Kinetics real-time

To study reaction kinetics, real-time analysis of a reaction mixture composition is commonly carried out by quenched-flow78,79 and stopped-flow80-83 methods. 

There are two main applications for such real-time analysis. The first is the detemiination of the chemical reaction kinetics. Wlien the sample temperature is ramped linearly with time, the data of thickness of fomied phase together with ramped temperature allows calculation of the complete reaction kinetics (that is, both the activation energy and tlie pre-exponential factor) from a single sample [6], instead of having to perfomi many different temperature ramps as is the usual case in differential themial analysis [7, 8, 9, 10 and H]. The second application is in detemiining the... 

Fatty Acid Transporters. Figure 2 Quencher-based real-time fatty acid uptake assay with a fluorescently labeled FFA analogue (C1-Bodipy-C12). Predominantly protein-mediated fatty acid uptake by 3T3-L1 adipocytes (diamonds) was compared with diffusion-driven uptake by fibroblasts (squares) using the QBT Fatty Acid Uptake reagent (Molecular Devices Corp., CA, USA), which contains C1-Bodipy-C12 as substrate in conjunction with a cell impermeable quencher. Uptake kinetics was recorded using a Gemini fluorescence plate reader. Error bars indicate the standard deviations from 12 independent wells. RFU relative fluorescence units. 

Real-time spectroscopic methods can be used to measure the binding, dissociation, and internalization of fluorescent ligands with cell-surface receptors on cells and membranes. The time resolution available in these methods is sufficient to permit a detailed analysis of complex processes involved in cell activation, particularly receptor-G protein dynamics. A description of the kinetics and thermodynamics of these processes will contribute to our understanding of the basis of stimulus potency and efficacy. 

MORTENSEN A, SKIBSTED L H (1997) Relative stability of carotenoid radical cations and homologue tocopheroxyl radicals. A real time kinetic study of antioxidant hierarchy. FFBS Letters, 417, 261-6. 

In a real-time spectroellipsometric measurement in which the kinetics of a-Si H deposition is studied, trajectories are recorded in the A-4 plane at various photon energies between 2 and 4 eV. These trajectories can be simulated and fitted to models that represent the growing a-Si H layer. Canillas et al. [347] have made a detailed study of the deposition of the first few layers of a-Si H on a NiCr/glass substrate. Similar results are obtained for a c-Si substrate. They have proposed several models to explain the data. One possible model is the hemispherical nu-cleation model, which describes a hexagonal network of spherical a-Si H nuclei located at an average distance d between them. The growth is represented by an... 

Matheson, I. B. C. The method of successive integration a general technique for recasting kinetic equations in a readily soluble form which is linear in the coefficients and sufficiently rapid for real time instrumental use. Anal Instrum. 1987, 16, 345-373. 

From the examples given above it is clear that the development of a novel approach for real time measurements of the dissociation kinetics of reaction intermediates would greatly assist the unravelling of complex multistep processes associated with the transformation of even simple molecules at transition metal centers. While techniques are available for such measurements over a limited range of times, none of the methods are sufficiently general to be useful for extensive measurements. 

Despite the considerable amount of information that has been garnered from more traditional methods of study it is clearly desirable to be able to generate, spectroscopically characterize and follow the reaction kinetics of coordinatively unsaturated species in real time. Since desired timescales for reaction will typically be in the microsecond to sub-microsecond range, a system with a rapid time response will be required. Transient absorption systems employing a visible or UV probe which meet this criterion have been developed and have provided valuable information for metal carbonyl systems [14,15,27]. However, since metal carbonyls are extremely photolabile and their UV-visible absorption spectra are not very structure sensitive, the preferred choice for a spectroscopic probe is time resolved infrared spectroscopy. Unfortunately, infrared detectors are enormously less sensitive and significantly slower... 

Atomic metal ion-hydrocarbon reactions bond dissociation energies for fragments, 15,16t endothermic reactions, 13,15,17f Atomic transition metal ion reactions development of approach for real-time measurements of dissociation kinetics, 39 ion beam apparatus, 12,14f studies of... 

Ramanujan, V. K., Biener-Ramanujan, E., Armmer, K., Centonze, V. E. and Herman, B. A. (2006). Spectral kinetics ratiometry A simple approach for real-time monitoring of fluorophore distributions in living cells. Cytometry A 69, 912-9. 

