New Approaches to Stretched Film Sample Alignment and Data Collection for Vibrational Linear Dichroism.

Rapid measurements of vibrational linear dichroism (VLD) infrared spectra are shown to be possible by using stretched polymer films and an extension of existing instrumentation designed for vibrational circular dichroism spectroscopy. Earlier techniques can be extended using additional inexpensive polymer substrates to record good-quality VLD spectra of a significantly wider range of compounds with comparatively short sample-preparation times. The polymer substrates used, polyethylene and polytetrafluoroethylene, are commonly available and inexpensive, and samples are more easily prepared than that for many earlier stretched-film and crystal studies. Data are presented for neutral hydrophobic organic molecules on hydrophobic films including acridine, anthracene, fluorene, and recently synthesized S-(4-((4-cyanophenyl)ethynyl)phenyl)ethanethioate. We extend the approach to polar or ionic species, including 2,2'-bipyridine, 1,10-phenanthroline, and sodium dodecyl sulfate, by oxidizing polyethylene films to change their wetting properties. The combination of new instrumentation and modified sample preparation methods is useful in basic spectroscopy for untangling and assigning complicated infrared spectra. Nevertheless, it is not a panacea as surface-adsorbed molecules are often not monodispersed, and higher analyte concentrations can lead to aggregation and resonance phenomena that have previously been observed for infrared spectra on surfaces. These effects can be assessed by varying the sample concentration. The focus of this paper is experimental, and detailed analysis of most of the spectra lies outside its scope, including some well-studied compounds such as acridine and anthracene that allow comparisons with earlier research.

Composting and vermicomposting of sewage sludge at various C/N ratios: Technological feasibility and end-product quality.

Even though sewage sludge (SS) contains a high level of pollutants, it is rich in essential plant nutrients and has the potential to enhance soil fertility. However, the SS must be further treated through pre-composting plus vermicomposting to make it safe for use on food crops. More research and data are needed to determine how different carbon-to-nitrogen ratios (C/N) affect the feasibility and quality of composting vs vermicomposting of SS. Therefore, in this study we comprehensively evaluated the feasibility and end-product quality of compost and vermicompost produced from SS under different C/N ratios. SS was mixed with pelletized wheat straw (PWS) at various proportions to produce C/N ratios of 6:1, 18:1, 28:1, and 38:1, then pre-composted for 14 days followed by vermicomposting using the earthworm Eisenia andrei for 120 days. Agrochemical properties were measured at 0, 30, 60, 90, and 120 days. Results revealed significantly higher levels of agrochemicals in vermicompost compared to compost, including total potassium (37-88%) and magnesium (4.3-12%), nitrate nitrogen (71-98%), available potassium (53-88%), available phosphorus (79%), available magnesium (54-453%), available boron (48-303%), and available copper (2.5-82%). However, lower levels of ammonium nitrogen by (59-85%), available iron (2.3-51.3%), available manganese (29.7-52.2%), available zinc (10.5-29.8%), total carbon (0.75-4.5%), and total nitrogen (1.6-22.2%) were measured. Comparison of the various C/N ratios, showed that vermicompost with an 18:1 C/N ratio outperformed compost and demonstrated the highest earthworm population (165 pieces/kg). Thus, vermicomposting SS at an 18:1 C/N ratio is strongly recommended as a sustainable technology for producing high-quality vermicompost from SS.

Concentration of the main persistent organic pollutants in sewage sludge in relation to wastewater treatment plant parameters and sludge stabilisation.

Concentration of 16 polycyclic aromatic hydrocarbons (PAHs), 7 polychlorinated biphenyls (PCBs), and 11 organochlorine pesticides (OCPs) in sewage sludge from 40 wastewater treatment plants (WWTPs) was investigated. Relationship between pollutant sludge contents, main WWTP parameters and type of sludge stabilisation was carefully evaluated. Average load of PAHs, PCBs, and OCPs in different sludges from Czech Republic was 3096, 95.7 and 76.1 µg/kg dry weight, respectively. There were moderate/strong correlations among the individual tested pollutants in sludge (r = 0.40-0.76). Relationship between total pollutant contents in sludge, common WWTP parameters and sludge stabilisation was not evident. Only individual pollutants such anthracene and PCB 52 correlated significantly (P < 0.05) with biochemical oxygen demand (r = -0.35) and chemical oxygen demand removal efficiencies (r = -0.35), suggesting recalcitrance to degradation during wastewater treatment. When sorted according to the design capacity, a linear correlation between WWTP size and pollutant contents in sludge was evident with growing WWTP capacity. Our study indicated that WWTPs with anaerobic digestion are prone to accumulate a statistically higher content of PAHs and PCBs (P < 0.05) in digested sludges compared to aerobically digested ones. The influence of anaerobic digestion temperature of treated sludge on tested pollutants was not evident.

