This dissertation illustrates an efficient purification method for citrus
limonoids and flavonoids, while examining their effects on cytochrome P450
inhibition and apoptotic induction on human neuroblastoma (SH-SY5Y) and
colonic adenocarcinoma (Caco-2) cells. The first study developed a bulk
purification method for limonoids, from seeds and molasses of citrus fruits, using
a combination of chromatographic techniques. This also resulted in an efficient
purification method for naringin and hesperidin from citrus byproducts.
The second study investigated the inhibitory effects of purified limonoids
and flavonoids on the enzymatic activities of different isoforms of human
cytochrome P450. O-Dealkylase and hydroxylase activities of CYP1A2,
CYP1B1, CYP3A4 and CYP19, using specific substrates such as
ethoxyresorufin (ethoxyresorufin O-dealkylase, EROD), methoxyresorufin
(methoxyresorufin O-dealkylase, MROD), and dibenzylfluorescein (DBF), were found to be significantly (P < 0.001) reduced at micromolar levels. A kinetic
analysis showed competitive and non-competitive modes of inhibition by
limonoids, on CYP19 hydroxylase activity. The results corroborate the active role
of limonoids in the redox cycling mechanisms.
The third study examined the antioxidant and apoptotic inducing ability of
limonoid glucosides on human neuroblastoma cells. Four limonoid glucosides,
LG (17beta-D glucopyranoside limonin), OG (obacunone 17beta-D glucopyranoside),
NAG (nomilinic acid 17beta-D glucopyranoside), and DNAG (deacetylnomilinic acid
17beta-D glucopyranoside), have shown superoxide scavenging at millimolar
levels. Micromolar amounts of LG and OG induced rapid necrosis of SH-SY5Y
cells. Cytotoxicity was correlated (P = 0.046) to a concentration and timedependent
increase in caspase 3/7 activity. Analyses of DNA content during the
S phase of the cell cycle indicated reductions of 86.6% for LG and 82.3% for OG
as compared to untreated. The results validate the antineoplastic distinctiveness
of limonoid glucosides.
In the fourth study, cytotoxic effects of limonoid aglycones and glucosides
were assessed on human SH-SY5Y neuroblastoma and colon carcinoma
(CaCo-2) cell lines and compared with the non-cancerous Chinese hamster
ovary (CHO) cells. Significant (P < 0.001) cytotoxic effects were observed only
on cancerous cells, over 24 to 36 h. The study revealed a marked increase in
the DNA content of aneuploidic cells, which results in cell cycle arrest. The
results confirm that glycosides are the most active apoptotic inducing form.
| Identifer | oai:union.ndltd.org:tamu.edu/oai:repository.tamu.edu:1969.1/4946 |
| Date | 25 April 2007 |
| Creators | Poulose, Shibu M. |
| Contributors | Patil, Bhimanagouda S., Pike, Leonard M. |
| Publisher | Texas A&M University |
| Source Sets | Texas A and M University |
| Language | en_US |
| Detected Language | English |
| Type | Book, Thesis, Electronic Dissertation, text |
| Format | 2468193 bytes, electronic, application/pdf, born digital |
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