Excitation Emission Spectroscopy Research Articles

Capability of detection and three-way data

Due to the possibility of making analytical determinations in the presence of non-modelled interferents and to identify the analyte of interest, calibrations based on scores of PARAFAC decomposition of three-way data are becoming increasingly important in routine analysis. Furthermore, the IUPAC and EU (European Decision 2002/657/EC) have accepted the definition given by the ISO 11843 for the capability of detection as the minimum net quantity detectable with a pre-set probability of false positive and false negative. What is more, recently our research group has generalised this definition of capability of detection, CCβ, to multivariate calibrations. In practice, CCβ is a good measure of the quality of the calibration because in its definition it brings together analytical sensitivity with precision in analytical determinations. This paper studies the effect of the pre-treatment of the sample, the signal/noise ratio and the second-order advantage on CCβ when using second-order signals modelled by PARAFAC. All of them are experimental factors which influence the quality of the calibration. Analytical pre-treatment is habitual in the analysis of real samples. Specifically, we analyse the effect of the extraction phase and the clean-up of milk samples on the determination of chlortetracycline by HPLC-DAD. It is shown that it is more efficient to do the joint PARAFAC decomposition of the pure standards with the milk samples. Secondly, the effect of asymmetry on CCβ, according to the path of the noise of the signals, is studied. Specifically, in the determination of naphthalene by excitation-emission spectroscopy, EEM, it is the emission spectrum which limits the capability of detection. It is shown that by eliminating the spectra with the poorest signal/noise ratio in this path, the capability of detection can be substantially improved. Thirdly, the impact on CCβ when the second-order advantage is used, that is when PARAFAC calibration is used over samples with an unknown interference not modelled in the calibration step. This is important to apply a PARAFAC calibration to routine analysis in the IUPAC and European Decision framework. Specifically, in the determination of enrofloxacine in poultry feeding water through excitation-emission fluorescence CCβ is evaluated when the PARAFAC is built only with calibration samples or with the calibration samples plus the test samples with uncalibrated and unknown interferent.

Laser-induced fluorescence and single vibronic level emission spectroscopy of chiral (R)-1-aminoindan and some of its clusters in a supersonic jet

Two low energy conformers of the chiral (R)-1-aminoindan molecule are identified in supersonic jet and their ground and excited states vibrational spectroscopy has been investigated by laser-induced fluorescence (LIF) excitation and single vibronic level (SVL) emission spectroscopy. Ab initio calculations confirm the existence of two lowest-energy structures, where the amino group is in equatorial position with its lone pair directed opposite to the aromatic electron cloud. Harmonic frequencies have been calculated for these two conformers at the DFT level with B3LYP functional. A low-frequency progression of 118 cm(-1) and 114 cm(-1), respectively, appears in the fluorescence excitation spectrum of the two conformers, with its ground state counterpart at approximately 147 cm(-1). It has been assigned to the puckering motion coupled with the ring flapping mode. The other calculated low-frequency mode corresponds to the puckering motion coupled with the ring twisting mode and its ground state frequency has been observed at 119 cm(-1) and 111 cm(-1) from SVL spectra. Both conformers form similar 1 : 1 water clusters, whose 0-0 transitions are shifted to the blue by 41 cm(-1) and 44 cm(-1), respectively, and whose SVL spectra are similar. Interestingly, one of the conformers seems to preferentially make complexes with (S)-methyllactate, while the other one shows selective complexation to (R)-methyllactate.

Concentration of colloidal silica suspensions using fluorescence spectroscopy

A simple detection method based on fluorescence excitation-emission spectroscopy has been developed to quantify the concentration of Ludox colloidal silica nanoparticles having diameters ranging from 7 to 22 nm. The technique works for suspensions of nominally negatively (SM30, LS, and TM40) and positively (CL) charged colloid silica particles. Without pretreatment, the method detection limits are 1.4 mg/L SiO 2 for CL (12 nm) and TM40 (22 nm) suspensions and 12 mg/L SiO 2 for SM30 (7 nm) and LS (12 nm) suspensions at an excitation wavelength of 308 nm and an emission wavelength of 318 nm. The fluorescence intensities of all the colloidal silica particle suspensions are linear ( R 2 > 0.98) with concentration in the range of 0–300 mg/L SiO 2. The fluorescence intensity of the negatively charged particle suspensions is constant between pH 3 and 9. The fluorescence intensity of suspensions of particles that are nominally positively charged is constant above pH 5.5, where the particles become negatively charged.

