Heptamethine cyanine dyes are attractive compounds for imaging purposes in biomedical applications because of their chemical and photophysical properties exhibited in the near-infrared region. A series of meso amino-substituted heptamethine cyanine dyes with indolenine, benz[e]indolenine and benz[c,d]indolenine heterocyclic moieties were synthesized and their spectral properties including fluorescence quntum yield were investigated in ethanol and ethanol/water mixture. Upon substitution with amines, the absorption maxima of the dyes shifted to the lower wavelength region (~600 nm), showed larger Stokes shifts and stronger fluorescence which can be attributed to an excited state intramolecular charge transfer (ICT). High quantum yields were observed for primary amine derivatives and lower quantum yields were observed for secondary amine derivatives. Fluorescence quantum yields are greater for dyes with 3H-indolenine terminal moieties than for dyes with benz[e]indolenine end groups. Benz[c,d]indolenine based heptamethine cyanine dyes exhibited the lowest quantum yield due to aggregation in solution. In general, the benz[e]indolenine hepatemethine cyanines showed high Stokes shifts compared to indolenine dyes. For the meso-chloro dyes, the absorption maxima for the dyes shifted bathochromically in the order of indolenine, benz[e]indolenine and benz[c,d]indolenine.

SecA is a central component of the general secretion system that is essential for growth and virulence of bacteria. A series of fluorescein analogs were tested against ATPase activities of Escherichia coli SecA. Rose Bengal (RB) and Erythrosin B are potent inhibitors abolishing the activities of three forms of SecA ATPase with IC50 in µM range. Both inhibit SecA intrinsic ATPase with two mechanisms depending on ATP concentrations, indicating they influence the two non-identical nucleotide binding sites differently. RB shows different inhibitory effects against three forms of SecA ATPase activities, suggesting that the inhibition is related to the conformation of SecA. RB with IC50 at sub-µM level is the most potent inhibitor of SecA ATPases and SecA-dependent protein translocation to date. The fluorescein analogs inhibit intrinsic ATPase of Bacillus subtilis SecA similarly, and also exhibit antibacterial effects in E. coli and B. subtilis. Our findings indicate the value of fluorescein analogs as probes for mechanistic studies of SecA and the potential development of new SecA-targeted antimicrobial agents. A series of SecA derivatives with truncated C-terminus within the first long α-helix of the helix-bundle extending the ATPase catalytic domain of N68 was analyzed. These SecA variants interact with lipids, and those containing the C-terminal portion of the long α-helix starting at residues #639 form the ring-like structure in liposomes, indicating the critical domains for forming the protein-conducting channel. The presence and length of the C-domain influence the response to RB of NBDII mutants and C-terminal truncates of SecA. Thus this region may interact with the inhibitors and is involved in the structure and regulation of SecA ATPase activity. B. subtilis SecA was analyzed for interspecies comparison. Despite sharing high homology, this SecA homolog cannot complement E. coli mutants with SecA defect. Phospholipids do not stimulate ATPase activities of B. subtilis SecA, but induce its conformational changes, leading to the lipid-specific domains and ring-like structures similar to E. coli SecA. These pore-ring structures may represent part of the protein-conducting channels. Therefore, the potential structural roles of SecA in the protein translocation machinery may be universal in both Gram-negative and Gram-positive bacteria.

In part A, the effect of varying short-chain alkyl substitution on the spectroscopic properties of cyanine dyes was examined. Molar absorptivities and quantum yields were determined for groups of pentamethine and heptamethine dyes for which the substitution of the indole nitrogen was varied. For both sets of dyes, increasing alkyl chain length did not significantly change quantum yield or molar absorptivity. These results may be useful in designing new cyanine dyes. In part B, the effect of structure on the suitability of 2,4,6-trisubstituted pyridines as color pH indicators was studied by determining spectral effects of protonation, molar absorptivities, pKa values, and the structural origin of the spectral behavior. Good color indicating properties result from aniline substitution at the 4 position of pyridine and electron donating substitution at the 2 and 6 positions of pyridine, which provide a strong red shift in the spectra and greater red shifted peak absorptivity, respectively.

