1 Ultrafialová a viditelná spektroskopie Brno 2015, Dominik HegerÚstav chemie, MU Elektronová spektroskopie 2 Sluneční světlo 4 What is UV-VIS spectroscopy measuring? Electronic transitions. l / nm 185 - 200 Vacuum-UV 200 – 380 Near-UV (quartz) 380 – 780 Visible Eint = Eel + Evib + Erot D E = h n 5 Spectrum Abscissa (x) ~ D E ~ ~ 1/l Ordinat (y) ~ intensity ~ probability of transition  the oscillator strength f ≡ 4.3e-9 ∫ e d ~ 4.3e-9 emax D 1/2n~ D E =hn =hc/l=hc n~ n~ n~ 6 • Překryvový integrál • Dipólový moment 7 Beer – Lambert – Bouguer law -log (Ft/F0) = -log ti = ecb = A(l) ti = Ft/F0 internal transmittance (transmission factor)  = Fr/F0 reflectance (reflection factor) a = Fa /F0 = 1 – t absorptance (absorption faktor) F0 Fa Fr Ft Term Symbol Unit Monochromatic radiant power F W (decadic) internal absorbance A 1 Molar (decadic) absorption coefficient e L mol-1 cm-1 Absorption pathlength b cm-1 Cell pathlength l cm-1 http://old.iupac.org/reports/V/spectro/partVII.pdf Extinction – sum of absoption, scattering and luminescence. Beer – Lambert – Bouguer law 8 0.000 0.015 0.030 0.045 0.060 0.075 0.090 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 Calibration Curve of Fe 2+ A510nm Fe2+ (mM) Equation y = a + b*x Weight No Weighting Residual Sum of Squares 5.95001E-4 Adj. R-Square 0.99971 Value Standard Error A 510nm Intercept 6.40621E-4 0.00143 A 510nm Slope 10809.93043 35.86739 9 Beer – Lambert – Bouguer law limitations • Chemical reasons – chemical equilibria (acid-base reaction, self-association, complexation, thermal reaction, photochemical reaction, inhomogenous samples) • Physical reasons – thermochromism, saturation effects – depletion of the ground state, incident radiation must be parallel Jak funguje spektrometr? 11 Single beam UV – VIS spectrophotometer with diod array detector 12 Single beam UV – VIS spectrophotometer 13 Double beam UV – VIS spectrophotometer 14 Double beam, double monochromators UV – VIS spectrophotometer 15 UV – VIS spectrophotometer Light source • Continuuu × spectral-line source • Hg-halogen lamp <330; 1200) nm • D2 lamp nm • Xe arc lamp <190; 1000> nm Spectrometer • Single beam × double beam • Sequential × simultaneous Cuvettes • shape • volume •material •use of matched cells Detectors • photo-emisive detectors (evacuated phototubes, photomultiplier...) • semiconuctive detectors (photodiods; detectors with a spacial resolution– CCD –charge-coupled device) http://www.hellmaoptik.com/en/kuevetten/ 16 Lamps Methyl-Naftalen 17 A t 0.001 0.998 0.010 0.977 0.046 0.900 0.100 0.794 0.430 0.370 0.500 0.316 1.000 0.100 1.500 0.032 2.000 0.010 3.000 0.001 4.0000 0.0001 -log (Ft/F0) = -log ti = ecb = A(l) 18 Franck-Condon Principle; Vertical excitation Benzene 19What is the vibrational progression in GS and in S1? Application of derivative spectroscopy to benzene 20 Franck-Condon Principle 21 220 230 240 250 260 270 0.0 0.2 0.4 A ~n / (m–1 ) 4.6 4.4 4.2 4.0 3.8 Benzene in hexane l / nm 22 Absorption bands of organic molecules Type of transition emax n  p* (R) < 100 p  p* (K) > 10 000 (B – benzenoid) ~ 500 (E - ethylenic) 2000 – 14 000 Effects hyperchromic hypochromic Shifts hypsochromic bathochromic 23 24 25 Measuring techniques • Kvantitativní analýza • Derivative spectroscopy-Finding of maxima– Antonov – Step by step filter (SBSF) – • Resolution of overlapping bands (x deconvolution) – Curve fitting – Gaussian or others – Nonparametric methods – Singular Value Decomposition, Target Factoral Analysis • Difference absorption spectroscopy • Absorbance matching 26 Derivative spectroscopie Antony J. Owen: Uses of Derivative Spectroscopy, Agilent Technologies, 1995 27 Derivative spectroscopie 1st derivative 2nd derivative 3th derivative 4th derivative 250 300 350 400 450 0 5000 10000 15000 20000 25000 l / nm -10 -5 0 5 ~n / (m–1 ) 4.5 4.0 3.5 3.0 2.5 e / (M-1 cm-1 ) 250 300 350 400 450 0 5000 10000 15000 20000 25000 l / nm -0.010 -0.005 0.000 0.005 ~n / (m–1 ) 4.5 4.0 3.5 3.0 2.5 e / (M-1 cm-1 ) 250 300 350 400 450 0 5000 10000 15000 20000 25000 l / nm -0.00002 -0.00001 0.00000 0.00001 0.00002 0.00003 ~n / (m–1 ) 4.5 4.0 