Optical Rotatory Dispersion Spectroscopy Research Articles

On the Optical Activity of Poly(l-lactic acid) (PLLA) Oligomers and Polymer: Detection of Multiple Cotton Effect on Thin PLLA Solid Film Loaded with Two Dyes

Optical rotatory dispersion (ORD) is a beautiful analytical technique for the study of chiral molecules and polymers. In this study, ORD was applied successfully to follow the degree of polycondensation of l-(+)-lactic acid toward the formation of poly(lactic acid) oligomers (PLAO) and high molecular weight poly(l-lactic acid) (PLLA) in a simple esterification reaction equipment. PLLA is a biodegradable polymer obtainable from renewable raw materials. The racemization of the intrinsically isotactic PLLA through thermal treatment can be easily followed through the use of ORD spectroscopy. Organic or molecular electronics is a hot topic dealing with the combination of π-conjugated organic compounds and polymers with specific properties (e.g., chirality) which can be exploited to construct optoelectronic devices, such as organic light-emitting diodes (OLEDs), organic photovoltaic (OPV) high efficiency cells, switchable chirality devices, organic field-effect transistors (OFETs), and so on. ORD spectroscopy was applied to study either the gigantic optical rotation of PLLA films, as well as to detect successfully the excitonic coupling, occurring in thin solid PLLA green film loaded with a combination of two dyes: SY96 (a pyrazolone dye) and PB16 (the metal-free phthalocyanine pigment). The latter compound PLLA loaded with SY96 and PB16 shows a really gigantic optical activity in addition to typical ORD signal due to exciton coupling and may be considered as a simple and easily accessible model composite of a chiral polymer matrix combined with π-conjugated dyes for molecular electronics studies.

Unfolding Dynamics of a Photoswitchable Helical Peptide

We present an atomistic force field for the azo-moiety of the photoswitchable FK-11-X peptide. We use the parameters to study the unfolding of the peptide through molecular dynamics simulations. The unfolded ensemble contains many different structures, ranging from a partially unfolded peptide to a fully unfolded structure. The averaged computed far-ultraviolet circular dichroism (CD) spectrum of the set of structures, which was simulated using the newly developed force field, agrees well with experiment. The rate of the simulated unfolding process was estimated to have a time constant of 5.80 ± 0.03 ns from the time evolution of the CD spectrum of the peptide, computed from the backbone conformations sampled over 40 simulated trajectories. Our estimated time constant is faster than, but not inconsistent with, previous experimental estimates from time-resolved infrared and optical rotatory dispersion spectroscopy.

Radiolysis and radioracemization of RNA ribonucleosides: implications for the origins of life

The RNA nucleosides, namely adenosine, cytidine, guanosine and uridine were γ-radioyzed in solid state and in vacuo at room temperature to a total dose of 3.2 MGy. Through electronic absorption spectroscopy, differential scanning calorimetry and through polarimetry and optical rotatory dispersion spectroscopy, it was found that the purine-based nucleosides (adenosine and guanosine) show a much higher radiolysis resistance than the pyrimidine-based nucleosides (cytidine and uridine). In an astrochemical/astrobiological context, these results may explain why purine nucleobases are found in practically all carbonaceous chondrite meteorites while the pyrimidine nucleobases are absent or below the detection limits of the current analytical techniques. In the hypothesis that both purines and pyrimidines nucleobases were present in certain bodies at the beginning of the solar system 4.6 × 109 years ago, the radiolysis due to radionuclides decay has destroyed more easily and completely the pyrimidine bases due to their much lower radiolysis resistance than the purine bases.

Chiroptical properties of cholesteric liquid crystals of chitosan phenylcarbamate in ionic liquids

