Excess molar enthalpies of methyl alkanoates + n-nonane at 298.15 k

Microcalorimetric measurements of excess enthalpies were carried out over the whole concentration range for mixtures of nine methyl alkanoates (from acetate to decanoate) with n-nonane at 298.15 K. From these data, information could be obtained about the interactions between both chemical species. All show positive excess molar enthalpies which decrease with increasing length of the aliphatic chain of the methyl alkanoate.     

Single molecule force spectroscopy using polyproteins

In recent years single molecule force spectroscopy has emerged as a powerful new tool to explore the mechanical stability and folding pathways of individual proteins. This technique is used to apply a stretching force between two points of a protein, unfolding the protein to an extended state. By measuring the unfolding and folding trajectories of individual proteins, insight can be gained into the physical mechanisms of protein folding. In this tutorial review we introduce the reader to single molecule force spectroscopy using the atomic force microscope (AFM), and explain the two main modes of operation of the AFM for force spectroscopy: force-extension and force-clamp. We introduce the approach of using polyproteins to obtain a clear mechanical fingerprint for monitoring the response of proteins to an applied mechanical force. In addition, we provide an informative and representative review of recent research on proteins using single molecule force spectroscopy. We focus on areas which have made a significant contribution to the single molecule protein folding field and highlight emerging areas of research which have wider implications for the general scientific community.     

Single molecule force spectroscopy of asphaltene aggregates

Asphaltene aggregation and deposition cause severe problems in nearly all phases of petroleum processing. To resolve those problems, understanding the aggregation mechanisms is a prerequisite and has attracted the interest of a great number of investigators. However, to date, the nature and extent of asphaltene aggregation remain widely debated. In the present study, we attempt to investigate asphaltene aggregation from a completely new perspective. The technique of single molecule force spectroscopy (SMFS) was used to investigate the response of single asphaltene aggregates under an external pulling force. Force curves representing the stretching of single asphaltene aggregates were obtained in simple electrolyte solutions (KCl and calcium) and organic solvents (toluene and heptane). These force curves were well-fitted by the modified worm-like chain model, indicating that those asphaltene aggregates acted like long-chain polymers under pulling by an external force. It was found that lower solution pH values and the presence of divalent cations resulted in a lower bending rigidity of the formed aggregates. The information retrieved from the force curves suggests that asphaltene molecules with a structure featuring small aromatic clusters connected by aliphatic chains do exist and that asphaltene aggregation could occur through a linear polymerization mechanism. The current study extends the application scope of SMFS.     

Single molecule modulation spectroscopy of conjugated polymers

This paper describes a new single molecule spectroscopy approach for the investigation of triplet-triplet and singlet-triplet interactions in conjugated polymers. The technique involves the irradiation of isolated single, mulitchromophoric, conjugated polymer molecules by a repetitive sequence of variable-intensity microsecond time scale excitation pulses. The fluorescence intensity is synchronously time-averaged for thousands of cycles of the pulse sequence to yield a high signal-to-noise fluorescence transient on the microsecond time scale. The transient can be analyzed with kinetic models to obtain quantitative information about the kinetics of triplet-triplet exciton annihilation and the quenching of singlet excitons by triplet excitons in conjugated polymers.     

Single molecule spectroscopy as a probe for dye-polymer interactions

Experimental (Single Molecule Spectroscopy) and theoretical (quantum-chemical calculations and Monte Carlo and molecular dynamics simulations) techniques are combined to investigate the behavior and dynamics of a polymer-dye molecule system. It is shown that the dye molecule of interest (1,1'-dioctadecyl-3,3,3',3'-tetramethylindo-dicarbocyanine) adopts two classes of conformations, namely planar and nonplanar ones, when embedded in a poly(styrene) matrix. From an in-depth analysis of the fluorescence lifetime trajectories, the planar conformers can be further classified according to the way their alkyl side chains interact with the surrounding poly(styrene) chains.     

