Second Edition of the Encyclopedia of Spectroscopy and Spectrometry pulls key information into a single source for quick access to answers and/or in-depth examination of topics. SPEC-2 covers theory, methods, and applications for researchers, students, and professionals - combining proven techniques and new insights for comprehensive coverage of the field. The content is available in print and online via ScienceDirect, the latter of which offers optimal flexibility, accessibility, and usability through anytime, anywhere access for multiple users and superior search functionality. No other work gives analytical and physical (bio)chemists such unprecedented access to the literature. With 30% new content, SPEC-2 maintains the 'authoritative, balanced coverage' of the original work while also breaking new ground in spectroscopic research. The key features of the book include: incorporates more than 150 color figures, 5,000 references, and 300 articles (30% of which are new), for a thorough examination of the field; highlights new research and promotes innovation in applied areas ranging from food science and forensics to biomedicine and health; and features a new co-editor: David Koppenaal of Pacific Northwest National Laboratory, Washington, USA, whose work in atomic mass spectrometry has been recognized internationally.

Determining accurate dissociation constants for equilibrium processes involving a fluorescent mechanism can prove to be quite challenging. Typically, titration curves and nonlinear least squares fitting of the data using computer programs are employed to obtain such constants. However, these approaches only consider the total fluorescence signal and often ignore other energy transfer processes within the system. The current model considers the impact on fluorescence from equilibrium binding (viz., metal-ligand, ligand-substrate, etc.), quenching, and resonance energy transfer. This model should provide more accurate binding constant as well as insights into other photonic processes. The equations developed for this model are discussed and are applied to experimental data from titrimetric experiments. Since the experimental data are generally in excess of the number of parameters that are needed to define the system, fitting is operated in an overdetermined mode and employs error minimization (either absolute or relative) to define goodness of fit. Examples of how changes in certain parameters affect the shape of the titrimetric curve are also presented. The current model does not consider chelation-enhanced fluorescence.

The coupling of separation by preparative ultracentrifugation and metal detection by inductively coupled plasma mass spectrometry (ICP-MS) has been explored for metal–protein equilibrium determinations. This study characterizes the stoichiometry as well as apparent (Kapp) and intrinsic (Kint) binding affinities of the metal–protein association for a model protein. In particular, the affinity of Cu2+ for the high affinity binding site in bovine serum albumin (BSA) is determined. Once equilibrium is established between Cu2+ and BSA, preparative ultracentrifugation moves the metalloprotein away from the meniscus, leaving unbound equilibrium copper in the protein free solution. Since the initial (total) concentrations of purified BSA and Cu2+ can be determined, the free copper concentration at equilibrium can also be determined by taking a small aliquot above the sedimenting boundary for analysis using ICP-MS. This analysis allows for the determination of free Cu2+ ion, which is identical to the equilibrium concentration prior to ultracentrifugation. From these data Kapp and Kint were determined at two different conditions, 100mM Tris(hydroxymethyl)aminomethane (Tris) at pH 9.53 and pH 7.93. log Kapp values of 17.6 and 14.6 were determined at pH 9.53 and pH 7.93, respectively. Furthermore, pH-independent log Kint values of −1.43 and −1.04 were determined at pH 9.53 and 7.93, respectively. While the log Kint at pH 9.53 was in good agreement with literature values obtained from alternative methods, Kint at pH 7.93 was about 2.5× larger than previously reported. BSA undergoes a structural rearrangement between pH 7–9, and the generally accepted pH-dependency of protein tertiary structure may be responsible for the variations in the “intrinsic” binding constant. The Cu–BSA binding affinity was also monitored in 100mM Tris 0.1% sodium dodecyl sulfate (SDS) solution at pH 7.93 in order to determine the effect of a denaturant on metal binding. Results for both log Kapp and log Kint were similar to those obtained in the absence of 0.1% SDS at pH 7.93. Overall, this study validates and shows the efficacy of combining preparative ultracentrifugation with ICP-MS detection for interrogating metal–protein associations while causing minimal equilibrium perturbations as a result of the separation and measurement processes. Advantages and disadvantages of this methodology are discussed as it relates to alternative methods for metal–protein studies.