3.5 3.0 2.5 e / (M-1 cm-1 ) 250 300 350 400 450 0 5000 10000 15000 20000 25000 l / nm -4.00E-008 0.00E+000 4.00E-008 ~n / (m–1 ) 4.5 4.0 3.5 3.0 2.5 e / (M-1 cm-1 ) 28 Photochromism 29 pH titration Naše aplikace • Přesné stanovení koncentrací (kvantové výtěžky) • Speciace v roztoku • Rovnovážné konstanty (pKa) • Solvatochromismus • První informace o molekule v excitovaném stavu (ES, pKa v excitovaném stavu) 30 Excimers 31Theodor Foerster and K. Kasper: Z. physik. Cehm. N.F. J. 275 1954 Excimers in Ice 32 Benzene Naphthalene Does Bz and Np shifts bathochromicaly on ice? 33 Perfect match between: gas phase vibrations and solid state spectrum! 34 Konečně nahoře! Legrace (fotochemie) teprve začíná. Rhodopsin = 11 – cis retinal + opsin ~ 200 fs Fotochemie vidění Mechanismus fototransdukce Lidské detektory světla • Tyčinky - rhodopsin • Čípky – photopsiny, trichromatické vidění Záblesková fotolýza M. Eigen; R. G. W. Norrish; G. Porter (NP 1967) A. Zewail (NP 1999) 300 400 500 0.0 0.1 0.2 0.3 l / nm DA 20 ns 10 s 0 5.10-5 1.10-4 1.10-4 0.0 0.1 0.2 t / s DA l = 390 nm Fotoaktivovatelné molekuly chránicí skupiny X NO2 HO O X O X O X OMeO O X HO O NO2 CH CH3 O P O O O P O O O N NN N NH2 O OHOH OP O O H NO O CH3 O P O O O P O O O N NN N NH2 O OHOH OP O O H + 2Na 2Na hn Fotoaktivovatelná cytostatika neurotransmiter Protecting group spoutané neurotrasmitery Deriváty p- hydroxyacetofenonu HO O O S O O HO O O HO O X hn -X HO O OH T1 O O HO O X O X OH hn T1 3 l / nm t / ps DA DA l / nm t / ps Fotochemické šití tkáně • Zesíťování kolagenových vláken • Rychlé a nedestruktivní • Bengálská červeň Fotochemické šití tkáně Tepna prasete – ex vivo Tepna krysy – in vivo Scientia est potentia! Dvoufotonová absorpce • 3D nanoobrábění = dvoufotonová fotopolymerizace 49 , Normalizovaná odezva lidských čípků Proč je mrkev červená? β-karoten Fotosyntetizujicí barviva Proč papír žloutne? Struktura ligninu Základní jednotky - lignoly Proč býval papír nažloutlý? • Fotodegradace ligninu O O • Oxidace Jak se bělí (modří!) prádlo v Itálii? • Modrá šmolka (hlinitokřemičitany sodné + síra) • S3 -. radikál způsobuje modrou barvu Fluorescence • látka emituje při delších vlnových délkách než absorbuje • zvýrazňovače, cedulky na lyžařských svazích Fluorescence fluoresceinu Fluorescence fluoresceinu 58 Literature • http://old.iupac.org/reports/V/spectro/partVII.pdf • Silverstein R. M. et all: Spectrometric indentification of organic compounds, John Wiley & Sons, inc. • Reichardt, C. (1994). "Solvatochromic Dyes as Solvent Polarity Indicators." Chemical Reviews 94(8): 2319-2358. • http://www.hellmaoptik.com/en/kuevetten/ • http://www.orgchm.bas.bg/~lantonov/ • UV-vis spectra – Heinz-Helmut Perkampus: UV-VIS Atlas of Organic Compounds – science-softCon "UV/Vis+ Spectra Data Base" (UV/Vis+ Photochemistry Database) 7th Edition (2010); ISBN 978-3-00-030970-0 – http://webbook.nist.gov/chemistry/name-ser.html Fotosyntetizujicí barviva Proč je obloha modrá a slunce žluté? Proč při pohledu na oblohu nevidíme tmu? • Rayleighův rozptyl ~ 1/λ4 61 solvatochromism change in the position, intensity, and shape of absorption bands due to the surrounding medium N O = perichromism (peri - around) surrounding medium - liquids, solids, glasses, and surfaces • negative solvatochromism a blue (hypsochromic) shift • positive solvatochromism a red (bathochromic) shift solvent polarity increases ET solvent polarity increases ET N+O– O 62 Reichardt’s dye betaine-30 2,6-diphenyl-4-(2,4,6-triphenyl-pyridium)phenolate N O N O hn S0 S1 dipol moment in 1,4-dioxane g = 14.8+-1.2 D e = 6.2+-0.3 D •dipol moment - dipol/dipol, dipol/induced dipol interaction • p electron system - dispersion interaction • phenolate oxygen - highly basic EPD ceter 63 ET(30) and ET N scale of solvent polarity ET - molar electronic transition energy [ET] = kcal/mol [ET N] = 1 ET(30) = hc maxNA = (2.8591 E -3)( max / cm-1) =28591/(lmax / nm) ET N = (ET(solvent)-ET(TMS))/(ET(water)-ET(TMS)) = (ET(solvent) - 30.7)/32.4 ET N (TMS) = 0.000 ET N (H2O) = 1.000 TMS = Tetramethylsilane n~n~ Látka ET N n-hexan 0.009 Dichlor methan 0.309 EtOH 0.608 Voda 1 64