Analytical Tool for Determination of traces of Cu (II)

Heavy metals are widely existent in the contaminated environments. Copper is an essential metal for plants, microorganisms, animals and human beings to perform specific biological functions. As a toxicant at elevated levels of biologically available form, it produces a physiological response. Hence there is a need for rapid and sensitive methods for the analytical determination of copper. The aim of this article is to propose a rapid, selective and sensitive method for the determination of trace amounts copper (II). We aim to develop paptodes based on RGB analysis for copper determination and removal. A new optical analytical method, “digital RGB Analysis” is proposed instead of the conventional optical method, “spectrophotometry”. MATLAB image processing tool can transform the color information into digital RGB values that can be treated as analytical information. The Paper optode has been prepared by immobilizing resorcinol and oxalic acid 1:1 solution on chromatographic (TLC) strip and heating for 15min at 80-900C. The obtained color pattern was analyzed using image processing tool of MATLAB software to determine copper (II). All parameters affecting intensity on optode have been optimized. The proposed sensor was linear in the range 0.012-8.4µg mL-1 {12 µL of 1-700 µg mL-1). The minimum detection limit was found 15ng mL-1.The proportionality in intensity of the spot color on the optodes loaded with varying amounts of copper suggests its potential applications for environmental monitoring. The paptode can also be used for pollutant check at home. Thus the paper optode has great potential for this purpose. Key Words: MATLAB, RGB analysis, Heavy metals, optical analytical method, TLC strip

Can biowarfare agents be defeated with light?

Biological warfare and bioterrorism is an unpleasant fact of 21st century life. Highly infectious and profoundly virulent diseases may be caused in combat personnel or in civilian populations by the appropriate dissemination of viruses, bacteria, spores, fungi, or toxins. Dissemination may be airborne, waterborne, or by contamination of food or surfaces. Countermeasures may be directed toward destroying or neutralizing the agents outside the body before infection has taken place, by destroying the agents once they have entered the body before the disease has fully developed, or by immunizing susceptible populations against the effects. A range of light-based technologies may have a role to play in biodefense countermeasures. Germicidal UV (UVC) is exceptionally active in destroying a wide range of viruses and microbial cells, and recent data suggests that UVC has high selectivity over host mammalian cells and tissues. Two UVA mediated approaches may also have roles to play; one where UVA is combined with titanium dioxide nanoparticles in a process called photocatalysis, and a second where UVA is combined with psoralens (PUVA) to produce “killed but metabolically active” microbial cells that may be particularly suitable for vaccines. Many microbial cells are surprisingly sensitive to blue light alone, and blue light can effectively destroy bacteria, fungi, and Bacillus spores and can treat wound infections. The combination of photosensitizing dyes such as porphyrins or phenothiaziniums and red light is called photodynamic therapy (PDT) or photoinactivation, and this approach cannot only kill bacteria, spores, and fungi, but also inactivate viruses and toxins. Many reports have highlighted the ability of PDT to treat infections and stimulate the host immune system. Finally pulsed (femtosecond) high power lasers have been used to inactivate pathogens with some degree of selectivity. We have pointed to some of the ways light-based technology may be used to defeat biological warfare in the future

Can nanotechnology potentiate photodynamic therapy?

Photodynamic therapy (PDT) uses the combination of nontoxic dyes and harmless visible light to produce reactive oxygen species that can kill cancer cells and infectious microorganisms. Due to the tendency of most photosensitizers (PS) to be poorly soluble and to form nonphotoactive aggregates, drug-delivery vehicles have become of high importance. The nanotechnology revolution has provided many examples of nanoscale drug-delivery platforms that have been applied to PDT. These include liposomes, lipoplexes, nanoemulsions, micelles, polymer nanoparticles (degradable and nondegradable), and silica nanoparticles. In some cases (fullerenes and quantum dots), the actual nanoparticle itself is the PS. Targeting ligands such as antibodies and peptides can be used to increase specificity. Gold and silver nanoparticles can provide plasmonic enhancement of PDT. Two-photon excitation or optical upconversion can be used instead of one-photon excitation to increase tissue penetration at longer wavelengths. Finally, after sections on in vivo studies and nanotoxicology, we attempt to answer the title question, “can nanotechnology potentiate PDT?”National Institutes of Health (U.S.) (RO1 AI050875)United States. Air Force (Medical Free Electron Laser Program (FA9550-04-1-0079)

