Industrial Crops and Products 74 (2015) 585591

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Industrial Crops and Products

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Inhibition of DNA oxidative damage and antimutagenic activity by

dichloromethane extract of Brassica rapa var. rapa L. seeds
Robin, Saroj Arora, Adarsh Pal Vig
Department of Botanical & Environmental Sciences, Guru Nanak Dev University, Amritsar, Punjab, 143005, India

a r t i c l e i n f o a b s t r a c t

Article history: The current study was designed to evaluate the inhibition of DNA oxidative damage and antimutagenic
Received 4 February 2015 activity by dichloromethane extract of Brassica rapa var. rapa L. (Turnip) seeds belonging to family Bras-
Received in revised form 5 May 2015 sicaceae, via DNA oxidative damage assay and Ames assay. The hydroxyl radical mediated oxidative
Accepted 18 May 2015
damage of DNA was effectively inhibited by the seed oil extract at 1000 g/ml concentration. The extract
was assessed for its antimutagenic activity against direct acting mutagens (sodium azide and 4 nitro-
Keywords:
o-phenylenediamine) and indirect acting mutagen (2-aminouorene), with TA100 and TA98 strain of
Brassica rapa var. rapa L.
Salmonella typhimurium. It was found that there was more signicant antimutagenic effect against S9
Turnip
Seed oil
dependent indirect acting mutagen than direct acting mutagens. The results showed better desmutagenic
DNA oxidative damage effect with pre-incubation mode of experimentation, as compared to the co-incubation mode, with lowest
Ames assay value of IC50 0.0008 g/0.1 ml/plate with TA98 strain against S9 dependent mutagen. The GCMS analysis
revealed the presence of two major compounds, namely phenylethylbrassinin and 3-phenylpropionitrile,
in the seed oil extract. These compounds might be the main contributors for inhibition of hydroxyl radical
mediated DNA strand breaking and antimutagenic effect by extract. The study indicates the presence of
strong antimutagenic factors and hydroxyl radical scavengers in Turnip seeds enabling its possible thera-
peutic effect. Also, to the extent of our knowledge this is the rst study that enlightens the antimutagenic
effects of extracted seed oil from B. rapa var. rapa L.
2015 Elsevier B.V. All rights reserved.

1. Introduction antioxidants and nutrients can helps in suppressing these genetic

Acquisitions of genetic mutations are the source of sequential
alteration in genetic makeup. These genetic alterations may lead to
the transformation of normal cells to cancerous form. Nowadays,
due to the increasing prevalence of cancer cases throughout the
world, there is an urgent need to overcome this issue through effec-
tive repair of DNA damage. These mutations can be triggered upon
DNA through induction of oxidative stress. Under oxidative stress,
DNA is the most signicant target and this damage contributes to
the expansion of cancer cells of breast, colon, rectum and prostate
(Halliwell and Gutteridge, 1999; Ames et al., 1993). DNA oxida-
tive damage enhances the chances of mutagenicity and may trigger
the expansion of cancer (Devasagayam et al., 2004). DNA damage
may vary among individuals and its repair depends upon genetic
and dietary inuences (Halliwell, 1998; Kasai, 1997; Rehman et al.,
1999; England et al., 2000; Van Zeeland et al., 1999; Halliwell, 1999;
Bianchini et al., 2000; Benhamou and Sarasin, 2000). So, diet rich in
Corresponding author. Tel.: +91 9417062796.
E-mail address: dr.adarshpalvig@gmail.com (A.P. Vig).
alterations through, prevention of mutations and helps in overcom-
ing the oxidative stress which can boost the effective DNA repair.
On the basis of mode of action, natural inhibitors of mutagenesis can
be bioantimutagenic and desmutagenic (Hung et al., 2009). Bioan-
timutagen stops the mutation, after damage of genes by mutagen
and desmutagen stops mutation before the mutagens effects on
genes.
The Brassicaceae family has been extensively studied in above
mentioned aspects. The plants belonging to this family are grown
and consumed worldwide because of their high fat, protein and
nutritive value for human and animal consumption (Jahangir et al.,
2009). The phytoconstituents in this family, such as indole phy-
toalexins, phenolics and glucosinolates, has been widely studied for
their biological activities against oxidative damage, carcinogenic
and cardiovascular diseases (Jahangir et al., 2009). Among these
the compounds of interest are the one belonging to the category
of glucosinolates. The importance has been due to their capac-
ity to exert protective effects either by impairing bioactivation of
promutagen in the presence of cytochrome P450 (Phase I) or by
induction of detoxication enzymes (Phase II) that act as chemo-
protectors against cancer (Fenwick et al., 1982, 1983, 1990; Carlson

