Intramolecular OH Research Articles

Characterization of the Intermediate in and Identification of the Repair Mechanism of (6-4) Photolesions by Photolyases.

Quantum mechanics/molecular mechanics calculations are employed to assign previously recorded experimental spectroscopic signatures of the intermediates occurring during the photo-induced repair of (6-4) photolesions by photolyases to specific molecular structures. Based on this close comparison of experiment and theory it is demonstrated that the acting repair mechanism involves proton transfer from the protonated His365 to the N3' nitrogen of the lesion, which proceeds simultaneously with intramolecular OH transfer along an oxetane-like transition state.

Characterization of the Intermediate in and Identification of the Repair Mechanism of (6‐4) Photolesions by Photolyases

AbstractQuantum mechanics/molecular mechanics calculations are employed to assign previously recorded experimental spectroscopic signatures of the intermediates occurring during the photo‐induced repair of (6‐4) photolesions by photolyases to specific molecular structures. Based on this close comparison of experiment and theory it is demonstrated that the acting repair mechanism involves proton transfer from the protonated His365 to the N3′ nitrogen of the lesion, which proceeds simultaneously with intramolecular OH transfer along an oxetane‐like transition state.

Stable Blue Phosphorescence Iridium(III) Cyclometalated Complexes Prompted by Intramolecular Hydrogen Bond in Ancillary Ligand.

Improvement of the stability of blue phosphorescent dopant material is one of the key factors for real application of organic light-emitting diodes (OLEDs). In this study, we found that the intramolecular hydrogen bonding in an ancillary ligand from a heteroleptic Ir(III) complex can play an important role in the stability of blue phosphorescence. To rationalize the role of intramolecular hydrogen bonding, a series of Ir(III) complexes is designed and prepared: Ir(dfppy)2(pic-OH) (1a), Ir(dfppy)2(pic-OMe) (1b), Ir(ppy)2(pic-OH) (2a), and Ir(ppy)2(pic-OMe) (2b). The emission lifetime of Ir(dfppy)2(pic-OH) (1a) (τem = 3.19 μs) in dichloromethane solution was found to be significantly longer than that of Ir(dfppy)2(pic-OMe) (1b) (τem = 0.94 μs), because of a substantial difference in the nonradiative decay rate (knr = 0.28 × 10(5) s(-1) for (1a) vs 2.99 × 10(5) s(-1) for (1b)). These results were attributed to the intramolecular OH···O═C hydrogen bond of the 3-hydroxy-picolinato ligand. Finally, device lifetime was significantly improved when 1a was used as the dopant compared to FIrpic, a well-known blue dopant. Device III (1a as dopant) achieved an operational lifetime of 34.3 h for an initial luminance of 400 nits compared to that of device IV (FIrpic as dopant), a value of 20.1 h, indicating that the intramolecular hydrogen bond in ancillary ligand is playing an important role in device stability.

Simulating two-dimensional infrared-Raman and Raman spectroscopies for intermolecular and intramolecular modes of liquid water.

Full classical molecular dynamics (MD) simulations of two-dimensional (2D) infrared-Raman and 2D Raman spectroscopies of liquid water were carried out to elucidate a mode-mode coupling mechanism using a polarizable water model for intermolecular and intramolecular vibrational spectroscopy (POLI2VS). This model is capable of describing both infrared and Raman spectra. Second-order response functions, which consist of one molecular polarizability and two molecular dipole moments for 2D IR-Raman and three molecular polarizabilities for 2D Raman spectroscopies, were calculated using an equilibrium-non-equilibrium hybrid MD approach. The obtained signals were analyzed using a multi-mode Brownian oscillator (BO) model with nonlinear system-bath interactions representing the intramolecular OH stretching, intramolecular HOH bending, hydrogen bonded (HB)-intermolecular librational motion and HB-intermolecular vibrational (translational) motion of liquid water. This model was applied through use of hierarchal Fokker-Planck equations. The qualitative features of the peak profiles in the 2D spectra obtained from the MD simulations are accurately reproduced with the BO model. This indicates that this model captures the essential features of the intermolecular and intramolecular motion. We elucidate the mechanisms governing the 2D signal profiles involving anharmonic mode-mode coupling, the nonlinearities of the polarizability and dipole moment, and the vibrational dephasing processes of liquid water even in the case that the 2D spectral peaks obtained from the MD simulation overlap or are unclear. The mode coupling peaks caused by electrical anharmonic coupling (EAHC) and mechanical anharmonic coupling (MAHC) are observed in all of the 2D spectra. We find that the strength of the MAHC between the OH-stretching and HB-intermolecular vibrational modes is comparable to that between the OH-stretching and HOH bending modes. Moreover, we find that this OH-stretching and HB-intermolecular vibrational coupling should be observed as off-diagonal cross peaks in the 2D spectra.