Investigation of organic matter dynamics during in-vessel composting of an aged coal–tar contaminated soil using fluorescence excitation–emission spectroscopy

In-vessel composting of an aged coal–tar contaminated soil from a manufactured gas plant site was investigated over 98 days using laboratory-scale in-vessel composting reactors. The composting reactors were operated at 18 different operational conditions using a logistic three-factor factorial design with three temperatures ( T = 38, 55 and 70 °C), four soil to green waste ratios (S:GW; 0.6:1, 0.7:1, 0.8:1 and 0.9:1 on a dry weight basis) and three moisture contents (MC; 40%, 60% and 80%). Excitation–emission matrix (EEM) fluorescence spectroscopy was used to investigate organic matter dynamics in the composting mixture. The results of this investigation indicated that formation of humic substances can be monitored by fluorescence excitation–emission matrix, and provided evidence of progressive mineralization or humification of the composting mixture. Peak excitation wavelength shifts and peak fluorescence intensity can both be used as indicators to monitor the humification or maturation of compost. Finally, the fluorescence index can be applied to investigate the origin of humic substances and fulvic acids, and the humification or maturation of compost.

Multistep Resonant Excitation of Erbium Ions in Thin Silicon Oxide Layers

AbstractUsing the site-selective technique of combined excitation emission spectroscopy, we have studied a variety of Er doped silicon oxide layers with and without silicon nanocrystals. This technique allows clear distinction of cluster defect sites that are created during thermal annealing and become dominant for higher Er ion concentration and are suppressed by the presence of nanocrystals. In several samples, we were able to observe fluorescence line narrowing under resonant excitation allowing defect-selective excitation.

Combined Excitation Emission Spectroscopy of Europium ions in GaN and AlGaN Films

AbstractSite-selective combined excitation emission spectroscopy studies have been performed on Eu-doped GaN and numerous sites have been identified. Relative numbers and broadening of these peaks has been investigated for different growth conditions and for increasing AL content of the AlxGa1-xN alloy.

Combined excitation‐emission spectroscopy of colour centres in lithium fluoride crystals

The spectral identification of several photoluminescent defects produced in LiF by ionising radiation is still uncertain because many optical transitions are possible and their different spectral features are often overlapping. Performing a conventional spectroscopic characterization requires a lot of time and intuition, but recently a more effective investigation method has been developed: combined excitation-emission spectroscopy (CEES). In this approach a large number of excitation (or emission) spectra is recorded at a sequence of emission (or excitation) wavelengths. The resulting three-dimensional data set of excitation intensities as a function of excitation and emission wavelengths, or energies, is visualized using a contour plot, where lines of equal emission intensities are drawn. For the first time this method has been successfully applied to coloured LiF crystals and films. As an example, the case of visible-emitting colour centres, absorbing in the M band spectral region in a gamma irradiated LiF crystal is reported and discussed. Spectroscopic features of well-known active defects have been confirmed, more complex centres clearly identified and other properties better clarified. (© 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

Changes in CDOM fluorescence from allochthonous and autochthonous sources during tidal mixing and bacterial degradation in two coastal estuaries