Synthesis and Photodynamic Effect of New Highly Photostable Decacationically Armed [60]- and [70]Fullerene Decaiodide Monoadducts To Target Pathogenic Bacteria and Cancer Cells

Novel water-soluble decacationically armed C-60 and C-70 decaiodide monoadducts, C-60- and C-70[>M(C3N6+C3)(2)], were synthesized, characterized, and applied as photosensitizers and potential nano-PDT agents against pathogenic bacteria and cancer cells. A high number of cationic charges per fullerene cage and H-bonding moieties were designed for rapid binding to the anionic residues displayed on the outer parts of bacterial cell walls. In the presence of a high number of electron-donating iodide anions as parts of quaternary ammonium salts in the arm region, we found that C-70[>M(C3N6+C3)(2)] produced more HO center dot than C-60[>M(C3N6+C3)(2)], in addition to O-1(2). This finding offers an explanation of the preferential killing of Gram-positive and Gram-negative bacteria by C-60[>M(C3N6+C3)(2)] and C-70[>M(C3N6+C3)(2)], respectively. The hypothesis is that O-1(2) can diffuse more easily into porous cell walls of Gram-positive bacteria to reach sensitive sites, while the less permeable Gram-negative bacterial cell wall needs the more reactive HO center dot to cause real damage.National Institutes of Health (NIH) [1R01CA137108]National Institutes of Health (NIH

Low-Level Laser Therapy Activates NF-kB via Generation of Reactive Oxygen Species in Mouse Embryonic Fibroblasts

Background Despite over forty years of investigation on low-level light therapy (LLLT), the fundamental mechanisms underlying photobiomodulation at a cellular level remain unclear. Methodology/Principal Findings In this study, we isolated murine embryonic fibroblasts (MEF) from transgenic NF-kB luciferase reporter mice and studied their response to 810 nm laser radiation. Significant activation of NF-kB was observed at fluences higher than 0.003 J/cm2 and was confirmed by Western blot analysis. NF-kB was activated earlier (1 hour) by LLLT compared to conventional lipopolysaccharide treatment. We also observed that LLLT induced intracellular reactive oxygen species (ROS) production similar to mitochondrial inhibitors, such as antimycin A, rotenone and paraquat. Furthermore, we observed similar NF-kB activation with these mitochondrial inhibitors. These results, together with inhibition of laser induced NF-kB activation by antioxidants, suggests that ROS play an important role in the laser induced NF-kB signaling pathways. However, LLLT, unlike mitochondrial inhibitors, induced increased cellular ATP levels, which indicates that LLLT also upregulates mitochondrial respiration. Conclusion We conclude that LLLT not only enhances mitochondrial respiration, but also activates the redox-sensitive NFkB signaling via generation of ROS. Expression of anti-apoptosis and pro-survival genes responsive to NFkB could explain many clinical effects of LLLT.National Institutes of Health (U.S.) (grant R01AI050875)Center for Integration of Medicine and Innovative Technology (DAMD17-02-2-0006)United States. Dept. of Defense (CDMRP Program in TBI, W81XWH-09-1-0514)United States. Air Force Office of Scientific Research (FA9950-04-1-0079

Comparison of Therapeutic Effects between Pulsed and Continuous Wave 810-nm Wavelength Laser Irradiation for Traumatic Brain Injury in Mice