Conformational analysis of amphetamine and methamphetamine: a comprehensive approach by vibrational and chiroptical spectroscopy.

After cannabis, the most commonly used illicit substance worldwide is amphetamine and its derivatives, such as methamphetamine, with an ever-increasing number of synthetic modifications. Thus, fast and reliable methods are needed to identify them according to their spectral patterns and structures. Here, we have investigated the use of molecular spectroscopy methods to describe the 3D structures of these substances in a solution that models the physiological environment. The substances were analyzed by Raman and infrared (IR) absorption spectroscopy and by chiroptical methods, vibrational circular dichroism (VCD) and Raman optical activity (ROA). The obtained experimental data were supported by three different computational approaches based on density functional theory (DFT) and molecular dynamics (MD). Successful interpretation relies on good agreement between experimental and predicted spectra. The determination of the conformer populations of the studied molecules was based on maximizing the similarity overlap of weighted conformer spectra by a global minimization algorithm. Very good agreement was obtained between the experimental spectra and optimized-population weighted spectra from MD, providing a detailed insight into the structure of the molecules and their interaction with the solvent. The relative population of three amphetamine and six methamphetamine conformers was determined and is consistent with a previous NMR study. However, this work shows that only a few isolated conformers are not sufficient for the successful interpretation of the spectra, but the entire conformational space needs to be sampled appropriately and explicit interaction with the solvent needs to be included.

Nutrient recovery and changes in enzyme activity during vermicomposting of hydrolysed chicken feather residue.

Chicken feathers are hazardous to the environment because of their poor digestibility and potential as a source of environmental contaminants. However, this waste contains valuable plant nutrients that can be recovered and used to improve soil fertility and agricultural productivity. The objectives of this study were to evaluate how effective vermicomposting is at recovering nutrients and changes in enzymatic activity during vermicomposting of hydrolysed chicken feather residues (HCFR). The study included four treatments with three replications at different HCFR and pelletized wheat straw (PWS) mixing proportions: (T1) 25% HCFR+75% PWS with earthworms, (T2) 25% HCFR+75% PWS without earthworms, (T3) 50% HCFR+50% PWS with earthworms, and (T4) 50% HCFR+50% PWS (w/w) without earthworms. Eisenia andrei was used in the experiment for 120 days. Earthworm treatments recovered more available plant nutrients than non-earthworm treatments by 14% N-NO3- (T1); 50% K (T3); 47% Mg (T3); 75% P (T3); 55% B (T3); 34% Cu (T3); 40% Fe (T1); 46% Mn (T3); 11% Zn (T1). However, N-NH4+ was significantly reduced by -80% (T1). Acid phosphatase, arylsulphatase, alanine aminopeptidase, and leucine aminopeptidase were more active in the treatments with earthworms and positively correlated with P and C: N ratio. Alanine aminopeptidase (3752 µmol AMCA.g-1.h-1) and leucine aminopeptidase (4252 µmol AMCL.g-1.h-1) had higher activities in T3 on day 60 of vermicomposting. As a result, the earthworm treatment recovers more plant nutrients than the non-earthworm treatments, and it can be recommended as a better vermicomposting approach for nutrient recovery from HCFR.

Low-frequency Raman optical activity provides insight into the structure of chiral liquids.

Vibrational frequencies of modes involving intermolecular motions in liquids are relatively small, in the Raman scattering close to the excitation frequency, and the bands may merge into a diverging uninterpretable signal. Raman optical activity (ROA) spectral shapes in this region, however, are structured more and may better reflect the nature of the studied systems. To understand the origin of the signal and its relation to the molecules, ROA spectra of six chiral neat liquids are recorded and analyzed on the basis of molecular dynamics and density functional theory computations. The theory of Raman scattering of liquids is discussed and adapted for modeling based on clusters and periodic boundary conditions. A plain cluster approach is compared to a crystal-like model. The results show that the low-frequency optical activity can be reliably modeled and related to the structure. However, momentary arrangement of molecules leads to large variations of optical activity, and a relatively large number of geometries need to be averaged for accurate simulations. The intermolecular modes are intertwined with intramolecular ones and start to dominate as the frequency goes down. The low-frequency ROA signal thus reflects the chemical composition and coupled with the modeling it provides a welcome means to study the structure and interactions of chiral liquids.

