O0001 On the efficiency of the peroxyoxalate system using a simple experimental and theoretical approach Felipe A. Augusto, Carolina P. Frias, Wilhelm J. Baader Instituto de Química, São Paulo, SP, Brazil The peroxyoxalate system is the most efficient intermolecular chemiluminescent reaction with chemiluminescence quantum yields reaching up to three orders of magnitude higher than other similar systems.1 It consists in the reaction of an oxalic ester with hydrogen peroxide, typically catalyzed by a base, forming a high-energy intermediate which interacts with a compound called activator, ACT.2,3 This interaction leads to the ACT in its singlet excited state in a mechanism that involves a charge/electron transfer initially from the ACT to the peroxide and then back to the ACT.2,3 Several important details have been experimentally determined about the mechanism, like the cyclization rate constant and the direct interaction between the high-energy intermediate and the ACT.4,5 However the identity of the high-energy intermediate is still a matter of discussion, with only a few possible intermediate structures being properly studied or definitely discarded.6 The study and characterization of the peroxyoxalate reaction with a relatively small ACT, like naphthalene, could allow a detailed theoretical study of the chemiexcitation step, contrarily to the case of the compounds normally used as ACTs which would involve prohibitive computational costs. Therefore, the reaction of bis(2,4-dinitrophenyl) oxalate (DNPO) with hydrogen peroxide catalyzed by imidazole (IMI-H) was studied using naphthalene as ACT. The observed rate constant (kobs) showed linear dependence with the [H2O2] (Fig., kH2O2 = 23 ± 1 L mol-1 s-1) and [IMI-H] (Fig., kIMI-H = 202 ± 7 L mol-1 s-1) and no dependence with [DNPO] or [ACT] (data not shown). The study of the peroxyoxalate system, employing naphthalene as electronically simple ACT, indicates the existence of a linear relationship between the kobs and the H2O2 as well as IMI-H concentration, in general agreement with the mechanism proposed for the initial steps of the transformation. This fact indicates a normal behavior of the system in the conditions utilized. Therefore, the peroxyoxalate reaction with naphthalene as ACT will now be subject to theoretical studies in order to elucidate the exact mechanism of the chemiexcitation step, with the intention to understand the reason for the extremely high efficiency of the system even so it involves, apparently, intermolecular electron transfer steps. References 1. Augusto FA, Souza GA, Souza Junior SP, Khalid M, Baader WJ. Photochem. Photobiol. 2013;89:1299. 2. Ciscato LFML, Augusto FA, Weiss D, Bartoloni FH, Albrecht S, Brandl H, Zimmermann T, Baader WJ. ARKIVOC 2012;2012:391. 3. Bartoloni FH, Bastos EL, Ciscato LFML, Peixoto MMdeM, Santos APF, Santos CS, Oliveira S, Augusto FA, Pagano APE, Baader W. J. Quim. Nova 2011;34:544. 4. Da Silva SM, Casallanovo F, Oyamaguchi KH, Ciscato LFML, Stevani CV, Baader WJ. Luminescence 2002;17:313. 5. Ciscato LFML, Bartoloni FH, Bastos EL, Baader WJ. J. Org. Chem. 2009;74:8974. 6. Stevani CV, Campos IPA, Baader WJ. J. Chem. Soc., Perkin Trans. 2 1996;1645. O0002 Kinetic studies on the sodium salicylate catalyzed peroxyoxalate reaction Glalci A. Souza, Wilhelm J. Baader Instituto de Química da Universidade de São Paulo, São Paulo-SP, Brazil The peroxyoxalate reaction is the only chemiluminescence reaction which appears to involve the intermolecular Chemically Initiated Electron Exchange Luminescence (CIEEL) mechanism in its chemiexcitation step that possess highest emission quantum yields of up to 60%.1-3 Detailed kinetic studies on this highly efficient CL system has been performed mainly using imidazole as base catalyst and the mechanistic elucidation of the complete reaction has shown that this compound is acting also as nucleophilic catalyst.4-5 However, it has been shown also that the base and nucleophilic catalyst imidazole leads to a decrease in the CL emission quantum yield, apparently due to its interaction with the