http://dx.doi.org/10.1016/j.indcrop.2015.05.038
0926-6690/ 2015 Elsevier B.V. All rights reserved.
586 Robin et al. / Industrial Crops and Products 74 (2015) 585591
et al., 1987; Rampal et al., 2010; Nestle, 1997; Fahey et al., 1997). In
plants of this family, enzyme myrosinase catalyzes the hydrolysis
of glucosinolate and cleaves off glucose group to form isothio-
cyanates (mustard oils), nitriles, epithionitriles thiocyanates and
oxazolidines (Rask et al., 2000; Bones and Rossiter, 2006; Burow
et al., 2007; Taveira et al., 2009). These compounds are formed upon
tissue disruption from protruding attack of herbivores or pathogens
are known as phytoanticipins (Morant et al., 2008). In reality phy-
toanticipins are toxic, but have been proved potentially benecial
to human health at very low concentrations and are of major inter-
est for the strengthening cell defenses against many diseases (Ames
et al., 1990).
In the present study we have used Brassica rapa var. rapa L.,
commonly known as Turnip. It is a root vegetable food crop grown
in temperate regions worldwide. The plant has been known for its
utilization in the production of edible/industrial oils (Taveira et al.,
2009). Since prehistoric times Turnip has been used for human con-
sumption and considered as one of the oldest food crop (Liang et al.,
2006a,b). For industrial use, the plant roots have been utilized for
the economical high yield of peroxidase enzyme (Musthapa et al.,
2004). These peroxidase enzymes help in the removal of organic
pollutants from wastewater and are also used to catalyze chemi-
cal reactions for the synthesis of various industrial products. The
seeds of this plant have been utilized for the production of canola
(oil with low erucic acid and glucosinolates level), to make it safe for
human consumption. The oils from the seeds have been also used
for, soap manufacturing, lamp oil, animal feed, biodiesel production
and industrial lubricants (Young-Mathews, 2012).
Although glucosinolates are studied widely for its therapeutic
values but, pertaining to meager studies on antimutagenic effects,
the present study aims to extract the oil from seeds of B. rapa var.
rapa L. upon hydrolysis of damaged seed tissue. The study enables
the DNA protective nature and effective mode of action of the
volatile seed oil of B. rapa var. rapa L. against induced mutagenesis
and oxidative stress, by employing Ames assay and DNA oxidative
damage assay. The study may help in providing essential data in
support of its medicinal and pharmaceutical importance.
2. Materials and methods
2.1. Collection of plant seeds
The plant seeds of purple top Turnip (B. rapa var. rapa L.) were
procured from local market of Amritsar, Punjab, India. Seeds were
stored at 25 C without any treatment and washing.
2.2. Extraction with dichloromethane
The extraction of hydrolytic products from seeds was done by
dichloromethane (DCM) using method described by Liang et al.
(2006a,b). The crushed seeds (500 g) of B. rapa var. rapa L. were
allowed to hydrolyze with water (500 ml) at room temperature
for 30 min. Finally, DCM extract was obtained by extraction with
dichloromethane and using 10 g of sodium sulfate to remove traces
of water, followed by drying at 30 C under vacuum on rotary evap-
orator.
2.3. DNA oxidative damage assay
Supercoiled pBR322 plasmid DNA was used to identify the
hydroxyl radical ( OH) scavenging ability of DCM extract using
assay as described by Lee et al. (2002). The destructive effect of OH
on DNA was performed through Fentons reaction. The experiment
was executed with agarose gel electrophoresis using 1% agarose gel