Structure of Zinc and Nickel Histidine Complexes in Solution Revealed by Molecular Dynamics and Raman Optical Activity.

The histidine residue has an exceptional affinity for metals, but solution structure of its complexes are difficult to study. For zinc and nickel complexes, Raman and Raman optical activity (ROA) spectroscopy methods to investigate the link between spectral shapes and the geometry were used. The spectra were recorded and interpreted on the basis of ionic equilibria, molecular dynamics, ab initio molecular dynamics, and density functional theory. For zwitterionic histidine the dominant tautomer was determined by the decomposition of experimental spectra into calculated subspectra. An octahedral structure was found to prevail for the ZnHis2 complex in solution, in contrast to a tetrahedral arrangement in the crystal phase. The solution geometry of NiHis2 is more similar to the octahedral structure found by X-ray. The Raman and ROA structural determinations of metal complexes are dependent on extensive computations, but reveal unique information about the studied systems.

Occurrence of synthetic polycyclic and nitro musk compounds in sewage sludge from municipal wastewater treatment plants.

Synthetic musk compounds (SMCs) are widely used as fragrances that can be released from different sources and through the sewer system, finally reaching wastewater treatment plants (WWTPs). In this study, 6 synthetic polycyclic and 5 nitro musk compounds were screened in 55 sewage sludge (SS) samples from 43 different WWTPs in the Czech Republic, and the effect of WWTP technology parameters on SMC content in SS was assessed. Galaxolide and Tonalide were predominant synthetic polycyclic musk compounds (SPMCs) detected in all SS tested and accounted for 99.5% of the average content of sludge SMCs (5518 µg/kg dw). The amount of synthetic nitro musk compounds (SNMCs) in SS samples was negligible. The Tonalide content in SS correlated significantly with the WWTP design capacity (r = 0.32, P < 0.05). The significant correlation between chemical oxygen demand (COD) removal efficiency and SMCs (r = -0.37, P < 0.05) partly suggests the recalcitrance of SMCs, mainly of Celestolide, Galaxolide and Tonalide, to biodegradation in WWTPs. A statistically lower SNMC content was found in anaerobically digested sludges than in aerobic ones. There was no significant difference (P > 0.05) between the digestion technology as well as the temperature of anaerobic digestion on the SPMC content in sewage sludge. The wastewater (WW) load percentage or WW hydraulic retention time had no influence on the SMC content in the resulting SS. Musk compounds did not change over time when the SS samples were analysed with a gap of two years, suggesting that sewage sludge for soil applications only needs to be analysed for musk compounds once a year. Our study indicates that the currently common WWTP technologies have only very limited potential to affect the accumulation of musk compounds in sewage sludge.

Chiral detection by induced surface-enhanced Raman optical activity.

Combination of optical activity with surface-enhanced Raman scattering has been a dream of physical chemists for a long time. We report a measurement protocol based on silver colloids and aromatic linkers where chiral acids could be detected in concentrations of about 10-5 M. We explain the mechanism by binding and self-assembly of the linkers into chiral aggregates on the silver surface. Following the "sergeants-and-soldiers" principle, the chirality is determined by the relatively minor acidic component. Such detection of biologically relevant molecules may be useful when other methods, such as electronic circular dichroism, are not sensitive enough. In the future, variations of the chemical structure of the linker or other conditions are needed to provide a more specific signal allowing one to better discriminate among the optically active molecules.

Thermal thickening of nitrified liquid phase of digestate for production of concentrated complex fertiliser and high-quality technological water.

Large production of liquid phase of digestate (LPD) characterised by low dry matter content increases the costs of LPD storage and transportation. Additionally, significant nitrogen losses result from LPD handling. The operation of biogas plants also requires large amounts of technological water, a scarce resource. The method of LPD nitrification and subsequent thermal thickening as a promising alternative treatment able to overcome those problems was investigated. The properties of thermally thickened nitrified LPD and of the distillate derived from vacuum evaporation of nitrified LPD were examined. The results show that over 99.9% of nitrogen was accumulated in thickened nitrified LPD. Total nitrogen (TN) concentration in thickened LPD ranged from 12 to 18 g/L depending on the distillate/thickened LPD volume ratio applied. The thickened LPD was also richer in other nutrients, which indicates its possible use as a concentrated complex fertiliser. TN concentration and conductivity in the distillate did not exceed 6 mg/L and 0.03 mS/m respectively, suggesting that the distillate may be useful as alternative source of technological water. It was concluded that the system consisting of nitrification and subsequent thermal thickening of nitrified LPD yields final products exhibiting significantly superior properties compared to thermal thickening of raw LPD, without requiring any additional chemical agents. The quality of these products is comparable to those obtained through advanced membrane processes.

