AN INTRODUCTION TO ULTRAVIOLET/VISIBLE ABSORPTION SPECTROSCOPY

1. AN INTRODUCTION TO ULTRAVIOLET/VISIBLE ABSORPTION SPECTROSCOPY CHAPTER 13, 14

2. PROBLEM SET Chapter 13 13-5, -7, -8, -10, -11, -12, -22 Chapter 14 14-1, 14-6, 14-7, 14-11 Due Tues., May 1, 2012

3. FIRST LONG EXAMINATIONCh 135 A/B/C May 3, Thursday. 1:30 – 4:00 PM, C114 Coverage: Chapters 1, 6-10, 13-17 (5th Ed., Skoog)

4. In this chapter, absorption by molecules, rather than atoms, is considered. • Absorption in the ultraviolet and visible regions occurs due to electronic transitions from the ground state to excited state. • Broad band spectra are obtained since molecules have vibrational and rotational energy levels associated with electronic energy levels. • The signal is either absorbance or percent transmittance of the analytesolution.

5. AN INTRODUCTION TO ULTRAVIOLET/VISIBLEMOLECULAR ABSORPTION SPECTROMETRY Absorption measurements based upon ultraviolet and visible radiation find widespread application for the quantitative determination of a large variety species. Beer’s Law: A = -log T = logP0/P = bc A = absorbance  = molar absorptivity [M-1 cm-1] c = concentration [M] P0= incident power P = transmitted power (after passing through sample)

9. MEASUREMENT OF TRANSMITTANCE AND ABSORBANCE: The power of the beam transmitted by the analyte solution is usually compared with the power of the beam transmitted by an identical cell containing only solvent. An experimental transmittance and absorbance are then obtained with the equations. P0 and P refers to the power of radiation after it has passed through the solvent and the analyte.

10. BEER’S LAW AND MIXTURES Each analyte present in the solution absorbs light! The magnitude of the absorption depends on its e A total = A1+A2+…+An A total = e1bc1+e2bc2+…+enbcn If e1 = e2 =en then simultaneous determination is impossible Need nl’s where e’s are different to solve the mixture

11. LIMITATIONS TO BEER’S LAW Real limitations Chemical deviations Instrumental deviations

12. REAL LIMITATIONS • Beer’s law is good for dilute analyte solutions only. High concentrations (>0.01M) will cause a negative error since as the distance between molecules become smaller the charge distribution will be affected which alter the molecules ability to absorb a specific wavelength. • The same phenomenon is also observed for solutions with high electrolyte concentration, even at low analyte concentration. The molar absorptivity is altered due to electrostatic interactions.

13. REAL LIMITATIONS In the derivation of Beer’s law we have introduced a constant (e). However, e is dependent on the refractive index and the refractive index is a function of concentration. Therefore, ewill be concentration dependent. However, the refractive index changes very slightly for dilute solutions and thus we can practically assume that eis constant.

14. REAL LIMITATIONS In rare cases, the molar absorptivity changes widely with concentration, even at dilute solutions. Therefore, Beer’s law is never a linear relation for such compounds, like methylene blue.

15. CHEMICAL DEVIATIONS • This factor is an important one which largely affects linearity in Beer’s law. • It originates when an analyte dissociates, associates, or reacts in the solvent. • For example, an acid base indicator when dissolved in water will partially dissociate according to its acid dissociation constant:

20. Chemical deviations from Beer’s law for unbuffered solutions of the indicator Hln. Note that there are positive deviations at 430 nm and negative deviations at 570 nm. At 430 nm, the absorbance is primarily due to the ionized In- form of the indicator and is proportional to the fraction ionized, which varies nonlinearly with the total indicator concentration. At 570 nm, the absorbance is due principally to the undissociated acid Hln, which increases nonlinearly with the total concentration.

21. CALCULATED ABSORBANCE DATA FOR VARIOUS INDICATOR CONCENTRATIONS

23. INSTRUMENTAL DEVIATIONS Beer’s law is good for monochromatic light only since e is wavelength dependent. It is enough to assume a dichromatic beam passing through a sample to appreciate the need for a monochromatic light. Assume that the radiant power of incident radiation is Po and Po’ while transmitted power is P and P’. The absorbance of solution can be written as:

26. The effect of polychromatic radiation on Beer’s law. In the spectrum at the top, the absorptivity of the analyte is nearly constant over Band A from the source. Note in the Beer’s law plot at the bottom that using Band A gives a linear relationship. In the spectrum, Band B corresponds to a region where the absorptivity shows substantial changes. In the lower plot, note the dramatic deviation from Beer’s law that results.

28. Therefore, the linearity between absorbance and concentration breaks down if incident radiation was polychromatic. • In most cases with UV-Vis spectroscopy, the effect small changes in wavelengths is insignificant since ediffers only slightly; especially at the wavelength maximum.

29. STRAY RADIATION Stray radiation resulting from scattering or various reflections in the instrument will reach the detector without passing through the sample. The problem can be severe in cases of high absorbance or when the wavelengths of stray radiation is in such a range where the detector is highly sensitive as well as at wavelengths extremes of an instrument. The absorbance recorded can be represented by the relation: A = log (Po + Ps)/(P + Ps) Where; Ps is the radiant power of stray radiation.

