Aspects of transition cow metabolomics-Part I: Effects of a metaphylactic butaphosphan and cyanocobalamin treatment on the metabolome in liver, blood, and urine in cows with different liver metabotypes.

Dairy cows in modern production systems are at risk to develop metabolic disorders during the transition period. Reasons for individual differences in susceptibility, as well as the underlying pathomechanisms, are still only partially understood. The development of metaphylactic treatment protocols is needed. In this context, an on-farm prospective 3-fold blinded randomized study involving 80 German Holstein cows was performed throughout 1 yr. The trial involved a thorough recording of the production and clinical traits, clinical chemistry, and liver biopsies and blood and urine sampling at d 14 (mean: 12 d, range: 1-26 d) antepartum (AP), and d 7 (7, 4-13) and 28 (28, 23-34) postpartum (PP) for metabolomics analyses. Two groups received a treatment with butaphosphan and cyanocobalamin (BCC) at either the dosage recommended by the manufacturer or the double dosage (5 or 10 mL/100 kg of body weight 10% butaphosphan and 0.005% cyanocobalamin (Catosal, Bayer Animal Health), n = 20 in each group, parity: 4.2 ± 2.0 and 3.4 ± 1.3, respectively (mean ± SD)] and one group a placebo treatment (NaCl 0.9%, n = 40, parity: 4.0 ± 1.9). The animals were treated at 6 time points (7, 6, and 5 d AP, and 1, 2, and 3 d PP) via intravenous injection. Mass spectroscopy-based targeted metabolomics analysis of blood plasma and liver samples were performed using the AbsoluteIDQ p180 kit (Biocrates Life Sciences), whereas the urine samples were analyzed by nuclear magnetic resonance spectroscopy. Statistical analysis was performed using multivariate [partial least squares discriminant analysis (PLS-DA)] and univariate methods (linear mixed model). Multivariate data analysis (PLS-DA plots) of the liver metabolome revealed 3 different metabotypes (A = medium, B = minor, C = large alterations in liver metabolome profile between AP and PP status). Metabotype B animals were characterized by higher PP lipomobilization (stronger PP body condition decrease and higher blood bilirubin, fatty acids, gamma-glutamyltransferase, and triglyceride levels) and a higher occurrence of transition cow diseases, compared with the animals in metabotype C. Analysis of the feeding data showed that the period of metabotype B animals (calving in a distinct time frame) was characterized by a decreased grass silage quality. The PP liver metabolome of the metabotype C animals was characterized by higher concentrations of AA, acylcarnitines, lysoPC and sphingomyelins compared with metabotype B. For the metaphylactic treatment with BCC a dose-dependent effect was confirmed, differing between the metabotypes. In all matrices and metabotypes at various time points significant treatment effects were observed, with different profiles in clinical chemistry and as well in metabolomics data. The most clear-cut treatment effect was observed in metabotype B in the liver at 7 d PP, characterized by an increase in several acylcarnitines and phosphatidylcholines, indicating a more efficient influx and oxidation of fatty acids in mitochondria and thereby an increase in energy supply and more efficient triglyceride export in the liver. The results from the liver metabolomics analysis support the application of an indication-based metaphylactic treatment with BCC.

Aspects of transition cow metabolomics-Part III: Alterations in the metabolome of liver and blood throughout the transition period in cows with different liver metabotypes.

