Deconvolution of fluorescence spectra: contribution to the structural analysis of complex molecules

Fluorescence spectroscopy is a sensitive analytical tool in the studies of both simple and complex molecular structures. In complex molecules, however, determining the number and position of components may give a specific insight into the structure, complementary to the other analytical techniques. We applied log–normal model to analyze fluorescence of simple monofluorophore molecule. In order to analyze spectra where both fluorophores and Raman emission bands were present, we developed a method obtained by combination of the symmetric, Gaussian, for Raman and asymmetric, log–normal model, for fluorescence, applicable to the molecules of different complexity. Technically, for each sample we varied excitation wavelength with 5 nm step and recorded the corresponding emission spectra. They were subsequently used for component analysis. Position of each component was plotted against the excitation wavelength. Applying this approach we could identify minimal number of components having stable positions, while their approximate probability density (APD) in a spectral series was correlated with the probable number of fluorophores in the molecule. The method was tested on molecules containing different number of fluorophores: monomers involved in the synthesis of plant polymer lignin—coniferyl alcohol (one fluorophore), ferulic acid (two fluorophores) and on lignin model compound produced from these monomers (many fluorophores). All investigated species belong to benzene-substituted class of compounds, and it is reasonable to assume that they have similar fluorescence band contour. We also report the results of environmental scanning electron microscopy (ESEM) studies showing multilayered dehydrogenative polymer (DHP) structure, in order to show complexity of the polymer. Our results present complementarity of these two approaches in the structural studies of the lignin model compound.     

A critique on the structural analysis of lignins and application of novel tandem mass spectrometric strategies to determine lignin sequencing

This review is devoted to the application of MS using soft ionization methods with a special emphasis on electrospray ionization, atmospheric pressure photoionization and matrix‐assisted laser desorption/ionization MS and tandem MS (MS/MS) for the elucidation of the chemical structure of native and modified lignins. We describe and critically evaluate how these soft ionization methods have contributed to the present‐day knowledge of the structure of lignins. Herein, we will introduce new nomenclature concerning the chemical state of lignins, namely, virgin released lignins (VRLs) and processed modified lignins (PML). VRLs are obtained by liberation of lignins through degradation of vegetable matter by either chemical hydrolysis and/or enzymatic hydrolysis. PMLs are produced by subjecting the VRL to a series of further chemical transformations and purifications that are likely to alter their original chemical structures. We are proposing that native lignin polymers, present in the lignocellulosic biomass, are not made of macromolecules linked to cellulose fibres as has been frequently reported. Instead, we propose that the lignins are composed of vast series of linear related oligomers, having different lengths that are covalently linked in a criss‐cross pattern to cellulose and hemicellulose fibres forming the network of vegetal matter. Consequently, structural elucidation of VRLs, which presumably have not been purified and processed by any other type of additional chemical treatment and purification, may reflect the structure of the native lignin. In this review, we present an introduction to a MS/MS top–down concept of lignin sequencing and how this technique may be used to address the challenge of characterizing the structure of VRLs. Finally, we offer the case that although lignins have been reported to have very high or high molecular weights, they might not exist on the basis that such polymers have never been identified by the mild ionizing techniques used in modern MS. Copyright © 2015 John Wiley & Sons, Ltd.     

A comparative study of the dioxane lignins ofAlthea

Summary 1. The dioxane lignins have been extracted from three species ofAlthea for the first time. On the basis of elementary and functional analyses, the semiempirical formulas of the phenylpropane structural units have been calculated.2. The molecular weight distributions have shown that the DLAs of these species ofAlthea are polydisperse, and in the case of the DLAs of the wild-growing species —A. rhyticarpa andA. nudiflora — the high-molecular weight fraction is predominant, while for the DLA ofA. rosea the low-molecular-weight fraction is predominant.3. In a study of the products of nitrobenzene oxidation it has been established that the DLA of all three species ofAlthea includes p-coumaryl, guaiacyl, and syringyl structural units.4. In the products of decomposition with metallic sodium in liquid ammonia 18 phenols have been identified the structures of which show the presence in the initial lignins of guaiacyl- and syringylpropane units with free OH groups in the side chains in the - and -positions to the aromatic nucleus.Institute of the Chemistry of Plant Substances, Academy of Sciences of the Uzbek SSR, Tashkent. Translated from Khimiya Prirodnykh Soedinenii, No. 1, pp. 93–100, January–February, 1977.     

