Chemistry Reference
In-Depth Information
1223
100
A2PE
100
*
80
60
40
20
0
80
*
1239
60
**
40
127
1287
1223
100
1255
A2PE
430 nm
*
20
80
60
40
20
0
100
80
60
40
20
0
1239
0
A2PE
430 nm
Zeaxanthin
Lutein
+
+
+
+
+
+
+
+
+
+
+
+
1223
α-Tocopherol
(B)
A2PE
430 nm
lutein
500
1239
*
1255
400
**
300
1223
100
80
60
40
20
0
A2PE
430 nm
zeaxanthin
200
100
1239
1255
0
A2E A2E
Zeaxanthin Lutein
Endoperoxide
α-Tocopherol
(A)
1160
1240
1320
1400
m/z
(C)
A2E+endoperoxide
FIGURE 16.5
Photooxidation of A2PE and is decreased by lutein and zeaxanthin. The singlet oxygen
quenching activity of lutein and zeaxanthin. (A) FAB-MS of nonirradiated A2PE, A2PE irradiated at 430 nm,
and A2PE illuminated at 430 nm in the presence of lutein or zeaxanthin. The molecular ion peak at mass-to-
charge (
m/z
) ratio 1223 corresponds to the molecular mass of A2PE. A2PE photooxidation is rel ected by the
presence of additional higher molecular weight peaks for example,
m/z
1239, 1255, 1271, 1287 in the irradiated
samples. Illumination in the presence of lutein and zeaxanthin reduces the formation of these photooxidation
products.
*
, matrix peak at
m/z
1245 {M + Na}. (B) Lutein and zeaxanthin protect against A2PE photooxida-
tion. A2PE (200 mM) with and without lutein and zeaxanthin (200 mM) or a-tocopherol (200 mM) was irradi-
ated at 430 nm. A2PE was quantii ed by reverse-phase HPLC and integrated peak areas were normalized to
an external standard of A2E. The loss of A2PE after 430 nm irradiation is indicative of A2PE photooxidation;
the attenuation of this loss in the presence of lutein and zeaxanthin indicates protection against A2PE photo-
oxidation. +, the presence of compound/irradiation. Values are mean ± SD of 4-7 experiments, 3 replicates
per experiment.
*
p
<0.01, for the indicated comparison;
**
p
< 0.001 for a-tocopherol versus zeaxanthin and
lutein values; ANOVA followed by the Newman Keul Multiple Comparison test. (C) Zeaxanthin, lutein, and
a-tocopherol quench singlet oxygen thus reducing A2E oxidation. The consumption of A2E that accompanies
oxidation was quantii ed by HPLC after A2E (500 μM) was exposed to singlet oxygen generated from the
endoperoxide of 1,4-dimethyl-naphthalene (1 mM) in the absence and the presence of zeaxanthin, lutein, and
a-tocopherol (1 mM). Bar height in the presence of antioxidant is positively correlated with quenching ability.
Values are mean ± SD of 3 experiments.
*
p
< 0.05, zeaxanthin versus lutein;
**
p
< 0.01 a-tocopherol versus
lutein and zeaxanthin. ANOVA followed by the Newman Keul Multiple Comparison test.
(Figure 16.5). By analyzing peak areas in reverse phase HPLC chromatograms to quantify the loss
of A2E as it reacts with singlet oxygen, it was shown that both zeaxanthin and lutein were able to
compete with A2E for the quenching of singlet oxygen. Again, zeaxanthin was a more efi cient
quencher than lutein and both more effectively protected A2E than did a-tocopherol. The ability of
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