Molecular alignment techniques for polarised spectroscopy

The alignment of the bonds in a molecule or the orientations that individual molecules take with respect to each other in a macromolecular structure is of significant importance to understanding the molecular mechanics and the nature of their interactions with their environment. There are a variety of different techniques available to investigate this matter.1,2 In comparison with other widely used techniques such as crystallography and fibre diffraction, linear dichroism (LD) is simpler to apply, less time-consuming and also gives useful information about the transition polarisations.3–5 In the preliminary stages of this work, we tried to optimise the technique in order to collect data of higher quality than had been previously possible for a wide range of different types of molecules with different characteristics. We invented a new method of orienting polar and slightly-polar molecules by changing the surface of polyethylene (PE) films to have oxygen groups on them,making PEOX. Then we tried to combine the improved orientation and LD spectroscopy techniques with fluorimetry to make fluorescence detected linear dichroism (FDLD) to increase the sensitivity and selectivity of our experiments. As all UV-visible spectroscopy techniques, including LD and FDLD, are limited by the small number of UV-active functional groups (chromophores) in molecules, we then turned to vibrational spectroscopy techniques. In particular, we have worked on a new type of polarised Raman spectroscopy - Raman Linear Difference (RLD) spectroscopy. The first RLD spectra had been published in 2011.6 In this work, we used our new PEOX films and the new Raman spectroscopic technique to study the alignment of molecules in the vibrational realm. After recording high resolution polarised vibrational data for a set of small molecules using a Raman microscopy system, we designed a new polarised Raman spectrometer which was built in the final stages of this project. We evaluated the newly built instrument by repeating polarised Raman experiments we had performed on the microscope system and found that the results were a significant improvement

Oxidized polyethylene films for orienting polar molecules for linear dichroism spectroscopy

Stretched polyethylene (PE) films have been used to orient small molecules for decades by depositing solutions on their surface and allowing the solvent to evaporate leaving the analyte absorbed on the polymer film. However, the non-polar hydrophobic nature of PE is an obstacle to aligning polar molecules and biological samples. In this work PE film was treated with oxygen plasma in order to increase surface hydrophilicity. Different treatment conditions were evaluated using contact angle measurement and X-ray photoelectron spectroscopy. Treated PE (PEOX) films are shown to be able to align molecules of different polarities including progesterone, 1-pyrenecarboxaldehyde, 4′,6-diamidino-2-phenylindole (DAPI) and anthracene. The degree of alignment of each molecule was studied by running series of linear dichroism (LD) experiments and the polarizations of electronic transition moments were determined. For the first time optimal conditions (such as stretching factor and concentration of the sample) for stretched film LD were determined. PEOX aligning ability was compared to that of normal PE films. Progesterone showed a slightly better alignment on PEOX than PE. 1-Pyrenecarboxaldehyde oriented differently on the two different films which enabled transition moment assignment for this low symmetry molecule. DAPI (which does not align on PE) aligned well on PEOX and enabled us to obtain better LD data than had previously been collected with polyvinyl alcohol. Anthracene alignment and formation of dimers and higher order structures were studied in much more detail than previously possible, showing a variety of assemblies on PE and PEOX films

Spectroscopic signatures of an Fmoc-tetrapeptide, Fmoc and fluorene

The hydrophobic Fmoc [N-(fluorenyl)-9-methoxycarbonyl] unit is often conjugated to peptides to confer amphiphilicity on the molecules, and to introduce the potential for aromatic stacking interactions. In this paper we report spectroscopic data for fluorene, Fmoc and Fmoc conjugated to a small peptide (GRDS in this case) to enable interpretation of spectroscopic data collected on complex structures that can be created with Fmoc–peptide molecules. We report absorbance data (ultra violet and infrared), circular dichroism, linear dichroism (both film and Couette flow orientation) and Raman spectra. Orientations of the Fmoc chromophores in the linear dichroism studies are deduced.5 page(s

Rational design, synthesis, and characterization of a solid Δ9-tetrahydrocannabinol (THC) nanoformulation suitable for “microdosing” applications