La industria textil produce grandes cantidades de agua residual con un alto contenido de materia orgánica y color. Debido a la amenaza ambiental que supone su naturaleza refractaria, surge la necesidad del tratamiento antes de la descarga. Los procesos biológicos, físicos y químicos convencionales resultan ineficaces en la decoloración y mineralización completa de estos efluentes. Como alternativa emergen los Procesos Avanzados de Oxidación (PAOs), basados principalmente en la generación de radicales hidroxilo (HO·) altamente reactivos. Éstos pueden aplicarse de forma exclusiva o combinados con tratamientos biológicos con el fin de reducir su elevado consumo de reactivos y energía. Entre los diferentes PAOs disponibles, los procesos de Fenton y, particularmente, los procesos de foto-Fenton (basados en la generación de HO· mediante la adición de peróxido de hidrógeno y una sal ferrosa en disolución acuosa) suponen la mejor opción debido a su elevada eficacia y bajo coste económico. Adicionalmente, los procesos de foto-Fenton pueden llevarse a cabo bajo radiación solar, ofreciendo nuevas ventajas económicas y medioambientales. Esta tesis doctoral se centra básicamente en la aplicación del proceso de foto-Fenton como etapa previa en la degradación de soluciones de azo colorantes reactivos biorrecalcitrantes, con el propósito de generar un nuevo efluente compatible con un tratamiento biológico aerobio posterior de menor coste e impacto ambiental que el primero. La evolución de parámetros clave como el color, el carbono orgánico total, la aromaticidad, la toxicidad, la biodegradabilidad, la naturaleza de los intermedios de degradación, así como el efecto de la fuente de irradiación (luz artificial, luz solar), la temperatura, el tiempo de reacción y la concentración de reactivos, proporcionan información acerca de las condiciones idóneas para el acoplamiento PAO/tratamiento biológico. Los resultados obtenidos confirman la idoneidad del PAO propuesto con este fin, tanto a escala de laboratorio como en planta piloto. Mediante el proceso de foto-Fenton aplicado bajo condiciones de oxidación suaves, las soluciones bajo estudio resultan incoloras, de naturaleza biodegradable y no tóxica. De este modo, el tratamiento combinado foto-Fenton/tratamiento biológico aerobio permite su degradación completa. La luz solar como fuente de radiación en el proceso de foto-Fenton proporciona los mejores resultados. Por otro lado, la secuencia oxidativa parece comenzar por la decoloración por rotura del grupo azo seguida de la degradación de aromáticos para formar ácidos carboxílicos alifáticos -dando lugar a la formación de CO2 y H2O- o el anillo triazina de naturaleza recalcitrante. Una parte importante de los heteroátomos presentes en la molécula original aparecen como productos finales de naturaleza inorgánica inocua.El trabajo está ampliado con un estudio económico y medioambiental del proceso secuencial foto-Fenton (luz artificial)/tratamiento biológico. La evaluación medioambiental se ha realizado mediante la herramienta de Análisis de Ciclo de Vida. En comparación con los procesos simples foto-Fenton bajo luz artificial y foto-Fenton bajo luz solar, el tratamiento combinado resulta la mejor opción en ambos contextos. Los mayores impactos se asocian, en este orden, al consumo de peróxido de hidrógeno y de energía para alimentar la luz artificial. En consecuencia, y atendiendo a los resultados obtenidos previamente, es posible concluir que el proceso de foto-Fenton asistido con luz solar como pre-tratamiento de un proceso biológico sería la mejor opción en términos de efectividad, impacto medioambiental y coste operacional. Finalmente, como alternativa a la estrategia de oxidación química/tratamiento biológico aerobio, el PAO se aplica como post-tratamiento a un proceso biológico anaerobio. Los PAOs considerados son la ozonización y el proceso de foto-Fenton. Los resultados obtenidos manifiestan la idoneidad de la secuencia propuesta, obteniéndose unos mejores niveles de degradación mediante el proceso con ozono. Estos resultados son de especial interés en futuras aplicaciones para el tratamiento de aguas residuales textiles reales. / The textile industry produces large quantities of wastewater that is highly coloured and contains large concentrations of organic matter. Due to the environmental threat that supposes its recalcitrant nature, the application of specific treatment is required prior discharge. Conventional biological, physical and chemical processes are quite inefficient to completely mineralise and decolourise these effluents. Alternatively appear the Advanced Oxidation Processes (AOPs), principally based on the generation of highly reactive hydroxyl radicals (HO·). They may be applied as exclusive processes or combined with biological treatments in an attempt to reduce their large chemicals and energy consumption. Among available AOPs, the Fenton and, particularly, the photo-Fenton processes (based on HO· generation by means of hydrogen peroxide and a ferrous salt addition in aqueous solution) are of special interest since they achieve high reaction yields with a lower operational cost. Additionally, the photoassisted reaction presents the possibility of be driven under solar irradiation, offering further economic and environmental advantages. This doctoral dissertation is basically centred in the performance of the photo-Fenton process as a previous step to treat biorecalcitrant solutions polluted with commercial reactive azo dyes, aiming to generate a new effluent amenable to aerobic biotreatment (with a lower economic and environmental impact than the AOP). The evolution of key parameters such as the colour, the total organic carbon, the aromatic compounds content, the toxicity, the biodegradability, the generated by-products nature, as well as the type of irradiation effect (artificial light, solar light), the temperature, the reaction time and the reagents concentrations, provide information about the most suitable conditions to carry out the AOP/biological coupled treatment. Obtained results evidence the aptness of the AOP proposed with this aim, either at laboratory and pilot plant scale. With a partial oxidation run under proper mild conditions, the studied dye solutions become decolourised, biodegradable as well as non toxic. In this way, the combined photo-Fenton/biological treatment allows the complete mineralisation. The solar light as a source of irradiation provides the best results. On the other hand, the oxidative sequence appears to begin with the hydroxyl radical attack to azo groups, giving place to the solutions decolourisation. The following aromatics degradation generates either short chain carboxylic acids -finally yielding CO2 and water- or the recalcitrant triazine moiety. An important part of the heteroatoms initially present in the molecule gradually appear as innocuous final products of inorganic nature.The work is extended with an economic and environmental study of the sequential photo-Fenton (artificial light)/biological treatment process. The environmental evaluation has been realised by means of the Life Cycle Assessment tool. In comparison with single artificial light photo-Fenton process and solar driven photo-Fenton process, the combined treatment supposes the best option in both contexts. The major burdens are mainly attributed to the hydrogen peroxide requirements and the electrical energy consumption to run the artificial light, in this order. Consequently, and attaining to previously obtained results, the solar driven photo-Fenton process as a pre-treatment of a biological treatment would be the best option in terms of effectiveness, environmental impact and operational cost. Finally, as an alternative to the chemical/aerobic biological treatment, the AOP is performed as a post-treatment of an ensuing anaerobic biotreatment. Ozonation and photo-Fenton processes have been chosen for this role. Obtained results manifest the suitability of the proposed sequence, attaining the best degradation levels by means of ozonation process. These results are of special interest for real wastewater applications.