31. INSTRUMENTAL NOISE AS A FUNCTION IN TRANSMITTANCE

34. Therefore, an absorbance between 0.2-0.7 may be advantageous in terms of a lower uncertainty in concentration measurements. At higher or lower absorbances, an increase in uncertainty is encountered. It is therefore advised that the test solution be in the concentration range which gives an absorbance value in the range from 0.2-0.7 for best precision.

35. EFFECT OF BANDWIDTH Effect of bandwidth on spectral detail for a sample of benzene vapor. Note that as the spectral bandwidth increases, the fine structure in the spectrum is lost. At a bandwidth of 10 nm, only a broad absorption band is observed.

36. Effect of slit width (spectral bandwidth) on peak heights. Here, the sample was s solution of praseodymium chloride. Note that as the spectral bandwidth decreases by decreasing the slit width from 1.0 mm to 0.1 mm, the peak heights increase.

37. EFFECT OF SCATTERED RADIATION AT WAVELENGTH EXTREMES OF AN INSTRUMENT • Wavelength extremes of an instrument are dependent on type of source, detector and optical components used in the manufacture of the instrument. Outside the working range of the instrument, it is not possible to use it for accurate determinations. However, the extremes of the instrument are very close to the region of invalid instrumental performance and would thus be not very accurate. An example may be a visible photometer which, in principle, can be used in the range from 340-780 nm. It may be obvious that glass windows, cells and prism will start to absorb significantly below 380 nm and thus a decrease in the incident radiant power is significant.

38. B: UV-VIS spectrophotometer A: VIS spectrophotometer EFFECT OF SCATTERED RADIATION Spectrum of cerium (IV) obtained with a spectrophotometer having glass optics (A) and quartz optics (B). The false peak in A arises from transmission of stray radiation of longer wavelengths.

39. The output from the source at the low wavelength range is minimal. Also, the detector has best sensitivities around 550 nm which means that away up and down this value, the sensitivity significantly decrease. However, scattered radiation, and stray radiation in general, will reach the detector without passing through these surfaces as well as these radiation are constituted from wavelengths for which the detector is highly sensitive. In some cases, stray and scattered radiation reaching the detector can be far more intense than the monochromatic beam from the source. False peaks may appear in such cases and one should be aware of this cause of such peaks.

40. INSTRUMENTATION • Light source •  - selection • Sample container • Detector • Signal processing • Light Sources (commercial instruments) • D2 lamp (UV: 160 – 375 nm) • W lamp (vis: 350 – 2500 nm)

41. SOURCESDeuterium and hydrogen lamps (160 – 375 nm) D2 + Ee → D2* → D’ + D’’ + h Excited deuterium molecule with fixed quantized energy Dissociated into two deuterium atoms with different kinetic energies Ee = ED2* = ED’ + ED’’ + hv Ee is the electrical energy absorbed by the molecule. ED2* is the fixed quantized energy of D2*, ED’ and ED’’ are kinetic energy of the two deuterium atoms.

43. SOURCES Deuterium lamp UV region (a) A deuterium lamp of the type used in spectrophotometers and (b) its spectrum. The plot is of irradiance Eλ (proportional to radiant power) versus wavelength. Note that the maximum intensity occurs at ~225 nm.Typically, instruments switch from deuterium to tungsten at ~350 nm.

44. VISIBLE AND NEAR-IR REGION (a) A tungsten lamp of the type used in spectroscopy and its spectrum (b). Intensity of the tungsten source is usually quite low at wavelengths shorter than about 350 nm. Note that the intensity reaches a maximum in the near-IR region of the spectrum (~1200 nm in this case).

45. The tungsten lamp is by far the most common source in the visible and near IR region with a continuum output wavelength in the range from 350-2500 nm. The lamp is formed from a tungsten filament heated to about 3000 oChoused in a glass envelope. The output of the lamp approaches a black body radiation where it is observed that the energy of a tungsten lamp varies as the fourth power of the operating voltage.

46. Tungsten halogen lamps are currently more popular than just tungsten lamps since they have longer lifetime. Tungsten halogen lamps contain small quantities of iodine in a quartz envelope. The quartz envelope is necessary due to the higher temperature of the tungsten halogen lamps (3500 oC). The longer lifetime of tungsten halogen lamps stems from the fact that sublimed tungsten forms volatile WI2 which redeposits on the filament thus increasing its lifetime. The output of tungsten halogen lamps are more efficient and extend well into the UV.

47. SOURCESTungsten lamps (350-2500 nm) Why add I2 in the lamps? W + I2 → WI2 Low limit: 350 nm Low intensity Glass envelope

48. Xenon Arc Lamps • Passage of current through an atmosphere of high pressured xenon excites xenon and produces a continuum in the range from 200-1000 nm with maximum output at about 500 nm. Although the output of the xenon arc lamp covers the whole UV and visible regions, it is seldom used as a conventional source in the UV-Vis. The radiant power of the lamp is very high as to preclude the use of the lamp in UV-Vis instruments. However, an important application of this source will be discussed in luminescence spectroscopy which will be discussed later

50. SAMPLE CONTAINERS • Sample containers are called cells or cuvettes and are made of either glass or quartz depending on the region of the electromagnetic spectrum. • The path length of the cell varies between 0.1 and 10 cm but the most common path length is 1.0 cm. Rectangular cells or cylindrical cells are routinely used. • In addition, disposable polypropylene cells are used in the visible region. The quality of the absorbance signal is dependent on the quality of the cells used in terms of matching, cleaning as well as freedom from scratches.