Page 220 - Vibrational Spectroscopic Imaging for Biomedical Applications

196 Cha pte r Se v e n

good biomarker for fruit and vegetable intake, Raman measurements of
skin carotenoid levels could be used as rapid optical method to assess
fruit and vegetable consumption in large populations.
For many decades, the standard technique for measuring carot-
enoids has been high-pressure liquid chromatography (HPLC). This
chemical method works well for the measurement of carotenoids in
serum, but it is time consuming, expensive, and highly invasive for
human tissue, since it requires biopsies of relatively large tissue vol-
umes. Additionally, serum antioxidant measurements are more indic-
ative of short-term dietary intakes of antioxidants rather than
steady-state accumulations in body tissues exposed to external oxi-
dative stress factors such as smoking and UV-light exposure.

7.2 Optical Properties and Resonance Raman
Scattering of Carotenoids
Carotenoids are π-electron conjugated carbon-chain molecules and are
similar to polyenes with regard to their molecular structure and optical
properties. Distinguishing features are the number of conjugated carbon
double bonds (C=C), the number of attached methyl side groups, and the
presence or absence and composition of attached end groups. The molecu-
lar structure of β-carotene is shown as an example in Fig. 7.1, which con-
sists of a linear, conjugated, carbon backbone with alternating carbon single
(C—C) and double bonds (C=C), two ione end groups, and 4 methyl side
groups. The structure is very similar to all other carotenoids of interest in
this chapter. Resonance Raman spectroscopy detects the vibrational stretch
frequencies of the carbon bonds as well as the rocking motion of the
attached methyl side groups. Also shown is a configuration coordinate
diagram for the three lowest lying energy states, and an indication of all
optical and nonradiative transitions connecting the states. The configura-
tion coordinate represents the displacement of a normal coordinate of the
molecule’s atoms its equilibrium position. Absorption, fluorescence, and
Raman transitions occur at fixed nuclear positions of the molecule’s atoms
(fixed configuration coordinate) as vertical transitions. A strong, electric-
dipole allowed absorption transition occurs between the molecule’s delo-
1
1
calized π-orbital from the 1 A singlet ground state to the 1 B singlet
g u
excited state. As illustrated in Fig. 7.2a, this gives rise to a broad absorption
band in the blue-green spectral region (peak at ~460 nm, spectral width
~100 nm) that features clearly resolved vibronic substructures (with spec-
1
−1
tral spacing of ~1400 cm ). After excitation to the 1 B state, the carotenoid
u
molecule relaxes very rapidly, within ~200 to 250 fs, via nonradiative
17
1
transitions, to the lower-lying 2 A excited state. Since this is a state with
g
gerade parity, a radiative transition to the ground state is parity-forbidden.
1
1
As a consequence, the luminescence transitions from the 1 B and 2 A
u g
states to the ground states are very weak (10 to 10 efficiency). It is this de
−5
−4
facto absence of intrinsic luminescence of carotenoids that allows one to