high-energy intermediate.4-6 In order to further elucidate the mechanism of the peroxyoxalate system, the kinetics of the reaction were studied with sodium salicylate as base catalyst. The CL emission obtained in the reaction of bis(2,4,6-trichlorophenyl) oxalate (TCPO) (0.1mM) with hydrogen peroxide, catalyzed by sodium salicylate, in the presence of 9,10-diphenylanthracene (DPA) (0.2mM) as activator, in ethyl acetate at 25°C, was measured in different conditions. When the reaction was performed varying the sodium salicylate concentration, the rate constants corresponding to the emission decay increased with the base concentration, showing a saturation curve like behavior. The decay rate constant also increased with increasing hydrogen peroxide concentrations and at low H2O2 concentrations the rate constants show a linear dependence on the hydrogen peroxide concentration allowing the determination of a bimolecular rate constant (kbim). These rate constants showed to depend also on the sodium salicylate concentration; kbim=(0.56±0.06) 10-3; (1.6±0.2) 10-3 and (2.04±0.06) 10-3Lmol-1s-1, for sodium salicylate concentrations of 0.3, 1.0 and 5.0mmolL-1, respectively. Upon variation at higher hydrogen peroxide concentrations the rate constants showed saturation curves, indicating a change in the rate-limiting step. The rate constants corresponding to the initial rise in the emission intensity have been shown independent of the hydrogen peroxide concentration showing that this reagent does not participate in the reaction step measured in this part of the kinetic curve. Similarly to the behavior observed with imidazole, the CL emission quantum yield showed to decrease with an increase in the sodium salicylate concentration. The maximum quantum yields obtained with sodium salicylate were smax=(1.24±0.06) 10-3 E mol-1 when the sodium salicylate concentration was 0.5mmolL-1. These data indicate that, like imidazole, also sodium salicylate appears to interaction with the high-energy intermediate in the peroxyoxalate reaction, which diminishes the CL emission quantum yields. Financial support; FAPESP, Capes, CNPq. 1. Stevani CV, Silva SM, Baader WJ. Eur. J. Org. Chem. 2000;4037. 2. Augusto FA, Souza GA, Souza Jr., SP, Khalid M, Baader WJ. Photochem. Photobiol. 2013, 89, 1299. 3. Ciscato LFML, Augusto FA, Weiss D, Bartoloni FH, Bastos EL, Albrecht S, Brandl H, Zimmermann T, Baader, WJ. Arkivoc;2012 (iii), 391. 4. Silva SM, Casallanovo F, Oyamaguchi KH, Ciscato LFLM, Stevani CV, Baader WJ, Luminescence 2002;17:313. 5. Stevani CV, Lima DF, Toscano VG, Baader WJ, J. Chem. Soc. Perk. Trans. 2 1996;989. 6. Ciscato LFML., Bartoloni FH, Bastos EL., Baader WJ, J. Org. Chem. 2009;74:8974. O0003 How can quantitative bioluminescence and in-situ fluorescence of firefly oxyluciferin in luciferase be compared with theoretical calculations? Hidefumi Akiyamaa, Yu Wanga,b, Miyabi Hiyamaa, Toshimitsu Mochizukia, Kanako Terakadoc, Toru Nakatsuc aUniversity of Tokyo, Kashiwa, Chiba 2778581, Japan bInstitute of Genetics and Developmental Biology, Beijing 100101, Japan cKyoto University, Sakyo-ku, Kyoto 6068501, China To investigate color-determination mechanisms from physicist points of view, we study quantitative spectra of firefly bioluminescence, and the in-situ absorption and fluorescence spectra of oxyluciferin contained in luciferase in a consumed reaction mixture, and intend to compare them with quantum-chemistry theoretical calculations. We have so far measured quantitative in-vitro firefly bioluminescence spectra influenced by pH, kinds of bivalent metal ions, temperatures, and mutant luciferase using our total-photon-flux spectrometer with our new light standards. We found that all the spectra were systematically and quantitatively decomposed into one environment-sensitive and two environment -insensitive Gaussian peaks, and that no intensity conversion between yellow-green and red emissions but mere intensity variation of the pH-sensitive green peak at 2.2eV causes the changes in apparent emission colors [1,2]. Therefore, answers for the