Single molecule spectroscopy studies of diffusion in mesoporous silica thin films

Single molecule spectroscopy is applied in studies of diffusion and surface adsorption in sol-gel-derived mesoporous silica thin films. Mesoporous films are obtained by spin casting surfactant-templated sols onto glass substrates. Small-angle X-ray diffraction results are consistent with hexagonally ordered mesophases in as-synthesized (i.e., surfactant-containing) films. Upon calcination, a 30% contraction and disordering of these structures occurs. Nile Red is used as a fluorescent probe of both the as-synthesized and calcined films. It is loaded into the samples at subnanomolar levels either prior to spin casting or after calcination. Fluorescence imaging and single-point fluorescence time transients show the dye molecules to be relatively mobile in the as-synthesized samples. In contrast, the molecules appear entrapped at fixed locations in dry calcined films. In calcined films rehydrated under high humidity conditions, the Nile Red molecules again become mobile. Time transients obtained from the as-synthesized and rehydrated samples provide clear evidence for frequent reversible adsorption of the dye to the silica surfaces. Autocorrelations of the time transients provide quantitative data on the mean diffusion coefficients (D = 2.4 x 10(-10) and 2.6 x 10(-10) cm2/s) and mean desorption times (1/k = 25 and 40 s) for the as-synthesized and rehydrated films, respectively. The results prove both water and surfactant play important roles in governing matrix interactions and mass transport.     

Single molecule atomic force microscopy and force spectroscopy of chitosan

Atomic force microscopy (AFM) and AFM-based force spectroscopy was used to study the desorption of individual chitosan polymer chains from substrates with varying chemical composition. AFM images of chitosan adsorbed onto a flat mica substrate show elongated single strands or aggregated bundles. The aggregated state of the polymer is consistent with the high level of flexibility and mobility expected for a highly positively charged polymer strand. Conversely, the visualization of elongated strands indicated the presence of stabilizing interactions with the substrate. Surfaces with varying chemical composition (glass, self-assembled monolayer of mercaptoundecanoic acid/decanethiol and polytetrafluoroethylene (PTFE)) were probed with chitosan modified AFM tips and the corresponding desorption energies, calculated from plateau-like features, were attributed to the desorption of individual polymer strands. Desorption energies of 2.0±0.3×10(-20)J, 1.8±0.3×10(-20)J and 3.5±0.3×10(-20)J were obtained for glass, SAM of mercaptoundecanoic/dodecanethiol and PTFE, respectively. These single molecule level results can be used as a basis for investigating chitosan and chitosan-based materials for biomaterial applications.     

Single molecule spectroscopy of conjugated polymer chains in an electric field-aligned liquid crystal

Using single molecule polarization spectroscopy, we investigated the alignment of a polymer solute with respect to the liquid crystal (LC) director in an LC device while applying an external electric field. The polymer solute is poly[2-methoxy-5-(2'-ethyl-hexyloxy)-1,4-phenylene vinylene] (or MEH-PPV), and the LC solvent is 5CB. The electric field induces a change in the LC director orientation from a planar alignment (no electric field) to a perpendicular (homeotropic) alignment with an applied field of 5.5 x 103 V/cm. We find that the polymer chains align with the LC director in both planar and homeotropic alignment when measured in the bulk of the LC solution away from the device interface. Single molecule polarization distributions measured as a function of distance from the LC device interface reveal a continuous change of the MEH-PPV alignment from planar to homeotropic. The observed polarization distributions are modeled using a conventional elastic model that predicts the depth profile of the LC director orientation for the applied electric field. The excellent agreement between experiment and simulations shows that the alignment of MEH-PPV follows the LC director throughout the LC sample. Furthermore, our results suggest that conjugated polymers such as MEH-PPV can be used as sensitive local probes to explore complex (and unknown) structures in anisotropic media.     

Desorption kinetic detection of different adsorption sites on opened carbon single walled nanotubes: The adsorption of n-nonane and CCl4

We show that thermal desorption kinetics clearly resolve adsorbates bound in different sites on single walled carbon nanotube bundles. The molecules n-C₉H₂₀ and CCl₄ were compared and it was found that the nanotube internal sites exhibited the highest desorption temperature, whereas external groove sites exhibited the next highest desorption temperature for both molecules. When n-C₉H₂₀ and CCl₄ coadsorb, the more strongly bound n-C₉H₂₀ quantitatively displaces CCl₄ from internal sites to groove sites. Molecular shape governs the capacity of the different sites for the two molecules.     

Single molecule kinetics. II. Numerical Bayesian approach

As discussed in the companion paper [J. B. Witkoskie and J. S. Cao, J. Chem. Phys. 121, 6361 (2004), preceding paper], quantitative extraction of information from single molecule experiments by several proposed indicators is difficult since the experiments only observe certain characteristics of the system, even though the indicators can contain all available information. This paper shows how one can circumvent the shortcomings of these indicators by combining information extracted from indicators with a numerical Bayesian statistical approach. The Bayesian approach determines the relative probability of various models reproducing the entire sequence of the single molecules trajectory, instead of binning and averaging over the data, which removes much of this information.     