Chitosan phenylcarbamate (CtsPC) samples were synthesized with different degrees of substitution (DS) ranging from 2.3 to 3.5, and their lyotropic liquid crystallinity in ionic liquids was investigated. 1-Ethyl-3-methylimidazolium dicyanamide ([C2Mim][N(CN)2]) was found to be a novel solvent for CtsPC, and the concentrated lyotropic system formed a cholesteric liquid-crystalline phase and mostly exhibited vivid colorations owing to selective reflection of visible light. Ultraviolet–visible–near-infrared spectroscopy and wide-angle X-ray diffractometry were used to evaluate the maximum wavelength of selective light reflection and the distance and rotational difference between adjacent nematic thin layers in the cholesteric mesophase, respectively. The cholesteric helical pitch increased with decreasing polymer concentration, increasing temperature or increasing DS. The helical sense remained right-handed under the adopted measurement conditions, although CtsPCs are known to form a left-handed cholesteric structure in polar organic solvents, such as N,N-dimethylformamide (DMF). Accordingly, an inversion of the cholesteric sense was achieved by combination of [C2Mim][N(CN)2] and DMF as solvent; circular dichroism and optical rotatory dispersion spectroscopy verified that the sense of CtsPC cholesterics changed from left-handed to right-handed with increasing [C2Mim][N(CN)2] content in the mixed solvent. Chitosan phenylcarbamate (CtsPC) samples were synthesized and their lyotropic liquid crystallinity in ionic liquids was investigated. 1-Ethyl-3-methylimidazolium dicyanamide ([C2Mim][N(CN)2]) was found to be a novel solvent for CtsPC, and the lyotropic system formed a cholesteric liquid-crystalline phase and exhibited vivid colorations owing to selective light reflection. The cholesteric helical pitch was controllable by polymer concentration, temperature or the degree of phenylcarbamoyl substitution. Although the helical sense of CtsPC/[C2Mim][N(CN)2] cholesterics remained right-handed under the adopted measurement conditions, an inversion of cholesteric sense was achieved by using a mixed solvent of [C2Mim][N(CN)2]/N,N-dimethylformamide.

Andhraxylocarpins A–E: Structurally Intriguing Limonoids from the True Mangroves Xylocarpus granatum and Xylocarpus moluccensis

Five new limonoids, including andhraxylocarpins A and B (1 and 2) which contain a 9-oxa-tricyclo[3.3.2.1(7, 10)]undecane-2-ene motif, andhraxylocarpins C and D (3 and 4), which contain a (Z)-bicyclo[5.2.1]dec-3-en-8-one substructure, and andhraxylocarpin E (5), which contains a tricyclo[3.3.1.1(3, 6)]decane-9-one scaffold, were isolated from the seeds of two true mangroves, Xylocarpus granatum and Xylocarpus moluccensis, that were collected in the estuaries of Andhra Pradesh, India. The absolute configurations of these compounds were determined by extensive NMR investigations, single-crystal X-ray diffraction analysis, and by circular dichroism and optical rotatory dispersion spectroscopy, in combination with quantum-chemical calculations. The pronounced structural diversity of limonoids from these mangroves might originate from environmental factors.

Solid state radiolysis of non-proteinaceous amino acids in vacuum: astrochemical implications

The analysis of the amino acids present in Murchison meteorite and in other carbonaceous chondrites has revealed the presence of 66 different amino acids. Only eight of these 66 amino acids are proteinaceous amino acids used by the present terrestrial biochemistry in protein synthesis, the other 58 amino acids are somewhat “rare” or unusual or even “unknown” for the current terrestrial biochemistry. For this reason in the present work a series of “uncommon” non-proteinaceous amino acids, namely, l-2-aminobutyric acid, R(−)-2-aminobutyric acid, 2-aminoisobutyric acid (or α-aminoisobutyric acid), l-norleucine, l-norvaline, l-β-leucine, l-β-homoalanine, l-β-homoglutamic acid, S(−)-α-methylvaline and dl-3-aminoisobutyric acid were radiolyzed in vacuum at 3.2 MGy a dose equivalent to that emitted in 1.05 × 109 years from the radionuclide decay in the bulk of asteroids or comets. The residual amount of each amino acid under study remained after radiolysis was determined by differential scanning calorimetry in comparison to pristine samples. For optically active amino acids, the residual amount of each amino acid remained after radiolysis was also determined by optical rotatory dispersion spectroscopy and by polarimetry. With these analytical techniques it was possible to measure also the degree of radioracemization undergone by each amino acid after radiolysis. It was found that the non-proteinaceous amino acids in general do not show a higher radiation and radioracemization resistance in comparison to the common 20 proteinaceous amino acids studied previously. The unique exception is represented by α-aminoisobutyric acid which shows an extraordinary resistance to radiolysis since 96.6 % is recovered unchanged after 3.2 MGy. Curiously α-aminoisobutyric acid is the most abundant amino acid found in carbonaceous chondrites. In Murchison meteorite α-aminoisobutyric acid represents more than 20 % of the total 66 amino acids found in this meteorite.