color-determination problem need not only the assignment of the peaks, but also the explanation of the intensity change of the green peak. We next measured in-situ absorption and fluorescence characteristics of oxyluciferin, still combined with luciferase in a consumed reaction mixture [3]. The in-situ absorption spectra indicated that neutral oxyluciferin was dominant, which was in weak pH-dependent equilibrium with oxyluciferin mono-anions. The neutral oxyluciferin was more dominant in luciferase environments than in bare water environments. The in-situ fluorescence spectra shown in Fig. clarified that the neutral oxyluciferin is a blue emitter and the oxyluciferin mono-anion is a green emitter (Fig.). Even in red-mutant luciferase environment, the in-situ fluorescence has shown strong blue and green fluorescent emissions (Fig.). In short, the spectra of in-situ fluorescence of oxyluciferin in luciferase and those of bioluminescence have shown significant discrepancy. Therefore, the above-mentioned discrepancy between bioluminescence and in-situ fluorescence cast an important question, how they can be consistently compared with quantum-chemistry theoretical calculations, which are recently published in a large numbers. The above results may suggest that enzyme microenvironment affects the transition state in bioluminescence chemical reaction, but not the oxyluciferin states after the reaction. Physicists need to discuss this issue with biologists and chemists in the bioluminescence research community. 1. Ando Y, Niwa K, Yamada N, Irie T, Enomoto T, Kubota H, Ohmiya Y, Akiyama H. Nature Photonics 2008;2:44-47. 2. Wang Y, Akiyama H, Terakado K, Nakatsu T. Scientific Reports 2013;3:2490. 3. Wang Y, Hayamizu Y, Akiyama H. J. Phys. Chem. B 2014;118:2070-2076. O0004 Determination of the total phenolic / antioxidant content in honey samples using formaldehyde / potassium permanganate chemiluminiscence system in a novel microfluidics device Butheina A. M. Al Haddabi, Haider A. J. Al Lawati, FakhrEldin O. Suliman, Gouri B. Varma Sultan Qaboos University, Al-Khod, Oman Microfluidc device has been explored as a tool for the estimation of the total phenolic content/ antioxidant content in honey using acidic potassium permanganate chemiluminescence (KMnO4-CL) detection system. Selected phenolic antioxidants including quercetin, catechin, gallic acid, caffeic acid and ferulic acid elicited analytically useful CL with detection limits ranging between 2.38nmolL-1 for galic acid and 33.9nmolL-1 O-coumaric acid for only 2µL injection volume. The parameters that affect the CL signal intensity of each antioxidant were carefully optimized. . It was observed that formaldehyde can enhance the CL signal intensity of phenolic compounds up to 27 times (Fig.). Additionally, it was observed that the chip volume and geometry both can play an important role in enhancing the CL signal intensity in this system. The CL signal intensity was enhanced five times when a spiral - flow split chip (SF) geometry was used, compared to the simple spiral chip (S) geometry commonly used (Fig.). Other parameters were also optimized, including pH and concentration of reagents used and the flow rates. The effect of solvents and surfactants on CL signal intensities was also studied. The method was applied on Omani honey samples. Nine different honey samples resulted in total phenolic / antioxidant level range between 40 and 772mg Kg-1 with respect to gallic acid. Folin Coicalteu reagent (FCR) resulted in a good correlation with the developed method which was found to be a selective, rapid and sensitive method to estimate total phenolic / antioxidant level in a good agreement with reported results for honey samples. References 1. Costin JW, Barnett NW, Lewis SW, McGillivery DJ. Analytica Chimica Acta 2003;499:47-56. 