Raman spectroscopy for the detection of AGEs/ALEs.

Raman spectroscopy is a noninvasive, nondestructive tool for capturing multiplexed biochemical information across diverse molecular species including proteins, lipids, DNA, and mineralizations. Based on light scattering from molecules, cells, and tissues, it is possible to detect molecular fingerprints and discriminate between subtly different members of each biochemical class. Raman spectroscopy is ideal for detecting perturbations from the expected molecular structure such as those occurring during senescence and the modification of long-lived proteins by metabolic intermediates as we age. Here, we describe the sample preparation, data acquisition, signal processing, data analysis and interpretation involved in using Raman spectroscopy for detecting age-related protein modifications in complex biological tissues.

Unexpected reversible pyrazine based methylation in a Ru(II) complex bearing a pyrazin-2'-yl-1,2,4-triazolato ligand and its effect on acid/base and photophysical properties.

The regioselective methylation of a ruthenium polypyridyl complex bearing both a 1,2,4-triazolato and a pyrazine moiety is reported. In contrast to previous studies in which methylation of the 1,2,4-triazolato ring was observed, in the present system methylation takes place exclusively at the non-coordinated nitrogen of the pyrazine ring. The monomethylation is confirmed by (1)H NMR spectroscopy and ESI-MS and the electronic properties of the methylated complexes are studied by UV/vis absorption, emission, surface enhanced, resonance and transient resonance Raman spectroscopy. Ligand deuteriation is used to simplify the (1)H NMR spectra and to assign definitively the Raman spectra. Acid-base studies show that the triazolato ring of the N-methylated complexes can be protonated at low pH and that at high pH the N-methyl group can be deprotonated reversibly. Furthermore it is shown that under conditions where the methyl group is deprotonated, demethylation occurs to recover the initial complex.

Profiling retinal biochemistry in the MPDZ mutant retinal dysplasia and degeneration chick: a model of human RP and LCA.

PURPOSE: Raman microscopy, a rapid nondestructive technique that profiles the composition of biological samples, was used to characterize retinal biochemistry in the retinal dysplasia and degeneration (rdd) and wild-type (wt) chick retina during retinogenesis and at hatching. METHODS: Embryonic day (E)13 and posthatch day (P)1 rdd and wt retinal cross-sections (n = 3 of each line at each age) were profiled using 633 helium-neon laser excitation. The biochemical composition was determined using computational analysis of the Raman spectra. In parallel histology, TUNEL and glial fibrillary acidic protein (GFAP) immunostaining were used to visualize retinal dysfunction. RESULTS: Principal component (PC) analysis of the Raman spectra identified 50 major biochemical profiles, but only PCs that made significant contributions to variation within rdd and wt retina were mapped. These significant PCs were shown to arise from DNA, various fatty acids, melanin, and a number of proteins. Distinct patterns of GFAP immunostaining and a larger population of TUNEL-positive nuclei were observed in the rdd versus wt retina. CONCLUSIONS: This study has demonstrated that Raman microscopy can discriminate between major retinal biomolecules, thus providing an unbiased account of how their composition varies due to the impact of the MPDZ null mutation in the rdd chick relative to expression in the normal wt retina.

Sclera as a surrogate marker for determining AGE-modifications in Bruch's membrane using a Raman spectroscopy-based index of aging.

PURPOSE: Raman spectroscopy is an effective probe of advanced glycation end products (AGEs) in Bruch's membrane. However, because it is the outermost layer of the retina, this extracellular matrix is difficult to analyze in vivo with current technology. The sclera shares many compositional characteristics with Bruch's membrane, but it is much easier to access for in vivo Raman analysis. This study investigated whether sclera could act as a surrogate tissue for Raman-based investigation of pathogenic AGEs in Bruch's membrane. METHODS: Human sclera and Bruch's membrane were dissected from postmortem eyes (n = 67) across a wide age range (33-92 years) and were probed by Raman spectroscopy. The biochemical composition, AGEs, and their age-related trends were determined from data reduction of the Raman spectra and compared for the two tissues. RESULTS: Raman microscopy demonstrated that Bruch's membrane and sclera are composed of a similar range of biomolecules but with distinct relative quantities, such as in the heme/collagen and the elastin/collagen ratios. Both tissues accumulated AGEs, and these correlated with chronological age (R(2) = 0.824 and R(2) = 0.717 for sclera and Bruch's membrane, respectively). The sclera accumulated AGE adducts at a lower rate than Bruch's membrane, and the models of overall age-related changes exhibited a lower rate (one-fourth that of Bruch's membrane) but a significant increase with age (P < 0.05). CONCLUSIONS: The results suggest that the sclera is a viable surrogate marker for estimating AGE accumulation in Bruch's membrane and for reliably predicting chronological age. These findings also suggest that sclera could be a useful target tissue for future patient-based, Raman spectroscopy studies.

