A strategy for high-speed countercurrent chromatography purification of specific antioxidants from natural products based on on-line HPLC method with radical scavenging assay
A strategy for high-speed countercurrent
chromatography purification of
specific antioxidants from natural products based on
on-line HPLC method
with radical scavenging assay
Koichi Inoue a,b,⇑, Erika Baba a,
Tomoaki Hino a,c, Hisao Oka a,c
a Department of
Physical and Analytical Chemistry, School of Pharmacy, Kinjo Gakuin University,
2-1723 Omori, Moriyama-ku, Nagoya 463-8521, Japan
b Laboratory of
Analytical and Bio-Analytical Chemistry, School of Pharmaceutical Sciences,
University of Shizuoka, Japan
c Graduate
School of Human Ecology, Human Ecology Major, Kinjo Gakuin University, Japan
a
b s t r a c t
We have
proposed a novel and first strategy of high-speed countercurrent chromatography
(HSCCC) purification for the efficient and effective discovery of antioxidant
from natural product based on on-line HPLC method with radical scavenging
assay. To achieve a strategy for HSCCC purification, the antioxidants in
materials are identified by on-line HPLC with DPPH radical scavenging assay.
Then, the optimal condition of target peaks would be investigated for the
two-phase solvent system, and purified by HSCCC. In this study, the specific
antioxidants in red cabbage, perilla and elderberry pigments were evaluated by on-line
HPLC with DPPH radical scavenging assay, and purified by HSCCC technique.
Specific antioxidants could be rapidly pinpointed in complex mixtures by
on-line HPLC with DPPH radical scavenging assay. Then, the optimal two-phase
solvent systems were investigated using these HPLC peaks. Finally, the
purification of these nine antioxidants form three mixtures were performed by
HSCCC. Using mass spectrometric analysis, these antioxidants were confirmed to
cyanidin-based anthocyanin from red cabbage and elderberry pigments, and
luteolin-based flavones from perrilla pigment. Due to the advantages derived from
on-line HPLC with DPPH radical scavenging assay and HSCCC technique, a rapid,
efficient and effective strategy has been developed for the discovery of antioxidants
from natural products.
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