Influence of Fluorination on the Conformational Properties and Hydrogen‐Bond Acidity of Benzyl Alcohol Derivatives

The effect of fluorination on the conformational and hydrogen-bond (HB)-donating properties of a series of benzyl alcohols has been investigated experimentally by IR spectroscopy and theoretically with quantum chemical methods (ab initio (MP2) and DFT (MPWB1K)). It was found that o-fluorination generally resulted in an increase in the HB acidity of the hydroxyl group, whereas a decrease was observed upon o,o′-difluorination. Computational analysis showed that the conformational landscapes of the title compounds are strongly influenced by the presence of o-fluorine atoms. Intramolecular interaction descriptors based on AIM, NCI and NBO analyses reveal that, in addition to an intramolecular OH⋅⋅⋅F interaction, secondary CH⋅⋅⋅F and/or CH⋅⋅⋅O interactions also occur, contributing to the stabilisation of the various conformations, and influencing the overall HB properties of the alcohol group. The benzyl alcohol HB-donating capacity trends are properly described by an electrostatic potential based descriptor calculated at the MPWB1K/6-31+G(d,p) level of theory, provided solvation effects are taken into account for these flexible HB donors.

Ultrafast Dynamics of Liquid Water: Energy Relaxation and Transfer Processes of the OH Stretch and the HOH Bend.

The vibrational energy relaxation and transfer processes of the OH stretching and HOH bending vibrations in liquid water are investigated via the theoretical calculation of the pump-probe spectra obtained from nonequilibrium molecular dynamics simulations with the TTM3-F interaction potential. The excitation of the OH stretch induces an instantaneous response of the high frequency librational motions in the 600-1000 cm(-1) range. In addition, the excess energy of the OH stretch of a water molecule quickly transfers to the OH stretches of molecules in its first hydration shell with a time constant of ∼50 fs, followed by relaxation to the HOH bends of the surrounding molecules with a time constant of 230 fs. The excitation of the HOH bend also results in the ultrafast excitation of the high frequency librational motions. The energy of the excited HOH bend of a water molecule decays, with a time constant of 200 fs, mainly to the relaxation of the HOH bends of its surrounding molecules. The energies of the HOH bends were found to transfer quickly to the intermolecular motions via the coupling with the high frequency librational motions. The excess energy of the OH stretch or the HOH bend relaxes to the high frequency intermolecular librational motions and eventually to the hot ground state with a time scale of ∼1 ps via the coupling with the librational and translational motions. The energy relaxation and transfer processes were found to depend on the local hydrogen bonding network; the relaxations of the excess energy of the OH stretch and the HOH bend of four- and five-coordinated molecules are faster than those of a three-coordinated molecule due to the delocalization of the vibrational motions of the former (four- and five-coordinated molecules) compared to those of the later (three-coordinated molecules). The present results highlight the importance of the high frequency intermolecular librational modes in facilitating the ultrafast energy relaxation process in liquid water via their strong nonlinear couplings with the intramolecular OH stretching and HOH bending vibrations.

Synthesis of a Tight Intramolecular OH···Olefin Interaction, Probed by IR, (1)H NMR, and Quantum Chemistry.

We have synthesized a molecule containing a tight hydrogen-bonding interaction between an alcohol and a nonconjugated π-system. The strength of this hydrogen bond results in a large red shift, nearly 189 cm(-1), on the alcohol stretching frequency in the IR spectrum in comparison to a free alcohol control. The interaction is notable in that it possesses a better defined intramolecular hydrogen bond compared to the usual molecules for which it is noted, such as syn-7-norbornenol. This interaction was studied through the use of IR and NMR spectroscopy, X-ray crystallography, and molecular modeling calculations.

