Introduction to Spectroscopy imcluding basics theory.

Introduction to Spectroscopy

1. D. P. Gangwar (M.Sc. Physics/Electronics) DIRECTOR, DP Forensic Lab (95690-99174) [email protected] Ex-Assistant Director & HOD (Physics & Cyber) CFSL, Chandigarh & Hyderabad, DFSS, New Delhi. Ex-Visiting Guest Faculty of PU, Chandigarh Ex-Forensic Expert, UPSIFS Introduction to Spectroscopy

2. Introduction to Spectroscopy Study of spectrum to identify substances Studies of the interaction of EMR energy with matter The branch of science concerned with the investigation and measurement of spectra produced when matter  interacts  with or  emits   electromagnetic  radiation. Study of radiation after interaction with material

3. Electromagnetic Radiation (EMR) Electromagnetic radiation (EMR): is a form of energy t hat travels through space as waves.  These waves consist of oscillating electric and magnetic fields

4. 1. Wave Properties: Transverse Wave: It oscillating electric and magnetic fields that are perpendicular to each other and to the direction the wave travels.  Wavelength and Frequency: Wavelength (distance between wave crests) and frequency (number of wave cycles per second) are inversely related; higher frequency means shorter wavelength and vice versa.  Speed of Light: EMR travels at the speed of light (approximately 3.0 x 10^8 m/s in a vacuum).  Superposition: Electromagnetic waves can overlap and combine, a phenomenon known as superposition.  Reflection, Refraction, Diffraction, and Interference: EMR exhibits these wave behaviors when interacting with matter.  EMR Properties

5. Radiation Properties EMR has particle nature, it travel in form of small energy particle called Photons: Photon is small particle has energy E=hf Photoelectric Effect: The frequency of electromagnetic radiation is equal to the energy of absorbed or emitted radiation. The atoms and molecules can emit and absorb a specific quantity of energy. The smallest amount of energy that can be absorbed or emitted is termed quantum.

6. Electromagnetic (EM) Spectrum

7. Interaction of Radiation with Material Scattering Refraction Reflection Transmission

8. Spectroscopy Normal Light-all wavelength Some WL absorbed Emit only particulat WL Present All wavelength Comes new wavelength Absent few wavelength

9. Atomic Absorption Spectrophotometer(AAS) trace metal analysis, environmental monitoring, and quality control  generally accurate for determining concentrations at the parts per billion (ppb) level

10. UV-Visible Spectrophotometer Drug Identification and Quantification: Quality Control: Biological and Biochemical Analysis: DNA/RNA Bacterial Culture Monitoring: Enzymatic Reaction Analysis: Water Quality Assessment, Air Quality Monitoring: Food and Beverage Analysis: 

11. Raman Spectroscopy Raman Spectroscopy is a spectroscopic technique which is used to analyze vibrational, rotational, and other low-frequency modes in a system . Sir C. V. Raman       Wikipedia Raman scattering - Wikipedia In chemistry and physi When light interacts with a sample, most of the light is either absorbed, transmitted, or reflected, but a small fraction is scattered inelastically, resulting in a shift in energy that corresponds to the vibrational energy levels of the molecules in the sample. This phenomenon can be categorized into  Stokes  and  anti-Stokes  scattering, depending on whether the scattered photons have lower or higher energy than the incident photons, respectively. Raman scattering is widely used in spectroscopy to provide information about molecular vibrations and chemical composition .

12. FTIR: Introduction Fourier Transform Infra Red M athematical representation of a periodic function as an infinite sum of sine and cosine waves It allows complex periodic functions to be broken down into simpler, constituent waves, which is particularly useful in fields like signal processing, physics, and engineering.  Mathematical process of converting raw data (an interferogram) into a spectrum EM Radiation wavelength range: 800nm to 130000nm

13. FTIR: Introduction Spectrum and Spectroscopy Spectrum: A pattern of different colors observed when the white light was dispersed through the prism The changing of light intensity as a function of frequency Breaking the EMR/light in its components

14. IR Spectroscopy: When the IR passed through the sample, it absorbed the IR and the remaining passed through the samples. The absorbance of IR depends on the nature of sample .i.e. bound and position of functional group. Two compound does not have identical IR Spectra FTIR Spectroscopy IR T.IR R.IR A.IR Detector Detect this