Multiplex analysis of age-related protein and lipid modifications in human Bruch's membrane.

Aging of the human retina is characterized by progressive pathology, which can lead to vision loss. This progression is believed to involve reactive metabolic intermediates reacting with constituents of Bruch's membrane, significantly altering its physiochemical nature and function. We aimed to replace a myriad of techniques following these changes with one, Raman spectroscopy. We used multiplexed Raman spectroscopy to analyze the age-related changes in 7 proteins, 3 lipids, and 8 advanced glycation/lipoxidation endproducts (AGEs/ALEs) in 63 postmortem human donors. We provided an important database for Raman spectra from a broad range of AGEs and ALEs, each with a characteristic fingerprint. Many of these adducts were shown for the first time in human Bruch's membrane and are significantly associated with aging. The study also introduced the previously unreported up-regulation of heme during aging of Bruch's membrane, which is associated with AGE/ALE formation. Selection of donors ranged from ages 32 to 92 yr. We demonstrated that Raman spectroscopy can identify and quantify age-related changes in a single nondestructive measurement, with potential to measure age-related changes in vivo. We present the first directly recorded evidence of the key role of heme in AGE/ALE formation.

Fe(II) complex with the octadentate btpa ligand: a DFT study on a spin-crossover system that reveals two distinct high-spin states.

Density functional theory calculations (DFT) were performed for the spin-crossover system [Fe(btpa)]2+ (btpa = N,N,N',N'-tetrakis(2-pyridylmethyl)-6,6'-bis(aminomethyl)-2,2'-bipyridine), and for the predominantly low-spin [Fe(b(bdpa))]2+ complex (in the solid state) (b(bdpa) = N,N'-bis(benzyl)-N,N'-bis(2-pyridylmethyl)-6,6'-bis(aminomethyl)-2,2'-bipyridine). The calculations confirmed that the former complex exhibits two high-spin isomers of the complexes, i.e. with C1 quasi hepta-coordinated (long-lived isomer) and C2 hexa-coordinated (short-lived isomer) structures that have been suggested previously based on time-resolved Raman and flash photolysis experiments. Application of B3LYP and B3LYP* functionals together with the CEP-31G basis yielded reasonable estimates of electronic energies (E(el) = E(el)(HS) - E(el)(LS)) for both isomers (calculated E(el) of ca. 24 and 31 kJ mol(-1) for long- and short-lived HS isomers, respectively, vs. the experimentally determined value of 27.5 kJ mol(-1)). Further calculations yielded the electronic structure of the low-spin isomer together with lowest lying singlet and triplet excited states of the [Fe(btpa)]2+ as well as the energy profile of the C2 <--> C1 isomerisation pathway for the high-spin [Fe(btpa)]2+ within the framework of the QST (quadratic synchronous transit) approach. The data obtained are discussed in relation to the observed ultrafast intersystem crossing in Fe(II) polypyridine complexes. The importance of ligand strain in relation to the destabilisation of the low-spin isomers is also discussed. In that context, calculations for a further 15 Fe(II) spin-crossover complexes of hexa-coordinating nitrogen-donor ligands have shown that the LS-HS conversion is associated with a release of ligand stress of 95 +/-16 kJ mol(-1), on average. On the basis of the calculations presented in this paper we propose that octahedral high-spin d-6 isomers are far more elastic regarding the angular distortions (equatorial and meridional strain, i.e. the declination of cis- and trans-L-M-L angle from the regular values of 90 and 180 degrees, respectively) than their low-spin counterparts.

Ex vivo diagnosis of lung cancer using a Raman miniprobe.

Lung cancer is the most common cause of cancer death. The conventional method of confirming the diagnosis is bronchoscopy, inspecting the airways of the patient with a fiber optic endoscope. A number of studies have shown that Raman spectroscopy can diagnose lung cancer in vitro. In this study, Raman spectra were obtained from ex vivo normal and malignant lung tissue using a minifiber optic Raman probe suitable for insertion into the working channel of a bronchoscope. Shifted subtracted Raman spectroscopy was used to reduce the fluorescence from the lung tissue. Using principal component analysis with a leave-one-out analysis, the tissues were classified accurately. This novel technique has the potential to obtain Raman spectra from tumors from patients with lung cancer in vivo.