Spatial arrangement and acid strength effects on acid–base cooperatively catalyzed aldol condensation on aminosilica materials

Weak acids are known to enhance the activity of amines in aldol condensation reactions on silica-based catalysts. The effects of acid strength and arrangement of the promoting site with respect to a secondary amine have been investigated in the aldol condensation of 4-nitrobenzaldehyde with acetone. Changing the substituent of this secondary amine from a methyl to an ethyl group decreases the activity. An intramolecular OH function provided by a primary alcohol incorporated on the β-carbon of the amine substituent exhibits a similar cooperativity as an intermolecular OH function provided by neighboring surface silanols. A maximum activity was achieved when the secondary amine with the same primary alcohol-containing substituent was surrounded by surface silanols, indicating the potential advantage of simultaneously activating both reactants by the formation of a hydrogen bond in contrast to the consecutive activation when there is only one promoting site in the vicinity of the amine. Changing the alcohol to stronger acids resulted in a reduced cooperativity with increasing acid strength. After removing the silanols from the surface, the activity of the catalysts which exhibit an intramolecular cooperativity retained about 68–83% of their activities while the activity of the conventional secondary amine was reduced by a factor of four compared to its intermolecularly cooperative counterpart.

Biscoumarin derivatives: Synthesis, crystal structure, theoretical studies and induced apoptosis activity on bladder urothelial cancer cell

In this study, five new biscoumarin derivatives (1–5) were synthesized and compound 4 inhibited the proliferation of the bladder urothelial cells (J82 cell line) obviously after 48h treatment at different concentration (1, 5 and 10μmol/L), and J82 cells were predominantly induced to apoptotic cell death after compound 4 treatment. Morphologic changes of bladder urothelial cancer cells were also observed under transmission electron microscopy (TEM) after compound 4 treatment. In addition, compound 4 had much less toxicity to human umbilical vein endothelial cells. To explore the possible anti-cancer mechanism of compound 4, two classical intramolecular OH⋯O hydrogen bonds (HBs) in their structures and the corresponding HB energies were performed with the density functional theory (DFT) [B3LYP/6-31G∗] method. Anti-bladder cancer activity of compound 4 is possible due to the intramolecular weakest HB energies.

Synthesis, spectroscopic characterization and in vitro antimicrobial, anticancer and antileishmanial activities as well interaction with Salmon sperm DNA of newly synthesized carboxylic acid derivative, 4-(4-methoxy-2-nitrophenylamino)-4-oxobutanoic acid

This paper stresses on the synthesis, characterization of novel carboxylic acid derivative and its application in pharmaceutics. Carboxylic acid derivatives have a growing importance in medicine, particularly in oncology. A novel carboxylic acid, 4-(4-methoxy-2-nitrophenylamino)-4-oxobutanoic acid, was synthesized and characterized by elemental analysis, FT-IR, NMR (1H, and 13C), mass spectrometry and single crystal X-ray structural analysis. The structure of the title compound, C11H12N2O6, shows the molecules dimerised by short intramolecular OH⋯O hydrogen bonds. The compound was screened for in vitro antimicrobial, anticancer, and antileishmanial activities as well as interaction with SS-DNA. The compound was also checked for in vitro anticancer activity against BHK-21, H-157 and HCEC cell lines, and showed significant anticancer activity. The compound was almost non-toxic towards human corneal epithelial cells (HCEC) and did not show more than 7.4% antiproliferative activity when used at the 2.0μg/mL end concentration. It was also tested for antileishmanial activity against the promastigote form of leishmania major and obtained attractive result. DNA interaction study exposes that the binding mode of the compound with SS-DNA is an intercalative as it results in hypochromism along with minor red shift. A new and efficient strategy to identify pharmacophores sites in carboxylic acid derivative for antibacterial/antifungal activity using Petra, Osiris and Molinspiration (POM) analyses was also carried out.

