Identification and detection of molecules
- 2. Why do we need to know the structure of a molecule? Because of the Knowing the structure simplifies drug designing as otherwise, it would solely be based on trial and error Structure helps predict the effects of a drug molecule (both desired and adverse effects) To discover more efficient analogues Knowing the structure also makes drug designing cost effective as in silico techniques can then be effectively employed
- 3. Most common techniques available Spectroscopy X- ray crystallography Electron microscopy Note: A combination of techniques must be adopted to effectively determine a molecule’s structure as different techniques often provide only part of the information provided.
- 6. Mass Spectrometry This is a technique which ionizes molecules and separates the ions based on their mass- to- charge ratio. Used to determine the constituents of a molecule
- 7. Mass Spectrometry- Brief Procedure 1. The molecule is ionized by bombarding with electrons 2. The ions are subjected to an electric field which alters their speed 3. They are then subject to a magnetic field which alters their direction- Lighter ions are deflected more than heavier ions 4. The deflected ions fall on a detector which will record their abundance
- 9. Mass Spectroscopy- Isotopes When an element has isotopes, the corresponding number of recordings are produced on the mass spectrum based on their m/z value. The relative intensities of the recordings will depend on their natural availability. Eg.: Cl is naturally present as 35Cl and 37Cl in the ration 3:1. Hence, two recordings will be produced with the recording for 35Cl 3 times more intense than that for 37Cl.
- 10. Absorption Spectroscopy This is a technique that measures the amount of radiation absorbed by a molecule. The amount varies depending on the frequency of the radiations used. Some physical properties like molecular mass and molecular geometry can be determined by comparing the molecule’s absorption spectrum with certain available models.
- 11. Absorption Spectroscopy- Brief Procedure 1. The spectrum of the radiation source is measured using the detector 2. The sample (molecule) is placed between the radiation source and detector and its absorption spectrum measured 3. Both the spectra are compared to determine the sample’s absorption spectrum
- 12. Absorption Spectroscopy- UV Visible Spectroscopy Radiations in the visible and near- visible regions of the electromagnetic spectrum are used. These radiations have right amount of energy to excite the electrons from one orbit to another.
- 13. Absorption Spectroscopy- UV Visible Spectroscopy Used to determine the type of bonds present The electrons most easily excited are those in the p bonds. Hence greater the number of p bonds, greater will be the absorption.
- 14. Absorption Spectroscopy- Infrared Spectroscopy Radiations in the infrared region are used. These rays have the right amount of energy to cause bonds to stretch and bend. Used to determine the functional groups present
- 15. Absorption Spectroscopy- Infrared Spectroscopy • Different bonds absorb different amounts of energy due to numerous factors like bond length, the atoms involved, etc. • The whole molecule can absorb a certain amount of energy to cause its vibration. This energy is generally <1500 cm-1. This is referred to as the fingerprint of the molecule and is unique for each molecule.
- 16. Fingerprint region Infrared spectrum of propan- 1- ol (CH3CH2CH2OH)
- 17. NMR Spectroscopy Based on the amount of radio waves absorbed by certain isotopes placed in a magnetic field. Used to determine the environment of certain isotopes
- 18. NMR Spectroscopy- The Basics • Isotopes with a nuclear spin of ½ are analyzed • When a magnetic field is applied, the magnetic field resulting from the spin can align either to (lower energy) or against (higher energy) the external field • When radio waves are provided, the nuclei in the lower energy state absorb some energy and jump to the higher energy state. The energy absorbed is what is measured. • The magnetic fields produced by the neighboring atoms also influence the energy absorbed by the nucleus in question
- 19. NMR spectrum of 2- ethyl phenol
- 21. X- ray Crystallography This is a technique for determining the structure of a molecule post crystallization by directing a beam of x- rays and studying the diffraction pattern. X- ray crystallography of bovine Cu, Zn superoxide dismutase
- 23. Electron microscopy This is a technique which detects the structure of a molecule using electrons. TEM SEM • A high voltage electron beam is focused onto the molecule using magnetic fields • Some electrons are transmitted through the molecule while some are scattered • The transmitted electrons are recorded and the structure, developed • An electron beam scans a rectangular area around the molecule • The electrons interact with the molecule and cause loss of energy as heat, light, x- rays, emission of low- energy secondary electrons and high- energy back scattered electrons • These signals are recorded and the structure, developed
- 24. Electron microscopy Note: SEM can only determine the surface structure of molecules while TEM can determine the internal structure as well. B. anthracis spores