The novel analytical application of the combination of an inline electrothermal vaporization (ETV) and nebulization source for inductively coupled plasma mass spectrometry (ICP-MS) has been studied. Wet plasma conditions are sustained during ETV introduction by 200mL/min gas flow through the nebulizer, which is merged with the ETV transport line at the torch. The use of a wet plasma with ETV introduction avoided the need to change power settings and torch positions that normally accompany a change from wet to dry plasma operating conditions. This inline-ETV source is shown to have good detection limits for a variety of elements in both HNO3 and HCl matrices. Using the inline-ETV source, improved limits of detection (LOD) were obtained for elements typically suppressed by polyatomic interferences using a nebulizer. Specifically, improved LODs for 51V and 53Cr suffering from Cl interferences (51ClO+ and 53ClO+ respectively) in a 1% HCl matrix were obtained using the inline-ETV source. LODs were improved by factors of 65 and 22 for 51V and 53Cr, respectively, using the inline-ETV source compared to a conventional concentric glass nebulizer. For elements without polyatomic interferences, LODs from the inline-ETV were comparable to conventional dry plasma ETV-ICP time-of-flight mass spectrometry results. Lastly, the inline-ETV source offers a simple means of changing from nebulizer introduction to inline-ETV introduction without extinguishing the plasma. This permits, for example, the use of the time-resolved ETV-ICP-MS signals to distinguish between an analyte ion and polyatomic isobar.

In a previous paper from this laboratory, 51 different elements were ratioed to each other under a variety of altered matrix and instrumental conditions. The objective was to isolate element pairs that made good internal standards for each other. The data were collected and evaluated using a nebulizer and an inductively coupled plasma time-of-flight mass spectrometer. Although results suggested that many optimal internal standards were correctly predicted, it was not clear whether these predicted internal standard pairs would remain good choices for this instrument over an extended time period and whether changing instrumental platforms would alter the internal standard selections. This article considers the choice of previously selected "good'' internal standards for several elements using the same GBC ICP(TOF)MS after several optimizations, a torch change and nebulizer replacement. It also considers the use of the previously determined "good'' internal standards for an Agilent ICP(quadrupole)MS with a different nebulizer system, torch, mass analyzer, etc. Interestingly, it was found that the internal standards predicted in the initial study continued to perform well on both platforms considered in the present study. In fact, overall errors were smaller in the second set of TOF data and on the quadrupole than those in the first set of TOF data. Mass and ionization potential trends were also similar to those from the previous study. Although there remains to be an equation allowing a priori selection of the ideal internal standard, the current results suggest that the prediction program developed in the previous study is effective over time and instrumental platforms.

Isotope ratio measurements are found to have systematic bias when using the analog detection mode on an inductively coupled plasma time-of-flight (TOF) mass spectrometer. This bias is dependent upon the value of the ratio, the intensity of the signal, and the gain of the electron multiplier tube. The error should not appear if ion counting is employed instead of analog detection, although analog detection with time-of-flight has other distinct advantages. The cause of this isotope ratio inaccuracy is rooted in disproportionate recording of the analog signal because of the need to filter out noise by blocking analog signals below a threshold voltage. This attenuates smaller signals to a greater degree than larger signals. This variable "detection efficiency" causes a larger systematic error in the isotopic ratio as the isotopic abundances become more disparate. Ratios close to unity are generally accurate within the precision of the measurement. The use of an increased gain on the detector leads to improved ratio accuracy, but at the cost of decreased detector lifetime. This research presents a method of analyzing solutions using natural, known isotopic ratios to produce an efficiency correction curve. The average error of several isotope ratios for a 500 ng/mL solution of various elements with ratios between 3.4 and 10 was found to be 6.5% without correction, 3.0% with increased detector gain, 1.1% with efficiency correction and 0.6% with both increased gain and efficiency correction. (C) 2008 Elsevier B.V. All rights reserved.

A new method of separating Rb and Sr by ETV-ICP-MS is demonstrated. An electrothermal vaporizer (ETV) is used for sample introduction and control of the ETV heating cycle provides temporal separation of the isobars at m/z 87, allowing 87Rb and 87Sr determination within the same sample. The method utilizes an inductively coupled plasma mass spectrometer (ICP-MS) for ionization and detection. A one or two vaporization stage heating method is used for the ETV separation, and initial methodology and feasibility experiments were conducted on an ETV-ICP-TOFMS. Final measurements were made using a MC-ICP-MS for data collection. The method uses minimal sample preparation and no front end chromatographic separation of Rb and Sr prior to analysis. The technique is applied to the analysis of a potassium feldspar sample (NIST SRM 607) and is demonstrated to be viable as a means of determining isotopic data for the Rb–Sr geochronological technique. The standard was determined to have a 87Sr/86Sr ratio of 1.1876 ± 0.011, a Sr concentration of 64.1 ± 4.7 µg g−1, and a Rb concentration of 557 ± 6 µg g−1 (95% CI). This corresponds to a model age of 1266 ± 186 million years, statistically similar to the certified value of 1409 ± 14 for the standard. Error analysis is presented for several key calculations and a discussion is presented regarding approaches to improve precision.