Ozonolysis of α-PINENE, β-PINENE, d- and l-Turpentine Oil Studied by Chirooptical Methods; Some Implications on the Atmospheric Chemistry of Biogenic Volatile Organic Compounds

The main constituents of biogenic volatile organic compounds (BVOC) released in the atmosphere from vegetation are terpenes. The most common terpenes are α-pinene and β-pinene. These molecules react with ozone, forming oxidized derivatives which have low vapor pressure and which nucleate into secondary organic aerosol (SOA). Such aerosol-forming reactions have been simulated in the gas phase on laboratory scale with the visualization of the formation of SOA. Electronic absorption spectroscopy shows that the UV peak of pinenes at 205 nm is shifted to 225 nm during and after ozonolysis and is accompanied by a series of spectral features in the near infrared region (between 850 nm and 1100 nm). Optical rotatory dispersion (ORD) spectra were recorded on pure β(+)pinene and β(−)pinene. After ozonolysis β-pinene enantiomers are converted into nopinone enantiomers. The ORD spectra of both (−)nopinone and (+)nopinone respectively were recorded. The former showed a peak at 453 nm, an inflection point at 430 nm and a trough at 405 nm. The ORD spectrum of (−)-nopinone appears completely symmetrical to that of (+)nopinone. Thus, ORD spectroscopy can distinguish between BVOC and oxidized BVOC quite easily at least in the case of β-pinene. The ORD spectra of α-pinene enantiomers are different from those of β-pinene enantiomers and although the ORD spectra of ozonized α-pinene are not easily distinguishable from those of the primary compounds, they are completely different from those of ozonized β-pinene enantiomers. A reasonable natural model of BVOC is offered by l- and d-turpentine oil whose composition reflects that of the BVOC from conifer forests. The ORD spectra of ozonized l- and d-turpentine has permitted to distinguish between the levorotatory and dextrorotatory oxidation products and showing the ketone peak at about 367–380 nm, thus permitting in a real model to distinguish between primary BVOC (turpentine) and oxidized BVOC (or SOA) corresponding to ozonized turpentine. It has been proposed that ORD spectra in the gas phase could be used to detect chiral BVOC and oxidized chiral BVOC.

Radioracemization and radiation-induced chiral amplification of chiral terpenes measured by optical rotatory dispersion (ORD) spectroscopy

For the first time the radioracemization of α(+)pinene and α(−)pinene, of turpentine and of R(−)- α-phellandrene has been studied by optical rotatory dispersion (ORD) spectroscopy. For all these compounds, the radioracemization implies a shift of the ORD curves toward lower levels of specific optical rotation. The radioracemization degree ( R R) has been defined and calculated for all the compounds studied. It has been found that for radiation dose of 1 MGy the radioracemization degree is about 4.5% for the compound with the highest optical purity and reaches 7–8% for the less optically pure compounds, demonstrating that impurities can affect greatly the radioracemization. In contrast with the general radioracemization effect exerted by high-energy radiation on chiral molecules, β(−)pinene, β(+)pinene when irradiated show an increment of their specific optical rotation. This fact has been measured for the first time by ORD spectroscopy and the amplification degree of chirality can reach 1000% in the near UV. This phenomenon is due to the formation of a chiral polymer, poly- β-pinene, which forms a solution with the monomer enhancing its optical activity. The implications for the theories of the origin of life of such unexpected phenomenon are discussed briefly.

A LOV Story: The Signaling State of the Phot1 LOV2 Photocycle Involves Chromophore-Triggered Protein Structure Relaxation, As Probed by Far-UV Time-Resolved Optical Rotatory Dispersion Spectroscopy

Light-, oxygen-, or voltage-regulated (LOV1 and LOV2) domains bind flavin mononucleotide (FMN) and activate the phototropism photoreceptors phototropin 1 (phot1) and phototropin 2 (phot2) by using energy from absorbed blue light. Upon absorption of blue light, chromophore and protein conformational changes trigger the kinase domain for subsequent autophosphorylation and presumed downstream signal transduction. To date, the light-induced photocycle of the phot1 LOV2 protein is known to involve formation of a triplet flavin mononucleotide (FMN) chromophore followed by the appearance of a FMN adduct within 4 micros [Swartz, T. E., Corchnoy, S. B., Christie, J. M., Lewis, J. W., Szundi, I., Briggs, W. R., and Bogomolni, R. A. (2001) J. Biol. Chem. 276, 36493-36500] before thermal decay back to the dark state. To probe the mechanism by which the blue light information is relayed from the chromophore to the protein, nanosecond time-resolved optical rotatory dispersion (TRORD) spectroscopy, which is a direct probe of global secondary structure, was used to study the phot1 LOV2 protein in the far-UV region. These TRORD experiments reveal a previously unobserved intermediate species (tau approximately 90 micros) that is characterized by a FMN adduct chromophore and partially unfolded secondary structure (LOV390(S2)). This intermediate appears shortly after the formation of the FMN adduct. For LOV2, formation of a long-lived species that is ready to interact with a receptor domain for downstream signaling is much faster by comparison with formation of a similar species in other light-sensing proteins.

