Fluorescence Excitation Spectroscopy Research Articles

Characterization and C-, N-disinfection byproduct formation of dissolved organic matter in MBR and anaerobic-anoxic-oxic (AAO) processes

This study investigated distinguishing characteristic of dissolved organic matter (DOM) in MBR and AAO effluents by UF fractionation, XAD-8 resin adsorption isolation, excitation and emission matrix fluorescence spectroscopy and fourier transform infrared spectroscopy analysis. Halogenated carbonaceous and nitrogenous disinfection byproducts (DBPs) especially N-nitrosodimethylamine (NDMA) formation reactivity of various DOM fractions were also analyzed. Results showed that although the MBR effluent contained lower DOM than the AAO effluent, more halogenated C, N-DBPs in the MBR effluent were formed than in the AAO effluent after chlorination due to its higher percentage of molecular weight (MW)>1kDa materials, fulvic acid-like substances and polysaccharides. For example, the content of TCM in MBR effluent was high as 526μg/mg-C, which was about 13 times as that in AAO effluent. In contrast, the NDMA yield of AAO effluent was more than that of MBR effluent because AAO contained higher percentage of MW<1kDa materials, aromatic proteins, polypeptides and amino acids especially secondary amines.

CW Optical Limiting Study in Disperse Yellow Dye-doped PMMA-MA Polymer Films

&lt;div&gt;&lt;p&gt;&lt;em&gt;Molecules with high two-photon absorption (TPA) cross section and the molecules which have reverse saturation absorption (RSA) cross section are of great interest today because of their application in three-dimensional optical data storage and photonic switches. During last forty years, several techniques have been developed to determine the two-photon absorption cross-sections and reverse saturation absorption cross-sections of the materials. This includes direct methods, such as nonlinear transmission and Z-scan methods and indirect methods, such as two-photon excited fluorescence and two-photon pump-probe transient absorption spectroscopy. During recent years, the organic dye-doped polymer films getting more attention due to their advantages to fabricate photonics devices. In this paper, we have studied the nonlinear optical properties like nonlinear absorption and nonlinear refraction of an azo dye Disperse yellow-7 (DY-7) doped in Polymethyl methacrylate-methacrylic acid (PMMA-MA) polymer matrix using open aperture and closed aperture Z-scan experimental methods using continuous wave (CW) laser. The optical limiting properties of these films are also studied in Type 1 and Type 2 configurations at different input power using continuous wave (CW) laser beams of 532 nm wavelength.&lt;/em&gt;&lt;/p&gt;&lt;/div&gt;

Structure-Dependent Electronic Interactions in Ethyne-Bridged Porphyrin Arrays Investigated by Single-Molecule Fluorescence Spectroscopy.

By using single-molecule fluorescence spectroscopy, we have investigated the electronic interaction of ethyne-bridged porphyrin arrays (ZNE) depending on their structure. The fluorescence dynamics of ZNE show a large amount of one-step photobleaching behaviors, indicating the high degree of π-conjugation. The ratio of one-step photobleaching behavior decreased as the number of porphyrin units increased. This behavior indicates that the linear and shortest Z2E shows a strong electronic coupling between constituent porphyrin moieties. Structural properties and orientation of ZNE were also measured by wide-field excitation fluorescence spectroscopy (ExPFS) and defocused wide-field imaging (DWFI). The ExPFS and DWFI show that the structure of absorbing and emitting units of Z2E and Z3E are linear. On the other hand, star-shaped pentamer with five porphyrins acts as an absorbing unit, but unidirectional trimer moiety acts as an emitting unit in the Z5E molecule. Collectively, these studies provide further information on the electronic interaction depending on their structure and length.

Reduction of disinfection by-product precursors in reservoir water by coagulation and ultrafiltration.

In this study, reservoir water intended for drinking water supply was treated by (i) ultrafiltration (UF) (ii) coagulation (CW) (iii) coagulation combined with ultrafiltration (CW-UF). To probe the influences of three treatment processes on disinfection byproduct (DBP) precursors in source water, the changes of dissolved organic matter (DOM) amounts and physicochemical properties, and disinfection byproduct (DBP) formation characteristics during chlorine disinfection were investigated. Both carbonaceous DBP (C-DBP) and nitrogenous DBP (N-DBP) formation and speciation were analyzed. The influence of chlorine dose, contact time on DBP formation and speciation were also studied to optimize the disinfection conditions to minimize the DBP formation. Compared with UF and CW alone, CW-UF improved the dissolved organic carbon (DOC) removal from about 20% to 59%. The three-dimensional excitation and emission matrix (3DEEM) fluorescence spectroscopy analysis showed that CW-UF had high removal efficiency in microbial products (Region IV), fulvic acid-like (Region III) and humic acid-like (Region V). The total C-DBP was determined by the formation of trihalomethanes and trichloromethane was the most abundant species (40%). The most abundant N-DBP species was dichloroacetonitrile (32.5%), followed by trichloroactetonitrile. CW-UF effectively reduced the risk of DBPs in drinking water supply by reducing 30.8% and 16.9% DBPs formation potential compared with UF and CW alone. Increasing contact time improved the yields of both C-DBPs and N-DBPs. Chlorine dosage had slight influence on DBP yield in this study.