2. Alvarez-Suarez JM, Gonzalez-Paramas AM, Santos-Buelga C, Battino M. J. Agric. Food Chem. 2010;58:9817-9824. O0005 Electrochemiluminescence sensor based on tris(2,2'bipyridyl)ruthenium(II)/poly(AHNSA) for chlorpheniramine maleate analysis Mohammed M. Alhinaai, Emad A. Khudaish Sultan Qaboos University, AlKhude, Muscat, Oman Since the discovery of impressive luminance property of tris(2,2'bipyridyl)ruthenium(II), [Ru(bpy)3]2+, it becomes one of intensive used reagent for chemiluminescence and electrochemiluminescence (ECL) analysis with a wide range of coreactants. Immobilizing of the expensive Ru(II)-complex on the electrode surface began early 1980s using Nafion as an exchanger polymer [1] is still a hot and continuous topic by many research groups [2]. The main objective of the present work was to develop a solid-state ECL sensor based on immobilizing [Ru(bpy)3]2+ on a conducting polymer for chlorpheniramine maleate (CPM) determination. The sensor was fabricated by composite electropolymerization of 2.0mM [Ru(bpy)3]2+ and 1.5mM of 4-amino-3-hydroxy-naphthalene sulfonic acid (AHNSA) in acidic medium via potentiodynamic repetitive cycles between -0.8 and +2.0V at 0.1Vs-1. The fabrication parameters were optimized carefully in order to produce a stable film and obtain an intense ECL signal. Figure shows the electrochemical characterization of the composite surface film where a redox peak of [Ru(bpy)3]2/3+ are well defined at 1180mV (anodic) and 980mV (cathodic), respectively. Impedance spectroscopy analysis showed that the charge transfer resistance of the composite film is greatly lowered by doping Ru(II)-complex on the moiety of the PAHNSA which suggests that Ru(II)-complex is acting as a charge transfer center [3]. This sensor exhibited excellent ECL behaviour toward CPM analysis as shown in Fig. with a good stability and reproducibility. The standard deviation of 16 measurements in flow stream was 2.35%. It also has a very good lifetime when stored in 5°C for two weeks where the recovery measurement approached 94%. The sensor was also applied to estimate CPM in pharmaceutical preparations. The linear dynamic range was from 0.1 to 32µg/mL (R2=0.9956). The detection limit was 23µg/L and the recovery of real sample analysis was from 102.0% to 98.50% for syrup and tablets respectively. Interference studies showed no effect of common compounds and the acceptable molar concentration ratios of foreign species to CPM were higher than1000-fold for Na+, K+, NO3-, SO32-, 100-fold for Mg2+, Al3+, NH4+, Cl-, lactose, sucrose, and glucose, and 10-fold for Fe3+ and Co2+. The analytical parameters such as pH, flow rate, buffer concentration were systematically tested. In conclusion, a novel composite polymeric film was fabricated using simple electrochemical method and applied for determination of CPM in real samples. The sensor showed a good stability and sensitivity regardless the matrix of the pharmaceutical preparation. References 1. Rubinstein I, Bard A. J. J. Am. Chem. SOC. 1980;102;6641-6842. 2. Su M, Wei W, Liu S. Analytica Chimica Acta 2011;704:16-32. 3. Zhang B, Shi S, Shi W, Sun Z, Kong X, Wei M, Duan X. Electrochimica Acta 2012;67:133-139. O0006 Experimental Evidence of the Occurrence of an Intermolecular Electron Transfer in the Catalyzed Decomposition of spiro-Alkyl-1,2-Dioxetanones Fernando Heering Bartoloni2,1, Marcelo Almeida de Oliveira1, Luiz Francisco Monteiro Leite Ciscato2,1, Felipe Alberto Augusto1, Erick Leite Bastos1, Wilhelm Josef Baader1 1Departamento de Química Fundamental do Instituto de Química da Universidade de São Paulo, Sao Paulo. SP, Brazil, 2Centro de Ciências Naturais e Humanas, Universidade Federal do ABC, Santo Andre, SP, Brazil Several chemical and biochemical reactions have light as a co-product and some of them can show high quantum efficiencies, include firefly bioluminescence, the peroxyoxalate system, and the induced decomposition of 1,2-dioxetanes.1 Cyclic peroxides have been frequently described as high-energy intermediates in the chemical formation of products in the electronic excited state because their decomposition fulfill both energetic and geometric criteria required for chemiexcitation. Nevertheless, the thermal decomposition of 1,2-dioxetanes and 1,2-dioxetanones results in inefficient chemiluminescence emission due to the preferential