Raman Optical Activity of Glucose and Sorbose in Extended Wavenumber Range.

Raman optical activity (ROA) is pursued as a promising method for structural analyses of sugars in aqueous solutions. In the present study, experimental Raman and ROA spectra of glucose and sorbose obtained in an extended range (50-4000 cm-1 ) are interpreted using molecular dynamics and density functional theory, with the emphasis on CH stretching modes. A reasonable theoretical basis for spectral interpretation was obtained already at the harmonic level. Anharmonic corrections led to minor shifts of band positions (up to 25 cm-1 ) below 2000 cm-1 , while the CH stretching bands shifted more, by ∼180 cm-1 , and better reproduced the experiment. However, the anharmonicities could be included on a relatively low approximation level only, and they did not always improve the harmonic band shapes. The dependence on the structure and conformation shows that the CH stretching ROA spectral pattern is a sensitive marker useful in saccharide structure studies.

Vibrational Optical Activity of Intermolecular, Overtone, and Combination Bands: 2-Chloropropionitrile and α-Pinene.

Spectroscopy of vibrational optical activity has been established as a powerful tool to study molecular structures and interactions. In most cases, only fundamental molecular transitions are analyzed. In the present study, we analyze a broader range of vibrational frequencies (40-4000 cm-1), which could be measured on a new Raman optical activity (ROA) instrument. An unexpectedly strong vibrational Raman optical activity of 2-chloropropionitrile has been observed within the low-frequency region (40-150 cm-1). On the basis of combined molecular dynamics and density functional theory simulations, it could be assigned to intermolecular vibrations. A detailed analysis also revealed connection between spectral shapes and molecular structure and flexibility, such as bending of the CCN group. At the other edge of the scale, within ∼1500-4000 cm-1, for the first time, many combination and overtone ROA bands have been observed for 2-chloropropionitrile and α-pinene. These were also partially assigned, using quantum-chemical computations. The band assignment was confirmed by a comparison with Raman, absorption, and vibrational circular dichroism spectra. The measurement in the broader vibrational range thus significantly extends the information that can be obtained by optical spectroscopy, including intermolecular interactions of chiral molecules and liquids.

Nitrification in a completely stirred tank reactor treating the liquid phase of digestate: The way towards rational use of nitrogen.

The nitrification of the liquid phase of digestate (LPD) was conducted using a 5L completely stirred tank reactor (CSTR) in two independent periods (P1 - without pH control; P2 - with pH control). The possibility of minimizing nitrogen losses during the application of LPD to the soil as well as during long-term storage or thermal thickening of LPD using nitrification was discussed. Moreover, the feasibility of applying the nitrification of LPD to the production of electron acceptors for biological desulfurization of biogas was assessed. Despite an extremely high average concentration of ammonia and COD in LPD reaching 2470 and 9080mg/L, respectively, nitrification was confirmed immediately after the start-up of the CSTR. N-NO3- concentration reached 250mg/L only two days after the start of P1. On the other hand, P1 demonstrated that working without pH control is a risk because of the free nitrous acid (FNA) inhibition towards nitrite oxidizing bacteria (NOB) resulting in massive nitrite accumulation. Up to 30.9mg/L of FNA was present in the reactor during P1, where the NOB started to be inhibited even at 0.15mg/L of FNA. During P2, the control of pH at 7.0 resulted in nitrogen oxidation efficiency reaching 98.3±1.5% and the presence of N-NO3- among oxidized nitrogen 99.6±0.4%. The representation of volatile free ammonia within total nitrogen was reduced more than 1000 times comparing with raw LPD under these conditions. Thus, optimum characteristics of the tested system from the point of view of minimizing the nitrogen losses as well as production of electron acceptors for the desulfurization of biogas were gained in this phase of reactor operation. Based on the results of the experiments, potential improvements and modifications of the tested system were suggested.

Anaerobic digestion of grass: the effect of temperature applied during the storage of substrate on the methane production.

Within this research, biogas production, representation of methane in biogas and volatile solids (VSs) removal efficiency were compared using batch tests performed with the samples of intensively and extensively planted grasses originating from public areas. Before the batch tests, the samples were stored at different temperatures achievable on biogas plants applying trigeneration strategy (-18°C, +3°C, +18°C and +35°C). Specific methane production from intensively planted grasses was relatively high (0.33-0.41 m3/kg VS) compared to extensively planted grasses (0.20-0.33 m3/kg VS). VSs removal efficiency reached 59.8-68.8% for intensively planted grasses and 34.6-56.5% for extensively planted grasses. Freezing the intensively planted grasses at -18°C proved to be an effective thermal pretreatment leading to high biogas production (0.61 m3/kg total solid (TS)), high representation of methane (64.0%) in biogas and good VSs removal efficiency (68.8%). The results of this research suggest that public areas or sport parks seem to be available, cheap and at the same time very effective feedstock for biogas production.

