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Investigation into the dominant strains of Clostridium difficile within hospitals and strategic cleaning regimesPatel, Krusha January 2013 (has links)
Clostridium difficile is a common and potentially fatal cause of antibiotic-associated diarrhoea and pseudomembranous colitis worldwide. It has been isolated from patients and their surroundings, in healthcare facilities and from the community. C. difficile is able to survive for many months on inanimate surfaces in the form of spores. PCR ribotyping is used in the UK to characterise and identify strain diversity. Investigating how the most problematic strains respond to cleaning regimes may influence the control of disease. This work used the University Hospitals of Leicester Trust as a case study for this purpose of understanding the epidemiology of this pathogen within healthcare facilities. Five individual agar media were compared based on their abilities to recover and resuscitate damaged ribotype 027 spores, a strain associated with disease outbreaks and increased severity. Controlled laboratory experiments with a sub-lethal dose of a germicide were conducted before C. difficile recovery from hospital wards. An additional two sampling campaigns acquired environmental strains. C. difficile isolation after routine cleaning demonstrated the inefficiency of the current recovery regime as C. difficile spores were recovered using direct contact plates, enrichment broths, and resuscitation media. This study used layering of non-selective agar over selective agar, identifying a potential link in the proportions of media following the use of sponges in environmental sampling. All strains were characterised by ribotyping; ribotype 027 was isolated from all sampling cohorts. A four-month epidemiological study was conducted into the ribotype prevalence and distribution from C. difficile-positive faecal specimens. A second survey investigated these effects with a modification of C. difficile detection from faecal samples. Hydrogen peroxide vapour is currently being explored as a means of decontamination of healthcare-associated infections. Inactivation kinetics of ribotype 027 spores were analysed in response to vapour and liquid exposure of hydrogen peroxide. No reports thus far have explored such kinetics and controlled decontamination with both clinical and non-clinical strains. Evidence strongly suggests spores can be inactivated with its application. Furthermore, this study revealed there appears to be significant differences in susceptibility and inactivation of different C. difficile ribotypes.
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Διερεύνηση των μηχανισμών ρύθμισης της έκφρασης του αυξητικού παράγοντα HARP από το υπεροξείδιο του υδρογόνου / Regulation of HARP gene expression by hydrogen peroxideΠολυτάρχου, Χρήστος 10 June 2007 (has links)
Τα τελευταία χρόνια μια πληθώρα ερευνητικών εργασιών συσχετίζει την παραγωγή των δραστικών μορφών οξυγόνου (ROS) με ποικίλες κυτταρικές λειτουργίες. Στο επίπεδο του κυττάρου, η παραγωγή ROS δύναται να προκληθεί τόσο υπό την επίδραση φυσιολογικών ερεθισμάτων (όπως ορμόνες και κυτταροκίνες) όσο και από εξωγενείς παράγοντες (όπως μικροοργανισμοί και ακτινοβολία). Η αυξημένη παραγωγή ROS ενέχεται στην παθογένεση διαφόρων ασθενειών, όπως του καρκίνου, της αθηροσκλήρωσης και της ρευματοειδούς αρθρίτιδας. Αν και σε υψηλές συγκεντρώσεις οι ROS καθίστανται κυτταροτοξικές, σε χαμηλά επίπεδα φαίνεται να δρουν ως