The changes in a and f> can be determined very rapidly, allowing changes in film thickness and optical properties to be determined in real time, hence the technique can be used to gain kinetic information. The speed of data collection is limited only by the response of the instrument and this may be as low as a few tens of microseconds. 

Polzius R., Diessel. E., Bier F., Bilitewski U., Real-Time Observation of Affinity Reactions Using Grating Couplers Determination of the Detection Limit and Calculation of Kinetic Rate Constants, Analyt. Biochem. 1997 249 269-276. 

The background problem can be further overcome when using a surface-confined fluorescence excitation and detection scheme at a certain angle of incident light, total internal reflection (TIR) occurs at the interface of a dense (e.g. quartz) and less dense (e.g. water) medium. An evanescent wave is generated which penetrates into the less dense medium and decays exponentially. Optical detection of the binding event is restricted to the penetration depth of the evanescent field and thus to the surface-bound molecules. Fluorescence from unbound molecules in the bulk solution is not detected. In contrast to standard fluorescence scanners, which detect the fluorescence after hybridization, evanescent wave technology allows the measurement of real-time kinetics (www.zeptosens.com, www.affinity-sensors.com). 

The PHSS method of real-time H2S measurement allows for investigating the potentially complex H2S kinetic responses of organs, tissues, cells, and mitochondria as levels of 02 and NO as well as metabolic state are adjusted within physiological limits. Kinetic changes in H2S concentration continuously reported by the PHSS, which are not seen with other H2S measurement techniques, suggest potentially complex interactions of H2S production and consumption mechanisms. H2S may likely exist as a cellular pool of free and labile persulfides able to rapidly respond to redox challenges with production and consumption pathways that operate to maintain the pool. This possible scenario reinforces the need for the PHSS as a valuable tool to provide a continual report of H2S throughout the course of an experimental treatment or to accurately determine H2S levels in situ. 

Vilan A, Ussyshkin R, Gartsman K, Cahen D, Naaman R, Shanzer A (1998) Real-time electronic monitoring of adsorption kinetics evidence for two-site adsorption mechanism of dicarboxylic acids on GaAs(100). J Phys Chem B 102(18) 3307-3309... 

The objective of the present work was to determine the influence of the light intensity on the polymerization kinetics and on the temperature profile of acrylate and vinyl ether monomers exposed to UV radiation as thin films, as well as the effect of the sample initial temperature on the polymerization rate and final degree of cure. For this purpose, a new method has been developed, based on real-time infrared (RTIR) spectroscopy 14, which permits to monitor in-situ the temperature of thin films undergoing high-speed photopolymerization, without introducing any additive in the UV-curable formulation 15. This technique proved particularly well suited to addressing the issue of thermal runaway which was recently considered to occur in laser-induced polymerization of divinyl ethers 13>16. 

Figure 3.1 shows a typical laboratory flow reactor for the study of catalytic kinetics. A gas chromatograph (GC, lower shelf) and a flow meter allow the complete analysis of samples of product gas (analysis time is typically several minutes), and the determination of the molar flow rate of various species out of the reactor (R) contained in a furnace. A mass spectrometer (MS, upper shelf) allows real-time analysis of the product gas sampled just below the catalyst charge and can follow rapid changes in rate. Automated versions of such reactor assemblies are commercially available. 

The most common methodology for measuring fast kinetics in real time is to perturb a system at equilibrium for a time duration that is much shorter than the relaxation kinetics that follow perturbation. This perturbation can be achieved by changing the concentration of chemicals through fast mixing (stopped-flow), changing the temperature of the solution (temperature jump), simultaneously changing the... 

In a typical SPR experiment real-time kinetic study, solution flows over the surface, so desorption of the guest immobilized on the surface due to this flow must be avoided.72 In the first stage of a typical experiment the mobile reactant is introduced at a constant concentration ([H]0) into the buffer flowing above the surface-bound reactant. This favors complex association, and the progress of complex formation at the surface is monitored. The initial phase is then followed by a dissociation phase where the reactant is removed from the solution flowing above the surface, and only buffer is passed over the surface to favor dissociation of the complex.72 74 The obtained binding curves (sensograms) contain information on the equilibrium constant of the interaction and the association and dissociation rate constants for complex formation (Fig. 9). 

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