Single carbon nanotube optical spectroscopy.

This Minireview discusses novel insights into the electronic structure of carbon nanotubes obtained using single-molecule fluorescence spectroscopy. Fluorescence spectra from single nanotubes are well described by a single, Lorentzian lineshape. Nanotubes with identical structures fluoresce with different energies due to local electronic perturbations. Carbon nanotube fluorescence unexpectedly does not-show any intensity or spectral fluctuations at 300 K The lack of intensity blinking or bleaching demonstrates that carbon nanotubes have the potential to provide a stable, single-molecule infrared photon source, allowing for the exciting possibility of applications in quantum optics and biophotonics.     

Single molecule force spectroscopy on poly(vinyl alcohol) by atomic force microscopy

The elastic properties of poly(vinyl alcohol) (PVA) were investigated on the nanoscale using the new technique of single molecule force spectroscopy by atomic force microscopy (AFM). It was found that the elastic properties of PVA molecules scale linearly with their contour lengths. This finding corroborates that the deformation of individual PVA chains is measured. The force spectra of PVA show a kink at around 200 pN and cannot be fitted by an extended Langevin function. The deviation of the elastic behavior of PVA from a freely jointed chain model may indicate the presence of a suprastructure of PVA in NaCl solution.     

Single molecule force spectroscopy reveals the context dependent folding pathway of the C-terminal fragment of Top7

Top7 is a de novo designed protein with atomic level accuracy and shows a folded structure not found in nature. Previous studies showed that the folding of Top7 is not cooperative and involves various folding intermediate states. In addition, various fragments of Top7 were found to fold on their own in isolation. These features displayed by Top7 are distinct from those of naturally occurring proteins of a similar size and suggest a rough folding energy landscape. However, it remains unknown if and how the intra-polypeptide chain interactions among the neighboring sequences of Top7 affect the folding of these Top7 fragments. Here we used single-molecule optical tweezers to investigate the folding–unfolding pathways of full length Top7 as well as its C-terminal fragment (CFr) in different sequence environments. Our results showed that the mechanical folding of Top7 involves an intermediate state that likely involves non-native interactions/structure. More importantly, we found that the folding of CFr is entirely dependent upon its sequence context in which it is located. When in isolation, CFr indeed folds into a cooperative structure showing near-equilibrium unfolding–folding transitions at ∼6.5 pN in OT experiments. However, CFr loses its autonomous cooperative folding ability and displays a folding pathway that is dependent on its interactions with its neighboring sequence/structure. This context-dependent folding dynamics and pathway of CFr are distinct from those of naturally occurring proteins and highlight the critical importance of intra-chain interactions in shaping the overall energy landscape and the folding pathway of Top7. These new insights may have important implications on the de novo design of proteins.

Optical tweezers experiments reveal that the folding of the C-terminal fragment of Top7 (cFr) is context-dependent. Depending on its neighboring sequence, cFr shows very different folding pathways and folding kinetics.      

Single molecule fluorescence correlation spectroscopy of single apoptotic cells using a red-fluorescent caspase probe

The detection of single molecules in single cells has enabled biochemical analyses to be conducted with high sensitivity and high temporal resolution. In this work, detection of apoptosis was studied by single molecule fluorescence correlation spectroscopy (FCS) in single living cells. Caspase activity was assayed using a new red fluorogenic probe that avoids the spectral overlap of green fluorescent probes and cell autofluorescence. This new probe, 2SBPO-Casp, was synthesized by coupling a water-soluble Nile Blue derivative (2SBPO) to an aspartic acid residue. Upon apoptosis induction and caspase activation, free 2SBPO dye is shown to accumulate inside the cell after probe cleavage. In previous work in our lab, single molecule fluorescence in single apoptotic cells was detected 45 min after induction using a rhodamine 110-based probe. However, significant statistical analysis was needed to exclude false positives. The use of 2SBPO-Casp overcomes the autofluorescence problem and offers a steady fluorescence signal. In our single molecule FCS measurements, Ramos cells were determined apoptotic on the basis of their correlation coefficient value (R(2)). Cells that contain an R(2) ≥ 0.65 were identified as highly correlated and therefore determined to be apoptotic. Single apoptotic cells identified in this manner were found as early as 30 min after induction and the number of apoptotic cells reached a peak value at the 3rd hour, which is consistent with other techniques. Using single molecule techniques and a new apoptosis probe, the temporal dynamics were elucidated with better sensitivity and resolution than in previous studies.     