The liver plays a central role in the postpartum (PP) energy metabolism of the transition dairy cow; however, studies describing the liver metabolome during this period were lacking. The aim of the presented study was therefore to compare the alterations in the liver and blood metabolome of transition dairy cows. For this purpose, an on-farm trial with 80 German Holstein cows (mean lactation number: 3.9; range: 2-9) was performed, with thorough documentation of clinical traits and clinical chemistry, as well as production data. Liver biopsies and blood samples were collected at d 14 (mean: 12 d, range: 1-26 d) antepartum (AP), d 7 (7, 4-13) and 28 (28, 23-34; mean, earliest-latest) PP for targeted mass spectroscopy-based metabolomics analysis using the AbsoluteIDQ p180 kit (Biocrates Life Sciences). Statistical analysis was performed using multivariate (partial least squares discriminant analysis) as well as univariate methods (linear mixed model). Multivariate data analysis of the liver metabolome revealed 3 different metabotypes (A = medium, B = minor, C = large alterations in the liver metabolome profile between AP and PP). In metabotype C, an increase of almost all acylcarnitines, lysophosphatidylcholines (lysoPC), sphingomyelins, and some phosphatidylcholines (PC, mainly at 7 d PP) was observed after calving. In contrast to metabotype C, the clinical data of the metabotype B animals indicated a higher PP lipomobilization and occurrence of transition cow diseases. The liver metabolome profile of these animals most likely mirrors a failure of adaptation to the PP state. This strong occurrence of metabotypes was much less pronounced in the blood metabolome. Additionally, differences in metabolic patterns were observed across the transition period when comparing liver and blood matrices (e.g., in different biogenic amines, acylcarnitines and sphingolipids). In summary, the blood samples at 7 d PP showed lower acylcarnitines and PC, with minor alterations and a heterogeneous pattern in AA, biogenic amines, and sphingomyelins compared with 14 d AP. In contrast to 7 d PP, the blood samples at 28 PP revealed an increase in several AA, lysoPC, PC, and sphingomyelins in comparison to the AP state, irrespective of the metabotype. In the liver biopsies metabotype B differed from metabotype C animals ante partum by following metabolites: higher α aminoadipic acid, lower AA, serotonin, taurine, and symmetric dimethylarginine levels, lower or higher concentrations of certain acylcarnitines (higher: C2, C3, C5, C4:1; lower: C12:1, C14:1-OH, C16:2), and lower lysoPC (a C16:0, C18:0, C20:3, C20:4) and hexose levels. In blood samples, fewer differences were observed, with lower serotonin, acylcarnitine C16:2, lysoPC (a C16:0, C17:0, C18:0 and C18:1), PC aa C38:0, and PC ae C42:2. The results show that the use of only the blood metabolome to assess liver metabolism may be hampered by the fact that blood profiles are influenced by the metabolism of many organs, and metabolomics analysis from liver biopsies is a more suitable method to identify distinct metabotypes. Future studies should investigate the stability and reproducibility of the metabotype and phenotypes observed, and the possible predictive value of the metabolites already differing AP between metabotype B and C.

Density Functional Study on the Intercalation of Fullerenes into AnE-PV Copolymer Layers.

We investigated the intercalation of C60 into poly(p-anthracene-ethynylene)-alt-poly(p-phenylene-vinylene) copolymers layers by density functional theory calculations in respect of crystal structures and electronic band structures. Based on the experimental observations, we found that the copolymer with branched side chains substituted next to the anthracene units and the linear side chains substituted to the vinylene units has a better tendency to intercalate with C60 than the reversely substituted copolymer. The calculated electronic band structures of the intercalated phase, featured by flat in-gap states resulting from C60 molecules, explain the experimentally observed variations of the photocurrent, photoluminescence, and electroluminescence yields with different ratio between PCBM and the two types of copolymers in the ternary blend.

Density Functional Study of Stacking Structures and Electronic Behaviors of AnE-PV Copolymer.

In this work, we report an in-depth investigation on the π-stacking and interdigitating structures of poly(p-anthracene-ethynylene)-alt-poly(p-phenylene-vinylene) copolymer with octyl and ethyl-hexyl side chains and the resulting electronic band structures using density functional theory calculations. We found that in the π-stacking direction, the preferred stacking structure, determined by the steric effect of the branched ethyl-hexyl side chains, is featured by the anthracene-ethynylene units stacking on the phenylene-vinylene units of the neighboring chains and vice versa. This stacking structure, combined with the interdigitating structure where the branched side chains of the laterally neighboring chains are isolated, defines the energetically favorable structure of the ordered copolymer phase, which provides a good compromise between light absorption and charge-carrier transport.

Effect of Side Chains on Molecular Conformation of Anthracene-Ethynylene-Phenylene-Vinylene Oligomers: A Comparative Density Functional Study With and Without Dispersion Interaction.

Using density functional calculations with and without dispersion interaction, we studied the effects of linear octyl and branched 2-ethylhexyl side chains on the oligomer conformation of the conjugated copolymer poly(p-anthracene-ethynylene)-alt-poly(p-phenylene-vinylene). With dispersion included, the branched side chains can cause significant bending of the oligomer backbone, while without dispersion they induce mainly torsional disorder. The oligomers with mainly linear side chains keep good planarity when optimized with and without dispersion. Despite their dramatically different conformations, the calculated absorption spectra of the oligomers with various side chain combinations are very similar, indicating that the conformation of the copolymer is not the main reason for the experimentally observed different spectra of ordered and disordered phases.