15. Infrared Spectroscopy An IR spectrum is a graph showing the amount of IR light absorbed by a sample at different frequencies The x-axis represents frequency (or wavelength) of the IR light. The y-axis represents the intensity of absorption (or transmittance, how much light passes through) Absorption peaks in the spectrum indicate which frequencies were absorbed, revealing which bonds and vibrations are present in the molecule.    when a molecule absorbs infrared radiation, causing its chemical bonds to vibrate.  The specific frequencies of light absorbed are unique to the molecule's structure and bonding, creating a "f ingerprint " that can be used for identification

16. Principle of IR Spectroscopy Any molecules, it is group of atoms are connected by bond 2. These are analogous to spring and not rigid in nature. 3. They continues move/vibrate with some frequency characteristic to evert portion of molecules( these natural f )

17. Principle of IR Spectroscopy 4. When energy applied (IR), molecules are excited to high energy state on absorption IR (selective WL) and vibrate the molecules. Absorption is corresponding to the functional group and bond of a compound. As every compound has unique FG, bonds hence, absorbed IR is unique. Hence, IR Spectra is unique. H2O Then f will Absorbed and It will absent in transmitted light

18. Despite the Typical Graphical Display of Molecular Structures, Molecules are Highly Flexible and Undergo Multiple Modes Of Motion Over a Range of Time-Frames Motions involve rotations, translations, and changes in bond lengths, bond angles, dihedral angles, ring flips, methyl bond rotations. Infrared Spectroscopy

19. Infrared Spectroscopy An IR spectrum is a graph showing the amount of IR light absorbed by a sample at different frequencies The x-axis represents frequency (or wavelength) of the IR light. The y-axis represents the intensity of absorption (or transmittance, how much light passes through) Absorption peaks in the spectrum indicate which frequencies were absorbed, revealing which bonds and vibrations are present in the molecule.    when a molecule absorbs infrared radiation, causing its chemical bonds to vibrate.  The specific frequencies of light absorbed are unique to the molecule's structure and bonding, creating a "f ingerprint " that can be used for identification

20. B) Theory of IR Absorption 1.) Molecular Vibrations i .) Harmonic Oscillator Model : - approximate representation of atomic stretching - two masses attached by a spring E = ½ ky 2 where: y is spring displacement k is spring constant Infrared Spectroscopy

21. Vibrational frequency given by: = 1/2 p p k / m where: n : frequency k : force constant (measure of bond stiffness) m : reduced mass – m 1 m 2 /m 1 +m 2 If know n and atoms in bond, can get k: Single bonds : k ~ 3x10 2 to 8 x10 2 N/m (Avg ~ 5x10 2 ) double and triple bonds ~ 2x and 3x k for single bond. n ~ p k So, vibration n occur in order: single < double < triple Infrared Spectroscopy

22. FTIR Spectroscopy Covers wavelengths from approximately Near-Infrared (NIR): 0.7 to 2.5 (µm) or 14,000 to 4,000 cm⁻¹ Often associated with overtone an d combination bands of molecular vibrations.  Used in applications like spectroscopy, remote sensing, and some medical imaging.  Mid-Infrared (MIR): 2.5 to 25 µm or 4,000 to 400 cm⁻¹ ( fundamental vibrational transitions of molecules Widely used in chemical analysis (infrared spectroscopy) to identify functional groups and structures Also used in thermal imaging and remote sensing.  Far-Infrared (FIR): Covers wavelengths from 25 to 1000 µm or 400 to 10 cm⁻¹ wavenumbers.  Involves low-frequency molecular vibrations, rotations, and intermolecular interactions .

23. FTIR Spectroscopy

24. FTIR Spectroscopy

25. Fundamental Vibrations Bond Stretching Bond Bending symmetric asymmetric In-plane rocking In-plane scissoring Out-of-plane wagging Out-of-plane twisting

26. FTIR Spectroscopy Wave number calculation

27. FTIR Spectroscopy

28. Fundamental Vibration : Non-linear (No. of Vibration mode= 3n-6) Linear: (3n-5) FTIR Spectroscopy 3N - 6 = 3× 3 - 6 = 3.

29. Molecular Vibrations

30. FTIR Spectroscopy

31. Fourier Transfer IR (FTIR) – alternative to Normal IR Based on Michelson Interferometer Principale : 1) light from source is split by central mirror into 2 beams of equal intensity 2) beams go to two other mirrors, reflected by central mirror, recombine and pass through sample to detector 3) two side mirrors. One fixed and other movable a) move second mirror, light in two-paths travel different distances before recombined b) constructive & destructive interference c) as mirror is moved, get a change in signal

32. Destructive Interference can be created when two waves from the same source travel different paths to get to a point. This may cause a difference in the phase between the two waves. If the paths differ by an integer multiple of a wavelength, the waves will also be in phase. If the waves differ by an odd multiple of half a wave then the waves will be 180 degrees out of phase and cancel out.

