Optical rotation: recent advances in determining the absolute configuration

The field of optical rotations is currently undergoing a renaissance, which is a direct result of the advances in quantum mechanics and the availability of faster desktop computers. In the last few years, the field of optical rotations has taken an important role for determining the three-dimensional molecular structure (absolute configuration and conformations) with confidence. The purpose of this review is to present the latest advances in this area so that practicing chemists can utilize them in their respective areas of research and development.     

Estimation of optical rotation of γ‐alkylidenebutenolide,cyclopropylamine, cyclopropyl‐methanol and cyclopropenone based compounds by a Density Functional Theory (DFT) approach

Computing the optical rotation of organic molecules can be a real challenge, and various theoretical approaches have been developed in this regard. A benchmark study of optical rotation of various classes of compounds was carried out by Density Functional Theory (DFT) methods. The aim of the present research study was to find out the best‐suited functional and basis set to estimate the optical rotations of selected compounds with respect to experimental literature values. Six DFT functional LSDA, BVP86, CAM‐B3LYP, B3PW91, and PBE were applied on 22 different compounds. Furthermore, six different basis sets, i.e., 3‐21G, 6‐31G, aug‐cc‐pVDZ, aug‐cc‐pVTZ, DGDZVP, and DGDZVP2 were also applied with the best‐suited functional B3LYP. After rigorous effort, it can be safely said that the best combination of functional and basis set is B3LYP/aug‐cc‐pVTZ for the estimation of optical rotation for selected compounds.     

Specific Optical Rotations and the Horeau Effect

The observation of nonequivalence of optical and enantiomeric purities, referred to as the Horeau effect, is thought to arise from molecular aggregation in liquid solutions. Although this effect was first observed in 1969, the conditions under which this effect may, or may not, be observable are not established. Considering the formation of dimers as the simplest form of aggregation, the expressions for specific optical rotations in the presence of homochiral and heterochiral monomer–dimer equilibria are presented. Analysis of these equations indicates that the Horeau effect will not be observable even in the presence of aggregation under either of the following two situations: 1) The specific optical rotation of the monomeric species is equal to that of the dimeric species; 2) The heterochiral equilibrium constant is twice that of the homochiral equilibrium constant. Chirality 28:181–185, 2016. © 2015 Wiley Periodicals, Inc.     

Ab initio prediction of optical rotation: comparison of density functional theory and Hartree-Fock methods for three 2,7,8-trioxabicyclo[3.2.1]octanes

We report ab initio calculations of the frequency-dependent electric dipole-magnetic dipole polarizabilities, beta(nu), at the sodium D line frequency and, thence, of the specific rotations, [alpha](D), of 2,7,8-trioxabicyclo[3.2.1]octane, 1, and its 1-methyl derivative, 2, using the Density Functional Theory (DFT) and Hartree-Fock/Self-Consistent Field (HF/SCF) methodologies. Gauge-invariant (including) atomic orbitals (GIAOs) are used to ensure origin-independent [alpha](D) values. Using large basis sets which include diffuse functions DFT [alpha](D) values are in good agreement with experimental values (175.8 degrees and 139.2 degrees for (1S,5R)-1 and -2, respectively); errors are in the range 25-35 degrees. HF/SCF [alpha](D) values, in contrast, are much less accurate; errors are in the range 75-95 degrees. The use of small basis sets which do not include diffuse functions substantially lowers the accuracy of predicted [alpha](D) values, as does the use of the static limit approximation: beta(nu) approximately beta(o). The use of magnetic-field-independent atomic orbitals, FIAOs, instead of GIAOs, leads to origin-dependent, and therefore nonphysical, [alpha](D) values. We also report DFT calculations of [alpha](D) for the 1-phenyl derivative of 1, 3. DFT calculations find two stable conformations, differing in the orientation of the phenyl group, of very similar energy, and separated by low barriers. Values of [alpha](D) predicted using two different algorithms for averaging over phenyl group orientations are in good agreement with experiment. In principle, the absolute configuration (AC) of a chiral molecule can be assigned by comparison of the optical rotation predicted ab initio to the experimental value. Our results demonstrate the critical importance of the choice of ab initio methodology in obtaining reliable optical rotations and, hence, ACs, and show that, at the present time, DFT constitutes the method of choice.     