Component analysis of the fluorescence spectra of a lignin model compound

In order to test whether lignin fluorescence originates from discrete fluorophores, fluorescence emission spectra of the lignin model dehydrogenative polymer (DHP) were analyzed by the band deconvolution method and time-resolved analysis of both the excitation and emission spectra. Two series of 22 fluorescence emission spectra of DHP in chloroform/methanol (3:1, v/v) solution, and as a solid suspension in water, were deconvoluted into three fluorescence and one Raman Gaussian components. Emission spectra were obtained by stepwise variation of the excitation wavelength from 360 to 465 nm. Deconvolution was performed by nonlinear fitting of all three Gaussian parameters: area, width and position. Position of all components in a series was treated as a random variable and its approximate probability distribution (APD) calculated from a series of histograms with increasing number of abscissa intervals. A five peak multimodal APD profile was obtained for both series of DHP emission spectra. The mean fluorescence lifetime varied with wavelength both in the emission and the excitation decay-associated spectra (DAS), where four kinetic components were resolved. The shapes of the excitation spectra of the four components were quite different and gradually shifted bathochromically. The multicomponent nature of the DHP emission spectra along with the changes in the mean fluorescence lifetime and the form of the excitation DAS of the four components give evidence of the heterogeneous origin of fluorescent species emitting in the visible.     

The mass spectra of TMS-ethers of deuterated polyalcohols. A contribution to the structural investigation of sugars

Aldoses, ketoses, deoxy-aldoses, deoxy-keto-aldoses, lactones, deoxy-lactones and other similar compounds are readily reduced by NaBD₄ to the corresponding polyalcohol. Their trimethylsilyl ethers show characteristic mass spectra which allow firm assignments of the position of deuteration and hence of the nature of the starting sugar compound. The g.c./m.s. coupling technique even allows trace analysis since full information can be derived from the most intense fragments of the mass spectra.     

A contribution to the theory of the structural glass transition

The equations characterizing the temperature dependence of the mobility of molecules in the glass transition region are compared and analysed. A relationship between the parameters of the relaxation theory of the glass transition and the concept of free volume is established. The Williams-Landel-Ferry equation is treated in terms of the relaxation theory of glass transition.     

Low temperature structural fluorescence spectra of tetrahydroxyanthraquinone adsorbed on silica

Adsorption of TOAQ on the surface of porous (silica gel) and non-porous (aerosil) silica modification does not cause rupture of intramolecular hydrogen bonds (IMHB). The fine structure spectra of TOAQ fluorescence on silica gel contain a set of frequencies that are characteristic of weakly perturbed molecules. A number of new spectral effects appear due to the existence of pores and centres of a nonhydroxyl character. Hydroxyl groups of aerosil take part in formation of intermolecular hydrogen bonding with the TOAQ C=O group. The bond energy of the four OH-groups of the TOAQ molecule changes non-uniformly, and the symmetry of an equilibrium nuclear configuration of TOAQ decreases.     

Vibrational spectroscopy studies of structural changes in lignin under microwave irradiation

Structural changes that occur in lignin surface-modified with nickel nanoparticles during microwave- assisted dry reforming (DR) are studied via vibrational spectroscopy. IR spectroscopy reveals that the nickel deposition has a considerable effect on the structural characteristics of lignin. It is found that nickel deposition from an acetate salt substantially reduces the intensity of absorption bands at 1700 cm^?1. This finding suggests that Ni(2+) interacts mostly with formate groups, which are subsequently oxidized to carboxylate groups. It is shown that with the deposition of metallic nickel particles from a colloidal nickel solution in toluene prepared via metal vapor synthesis, the nickel particles do not interact with the surface functional groups of the lignin. Deep conversion of an organic mass of lignin by DR to form synthesis gas reduces the intensity of the absorption bands of the identified functional groups and raises the intensity of the absorption bands of the aromatic rings. Raman spectroscopy shows that during lignin conversion, the aromatic rings condense partially to form amorphized graphite. In operando studies reveal that the DR of nickel-modified lignin heated to 200–400°C results in the isolation of vanillic oxygenates that are probably intermediate products of reforming.     