Unconventional redox properties of hydroquinones with intramolecular OH−N hydrogen bonds

The redox (chemical and electrochemical) properties of several hydroquinones are reported in which the OH protons are engaged in intramolecular hydrogen bonds to a nitrogen-based acceptor (pyridine or amine). The 1,4-hydroquinones generally undergo reversible oxidation to quinones in which both OH protons have transferred to the pendant bases; the oxidation processes are generally chemically and electrochemically reversible, in stark contrast with normal hydroquinones, which are oxidized irreversibly (via proton loss) to quinones. The oxidation processes, believed to occur in concerted proton/electron transfer steps, are at much lower potentials for the hydrogen-bonded derivatives relative to unsubstituted derivatives. In contrast, isomeric 1,3-hydroquinones (resorcinols) are oxidized irreversibly at relatively high potentials. The stability of some of the 1,4-hydroquinone oxidized species permits their isolation and characterization both spectroscopically and structurally. Somewhat surprisingly, in the oxidized species in which the proton is now located on the nitrogen base, the characterization data indicate that there is no NH−O hydrogen bond. Relationships between the particulars of the redox properties of the hydroquinones (potentials, reversibility/stability) and molecular structure are discussed.

Structural, spectroscopic and DFT study of 4-methoxybenzohydrazide Schiff bases. A new series of polyfunctional ligands

Five Schiff bases obtained from condensation of 4-methoxybenzohydrazide with related aldehydes, namely o-vanillin, vanillin, 5-bromovanillin, 5-chlorosalicylaldehyde and 5-bromosalicylaldehyde were prepared. A detailed structural and spectroscopic study is reported. The crystal structures of four members of the family were determined and compared with one another. The hydrazones obtained from 5-chlorosalicylaldehyde and 5-bromosalicylaldehyde resulted to be isomorphic to each other. The solid-state structures are stabilized by intra-molecular OH⋯N interactions in salicylaldehyde derivatives between the OH moiety from the aldehyde and the hydrazone nitrogen atom. All crystals are further stabilized by inter-molecular H-bonds mediated by the crystallization water molecule. A comparative analysis between experimental and theoretical results is presented. The conformational space was searched and geometries were optimized both in gas phase and including solvent effects. The structure is predicted for the compound for which the crystal structure was not determined. Infrared and electronic spectra were measured and assigned with the help of data obtained from computational methods based on the Density Functional Theory.

Characteristics of hydrogen bond revealed from water clusters

The hydrogen bond network is responsible for the exceptional physical and chemical properties of water, however, the description of hydrogen bond remains a challenge for the studies of condensed water. The investigation of structural and binding properties of water clusters provides a key for understanding the H-bonds in bulk water. In this paper, a new set of geometric parameters are defined to describe the extent of the overlap between the bonding orbital of the donor OH and the nonbonding orbital of the lone-pair of the acceptor molecule. This orbital overlap plays a dominant role for the strength of H-bonds. The dependences of the binding energy of the water dimer on these parameters are studied. The results show that these parameters properly describe the H-bond strength. The ring, book, cage and prism isomers of water hexamer form 6, 7, 8 and 9 H-bonds, and the strength of the bonding in these isomers changes markedly. The internally-solvated and the all-surface structures of (H2O)n for n = 17, 19 and 21 are nearly isoenergetic. The internally-solvated isomers form fewer but stronger H-bonds. The hydrogen bonding in the above clusters are investigated in detail. The geometric parameters can well describe the characters of the H-bonds, and they correlate well with the H-bond strength. For the structures forming stronger H-bonds, the H-bond lengths are shorter, the angle parameters are closer to the optimum values, and their rms deviations are smaller. The H-bonds emanating from DDAA and DDA molecules as H-donor are relatively weak. The vibrational spectra of (H2O)n (n = 17, 19 and 21) are studied as well. The stretching vibration of the intramolecular OH bond is sensitive to its bonding environment. The H-bond strength judged from the geometric parameters is in good agreement with the bonding strength judged from the stretching frequencies.

Conformational isomerism of pyridoxal. Infrared matrix isolation and theoretical studies

A combined matrix isolation FTIR and theoretical DFT/B3LYP/6–311++G(2p,2d) study of pyridoxal was performed. The calculations resulted in five stable PLHB conformers stabilized by intramolecular OH⋯O bonding between phenolic OH and carbonyl CO groups and another thirteen conformers in which OH or/and aldehyde groups are rotated by 180° around CO or/and CC bonds leading, respectively, to formation of PLO, PLA and PLOA conformers. The analysis of the spectra of the as-deposited matrix indicated that two most stable PLHB1 and PLHB2 conformers with intramolecular hydrogen bond are present in the matrix. The exposure of the PL/Ar matrix to mercury lamp radiation (λ>345nm) induced conformational change of PLHB isomers to PLOA ones.

