression increased significantly after LaCl3 exposure. Besides, LaCl3 exposure increased GRP78, GRP94, GADD153 and p-JNK expression, promoted the activation of caspase-3, caspase-9 and caspase-12, induced PARP cleavage and caused excessive apoptosis. These results indicate that LaCl3 increases glutamate level, [Ca2+]i and ratio of Bax and Bcl-2 expression, which cause excessive apoptosis by the mitochondrial and endoplasmic reticulum stress-induced pathway, and thus neuronal damages in the hippocampus.
Keywords: Lanthanum; Glutamate; Intracellular calcium; Caspases; Hippocampus

Cisplatin is one of the most active cytotoxic agents used in the treatment of cancer. However, cisplatin therapy is also associated with severe side effects like nephrotoxicity and genotoxicity. Free oxygen radicals are known to play a major role in cisplatin induced toxicities. Selenium is believed to be an important trace element and dietary antioxidant because of its ability to scavenge free oxygen radicals, thereby preventing cells from oxidative stress. The purpose of this study is to evaluate the protective role of a novel naphthalimide based organoselenium compound 2-(5-selenocyanato-pentyl)-benzo[de]isoquinoline 1,3-dione against cisplatin induced toxicities in Swiss albino mice. Cisplatin was administered intraperitoneally (5 mg/kg b.w.) and the organoselenium compound was given by oral gavages (3 mg/kg b.w.) in concomitant and pretreatment schedule. The results showed that the test compound substantially reduced cisplatin induced reactive oxygen species generation and lipid peroxidation in kidney as well as blood urea nitrogen and creatinine levels in serum. Treatment with organoselenium compound was also able to restore the renal antioxidant system by modulating the cisplatin induced depleted activities of glutathione S-transferase, thioredoxin reductase, superoxide dismutase, catalase, glutathione peroxidase and reduced glutathione level. In addition, the organoselenium compound could efficiently minimize cisplatin induced chromosomal aberrations in bone marrow cells and extent of DNA damage in lymphocytes. Furthermore, the chemoprotective efficacy of the compound against cisplatin induced toxicity was confirmed by histopathological evaluation. The results suggest that the organoselenium compound has the potential to protect against cisplatin induced nephrotoxicity and genotoxicity in part by scavenging reactive oxygen species and by up regulating the antioxidant enzyme system.
Keywords: Organoselenium; Cisplatin; Nephrotoxicity; Reactive oxygen species; Antioxidant enzyme system; DNA damage

An EPR, thermostability and pH-dependence study of wild-type and mutant forms of catechol 1,2-dioxygenase from Acinetobacter radioresistens S13 by Raffaella Caglio; Enrica Pessione; Francesca Valetti; Carlo Giunta; Elena Ghibaudi (75-84).
Intradiol dioxygenase are iron-containing enzymes involved in the bacterial degradation of natural and xenobiotic aromatic compounds. The wild-type and mutants forms of catechol 1,2-dioxygenase Iso B from Acinetobacter radioresistens LMG S13 have been investigated in order to get an insight on the structure–function relationships within this system. 4K CW-EPR spectroscopy highlighted different oxygen binding properties of some mutants with respect to the wild-type enzyme, suggesting that a fine tuning of the substrate-binding determinants in the active site pocket may indirectly result in variations of the iron reactivity. A thermostability investigation by optical spectroscopy, that reports on the state of the metal center, showed that the structural stability is more influenced by the type rather than by the position of the mutation. Finally, the influence of pH and temperature on the catalytic activity was monitored and discussed in terms of perturbations induced on the tertiary contact network of the enzyme.
Keywords: EPR spectroscopy; Structure–function relationships; Intradiol dioxygenase; Aromatic compound degradation; Protein stability; Iron-containing enzyme