Fluorescence-excitation and Emission Spectroscopy on Single FMO Complexes.

In green-sulfur bacteria sunlight is absorbed by antenna structures termed chlorosomes, and transferred to the RC via the Fenna-Matthews-Olson (FMO) complex. FMO consists of three monomers arranged in C3 symmetry where each monomer accommodates eight Bacteriochlorophyll a (BChl a) molecules. It was the first pigment-protein complex for which the structure has been determined with high resolution and since then this complex has been the subject of numerous studies both experimentally and theoretically. Here we report about fluorescence-excitation spectroscopy as well as emission spectroscopy from individual FMO complexes at low temperatures. The individual FMO complexes are subjected to very fast spectral fluctuations smearing out any possible different information from the ensemble data that were recorded under the same experimental conditions. In other words, on the time scales that are experimentally accessible by single-molecule techniques, the FMO complex exhibits ergodic behaviour.

Advanced landfill leachate treatment using iron-carbon microelectrolysis- Fenton process: Process optimization and column experiments

A novel hydrogen peroxide-enhanced iron-carbon (Fe-C) microelectrolysis reactor was proposed for the pretreatment of mature landfill leachate. This reactor, combining microelectrolysis with Fenton process, revealed high treatment efficiency. The operating variables, including Fe-C dosage, H2O2 concentration and initial pH, were optimized by the response surface methodology (RSM), regarding the chemical oxygen demand (COD) removal efficiency and biochemical oxygen demand: chemical oxygen demand (BOD5/COD) as the responses. The highest COD removal (74.59%) and BOD5/COD (0.50) was obtained at optimal conditions of Fe-C dosage 55.72g/L, H2O2 concentration 12.32mL/L and initial pH 3.12. Three-dimensional excitation and emission matrix (3D-EEM) fluorescence spectroscopy and molecular weight (MW) distribution demonstrated that high molecular weight fractions such as refractory fulvic-like substances in leachate were effectively destroyed during the combined processes, which should be attributed to the combination oxidative effect of microelectrolysis and Fenton. The fixed-bed column experiments were performed and the breakthrough curves at different flow rates were evaluated to determine the practical applicability of the combined process. All these results show that the hydrogen peroxide-enhanced iron-carbon (Fe-C) microelectrolysis reactor is a promising and efficient technology for the treatment of mature landfill leachate.

Structure of Light-Harvesting Aggregates in Individual Chlorosomes.

Among all photosynthetic organisms, green bacteria have evolved one of the most efficient light-harvesting antenna, the chlorosome, that contains hundreds of thousands of bacteriochlorophyll molecules, allowing these bacteria to grow photosynthetically by absorbing only a few photons per bacteriochlorophyll molecule per day. In contrast to other photosynthetic light-harvesting antenna systems, for which a protein scaffold imposes the proper positioning of the chromophores with respect to each other, in chlorosomes, this is accomplished solely by self-assembly. This has aroused enormous interest in the structure-function relations of these assemblies, as they can serve as blueprints for artificial light harvesting systems. In spite of these efforts, conclusive structural information is not available yet, reflecting the sample heterogeneity inherent to the natural system. Here we combine mutagenesis, polarization-resolved single-particle fluorescence-excitation spectroscopy, cryo-electron microscopy, and theoretical modeling to study the chlorosomes of the green sulfur bacterium Chlorobaculum tepidum. We demonstrate that only the combination of these techniques yields unambiguous information on the structure of the bacteriochlorophyll aggregates within the chlorosomes. Moreover, we provide a quantitative estimate of the curvature variation of these aggregates that explains ongoing debates concerning the chlorosome structure.

Evaluation of cell and matrix mechanics using fluorescence excitation spectroscopy: Feasibility study in collagen gels containing fibroblasts.