Background and Objective Transcranial low-level laser therapy (LLLT) using near-infrared light can efficiently penetrate through the scalp and skull and could allow non-invasive treatment for traumatic brain injury (TBI). In the present study, we compared the therapeutic effect using 810-nm wavelength laser light in continuous and pulsed wave modes in a mouse model of TBI. Study Design/Materials and Methods TBI was induced by a controlled cortical-impact device and 4-hours post-TBI 1-group received a sham treatment and 3-groups received a single exposure to transcranial LLLT, either continuous wave or pulsed at 10-Hz or 100-Hz with a 50% duty cycle. An 810-nm Ga-Al-As diode laser delivered a spot with diameter of 1-cm onto the injured head with a power density of 50-mW/cm2 for 12-minutes giving a fluence of 36-J/cm2. Neurological severity score (NSS) and body weight were measured up to 4 weeks. Mice were sacrificed at 2, 15 and 28 days post-TBI and the lesion size was histologically analyzed. The quantity of ATP production in the brain tissue was determined immediately after laser irradiation. We examined the role of LLLT on the psychological state of the mice at 1 day and 4 weeks after TBI using tail suspension test and forced swim test. Results The 810-nm laser pulsed at 10-Hz was the most effective judged by improvement in NSS and body weight although the other laser regimens were also effective. The brain lesion volume of mice treated with 10-Hz pulsed-laser irradiation was significantly lower than control group at 15-days and 4-weeks post-TBI. Moreover, we found an antidepressant effect of LLLT at 4-weeks as shown by forced swim and tail suspension tests. Conclusion The therapeutic effect of LLLT for TBI with an 810-nm laser was more effective at 10-Hz pulse frequency than at CW and 100-Hz. This finding may provide a new insight into biological mechanisms of LLLT.National Institutes of Health (U.S.) (NIH grant R01AI050875)Center for Integration of Medicine and Innovative Technology (DAMD17-02-2-0006)United States. Dept. of Defense. Congressionally Directed Medical Research Programs (W81XWH-09-1-0514)United States. Air Force Office of Scientific Research (Military Photomedicine Program (FA9950-04-1-0079))Japan. Ministry of Education, Culture, Sports, Science and TechnologyJapan Society for the Promotion of Scienc

Evaluation of chlorin p6 for photodynamic treatment of squamous cell carcinoma in the hamster cheek pouch model

We studied pharmacokinetics and tumor response to photodynamic therapy (PDT) using chlorin p6 (CP6) in hamster cheek pouch model. CP6 was administered either intraperitoneally (IP) at a dose of 1.5mg/kg body weight or applied topically at 1.0mg/kg body weight and its accumulation in tumor, normal mucosa, and abdominal skin was measured by optical fiber-based fluorescence spectroscopy. Photodynamic therapy was performed by superficial illumination of tumor with 660nm (±25nm) light at a fluence rate of 100J/cm2 and tumor response to PDT was analyzed by histological examination. CP6 accumulation was higher in tumors as compared to adjoining tissue and normal mucosa at 4-6h after its IP administration. For relatively large tumors (size >8mm) topical application was observed to be more effective than IP. The level of CP6 in tumor, surrounding tissue, normal mucosa and skin was seen to decrease rapidly within 24h after its administration and was undetectable at longer time (>72h) intervals. PDT of small tumors at 4h after IP injection of CP6 resulted in complete tumor necrosis. Whereas, PDT of large tumors receiving CP6 topically caused necrosis in 300-800 μm superficial region of the tumor. In one animal kept for follow up in each treatment group, it was observed that small tumors disappeared completely leaving scar tissue, while large tumor had significant reduction in tumor size. The use of CP6 for PDT of oral cancer is suggested

Pharmacokinetics and phototoxicity of purpurin-18 in human colon carcinoma cells using liposomes as delivery vehicles