光学相干层析分子成像方法初探

介绍了分子对比剂在光学相干层析成像(optical coherence tom ography,OCT)技术中的研究现状,概述了迄今出现的几种不同的光学相干层析分子成像方法(m olecu lar contrast OCT,简称为MCOCT),讨论了MCOCT的几个重要的实际问题:对比剂的选择范围、激发光强的限制、各种方法灵敏度比较以及MCOCT应用于临床与生物学领域需要考虑的因素。     

量子点在生物学中的研究进展

量子点作为一种新型的荧光标记物近年来已在生物学中获得广泛应用。本文总结了量子点的主要光学特性,其中包括荧光激发和发射光谱特性、量子产额、光漂白特性和荧光寿命等。重点综述了量子点在细胞标记、活体和组织成像、组合标记和光动力学治疗等生物学中的应用及其最新研究进展。同时讨论了量子点在应用中可能存在的细胞毒性等主要问题,最后对量子点在生物学中的应用前景作了展望。     

Simultaneous measurement of optical rotation dispersion and absorption spectra for chiral substances

We present a new method based on optical null methods for simultaneously measuring the optical rotatory dispersion (ORD) and absorption spectra of chiral substances. The optical rotation angle at a specific wavelength can be obtained from the optical nulls of the Malus curves with and without the sample. We use the optical nulls of the two curves as benchmark points and the readings to the right of the benchmark points by a certain angular offset to eliminate the influence of the analyzer on the light intensity and obtain the absorbance of the chiral substance at a specific wavelength. The 4096 pixels of the line scan CCD can measure multiple wavelengths simultaneously so that continuous ORD and absorption spectra can be obtained. The experimental results show that the standard deviation of the specific optical rotation is 0.11 deg mL g⁻¹ dm⁻¹, the standard deviation of the maximum absorption wavelength is 0.45 nm, and that absorbance of the sample varies linearly with the concentration. This method is helpful for simplifying the experiment and has a profound influence on the analysis of the contents and molecular configurations of chiral substances in the future.     

The design and use of an optical mapping system for the study of intracardiac electrical signaling

Fluorescent optical mapping of electrically active cardiac tissues provides a unique method to examine the excitation wave dynamics of underlying action potentials. Such mapping can be viewed as a bridge between cellular level and organ systems physiology, e.g., by facilitating the development of advanced theoretical concepts of arrhythmia. We present the design and use of a high-speed, high-resolution optical mapping system composed entirely of "off the shelf" components. The electrical design integrates a 256 element photodiode array with a 16 bit data acquisition system. Proper grounding and shielding at various stages of the design reduce electromagnetic interference. Our mechanical design provides flexibility in terms of mounting positions and applications (use for whole heart or tissue preparations), while maintaining precise alignment between all optical components. The system software incorporates a user friendly graphical user interface, e.g., spatially recorded action potentials can be represented as intensity graphs or in strip chart format. Thus, this system is capable of displaying cardiac action potentials with high spatiotemporal resolution. Results from cardiac action potential mapping with intact mouse hearts are provided. It should be noted that this system could be readily configured to study isolated myocardial biopsies (e.g., isolated ventricular trabeculae). We describe the details of a versatile, user-friendly system that could be employed for a magnitude of study protocols.     