Collagen gels containing cells are commonly used in tissue engineering, wound healing, and cancer research to investigate the interplay between cells and the extracellular matrix (ECM), as changes in the density and stiffness of the microenvironment are known to play a role in many diseases or pathological conditions. In these gels, the stiffness is regularly determined using destructive methods, such as indentation and tensile tests. Certain molecules native to cells and the ECM display fluorescence upon irradiation with ultraviolet light. The objective of the present study was to investigate the feasibility of using the endogenous, or innate, fluorescence of collagen gels containing fibroblasts as an optical marker to evaluate changes in the mechanical properties of the ECM. Human foreskin fibroblasts cells at concentrations of 50,000 and 100,000 cells/ml were cultured in three-dimensional gels of type I collagen for 16 days. Fibroblast cells remodeled the ECM, contracting and increasing the stiffness of the gel. During this remodeling process, changes in mechanical properties and fluorescence were measured with an indentation test and a spectrofluorometer, respectively. Force and displacement measurements from the indentation test were used to calculate the elastic modulus of the gel. Maps of fluorescence intensity, at excitation/emission of 240-520/290-530 nm, were used to identify the wavelengths of interest. Fluorescence excitation/emission maps exhibited two distinct excitation/emission bands whose intensities increased as the fibroblasts remodeled and increased the stiffness of the ECM: The 290/340 nm band ascribed to tryptophan and the 330/390 nm band ascribed to cross-links of collagen (pepsin-digestible cross-links). A Spearman correlation analysis, between the elastic modulus of the gel containing fibroblasts and the fluorescence of cross-links of collagen, resulted in R = 0.95 (P < 0.05) and R = 0.77 (P = 0.12) for 50,000 and 100,000 cells/ml, respectively. The endogenous fluorescence intensity ascribed to pepsin-digestible cross-links of collagen may serve as an optical marker to evaluate changes in the mechanical properties of the ECM; this is relevant to collagenous tissues for which pathological states are related to mechanical alterations, such as keratoconus in cornea and osteoarthritis in articular cartilage.

Review of applications of fluorescence excitation spectroscopy to dermatology.

Endogenous molecules that exhibit fluorescence hold the potential to serve as reporters of tissue structure, activity and physiology. Fluorescence excitation spectroscopy is one means to measure and express tissue's innate fluorescence. This review focuses on the application of endogenous fluorescence ultraviolet excitation spectroscopy to dermatology.

An intact light harvesting complex I antenna system is required for complete state transitions in Arabidopsis.

Efficient photosynthesis depends on maintaining balance between the rate of light-driven electron transport occurring in photosystem I (PSI) and photosystem II (PSII), located in the chloroplast thylakoid membranes. Balance is achieved through a process of 'state transitions' that increases energy transfer towards PSI when PSII is overexcited (state II), and towards PSII when PSI is overexcited (state I). This is achieved through redox control of the phosphorylation state of light-harvesting antenna complex II (LHCII). PSI is served by both LHCII and four light-harvesting antenna complex I (LHCI) subunits, Lhca1, 2, 3 and 4. Here we demonstrate that despite unchanged levels of LHCII phosphorylation, absence of specific Lhca subunits reduces state transitions in Arabidopsis. The severest phenotype-observed in a mutant lacking Lhca4 (ΔLhca4)-displayed a 69% reduction compared with the wild type. Yet, surprisingly, the amounts of the PSI-LHCI-LHCII supercomplex isolated by blue native polyacrylamide gel electrophoresis (BN-PAGE) from digitonin-solubilized thylakoids were similar in the wild type and ΔLhca mutants. Fluorescence excitation spectroscopy revealed that in the wild type this PSI-LHCI-LHCII supercomplex is supplemented by energy transfer from additional LHCII trimers in state II, whose binding is sensitive to digitonin, and which are absent in ΔLhca4. The grana margins of the thylakoid membrane were found to be the primary site of interaction between this 'extra' LHCII and the PSI-LHCI-LHCII supercomplex in state II. The results suggest that the LHCI complexes mediate energetic interactions between LHCII and PSI in the intact membrane.

