Excitation-emission fluorescence matrices

Guimet et al. used two potential multiway methods for the discrimination between virgin olive oils and pure olive oils the unfold principal component analysis (U-PCA) and parallel factor analysis (PARAFAC), for the exploratory analysis of these two types of oils. Both methods were applied to the excitation-emission fluorescence matrices (EEM) of olive oils and followed the comparison of the results with the ones obtained with multivariate principal component analysis (PCA) based on a fluorescence spectrum recorded at only one excitation wavelength. 

Patel-Sorrentino, N., S. Mounier, and J. Y. Benaim. 2002. Excitation-emission fluorescence matrix to study pH influence on organic matter fluorescence in the Amazon basin rivers. Water Research 36, no. 10 2571-2581. doi 10.1016/S0043-1354(01)00469-9. 

The simultaneous fluorometric determination of PG and BHA in cosmetics has also been described using excitation-emission fluorescence matrix data, which were processed by applying the second-order calibration method based on self-weighted alternating normalized residue fitting algorithm [80]. The detection limit obtained for PG was 2.2 ng/mL. 

There are several other chemometric approaches to calibration transfer that will only be mentioned in passing here. An approach based on finite impulse response (FIR) filters, which does not require the analysis of standardization samples on any of the analyzers, has been shown to provide good results in several different applications.81 Furthermore, the effectiveness of three-way chemometric modeling methods for calibration transfer has been recently discussed.82 Three-way methods refer to those methods that apply to A -data that must be expressed as a third-order data array, rather than a matrix. Such data include excitation/emission fluorescence data (where the three orders are excitation wavelength, emission wavelength, and fluorescence intensity) and GC/MS data (where the three orders are retention time, mass/charge ratio, and mass spectrum intensity). It is important to note, however, that a series of spectral data that are continuously obtained on a process can be constructed as a third-order array, where the three orders are wavelength, intensity, and time. 

There are two competing and equivalent nomenclature systems encountered in the chemical literature. The description of data in terms of ways is derived from the statistical literature. Here a way is constituted by each independent, nontrivial factor that is manipulated with the data collection system. To continue with the example of excitation-emission matrix fluorescence spectra, the three-way data is constructed by manipulating the excitation-way, emission-way, and the sample-way for multiple samples. Implicit in this definition is a fully blocked experimental design where the collected data forms a cube with no missing values. Equivalently, hyphenated data is often referred to in terms of orders as derived from the mathematical literature. In tensor notation, a scalar is a zeroth-order tensor, a vector is first order, a matrix is second order, a cube is third order, etc. Hence, the collection of excitation-emission data discussed previously would form a third-order tensor. However, it should be mentioned that the way-based and order-based nomenclature are not directly interchangeable. By convention, order notation is based on the structure of the data collected from each sample. Analysis of collected excitation-emission fluorescence, forming a second-order tensor of data per sample, is referred to as second-order analysis, as compared with the three-way analysis just described. In this chapter, the way-based notation will be arbitrarily adopted to be consistent with previous work. 

Any analytical data obtained by hyphenated instruments or by two-way spectroscopic techniques such as excitation-emission fluorescence spectroscopy are bilinear ones. The bilinear data matrix has a very useful property, namely the rank of such matrix obtained with any chemical mixture is equal to the number of chemical components in the mixture. Thus, theoretically, the rank of a data matrix of any pure chemical component is unit. It can be expressed by the product of two vectors ... 

Lee, S. and Ahn, K.H. (2004). Monitoring of COD as an organic indicator in wastewater and treated effluent by fluorescence excitation-emission (FEEM) matrix characterization. Water Sci. TechnoL, 50 (8), 57-63. 

Room-temperature fluorescence (RTF) has been used to determine the emission characteristics of a wide variety of materials relative to the wavelengths of selected Fraunhofer lines in support of the Fraunhofer luminescence detector remote-sensing instrument. RTF techniques are now used in the compilation of excitation-emission-matrix (EEM) fluorescence "signatures" of materials. The spectral data are collected with a Perkin-Elraer MPF-44B Fluorescence Spectrometer interfaced to an Apple 11+ personal computer. EEM fluorescence data can be displayed as 3-D perspective plots, contour plots, or "color-contour" images. The integrated intensity for selected Fraunhofer lines can also be directly extracted from the EEM data rather than being collected with a separate procedure. Fluorescence, chemical, and mineralogical data will be statistically analyzed to determine the probable physical and/or chemical causes of the fluorescence. 

