The liquid structure of acetic acid was studied experimentally and theoretically. Experimentally, the Raman spectra of acetic acid at various temperatures between 287 and 348 K were measured in the region 15−3700 cm-1. Theoretically, ab initio molecular orbital calculations were performed on the Raman activities of seven cluster species of acetic acid molecules. The Raman spectrum (in R(ν̄) representation) of crystalline acetic acid at 287 K shows six distinct bands in the 15−300 cm-1 region. These bands broaden on the melting of the crystal, whereas their peak positions remain almost unchanged on melting. These spectral changes are reproduced in the case where the liquid spectrum mainly arises from a variety of sizes of chain clusters as the fragments of the crystalline networks. The CO stretching band becomes broadened toward higher wavenumbers and exhibits an asymmetric shape with increasing temperature. The wavenumbers calculated for the CO stretching vibrations suggest that the strongly hydrogen-bonded CO groups of the chain clusters show the prominent CO band and its asymmetric shape is due to the presence of weakly hydrogen-bonded CO groups of the same cluster species. The spectral analyses in both the low wavenumber and the CO stretching regions suggest that liquid acetic acid is mainly composed of the chain clusters, not the cyclic dimer. Assignments of the low-frequency Raman bands observed in the vapor and crystalline states are discussed on the basis of the calculated wavenumbers.

We have studied physiological parameters in a living cell using fluorescence lifetime imaging of endogenous chromophores. In this study, pH dependence of the fluorescence lifetime of flavin adenine dinucleotide (FAD), that is a significant cofactor exhibiting autofluorescence, has been investigated in buffer solution and in cells. The fluorescence lifetime of FAD remained unchanged with pH 5 to 9 in solution. However, the fluorescence lifetime in HeLa cells was found to decrease with increasing intracellular pH, suggesting that pH in a single cell can be estimated from the fluorescence lifetime imaging of FAD without adding exogenous fluorescent probes.

Articles you may be interested inCommunication: Ultrafast time-resolved ion photofragmentation spectroscopy of photoionization-induced proton transfer in phenol-ammonia complex Ultrafast nonradiative dynamics in electronically excited hexafluorobenzene by femtosecond time-resolved mass spectrometry Ultrafast processes in photoexcited N-salicylideneaniline have been investigated with femtosecond time-resolved resonance-enhanced multiphoton ionization spectroscopy. The ion signals via the S 1 (n,*) state of the enol form as well as the proton-transferred cis-keto form emerge within a few hundred femtoseconds after photoexcitation to the first S 1 (,*) state of the enol form. This reveals that two ultrafast processes, excited-state intramolecular proton transfer ͑ESIPT͒ reaction and an internal conversion ͑IC͒ to the S 1 (n,*) state, occur on a time scale less than a few hundred femtoseconds from the S 1 (,*) state of the enol form. The rise time of the transient corresponding to the production of the proton-transferred cis-keto form is within 750 fs when near the red edge of the absorption is excited, indicating that the ESIPT reaction occurs within 750 fs. The decay time of the S 1 (,*) state of the cis-keto form is 8.9 ps by exciting the enol form at 370 nm, but it dramatically decreases to be 1.5-1.6 ps for the excitation at 365-320 nm. The decrease in the decay time has been attributed to the opening of an efficient nonradiative channel; an IC from S 1 (,*) to S 1 (n,*) of the cis-keto form promotes the production of the trans-keto form as the final photochromic products. The two IC processes may provide opposite effect on the quantum yield of photochromic products: IC in the enol form may substantially reduce the quantum yield, but IC in the cis-keto form increase it.

With the addition of water into liquid acetic acid, the CO stretching vibration band of acetic acid shows a high-frequency shift from 1665 to 1715 cm-1. This means that the hydrogen bond of the CO group of acetic acid is not as strong as those seen in liquid acetic acid or in CCl4 solution (in which the band appears at 1668 cm-1). A bent type hydrogen bond is accountable for this observation. On the other hand, the increase of acetic acid in water drastically decreases the intensity of the hydrogen-bonded O−H stretching Raman band of water at 3200 cm-1. This suggests that acetic acid breaks the hydrogen-bond networks of water. Low-frequency R(ν̄) spectra of acetic acid/water binary solutions are re-examined with new experimental data and ab initio molecular orbital analysis of intermolecular vibrational modes. The R(ν̄) spectrum of the aqueous mixture at x A = 0.5 bears a very close resemblance to that of the acetic acid/methanol mixture with x A = 0.5, indicating that the molecular complexes responsible for the Raman spectra are acetic acid clusters. The calculated low-frequency Raman feature of a side-on type dimer with bent-type hydrogen bonds based on ab initio molecular orbital theory reproduces the observed Raman pattern nicely. Any evidence of the formation of stable acid−water pairs is not found in the low-frequency Raman spectra. Furthermore, an isosbestic point is seen in the region of 0.1 ≤ x A (mole fraction of acetic acid) ≤ 0.5, and another one is also observed in 0.5 ≤ x A ≤ 1.0. The observed spectra in the region of 0 < x A < 0.5 are reproduced simply by linear combinations of the pure water spectrum and the spectrum at x A = 0.5. These results strongly suggest the presence of the two microphases with homogeneously associated molecules:  a water cluster phase and an acetic acid cluster phase. The spectral change in 0.5 < x A < 1.0 is attributed to the coexistence of the acetic acid cluster phase in aqueous environment and the acid associated phase characteristic of liquid acetic acid.

