Three-dimensional Fluorescence Data Research Articles

Determination of monophenolase activity based on backpropagation neural network analysis of three-dimensional fluorescence spectroscopy

Tyrosinase is pivotal for melanin formation. Measuring monophenolase activity is of great importance for both fundamental research and industrial applications. For the first time, a backpropagation (BP) artificial neural network with three-dimensional fluorescence spectroscopy was applied for the real-time determination of tyrosinase monophenolase activity. Principal component analysis (PCA) was utilized for the dimension reduction of three-dimensional fluorescence data. The four principal components served as inputs for the neural network. Network parameters were optimized using a genetic algorithm (GA). BP learning algorithm was applied to train the network model to determine tyrosine levels in a binary mixture containing tyrosine and L-DOPA without any chemical separation. The time course of tyrosine consumption by monophenolase was determined to calculate the initial velocity of the enzymatic reaction. The limit of detection of the monophenolase assay was 0.0615 U·mL−1. This combined strategy of PCA, GAs, and BP artificial neural networks for three-dimensional fluorescence spectroscopy was efficient for the real-time and in-situ determination of monophenolase activity in a cascade reaction.

Measurement report: Particle-size-dependent fluorescence properties of water-soluble organic compounds (WSOCs) and their atmospheric implications for the aging of WSOCs

Abstract. Water-soluble organic compounds (WSOCs) play important roles in atmospheric particle formation, migration, and transformation processes. Size-segregated atmospheric particles were collected in a rural area of Beijing. Three-dimensional fluorescence spectroscopy was used to investigate the optical properties of WSOCs as a means of inferring information about their atmospheric sources. Sophisticated analysis on fluorescence data was performed to characteristically estimate the connections among particles of different sizes. WSOC concentrations and the average fluorescence intensity (AFI) showed a monomodal distribution in winter and a bimodal distribution in summer, with the dominant mode in the 0.26–0.44 µm size range in both seasons. The excitation–emission matrix (EEM) spectra of WSOCs varied with particle size, likely due to changing sources and/or the chemical transformation of organics. Size distributions of the fluorescence regional integration (regions III and V) and humification index (HIX) indicate that the humification degree or aromaticity of WSOCs was the highest in the particle size range of 0.26–0.44 µm. The Stokes shift (SS) and the harmonic mean of the excitation and emission wavelengths (WH) reflected that π-conjugated systems were high in the same particle size range. The parallel factor analysis (PARAFAC) results showed that humic-like substances were abundant in fine particles (< 1 µm) and peaked at 0.26–0.44 µm. All evidence supported the fact that the humification degree of WSOCs increased with particle size in the submicron mode (< 0.44 µm) and then decreased gradually with particle size, which implied that the condensation of organics occurred in submicron particles, resulting in the highest degree of humification in the particle size range of 0.26–0.44 µm rather than in the < 0.26 µm range. Synthetically analyzing three-dimensional fluorescence data could efficiently reveal the secondary transformation processes of WSOCs.

Rapid Assessment of Total Polar Material in Used Frying Oils Using Manganese Tetraphenylporphyrin Fluorescent Sensor with Enhanced Sensitivity

Probe-based three-dimensional fluorescence spectroscopy was developed for the rapid assessment of frying oil quality. Tetraphenylporphyrin manganese (MnTPP) was selected due to its sensitivity and selectivity in response to dissolved organic matter, which is typically produced during the frying process of oil analyte. Parallel factor analysis (PARAFAC) was then performed to decompose three-dimensional fluorescence data for model construction. The digital data representing the change of the MnTPP-base fluorescent probe before and after exposure to dissolved organic matter were analyzed using support vector regression (SVR). SVR was then used for the construction of the regression model between the fluorescence data and the total polar material (TPM). The coefficients of determination (R2) in the training set (R2 = 0.9719) and the test set (R2 = 0.9699) were achieved to evaluate the robustness of the best models for TPM. This study demonstrated that the probe-based three-dimensional fluorescence spectroscopy is useful for assessing the quality of frying oil and perhaps could be used for other food products with high oil contents.

Binding interactions between pepsin and 3,3′,4,4′,5-pentachlorobiphenyl

One member of the polychlorinated biphenyls that is linked to a number of human diseases is 3,3′,4,4′,5-pentachlorobiphenyl. Here, density functional theory calculation, molecular simulations and multi-spectroscopic techniques were utilized to evaluate the interaction mechanism of 3,3′,4,4′,5-pentachlorobiphenyl with the pepsin, the digestive enzyme. The results of density functional theory and molecular simulations demonstrated that 3,3′,4,4′,5-pentachlorobiphenyl could be a good candidate for its availability toward pepsin with a frontier orbital gap of 0.18109 eV, and bound to the hydrophobic cavity of pepsin. The steady-state fluorescence and time-resolved fluorescence revealed that the quenching was initiated by complexation of pepsin and 3,3′,4,4′,5-pentachlorobiphenyl with the binding constant of 105 order. The UV-absorption, synchronous fluorescence spectroscopy, Fourier transform infrared spectroscopy, circular dichroism spectroscopy, and three-dimensional fluorescence data further confirmed that the changes in the secondary structure of pepsin were induced on the presence of 3,3′,4,4′,5-pentachlorobiphenyl, and a quantitative analysis of the secondary structure elements in pepsin was obtained.