Synthesis, spectroscopic characterization, crystal structure, DNA interaction study and invitro biological screenings of 4-(5-chloro-2-hydroxyphenylamino)-4-oxobut-2-enoic acid

The titled compound, 4-(5-chloro-2-hydroxyphenylamino)-4-oxobut-2-enoic acid was synthesized and characterized by various techniques like elemental analyses, FT-IR, NMR (1H, and 13C) and single crystal X-ray structural analysis. The appearance of the OH peak of the carboxylic acid in the FT-IR and NMR spectra conform the formation of the compound. A good agreement was found between the calculated values of C, H, N and found values in elemental analysis that show the purity of the compound. Protons H2 and H3 are in cis conformation with each other as conformed both from 1H NMR as well as from single crystal X-ray analysis. The molecular structure of the title compound, C10H10NO3Cl, is stabilized by short intramolecular OH---O hydrogen bonds within the molecule. In the crystal structure, intermolecular NH---O hydrogen bonds link molecules into zigzag chains resulting in a dendrimer like structure. The title compound was screened for biological activities like interaction with DNA, cytotoxicity, antitumor and antioxidant activities. DNA interaction study reveals that the binding mode of interaction of the compound with SS-DNA is intercalative as it results in hypochromism along with significant red shift of 5nm. It was also found to be effective antioxidant of 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) and show almost comparable antioxidant activity to that of the standard and known antioxidant, ascorbic acid, at higher concentration. The antitumor activity data of the compound shows that it can be used as potent antitumor agent.

A study on the influence of intramolecular O[sbnd]H⋯X (X = O and S) hydrogen bond formation on the aromaticity of heptafulvene derivatives, in which the methylene is replaced by [formula omitted][formula omitted], NH and O groups

The influence of the intramolecular OH⋯O and OH⋯S hydrogen bond on the aromaticity of some heptafulvene derivatives, in which the methylene group is replaced successively by AlH2-, BH2-, NH and O groups, is investigated using magnetic (NICS(0), NICS(1) and NICS(1)zz), geometrical (HOMA) and electronic (FLU and SA) criteria to consider the multidimensional character of aromaticity. It is found that the formation of intramolecular hydrogen bond changes aromatic character of heptafulvene derivatives. The variations of NICS indices due to the formation of intramolecular H-bonding significantly correlate with the properties of the considered hydrogen bonds, while the other indices do not give any meaningful results. It seems that the hydrogen bond influences the magnetic dimension of aromaticity in heptafulvenes. The influence of the OH⋯O and OH⋯S hydrogen bond on the aromaticity of these rings are also compared with each other.

Experimental (X-ray, FT-IR and UV–vis spectra) and theoretical methods (DFT study) of (E)-3-methoxy-2-[(p-tolylimino)methyl]phenol

A suitable single crystal of (E)-3-methoxy-2-[(p-tolylimino)methyl]phenol, formulated as C15H15N1O2, reveals that the structure is adopted to its E configuration about the azomethine CN double bond. The compound adopts a enol–imine tautomeric form with a strong intramolecular OH⋯N hydrogen bond. The single crystal X-ray diffraction analysis at 296K crystallizes in the monoclinic space group P21/c with a=13.4791(11) Å, b=6.8251(3) Å, c=18.3561(15) Å, α=90°, β=129.296(5)°, γ=90° and Z=4. Comprehensive theoretical and experimental structural studies on the molecule have been carried out by FT-IR and UV–vis spectrometry.Optimized molecular structure and harmonic vibrational frequencies have been investigated by DFT/B3LYP method with 6-31G(d,p) basis set. Stability of the molecule, hyperconjugative interactions, charge delocalization and intramolecular hydrogen bond has been analyzed by using natural bond orbital (NBO) analysis.Electronic structures were discussed by TD-DFT method and the relocation of the electron density were determined. The energetic behavior of the title compound has been examined in solvent media using polarizable continuum model (PCM). Molecular electrostatic potential (MEP), Mulliken population method and natural population analysis (NPA) have been studied. Nonlinear optical (NLO) properties were also investigated. In addition, frontier molecular orbitals analysis have been performed from the optimized geometry. An ionization potential (I), electron affinity (A), electrophilicity index (ω), chemical potential (μ), electronegativity (χ), hardness (η), and softness (S), have been investigated.