Fluorescence lifetime images of reduced nicotinamide adenine dinucleotide (NADH) that is a key cofactor in cellular metabolism were obtained in a cell at various values of intracellular pH. The average fluorescence lifetime of NADH is found to become shorter monotonically with increasing pH, indicating that pH in a single cell can be determined by fluorescence lifetime imaging of NADH without adding exogenous fluorescent probes. The magnitude of the pH-induced lifetime change is higher in cells than that in buffer solution. The fluorescence lifetime of NADH is not uniform inside a cell; the fluorescence lifetime of nuclear NADH is shorter than that of mitochondrial NADH at each pH, and the magnitude of the pH-induced change is larger in nuclei than in other areas. The local electric field effect on the fluorescence lifetime is discussed since this effect may be one of the strong possibilities for the nonuniformity of the autofluorescence lifetime of NADH in cells.

We have shown that the intracellular pH of a single HeLa cell expressing the enhanced green fluorescent protein (EGFP) can be imaged using the fluorescence lifetime of EGFP, which can be interpreted in terms of the pH-dependent ionic equilibrium of the p-hydroxybenzylidene-imidazolidinone structure of the chromophore of EGFP.

Fluorescence decays of flavin adenine dinucleotide (FAD) that is a typical autofluorescent species in cells and tissues have been measured in a mixture of alcohol and water in the femtosecond and nanosecond time range. The fluorescence lifetimes of both the stacked conformation between the isoalloxazine and adenine moieties in close proximity and the extended open conformation in water are affected by the addition of alcohol. The nanosecond fluorescence lifetime of the open conformation increases with decreasing dielectric constant of the medium, contributing to the enhancement of the fluorescence intensity of FAD in less dielectric media. The fluorescence lifetime of the open conformation is also affected by medium viscosity, suggesting that the photoexcited open conformation is quenched by the dynamic interaction between the two aromatic rings. The fluorescence component decaying in tens of picoseconds is attributed to the stacked conformation that shows the efficient fluorescence quenching due to the intramolecular electron transfer. The picosecond fluorescence lifetime of the stacked conformation increases with decreasing dielectric constant, suggesting the shift of the distribution of the stacked conformation to a longer intramolecular distance between the two aromatic rings in less dielectric media. The pre-exponential factor of the picosecond decaying component relative to that of the nanosecond one decreases with the increase of the alcohol concentration in the femtosecond time-resolved fluorescence, which demonstrates the increase in the population of the open conformation with the reduction of the dielectric constant. The possibility to evaluate the polar environment in a cell by the fluorescence lifetime of FAD is discussed based on the results obtained.

Electric-field-induced changes in absorption and emission spectra of colloidal CdS nanoparticles ranging in size from 1.0 to 5.0 nm in diameter have been measured by using electric field modulation spectroscopy. The analysis of the electroabsorption spectra indicates that the dipole moment in the first exciton state becomes larger with increasing particle size. The presence of the large dipole moment following photoexcitation into the first exciton band suggests that the CdS nanoparticles have large CT character in the first exciton state. The quantum yields both of the exciton emission and of the trap emission are markedly reduced by application of an electric field. On the basis of the direct measurements of the field-induced change in emission decay profile, it is suggested that the field-induced de-enhancement of these emission yields results from the field-induced decreases both in lifetime and in initial population of each emitting state. It is also found that the emission intensity of CdS nanoparticles increases under the UV light irradiation in air and decreases in a vacuum condition and that fluorescence lifetime in the former case is longer than that in the latter. This enhancement and de-enhancement process in emission intensity is almost reversible at least in several cycles.