Single molecule spectroscopy of plasmonically enhanced single and multi chromophoric systems

Evans, Owen 2025. Single molecule spectroscopy of plasmonically enhanced single and multi chromophoric systems. PhD Thesis, Cardiff University. Item availability restricted.

Preview	PDF - Accepted Post-Print Version Available under License Creative Commons Attribution Non-commercial No Derivatives. Download (74MB) | Preview
	PDF (Cardiff University Electronic Publication Form) - Supplemental Material Restricted to Repository staff only Download (159kB)

Abstract

This thesis primarily explores the low-temperature excited state dynamics of single molecules by utilising plasmonic nanoparticles to enhance emission in single-molecule spectroscopy. We use this method of investigation to study single fluorophores and single multichromophoric systems, and observe sharp spectral features that are sig nificant compared to the background, allowing for an in-depth analysis of the spectral lines. The utilisation of plasmonics in single-molecule spectroscopy is an emerging f ield as their unique and tunable optical response is becoming increasingly recog nised as a tool for enhanced light-matter interactions. In our studies, we use gold nanorods (GNR) and silver nanocubes (SNC). We present a computational study of the optical properties of these plasmonic nanoparticles by examining how their physical characteristics, configurations, and local environment influence the optical response. This investigation supports the other studies in this thesis, providing in sight into the requirements for strong plasmonic enhancement of emission for the molecules investigated. We present a low-temperature single-molecule spectroscopy study of dibenzoterrylene (DBT) embedded in anthracene. Firstly, we detail a sam ple preparation procedure to embed DBT molecules in anthracene nanocrystals that form around GNRs that act as nucleation sites. By developing this novel sample preparation procedure to localise DBT molecules to the ’hotpots’ of GNRs and sim ulating the optical response with specific sample properties, we are able to meet the sample requirements for plasmonic enhancement of single DBT molecules and observe enhanced emission in their low-temperature spectra. We conduct spectral time-trace measurements that allow the excited state dynamics to be studied by analysing the spectral lines. The emission observed is at higher energies than pub lished in the literature, and we hypothesise that this is caused by strain on the anthracene crystal as a result of crystal distortion from the gold nanorod. More over, by analysing the emission lineshapes and spectral wandering, we categorise the emission into enhanced fluorescence and surface-enhanced Raman emission. We examine the Debye-Waller factor from the zero-phonon line and phonon-sideband in the fluorescence emission and conduct non-negative matrix factorisation on the Raman data to further analyse the data. DBT is a well-studied molecule after gaining interest for its application as a single-photon source in quantum comput ing. In this study, we demonstrate the utility of plasmonic nanoparticles to gain enhanced spectral data from DBT, which opens avenues for more in-depth stud ies of this molecule and its integration into nanophotonic platforms. We present a similar study on a newly developed fluorescent protein, mRhubarb720, embed ded in polyvinyl alcohol (PVA). We detail the sample preparation procedure devel oped to localise mRhubarb720 molecules to the ’hotspots of GNRs and we conduct simulations with the specific sample characteristics to ensure the requirements for plasmonic enhancement are met. The low temperature time-trace emission spec-– i tra of single mRhubarb720 molecules show characteristics indicative of fluorescence emission and resonant Raman emission. We analyse the Raman spectral lines using non-negative matrix factorisation analysis to resolve the spectral lines into com ponents. We further analyse the Raman spectral lines by calculating correlation coefficients for the peak positions, widths and areas. We also present a temperature dependence of the Raman spectra. mRhubarb720 was developed towards a goal of engineering an ideal fluorescent probe. One criterion for an idea fluorescent probe is that its far-red emitting to reduce absorption in biological samples, so contrast is increased in fluorescence imaging. mRhubarb720 meets other criteria for ideal fluo rescent probes and is the farthest red-emitting fluorescent protein developed to date. However, research into far-red emitting fluorescent proteins may provide insight into engineering further-red emitting fluorescent proteins. This study provides a method to gain spectral data on individual fluorescent proteins and is the first to investi gate mRhubarb720. We also provide a protocol for in-depth analysis of the spectral lines and provide a benchmark for further analysis. We extend the investigation to single multichromophoric systems. Multichromophoric systems exhibit complex excited-state dynamics due to the energy transfer mechanisms between the fluo rophores. In nature, multichromophoric systems often have a functional role. Some light-harvesting complexes in plants, bacteria and algae are capable of transferring sunlight energy with high efficiency. Studies have shown this process arises from quantum effects that manifest on a molecular scale. Optical studies to gain informa tion on the excited state dynamics of light-harvesting complexes are intriguing and by utilising GNRs, we provide a method of studying these systems in more depth. Westudy the light-harvesting complex, LH2, from purple bacteria. We use the same sample preparation procedure as with mRhubarb720, except using GNRs with dif ferent aspect ratios and show simulations with the sample-specific conditions. LH2 exhibits two emission peaks due to the two bacteriochlorophyll rings in the structure, the B850 ring and the B800 ring. By choosing GNRs with different aspect ratios and therefore plasmon resonance peaks, we probe both bacteriochlorophyll rings. In the spectral time-trace data for the B850 band, we observe zero-phonon lines with significant spectral wandering. The spectral width of these zero phonon lines is as narrow as 0.073meV, significantly sharper than previously reported. In the spectral time-trace data for the B800 band, we observe similar spectra with extremely sharp zero-phonon lines as narrow as 0.083meV. To our knowledge, sharp spectral lines from this emission band have not been reported in the literature. Finally, we present a study on an artificial multichromophoric system. Biomimicking light-harvesting complexes to create a selective and tunable artificial system allows for a controlled study that can provide valuable information. We study a dye-conjugated virus called OGBMV,which consists of 272 Oregon Green 488 fluorophores bioconjugated to the surface of a 28nm virus surface. Firstly, we conduct a room temperature fluores cence time trace study of single molecules and use an in-house developed image analysis software to determine the emission time-trace of individual molecules. We observe photobleaching steps and we hypothesise that these are from ’super’ emitters in the system which are photobleaching. We also observe steps in which emission increases, which we hypothesise are a result of a decrease in the non-radiative de cay from energy sinks in the system due to a disruption of the Förster transport chain to the sinks. We then present a low-temperature single-molecule spectroscopy study of OGBMV coupled to SNCs embedded in Polyvinyl pyrrolidone. We use the same sample preparation procedure as used for LH2, and we present system-specific– ii simulations of the optical properties of the system. In the spectral time-trace data, we observe zero-phonon lines, which rapidly photobleach and photobleaching steps. To provide insight into the excited state dynamics of this system, we developed a Monte Carlo simulation. Using x-ray-diffraction data to provide the coordinates of Oregon Green 488 conjugation sites, and providing rate values for processes, we sim ulate the excited-state dynamics over a few nanoseconds. We include energy sinks in the system and test the hypothesis over various parameter sweeps. We find that energy sinks get saturated at high excitation rates and high fluorophore densities, producing a radiation brightening effect.

Item Type:	Thesis (PhD)
Date Type:	Completion
Status:	Unpublished
Schools:	Schools > Physics and Astronomy
Subjects:	Q Science > QC Physics
Uncontrolled Keywords:	Plasmonics, quantum optics, spectroscopy, fluorescence, multichromophoric systems, low temperature, biophysics, nanophysics, single-molecule, surface enhanced Raman spectroscopy, SERS.
Funders:	EPSRC Standard Research (DTP)
Date of First Compliant Deposit:	15 April 2026
Last Modified:	15 Apr 2026 13:57
URI:	https://orca.cardiff.ac.uk/id/eprint/186427

Actions (repository staff only)

Edit Item