Paclitaxel conjugated Fe3O4@LaF3:Ce3+,Tb3+ nanoparticles as bifunctional targeting carriers for Cancer theranostics application

The bi-functional Chitosan functionalized magnetite doped luminescent rare earth nanoparticles (Fe3O4@LaF3: Ce3+,Tb3+/chi NPs) as a carrier of paclitaxel (PTX) drug was designed using a co-precipitation and facile direct precipitation method. The synthesized nanoparticles are spherical in shape with a typical diameter of 19–37nm respectively. They are water soluble, super paramagnetic and biocompatible, in which the amino groups on the nanoparticles surface are used for the conjugation with an anticancer drug, paclitaxel. The nature of PTX binding with Fe3O4@LaF3: Ce3+,Tb3+/chi nanoparticles were studied using X-ray diffraction, vibrating sample magnetometer and scanning electron micrograph. The nature of interactions between PTX and Fe3O4@LaF3: Ce3+,Tb3+/chi NPs due to complex formation were conceded out by various spectroscopic methods viz., UV–visible, steady state and excited state fluorescence spectroscopy. The photo-physical characterization reveals that the adsorption and release of PTX from Fe3O4@LaF3:Tb3+/chi nanoparticles is quicker when compared with other nanoparticles and also confirms that this may be due to the hydrogen bond formation between the hydroxyl group of drug and amino group of nanoparticles respectively. The maximum loading capacity and entrapment efficiency of 83.69% and 80.51% were attained at a ratio of 5:8 of PTX and Fe3O4@LaF3: Ce3+,Tb3+/chi NPs respectively. In addition with that, antitumoral activity study of PTX conjugated Fe3O4@LaF3:Tb3+/chi nanoparticles exhibits increased cytotoxic effects on A549 lung cancer cell lines than that of unconjugated PTX.

Chirality-dependent mechanical response of empty and water-filled single-wall carbon nanotubes at high pressure

The mechanical stability of single-wall carbon nanotubes (SWCNTs) at high pressure was studied by high-resolution resonant Raman and wavelength-dependent fluorescence-excitation (PLE) spectroscopy resolving the vibrational and electronic resonances of 18 individual chiralities and furthermore even resolving the different behaviour of empty (closed, pristine) and water-filled (opened) SWCNTs (diameter range = 0.6–1.42 nm). We find that water-filling exerts a stabilizing counter-pressure on the SWCNT walls, leading to an increasing difference between the radial breathing mode frequencies of water-filled and empty SWCNTs at elevated pressures. For small diameter SWCNTs (d < 1 nm) with a chiral angle of ∼12°, in particular for the (7,2) chirality, an anomalous behaviour is observed, revealing an increased mechanical instability for these SWCNTs. We furthermore ascribe the longstanding contradiction between experiments and theory on the collapse pressure of SWCNTs to the presence of filling in most experiments to date, while empty SWCNTs follow the theoretically predicted collapse behaviour.

Moving in on the Action: An Experimental Comparison of Fluorescence Excitation and Photodissociation Action Spectroscopy.

Photodissociation action spectroscopy is often used as a proxy for measuring gas-phase absorption spectra of ions in a mass spectrometer. Although the potential discrepancy between linear optical and photodissociation spectra is generally acknowledged, direct experimental comparisons are lacking. In this work, we use a quadrupole ion trap that has been modified to enable both photodissociation and laser-induced fluorescence to assess how closely the visible photodissociation action spectrum of a fluorescent dye reflects its fluorescence excitation spectrum. Our results show the photodissociation action spectrum of gaseous rhodamine 110 is both substantially narrower and slightly red-shifted (∼120 cm(-1)) compared to its fluorescence excitation spectrum. Power dependence measurements reveal that the photodissociation of rhodamine 110 requires, on average, the absorption of three photons whereas fluorescence is a single-photon process. These differing power dependences are the key to interpreting the differences in the measured spectra. The experimental results provide much-needed quantification and insight into the differences between action spectra and linear optical spectra, and emphasize the utility of fluorescence excitation spectra to provide a more reliable benchmark for comparison with theory.

Trihalomethanes (THMs) precursor fractions removal by coagulation and adsorption for bio-treated municipal wastewater: Molecular weight, hydrophobicity/hydrophily and fluorescence