ειδικά μόρια που συμμετέχουν στη μεταγωγή σήματος. Ανάμεσα στις ROS, ιδιαίτερο ενδιαφέρον προσελκύει το Η2Ο2, το οποίο λόγω της σχετικής σταθερότητας, της έλλειψης πολικότητας και του μικρού μοριακού μεγέθους, είναι ιδανικός υποψήφιος μεσολαβητής στη μεταγωγή σήματος στο εσωτερικό των κυττάρων. Η heparin affin regulatory peptide (HARP) είναι αυξητικός παράγοντας με σημαντικό ρόλο στην ανάπτυξη του καρκίνου και στην αγγειογένεση. Στην παρούσα εργασία μελετήθηκε η επίδραση του Η2Ο2 στον πολλαπλασιασμό και τη μετανάστευση των ανθρώπινων καρκινικών κυττάρων προστάτη LNCaP και των ενδοθηλιακών κυττάρων από φλέβα ομφάλιου λώρου (HUVEC), καθώς και στην αγγειογένεση in vivo στο μοντέλο της χοριοαλλαντοϊκής μεμβράνης (CAM) εμβρύου όρνιθας. Μελετήθηκε επίσης η πιθανή εμπλοκή της HARP στη δράση του Η2Ο2. Βρέθηκε ότι εξωγενώς χορηγούμενο Η2Ο2 επάγει τον πολλαπλασιασμό και τη μετανάστευση των κυττάρων HUVEC και την αγγειογένεση in vivo στην CAM εμβρύου όρνιθας. Επιπρόσθετα, το Η2Ο2 αύξησε τα επίπεδα της πρωτεΐνης HARP in vitro και in vivo. Το U0126 και η κουρκουμίνη ανέστειλαν τόσο την αύξηση των επιπέδων της πρωτεΐνης HARP, όσο και τον πολλαπλασιασμό των HUVEC που επάγονται από το Η2Ο2. Σε συμφωνία με τα παραπάνω, βρέθηκε ότι το Η2Ο2 επάγει τον πολλαπλασιασμό και τη μετανάστευση των κυττάρων LNCaP. Επιπλέον, προκάλεσε αύξηση των επιπέδων του mRNA και της πρωτεΐνης HARP, με τρόπο εξαρτώμενο από τη συγκέντρωση και το χρόνο. Η HARP φαίνεται να μεσολαβεί στις δράσεις του Η2Ο2, καθώς σε κύτταρα LNCaP στα οποία έχει κατασταλεί η έκφρασή της δεν εκδηλώθηκαν οι διεγερτικές δράσεις του Η2Ο2. Η κουρκουμίνη και ολιγονουκλεοτίδια-δολώματα έναντι του AP-1 ανέστειλαν τόσο την έκφραση της HARP, όσο και τον πολλαπλασιασμό και τη μετανάστευση των κυττάρων LNCaP που επάγονται από το Η2Ο2. Μετά από εισαγωγή της ρυθμιστικής περιοχής του γονιδίου της HARP σε πλασμιδιακό φορέα ανοδικά γονιδίου αναφοράς, το Η2Ο2 αύξησε με χρόνο-εξαρτώμενο τρόπο τη δραστηριότητα του υποκινητή του συγκεκριμένου γονιδίου. Η επαγωγική αυτή δράση ανεστάλη όταν εισήχθησαν σημειακές μεταλλαγές σε τουλάχιστον μία από τις δύο πιθανές αλληλουχίες δέσμευσης του AP-1. Η επίδραση του Η2Ο2 φαίνεται να οφείλεται στη δέσμευση των Fra-1, JunD και της ενεργού μορφής της c-Jun στη ρυθμιστική περιοχή του γονιδίου της HARP. Συμπερασματικά, στην παρούσα εργασία ενισχύεται η άποψη ότι η HARP διαδραματίζει σημαντικό ρόλο στον πολλαπλασιασμό και τη μετανάστευση των κυττάρων. Επίσης, προτείνεται ότι το γονίδιο της HARP παρουσιάζει ευαισθησία σε μεταβολές της οξειδωτικής κατάστασης του κυττάρου, καθώς η έκφρασή της επάγεται σε χαμηλές συγκεντρώσεις Η2Ο2. Τέλος, αναδεικνύεται η εμπλοκή συγκεκριμένων συμπλόκων του AP-1 στη ρύθμιση της μεταγραφικής δραστηριότητας του γονιδίου της HARP και των επακόλουθων βιολογικών δράσεων σε απόκριση στο Η2Ο2. / A growing body of evidence correlates the production of Reactive Oxygen Species (ROS) with many aspects of cell functions. Cellular ROS production can be augmented by the action of hormones, cytokines, and other physiological stimuli and also by external factors, such as xenobiotics or irradiation. Increased ROS production has been implicated in the pathogenesis of several diseases, such as cancer, atherosclerosis, rheumatoid arthritis and ischemia/reperfusion injury. Although ROS may be cytotoxic when produced in excess, at moderate levels they may act as specific signalling molecules. Among different ROS, H2O2 possesses the properties of relative stability, neutrality and small size, which