The earliest events in protein folding: a structural requirement for ultrafast folding in cytochrome C.

The folding dynamics of reduced cytochrome c (redcyt c) obtained from tuna heart, which contains a tryptophan residue at the site occupied by His33 in horse heart cytochrome c, was studied using nanosecond time-resolved optical rotatory dispersion spectroscopy. As observed previously for horse heart redcyt c, two time regimes were observed for secondary structure formation in tuna redcyt c: a fast (microseconds) and a slow (milliseconds) phase. However, the fast phase of tuna redcyt c folding was much slower and smaller in amplitude than the same phase in horse. The differences in the fast folding phases suggest that for horse heart redcyt c, the conformers that undergo the fastest observed folding have the His18-Fe-His33 heme configuration, which appears to be necessary, but not sufficient, to poise an unfolded chain conformation for fastest folding in redcyt c.

Earliest Events in Protein Folding: Submicrosecond Secondary Structure Formation in Reduced Cytochrome c,

Protein folding is uniquely characterized as a dynamic process by the tremendous conformational heterogeneity of the unfolded state. This conformational heterogeneity implies the possibility of significant kinetic heterogeneity in the folding dynamics, yet many folding reactions appear to proceed homogeneously, i.e., from unfolded conformations that are in equilibrium with each other and a transition state. At what point in time on the way to the folded state the kinetic heterogeneity of the unfolded state is lost to conformational equilibration has been an open question. We present evidence in this regard obtained from nanosecond far-UV optical rotatory dispersion spectroscopy of the fastest folding process observed in photoreduced cytochrome c, a secondary structure formation process proceeding on a submicrosecond time scale at high denaturant concentration (observed as a “burst” phase in millisecond stopped-flow circular dichroism studies). The kinetics of this fast folding process imply that it proceeds from a conformational ensemble that is not in equilibrium with the bulk of protein conformers during the time required to completely reduce the sample, ∼100 μs. We thus determine a lower limit on the time required for conformational equilibration of ∼10-4 s, in agreement with the previous estimate from time-resolved magnetic circular dichroism measurements on photolyzed cytochrome c-CO (Goldbeck, R. A.; Thomas, Y. G.; Chen, E.; Esquerra, R. M.; Kliger, D. S. Proc. Natl. Acad. Sci. 1999, 96, 2782−2787). Combined with an upper limit from stopped-flow measurements (Lyubovitsky, J. G.; Gray, H. B.; Winkler, J. R. J. Am. Chem. Soc. 2002, 124, 5481−5485), this allows us to bracket the conformational diffusion time, i.e., the time interval over which the kinetic heterogeneity of this protein's unfolded state is lost through conformational equilibration, within the range ∼10-4−10-3 s. This estimate of the conformational diffusion time implies that the earliest folding events, helix formation and possibly the collapse of extended conformations, proceed under an energy landscape regime wherein conformational diffusion is slow, whereas the final (milliseconds) folding phase proceeds along a classical kinetic pathway.

Nanosecond optical rotatory dispersion spectroscopy: application to photolyzed hemoglobin-CO kinetics

A standard technique for static optical rotatory dispersion (ORD) measurements is adapted to the measurement of ORD changes on a nanosecond (ns) time scale, giving approximately a million-fold improvement in time-resolution over conventional instrumentation. The technique described here is similar in principle to a technique recently developed for ns time-resolved circular dichroism (TRCD) spectroscopy, although the time-resolved optical rotatory dispersion (TRORD) technique requires fewer optical components. As with static ORD, TRORD measurements may be interpreted by empirical comparisons or may be transformed, via the Kramers-Kronig relations, to more easily interpreted TRCD spectra. TRORD can offer experimental advantages over TRCD in studying kinetic processes effecting changes in the chiral structures of biological molecules. In particular, the wider dispersion of ORD bands compared with the corresponding CD bands means that ORD information may often be obtained outside of absorption bands, a signal-to-noise advantage for multichannel measurements. Demonstration of the technique by its application to ns TRORD and the transform-calculated TRCD of carboxy-hemoglobin (Hb-CO) after laser photolysis is presented.