Fig. 4.8. Fluorescence lifetime of a stained section of Convallaria resolved with respect to lifetime, excitation and emission wavelength (A) intensity image integrated over the time-resolved excitation-emission matrix (EEM) (B, D) time-integrated EEM from areas A and B respectively in (A) (C) fluorescence decay profile for /ex 490 nm and Aem 700 nm corresponding to area A (E) fluorescence decay profile for Aex 460 nm and /em 570 nm corresponding to area B.

More fluorescence features than just the emission intensity can be used to develop luminescent optosensors with enhanced selectivity and longer operational lifetime. The wavelength dependence of the luminescence (emission spectmm) and of the luminophore absorption (excitation spectrum) is a source of specificity. For instance, the excitation-emission matrix has shown to be a powerful tool to analyze complex mixtures of fluorescent species and fiber-optic devices for in-situ measurements (e.g. 

Baker, A. 2001. Fluorescence excitation-emission matrix characterization of some sewage- impacted rivers. Environmental Science Technology, 35, 948-953. 

Bidimensional fluorescence spectra are commonly obtained in the three modes of emission, excitation, and synchronous-scan excitation, whereas tridimensional fluorescence (or total luminescence) spectra are obtained in the form of excitation-emission matrix (EEM) plots by measuring the fluorescence intensity emitted as a function of the wavelength over a range of excitation wavelengths. This technique allows to obtain more detailed information than that obtained by using conventional monodimensional fluorescence (Mobed et al., 1996). Fluorescence spectroscopy has provided valuable information on the molecular structure, functionalities, conformation, and intramolecular and intermolecular interactions of HS from organic amendments and unamended and amended soils (Senesi et al., 1990,1996, 2007 Mobed et al., 1996 Chen et al., 2003 Senesi and Plaza, 2007). 

Figure 4.5. Fluorescence excitation-emission matrix spectra of humic acids (HAs) isolated from sewage sludge (B) and two soils either unamended (TH1 and TH2, respectively) or amended with 3901 ha-1 of sewage sludge (THB1 and THB2, respectively) sampled from the surface (s, 0-25cm) and subsurface (ss, 25-50cm) layers. Reprinted from Bertoncini, E. I., D Orazio, V., Senesi, N., and Mattiazzo, M. E. (2005). Fluorescence analysis of humic and fulvic acids from two Brazilian oxisols as affected by biosolid amendment. Anal. Bioanal. Chem. 381,1281-1288, with permission from Springer.

Figure 16.41. Fluorescence excitation-emission matrix spectra of humic acids (HA) isolated from municipal solid waste compost (MSWC), from soil amended with MSWC at 40tha 1yr 1 (MSWC40), and from the corresponding unamended control soil (MSWC0), in the absence and presence of Cu2+, Zn2+, Cd2+, and Pb2+ ions at a total concentration of 40 xmol liter-1. EEWPmax denotes the excitation/emission wavelength pairs at maximum fluorescence intensity (Plaza et al., 2006).

Molecular fluorescence spectroscopy is a commonly employed analytical method that is sensitive to certain chemical properties of FA (9-13). Fulvic acid s molecular fluorescence is principally due to conjugated unsaturated segments and aromatic moieties present in the macromolecule (14). Several types of fluorescence spectra can be measured, including an excitation emission matrix or total luminescence spectrum, constant offset synchronous fluorescence, excitation spectra, and emission spectra, furnishing the researcher with useful data. The ability to resolve and select multiple fluorescent species makes these approaches extremely useful for studying FA relative to its chemical reactivity. 

According to the initial results reported here, a fluorescence enhancement effect for humic material binding with aluminum can be observed, while under different conditions a quenching of fluorescence predominated. Another very interesting fluorescence phenomenon described here is that after complexation with AE, the maximum intensity of the humic material fluorescence shifts to longer excitation wavelengths and to shorter emission wavelengths in an excitation emission matrix (EEM). 

Figure 4. Excitation emission matrix contour plot of 15.0 mg/L soil fulvic acid in 0.1M NaC104 at pH 4.00. The contour lines give fluorescence intensity in arbitrary units.

