Spectroscopic techniques: I Spectrophotometric techniques

Abstract

INTRODUCTION Spectroscopic techniques employ light to interact with matter and thus probe certain features of a sample to learn about its consistency or structure. Light is electromagnetic radiation, a phenomenon exhibiting different energies, and dependent on that energy, different molecular features can be probed. The basic principles of interaction of electromagnetic radiation with matter are treated in this chapter. There is no obvious logical dividing point to split the applications of electromagnetic radiation into parts treated separately. The justification for the split presented in this text is purely pragmatic and based on ‘common practice’. The applications considered in this chapter use visible or UV light to probe consistency and conformational structure of biological molecules. Usually, these methods are the first analytical procedures used by a biochemical scientist. The applications covered in Chapter 13 present a higher level of complexity in undertaking and are employed at a later stage in biochemical or biophysical characterisation. An understanding of the properties of electromagnetic radiation and its interaction with matter leads to an appreciation of the variety of types of spectra and, consequently, different spectroscopic techniques and their applications to the solution of biological problems. Properties of electromagnetic radiation The interaction of electromagnetic radiation with matter is a quantum phenomenon and dependent upon both the properties of the radiation and the appropriate structural parts of the samples involved. This is not surprising, since the origin of electromagnetic radiation is due to energy changes within matter itself.