Photophysical and electrochemical properties of meso-substituted thien-2-yl Zn(II) porphyrins.

The influence of the thiophene ring on the ground and excited state properties of the porphyrin ring is investigated, when substituted at the meso-position. A series of mono-, di-, tri-, and tetra- meso-thien-2-yl porphyrins are studied and discussed with respect to the reference compounds zinc(II)-5,10,15,20-tetra(thien-2'-yl)porphyrin ( 1a) and zinc(II)-5,10,15,20-tetraphenylporphyrin (ZnTPP). The extended conjugated system zinc(II)-5-(5'-(5''-ethynyl-2''-thiophenecarboxaldehyde)thien-2'-yl)-10,15,20-triphenylporphyrin ( 4d) is also studied and shows enhanced charge transfer character due to the presence of the terminal aldehyde accepting group. A detailed analysis of ground and excited state UV-vis absorption, steady-state and time-resolved fluorescence, laser flash photolysis, and electrochemical data all point toward substantial electronic communication between the central Zn(II) porphyrin ring and the meso-thien-2-yl substituents, which is evident from excited state charge transfer character.

Sub-picosecond time resolved infrared spectroscopy of high-spin state formation in Fe(ii) spin crossover complexes.

The photoinduced low-spin (S = 0) to high-spin (S = 2) transition of the iron(ii) spin-crossover systems [Fe(btpa)](PF(6))(2) and [Fe(b(bdpa))](PF(6))(2) in solution have been studied for the first time by means of ultrafast transient infrared spectroscopy at room temperature. Negative and positive infrared difference bands between 1000 and 1065 cm(-1) that appear within the instrumental system response time of 350 fs after excitation at 387 nm display the formation of the vibrationally unrelaxed and hot high-spin (5)T(2) state. Vibrational relaxation is observed and characterized by the time constants 9.4 +/- 0.7 ps for [Fe(btpa)](PF(6))(2)/acetone and 12.7 +/- 0.7 ps for both [Fe(btpa)](PF(6))(2)/acetonitrile and [Fe(b(bdpa)](PF(6))(2)/acetonitrile. Vibrational analysis has been performed via DFT calculations of the low-spin and high-spin state normal modes of both compounds as well as their respective infrared absorption cross sections. The simulated infrared difference spectra are dominated by an increase of the absorption cross section upon high-spin state formation in accordance with the experimental infrared spectra.

Advanced glycation as a basis for understanding retinal aging and noninvasive risk prediction.

The retina is exquisitely sensitive to age-related processes, and, in many cases, these can precipitate progressive and profound loss of vision. Many asymptomatic abnormalities that accrue in the outer retina as we get older can serve as a sinister preamble to age-related macular degeneration (AMD). This condition remains the leading cause of irreversible blindness in industrialized countries, but its precise pathogenesis has yet to be completely elucidated. Over recent years, increasing evidence has suggested that the accumulation of advanced glycation end products (AGEs) and activation of the receptor for AGEs in the outer retina could play a significant role in the initiation and progression of AMD. The current review outlines this evidence and indicates how products of Maillard chemistry could be used as robust markers for outer retinal aging and susceptibility to AMD. The utility of Raman spectroscopy to measure AGE adducts in human tissues is presented. The methodology reinforces the association between AGE formation and retinal aging and provides exciting possibilities for assessing these pathogenic agents in the living eye and, perhaps, for providing a valuable index for AMD susceptibility.

The early picosecond photophysics of Ru(II) polypyridyl complexes: a tale of two timescales.

