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Medicinal Plant Research 2014, Vol.4, No.3, 30
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2 Discussion
Antioxidant capacity of
P. peruviana
was reported
previous studies (Wu et al., 2006; Ramadan and
Moersel 2007; Vasco et al., 2008; Puente et al., 2011).
It was demonstrated that
P. peruviana
leaves have
potent antioxidant activity (Wu et al., 2006). Ramadan
and Moersel (2007) investigated the antioxidant
activity of juice of
P. peruviana
with respect to
reaction time with DPPH. They noted that the
antioxidant capacity of this fruit is related to its
fat-soluble bioactive contents such as tocopherols,
sterols and carotenoids. Vasco et al. (2008) analysed
antioxidant capacities of different types of major fruits
from Ecuador and determined that
P. peruviana
has
antioxidant activity at low level among other fruits. In
this study, observations showed that antioxidant
activity of
P. peruviana
fruits’ crude extract is at low
level.
Beside its antioxidant capacity, IC
50
values of
P.
peruviana
for 48 h for SaOS-2, HT-29, Hep3B and
SH-SY5Y cells are 15.44, 40.79, 24.92 and 44.24
µg/mL respectively. Wu et al. (2004) evaluated
anticancer activity of
P. peruviana
extracts (whole
plant) on Hep3B and HepG2 cell lines and they have
reported a less cytotoxic value (IC
50
41.25±1.40 µg/ml)
for Hep3B cell line. According to these results
P.
peruviana
fruits’ crude extract is more cytotoxic than
the extract from whole plant.
Mechanism of apoptosis’ induction in HepG2 cells has
been also studied by Wu and co-workers (2004). The
inhibition pathways for other cell lines used in our
study should also be studied in molecular level
because non-cancerous cell line Vero, was not affected
with all mentioned concentrations with two cancer cell
lines, MCF-7 and LNCap.
For further studies, different extraction methods such
as; supercritical CO
2
extraction may be studied for
P.
peruviana
fruit. The extraction efficiency may vary
depending on extraction technique. Different extra-
ction methods can also result in the difference pattern
of the active compounds may affect the capacity of the
pharmacological properties.
Additionally active molecules, which take place in
molecular mechanism of cytotoxicity needs to be
clarified for used in this experiment and other cancer
cell lines besides normal cell lines. Controversially
after 24 h treatment, induction of the proliferating
cancer cells has to be studied (data not given) also.
Importance of this effect is related with the possibility
of
P. peruviana
extracts’ medical usage. Purification
of active molecules from raw fruit extract will help to
determine reasons of cytotoxic and proliferative
effects.
3
Material and Methods
3.1 Sample preparation and extraction
Fruits of
P. peruviana
were purchased from a local
supplier from Antalya in March 2012. Fruits were
washed and air-dried before the extraction step. Two
hundred grams of fruits were homogenized in a
domestic blender and extracted with 600 mL of 80%
ethanol at 28°C at 150 rpm using an orbital shaker
overnight. The extract was filtered with Whatman No
1 filter paper and the residue was re-extracted with the
same conditions day long. After pooling all filtrates,
the solvent was evaporated under vacuum at 40°C and
remaining sample was freeze-dried. (Moisture content
of fruits was determined as 82%). The extract was
stored at
-
20°C until use.
3.2 DPPH radical scavenging activity
Antioxidant activity of the extract was determined
according to Rodriguez et al. (2011). This assay is
used for determining the antioxidant capacity of
compounds by testing the ability of them to act as free
radical scavengers or hydrogen donors (Reddy et al.,
2010). The colour of DPPH (1,1-diphenyl-2-
picrylhidrazil) solution turns from purple into yellow
because of forming the reduced DPPH-H by pairing
with a hydrogen from a free radical scavenging
antioxidant. Ascorbic acid was used as a standard
compound. Different concentrations of extract and
standard were prepared with 80% methanol. 100 µL of
samples were mixed with 2.9 ml of 60 µM solution of
DPPH (Sigma Aldrich, Germany) reagent in 80%
methanol. The reaction mixture was allowed to stand
at room temperature for 30 minutes in dark.
Absorbance was measured at 515 nm using
spectrophotometer (Jenway 6400 Bibby Scientific,
UK) against 80% methanol solution as blank. Each