Reactions of copper macrocycles with antioxidants and HOCl: potential for biological redox sensing by Rebecca J. Sowden; Katherine D. Trotter; Lynsey Dunbar; Gemma Craig; Omer Erdemli; Corinne M. Spickett; John Reglinski (85-96).
A series of simple copper N2S2 macrocycles were examined for their potential as biological redox sensors, following previous characterization of their redox potentials and crystal structures. The divalent species were reduced by glutathione or ascorbate at a biologically relevant pH in aqueous buffer. A less efficient reduction was also achieved by vitamin E in DMSO. Oxidation of the corresponding univalent copper species by sodium hypochlorite resulted in only partial (~65 %) recovery of the divalent form. This was concluded to be due to competition between metal oxidation and ligand oxidation, which is believed to contribute to macrocycle demetallation. Electrospray mass spectrometry confirmed that ligand oxidation had occurred. Moreover, the macrocyclic complexes could be demetallated by incubation with EDTA and bovine serum albumin, demonstrating that they would be inappropriate for use in biological systems. The susceptibility to oxidation and demetallation was hypothesized to be due to oxidation of the secondary amines. Consequently these were modified to incorporate additional oxygen donor atoms. This modification led to greater resistance to demetallation and ligand oxidation, providing a better platform for further development of copper macrocycles as redox sensors for use in biological systems.
Keywords: Copper macrocycles; Oxidation–reduction; Glutathione; Ascorbate; Vitamin E; Oxidative stress

The effects of increasing concentrations of nickel sulfate, NiSO4 (200 and 400 μM) in the growth medium on the content of starch and sugars and activity levels of enzymes involved in starch and sugar metabolism were examined in seedlings of the two Indica rice cvs. Malviya-36 and Pant-12. During a 5–20 day growth period of seedlings in sand cultures, with Ni treatment, no definite pattern of alteration in starch level could be observed in the seedlings. In both roots and shoots of the seedlings Ni treatment led to a significant decrease in activities of starch degrading enzymes α-amylase, β-amylase, whereas starch phosphorylase activity increased. The contents of reducing, non-reducing, and total sugars increased in Ni-treated rice seedlings with a concomitant increase in the activities of sucrose degrading enzymes acid invertase and sucrose synthase. However, the activity of sucrose synthesizing enzyme sucrose phosphate synthase declined. These results suggest that Ni toxicity in rice seedlings causes marked perturbation in metabolism of carbohydrates leading to increased accumulation of soluble sugars. Such perturbation could serve as a limiting factor for growth of rice seedlings in Ni polluted environments and accumulating soluble sugars could serve as compatible solutes in the cells under Ni toxicity conditions.
Keywords: Nickel toxicity; Carbohydrate metabolism; Starch; Sugars; Invertase; Sucrose synthase; Oryza sativa L.

Expression and characterization of recombinant bovine lactoferrin in E. coli by Isui García-Montoya; Jose Salazar-Martínez; Sigifredo Arévalo-Gallegos; Sugey Sinagawa-García; Quintin Rascón-Cruz (113-122).
Lactoferrin is a member of the transferrin family of iron-binding proteins with a number of properties, including antibacterial activity against a broad spectrum of Gram-negative and Gram-positive bacteria. bovine lactoferrin cDNA was isolated, cloned and expressed as a fusion protein. The amino acid sequence of the fusion was analyzed and compared with other species. Crystallographic data were used to compare structural differences between bovine and human lactoferrin in 3-D models. A thioredoxin fusion protein was expressed and shown to have a different molecular weight compared with native bLf. After purification using Ni-NTA, the yield of recombinant bovine lactoferrin was 15.3 mg/l with a purity of 90.3 %. Recombinant bLf and pepsin-digested rbLf peptides demonstrated antibacterial activity of 79.8 and 86.9 %, respectively. The successful expression of functional, active and intact rbLf allows us to study the biochemical interactions of antimicrobial proteins and peptides and will facilitate their study as immunomodulators.
Keywords: Bovine lactoferrin; Antibacterial activity; Thrombin; Fusion expression; Affinity chromatography