33. observe a plot of Intensity vs. Distance (interferograms) convert to plot of Intensity vs. Frequency by doing a Fourier Transform resolution Dn = 1/ Dd (interval of distance traveled by mirror)

34. Advantages of FTIR compared to Normal IR : 1) much faster, seconds vs. minutes 2) use signal averaging to increase signal-to-noise (S/N) increase S/N % r number scans 3) higher inherent S/N – no slits, less optical equipment, higher light intensity 4) high resolution (<0.1 cm -1 ) Disadvantages of FTIR compared to Normal IR : 1) single-beam, requires collecting blank 2) can’t use thermal detectors – too slow In normal IR, scan through frequency range. In FTIR collect all frequencies at once.

35. Bond Type of Compound Frequency Range, cm -1 Intensity C-H Alkanes 2850-2970 Strong C-H Alkenes 3010-3095 675-995 Medium strong C-H Alkynes 3300 Strong C-H Aromatic rings 3010-3100 690-900 Medium strong 0-H Monomeric alcohols, phenols Hydrogen-bonded alchohols, phenols Monomeric carboxylic acids Hydrogen-bonded carboxylic acids 3590-3650 3200-3600 3500-3650 2500-2700 Variable Variable, sometimes broad Medium broad N-H Amines, amides 3300-3500 medium C=C Alkenes 1610-1680 Variable C=C Aromatic rings 1500-1600 Variable Alkynes 2100-2260 Variable C-N Amines, amides 1180-1360 Strong Nitriles 2210-2280 Strong C-O Alcohols, ethers,carboxylic acids, esters 1050-1300 Strong C=O Aldehydes, ketones, carboxylic acids, esters 1690-1760 Strong NO 2 Nitro compounds 1500-1570 1300-1370 Strong Abbreviated Table of Group Frequencies for Organic Groups

36. iii.) Fingerprint Region (1200-700 cm -1 ) - region of most single bond signals - many have similar frequencies, so affect each other & give pattern characteristics of overall skeletal structure of a compound - exact interpretation of this region of spectra seldom possible because of complexity - complexity  uniqueness Fingerprint Region

37. FTIR Spectroscopy IR Source: Silicon Carbide ( Globar ): MIR Tungsten Filament: NIR Mercury: FIR Beam Splitter: KBR for MIR 632.8 nm

38. Reference Signal : FTIR spectrometers use a laser (typically a H elium-Neon laser ) to generate a stable, monochromatic beam that serves as a reference signal.  Mirror Movement Tracking: /I nterferometer Alignment The laser beam passes through the interferometer, and its interference pattern (an interferogram) is used to track the precise position of the moving mirror.  Data Acquisition Timing : The laser's interferogram provides a timing reference, allowing the spectrometer to acquire data points at equal intervals of the mirror's path difference. This ensures accurate digitization of the infrared signal for the subsequent Fourier transform.  Accuracy and Precision / Reference Signal : By precisely measuring the mirror's movement, the laser enables accurate wavenumber (frequency) determination of the infrared spectrum and high-resolution measurements.  Internal Calibration : he laser provides an internal calibration standard, allowing for reliable spectral comparisons, even when measurements are taken at different times.  Wavelength Stability : Helium-Neon lasers  are favored for their excellent wavelength stability, which is critical for accurate wavenumber measurements.  Use of Laser in FTIR 632.8 nm

39. FTIR: Application in Forensic Science All form of material---- solid, Liquid, Gas All types of material—Inorganic, organic, etc. It may include i . Drug, Explosive, Alcohol, Poison, etc. ii. Polymer, plastic, rubber, coatings, etc. iii. Hair and Fiber Analysis, Feathers other biological items iv. Explosives and Gunshot Residue Analysis (GSR) v. Ink, Paper, etc.

40. Sample Preparation Nature of sample Solid , Liquid, Gas Mode to Record the IR Spectra Transmission, Reflectance Solid Reflection mode: Benefit-No sampling prepation,100% non- destrictive Shortcomings: less accuracy quality of spectra Transmission Mode: sample allow to pass IR, Make KBr pellet:

41. KBR pellet Preparation

42. Use of Laser in FTIR

43. How to record FTIR Select No of scan Resolution generally 4cm -1 Take BKD spectra Scan sample

44. Analysis of Paint using FTIR

45. FTIR Spectra of two paint samples

46. FTIR Spectra Explosive material

47. FTIR Spectra of paint sample and library matching

48. FTIR Spectra Heroin sample

49. Questions Please