Quantum chemical insights into the dependence of porphyrin basicity on the meso-aryl substituents: thermodynamics, buckling, reaction sites and molecular flexibility.

The chemical and sensing properties of porphyrins are frequently tuned via the introduction of peripheral substituents. In the context of the exceptionally fast second protonation step in the case of 5,10,15,20-tetraphenylporphyrin (TPP), as compared to porphin and 5,10,15,20-tetramesitylporphyrin (TMesP), we investigated the macrocycle-substituent interactions of these three porphyrin derivatives in detail. Using quantum chemical thermodynamics calculations, the analysis of geometric structures, torsional profiles, electrostatic potential distributions, and particularly the analysis of molecular flexibilities via ab initio molecular dynamics simulations, we obtained a comprehensive picture of the reactivities of the studied porphyrins and how these are influenced by the meso-substituents. As compared to porphin and TMesP the second protonation of TPP is energetically more favorable and is particularly energetically comparable to its first protonation, instead of being significantly less favorable like in the case of porphyrin and TMesP. Additionally, the second TPP protonation is facilitated by an interplay between out-of-plane (oop) distortion of the protonation site and a pronounced electrostatic binding spot at the protonation site. Furthermore, the second protonation is particularly facilitated in the case of TPP by the large oop-flexibility of the diprotonated species as unraveled by ab initio molecular dynamics simulations.

Sub-bandgap absorption in organic solar cells: experiment and theory.

Most high-performance organic solar cells involve bulk-heterojunctions in order to increase the active donor-acceptor interface area. The power conversion efficiency depends critically on the nano-morphology of the blend and the interface. Spectroscopy of the sub-bandgap region, i.e., below the bulk absorption of the individual components, provides unique opportunities to study interface-related properties. We present absorption measurements in the sub-bandgap region of bulk heterojunctions made of poly(3-hexylthiophene-2,5-diyl) as an electron donor and [6,6]-phenyl-C61-butyric acid methyl ester (P3HT:PCBM) as an electron acceptor and compare them with quantum-chemical calculations and recently published data on the external quantum efficiency (EQE). The very weak absorption of the deep sub-bandgap region measured by the ultra-sensitive Photothermal Deflection Spectroscopy (PDS) features Urbach tails, polaronic transitions, conventional excitons, and possibly charge-transfer states. The quantum-chemical calculations allow characterizing some of the unsettled spectral features.

Surface electronic structure of [XMIm]Cl probed by surface-sensitive spectroscopy.

We apply electron spectroscopy methods with different surface sensitivities to elucidate the DOS of the surface and the near-surface region of [XMIm]Cl (X=octyl, hexyl, butyl, and ethyl alkyl chain) ionic liquids. Using metastable induced electron spectroscopy (MIES) we are able to detect the density of states in front of the outermost surface, whereas ultraviolet and X-ray photoelectron spectroscopy (UPS and XPS) measurements provide lower surface sensitivity. The assignment of certain structures in the valence band spectra to particular atoms/functional groups of the ionic liquid based on DFT calculations and the reconstruction of PES spectra enables us to obtain information on the dominating groups at the surface, or in other words, on the molecular/ionic arrangement and orientation at the surface. From angular resolved XPS it is concluded that the alkyl chains dominate at the outermost surface. In agreement with this a decreasing chlorine signal is observed in the UPS spectra for ionic liquids with increasing alkyl chain length. The analysis of the MIES data shows that in case of [OMIm]Cl--in contrast to UPS and XPS--no Cl-induced features are visible in the MIES spectra at all and that the MIES spectra are dominated by the [OMIm](+) alkyl chain.

Theoretical reconstruction and elementwise analysis of photoelectron spectra for imidazolium-based ionic liquids.

We have recently measured core level and valence band XPS, UPS, and MIES spectra of two room temperature ionic liquids composed of bis(trifluoromethylsulfonyl)imide anions ([Tf(2)N](-)) and either 1-ethyl-3-methyl-imidazolium ([EMIm](+)) or 1-octyl-3-methyl-imidazolium cations ([OMIm](+)). [T. Ikari, A. Keppler, M. Reinmöller, W. J. D. Beenken, S. Krischok, M. Marschewski, W. Maus-Friedrichs, O. Höfft and F. Endres, e-J. Surf. Sci. Nanotechnol., 2010, 8, 241.] In the present work we analyze these spectra by means of partial density of states (pDOS) as calculated from a single ion pair of the respective ionic liquid using density functional theory (DFT). Subsequently we reconstruct the XPS and UPS spectra by considering photoemission cross sections and analyze the MIES spectra by pDOS, which provides us decisive hints to the ionic liquid surface structure.