Chromophoric dissolved organic matter (CDOM) was collected and concentrated using 1 kDa cutoff tangential flow filtration (TFF) from marine (∼33 salinity), mid-estuarine (∼15 salinity), and freshwater (<1 salinity) portions of the Chesapeake and San Francisco Bays. Natural bacterioplankton were also collected during the same transects on 0.22-μm pore size filters. TFF permeates from freshwater, mid-estuarine and marine stations were used to create a series of salinity samples ranging from 0 to 33 by increments of 3. Freshwater CDOM was added in the same proportion to each salinity sample to determine changes in spectral signals during simulated estuarine mixing. A series of incubations was conducted in which concentrated CDOM was added to TFF permeates (<1 kDa, low fluorescence) in a nine-membered matrix such that each station's CDOM was added to each station's TFF permeate. Each incubation was then inoculated with a filter from its respective collection location. Subsamples from bacterial incubations were collected at various times and analyzed by high resolution three-dimensional fluorescence excitation–emission spectroscopy (EEMs) to determine if changes in ionic strength encountered during estuarine mixing affect the bioavailability and optical properties of CDOM. Five EEMs peaks were identified for each mixing experiment and microbial subsample; Ex max: 330–350 nm/ Em max: 420–480 nm, Ex max: 250–260 nm/ Em max: 380–480 nm, Ex max: 310–320 nm/ Em max: 380–420 nm, Ex max: 270–280 nm/ Em max: 300–320 nm, and Ex max: 270–280 nm/ Em max: 320–350 nm. These peak ratios were monitored over the time course of the experiment. Changes in several spectral properties during the simulated estuarine mixing were observed indicating CDOM conformational changes as it moves through the estuary. We hypothesized these changes may impact the biodegradability of CDOM as it moves from upland sources to the coastal ocean. Changes in DOC concentration during incubation indicated that allochthonous CDOM was a more utilizable substrate for estuarine and marine bacteria. There were also differences in peak ratios observed during incubation with allochthonous and autochthonous CDOM. There were Em max peak shifts dependent on the source of CDOM and bacteria, with more red shifting (toward higher wavelengths) in upper reaches of the estuary and more blue-shifting at the oceanic end-member. We conclude that bacterial degradation of specific components of autochthonous and allochthonous CDOM may impact the spectral characteristics observed throughout an estuary and that CDOM optical properties are partially a function of the CDOM's origin and mixing history.

Site-selective spectroscopy of Er in GaN

We investigated different Er3+ defect sites found in Er-doped GaN layers by site-selective combined excitation-emission spectroscopy and studied the role of these sites in different direct and multistep excitation schemes. The layers were grown by molecular beam epitaxy and were 200 nm thick. Two majority sites were found along with several minority sites. The sites strongly differ in excitation and energy transfer efficiencies as well as branching ratios during relaxation. For this reason, relative emission intensities from these sites depend strongly on emission and excitation. The sites were identified for several transitions and a comprehensive list of energy levels has been compiled. One of the minority sites appears strongly under ultraviolet excitation above the GaN band gap suggesting that this site is an excellent trap for excitation energy of electron-hole pairs.

Static and time-resolved fluorescence investigations of tryptophan analogues--a solvent study.

The fluorescence properties of tryptophan, polytryptophan and seven of its analogues (7-azatryptophan, 5-hydroxytryptophan, 5-methoxytryptophan, 5-fluorotryptophan, 5-methyltryptophan, 5-bromotryptophan, and 6-fluorotryptophan) are studied using two novel fluorescence spectroscopic techniques for a wide range of solvent polarities. Two-dimensional mapping of all emission and all fluorescence spectra using excitation-emission spectroscopy (EES) has been used to determine quantum yields, positions of emission maxima, full widths at half maximum (FWHMs) as well as Stokes' shifts. Additionally, fluorescence lifetimes obtained from time-resolved experiments using a picosecond laser system are presented and compared with the data acquired from the static setup. This systematic study of the fluorescence characteristics is a prerequisite to assess the potential of these analogues to act as structure-conserving substitutes for tryptophan in protein fluorescence experiments. The potential of these analogues, to act as probes for the local environment, and allow estimation of the polarity in the vicinity of the fluorophore and its exposure to the solvent, is discussed.