光钳及其在生物学中的应用

本文综述了光钳的原理及其在细胞生物学、免疫学及分子遗传学中的应用。将光钳和激光微束系统联合使用可以进行显微操作。其特点是操作简便,无机械接触並且绝对无菌。光钳和微束系统可用来切割染色体,收集染色体片段,融合细胞以及改变细胞和细胞器的行为。     

Polarimetric detection in high-performance liquid chromatography

Chiroptical detection for HPLC is particularly useful as a selective detection method for chiral molecules, and in enantiomeric purity determination with partial chiral separation or without chiral separation. The recent development of laser-based polarimeters with microdegree sensitivity has increased the applicability of optical rotation detection in HPLC. The detection limit of these instruments is submicrogram on-column for many chiral compounds in analytical HPLC. A variety of applications of the selective detection of optically active molecules are reviewed. The use of polarimetric detection with partial chiral separation is considered, both as an aid to method development and for enantiomeric purity determination. Finally applications to enantiomeric purity determination without chiral separation are reviewed, with the dual use of nonchirally selective and chiroptical detectors to determine the total amount and optical purity of the analyte. Determinations of chiral purity for samples of high enantiomeric excess are described, which with laser-based instrumentation may give accuracies of better than +/- 1% with sample loadings of 50 micrograms on an achiral column. Applications to the study of enantioselective reactions are also considered, with determination of enantiomeric excess in near-racemates to better than +/- 0.1%.     

Conformational aspects of polypeptides. XVIII. Conformational studies of oligopeptides derived from L-alanine

The conformations of oligopeptides derived from L -alanine and co-oligomers of L -alanine with γ-methyl-L -glutamate were studied in several solvents via optical rotation and far-ultraviolet spectroscopy. Calculated values for optical rotation based on model compounds were compared with experimental values for the oligomers. In trifluoroacetic and dichloroacetic acids, the oligomers and co-oligomers exhibit rotations in close agreement with predicted values based on model compounds. Thus, in these solvents only nonhelical conformations exist. In trifluoroethanol, the experimental points of molar rotation for the pentamer and larger oligomers no longer follow the predicted values. In addition, the benzyloxycarbonyl and acetyl cononamers show b₀ values of about ?150, which demonstrates the presence of stable helical forms for these peptides. We also examined the molar extinction coefficients of oligopeptides in the 190 mμ region and determined the values for nonhelical peptide groups. The molar extinction coefficients per amide bond for the benzyloxycarbonyl and acetyl cononamers show extensive hypo-chromism, once again indicating the presence of stable helices for these compounds in trifluoroethanol.     

Conformational studies on polynucleotide chains. IV. Backbone structure of ribonucleic acids.

In preceding papers the energies associated with the internal rotations in the sugar–phosphate–sugar complex were described with an analytical potential consisting of a Lennard-Jones 6–12 term and an intrinsic torsional term and representing the best fit to a large number of energies computed with a quantum mechanical ab initio technique. The complex considered there (of 37 atoms and with the chemical formula C₁₀H₁₈O₈P) is repesentative of deoxyribonucleic acids. In this paper we apply our potential to evaluating the intramolecular energies of the 39-atom complex C₁₀H₁₈O₁₀P, representative of the ribonucleic acids. The potential energies for the internal rotations (considered independent from one another) and the energy maps for rotations about consecutive bonds of the backbone chain are critically compared, both with those obtained for the deoxy system and with those obtained from different theoretical approaches as available from literature. It is shown that, at least for certain combinations of the internal rotation angles, the choice of the starting geometry for the sugarphosphate–sugar molecule (bond lengths and valence angles) strongly affects the value of the computed energy. If a proper geometry is used, very low energies are predicted by our potential in correspondence of the sets of torsional angles found in various RNAs by x-ray crystallography.     

