Raman and IR Spectroscopic Studies of Bacteria, Endotoxin, DNA and Lipoproteins
In the past few decades, vibrational spectroscopy has emerged as a powerful technique in biomedical research. Raman and IR spectroscopy, which are based on signatures specific to molecular vibrations, has found a wide variety of applications ranging from fundamental research problem to applications in clinics. These two techniques combined with chemometrics can provide molecular insight to any biochemical processes. The techniques are label free, non-invasive and require minimal sample preparation allowing the study of not only simple biomolecules but also whole organisms like cells and bacteria and body fluids from human. In this thesis, important life processes including bacterial growth and stress, detection of endotoxin, prediction of metabolic disorder and DNA conformational changes has been investigated with the help of vibrational spectroscopy and chemometrics. The thesis begins with a brief introduction to Raman and IR spectroscopy and the parameters to consider for biological applications of these two techniques (Chapter 1 and 2). In the bacterial study, differences in bacterial growth have been investigated under different growth conditions with an aim to understand survival mechanism of bacteria in adverse conditions. In this thesis, the univariate, ratiometric and multivariate markers for growth under different conditions has been summarized (Chapter 3). In continuation to this, a novel resonance Raman marker band for cytochrome in E. coli has been identified. The serendipitous discovery of a new peak and its plausible applications in understanding bacterial growth and stress has been discussed (Chapter 4). Another clinically important biomolecule, which is released from the cell wall of bacteria is lipopolysaccharides (commonly known as endotoxin). Detection of endotoxin in ultralow concentration is important as these toxic substances can be lethal even when present in ng/Kg of bodyweight. Our effort to detect endotoxin with a detection limit of pg/mL concentration, with the help of Surface Enhanced Raman Spectroscopy, has been shown (Chapter 5). In the next part, potential of IR and Raman spectroscopy in studying clinically isolated human LDL samples targeting cardiovascular risk has been be demonstrated (Chapter 6). Lastly, it has been shown how Raman optical activity can be useful in understanding DNA conformational changes, a very fundamental life process (Chapter 7). Finally, the work done in the thesis and the future scope of the work has been briefed (Chapter 8). Overall, the thesis describes the applicability of vibrational spectroscopy ranging from basic research to clinics.