make it an ideal candidate for a signalling molecule. Heparin affin regulatory peptide (HARP) is a mitogenic and angiogenic growth factor and plays a significant role on tumor growth and angiogenesis. In the present study we found that exogenous H2O2 significantly induced human umbilical vein endothelial cell (HUVEC) proliferation and migration, and angiogenesis in the in vivo chorioallantoic membrane assay. In addition, it increased HARP protein amounts in vitro and in vivo. U0126 and curcumin abrogated both H2O2-induced HARP protein amounts increment and HUVEC proliferation. In the same line, exogenous H2O2 significantly induced human prostate cancer LNCaP cell proliferation and migration. Moreover, H2O2 significantly increased HARP mRNA and protein amounts in a concentration- and time-dependent manner. HARP seems to be involved in the stimulatory effect of H2O2, since the latter had no effect on stably transfected LNCaP cells that did not express HARP. Curcumin and AP-1 decoy oligonucleotides abrogated both H2O2-induced HARP expression and LNCaP cell proliferation and migration. H2O2 increased luciferase activity of the 5’-flanking region of the HARP gene introduced in a reporter gene vector, an effect that was abolished when even one of the two putative AP-1 binding sites of the HARP promoter was mutated. The effect of H2O2 seems to be due to binding of Fra-1, JunD and phospho-c-Jun to the HARP promoter. In conclusion, H2O2 up-regulates HARP expression through the induction of specific AP-1 dimers, leading to increased cell proliferation and migration.
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Study of electrocatalytic processes at Prussian blue modified glassy carbon electrode / Elektrokatalizinių procesų tyrimas ant Berlyno mėlynuoju modifikuoto stiklo anglies elektrodoAraminaitė, Rūta 13 February 2010 (has links)
The main purpose of this work is study of electrochemical hydrogen peroxide and ascorbate reactions on electrodes modified by Prussian blue (PB), with the aim to apply these electrodes in creation of sensors and biosensors. For this purpose, a detailed study of electrochemical reduction of hydrogen peroxide, as well as of oxidation of ascorbate at Prussian blue modified rotating disk electrode. In view of the results obtained, a mechanism for hydrogen peroxide reduction at PB modified electrode has been proposed. In accordance with this mechanism, electron transfer appears to be rate-limiting step. The kinetics of decomposition of PB modified electrode in the course of a cathodic reduction of hydrogen peroxide has been studied, and the influence of different factors to this process has been determined. Prototypes of sensors and biosensors, for different analytes have been elaborated and tested. / Darbo tikslas yra elektrocheminių vandenilio peroksido ir askorbato reakcijų tyrimas ant Berlyno mėlynuoju (BM) modifikuotų elektrodų, siekiant pritaikyti šiuos elektrodus jutiklių ir biojutiklių kūrimui. Ištirta vandenilio peroksido redukciją ir askorbato oksidaciją naudojant sukamojo disko elektrodą. Gauti rezultatai galimai įrodo stadijinį vandenilio peroksido katodinės redukcijos mechanizmą vykstantį ant BM modifikuoto elektrodo. Detaliai ištirta BM sluoksnio irimo kinetika vandenilio peroksido elektroredukcijos metu, ir nustatyti faktoriai, įtakojantys irimo proceso greitį. Sukurti jutiklių ir biojutiklių prototipai, kurie galėtų būti panaudoti biologiškai aktyvių medžiagų (vandenilio peroksido, askorbato, gliukozės) nustatymui.