Anomalous energy transfer dynamics due to torsional relaxation in a conjugated polymer.

In isolated conjugated polymers two explanations are in discussion for the redshift of the emission on a picosecond time scale-exciton energy transfer (EET) between conjugated segments along the chains and conformational changes of these segments themselves, i.e., torsional relaxation. In order to resolve this question we perform femtosecond time-resolved transient absorption measurements of the energy relaxation of poly[3-(2,5-dioctylphenyl)thiophene] in toluene solution. We show that torsional relaxation can be distinguished from EET by site-selectively exciting low-energy conjugated segments. We present a unified model that integrates EET and torsional dynamics. In particular, comparison to ultrafast depolarization measurements shows that torsional dynamics cannot be neglected when analyzing EET dynamics and furthermore reveals that the exciton extends itself by about 2 monomer units during torsional relaxation.

Conformational disorder of conjugated polymers.

Conformational disorder of conjugated polymers is an important issue to be understood and quantified. In this paper we present a new method to assess the chain conformation of conjugated polymers based on measurements of intrachain energy transfer. The chain conformation is modeled on the basis of monomer-monomer interactions, such as torsion, bending, and stretching of the connecting bond. The latter two potentials are assumed to be harmonic, while the torsional potential was calculated by density functional theory using B3-LYP functional with the SVP basis set. The energy transfer dynamics of excitons on these chains are quantitatively simulated using Forster-type line-dipole energy transfer. This allows us to compare the simulated ground state conformation of single polymer chains to ultrafast depolarization experiments of poly [3-(2,5-dioctylphenyl)thiophene] in solution. We identify torsional rotation as the main contributor to conformational disorder and find that this disorder is mainly controlled by the energy difference between syn and anti bonds.

Control of emission by intermolecular fluorescence resonance energy transfer and intermolecular charge transfer.

Control of emission by intermolecular fluorescence resonant energy transfer (IFRET) and intermolecular charge transfer (ICT) is investigated with the quantum-chemistry method using two-dimensional (2D) and three-dimensional (3D) real space analysis methods. The work is based on the experiment of tunable emission from doped 1,3,5-triphenyl-2-pyrazoline (TPP) organic nanoparticles (Peng, A. D.; et al. Adv. Mater. 2005, 17, 2070). First, the excited-state properties of the molecules, which are studied (TPP and DCM) in that experiment, are investigated theoretically. The results of the 2D site representation reveal the electron-hole coherence and delocalization size on the excitation. The results of 3D cube representation analysis reveal the orientation and strength of the transition dipole moments and intramolecular or intermolecular charge transfer. Second, the photochemical quenching mechanism via IFRET is studied (here "resonance" means that the absorption spectrum of TPP overlaps with the fluorescence emission spectrum of DCM in the doping system) by comparing the orbital energies of the HOMO (highest occupied molecular orbital) and the LUMO (lowest unoccupied molecular orbital) of DCM and TPP in absorption and fluorescence. Third, for the DCM-TPP complex, the nonphotochemical quenching mechanism via ICT is investigated. The theoretical results show that the energetically lowest ICT state corresponds to a pure HOMO-LUMO transition, where the densities of the HOMO and LUMO are strictly located on the DCM and TPP moieties, respectively. Thus, the lowest ICT state corresponds to an excitation of an electron from the HOMO of DCM to the LUMO of TPP.

Spectral broadening and diffusion by torsional motion in biphenyl.

We have studied biphenyl by time-dependent density-functional theory. In particular, we have analyzed the dependence of singlet excitation energies and transition dipoles on the torsional angle between the phenyl groups. The torsional spectrum has been computed quantum mechanically as well as semiclassically in order to understand how this influences the broadening of absorption and luminescence spectra. Our results are in best agreement with supersonic jet spectroscopy data, but also fit astonishingly well to spectra of biphenyl in condensed phase. Furthermore, we compare the torsional and vibrational relaxation and discuss qualitatively the general consequences for poly-para-phenylenes and related conjugated polymers as poly-thiophenes, considering, in particular, how side chains and solvents may affect the optical spectra.

The electronic states of polyfluorene copolymers with alternating donor-acceptor units.