Sub-doppler spectroscopy of the NaCs molecule

Doppler-free polarization spectroscope in combination with double-resonance techniques and laser-induced fluorescence spectroscope has been used to det     

Single molecule kinetics. I. Theoretical analysis of indicators

Single molecule experiments reveal intriguing phenomenon in chemical and biological systems. Several indicators of complex dynamics, including "intensity" correlations, "event" correlations, and characteristic functions have been proposed, but extraction of information from these indicators can be difficult since these indicators only observe certain characteristics of the system. Generally, for systems that follow Poisson kinetics, all of these indicators contain similar information about the relaxation times of the system and the connections between different relaxation times, but the information is convoluted in different ways so the strength of various indicators is system specific. The paper discusses the theoretical implications and information content of various data analysis methods for single molecule experiments and demonstrates the relationships between indicators. Under certain conditions, common indicators contain all available information about systems with Poisson kinetics between degenerate states, but extraction of this information is generally not numerically feasible. The paper also discusses practical issues associated with these analyses, which motivates a numerical study based on Bayes' formula in the companion paper [J. Witkoskie and J. S. Cao, J. Chem. Phys. 121, 6373 (2004), following paper].     

Reverse Intersystem Crossing of Single Deuterated Perylene Molecules in a Dibenzothiophene Matrix

Intersystem crossing to the long-lived metastable triplet state is often a strong limitation on fluorescence brightness of single molecules, particularly for perylene in various matrices. In this paper, we report on a strong excitation-induced reverse intersystem crossing (rISC), a process where single perylene molecules in a dibenzothiophene matrix recover faster from the triplet state, turning into bright emitters at saturated excitation powers. With a detailed study of single-molecule fluorescence autocorrelations, we quantify the effect of rISC. The intrinsic lifetimes found for the two effective triplet states (8.5±0.4 ms and 64±12 ms) become significantly shorter, into the sub-millisecond range, as the excitation power increases and fluorescence brightness is ultimately enhanced at least fourfold. Our results are relevant for the understanding of triplet state manipulation of single-molecule quantum emitters and for markedly improving their brightness.     

Single DNA molecule isolation and trapping in a microfluidic device.

This paper presents a systematic method to isolate and trap long single DNA segments between integrated electrodes in a microfluidic environment. Double stranded lambda-DNA molecules are introduced in a microchip and are isolated by electrophoretic force through microfluidic channels. Downstream, each individual molecule is extended and oriented by ac dielectrophoresis (900 kHz, 1 MV m(-1)) and anchored between aluminium electrodes. With a proper design, a long DNA segment (up to 10 microm) can be instantly captured in stretched conformation, opening way for further assays.     

Structural evolutions of the n-heneicosane and n-tricosane molecular alloys at 293 K

A structural study of odd-numbered n-alkane (C_n) binary mixtures (C₂₁ : C₂₃) was carried out on powder samples using a Guinier-de Wolff camera with increasing concentration of n-C₂₃ at 293 K.

Despite the reports in the literature, these molecular alloys do not form an orthorhombic continuous homogeneous solid solution to C₂₁ from C₂₃ at “low temperature”. Instead, as already observed in two even-numbered C_n systems, X-ray diffraction results show the existence of seven solid solutions as the molar concentration of C₂₃ increases: four terminal solid solutions, denoted β₀(C₂₁)β₀(C₂₃), isostructural with the “low temperature” phase of pure C₂₁ and C₂₃ (Pbcm), β′₀(C₂₁) and β′₀(C₂₃), identical to the phase β′₀ which appears in pure C₂₃ above the δ transition, and three orthorhombic intermediate solid solutions, designated β″₁, β′₁ and β″₂.

On the basis of powder X-ray photographs, the phases β″₁ and β″₂ (C₂₁ : C₂₃) are indistinguishable, and they are isostructural with the intermediate solid solution β″ of the even-numbered C_n binary systems (C₂₂ : C₂₄) and (C₂₄ : C₂₆). The phase β′₁(C₂₁ : C₂₃) is also isostructural with the two indistinguishable intermediate solid solutions β′₁ and β′₂ of the molecular alloys (C₂₂ : C₂₄) and (₂₄ : C₂₆).

From this study and our other laboratory results, the sequences of appearance of the solid solutions and the structural identities between these phases are established at “low temperature” for all the binary molecular alloys of consecutive C_n (odd-odd, even-even or odd-even: 19 < n < 27) when increasing the solute concentration.