Dynamic Properties of Molecular Tweezers with a Bis(2‐hydroxyphenyl)pyrimidine Backbone

4,6-Bis(2-hydroxyphenyl)-2-alkylpyrimidines with two anthryl or 9-ethylnylanthryl substituents at the positions para to the OH groups prefer a U-shaped conformation supported by two intramolecular OH⋅⋅⋅N hydrogen bonds in the solid state and in CDCl3 solution. The compound with a hexyl substituent on the pyrimidine group and two 9-ethynylanthryl arms at the hydroxyphenyl groups forms a 1:1 complex with 2,4,7-trinitrofluorenone. Its association constant K(a) was estimated to be 2100 M(-1) at 298 K, which is larger than those of other molecular tweezers (K(a) < 1000 M(-1)). DFT calculations suggested that the complex adopts a stable conformation supported by intramolecular hydrogen bonds among the OH groups and the pyrimidine ring as well as by intermolecular π-π interaction between the anthryl groups and 2,4,7-trinitrofluorenone. Addition of nBu4NF to a solution of the molecular tweezers or their complexes causes the cleavage of one or two OH⋅⋅⋅N hydrogen bonds, formation of new O⋅⋅⋅HF hydrogen bonds, and changes in the molecular conformation. The resulting structure of the molecular tweezers contains nonparallel anthryl groups, which do not bind the guest molecule. Photochemical measurements on 4,6-bis(2-hydroxyphenyl)-2-methylpyrimidine with two anthryl substituents showed negligible luminescence (quantum yield ϕ<0.01), owing to photoinduced electron transfer of the molecule with a U-shaped structure. However, the O-hexylated compound exhibits emission from the anthryl groups with ϕ=0.39.

Crystal structure and spectroscopic characterization of (E)-2-(((4-bromo-2-(trifluoromethoxy)phenyl)imino)methyl)-4-nitrophenol: A combined experimental and computational study

A novel compound crystallizes in the triclinic space group P−1 with a=7.674(4) Å, b=12.584(6) Å, c=15.921(6) Å, α=89.62(4)°, β=84.34(4)°, γ=73.77(4)° and Z=4. This compound contains Schiff base and rings of molecule has (E) configuration with respect to the central CN double bond. The crystal structure has the intramolecular OH⋯N and the intermolecular CH⋯O hydrogen bonds. Molecular modeling of the title compound was done by using density functional theories (DFT). Detailed vibrational assignments have been made on the basis of potential energy distribution (PED). Additionally, chemical shift assignments, investigations of thermodynamical parameters and plotting of molecular electrostatic potential surfaces have been performed with the help of DFT method. In order to understand the electronic transitions of the title compound, time dependent DFT (TD-DFT) calculations were performed in gas phase. The dipole moment, linear polarizabilities, anisotropy and first hyperpolarizabilities values have been also computed using the same method.

Growth, molecular structure, NBO analysis and vibrational spectral analysis of l-tartaric acid single crystal

Single crystal of l-tartaric acid (LTA) has been grown by slow evaporation technique. The experimental and theoretical studies on molecular structure, vibrational spectra, electronic absorption spectra and non-linear optical property of the crystal are studied. The FT-IR, FT-Raman and UV–Vis–NIR experimental spectra of LTA crystal have been recorded in the range 400–4000cm−1, 100–3700cm−1 and 190–1100nm, respectively. Density functional theory calculations with B3LYP/6-311++G(d,p) basis sets was used to determine ground state molecular geometries, vibrational frequencies, ICT interactions, Mulliken population analysis on atomic charge, HOMO–LUMO analysis, non-linear optical response properties and thermodynamic properties for LTA and the results were discussed. Vibrational analysis confirms the formation of intramolecular OH⋯O hydrogen bonding. The stability of the molecule has been analyzed using NBO analysis. The results of electronic absorptions in gas phase and water phase LTA were calculated using TD-DFT method. The third-order nonlinear absorption behaviour of LTA was studied using open aperture Z-scan technique, with 5ns laser pulses at 532nm and the nonlinear absorption coefficient of the grown crystal was measured. The predicted NLO properties, UV absorption and Z-scan studies indicate that LTA is an attractive material for laser frequency doubling and optical limiting applications.