El objeto de la presente tesis es el desarrollo de metodologías analíticas rápidas para optimizar los procesos de degradación de colorantes orgánicos presentes en aguas residuales. Se ha llevado a cabo una revisión bibliográfica acerca de dichos procesos y las técnicas analíticas utilizadas para su monitorización y la identificación de posibles intermedios. Se han establecido nuevas metodologías analíticas basadas en herramientas quimiométricas tales como diseño de experimentos para optimizar procesos de fotodegradación con y sin catalizador. Se ha empleado resolución de curvas multivariante para la determinación simultánea de colorantes e intermedios durante estos procesos. Dado que la velocidad de degradación es el parámetro mas utilizado para evaluar la eficiencia de las fotodegradaciones se han evaluado todas las constantes involucradas en la degradación y/o la adsorción sobre el catalizador mediante HS-MCR-ALS. Se ha establecido una metodología analítica rápida para la determinación de colorantes basada en cromatografía de inyección secuencial (SIC). / The main aim of this thesis is the development of rapid analytical methodologies to optimize the photodegradation processes of organic azo dyes from industrial wastewater. A bibliographic review has been done concerning the most employed processes for dye removal and the analytical techniques used for the monitoring and the identification of intermediates. New analytical methodologies based on chemometric tools such as experimental designs have been established to optimize photodegradation processes with and without using heterogeneous catalysts. Multivariate curve resolution has been employed to determine simultaneously different dyes and intermediates along degradation processes. Since the degradation rate is the most employed parameter to evaluate the photodegradation efficiency all the kinetic constants involved in the degradation and/or the adsorption of the dye onto the catalyst have been evaluated by HS-MCR-ALS. Furthermore a rapid analytical methodology for dye determination based on sequential injection chromatography (SIC) has been established.