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Elektrokatalizinių procesų tyrimas ant Berlyno mėlynuoju modifikuoto stiklo anglies elektrodo / Study of electrocatalytic processes at Prussian blue modified glassy carbon electrodeAraminaitė, Rūta 13 February 2010 (has links)
Darbo tikslas yra elektrocheminių vandenilio peroksido ir askorbato reakcijų tyrimas ant Berlyno mėlynuoju (BM) modifikuotų elektrodų, siekiant pritaikyti šiuos elektrodus jutiklių ir biojutiklių kūrimui. Ištirta vandenilio peroksido redukciją ir askorbato oksidaciją naudojant sukamojo disko elektrodą. Gauti rezultatai galimai įrodo stadijinį vandenilio peroksido katodinės redukcijos mechanizmą vykstantį ant BM modifikuoto elektrodo. Detaliai ištirta BM sluoksnio irimo kinetika vandenilio peroksido elektroredukcijos metu, ir nustatyti faktoriai, įtakojantys irimo proceso greitį. Sukurti jutiklių ir biojutiklių prototipai, kurie galėtų būti panaudoti biologiškai aktyvių medžiagų (vandenilio peroksido, askorbato, gliukozės) nustatymui. / The main purpose of this work is study of electrochemical hydrogen peroxide and ascorbate reactions on electrodes modified by Prussian blue (PB), with the aim to apply these electrodes in creation of sensors and biosensors. For this purpose, a detailed study of electrochemical reduction of hydrogen peroxide, as well as of oxidation of ascorbate at Prussian blue modified rotating disk electrode. In view of the results obtained, a mechanism for hydrogen peroxide reduction at PB modified electrode has been proposed. In accordance with this mechanism, electron transfer appears to be rate-limiting step. The kinetics of decomposition of PB modified electrode in the course of a cathodic reduction of hydrogen peroxide has been studied, and the influence of different factors to this process has been determined. Prototypes of sensors and biosensors, for different analytes have been elaborated and tested.
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Cellular senescence response to hydrogen peroxide at different cell cycle stagesHunt, Andrea 02 September 2011 (has links)
The development of cellular senescence is not consistent in cultured bovine fibroblasts. Numerous factors may be contributing to the variable onset of cellular senescence, including oxidative stress, telomere shortening and DNA damage. Recent results indicate that cellular senescence is also associated with markers of DNA replication. This suggests that cells may be more likely to enter a senescent state depending on their cell cycle progression. The purpose of this study was to determine the effect of cell cycle phase on the development of stress-induced premature cellular senescence (SIPS). Bovine fibroblasts were synchronized at various cell cycle phases, followed by treatment with increasing doses of H2O2. Senescent cells were detected using SA-β-galactosidase staining assay. As H2O2 dosage increased, the amount of cell death by necrosis increased in both unsynchronized and cell cycle synchronized groups, while the amount of senescence varied depending on cell cycle phase. Our results suggest that the S phase of the cell cycle is the most resistant to oxidative damage, the G2/M phase is the most susceptible, and the G1/S phase is the most likely to enter senescence as a protective measure following low doses of H2O2. Increased senescence also resulted from an increased recovery time post-H2O2 treatment, and gene expression studies suggest SIPS bovine fibroblasts senesce via the p53-independent pathway. An improved understanding of SIPS has important implications for improving cloning efficiency and understanding age-related diseases.
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An in vitro study of post-restorative bleaching : effect on microleakageManni, Hassan January 2010 (has links)
<p>Aim and Objectives: To assess the effect of bleaching on the marginal integrity of Class V composite resin restorations. To determine the effect of a 6% hydrogen peroxide over the counter and a 38% hydrogen peroxide in-office vital bleaching treatment products on the microleakage of Class V composite restorations.</p>
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Nanostructured Metal Electrodes for Wool Processing and ElectroanalysisCruickshank, Amy Clare January 2007 (has links)
The research presented in this thesis firstly concerns the use of electrochemical techniques to develop approaches to wool processing which have a lower impact on the environment than conventional chemical methods. Wool is a sulfur rich substrate and current methods used in wool processing often rely on sulfur-based reducing agents such as metabisulfite. However, due to increasing concern over the environmental impacts of metabisulfite, alternative methods are of interest. Electrochemical techniques have been applied to the process of wool setting in the presence of thiol setting agents. Wool disulfide bonds are reduced during this process and the thiol setting agent is converted to the disulfide. Efficient conversion of the disulfide back to the thiol setting agent would allow catalytic amounts of thiols to be used in wool setting. The electroreduction of cystine and 2-hydroxyethyl disulfide has been examined at a range of metal and carbon electrodes to find efficient methods of generating the corresponding thiols, cysteine and 2-mercaptoethanol respectively. Gold and silver were identified as the most efficient electrode materials. In industrial wool processing, the use of large-scale metal electrodes is expensive and therefore, high surface area gold and silver nanoparticle electrodes were fabricated by electrochemically depositing the metals onto low-cost carbon substrates. The most efficient electrochemical system for generating the thiol setting agent involved the electroreduction of cystine at the gold nanoparticle electrode and this system was used to successfully demonstrate that wool setting can be achieved using relatively low concentrations of cysteine. Further research was carried out to investigate methods for the controlled preparation of metal nanoparticle electrodes and their utility for detecting hydrogen peroxide was examined. A simple and versatile approach for the preparation of tethered gold nanoparticle assemblies was developed by exploiting electrostatic interactions between citrate-capped gold nanoparticles and amine tether layers attached to carbon surfaces. The nanoparticle assemblies were optimised for the detection of hydrogen peroxide by selecting the size and density of electrostatically assembled nanoparticles. The number of amine functionalities on the surface and the assembly conditions controlled the nanoparticle density. Nanostructured palladium electrodes fabricated using vapour deposition methods to immobilise palladium nanoparticles directly onto carbon substrates were also examined for the electroanalysis of hydrogen peroxide.