Bioconjugated quantum dots for multiplexed and quantitative immunohistochemistry

Bioconjugated quantum dots (QDs) provide a new class of biological labels for evaluating biomolecular signatures (biomarkers) on intact cells and tissue specimens. In particular, the use of multicolor QD probes in immunohistochemistry is considered one of the most important and clinically relevant applications. At present, however, clinical applications of QD-based immunohistochemistry have achieved only limited success. A major bottleneck is the lack of robust protocols to define the key parameters and steps. Here, we describe our recent experience, preliminary results and detailed protocols for QD-antibody conjugation, tissue specimen preparation, multicolor QD staining, image processing and biomarker quantification. The results demonstrate that bioconjugated QDs can be used for multiplexed profiling of molecular biomarkers, and ultimately for correlation with disease progression and response to therapy. In general, QD bioconjugation is completed within 1 day, and multiplexed molecular profiling takes 1-3 days depending on the number of biomarkers and QD probes used.     

Calculation of conformational energies and optical rotation of the most simple chiral alkane

Quantum chemical calculations have been performed to investigate the conformer distribution of 4-ethyl-4-methyloctane and its optical rotation. With the reference methods MP2 and SCS-MP2, the energies of seven conformers are found within a range of about 1.5 kcal mol(-1). It is demonstrated that the relative energies cannot be reliably predicted with conventional GGA or hybrid density functionals, Hartree-Fock, semiempirical AM1, and classical force field (MM3) calculations. An empirical dispersion correction to GGA (PBE-D), hybrid (B3LYP-D), or double hybrid (B2PLYP-D) functionals corrects these errors and results in very good agreement with the reference energies. Optical rotations have been calculated for all seven conformers at the TDDFT(BHLYP/aTZV2P) level. The computed macroscopic rotation is derived from a classical Boltzmann average. The result (1.9-3.2 deg dm(-1) (g/mL)(-1)) is very close to the experimental value of 0.2 deg dm(-1) (g/mL)(-1) for the (R)-enantiomer and has the right sign. Because six conformers are significantly populated at room temperature and the rotations of individual conformers differ in sign and magnitude, the calculated average rotation is rather sensitive to the conformer population used. From the electronic structure point of view, this example emphasizes the effect of long-range dispersion effects for the evaluation of population averaged quantities in large molecules. Computations based on free enthalpies are in worse agreement with experiment that is attributed to artefacts of the harmonic approximation used to compute the vibrational entropy terms.     

Design and validation of surface-marker clusters for the quantification of joint rotations in general movements in early infancy

Lack of complexity in general movements in early infancy is an important marker of potential motor disorders of neurological origin, such as cerebral palsy. Quantitative approaches to characterising this complexity are hampered by experimental difficulties in recording from infants in their first few months of life. The aim of this study was to design and validate bespoke surface-marker clusters to facilitate data acquisition and enable full quantification of joint rotations. The clusters were validated by recording the controlled movements of a soft-body dummy doll simultaneously with an optical (Qualisys) and inertial (XSens) motion capture system. The angles estimated from the optical system were compared with those measured by the inertial system. We demonstrate that the surface-marker based approach compares well with the use of an inertial system to obtain "direct" readings of the rotations whilst alleviating the issues associated with the use of an optical motion capture system. We briefly report use of this technique in 1-5 month old infants. By enabling full quantification of joint rotation, use of the custom made markers could pave the way for early diagnosis of movement disorders.     

Assessment of flexible backbone protein design methods for sequence library prediction in the therapeutic antibody Herceptin-HER2 interface

Computational protein design methods can complement experimental screening and selection techniques by predicting libraries of low-energy sequences compatible with a desired structure and function. Incorporating backbone flexibility in computational design allows conformational adjustments that should broaden the range of predicted low-energy sequences. Here, we evaluate computational predictions of sequence libraries from different protocols for modeling backbone flexibility using the complex between the therapeutic antibody Herceptin and its target human epidermal growth factor receptor 2 (HER2) as a model system. Within the program RosettaDesign, three methods are compared: The first two use ensembles of structures generated by Monte Carlo protocols for near-native conformational sampling: kinematic closure (KIC) and backrub, and the third method uses snapshots from molecular dynamics (MD) simulations. KIC or backrub methods were better able to identify the amino acid residues experimentally observed by phage display in the Herceptin-HER2 interface than MD snapshots, which generated much larger conformational and sequence diversity. KIC and backrub, as well as fixed backbone simulations, captured the key mutation Asp98Trp in Herceptin, which leads to a further threefold affinity improvement of the already subnanomolar parental Herceptin-HER2 interface. Modeling subtle backbone conformational changes may assist in the design of sequence libraries for improving the affinity of antibody-antigen interfaces and could be suitable for other protein complexes for which structural information is available.     