The interactions of toxic metals with essential metals may result in disturbances in the homeostasis of essential elements. However, there are few reports about toxic effect of arsenic (As) on the levels of essential trace elements in the central nervous system. To investigate whether subchronic exposure to As disturbs levels of main essential trace elements in the brain of mice and whether the gender difference in the response to As are altered, the concentrations of As, Iron (Fe), copper (Cu), selenium (Se), zinc (Zn) and Chromium (Cr) in the cerebrum and cerebellum of mice exposed to As subchronically were examined by inductively coupled plasma-mass spectrometry (ICP-MS). The gender difference in the changed levels of these essential trace elements was also statistically analyzed. The concentration of As was significantly higher in the cerebrum or cerebellum of mice exposed to As than that in control group (P < 0.05). It indicates that As can accumulate in brain of mice after subchronic exposure. The concentrations of Fe, Se and Cr in the cerebrum or cerebellum were significantly lower in mice exposed to As than those in control group (P < 0.05). On the contrary, the concentration of Cu in the cerebrum or cerebellum was significantly higher in mice exposed to As (P < 0.05). Our results indicate that subchronic exposure to As may decrease the levels of Fe, Se and Cr or increase the level of Cu in the brain of mice. Moreover, the significant gender difference was found relative to the effect of As on concentration of Se in cerebrum and concentrations of Cu and Se in cerebellum of mice. Therefore, more experiments are required to further understand mechanisms whereby As interacts with essential elements in brain and induces the gender difference.
Keywords: Subchronic exposure to arsenic; Essential element; Brain; Gender difference

Dietary zinc depletion and repletion affects plasma proteins: an analysis of the plasma proteome by Arthur Grider; Kathie Wickwire; Emily Ho; Carolyn S. Chung; Janet King (133-140).
Zinc (Zn) deficiency is a problem world-wide. Current methods for assessing Zn status are limited to measuring plasma or serum Zn within populations suspected of deficiency. Despite the high prevalence of Zn deficiency in the human population there are no methods currently available for sensitively assessing Zn status among individuals. The purpose of this research was to utilize a proteomic approach using two-dimensional gel electrophoresis (2DE) and mass spectrometry to identify protein biomarkers that were sensitive to changes in dietary Zn levels in humans. Proteomic analysis was performed in human plasma samples (n = 6) obtained from healthy adult male subjects that completed a dietary Zn depletion/repletion protocol, current dietary zinc intake has a greater effect on fractional zinc absorption than does longer term zinc consumption in healthy adult men. Chung et al. (Am J Clin Nutr 87 (5):1224–1229, 2008). After a 13 day Zn acclimatization period where subjects consumed a Zn-adequate diet, the male subjects consumed a marginal Zn-depleted diet for 42 days followed by consumption of a Zn-repleted diet for 28 days. The samples at baseline, end of depletion and end of repletion were pre-fractionated through immuno-affinity columns to remove 14 highly abundant proteins, and each fraction separated by 2DE. Following staining by colloidal Coomassie blue and densitometric analysis, three proteins were identified by mass spectrometry as affected by changes in dietary Zn. Fibrin β and chain E, fragment double D were observed in the plasma protein fraction that remained bound to the immunoaffinity column. An unnamed protein that was related to immunoglobulins was observed in the immunodepleted plasma fraction. Fibrin β increased two-fold following the Zn depletion period and decreased to baseline values following the Zn repletion period; this protein may serve as a viable biomarker for Zn status in the future.
Keywords: Fibrin β; Zn biomarker; Marginal Zn deficiency; Dietary Zn depletion and repletion; Two-dimensional gel electrophoresis; Hemostasis

