Dissertation > Biological Sciences > Biochemistry > General issues > Biochemical techniques

Fabrication of SERS-active Substrates and Their Utilization for Detection of Amino Acids and Bacteria Biomarkers

Author ChengHanWen
Tutor HuanShuangYan
School Hunan University
Course Analytical Chemistry
Keywords Surface-enhanced Raman scattering L-Histidine Gold nanoparticles SERS substrates DPA (Dipicolinic acid)
CLC Q503
Type Master's thesis
Year 2009
Downloads 88
Quotes 0
Download Dissertation

Surface-enhanced Raman scattering (SERS) is a powerful technique for applications in chemical and biological analyses. SERS can greatly magnify the Raman signals of the adsorbed molecules by a factor of 106 to 1014. Important attributes such as narrow bands, high resolution, low SERS intensity for water, and high stability have enabled SERS to find a wide range of applications in materials science, surface science, biomedical detection and diagnostics. Recently, SERS technique has been used as highly-sensitive technique for the detection of structural conformation adsorbed proteins and biomarkers in bacterial spores. Since Raman signals of proteins originate mainly from the side chains of amino acids that are the building blocks of proteins, it is essential to investigate the SERS spectra and structural confirmation of small molecules of amino acids. Dipicolinic acid (DPA) is a biomarker of bacterial spores. Earlier SERS studies have mainly focused on substrates derived from silver nanoparticles, but there are some drawbacks in using silver nanoparticles as substrate, including difficulty in quantitation and uncertainty in determining detection limit. It is therefore a challenging task to develop rapid and sensitive analytical techniques for the detection of DPA. The details are summarized as follows:In the second chapter, we describe results from an investigation that focuses on combining the SERS technique and density function computation method for the study of the self-assembly of L-Histidine, which is an essential amino acid for infants that differs from the necessary amino acid for adults. Using electrochemically-roughened surfaces of silver as SERS substrates, the absorption process of L- Histidine on silver surfaces was analyzed. The results showed that L-Histidine could complete the self-assembly process after an assembly time of 10.5 h. The theoretical vibrational value of L- Histidine was calculated using B1LYP/ 6-31G method. The comparison of the calculated results with the experimentally observed ring-related vibrational bands and the SERS bands involving N8 and O of the assembled L-Histidine molecule lead us to the conclusion that N1, N3, N8 and O atoms of the adsorbed molecule were close to the surface of silver.In the third chapter, we describe the preparation of highly-sensitive SERS substrates using gold nanoparticles, polyvinylpyrrolidone, and a gold electrode, which produces a gold-nanoparticle/polyvinylpyrrolidone/gold substrate (AuNPs/PVP/Au). The SERS substrate’s activity was correlated with particle sizes for Au nanoparticles of different sizes (50– 70 nm). The size and morphology of the gold nanoparticles were characterized using UV-Vis spectrophotometric technique, and FE-SEM technique. The observed shifts of the band positions of the surface plasmon resonance band for Au NPs of different sizes and their thin films on the Au substrates were correlated with the particle size effect and the partial particle aggregation effect. The experimental results have shown that a maximum SERS intensity was observed for the 60-nm particle size which was independent on the concentration of the analyte. Control experiments have also shown that in addition to particle-particle coupling effect, the particle-substrate coupling has played an important role in the observed SERS effect.In the fourth chapter, we describe the results of the an investigation of the AuNPs/PVP/Au substrates for the detection of dipicolinic acid (DPA), a biomarker for bacterial spores including Bacillus anthracis. The correlation between the SERS intensity of the diagnostic band at 1000cm-1 and the DPA concentration (0.1 ppb– 100 ppm) was shown to exhibit two linear regions, i.e., the low- (<0.01 ppm) and high-concentration (> 1 ppm) regions, with an intermediate region in between. The presence of such a linear relationship in the low-concentration region was observed for the first time in SERS detection of DPA. A detection limit of 0.1 ppb was obtained from the substrates with 60-nm sized Au NPs, which is to our knowledge the lowest detection limit reported for DPA using this type of SERS substrate. This finding was also supported by the estimated enhancement factor (106) and a large adsorption equilibrium constant for the low-concentration region (1.7x107 M-1). The adsorption characteristics of DPA on the SERS substrates were analyzed by modeling both monolayer and multilayer adsorption isotherms for the correlation between the SERS intensity and the DPA concentration. The observed transition from the low- to high-concentration linear regions was found to correspond to the transition from a monolayer to multilayer adsorption isotherm, which was in agreement with the estimated minimum DPA concentration for a monolayer coverage (0.01 ppm).

Related Dissertations
More Dissertations