Textile dyeing and finishing industry uses dryers/tenters for drying and heat-setting fabrics. A very large fraction of the heating value of the fuel consumed in the burner ends up as waste in the dryer exhaust. An initial calculation showed that up to 90% of the energy consumed in the tenter is wasted. Therefore, quantifying the energy waste and determining drying characteristics are vitally important to optimizing the tenter and dryer operations. This research developed a portable off-line gas chromatography-based characterization system to assess the excess energy consumption. For low-demanding heat-setting situations, energy savings can be realized quickly. On the other hand, there are demanding situations where fabric drying represents the production bottleneck. The drying rate may be governed either by the rate of heat transport or by the rate of moisture transport. A mathematical model is being developed that incorporates both these processes. The model parameters are being obtained from bench-scale dryer studies in the laboratories. The model will be validated using production scale data. This will enable one to predict optimization dryer operation strategies.

Anthraquinone dyes constitute the second largest class of textile dyes, and are used extensively in the textile industry. A high fraction of the initial reactive dye mass used in the dyeing process remains in the spent dyebath. Reactive dyes are not readily removed by typical wastewater treatment processes and the high salt concentration typical of reactive dyeing further complicates the management of spent reactive dyebaths. Investigation of the reductive transformation of reactive anthraquinone dyes and their decolorization products has been very limited. Additionally, very limited research has been conducted on the decolorization of spent reactive dyebaths. Research was conducted to investigate the key operational parameters of batch and continuous-flow ZVI decolorization of a reactive anthraquinone dye, Reactive Blue 4 (RB4), under anoxic conditions, as well as the potential for the biodegradation of its decolorization products in a halophilic culture under aerobic conditions. The effect of two operational parameters, such as mixing intensity and initial dye concentration, on the ZVI batch decolorization kinetics indicates that ZVI decolorization of RB4 is a surface-catalyzed, mass transfer-limited process. The high salt and base concentrations enhanced the rate of RB4 decolorization. Based on parameters such as porosity, hydraulic conductivity, pore water velocity, and dispersion coefficient, non-ideal transport characteristics were observed in a continuous-flow ZVI column. The results of a long-term continuous-flow ZVI decolorization kinetics demonstrated that continuous-flow ZVI decolorization is feasible. However, column porosity losses and a shift of reaction kinetics occur in long-term column operation. ZVI decolorization of RB4 was successfully described with a pseudo first-order or a site saturation model. Lastly, the RB4 decolorization products generated by ZVI treatment had no inhibitory effect on the halophilic culture. However, biodegradation and/or mineralization of RB4 decolorization products was not observed after a long-term incubation of the culture. This research demonstrated the feasibility of ZVI decolorization of reactive anthraquinone dyes, which will help in the development of a continuous-flow, dyebath decolorization process and the possible reuse of the renovated dyebath in the dyeing operation. Such a system could lead to substantial reduction of water usage, as well as a decrease of salt and dye discharges.

Supramolecular chemistry covers intermolecular interactions where non-covalent bonds are involved, and many of them are based on host-guest interactions. Cyclodextrins (CDs) are cyclic oligosaccharides consisting of 6-, 7- or 8-glucose units, which are called alpha-, beta- or gamma-CDs, respectively. They have hydrophobic interior and hydrophilic exterior, and are widely being used as hosts for various organic molecules. The formation of CD inclusion complexes with a variety of dyes has continuously drawn our interests, since CDs are readily available and have ability to include dye molecules altering their properties. The present thesis covers the study of inclusion complexes of CDs and chromophore dyes, largely in two ways; rotaxane and pseudorotaxane. The stable rotaxane structure is achieved with the synthesis of dye rotaxane. The introduction of CD ring around azo chromophore provides a simple way to improve the solubility and stability of azo dye. We have shown that by incorporating proper compounds as a coupler, azo dye rotaxanes can be used as pH indicators and metal ion sensors. We have described the synthesis of novel acetylene dye rotaxane using the Pd-catalyzed reaction of Heck-Cassar-Sonogashira-Hagihara type. Its fluorescence properties in the solid state as well as in solutions are examined and compared with those of free dye. Free dye, which has tetra-carboxylic groups, is found to be highly sensitive to various metal ions, exhibiting high Stern-Volmer constants, K(SV). On the contrary, acetylene dye rotaxane exhibits much less quenching against various quenchers. The appearance of fluorescent anisotropic structure has been observed by the formation of inclusion complex between acetylene dye and gamma-CD. Its structural nature is studied by various techniques, including fluorescence, fluorescence anisotropy, wide angle X-ray scattering (WAXD) and differential scanning calorimetry (DSC) measurements. Methyl orange, an acid azo dye, forms a dimeric inclusion complex with gamma-CD, resulting in the formation stable anisotropic aggregates. Several other azo dyes are found to form anisotropic supramolecule in the presence of gamma-CD, and their structural characteristic has been discussed in terms of the number and position of solubilizing groups.