formation of products in the non-emissive triplet-excited state (ΦS<10-4 E mol-1 vs. ΦT>0.1 E mol-1). ).2,3 However, it has been reported that fluorescent polycyclic aromatic hydrocarbons with low oxidation potentials (referred to as activators, ACT) catalyze the decomposition of 3,3-dimethyl-1,2-dioxetanone (1), resulting in noticeable increase in light emission intensity, and high singlet chemiexcitation quantum yields (ΦS=0.1 E mol-1).4 Contrarily, our group found recently that the ΦS for the catalyzed decomposition of 1,2-dioxetanones, the model intermediate in firefly bioluminescence, were overestimated in several orders of magnitude.5 Consequently, the validity of this system as a model for excited states formation might be questioned. Therefore, we report here our results of a kinetic study on the catalyzed decomposition of the spiro-substituted 1,2-dioxetanone derivatives, spiro-adamantyl-1,2-dioxetanone (2) and spiro-cyclopentyl-1,2-dioxetanone (3) by several activators and confirmed the occurrence of an intermolecular electron or charge transfer in this transformation. The 1,2-dioxetanone derivatives 2 and 3 were prepared, purified and handled as described elsewhere;6 kinetic assays and data treatment were performed as detailed before.5 The kobs values for the decomposition of 2 and 3, determined in toluene in the absence and in the presence of different ACTs, do not depend on the nature and concentration of the ACT (kobs(2, 50°C)=(6±1) 10-3s-1 and kobs(3, 25°C)=(9±3)×10-4s-1). Therefore, the bimolecular rate constant (kCAT) cannot be determined directly; however, the kCAT/kD ratios and the chemiexcitation quantum yields at the infinite ACT concentration (ΦS[infin]) can be calculated for each ACT from the double reciprocal plots of the singlet quantum yields (ΦS) versus the ACT concentrations.5 The kCAT/kD values show linear free-energy relation with the ACT's oxidation potential, indicating the importance of an intermolecular electron transfer from the ACT to the peroxide in the chemiexcitation step. The low efficiency in excited states formation in the catalyzed decomposition of these cyclic peroxides are rationalized by steric interactions between the activator and the bulky alkyl substituents on the peroxidic ring, thereby lowering the charge-transfer complex formation constant between the peroxide and the activator. Financial support; FAPESP, Capes, CNPq. 1. Augusto FA, Souza GA, Souza Junior SP, Khalid M, Baader WJ. Photochem. Photobiol. 2013;89:1299. 2. Adam W, Baader WJ. J. Am. Chem. Soc. 1985;107:416. 3. Adam W, Baader WJ. Angew. Chem. Int. Ed. Engl. 1984;23:166. 4. Schuster GB, Schmidt SP. Adv. Phys. Org. Chem. 1982;18:187. 5. de Oliveira MA, Bartoloni FH., Augusto FA, Ciscato LFML, Bastos EL, Baader WJ. J. Org. Chem. 2012;77;10537. 6. Bartoloni FH, de Oliveira MA, Augusto FA, Ciscato LFML, Bastos EL, Baader JW. J. Braz. Chem. Soc. 2012;23:2093. O0007 Chemiluminescent detection of Nitric Oxide Martina Bancirova Palacký University, Olomouc, Czech Republic The chemistry of nitric oxide inside humans and other mammals is perhaps the most interesting aspect of this simple molecule's behaviour. NO is involved in controlling blood pressure; transmitting nerve signals and a variety of other signalling processes. When tissues in the body become inflamed for long periods of time, the concentration of nitric oxide within them increases and this can be used to diagnose disease. But also NO secreted by activated cells appears to be a complex cocktail" of substances (see Fig..)