containing 0.50 g/ml ethidium bromide solution. The procedure
involve transfer of 0.5 g of supercoiled DNA in 10 l of Fentons
reagent (30 mM H2 O2 , 50 M ascorbic acid and 80 M FeCl3 ) fol-
lowed by addition of extract at different concentrations (1000, 500,
250, 125 and 62.5 g/ml) and raising nal volume to 20 l using
deionised water. Deionised water and rutin were used as a nega-
tive and positive control, respectively. The mixture was incubated
at 37 C for 30 min, loaded in agarose gel and electrophoresis was
accomplished. Subsequently, the analysis was performed on gel
documentation system (Gel Doc XR, Bio-Rad USA) for radical scav-
enging ability of extract.
2.4. Ames assay
Ames test was performed according to the recommendations of
Maron and Ames (1983) with slight modications given by Bala and
Grover (1989) to test the antimutagenicity of DCM extract using
two strains of Salmonella typhimurium i.e., TA98 and TA100. 4-
Nitro-o-phenylenediamine (NPD, 20 g/0.1 ml/plate) and sodium
azide (NaN3 , 2.5 g/0.1 ml/plate) were used as direct acting muta-
gens for TA98 and TA100, respectively. Different concentrations
(12.5, 25, 50 and 100 g/0.1 ml/plate) of extract were prepared
in dimethyl sulfoxide (DMSO). The extract was analyzed by two
experimental procedures viz. co-incubation and pre-incubation,
to differentiate its bioantimutagenicity and desmutagenicity. The
procedure of co-incubation involves 100 l of bacterial culture,
mutagen and different concentrations of DCM extract in 2 ml of
top agar. Whereas, pre-incubation involves 30 min incubation of
mixture of DCM extract and mutagen in equal volume at 37 C. Sub-
sequently, 200 l of this mixture was mixed in 2 ml of top agar
along with 100 l of bacterial culture. The top agar was evenly
spread on minimal glucose agar plate and revertant colonies were
counted after incubation at 37 C for 48 h. The DCM extract of
B. rapa var. rapa L. was assayed for antimutagenicity in tripli-
cates for each concentration. Simultaneously, spontaneous (100 l
bacterial culture in 2 ml top agar), positive control (100 l bacte-
rial culture and mutagen in 2 ml top agar) and negative control
(100 l bacterial culture and extract concentrations in 2 ml top
agar) were also assessed. Concurrently, the antimutagenicity of
DCM extract was also assayed in the presence of S9 fraction from rat
liver homogenate using 2-Aminouorene (AF, 20 g/0.1 ml/plate)
promutagen. The promutagen gets activated in the presence of
cytochrome based P450 metabolic activation system (Phase I)
present in liver. The post-mitochondrial supernatant S9 fraction
was prepared according to the recommendations of Garner et al.
(1972). The antimutagenicity of DCM extract was checked by using
S9-mix (comprising NADP, MgCl2 , KCl, glucose-6-phosphate, phos-
phate buffer and S9 fraction) in top agar. The procedure involves
same process as already discussed for co-incubation and pre-
incubation previously, only with addition of 0.5 ml of S9-mix in
2 ml of top agar along with other constituents. The antimutagenic-
ity of DCM extract was evaluated as decrease of reverse mutation
in percentage as:
ab
Inhibition (%) = 100
ac
where,
a = his+ revertants induced in presence of mutagen alone (Posi-
tive control).
b = his+ revertants induced by mutagen in presence of extract
concentration.
c = his+ revertants induced with extract concentration in
absence of mutagen (Negative control).
2.5. Gas chromatography-mass spectrometry (GCMS) analysis
Analysis was performed on a Shimadzu GC (model 2010) with a
mass selective detector (model QP2010; Shimadzu) equipped with
 Robin et al. / Industrial Crops and Products 74 (2015) 585591 587