Preparation and study of magnesium deuteroporphyrin myoglobin and hemoglobin species

The myoglobin and hemoglobin species containing magnesium deuteroporphyrin have been prepared and studied by electronic, circular dichroism and optical rotatory dispersion spectroscopy. The results are compared with those obtained for corresponding magnesium protoporphyrin and magnesium mesoporphyrin complexes. In all cases the magnesium-apomyoglobin species show additional band splittings. These may arise directly from differences in the protein environment or indirectly through water coordination to magnesium which is facilitated by features of the myoglobin heme pocket but inhibited in the hemoglobin complexes. The availability of results for three different porphyrins enables a red shift of spectral bands, observed in particular for MgPP-Mb **, to be specifically associated with the presence of side-chain vinyl groups.

Spectral studies of magnesium porphyrin-apomyoglobin and apohemoglobin complexes

Complexes of magnesium protoporphyrin and magnesium mesoporphyrin with apomyoglobin and apohemoglobin have been prepared and studied by electronic, circular dichroism, and optical rotatory dispersion spectroscopy. The myoglobin complexes show prominent splittings and red shifts of the visible absorption bands, with respect to those for the hemoglobin analogs. Comparisons are made with other heme protein systems that display similar spectral features. Different reasons for the observations are considered, including multiple conformer formation, polarity effects of the protein environment, and the formation of six-coordinate magnesium species through H 2O coordination for the Mb complexes (the latter explanation being favored).

Observation of induced cotton effects for magnesium porphyrin-L-histidine species

Complexes of magnesium protoporphyrin and magnesium mesoporphyrin with L-histidine have been prepared in solution and studied by circular dichroism and optical rotatory dispersion spectroscopy. Large induced Cotton effects are observed, in particular for the Soret transitions. These are interpreted in terms of the formation of six-coordinate magnesium porphyrin-(L-histidine) 2 species where the asymmetry of the L-histidine groups is coupled to the magnesium porphyrin electronic transitions. The major, five-coordinate components of these solutions do not produce Cotton effects. These results, when compared with data for corresponding protein complexes, lend support for the coupled oscillator description of the origin of optical activity in hemoproteins. The observation of Cotton effects for porphyrin complexes of the type reported here may enable the existence of six-coordinate species to be more clearly established in instances where the major equilibrium product is a five-coordinate species.

Determination of enantiomeric homogeneity (optical purity) of cyclophosphamide by nuclear magnetic resonance spectroscopy.

The enantiomeric homogeneity of resolved samples of the chiral anticancer drug cyclophosphamide was evaluated directly by 1H and 31P nuclear magnetic resonance spectroscopy with the use of the optically active shift reagent tris-[3-(trifluoromethylhydroxymethylene)-d-camphorato]europlum(III). These measurements, in concert with optical rotatory dispersion spectroscopy, established that, for optically pure cyclophosphamide, [alphaD] = 2.3 +/- 0.2 degrees.

Residue analyses on 2-amino-4-phenylthiazole, a piscine anesthetic, in fishes. III. Metabolism in rainbow trout and carp.

The major biotransformation product of 2-amino-4-phenylthiazole in rainbow trout (salmo gairdneri irideus) was isolated from water following exposure of fish to the anesthetic. The isolated crystalline metabolite was shown by means of ultraviolet, infrared and optical rotatory dispersion spectroscopy and gas chromatography to be identical to 2-amino-4-pphenylthiazole-2-N-β-mono-D-glucopyranosiduronic acid, the major biotransformation product previously found in medaka (killifish, Oryzias latipes). The major biotransformation product in carp (Cyprinus carpio) was also identified as 2-amino-4-phenylthiazole-2-N-ββ-mono-D-glucopyranosiduronic acid by molecular sieve, thin layer and gas chromatography. Conversion of 2-amino-4-phenylthiazole to the N-glucuronyl conjugate was 8 and 12%, respectively, in rainbow trout and carp as shown by thin layer chromatography of extracts from fish treated with 3H-labeled anesthetic. In addition, a minor metabolite of the anesthetic in rainbow trout was isolated as a yellowish-white crystalline powder and identified as 2-acetamido-4-(4'-hydroxyphenyl)-thiazole by means of ultraviolet and infrared spectroscopy, NMP and mass spectrometry. Chromatography suggested that this same metabolite was also formed in carp but in concentrations to low for isolation and definitive identification.

Model polytripeptides for collagen

Poly[ala-glu(OEt)-gly], poly[glu(OEt)-gly-gly], and poly [ala-gly-gly] have been prepared from their respective tripeptide activated esters. Preliminary characterization by X-ray diffraction, infrared, circular dichroism, and optical rotatory dispersion spectroscopy indicate that poly[ala-glu(OEt)-gly], as obtained from an aqueous suspension or film cast from formic acid, is in an anti-parallel β sheet while the other two polymers are obtained in a random conformation from their aqueous solutions.