The early picosecond time scale excited-state dynamics of the paradigm tris(2,2'-bipyridyl)Ruthenium(II) ([Ru(bpy)(3)](2+)) and related complexes have been examined by picosecond Kerr-gated time-resolved resonance Raman (ps-TR(3)) spectroscopy. The evolution of the signature Raman bands of the lowest thermally equilibrated excited (THEXI) state under two-color pump/probe conditions show that this state is not fully populated within several hundred femtoseconds as proposed previously but rather only within the first 20 ps following excitation. In addition to an emission observed within the instrument rise time (τ < 3 ps), the early picosecond dynamics are characterized by a rise in the intensity of the Raman marker bands of the THEXI-(3)MLCT state, a rise time which, within experimental uncertainty, is not influenced by either partial or complete ligand deuteriation or the presence of ligands other than bpy, as in the heteroleptic complexes [Ru(bpy)(2)(L1)](+) and [Ru(bpy)(2)(Hdcb)](+) (where H(2)dcb is 4,4'-dicarboxy-2,2'-bipyridine and L1 is 2,-(5'-phenyl-4'-[1,2,4]triazole-3'-yl)pyridine). Overall, although the results obtained in the present study are consistent with those obtained from examination of this paradigm complex on the femtosecond timescale, regarding initial formation of the vibrationally hot (3)MLCT state by ISC from the singlet Franck-Condon state, the observation that the THEXI-(3)MLCT state reaches thermal equilibration over a much longer time period than previously suggested warrants a re-examination of views concerning the rapidity with which thermal equilibration of transition metal complex excited states takes place.

Raman microscopy of porcine inner retinal layers from the area centralis.

PURPOSE: To characterize the Raman spectra of porcine inner retinal layers, specifically, the inner nuclear, inner plexiform, ganglion cell, and nerve fiber layers. METHODS: Raman microscopy was employed at three excitation wavelengths, 785, 633, and 514 nm to measure Raman spectra in a high resolution grid across the inner layers of 4% paraformaldehyde cryoprotected porcine retina. Multivariate statistics were used to summarize the principal spectral signals within those layers and to map the distribution of each of those signals. RESULTS: The detected Raman scattering was dominated by a signal characteristic of the protein population present in each layer. As expected, a significant nucleotide contribution was observed in the inner nuclear layer, while the inner plexiform layer displayed a minor contribution from fatty acid based lipid, which would be characteristic of the axonal and synaptic connection resident in this layer. Blood vessels were readily characterized by their distinct heme-derived spectral signature, which increased at 633 and 514 nm excitation compared to 785 nm. Discrete isolated nucleotide signals were identified in the ganglion cell layer, while the nerve fiber layer exhibited a homogenous profile, which is indicative of its broadly uniform axonal and cytoplasmic Muller cell components. CONCLUSIONS: The present study demonstrated the potential of Raman microscopy as a tool to study the biochemical composition of pathologically normal retina. Specifically, the method allowed a unique method of analyzing the network of neurons involved in relaying information from the photoreceptor population to the ganglion cell derived nerve fiber layer. The study has demonstrated the ability of Raman microscopy to generate simultaneously information on a range of specific biochemical entities within the stratified normal retina.

Confocal Raman microscopy can quantify advanced glycation end product (AGE) modifications in Bruch's membrane leading to accurate, nondestructive prediction of ocular aging.

The modification of proteins by nonenzymatic glycation leading to accumulation of advanced glycation end products (AGEs) is a well-established phenomenon of aging. In the eyes of elderly patients, these adducts have been observed in retinal pigment epithelium (RPE), particularly within the underlying pentalaminar substrate known as Bruch's membrane. AGEs have also been localized to age-related subcellular deposits (drusen and basal laminar deposits) and are thought to play a pathogenic role in progression of the major sight-threatening condition known as age-related macular degeneration (AMD). The current study has quantified AGEs in Bruch's membrane from postmortem eyes and established age-related correlations. In particular, we investigated the potential of confocal Raman microscopy to identify and quantify AGEs in Bruch's membrane in a nondestructive, analytical fashion. Bruch's membrane and the innermost layers of the underlying choroid (BM-Ch) were dissected from fresh postmortem eye-cups (n=56). AGE adducts were quantified from homogenized tissue using reverse-phase HPLC and GC/MS in combination with immunohistochemistry. For parallel Raman analysis, BM-Ch was flat-mounted on slides and evaluated using a Raman confocal microscope and spectra analyzed by a range of statistical approaches. Quantitative analysis showed that the AGEs pentosidine, carboxymethyllysine (CML), and carboxyethyllysine (CEL) occurred at significantly higher levels in BM-Ch with age (P<0.05-0.01). Defined Raman spectral "fingerprints" were identified for various AGEs and these were observed in the clinical samples using confocal Raman microscopy. The Raman data set successfully modeled AGEs and not only provided quantitative data that compared with conventional analytical approaches, but also provided new and complementary information via a nondestructive approach with high spatial resolution. It was shown that the Raman approach could be used to predict chronological age of the clinical samples (P<0.001) and a difference in the Raman spectra between genders was highly significant (P<0.000001). With further development, this Raman-based approach has the potential for noninvasive examination of AGE adducts in living eyes and ultimately to assess their precise pathogenic role in age-related diseases.