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TARGETING THE METAL CHELATOR D-PENICILLAMINE TO EXPLOIT THE ELEVATED COPPER AND OXIDATIVE STRESS ASSOCIATED WITH CANCERGupte, Anshul 01 January 2008 (has links)
The significantly increased copper and oxidative stress levels are characteristic hallmarks of cancer cells. These differences provide a unique opportunity for selective targeting of cancer cells. D-penicillamine (D-pen) has been proposed to generate reactive oxygen species (ROS) in presence of copper. Therefore, these studies were aimed at investigating the potential application of a currently marketed copper chelator, D-pen, as a novel cytotoxic anti-cancer agent. D-pen was shown to produce ROS, specifically hydrogen peroxide (H2O2), in the presence of cupric sulfate through a copper catalyzed oxidation reaction. During this process D-pen was converted to D-pen disulfide. The experimental proof of the H2O2 generation was conclusively shown with the aid of a novel High Performance Liquid Chromatography (HPLC) assay. The in-vitro cytotoxicity of D-pen co-incubated with cupric sulfate was examined in human beast cancer (MCF-7 and BT474) and leukemia cells (HL-60, HL-60/VCR, and HL-60/ADR). D-pen was shown to cause concentration dependent cytotoxicity in both leukemia and breast cancer cells. A direct correlation between the detection of intracellular ROS and cytotoxicity was established. The treatment of D-pen plus cupric sulfate resulted in a significant reduction in the intracellular thiol content. D-pen is highly hydrophilic and is rapidly eliminated from the body; therefore to improve the intracellular uptake and to protect the thiol group of D-pen, we carried out the synthesis and the in-vitro characterization of a novel gelatin-D-pen conjugate. It was shown that D-pen alone does not enter cells. Confocal microscopy was employed to exhibit the uptake of the novel gelatin-D-pen conjugate by cancer cells. As the cancer cells in-vitro do not accumulate the same levels of copper as reported for cancer cells in-vivo, cancer cells were pre-treated with cupric sulfate to simulate the elevated copper levels. The cupric sulfate pretreatment resulted in reduced thiol level and significantly increased cellular copper content compared to untreated cells. Whereas both free D-pen and gelatin-D-pen conjugate lacked cytotoxicity in un-treated cells, both agents caused concentration dependent cytotoxicity in cupric sulfate pre-treated leukemia cells. Therefore, it was shown that the administration of D-pen as polymer conjugate would potentially provide cytotoxicity and specificity in the treatment of cancer.