(1) So is necessary to take in account the presence of ROS during the determination of nitric oxide. One of the chemiluminescent detections is based upon the chemiluminescence reaction between NO and the luminol (5-amino-2,3-dihydro-1,4-phthalazinedione)-H2O2 system. The luminol-H2O2 system is specifically reactive to NO, so that other nitrogen-containing compounds (organic nitrite, organic nitrate, and thio-nitroso compounds do not interfere. (2) The light emission of luminol as detection of bloodstains is a complex process based on hydrogen peroxide decomposition catalyzed by haemoglobin. Three common known methods of bloodstains; detection according to Grodsky, Weber and by Bluestar Forensic reagent(3), are based on the luminol chemiluminescence (complex process based on hydrogen peroxide decomposition catalyzed by haemoglobin). The Bluestar Forensic Magnum was chosen because of its declared stability as a "new" detection system for nitric oxide. The different dilutions (up to three orders) of the Bluestar Forensic Magnum were used. The experiments were done by using luminometer (type FB 12, Berthold Detection Systems, Germany) in the total volume of 1mL. Sodium azide was used as a specific quencher of singlet oxygen to prove its presence (see Fig..) 1. Kroncke KD, Fehsel K, Kolb-Bachofen V. Nitric Oxide; Cytotoxicity versus Cytoprotection--How, Why, When, and Where?, NITRIC OXIDE; Biology and Chemistry 1997;1:107-120. 2. Kikuchi K, Nagano T, Hayakawa H, Hirata Y, Hirobe M. Detection of Nitric Oxide Production from a Perfused Organ by a Luminol-H202 System. Analytical Chemistry 1993;65:1794-1799. 3. Blum LJ, Esperança P, Rocquefelte S. A new high-performance reagent and procedure for latent bloodstain detection based on luminol chemiluminescence. Canadian Society of Forensic Science Journal. 2006;39:81-100. Financial support from the Czech Science Foundation, project 301/11/0767. O0008 An In-depth study on Blue electrochemiluminescent Iridium(III) complexes Gregory Barbantea, Egan Doevena, Paul Francisa, Timothy Connellc, Paul Donnellyc, Conor Hoganb, David Wilsonb aDeakin University, Geelong, Victoria, Australia bLa Trobe University, Melbourne, Victoria, Australia cMelbourne University, Melbourne, Victoria, Australia Electrogenerated chemiluminescence (ECL) is a form of luminescence produced by high-energy reactions between electrogenerated precursors,[1,2] in which the electronically excited states responsible for the emission of light can be generated through the annihilation between oxidised and reduced forms of the same species, or by using a sacrificial co-reactant. The application of a co-reactant ECL as a highly sensitive mode of detection has been predominantly based on the use of tris(2,2'-bipyridine)ruthenium(II) ([Ru(bpy)3]2+), and related polyimine-ruthenium(II) complexes, with characteristic orange/red emissions (max ca. 590-700nm).[1,2] Over the last decade, however, numerous researchers have begun to explore chemiluminescence and ECL reactions with cyclometalated iridium(III) complexes exhibiting a wide range of electrochemical properties and emission maxima that can be tuned through subtle changes in the structure of one or more ligands.[3] These complexes have created new possibilities for multiplexed ECL detection systems.[4] However, in contrast to the vast range of orange/red-emitting metal complex electrochemiluminophores,[5,6] relatively few blue emitters are available, and the most effective design of blue-emitting complexes for ECL detection is yet to be fully elucidated. We have therefore used electrochemical, spectroscopic and computational techniques to explore a series of blue-emitting iridium(III) complexes (see Fig.) that exhibit various potentially attractive structural attributes for conventional and multiplexed ECL detection. Theoretical and experimental studies reveal the most effective strategies for the design of blue-shifted iridium(III) complexes for efficient electrogenerated chemiluminescence. Stabilisation of the HOMO while only moderately stabilising the LUMO increases the energy gap, thus ensuring favourable thermodynamics and kinetics for the reaction leading to the excited state. Of the iridium(III) complexes examined, [Ir(df-ppy)2(ptb)]+ was most attractive as a blue-emitter for ECL detection, featuring a large hypsochromic shift (max=454 and 484nm), superior co-reactant ECL intensity than the archetypal homoleptic green and blue emitters; [Ir(ppy)3] and [Ir(df-ppy)3] (by over 16-fold and threefold, respectively), and greater solubility in polar solvents. We have therefore used electrochemical, spectroscopic