Tracing dissolved organic matter in aquatic environments using a new approach to fluorescence spectroscopy

Dissolved organic matter (DOM) is a complex and poorly understood mixture of organic polymers that plays an influential role in aquatic ecosystems. In this study we have successfully characterised the fluorescent fraction of DOM in the catchment of a Danish estuary using fluorescence excitation–emission spectroscopy and parallel factor analysis (PARAFAC). PARAFAC aids the characterisation of fluorescent DOM by decomposing the fluorescence matrices into different independent fluorescent components. The results reveal that at least five different fluorescent DOM fractions present (in significant amounts) in the catchment and that the relative composition is dependent on the source (e.g. agricultural runoff, forest soil, aquatic production). Four different allochthonous fluorescent groups and one autochthonous fluorescent group were identified. The ability to trace the different fractions of the DOM pool using this relatively cheap and fast technique represents a significant advance within the fields of aquatic ecology and chemistry, and will prove to be useful for catchment management.

Ferroelectric domain imaging by defect-luminescence microscopy

In order to study the role of defects within the domain inversion process in ferroelectric LiNbO3 crystals, we investigated the optical properties of intentionally introduced Er3+ defect complexes across a 180° domain wall produced at room temperature by electric-field poling. Using site-selective excitation-emission spectroscopy for well chosen excitation energies, we found drastic differences in the Er3+ emission, which are due to a rearrangement of the defect complexes. We used these changes in a confocal luminescence microscope to image ferroelectric-domain structures. This powerful imaging method with a 700 nm, 50 ms spatial and temporal resolution can be used to study real-time dynamics of domain walls.

Structural changes in the Ras protein revealed by fluorescence spectroscopy

The Y32W mutant of the Ras protein which has a tryptophan residue close to the guanine nucleotide binding site is studied using two fluorescence spectroscopic techniques. Two-dimensional mapping of all emission and all fluorescence spectra using excitation–emission spectroscopy (EES) in conjunction with time-resolved laser-induced fluorescence (LIF) is used to analyze and assign the contribution of the different fluorophores to the total fluorescence. Time-resolved LIF is shown to be a method that allows to follow the slight conformational changes of Ras binding to the nucleotides GDP, GTP, or the non-hydrolyzable analogues GppNHp, GppCH2p and GTP-γS and allows to distinguish between the active and inactive form. Additionally, a variant of the EES technique is used for the investigation of the intrinsic GTPase function of Ras and the determination of kinetic constants for this reaction.

Rearrangement of Rare Earth Defects Under Domain Inversion in LiNbO3

Using high resolution excitation-emission spectroscopy we investigated the changes occurring in the optical transition of Er3☎ ions in LiNbO3 during inversion of the ferroelectric axis. In stoichiometric LiNbO3 we find that a drastic reconfiguration among the different defect sites takes places favoring those centers which have already been dominant in the as-grown sample. The reconfiguration is attributed to changes in the arrangement of the local charge compensators. Furthermore, we find a small shift of the emission transition energy, which is consistent with an increase of the intrinsic electric field. These findings make the Er3☎ ions very suitable probes for 3D imaging of domain structures and for in-situ studies of the dynamics of the domain inversion processes and the defect/domain wall interaction.

Electronic and vibrational spectroscopy of jet-cooled complexes of o-cyanophenol

Hydrogen-bonded complexes of o-cyanophenol ( o-CNP) with a series of protic and non-protic solvents have been investigated in supersonic jet by means of laser-induced fluorescence excitation and dispersed emission spectroscopy. All the complexes exhibit a large red shift of the S 0–S 1 transition extending from −674 cm −1 for acetonitrile to −1321 cm −1 for the dimer of methanol. This shift is shown to correlate with the proton affinity of the protic solvents and has been related to the formation of cyclic hydrogen bond networks. For a non-protic solvent such as diethylether, the presence of a strong H bond has been confirmed by the study of the OH stretch using IR–UV double resonance spectroscopy. The low energy intermolecular vibrations observed in the cyclic complexes of o-CNP with H 2O and CH 3OH were assigned by comparing the experimental frequencies observed in the S 0 and S 1 states with the results of DFT calculations. The complexes with TEA undergo proton transfer as evidenced from the observation of the fluorescence attributable to the deprotonated form of o-CNP.