We calculate the electronic states of the low bandgap polyfluorene-based copolymer DiO-PFDTBT, which consists of alternating 9,9-dioctyl-9H-fluorene and 4,7-di-thiophen-2-ylbenzo[1,2,5]thiadiazole (TBT) units, and compare with the steady-state absorption, emission, and excitation spectrum. Using the semiempirical quantum-chemical (ZINDO) method we can assign the characteristic bands of the "camel-back" absorption spectrum to one charge transfer state at lower energy localized on the TBT unit, and one delocalized excitonic state at higher energy corresponding to the pi-conjugated electron system. Additional "dark" charge transfer states in the gap between these bands have been revealed. Calculations are also made on the red light emitting polyfluorene-based copolymer poly(fluorene-co-benzothiadiazole) (F8BT), which contains benzo[1,2,5]thiadiazole instead of TBT. The nature of the electronic states in F8BT and DiO-PFDTBT are found to be qualitatively the same.

Excitonic coupling in polythiophenes: comparison of different calculation methods.

In conjugated polymers the optical excitation energy transfer is usually described as Forster-type hopping between so-called spectroscopic units. In the simplest approach using the point-dipole approximation the transfer rate is calculated based on the interaction between the transition dipoles of two spectroscopic units. In the present work we compare this approach with three others: The line-dipole approximation, the Coulomb integral between the transition densities, and a quantum-chemical calculation of the interacting dimer as entity. The latter two approaches are based on the semiempirical method ZINDO. The line-dipole approximation is an attractive compromise between computational effort and precision for calculations of the excitonic coupling in extended conjugated polymers.

In vitro self-assembly of the light harvesting pigment-protein LH2 revealed by ultrafast spectroscopy and electron microscopy.

Controlled ensemble formation of protein-surfactant systems provides a fundamental concept for the realization of nanoscale devices with self-organizing capability. In this context, spectroscopic monitoring of pigment-containing proteins yields detailed structural information. Here we have studied the association behavior of the bacterial light-harvesting protein LH2 from Rhodobacter spheroides in an n,n-dimethyldodecylamine-n-oxide/water environment. Time-resolved studies of the excitation annihilation yielded information about aggregate sizes and packing of the protein complexes therein. The results are compared to transmission electron microscopy images of instantaneously frozen samples. Our data indicate the manifestation of different phases, which are discussed with respect to the thermodynamic equilibrium in ternary protein-surfactant-water systems. Accordingly, by varying the concentration the formation of different types of aggregates can be controlled. Conditions for the appearance of isolated LH2 complexes are defined.

Spectroscopic units in conjugated polymers: a quantum chemically founded concept?

In conjugated polymers the concept of spectroscopic units belonging to different spatial segments of the chain, which are responsible for the spectroscopic properties of the polymer, has been used to explain the spectral heterogeneity and the excitation migration by (Förster type) hopping transfer. In the present work we study the possible mechanism of segmentation of polythiophene into spectroscopic units by using quantum-chemical methods (ZINDO). We found that static geometric defects such as kinks or torsions do not result in a significant localization of the excited states to a certain segment. Hence, we propose that a dynamic localization of excitation due to the interaction between the nuclear and electronic degrees of freedom is responsible for the formation of the spectroscopic units.

Excitonic coupling of chlorophylls in the plant light-harvesting complex LHC-II.

Manifestation and extent of excitonic interactions in the red Chl-absorption region (Q(y) band) of trimeric LHC-II were investigated using two complementary nonlinear laser-spectroscopic techniques. Nonlinear absorption of 120-fs pulses indicates an increased absorption cross section in the red wing of the Q(y) band as compared to monomeric Chl a in organic solution. Additionally, the dependence of a nonlinear polarization response on the pump-field intensity was investigated. This approach reveals that one emitting spectral form, characterized by a 2.3(+/-0.8)-fold larger dipole strength than monomeric Chl a, dominates the fluorescence spectrum of LHC-II. Considering available structural and spectroscopic data, these results can be consistently explained assuming the existence of an excitonically coupled dimer located at Chl-bindings sites a2 and b2 (referring to the original notation of W. Nühlbrandt, D.N. Wang, and Y. Fujiyoshi, Nature, 1994, 367:614-621), which must not necessarily correspond to Chls a and b). This fluorescent dimer, terminating the excitation energy-transfer chain of the LHC-II monomeric subunit, is discussed with respect to its relevance for intra- and inter-antenna excitation energy transfer.