Due to concerns over health risk of disinfection byproducts (DBPs), removal of trihalomethanes (THMs) precursor from bio-treated wastewater by coagulation and adsorption was investigated in this study. Ultrafiltration (UF) membranes and nonionic resins were applied to fractionate THMs precursor into various molecular weight (MW) fractions and hydrophobic/hydrophilic fractions. Characteristics of coagulated water and adsorbed water were evaluated by the three-dimensional excitation and emission matrix (3DEEM) fluorescence spectroscopy. Results showed that coagulation and adsorption were suitable for removing different hydrophobic/hydrophilic and fluorescent fractions. Coagulation decreased THMs concentration in hydrophobic acids (HoA) fraction from 59μg/L to 39μg/L, while the lowest THMs concentration (9μg/L) in hydrophilic substances (HiS) fraction was obtained in adsorbed water. However, both coagulation and adsorption were ineffective for removing fractions with MW<5kDa. Although coagulation and adsorption processes could reduce THMs formation, some specific THMs formation potential (STHMFP) in residual dissolved organic matter (DOM) fractions increased in this study. Hydrophobic acid and hydrophilic fractions increased after coagulation treatment, and low MW and hydrophobic fractions increased after adsorption treatment. In addition, active carbon adsorbed more organic matter than coagulant, but brominated disinfection byproducts (Br-DBPs) in adsorbed water turned to the major THMs species after chlorination.

Bench-top aqueous two-phase extraction of isolated individual single-walled carbon nanotubes

Isolation and purification of single-walled carbon nanotubes (SWCNTs) are prerequisites for their implementation in various applications. In this work, we present a fast (∼5 min), low-cost, and easily scalable bench-top approach to the extraction of high-quality isolated SWCNTs from bundles and impurities in an aqueous dispersion. The extraction procedure, based on aqueous two-phase (ATP) separation, is widely applicable to any SWCNT source (tested on samples up to 1.7 nm in diameter) and independent of defect density, purity, diameter, and length. The extracted dispersions demonstrate that the removal of large aggregates, small bundles, and impurities is comparable to that by density gradient ultracentrifugation, but without the need for high-end instrumentation. Raman and fluorescence-excitation spectroscopy, single-nanotube fluorescence imaging, atomic force and transmission electron microscopy, and thermogravimetric analysis all confirm the high purity of the isolated SWCNTs. By predispersing the SWCNTs without sonication (only gentle stirring), full-length, pristine SWCNTs can be isolated (tested up to 20 μm). Hence, this simple ATP method will find immediate application in the generation of SWCNT materials for all levels of nanotube research and applications, from fundamental studies to high-performance devices.

Fabrication of fluorescent holographic micropatterns based on the rare earth complexes using azobenzene-containing poly(aryl ether)s as macromolecular ligands

In this work, on the basis of photoinduced surface relief gratings (SRGs) with the rare earth complexes using azo-polymers as macromolecular ligands, a series of novel materials for fabricating rewritable fluorescent two-dimensional micropatterns, whose color can be easily adjusted by changing the species of the rare earth ions, are demonstrated. The rare earth complexes are prepared using a series of poly(aryl ether)s containing azobenzene chromophores and carboxyl group as macromolecular ligands and 1,10-phenanthroline as co-ligands. The fluorescence properties of the rare earth complexes and the influence of the contents of azobenzene chromophores on the fluorescent intensity are investigated by means of fluorescence excitation and emission spectroscopy. By exposing the films of the rare earth complexes to an interference pattern laser beam, SRGs can be formed on the films. Under the excitation, fluorescent patterns of the SRGs can be observed by the measurement of fluorescence microscopy. © 2015 Wiley Periodicals, Inc. J. Polym. Sci. Part A: Polym. Chem. 2015, 53, 936–943

Intramolecular proton transfer in 2-(2'-hydroxyphenyl)oxazolo[4,5-b]pyridine: evidence for tautomer in the ground state.

The intramolecular proton transfer in a newly synthesized molecule, 2-(2'-hydroxyphenyl)oxazolo[4,5-b]pyridine (HPOP) is studied using UV-visible absorption, fluorescence emission, fluorescence excitation and time-resolved fluorescence spectroscopy. In the ground state, the molecule exists as cis- and trans-enol in all the solvents. However, in dioxane, alcohols, acetonitrile, dimethylformamide and dimethylsulfoxide the keto tautomer is also observed in the ground state. Dual fluorescence is observed in HPOP where the large Stoke shifted emission is due to emission from the excited-state intramolecular proton transfer product, whereas the other emission is the normal emission from enol form. The fluorescence (both normal and tautomer emission) of HPOP is less than those of corresponding benzoxazole and imidazopyridine derivatives. This reveals that the nonradiative decay becomes more efficient upon substitution of electronegative atom on the charge acceptor group. The pH studies substantiate the conclusion that (unlike in its imidazole analog) the third ground state species is the keto tautomer and not the monoanion. The effect of temperature on cis-enol-trans-enol-keto equilibrium and the nonradiative deactivation from the excited state are also investigated.

Native fluorescence spectroscopic characterization of DMBA induced carcinogenesis in mice skin for the early detection of tissue transformation.