Isolated Building Blocks of Photonic Materials: High-Resolution Spectroscopy of Excited States of Jet-Cooled Push-Pull Stilbenes

The lowest excited singlet states of 4-(dimethylamino)-4‘-cyanostilbene (DCS), 4-(dimethylamino)-4‘-nitrostilbene (DANS), and 4-di(hydroxy-ethyl)amino-4‘-nitrostilbene (DANS-diol) have been investigated by high-resolution fluorescence excitation and dispersed emission spectroscopy on samples seeded in supersonic expansions. Using ab initio calculations of the harmonic force fields of the electronic ground state as a starting point for the analysis of vibrational modes and frequencies, detailed vibrational assignments of the rich line structure of the S1 ← S0 transition in these three push−pull stilbenes are reported. From the experimental and theoretical analysis, it becomes apparent that the excitation spectra are dominated by low-frequency vibrational modes of the stilbene-like backbone, albeit that a characterization in terms of trans-stilbene vibrational terminology is not very appropriate. The fluorescence excitation and emission spectra show significant Franck−Condon factors for transitions involvin...

The fine structure of spectral response of Cr3+ sites in LiNbO3

We studied Cr3+ centers in congruent LiNbO3 crystals using the method of combined excitation emission spectroscopy. Performing a coordinate transformation of the measured experimental data the inhomogeneous broadening of the 2E → 4A2 transitions of Cr3+ was directly separated into contributions from the variations of the low symmetry crystal field and of free ion parameters. At liquid helium temperatures fine structure in the spectral response of Cr3+ “defect” centers in high crystal field sites was observed. Our results are discussed on the basis of a microscopic hierarchic model for perturbed Cr3+ ions in the lithium niobate crystal lattice.

Crystal field analysis of different Er3+ sites in LiNbO3

The Er3+ defect in lithium niobate (LiNbO3), and the slightly different lattice environments (“sites”) it experiences, have been investigated with combined excitation-emission spectroscopy under application of hydrostatic pressure and electric fields. The results give a solid basis for a crystal field analysis which yields site-specific crystal field parameters. We found that the Er3+ sites can be classified into two groups. Firstly, the sites are distinct due to different relative arrangements of the nearest-neighbor oxygens and secondly they encounter to different local electric fields which are caused by charge compensators and other defects in a more distant neighborhood.

Double-resonance excitation-emission spectroscopy on Er:Ti:LiNbO3 waveguides

The waveguide-specific optical properties of Er3+ in Ti:LiNbO3 have been investigated. Using double-resonance excitation-emission spectroscopy a drastic fluorescence line narrowing is observed which allows resolution of the strongly inhomogeneously broadened emission lines. This way, the Er3+ ions in this strongly perturbed environment could be studied selectively. In particular, it is found that the main influence of the Ti4+ diffusion profile within the wave-guiding zone is of electrostatic nature and is due to a decrease of the internal local field with increasing [Ti4+]-concentration.

Site-selective spectroscopy of Er3+:Ti:LiNbO3 waveguides

Starting from previous investigations in LiNbO3 bulk crystals, we studied the optical properties of Er3+ ions in Ti:LiNbO3 channel waveguides and investigated the waveguide-specific lattice environment of the Er3+ ions (“sites”) caused by the doping method used and the presence of a large number of Ti4+ ions. For that purpose the method of combined excitation–emission spectroscopy was applied for the first time to waveguides at low temperatures. Comparing the spectroscopic results obtained for the green, red, and near-IR luminescence (λ≈550, ≈650 and ≈980 nm) under direct (450 nm), 2-step (980 nm), and 3-step (1.5 μm) laser excitation, we found several distinguishable Er3+ sites which in terms of energy levels and relative numbers are similar to those in bulk material, but exhibit significantly different up-conversion efficiencies and strongly inhomogeneously broadened transitions. Moreover, we were able to distinguish isolated and cluster Er3+ sites by their characteristic excitation and emission transition energies and studied the respective excitation/relaxation channels. The cluster sites are most efficient in the up-conversion process, especially under 3-step excitation. Using accepted microscopic models for Er3+ and Ti4+ incorporation into the LiNbO3 crystal lattice, the site distribution and up-conversion mechanisms are elucidated and their consequences for laser applications in different spectral regions are discussed.