Investigation into the subambient behavior of aqueous mannitol solutions using temperature-controlled Raman microscopy.

The aim was twofold; to demonstrate the ability of temperature-controlled Raman microscopy (TRM) to locate mannitol within a frozen system and determine its form; to investigate the annealing behavior of mannitol solutions at -30 degrees C. The different polymorphic forms of anhydrous mannitol as well as the hemihydrate and amorphous form were prepared and characterized using crystal or powder X-ray diffractometry (XRD) as appropriate and Raman microscopy. Mannitol solutions (3% w/v) were cooled before annealing at -30 degrees C. TRM was used to map the frozen systems during annealing and was able to differentiate between the different forms of mannitol and revealed the location of both beta and delta polymorphic forms within the structure of the frozen material for the first time. TRM also confirmed that the crystalline mannitol is preferentially deposited at the edge of the frozen drop, forming a rim that thickens upon annealing. While there is no preference for one form initially, the study has revealed that the mannitol preferentially transforms to the beta form with time. TRM has enabled observation of spatially resolved behavior of mannitol during the annealing process for the first time. The technique has clear potential for studying other crystallization processes, with particular advantage for frozen systems.

Resonance Raman and lifetime studies on regioselectively deuteriated ruthenium(II) polypyridyl complexes.

Two series of ruthenium(II) polypyridyl complexes [Ru(bipy)(2)(phpytr)](+) and [Ru(bipy)(2)(phpztr)](+) (where Hphpytr = 2-(5-phenyl-1H-[1,2,4]triazol-3-yl)-pyridine and Hphpztr = 2-(5-phenyl-1H-[1,2,4]triazol-3-yl)-pyrazine) are examined by electrochemistry, UV/Vis, emission, resonance Raman, transient resonance Raman and transient absorption spectroscopy, in order to obtain a more comprehensive understanding of their excited state electronic properties. The interpretation of the results obtained is facilitated by the availability of several isotopologues of each of the complexes examined. For the pyridine-1,2,4-triazolato based complex the lowest emissive excited state is exclusively bipy based, however, for the pyrazine based complexes excited state localisation on particular ligands shows considerable solvent and pH dependency.

The effects of ligand substitution and deuteriation on the spectroscopic and photophysical properties of [Ru(LL)(CN)4]2- complexes.

The spectroscopic characterisation of a series of [Ru(LL)(CN)(4)](2-) complexes, where LL = 1,10-phenanthroline (phen) and its methyl- and phenyl-substituted derivatives and several deuteriated isotopologues are reported. The optical and vibrational properties of these complexes are compared with that of the series of 2,2'-bipyridine (bipy) derivatives and analogous [Ru(LL)(3)](2+) complexes. It has been demonstrated that substitution at the 4,4' positions of bipy and 4,7-positions of phen by electron donating (CH(3)) and withdrawing (C(6)H(5), COO(-)) groups induces a pronounced blue and red shift, respectively, in the lowest energy (1)MLCT absorption band of [Ru(LL)(CN)(4)](2-). The energy of the emission originating from the (3)MLCT excited state is found to be dependant on the nature of the vibrational modes of the aromatic rings and the electron donating and/or withdrawing properties of the substituents. Single-mode Franck-Condon analysis indicates that methyl substitution leads to a significant increase in the Huang-Rhys factor (S(M)), while phenyl substitution results in a decrease in S(M) for both series (bipy and phen) of complexes. The rate of non-radiative (k(nr)) and radiative decay (k(ph)) to the ground state and the parameters of thermally activated deactivation pathways (A(4th), DeltaE(4th) and A(dd), DeltaE(dd)) were estimated from the temperature dependence of luminescence quantum yields and lifetimes. It has been demonstrated that the non-radiative decay rate and the temperature dependent decay processes are more efficient for bipy complexes than for phen derivatives due to the rigidity of the latter ligand.

The use of Raman microscopy to determine and localize vitamin E in biological samples.