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Internal standards are routinely used with inductively coupled plasma mass spectrometry (ICPMS) to minimize the impact of signal instability. Previous studies indicate that internal standard choice should be directed by similarities in mass, but they neglect to address the possibilities of exceptions or what impact a poor internal standard choice may have on quantitative results. A 51 element suite was run under varying instrument and matrix perturbations on an ICPMS instrument equipped with a time of flight mass spectrometer. Each element was ratioed to every other element for perturbed conditions that were intended to simulate variations in matrix composition or drift in operating parameters. These included changing sample delivery rate, matrices (NaCl and acetic acid) and horizontal torch position. The %RSDs of each analyte-to-internal standard ratio were used to rank the relative quality of each analyte-internal standard pair and their response to changing conditions. Multiple ordinary least squares equations were calculated to evaluate various chemical and physical properties that may be predictors of an optimal analyte-internal standard combination. Overall, similarity of masses was found to be the most important predictor of a good internal standard. However, exceptions did exist under various perturbations and among different elements. In a final study, nine analytes were run under perturbed experimental conditions and in two complex matrices (NaCl and acetic acid), but quantitation was attempted using simple aqueous standards. Accuracy and precision were evaluated for several internal standards for each of the nine elements as well for the nine analytes where no internal standard was employed. From this evaluation, it was clear that internal standard choice can have substantial effects on analytical accuracy and the improvement of analytical precision, but there is no single physical or chemical parameter that reliably allows a priori selection of a “good” internal standard for any given analyte.

Poly-l-cysteine (PLCysn) (n=20) was immobilized onto the surface of commercially available magnetic γ-Fe2O3 nanoparticles, and its use as a selective heavy metal chelator was demonstrated. Magnetic nanoparticles are an ideal support because they have a large surface area and can easily be retrieved from an aqueous solution. PLCysn functionalization was confirmed using FTIR and the quantitative Ellman's test. Metal binding capacities for As(III), Cd(II), Cu(II), Ni(II), Pb(II) and Zn(II) were determined at pH 7.0 and compared to adsorption capacities for unfunctionalized γ-Fe2O3 nanoparticles. The effect of pH on the PLCysn functionalized nanoparticles was also investigated. For all of the metals examined, binding capacities (μmol metal/g support) were more than an order of magnitude higher than those obtained for PLCysn on traditional supports. For As(III), Cu(II), Ni(II) and Zn(II), the binding capacities were also higher than the metal adsorption capacities of the unfunctionalized particles. Metal uptake was determined to be rapid (<2.5min) and metal recoveries of >50% were obtained for all of the metals except As(III). PLCysn, which has a general metal selectivity towards soft metals acids, was chosen to demonstrate the proof of concept. Greater metal selectivity may be achievable through the use of combinatorial peptide library screening or by using peptide fragments based on known metal binding proteins.

A new fluorescent peptidyl chemosensor based on the mercury binding MerP protein with fluorescence resonance energy transfer (FRET) capabilities has been synthesized via Fmoc solid-phase peptide synthesis. The metal chelating unit, which is flanked by the fluorophores tryptophan (donor) and dansyl (acceptor), contains amino acids from MerP's metal binding loop (sequence: dansyl-Gly-Gly-Thr-Leu-Ala-Val-Pro-Gly-Met-Thr-Cys-Ala-Ala-Cys-Pro-Ile-Thr-Val-Lys-Lys-Gly-Gly-Trp-CONH2). A FRET enhancement or 'turn-on' response was observed for Hg2+ as well as for Zn2+, Cd2+ and Ag+ in a pure aqueous solution at pH 7.0. The emission intensity of the acceptor was used to monitor the concentration of these metals ions with detection limits of 280, 6, 103 and 496 mu g L-1, respectively. No response was observed for the other transition, alkali and alkaline earth metals tested. The fluorescent enhancement observed is unique for Hg2+ since this metal generally quenches fluorescence. The acceptor fluorescence increase resulting from metal binding-induced FRET suggests a sensor that is inherently more sensitive than one based on quenching by the binding event.