The objective of the study is to characterize the endogenous porphyrin fluorescence in a dimethylbenz(a)anthracene (DMBA) induced mouse skin tumor model using native fluorescence emission and excitation spectroscopy. Two intensity ratio parameters I580/I635 and I420/I515 were selected to represent the key fluorophore of endogenous porphyrins from emission and excitation spectra recorded in vivo from 31 DMBA treated animals and 5 control animals. In the emission spectrum, the endogenous porphyrin was elevated at 635 nm in different transformation lesions such as hyperplasia, papilloma, dysplasia, ESCC and WDSCC. This is corroborated by the endogenous porphyrin elevation at 420, 515, 550 and 588 nm in the WDSCC lesions from the excitation spectra. The elevation of endogenous porphyrin, probably protoporphyrin IX (PpIX), is due to biochemical and metabolic alterations in epithelial cells during tissue transformation. The loss of ferrochelatase activity might be responsible for enhanced PpIX in the transformed tissues. The sensitivity and specificity were determined for different lesion pairs from the scatter plot based on the discrimination value by validation with histopathological results. The emission intensity ratio I580/I635 at 405 nm excitation was selected to discriminate normal from hyperplasia, hyperplasia from papilloma, papilloma from dysplasia, dysplasia from early squamous cell carcinoma (ESCC), and ESCC from well differentiated squamous cell carcinoma (WDSCC) with specificities of 100%, 88%, 100%, 86%, and 100% and sensitivities of 100%, 80%, 100%, 100% and 100% respectively. Similarly, the excitation intensity ratio I420/I515 for 635 nm emission used to discriminate between WDSCC lesions and normal tissue gives 100% specificity and 100% sensitivity.

Study of NaCl:Mn2+ nanostructures in the Suzuki phase by optical spectroscopy and atomic force microscopy

NaCl:Mn2+ nanostructures in the Suzuki phase have been studied by fluorescence (emission and excitation) spectroscopy and atomic force microscopy (AFM) as a function of temperature. The “as-grown” samples give rise to two broad emission bands that peak at 508 (green emission) and 610nm (red emission). The excitation spectrum shows peaks at 227nm and 232nm for emission wavelengths at 508nm and 610nm, respectively. When the samples are heated continuously from room temperature up to 220°C, the green emission (associated to the excitation peak at 227nm) disappears at a temperature close to 120°C, whilst only the red emission remains, which is characteristic of manganese ions. AFM images on the (001) surface (freshly cleaved) show several conformations of nanostructures, such as disks of 20–50nm in diameter. Particularly, the images also reveal nanostructures with rectangular shape of ~280×160nm2 and ~6nm height; these are present only in samples with green emission associated to the Suzuki phase. Then, the evidence suggests that this topographic configuration might be related to the interaction with the first neighbors and the next neighbors, according to the configuration that has been suggested for the Suzuki phase.

Relationship between soluble microbial products (SMP) and effluent organic matter (EfOM): Characterized by fluorescence excitation emission matrix coupled with parallel factor analysis

Effluent organic matter (EfOM) originating from wastewater treatment plant (WWTP) is of significant concern, as it not only influences the discharge quality of WWTP but also exerts a significant effect on the efficiency of the downstream advanced treatment facilities. Soluble microbial products (SMP) is a major part of EfOM. In order to further understand the relationship between soluble microbial products (SMP) and EfOM, and in turn, to propose measures for EfOM control, the formation of SMP and EfOM in identical activated sludge sequencing batch reactors (SBR) with different feed water was investigated using fluorescence excitation and emission spectroscopy matrix coupled with parallel factor analysis (EEM–PARAFAC) as well as other organic matter quantification tools. Results showed that EfOM contained not only SMP but also a considerable amount of allochthonous organic matter that derived not merely from natural organic matter (NOM). Four components in EfOM/SMP were identified by EEM–PARAFAC. Tyrosine-like substances in EfOM (Component 3, λex/em=270/316nm) were mainly originated from utilization associated products (UAP) of SMP. Tryptophan-like substances (Component 2, λex/em=280/336nm) as well as fulvic-like and humic-like substances in EfOM (Component 1, λex/em=240(290)/392nm and Component 4, λex/em=260(365)/444nm) were majorly derived from the refractory substances introduced along with the influent, among which Component 2 was stemmed from sources other than NOM. As solid retention time (SRT) increased, Component 2 and polysaccharides in SMP/EfOM decreased, while Component 4 in SMP increased.