Alpha-tocopherol (aT), the predominant form of vitamin E in mammals, is thought to prevent oxidation of polyunsaturated fatty acids. In the lung, aT is perceived to be accumulated in alveolar type II cells and secreted together with surfactant into the epithelial lining fluid. Conventionally, determination of aT and related compounds requires extraction with organic solvents. This study describes a new method to determine and image the distribution of aT and related compounds within cells and tissue sections using the light-scattering technique of Raman microscopy to enable high spatial as well as spectral resolution. This study compared the nondestructive analysis by Raman microscopy of vitamin E, in particular aT, in biological samples with data obtained using conventional HPLC analysis. Raman spectra were acquired at spatial resolutions of 2-0.8 microm. Multivariate analysis techniques were used for analyses and construction of corresponding maps showing the distribution of aT, alpha-tocopherol quinone (aTQ), and other constituents (hemes, proteins, DNA, and surfactant lipids). A combination of images enabled identification of colocalized constituents (heme/aTQ and aT/surfactant lipids). Our data demonstrate the ability of Raman microscopy to discriminate between different tocopherols and oxidation products in biological specimens without sample destruction. By enabling the visualization of lipid-protein interactions, Raman microscopy offers a novel method of investigating biological characterization of lipid-soluble compounds, including those that may be embedded in biological membranes such as aT.

Vibrational spectrum of the spin crossover complex [Fe(phen)(2)(NCS)(2)] studied by IR and Raman spectroscopy, nuclear inelastic scattering and DFT calculations.

The vibrational modes of the low-spin and high-spin isomers of the spin crossover complex [Fe(phen)(2)(NCS)(2)] (phen = 1,10-phenanthroline) have been measured by IR and Raman spectroscopy and by nuclear inelastic scattering. The vibrational frequencies and normal modes and the IR and Raman intensities have been calculated by density functional methods. The vibrational entropy difference between the two isomers, DeltaS(vib), which is--together with the electronic entropy difference DeltaS(el)--the driving force for the spin-transition, has been determined from the measured and from the calculated frequencies. The calculated difference (DeltaS(vib) = 57-70 J mol(-1) K(-1), depending on the method) is in qualitative agreement with experimental values (20-36 J mol(-1) K(-1)). Only the low energy vibrational modes (20% of the 147 modes of the free molecule) contribute to the entropy difference and about three quarters of the vibrational entropy difference are due to the 15 modes of the central FeN(6) octahedron.

Raman microscopy for the chemometric analysis of tumor cells.

Raman spectroscopy is recognized as a tool for chemometric analysis of biological materials due to the high information content relating to specific physical and chemical qualities of the sample. Thirty cells belonging to two different prostatic cell lines, PNT1A (immortalized normal prostate cell line) and LNCaP (malignant cell line derived from prostate metastases), were mapped using Raman microscopy. A range of spectral preprocessing methods (partial least-squares discriminant analyses (PLSDAs), principal component analyses (PCAs), and adjacent band ratios (ABRs)) were compared for input into linear discriminant analysis to model and classify the two cell lines. PLSDA and ABR were able to correctly classify 100% of cells into benign and malignant groups, while PLSDA correctly classified a greater proportion of individual spectra. PCA was used to image the distribution of various biochemicals inside each cell and confirm differences in composition/distribution between benign and malignant cell lines. This study has demonstrated that PLSDAs and ABRs of Raman data can identify subtle differences between benign and malignant prostatic cells in vitro.

Raman scattering and FT-IR spectroscopic studies on dithienylethene switches--towards non-destructive optical readout.

The non-destructive readout of photochromic memory materials based on the dithienylethene unit both by IR spectroscopy and Raman scattering is explored. A representative series of C5-substituted thienyl hexahydro- and hexafluoro-cyclopentene based photochromes was investigated to explore the effect and potential usefulness of substitution for the development of multicomponent memory materials. The effect of the deposition method on the photochemistry of solid materials containing photochromic dithienylcyclopentene switches was also explored. Photoconversion in the solid state to the closed form was found to be low when starting from the open form, but, in contrast, ring opening to the open state from the closed form was found to be complete. The effect was found to be due to inner filter rather than conformational phenomena. Characteristic vibrational bands for the central dithienyl core are assigned and a comparison made of the vibrational spectroscopic properties of the perhydro- and perfluoro switches. The data enable the determination of the photoconversion achievable in the solid state as well as some assessment of the influence of the deposition method on the photoconversion. The potential of Raman spectroscopy as a method of achieving non-destructive optical readout is demonstrated through the large differences in absolute Raman scattering intensity between the open and closed states, when monitored at wavelengths which do not result in photochemical ring opening.