PMR spectra of the lignins ofCystoseira barbata at different ages

The PMR spectra of the dioxane lignins from the brown algaCystoseira have been studied. It has been established that the lignins isolated from specimens ofCystoseira of different ages have different degrees of substitution of the C₃ side chain and differ in their degree of condensation. The most highly condensed is the dioxane lignin of young specimens ofCystoseira (age up to one year) and the least condensed the Björkman lignin ofCystoseira aged more than three years. All the PMR spectra of preparations of the lignin of the alga under investigation contain the signals of protons present in coumaran structures, their amount being the greatest in the lignins of the youngest samples and decreasing with increasing age of the alga.Odessa Institute of Structural Engineering. Translated from Khimiya Prirodnykh Soedinenii, No. 1, pp. 91–94, January–February, 1983.     

A comparison of the surface properties of lignin and sulfonated lignins by FTIR spectroscopy and wicking technique

The comparison of the FTIR spectra indicated that the sulfonation without caused changes in the main structure of lignin and only in the lignin chains. The surface properties, e.g. the surface free energy and related components, e.g. the Lifshitz–van der Waals and Lewis acid–base interactions components, of lignin and lignosulfonates, LGSs, with different ions, e.g. Na⁺–, Mg²⁺–, and Ca²⁺–, studied and compared by wicking technique indicated that the surface free energy of lignin is lower than that of all used LGSs. This suggests that the sulfonation can enhance the surface property for lignin by increasing of the Lifshitz–van der Waals component and Lewis base component of lignin.     

A comparative study of LED-induced fluorescence and laser-induced fluorescence in SDS-CGE: application to the analysis of antibodies

LEDs present an alternative to lasers in LIF detection with CE, resulting in LED-induced fluorescence (LEDIF). LEDs are much less expensive, consume less energy and are more stable. In addition, LED light sources allow a greater range of wavelengths to better match the maximum wavelength for the fluorescence of the dye. Antibodies were largely studied in SDS capillary gel electrophoresis (SDS-CGE) and LIF detection with different dyes to label the proteins. In this work, our goal is to show that LEDs can advantageously replace lasers. We used 5-carboxytetramethylrhodamine succinimidyl ester (5-TAMRA.SE), 3-(2-furoyl)-quinoline-2 carboxaldehyde (FQ), and naphthalene-2,3-dialdehyde (NDA) to label IgG and we compared the LIF sensitivity with that obtained from LEDIF. We measured that the LOD values of LEDIF are identical to that obtained with the wavelength equivalent laser, and for 5-TAMRA.SE analysis, LOD values are about six times better than when the classical 488 nm laser was used.     

Thermal and structural studies of polypropylene blended with esterified industrial waste lignin

Microwave-assisted chemical modification of lignin was achieved through esterification using maleic anhydride. Modified lignin (ML) was blended in different proportions up to 25 mass% with polypropylene (PP) using Brabender electronic Plasticorder at 190 °C. The structural and thermal properties of blends were investigated by thermogravometric analysis (TG), differential scanning calorimetry (DSC), wide-angle X-ray diffraction (WAXD) and scanning electron microscopy (SEM). TG analysis showed increased thermal stability of blends due to antioxidant property of ML, which opposed oxidative degradation of PP. DSC analysis indicted slight depression in a glass transition temperature and melting temperature of blends due to partial miscible blend behavior between PP and ML. All blends showed higher crystallization temperatures and continuously reducing percentage crystallinity with increasing ML proportion in the blends. WAXD analysis indicated that PP crystallized in β polymeric form in addition to α-form in the presence of ML. However, proportion of β-form did not show linear relation with increase in ML proportion, thus ML acts as β nucleating agent in the PP matrix. SEM analysis showed good dispersion/miscibility in PP matrix indicating modification in lignin is useful.     

Effect of ionic liquid treatment on the structures of lignins in solutions: molecular subunits released from lignin

The solution structures of three types of isolated lignin-organosolv (OS), Kraft (K), and low sulfonate (LS)-before and after treatment with 1-ethyl-3-methylimidazolium acetate were studied using small-angle neutron scattering (SANS) and dynamic light scattering (DLS) over a concentration range of 0.3-2.4 wt %. The results indicate that each of these lignins is comprised of aggregates of well-defined basal subunits, the shapes and sizes of which, in D(2)O and DMSO-d(6), are revealed using these techniques. LS lignin contains a substantial amount of nanometer-scale individual subunits. In aqueous solution these subunits have a well-defined elongated shape described well by ellipsoidal and cylindrical models. At low concentration the subunits are highly expanded in alkaline solution, and the effect is screened with increasing concentration. OS lignin dissolved in DMSO was found to consist of a narrow distribution of aggregates with average radius 200 ± 30 nm. K lignin in DMSO consists of aggregates with a very broad size distribution. After ionic liquid (IL) treatment, LS lignin subunits in alkaline solution maintained the elongated shape but were reduced in size. IL treatment of OS and K lignins led to the release of nanometer-scale subunits with well-defined size and shape.     