and computational techniques to explore a series of blue-emitting iridium(III) complexes (see Fig.) that exhibit various potentially attractive structural attributes for conventional and multiplexed ECL detection. Theoretical and experimental studies reveal the most effective strategies for the design of blue-shifted iridium(III) complexes for efficient electrogenerated chemiluminescence. Stabilisation of the HOMO while only moderately stabilising the LUMO increases the energy gap, thus ensuring favourable thermodynamics and kinetics for the reaction leading to the excited state. Of the iridium(III) complexes examined, [Ir(df-ppy)2(ptb)]+ was most attractive as a blue-emitter for ECL detection, featuring a large hypsochromic shift (max=454 and 484nm), superior co-reactant ECL intensity than the archetypal homoleptic green and blue emitters; [Ir(ppy)3] and [Ir(df-ppy)3] (by over 16-fold and threefold, respectively), and greater solubility in polar solvents. 1. Bard AJ. Electrogenerated Chemiluminescence. Marcel Dekker; New York, 2004 2. Forster RJ, Keyes TE. Neuromethods 2013;80:347-367 3. Zanarini S, Felici M, Valenti G, Marcaccio M, Prodi L, Bonacchi S, Contreras-Carballada P, Williams RM, Feiters MC, Nolte RJM, De Cola L, Paolucci F. Chem. Eur. J. 2011;17:4640-4647 4. Doeven EH, Zammit EM, Barbante GJ, Hogan CF, Barnett NW, Francis PS. Angew. Chem. 2012;124:4430-4433 5. Barbante GJ, Hogan CF, Wilson DJD, Lewcenko NA, Pfeffer FM, Barnett NW, Francis PS. Analyst 2011;136:1329-1338 6. Gorman BA, Francis PS, Barnett NW. Analyst 2006;131:616-639 O0009 Chaetopterus variopedatus tissue autofluorescence spectral characteristics Anna Belousovaa, Fyodor Kondrashovb, Maria Plyuschevab aMoscow State University, Biological Faculty, Invertebrate Zoology Department, Moscow, Russia bCentre de Regulació Genòmica (CRG), Barcelona, Spain Chaetopterus variopedatus is a sedentary marine polychaete that lives in a parchment U-shaped tube. It has a body with three distinct regions, each region contains different morphologically developed segments. [3] The polychaete is famous for being capable of emitting light with its epithelium and producing blue luminous mucous. Studies on anatomy and morphology of phenomenon of its epithelial bioluminescence have pointed out some special areas of intensity, such as notopodial structures of middle and posterior regions. [2] Referring to the chemical approach, previous research on the chemical compounds of Chaetopterus bioluminescent system have proved that the photoprotein takes part in the reaction. [1] As for the whole biochemical pathway of the Chaetopterus bioluminescence - it is still a question to solve. Photoproteins that are involved in the luminescence reaction are known for becoming fluorescent after the reaction of luminescence takes place. [1] Therefore, the distribution of the products of the bioluminescence reaction can be studied using the confocal light microscopy methods. The autofluorescence itself is usually relatively stable either continious, or intensive. The distribution of fluorescence of different wavelengths was observed with a confocal microscope on several cross-sections of the worm's body and on epithelium of various parts of the body. The specimens of the worms tissue exhibit fluorescence of some ranges of wavelengths - excited by lasers from 405 to 633nm. Confocal observations have shown that the UV excitation of a tissue results most efficiently in the strong fast-bleaching fluorescence in far-red (630nm) spectrum area. Purified far-red autofluorescent component which is supposed to be a part of bioluminescence reaction [4] is stored in the vesicles which can be visualised with the confocal microscopy. Though having a strong intensity, the far-red fluorescence is bleaching very fast which makes it more challenging to observe. References 1. Shimomura O. Bioluminescence; chemical principles and methods. - World Scientific, 2012. 2. Anctil M. The epithelial luminescent system of Chaetopterus variopedatus //Canadian Journal of Zoology. - Т. 57. - . 6. - С, 1979;1290-1310. 