We developed two kinds of multidimensional fluorescence spectroscopic systems the time-gated excitation-emission matrix spectroscopic system and the time- and spectrally resolved fluorescence microscopic system. The former acquires the fluorescence intensities as a function of excitation wavelength (Ex), emission wavelength (Em), and delay time (x) after impulsive photoexcitation, while the latter acquires the fluorescence intensities as a function of Em, x, and spatial localization (%-, y-positions). In both methods, efficient acquisition of a whole data set is achieved based on line illumination by the laser beam and detection of the fluorescence image by a 2D image sensor, that is, a charge-coupled device (CCD) camera. 

Figure 32.1 (a) A schematic illustration of time-gated excitation-emission matrix spectroscopy, (b) A typical example of the 3D fluorescence data measured for Rhodamine 590 in ethanol... 

Summarizing this section, we developed the time-gated excitation-emission matrix spectroscopic system and applied it to the decomposition of a mixed solution of a number of fluorescent dyes. We demonstrated that our approach, which was based on unique optical configuration, efficient acquisition of a multidimensional data set, and decomposition of unknown fluorescent components by using the PARAFAC model, was effective for the analysis ofunknown multi-component targets. 

Figure 1 illustrates the total fluorescence matrix in contour format for BuPBD euid poly (VBuPBD) at 298K. These spectra graphically illustrate the above conclusions and the symmetry of the broadened profile of the polymer emission with respect to the emission/excitation axes is convincing evidence for the excimeric nature of the species responsible for the low energy intensity component. In addition the emission of both polymers reduces to that of the unassociated monomeric chromophore upon dispersal in a glassy (MeTHF) matrix at 77K. The effect is illustrated for poly (VBuPBD) in Figure 2. This observation is consistent with the formation of intramolecular excimers by a conformational sampling mecheutism.

The excitation-emission matrix spectra (EEMS) of CDOM fluorescence also are altered on exposure to solar radiation, generally with reductions in fluorescence that approximately parallel absorption losses [36-38]. Hypsochromic shifts (shifts to shorter wavelengths) occur in both excitation and emission maxima on irradiation [36-38]. Interactions between photochemical and microbial degradation [38] are involved. 

Figure 6.8. Example of the use of rank annihilation factor analysis for determining the concentration of tryptophane using fluorescence excitation-emission spectroscopy. In the top left plot the unknown sample is shown. It contains three different analytes. The standard sample (only tryptophane) is shown in the top right plot. In the lower right plot, it is shown that the smallest significant (third from top) singular value of the analyte-corrected unknown sample matrix reaches a clear minimum at the value 0.6. In the lower left plot the unknown sample is shown with 0.6 times the standard sample subtracted. It is evident that the contribution from the analyte is practically absent.

Jiji RD, Andersson GG, Booksh KS, Application of PARAFAC for calibration with excitation-emission matrix fluorescence spectra of three classes of environmental pollutants, Journal of Chemometrics, 2000, 14, 171-185. 

Zhu SH, Wu HL, Xia AL, Han QJ, Zhang Y, Yu RQ. Quantitative analysis of hydrolysis of carbaryl in tap water and river by excitation-emission matrix fluorescence coupled with second-order calibration. Talanta 2008 74 1579-5. 

Trevisan, M.G. and Poppi, R.J. (2003) Determination of doxorubicin in human plasma by excitation-emission matrix fluorescence and multi-way analysis. Anal Chim Acta, 493 (1), 69-81. 

Wu, F. C., Evans, R. D., and Dillon, P. J., Separation and characterization of NOM by high performance hquid chromatography and online three dimensional excitation emission matrix fluorescence detection. Environ. Sci. TechnoL, 37, 3687-3693, 2003. 

Y.G. Chung, J.A. Schwartz, C.M. Gardner, R. Sawya, S.L. Jaques (1994). Fluorescence of normal and cancerous brain tissue the excitation/emission matrix. In Robert R. Alfano (Ed.), Advances in Laser and Light Spectroscopy to Diagnose Cancer and other Diseases (SPIE Proc. Vol. 2135, pp. 66-75). 

Up to this point, regression has been restricted to two blocks of two-way data Y and X. In chemical analysis, however, a growing number of problems can be cast as three-way regression analysis. Consider the calibration of chemical constituents on the basis of their fluorescence excitation/emission spectrum or of gas chromatography/mass spectrometry (GC/MS) data. For each sample, two-dimensional measurements are available that constitute a three-way data array, X. This data array has to be related to sample concentrations of one, vector y, or several analytes, matrix Y. Cases can be imagined where even the matrix Y constitutes a three-way data array. 

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