1. Chapter 18 Let There Be Light Spectrophotometry The use of electromagnetic radiation to measure chemical concentrations

2.  UV absorption The Ozone Hole

3. Why south pole ? See p.392

4. 18.1 Properties of light -1 1) 2)Electromagnetic Spectrum (Fig 18.2) 3)Absorption vs Emission of light

5. change of nuclear configuration γ-ray change of e distribution X-ray uv.vis change of configuration IR change of orientation microwave change of spin NMR EPR

6. 18.2 Absorption of light -1 1)Spectrophotometer

7. 18.2 Absorption of light -2 2). When no light is absorbed, P=P 0 and A=0

8. Absorbance is proportional to the concentration of light-absorbance molecules in the sample. A =  bc  : molar absorptivity (M -1 cm -1 )

9. 18.2 Absorption of light -3 (ex) How effective is sunscreen ? at the peak absorbance near 300 nm ? A ~ 0.35 T = 10 -A = 10 -0.35 = 0.45 = 45%  55% UV-B is absorbed.

10. Observed color vs. absorbed color

11. 18.3 Practical Matters -1 1) Sample is usually contained in a cell called a cuvet (Figure 18-7), which has flat, fused-silica faces.  A glass made of SiO 2 : Vis. UV.  Plastics & ordinary glass: Vis  NaCl(s) KCl(s) : IR

12. 18.3 Practical Matters -2 2) Good Operating Techniques : a) Cuvets handle: systematic errors/ random errors b) Most accurate at A~ 0.3-2 Too little light : (high A), P is small & hard to measure Too much light : (low A), it is hard to distinguish P from P 0

13. 18.3 Practical Matters -3 c) Old vs. new curves d) Greatest sensitivity: λ max e) Baseline correction

14. (a) Proteins at 280 nm: tyr, phe, trp. (b) A colorimetric reagent to detect phosphate 18.4 Using Beer’s Law -1

15. 18.4 Using Beer’s Law -2 Ex.1 : Bezene: find molar absorptivity (  ) at p.404

16. 18.4 Using Beer’s Law -2 Ex.2 : Nitrite in an aquarium (using a standard curve)  543 nm

17. 18.4 Using Beer’s Law -3 (toxic when > 1 ppm) NH 3  animals & plant (Box 6-1) (toxic when > 1 ppm) [O] NO 3 -

18. 18.4 Using Beer’s Law -4 2) Standard Nitrite ex. at p407

19. 18.4 Using Beer’s Law -5 from least square (4.4) A = 0.1769 [ppm] + 0.0015

20. Enzyme-based nitrate Analysis - A Green Idea P.408 NO 3 - NO 2 - NO 3 - + NADH + H + NO 2 - + NAD + + H 2 O Cd Nitrate reductase pH 7