PMR spectra of the dioxane lignins of some plants of the family Malvaceae

Analysis of PMR spectra of the dioxane lignins of three species ofAthea, of kenaf, and of the cotton plant has shown that the number of free aromatic protons does not correlate with the number of OCH₃ groups, which indicates different degrees of condensation of the dioxane lignins studied. In all the preparations a very small amount of coumarane structures were found. The aliphatic OH groups have been distinguished as primary (γ-OH) and secondary (α-OH) and their quantitative estimation has been carried out from the PMR spectra.     

PMR spectra of the dioxane lignins of some plants of the family Malvaceae

Analysis of PMR spectra of the dioxane lignins of three species ofAthea, of kenaf, and of the cotton plant has shown that the number of free aromatic protons does not correlate with the number of OCH₃ groups, which indicates different degrees of condensation of the dioxane lignins studied. In all the preparations a very small amount of coumarane structures were found. The aliphatic OH groups have been distinguished as primary (-OH) and secondary (-OH) and their quantitative estimation has been carried out from the PMR spectra.Institute of the Chemistry of Plant Substances, Academy of Sciences of the Uzbek SSR, Tashkent. Translated from Khimiya Prirodnykh Soedinenii, No. 4, pp. 560–563, July–August, 1980.     

Relevance of the electric-dipole--electric-quadrupole contribution to Raman optical activity spectra

We examine the importance of the electric-dipole--electric-quadrupole polarizability tensor in the intensity theory of Raman optical activity (ROA) spectra. Using density functional theory, ROA spectra of organic cyclic compounds, alanine, oligoalanines, and examples from the literature are analyzed in detail, and a statistical investigation is performed. It is found that the contribution of the electric-dipole--electric-quadrupole tensor is often small, except for some special cases that involve C-H stretching vibrations.     

Captopril and its dimer captopril disulfide: comparative structural and conformational studies

The crystal structures of captopril {systematic name: (2S)‐1‐[(2S)‐2‐methyl‐3‐sulfanylpropanoyl]pyrrolidine‐2‐carboxylic acid}, C₉H₁₅NO₃S, (1), and its dimer disulfide metabolite, 1,1′‐{disulfanediylbis[(2S)‐2‐methyl‐1‐oxopropane‐3,1‐diyl]}bis‐L‐proline, C₁₈H₂₈N₂O₆S₂, (2), were determined by single‐crystal X‐ray diffraction analysis. Compound (1) crystallizes in the orthorhombic space group P2₁2₁2₁, while compound (2) crystallizes in the monoclinic space group P2₁, both with one molecule per asymmetric unit. The molecular geometries of (1) and (2) are quite similar, but certain differences appear in the conformations of the five‐membered proline rings and the side chains containing the sulfhydryl group. The proline ring adopts an envelope conformation in (1), while in (2) it exists in envelope and slightly deformed half‐chair conformations. The conformation adopted by the side chain is extended in (1) and folded in (2). A minimum‐energy conformational search using Monte Carlo methods in the aqueous phase reveals that the optimized conformations of the title compounds differ from those determined crystallographically, which depend on their immediate environment. Intermolecular O—H...O and relatively weak C—H...O interactions seem to be effective in both structures and, together with S—H...O and C—H...S contacts, they create three‐dimensional networks.     

A structural comparative approach to identifying novel antimalarial inhibitors

Malaria continues to affect millions of people annually. With the rise of drug resistant strains, the need for alternative treatments has become increasingly urgent. Recently, PfUCHL3 was identified as an essential deubiquitinating enzyme. The increasing number of drug target structures being solved has increased the feasibility of utilizing a structural comparative approach to identifying novel inhibitors. Using AutoDock Vina, we recently screened the NCI library of about 320,000 compounds against the crystal structure of PfUCHL3. The top hits were subsequently screened against its human ortholog UCHL3 as to identify compounds that could specifically target the PfUCHL3 over its human counterpart. This method was used to identify small molecule inhibitors that can preferentially inhibit the parasitic enzyme. Several compounds were identified that demonstrated significant binding affinity preference for the malaria target over the human enzyme. Two of these compounds demonstrated ng/mL activity.