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Antibacterial Strategies for Titanium BiomaterialsUnosson, Erik January 2015 (has links)
Titanium and titanium based alloys are widely used in dentistry and orthopedics to replace hard tissue and to mend broken bones. It has become a material of choice due to its low density, high strength, good biocompatibility and its capacity to integrate closely with the bone. Today, modern materials and surgical techniques can enable patients to live longer, and aid in maintaining or regaining mobility for a more fulfilling life. There are, however, instances where implants fail, and one of the primary causes for implant failure is infection. This thesis deals with two possible ways of reducing or eliminating implant associated infections; TiO2 photocatalysis, where a surface can become antibacterial upon irradiation with UV light; and incorporation of silver, where a subsequent release of silver metal ions result in an antibacterial effect. For the TiO2 photocatalysis strategy, a simple and cost effective chemical oxidation technique, using hydrogen peroxide (H2O2) and water, was used to create an active TiO2 surface on titanium substrates. This surface was shown to effectively degrade an organic model substance (rhodamine B) by generating reactive oxygen species (ROS) under UV illumination. However, it was shown that Ti-peroxy radical species remaining in the surface after the H2O2-oxidation process, rather than generation of ROS from a heterogeneous photocatalytic process, was responsible for the effect. This discovery was further exploited in a TiO2/H2O2/UV system, which demonstrated synergy effects in both rhodamine B degradation tests and in antibacterial assays. For the silver ion release strategy, a combinatorial materials science approach was employed. Binary Ag-Ti oxide gradients were co-deposited in a reactive (O2) environment using a custom built physical vapor deposition system, and evaluated for antibacterial properties. The approach enabled synthesis and composition-structure-property evaluation unlikely to have been achieved by traditional means, and the gradient coatings demonstrated antibacterial properties against both S. aureus and S. epidermidis according to silver ion release. The release was shown to depend more on structural features, such as surface area, crystallinity and oxidation state, than on composition. Ag-Ti oxide gradients were also evaluated under UV illumination, as Ag deposits on crystalline TiO2 can enhance photocatalytic properties. In this work, however, the TiO2 was amorphous and UV illumination caused a slight reduction in the antibacterial effect of silver ions. This was attributed to a UV-induced SOS response in the S. epidermidis bacteria. The results of this thesis demonstrate that both TiO2 photocatalysis, or UV induced activation of Ti-peroxy radical species, as well as incorporation of silver are viable antibacterial strategies for titanium biomaterials. However, their clinical applications are still pending risk-benefit analyses of potential adverse host tissue responses.
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Advanced Water Treatment Strategies for the Removal of Natural and Synthetic Organic ContaminantsHalevy, Patrick January 2014 (has links)
Prior to full-scale implementation of process modifications at the Brantford WTP, a pilot-scale treatability study was conducted to investigate intermediate ozonation/AOP and to determine the most suitable granular media (anthracite, GAC, and Filtralite®) for deep-bed biological filtration. The primary objectives of this research were to provide insight into the destruction of natural and synthetic organics and assess ozonated and halogenated DBP formation.
Ozone alone was unable to achieve the 1-log removal target for geosmin or MCPA, unless disinfection-level dosages were applied. No improvement was observed when adding hydrogen peroxide.
A major obstacle to the implementation of ozonation in bromide-laden source waters is the formation of bromate. There is a direct correlation between ozone dose and bromate formation and by applying ozone dosages at disinfection levels, bromate is likely to exceed regulatory limits. However, adding hydrogen peroxide reduced the amount of bromate formed, and in most cases levels fell below regulatory limits. A linear correlation was established between bromate inhibition and increasing H2O2/O3 ratio at constant ozone dose.
Amongst the three filtration media investigated, only GAC achieved 1-log removal for geosmin and MCPA. The superiority of GAC over anthracite and Filtralite® was attributed to its adsorption affinity. Filtralite® and anthracite media were both ineffective for MCPA removal due to its non-biodegradable nature under conventional water treatment conditions.
At a 1 mg/L-ozone dose, GAC and Filtralite® filters achieved a 1-log geosmin removal. In contrast, a 1.44 mg/L ozone dose was required to meet this target with anthracite.
The tandem of ozone followed by biological filtration was very effective for the control of distribution system TTHM production regardless of filter media, with levels well below current and anticipated provincial regulatory limits.
The combination of intermediate ozonation followed by deep-bed biological filtration is well suited for treating Grand River water. Scale-up considerations include pairing the proper filter media to the size of the ozone generator. The best two treatment scenarios were: Option 1: select GAC media and size the ozone generator to produce a 1 mg/L dose. Option 2: select anthracite media and size the ozone generator to deliver a 2 mg/L dose.
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