Analytical studies on the incorporation of aluminium in the cell walls of the marine diatom Stephanopyxis turris by Susanne Machill; Lydia Köhler; Susanne Ueberlein; René Hedrich; Marco Kunaschk; Silvia Paasch; Renate Schulze; Eike Brunner (141-150).
The eukaryotic diatoms are unicellular algae. They are well known for their filigree micro- and nanostructured cell walls which mainly consist of amorphous silica as well as various organic compounds. However, diatoms are also known to incorporate certain amounts of aluminium into their cell walls. Unexpectedly, enhanced Al concentrations in the Southern Yellow Sea were found to be correlated with a diatom spring bloom. Therefore, we have analyzed the influence of strongly enhanced Al concentrations in the culture medium upon the growth behaviour of the diatom Stephanopyxis turris (S. turris). The uptake and incorporation of Al into the cell walls was monitored. It turned out that S. turris survives aluminium concentrations up to 105.5 μM (2.85 mg/l) in the culture medium. Under the applied conditions, this corresponds to an Al/Si ratio of 1:1. These large amounts of Al had to be offered in the form of bis–tris-chelates in order to prevent uncontrolled precipitation. Under these conditions, the Al/Si ratio in the cell walls could be increased up to about 1:15 as determined by ICP-OES, the highest amount of aluminium found in diatom cell walls yet. Structural characterization of the biosilica by ATR-FTIR and solid-state 27Al NMR spectroscopy revealed that an amorphous aluminosilicate phase is formed where the aluminium exists as four- and sixfold-coordinated species.
Keywords: Diatom biosilica; Aluminium incorporation; ICP-OES; Solid-state NMR; FTIR

Gallium(III) complexes with 2-acetylpyridine-derived thiosemicarbazones: antimicrobial and cytotoxic effects and investigation on the interactions with tubulin by Josane A. Lessa; Marcella A. Soares; Raquel G. dos Santos; Isolda C. Mendes; Lívia B. Salum; Hikmat N. Daghestani; Adriano D. Andricopulo; Billy W. Day; Andreas Vogt; Heloisa Beraldo (151-165).
Complexes [Ga(2Ac4pFPh)2]NO3 (1), [Ga(2Ac4pClPh)2]NO3 (2), [Ga(2Ac4pIPh)2]NO3 (3), [Ga(2Ac4pNO2Ph)2]NO3·3H2O (4) and [Ga(2Ac4pT)2]NO3 (5) were obtained with 2-acetylpyridine N(4)-para-fluorophenyl-(H2Ac4pFPh), 2-acetylpyridine N(4)-para-chlorophenyl-(H2Ac4pClPh), 2-acetylpyridine N(4)-para-iodophenyl-(H2Ac4pIPh), 2-acetylpyridine N(4)-para-nitrophenyl-(H2Ac4pNO2Ph) and 2-acetylpyridine N(4)-para-tolyl-(H2Ac4pT) thiosemicarbazone. 1–5 presented antimicrobial and cytotoxic properties. Coordination to gallium(III) proved to be an effective strategy for activity improvement against Pseudomonas aeruginosa and Candida albicans. The complexes were highly cytotoxic against malignant glioblastoma and breast cancer cells at nanomolar concentrations. The compounds induced morphological changes characteristic of apoptotic death in tumor cells and showed no toxicity against erythrocytes. 2 partially inhibited tubulin assembly at high concentrations and induced cellular microtubule disorganization, but this does not appear to be the main mechanism of cytotoxic activity.
Keywords: Thiosemicarbazones; Gallium(III) complexes; Antimicrobial activity; Cytotoxicity; Tubulin