Small silver nanoclusters are synthesized using polycytosines as matrices. Different size silver nanoclusters ranging from Ag1 to Ag7 exhibit bright emission maxima at blue (480nm), green (525nm), red (650nm), and IR (720nm) wavelengths with varying the excitation wavelengths. With electrophoresis, correlation of emission with mass spectra, the Ag cluster sizes are identified with blue emitters as Ag5, green emitters as Ag4, red emitters as Ag3, and IR emitters as Ag2. Ag4 and Ag5 appear to be partially oxidized while Ag2 and Ag3 are likely fully reduced. Silver cluster stability and their dynamics are observed from silver clusters encapsulated by polycytosine (Cm:Agn). From length study of polycytosine, the longer the polycytosine is, the more stable the larger clusters such as Ag5 are. In time-dependent optical measurements, isosbestic points are observed from Cm:Agn by converting red and IR species into blue and green species, while in the case of temperature-dependent optical properties, with increasing temperature, the blue (oxidized Ag5) and green (oxidized Ag4) emitters convert into the red (Ag3) and IR (Ag2) emitters. NaCl-dependent optical measurements support the assignments of oxidized and fully reduced silver emitters. Circular dichroism (CD) is used to investigate conformational changes in Cm and Cm:Agn with varying conditions (time, temperature and NaCl) and the studies indicate that no conformational changes in Cm:Agn are observed from the time and temperature, while the conformational changes in Cm:Agn are observed from the NaCl studies. From pH-dependent emission study of Cm:Agn, the silver nanocluster dynamics slow down at high pH. Using confocal microscopy technique, single molecules on IR species, C12:Ag2 are investigated and demonstrate that C12:Ag2 is brighter and more photostable than Cy5 which is known to be one of the best IR dyes. With low excitation power, molecules can be monitored for hours, giving bright blinking free, stable fluorescence. The photophysics of this new dye make it a promising candidate for single molecule studies in biological applications.

New methodologies for the efficient syntheses of 4,4-difluoro-4-bora-3a,4adiaza- s-indacenes (BODIPYs) and rosamines were developed. A serendipitous discovery led to a new reaction which afforded BODIPYs in high yields. Systematic studies of the kinetics and mechanisms of the new reaction were performed. A series of BODIPYs were successfully prepared using the new approach. A simple and efficient synthesis of rosamines with cyclic-amine substituents was devised. These new rosamines showed interesting anti-tumor activities. Several types of novel fluorescent compounds were prepared. Highly fluorescent GFP-chromophore analogs were designed and synthesized. The correlation between the optical properties and the structures was investigated. New pyronin dyes with mesoheteroatom substituents were efficiently prepared. The fluorescence properties of these compounds were highly dependent on the nature of the meso-substituents. A set of BODIPY dyes that fluoresce brightly above 600 nm were made. They were then used as acceptors to prepare water-soluble through-bond energy transfer cassettes. All the cassettes had complete energy transfer and high quantum yields in MeOH. A few also had good fluorescence properties in aqueous media and even on proteins. The through-bond energy transfer cassettes were used to monitor protein-protein interactions. In order to test our hypothesis, an artificial protein interaction system was built by utilizing the biotin/(strept)avidin interactions. Thus Atto425-BSA-biotin, streptavidin-cassette1 and avidin-cassette2 were prepared. The interactions between Atto425-BSA-biotin and cassette labeled (strept)avidin were successfully detected in vitro and in living cells by fluorescence techniques.