3. Harvey EN. Bioluminescence. - Academic Press, 1952. 4. Branchini BR, et al. Chemical analysis of the luminous slime secreted by the marine worm Chaetopterus (Annelida, Polychaeta) //Photochemistry and photobiology. - Т. 90. - .1. - С, 2014;247-251. O0010 Construction of lux operon of the ancestor of bioluminescent bacteria and proposal of evolutionary hypothetical theories on the origin and propagation of luminescent bacterial species Ramesh CH, Mohanraju R Pondicherry University, Pondicherry, India Using the processes, "Horizontal gene transfer (HGT) or Chromosomal exchange mechanism (CEM) or Acquisition, Plasmid DNA exchange, genetical processes like Interchromosomal rearrangements (ICR), and geological processes" we assembled lux genes together and constructed the luminescent bacterial ancestor that might have not yet been isolated or extincted in the biological evolution during the geological processes. The construction of lux operon of this luminescent ancestor was carried out with different lux genes such as the regulatory and structural genes, and with other genes which are flanking towards upstream and downstream of the lux operon of different luminescent bacterial species. The lux operon of this ancestor was constructed based on the approximate base pairs of different lux genes. The approximate order of lux operon is as follows luxZYLOPUMNQRSTICDABFEGH-ribEBHA. Where rib genes of rib operon are linked to downstream of the lux operon. Hypothetically it is possible to construct this luminescent bacteria ancestor, while we expect possibilities of finding of this luminescent ancestor. The processes "Insertion, Deletion, Horizontal gene transfer (HGT) or Chromosomal exchange mechanism (CEM) and Interchromosomal rearrangements (ICR), geological processes and Plasmid DNA exchange (PDE) might have given origin for modern luminescent bacteria. These processes provides base for this ancestor construction and supports for the presence or possibility in constructing the ancestor of luminescent bacteria. We also propose new strong hypothetical theories which speak about the existence of luminescent ancestor and origin of modern luminescent bacterial species. O0011 Ecological functions of shark luminescence Julien Claes, Jérôme Mallefet Laboratoire de Biologie Marine, Earth and Life Institute, Université catholique de Louvain, Louvain-la-Neuve, Belgium Introduction; Sharks from the Etmopteridae and Dalatiidae families are among the most enigmatic bioluminescent organisms. Although they encompass about 12% of current shark diversity, with over 50 described species, their luminescence is rarely observed [1]. Moreover, contrary to the situation encountered in other animals, their intrinsic light organs (photophores) are primarily controlled by hormones rather than by nerves [2,3] and form a diversity of patterns whose adaptive advantage has long remained obscure. This work aims to synthetize recent advances made in the field of shark luminescence ecology as well as to present novel experimental data in order to inspire future research. It involves various techniques such as in vivo luminescence measurements [via an optic fibre coupled to a luminometer (Berthold FB12)], spectrophotometry [with a mini-spectrometer (Hamamatsu Photonics C10083CA)], stereology and visual modelling. Results and discussion; In the last five years, we observed and characterized the spontaneous luminescence of one dalatiid and three etmopterid shark species. At 0.5 prepelvic length, their luminescence [downward emission; λmax at 457-488nm; ventral intensity=0.34-130.78 Mq s-1mm-2 (n=31)] appears physically similar to the residual downwelling light present in their environment, supporting a function of camouflage by counterillumination [1,4]. However, digital photography revealed that etmopterid luminescence (i) was not homogeneous on the ventral side (Fig.A), a result confirmed by stereological analysis of photophore distribution; and (ii) was also present dorsally, as dim glows underlying several structures including fin spines, eyes and nostril
View source