Pharmacokinetics and phototoxicity of purpurin-18 (Pp18) in human colon carcinoma cells (Colo-205) was studied using liposomes as delivery vehicles. Cytotoxicity was measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and neutral red uptake assay, and mode of cell death was assessed by the study of cell morphology and nuclear staining with Hoechst 33342-propidium iodide. Pp18 solubilized in dimethyl sulfoxide saline solution was observed to aggregate (Q-band absorption 740 nm), resulting in very poor cellular uptake. Pp18 incorporated in liposome remained in monomeric form (Q-band absorption 695 nm), but due to the presence of an anhydride ring in the molecule it readily yielded another photosensitizer, chlorin p6 (Q-band absorption 662 nm). Measurements at various pH showed that Pp18 in liposome was stable at acidic pH (6.5). Incubation of cells with 6.0 μM Pp18 in liposome at pH 6.5 showed a rapid cellular uptake. Spectrofluorometric measurements showed the presence of both Pp18 and chlorin p6, indicating conversion of some amount of Pp18 into chlorin p6 in the cells. Fluorescence microscopy revealed that the fluorescence was localized mainly in the cytoplasm, sparing the nucleus. Illumination of cells to white light after 4-h incubation with Pp18 liposome preparation was observed to lead to dose-dependent decrease in cell viability. At low irradiation time, cells displayed formation of plasma membrane blebs and micronuclei typical of apoptotic cell death. In contrast, at higher irradiation time, cell swelling and vacuolization in nucleus was observed, suggesting cell death due to necrosis. Irradiation with narrow bandwidth light showed that at low pH, the relative phototoxicity due to pp18 was higher than that due to chlorin p6. It is suggested that the pH-dependent conversion of pp18 to chlorin p6 can be exploited to increase PDT selectivity

Biphasic Dose Response in Low Level Light Therapy – An Update

Low-level laser (light) therapy (LLLT) has been known since 1967 but still remains controversial due to incomplete understanding of the basic mechanisms and the selection of inappropriate dosimetric parameters that led to negative studies. The biphasic dose-response or Arndt-Schulz curve in LLLT has been shown both in vitro studies and in animal experiments. This review will provide an update to our previous (Huang et al. 2009) coverage of this topic. In vitro mediators of LLLT such as adenosine triphosphate (ATP) and mitochondrial membrane potential show biphasic patterns, while others such as mitochondrial reactive oxygen species show a triphasic dose-response with two distinct peaks. The Janus nature of reactive oxygen species (ROS) that may act as a beneficial signaling molecule at low concentrations and a harmful cytotoxic agent at high concentrations, may partly explain the observed responses in vivo. Transcranial LLLT for traumatic brain injury (TBI) in mice shows a distinct biphasic pattern with peaks in beneficial neurological effects observed when the number of treatments is varied, and when the energy density of an individual treatment is varied. Further understanding of the extent to which biphasic dose responses apply in LLLT will be necessary to optimize clinical treatments

Can dendritic cells see light?

There are many reports showing that low-level light/laser therapy (LLLT) can enhance wound healing, upregulate cell proliferation and has anti-apoptotic effects by activating intracellular protective genes. In the field of immune response study, it is not known with any certainty whether light/laser is proinflammatory or anti-inflammatory. Increasingly in recent times dendritic cells have been found to play an important role in inflammation and the immunological response. In this study, we try to look at the impact of low level near infrared light (810-nm) on murine bone-marrow derived dendritic cells. Changes in surface markers, including MHC II, CD80 and CD11c and the secretion of interleukins induced by light may provide additional evidence to reveal the mystery of how light affects the maturation of dendritic cells as well how these light-induced mature dendritic cells would affect the activation of adaptive immune response.United States National Institutes of Health (Grant R01AI050875)Center for Integration of Medicine and Innovative Technology (DAMD17-02-2-0006)United States Department of Defense. Office of the Congressionally Directed Medical Research Programs (Program in TBI (W81XWH-09-1-0514)