The metal binding capacities, conditional stability constants, and secondary structure of immobilized polyaspartic acid (PLAsp) (n = 6, 20, and 30) on TentaGel resin were determined when binding Mg2+, Co2+, Cd2+, and Ni2+. Metal binding to the synthesized peptides was evaluated using breakthrough curves from a packed microcolumn and flame atomic absorption spectrophotometry (FAAS) detection. The metal capacities reached values of 590, 2160, and 3710 mumol of metal/g of resin for the 6-mer, 20-mer, and 30-mer, respectively, and this resulted in 2-3 residues per metal for all peptides and metals tested. Surprisingly, the concentrated environment of the resin along with the spatial distribution of attachment groups allowed for most residues to participate in metal binding regardless of the peptide length. Conditional stability constants calculated using single metal binding isotherms indicated that binding strength decreased as the chain length increased on the resin. Raman microscopy on single beads was used to determine PLAsp secondary structure, and all peptides were of a mixed conformation (i.e., beta-sheets, alpha-helices, random chain, etc.) during neutral conditioning and metal binding. Uniquely, the longer 20-mer and 30-mer peptides showed a distinct change from a mixed conformation to beta-sheets and alpha-helices during metal release with acid. This study confirms that metal release by longer immobilized peptides is often assisted by a conformational change, which easily spoils the binding cavity, while shorter peptides may release metal primarily by H+ displacement.

A new method for screening split-pool combinatorial libraries for catalytic activity is described. Site-selective detection of catalytic activity for solution-based reactions was made possible without cofunctionalizing beads or adding diffusion-limiting matrixes. This was done by spatially separating resin-bound catalysts on an adhesive array on a microscope slide and introducing the reacting liquid to the top of the slide. Convective mixing and evaporation was controlled using a cover slide and imaging both the formation of products within active beads and the diffusion of products out of the beads. Colored reaction products and pH-sensitive indicators were used to visually detect catalytically active beads in the presence of inactive ones. Quantitative analyses of the images support the assumption that color intensities can be used to assess the quality of hits from a combinatorial screen. The Knoevenagel condensation reaction catalysis as well as esterase screening using methyl red were used to validate the approach. Using the esterase data, it was shown that some information on activity could also be extracted from the colored plume surrounding individual beads although the precision is not as good as that from direct measurement of absorbance through the bead. It was also found that the distribution of products within a single bead can also be gleaned from the absorbance data for different-sized beads.

One of the largest obstacles in miniaturizing traditional atomic spectroscopic sources is the need for a thermal/electrical source for free atom production. A single article in the literature has demonstrated atomic absorption detection of Ag, Cu, and Pd in solution at room temperature for atoms in the gas phase, which may ultimately permit miniaturization. Unfortunately, several laboratories have found that reproducing the phenomenon has been difficult. Without a sound fundamental explanation of the processes leading to the signal, one must conclude that it can be done, but some unsuspected and unknown design/methodological nuances are responsible for only a single reported success. Gas phase atoms could exist at room temperature "in solution" if the atoms were trapped in very small bubbles. In the current study, submicrometer-sized bubbles were created in a flow-through cell during the mixing of an alcohol-water solution containing a reducing agent with water containing the analyte. A repeatable atomic absorption signal was produced. Replacement of ethanol with 1-propanol and use of a surfactant increased the signal. Limits of detection of approximately 100 ppb in Pd were achieved, and it is estimated that approximately 0.4% of the Pd initially added is contained within the bubbles as gaseous atoms. The paper discusses the fundamental processes needed to achieve a repeatable signal.

A solventless, multi-element, steady-state optimization system is presented. The device uses three parallel conduits that contain solid NbF5 and SnBr4 as well as a low wattage tungsten filament loaded with Pb to deliver dry vapors to an ICP. These are used to create a steady-state signal containing the following ions that can be used for tuning: 28Si+, 79Br+, 120Sn+, 184W+, and 208Pb+. In comparison to using a nebulizer for optimization, the dry plasma optimization produced an average enhancement of 4.5(±0.4)-fold for 26 elements when using electrothermal vaporization sample introduction.