Background: This paper highlights the formulation of a solid THC-loaded ingestible prepared from pure THC distillate. A THC ethanol-assisted cannabinoid nanoemulsion (EACNE) was created without the need for specialized emulsification equipment such as a high-pressure homogenizer or a microfluidizer. Stress-testing was performed on the EACNE to evaluate its chemical and colloidal stability under the influence of different environmental factors, encompassing both physical and chemical stressors. Subsequently, the EACNE was converted to a solid powdery material while still retaining its THC potency, and suited for “microdosing” applications. Methods: An ethanol-assisted emulsification method was used to generate a THC nanoemulsion. The EACNE was fully characterized, imaged, and subjected to stress-tests. The EACNE was then mixed with a solid matrix material post facto and lyophilized to create a solid ingestible substance. Upon ball-milling, a dense powdery material was obtained. Flow properties and thermal properties of this material were recorded. Potency of the material was evaluated in triplicate using HPLC and correlated with the potency of the starting EACNE. Results: EACNE had an average lipid droplet size of ca. 190 nm, with a polydispersity index (PDI) of 0.15, and an average droplet zeta potential of -49±10 mV. The nanoemulsion was colloidally stable for at least 6 weeks, with no meaningful change in cannabinoid potency over the experimental period, as determined by HPLC analysis. The EACNE remained stable when subjected to physical stresses such as heat, freeze/thaw cycles, carbonation, dilution to beverage concentrations, high sucrose concentrations, and a pH range between 5-8. The effect of undesirable events during the lyophilization of the EACNE were minimized by ball-milling the resulting solid. The microencapsulated EACNE demonstrated limited free-flowing behaviour but was freely redispersible in water without any visible phase separation. Conclusions: A solvent-mediated emulsification protocol creates a THC-loaded nanoemulsion that can subsequently be converted to a water-soluble powder. These materials are particularly suited for THC “microdosing”, a practice that might decouple the health benefits of THC from its psychotropic effects

Experimental and Theoretical Polarized Raman Linear Difference Spectroscopy of Small Molecules with a New Alignment Method Using Stretched Polyethylene Film

This paper reports the development of the new technique of Raman linear difference (RLD) spectroscopy and its application to small molecules: anthracene and nucleotides adenosine-5′-monophosphate, thymidine-5′-monophosphate, guanosine-5′-monophosphate, and cytidine-5′-monophosphate. In this work we also present a new alignment method for Raman spectroscopy where stretched polyethylene films are used as the matrix. Raman spectra using light polarized along the orientation direction and perpendicular to it are reported. The polyethylene (PE) film spectra are consistent with powder samples and films deposited on quartz. RLD spectra determined from the difference of the parallel and perpendicular polarized light Raman spectra are also reported. The equations describing RLD are derived, and RLD spectra of anthracene and thymidine are calculated from these equations using Density Functional Theory and assuming perfect orientation of the samples. Because of the wealth of spectroscopic information in the vibrational spectra of biomolecules together with our ability to calculate spectra as a function of orientation, we conclude that RLD has the potential to provide structural information for biological samples that currently cannot be extracted from any other method

Experimental and theoretical polarized Raman linear difference spectroscopy of small molecules with a new alignment method using stretched polyethylene film

This paper reports the development of the new technique of Raman linear difference (RLD) spectroscopy and its application to small molecules: anthracene and nucleotides adenosine-5′-monophosphate, thymidine-5′-monophosphate, guanosine-5′-monophosphate, and cytidine-5′-monophosphate. In this work we also present a new alignment method for Raman spectroscopy where stretched polyethylene films are used as the matrix. Raman spectra using light polarized along the orientation direction and perpendicular to it are reported. The polyethylene (PE) film spectra are consistent with powder samples and films deposited on quartz. RLD spectra determined from the difference of the parallel and perpendicular polarized light Raman spectra are also reported. The equations describing RLD are derived, and RLD spectra of anthracene and thymidine are calculated from these equations using Density Functional Theory and assuming perfect orientation of the samples. Because of the wealth of spectroscopic information in the vibrational spectra of biomolecules together with our ability to calculate spectra as a function of orientation, we conclude that RLD has the potential to provide structural information for biological samples that currently cannot be extracted from any other method