Molecular dynamics simulations of forced conformational transitions in 1,6-linked polysaccharides

Recent atomic force microscopy stretching measurements of single polysaccharide molecules suggest that their elasticity is governed by force-induced conformational transitions of the pyranose ring. However, the mechanism of these transitions and the mechanics of the pyranose ring are not fully understood. Here we use steered molecular dynamics simulations of the stretching process to unravel the mechanism of forced conformational transitions in 1,6 linked polysaccharides. In contrast to most sugars, 1,6 linked polysaccharides have an extra bond in their inter-residue linkage, C5-C6, around which restricted rotations occur and this additional degree of freedom increases the mechanical complexity of these polymers. By comparing the computational results with the atomic force microscopy data we determine that forced rotations around the C5-C6 bond have a significant and different impact on the elasticity of alpha- and beta-linked polysaccharides. Beta-linkages of a polysaccharide pustulan force the rotation around the C5-C6 bonds and produce a Hookean-like elasticity but do not affect the conformation of the pyranose rings. However, alpha-linkages of dextran induce compound conformational transitions that include simultaneous rotations around the C5-C6 bonds and chair-boat transitions of the pyranose rings. These previously not-recognized transitions are responsible for the characteristic plateau in the force-extension relationship of dextran.     

Single-molecule Analysis of Inhibitory Pausing States of V1-ATPase

V(1)-ATPase, the hydrophilic V-ATPase domain, is a rotary motor fueled by ATP hydrolysis. Here, we found that Thermus thermophilus V(1)-ATPase shows two types of inhibitory pauses interrupting continuous rotation: a short pause (SP, 4.2 s) that occurred frequently during rotation, and a long inhibitory pause (LP, >30 min) that terminated all active rotations. Both pauses occurred at the same angle for ATP binding and hydrolysis. Kinetic analysis revealed that the time constants of inactivation into and activation from the SP were too short to represent biochemically predicted ADP inhibition, suggesting that SP is a newly identified inhibitory state of V(1)-ATPase. The time constant of inactivation into LP was 17 min, consistent with one of the two time constants governing the inactivation process observed in bulk ATPase assay. When forcibly rotated in the forward direction, V(1) in LP resumed active rotation. Solution ADP suppressed the probability of mechanical activation, suggesting that mechanical rotation enhanced inhibitory ADP release. These features were highly consistent with mechanical activation of ADP-inhibited F(1), suggesting that LP represents the ADP-inhibited state of V(1)-ATPase. Mechanical activation largely depended on the direction and angular displacement of forced rotation, implying that V(1)-ATPase rotation modulates the off rate of ADP.     

Motion from the past. A new method to infer vestibular capacities of extinct species

The vestibular system detects head movement in space and maintains visual and postural stability. The semicircular canal system is responsible for registering head rotation. How it responds to head rotation is determined by the rotational axis and the angular acceleration of the head, as well as the sensitivity and orientation of each semicircular canal. The morphological parameters of the semicircular canals are supposed to allow an optimal detection of head rotations induced by some behaviours, especially locomotor. We propose a new method of semicircular canal analysis, based on the computation of central streamlines of virtually reconstructed labyrinths. This method allows us to ascertain the functional structure of the semicircular canal system and to infer its capacity to detect particular head rotations, induced by particular behaviours. In addition, this method is well-suited for datasets provided by any kind of serial sectioning methods, from MRI to μCT scanning and even mechanical serial sectioning, of extant and extinct taxa.