Fluorometric and molecular docking investigation on the binding characteristics of SB202190 to human serum albumin

The interaction of SB202190, a p38 mitogen-activated protein kinase inhibitor with the main drug transporter in human circulation, human serum albumin (HSA) was studied using fluorescence spectroscopy and in silico docking methods. The association constant, Ka of the binding reaction was determined to be 3.24±0.07×104M−1 at 25°C based on fluorescence quenching titration results. The values of enthalpy change and entropy change for the interaction were found as −8.54kJmol−1 and 58.01Jmol−1K−1, respectively. Both thermodynamic data and docking results suggested the involvement of hydrophobic and van der Waals forces in the complex formation. Three-dimensional fluorescence data of SB202190–HSA complex demonstrated significant changes in the microenvironment around the protein fluorophores upon drug binding. Comparison of HSA thermograms obtained in the absence and the presence of SB202190 suggested improved protein thermal stability upon complexation with the drug. Competitive drug displacement results as well as modeling data concluded the preferred binding site of SB202190 on HSA as Sudlow׳s site I.

Exploring the interaction between the antiallergic drug, tranilast and human serum albumin: Insights from calorimetric, spectroscopic and modeling studies

The interaction of tranilast (TRN), an antiallergic drug with the main drug transporter in human circulation, human serum albumin (HSA) was studied using isothermal titration calorimetry (ITC), fluorescence spectroscopy and in silico docking methods. ITC data revealed the binding constant and stoichiometry of binding as (3.21±0.23)×106M−1 and 0.80±0.08, respectively, at 25°C. The values of the standard enthalpy change (ΔH°) and the standard entropy change (ΔS°) for the interaction were found as −25.2±5.1kJmol−1 and 46.9±5.4Jmol−1K−1, respectively. Both thermodynamic data and modeling results suggested the involvement of hydrogen bonding, hydrophobic and van der Waals forces in the complex formation. Three-dimensional fluorescence data of TRN–HSA complex demonstrated significant changes in the microenvironment around the protein fluorophores upon drug binding. Competitive drug displacement results as well as modeling data concluded the preferred binding site of TRN as Sudlow’s site I on HSA.

A novel pretreatment method of three-dimensional fluorescence data for quantitative measurement of component contents in mixture

Three-dimensional fluorescence technique is commonly used for the determination of component contents in the mixture. Fluorescence intensity data are used directly in the fluorescent spectrum data processing method. The relationship between fluorescence intensity values and concentrations is linear. Random noise is inevitable in the process of measuring due to fluorescence spectrometer. The measurement accuracy is reduced due to the existence of noise. To reduce random noise and improve the measurement sensitivity, a novel pretreatment method of three-dimensional fluorescence data is proposed. The method is based on Quasi-Monte-Carlo integral. Due to the increased slope of fluorescence intensity data during the integral, the measurement sensitivity is improved. At the same time, the sum of different exponentials of fluorescence intensity at the points reduces the random noise, so the measurement sensitivity is improved more. The recovery rates of the mixture mixed by gasoline, kerosene and diesel oil are calculated to validate the effectiveness of the method.

Nonequilibrium Mechanics of the Mitotic Spindle Measured in Living Cells

To carry out its life cycle and produce viable progeny through cell division, a cell must successfully coordinate and execute a number of complex non-equilibrium processes with high fidelity, in an environment dominated by thermal noise. One important example of such a process is the assembly of and maintenance of tension across the mitotic spindle, a nonequilibrium composite material including polymers and motor proteins that is responsible for organizing and separating the genetic material during cell division. The intrinsic microtubule dynamics and different motor proteins provide the forcing required for this dynamic process. We use high-resolution fluorescence confocal microscopy to observe and analyze the real space dynamics of the mitotic spindle in budding yeast and Drosophila melanogaster S2 cells. Centrosome trajectories are reconstructed from the three-dimensional fluorescence data and quantified in a coordinate system relevant to the cell division using specially-developed image analysis methods. The roles of specific motor proteins are isolated by altering their functionality through genetic and chemical means using a microfluidic device. We use the fluctuations in pre-anaphase centrosome positions to show how nonequilibrium motor activity controls the mechanical properties of an in vivo cytoskeletal network.