Application of Zinpyr-1 for the investigation of zinc signals in Escherichia coli by Hajo Haase; Silke Hebel; Gabriela Engelhardt; Lothar Rink (167-177).
Changes of the pico- to nanomolar concentration of free intracellular Zn2+ are part of the signal transduction in mammalian cells. These zinc signals regulate the enzymatic activity of target proteins such as protein tyrosine phosphatases. For Escherichia coli, previous studies have reported diverging concentrations from femto- to picomolar, raising the question if Zn2+ could also have a function in bacterial signaling. This manuscript explores the use of the low molecular weight fluorescent probe Zinpyr-1 in E. coli. The probe detects free Zn2+ in these bacteria. Comparable to mammalian cells, other metal ions, especially Hg2+ and Cd2+, interfere with the detection of Zn2+. Moreover, experiments in E. coli were particularly prone to artifacts based on cellular autofluorescence, necessitating corrections that are not required in mammalian cells. Based on measurements in lysates of E. coli and the mammalian cell line Jurkat, similar values between 0.1 and 0.2 nM free Zn2+ were found. For E. coli, this corresponds to less than one free zinc ion per cell. Moreover, phosphatase inhibition by Zn2+ was only observed in Jurkat, but not E. coli. This excludes a function for zinc signals as a regulator of bacterial phosphatases. Still, changes in the free Zn2+ concentration were observed in response to elevated extracellular Zn2+ and pH, or to addition of the detergent NP-40, suggesting that other processes could be controlled by the free intracellular Zn2+ concentration.
Keywords: Zinc; E. coli ; Zinpyr-1; Signal transduction

Altered transition metal homeostasis in mice following manganese injections for manganese-enhanced magnetic resonance imaging by Nataliya Moldovan; Alia Al-Ebraheem; Nelson A. Miksys; Michael J. Farquharson; Nicholas A. Bock (179-187).
In manganese-enhanced magnetic resonance imaging (MEMRI), the paramagnetic divalent ion of manganese (Mn2+) is injected into animals to generate tissue contrast, typically at much higher exposures than have been previously used in studies of Mn toxicity. Here we investigate the effect of these injections on the homeostasis of the transition metals iron and copper in mice to see if there are disruptions which should be considered in MEMRI studies. Manganese shares transport proteins with other transition metals including iron and copper, so it is possible that changes in manganese levels in tissue following injections of the metal may affect other metal levels too. This in turn may affect MRI contrast or the investigation of disease processes in the animal models being imaged. In this study, we measured manganese, iron, and copper concentrations in the blood, kidney, liver and in brain regions in mice treated with four injections of 30 mg/kg MnCl2 4H2O (dry chemical weight/body weight)—a common dose used in MEMRI. In addition to the expected increases in manganese in tissues, we noted a statistically significant reduction in copper in the kidney and liver. Also, we noted a statistically significant decrease in concentration of iron in the thalamus of the brain. These findings suggest that the high doses of manganese injected in MEMRI studies can disrupt the homeostasis of other transition metals in mice.
Keywords: Manganese enhanced magnetic resonance imaging (MEMRI); Mouse brain; Iron; Copper; X-ray fluorescence (XRF); Neutron activation analysis (NAA)

The effect of temperature on antibacterial activity of biosynthesized silver nanoparticles by Parastoo Pourali; Majid Baserisalehi; Sima Afsharnezhad; Javad Behravan; Rashin Ganjali; Nima Bahador; Sepideh Arabzadeh (189-196).
The purpose of this study was the evaluation of two different temperatures on antibacterial activity of the biosynthesized silver nanoparticles. 38 silver nanoparticles-producing bacteria were isolated from soil and identified. Biosynthesis of silver nanoparticles by these bacteria was verified through visible light spectrophotometry. Two strains were relatively active for production of silver nanoparticles. These strains were subjected for molecular identification and recognized as Bacillus sp. and Acinetobacter schindleri. In the present study, the effect of temperatures was evaluated on structure and antimicrobial properties of the silver nanoparrticles by transmission electron microscopy (TEM), X-ray diffraction (XRD) analysis and antimicrobial Agar well diffusion methods. The silver nanoparticles showed antibacterial activity against all the pathogenic bacteria; however, this property was lost after treatment of the silver nanoparticles by high temperatures (100 and 300 °C). TEM images showed that the average sizes of heated silver nanoparticles were >100 nm. However, these were <100 nm for non-heated silver nanoparticles. Although, XRD patterns showed the crystalline structure of heated silver nanoparticles, their antibacterial activities were less. This was possible because of the sizes and accordingly less penetration of the particles into the bacterial cells. In addition, elimination of the capping agents by heat might be considered another reason.
Keywords: Biosynthesized; Silver nanoparticles; Temperature; Antibacterial activity