Changes in Membrane Cholesterol Differentially Influence Preferential and Non-preferential Signaling of the M1 and M3 Muscarinic Acetylcholine Receptors.

We have found earlier that changes in membrane cholesterol content have distinct impact on signaling via the M1, M2, or M3 receptors expressed in CHO cells (CHO-M1 through CHO-M3). Now we investigated whether gradual changes in membrane cholesterol exerts differential effects on coupling of the M1 and M3 muscarinic receptors to preferential signaling pathways through Gq/11 and non-preferential Gs G-proteins signaling. Changes in membrane cholesterol resulted in only marginal alterations of antagonist and agonist affinity of the M1 and M3 receptors, and did not influence precoupling of either subtype. Changes in membrane cholesterol did not influence parameters of carbachol-stimulated GTP-γ(35)S binding in CHO-M1 membranes while reduction as well as augmentation of membrane cholesterol lowered the efficacy but increased the potency of carbachol in CHO-M3 membranes. Gradual increase or decrease in membrane cholesterol concentration dependently attenuated agonist-induced inositolphosphates release while only cholesterol depletion increased basal values in both cell lines. Similarly, membrane cholesterol manipulation modified basal and agonist-stimulated cAMP synthesis via Gs in the same way in both cell lines. These results demonstrate that changes in membrane cholesterol concentration differentially impact preferential and non-preferential M1 and M3 receptor signaling. They point to the activated G-protein/effector protein interaction as the main site of action in alterations of M1 receptor-mediated stimulation of second messenger pathways. On the other hand, modifications in agonist-stimulated GTP-γ(35)S binding in CHO-M3 membranes indicate that in this case changes in ligand-activated receptor/G-protein interaction may also play a role.

Membrane cholesterol content influences binding properties of muscarinic M2 receptors and differentially impacts activation of second messenger pathways.

We investigated the influence of membrane cholesterol content on preferential and non-preferential signaling through the M(2) muscarinic acetylcholine receptor expressed in CHO cells. Cholesterol depletion by 39% significantly decreased the affinity of M(2) receptors for [(3)H]-N-methylscopolamine ([(3)H]-NMS) binding and increased B(max) in intact cells and membranes. Membranes displayed two-affinity agonist binding sites for carbachol and cholesterol depletion doubled the fraction of high-affinity binding sites. In intact cells it also reduced the rate of agonist-induced receptor internalization and changed the profile of agonist binding from a single site to two affinity states. Cholesterol enrichment by 137% had no effects on carbachol E(max) of cAMP synthesis inhibition and on cAMP synthesis stimulation and inositolphosphates (IP) accumulation at higher agonist concentrations (non-preferred pathways). On the other hand, cholesterol depletion significantly increased E(max) of cAMP synthesis inhibition or stimulation without change in potency, and decreased E(max) of IP accumulation. Noteworthy, modifications of membrane cholesterol had no effect on membrane permeability, oxidative activity, protein content, or relative expression of G(s), G(i/o), and G(q/11) alpha subunits. These results demonstrate distinct changes of M(2) receptor signaling through both preferential and non-preferential G-proteins consequent to membrane cholesterol depletion that occur at the level of receptor/G-protein/effector protein interactions in the cell membrane. The significant decrease of IP accumulation by cholesterol depletion was also observed in cells expressing M(3) receptors and by both cholesterol depletion and enrichment in cells expressing M(1) receptors indicating relevance of reduced G(q/11) signaling for the pathogenesis of Alzheimer's disease.

Modelling the consequences of receptor-G-protein promiscuity.

Many G-protein-coupled receptors interact with more than one type of G protein, giving rise to extreme variability in the effects of receptor activation, depending on, for example, receptor density and desensitization, efficacy of agonists, and availability of specific G proteins. This leads to errors in interpretation of data. To facilitate understanding the consequences of receptor-G-protein promiscuity, we use two simplified models to simulate such consequences. Applied to the regulation of adenylyl cyclase and phosphoinositidase, the models predict seemingly paradoxical situations and explain some phenomena that, at first sight, might seem to require the induction of agonist-specific (G-protein-selective) receptor conformations.