Fig. 1. Effect of DCM extract of Brassica rapa var. rapa L. in DNA oxidative damage assay.
Lane 1: Negative control (deionised water); lane 2: Fentons reagent; lane 3: Positive control (Rutin); lane 48: Fentons reagent + different concentrations of Brassica rapa
var. rapa L. extract (1000, 500, 250, 125 and 62.5 g/ml); form I-supercoiled plasmid DNA; form II-open circular; form III-linear.

DB-5MS capillary column. Helium was used as carrier gas at a ow that there was formation of nicked DNA (form II and III) due to
rate of 1.10 ml min1 and sample injection volume of 2 l. The oven hydroxyl radicals generated through Fentons reaction. However,
temperature of column was programmed initially at 70 C for 5 min, with the addition of different concentration of DCM extract, the
rising at the rate of 4 C/min to 230 C and then held isothermal for OH-mediated strand breaking and conversion of supercoiled DNA

4 min. Ionization voltage of 70 eV, ion source temperature of 200 C
and interface temperature of 250 C was maintained. The spectrum
was scanned from m/z 50 to 500 amu. Individual peaks were iden-
tied by comparing their retention indices (RI), mass spectra (MS)
with literature data, as well as by computer matching against the
Wiley 275-library spectra database.
2.6. Statistical analysis
Statistical analysis was carried out using Microsoft Excel 2007.
The results were expressed as mean standard error. The 2-way
ANOVA was performed for the analysis of differences and interac-
tions at 5% level of signicance by program developed on Microsoft
Excel 2007.
3. Results and discussion
The study assessed the inhibition of oxidative damage and
antimutagenic effects of DCM extract by B. rapa var. rapa L. seeds
and revealed the following results:
3.1. DNA oxidative damage assay
Inhibition of DNA oxidative damage by DCM extract at differ-
ent concentrations viz. 1000, 500, 250, 125 and 62.5 g/ml has
been shown in Fig. 1. It was observed that the extract exhibit very
good radical scavenging activity on pBR322 plasmid DNA with
highest activity at 1000 g/ml concentration. The assay revealed
(form I) to forms II (open circular) and III (linear) was reduced in a
dose dependent manner.
3.2. Ames assay
The DCM extract of B. rapa var. rapa L. was analyzed for inhibi-
tion of mutagenicity of sodium azide and 2-aminouorene in TA100
strain of S. typhimurium in Ames assay, without and with S9 mix,
respectively (Table 1). The DCM extract through TA100 without
S9 mix has been found to show maximum percentage inhibition
(74.5% at 100 g/0.1 ml/plate) as compared to TA98 without S9
mix (Table 1). The extract showed effective antimutagenicity on all
applied concentrations in the assay with pronounced effect in pre-
incubation mode than co-incubation mode of experimentations in
TA100 (Fig. 2). It was also observed by percentage inhibitions and
IC50 values that, assay with S9 mix turn out to be more effective
in the inhibition of mutagenicity than without S9 mix in TA100
(Tables 1 and 2). Simultaneously, the analysis was performed for
inhibition of mutagenicity of 4-nitro-o-phenylenediamine and 2-
aminouorene in TA98 strain of S. typhimurium in Ames assay,
without and with S9 mix, respectively. The extract exhibited
antimutagenicity on all applied concentrations in the assay with
pronounced effect in pre-incubation mode than co-incubation
mode of experimentations in TA98 (Fig. 2). Overall, the obtained
values of IC50 revealed the maximum inhibition of mutagenic-
ity in the assay with lowest value of IC50 (0.0008 g/0.1 ml/plate)
with TA98 (with S9) in pre-incubation mode of experimentation
(Table 2).

Table 1
Percentage inhibition of mutagenecity by DCM extract of B. rapa var. rapa L. seeds in TA98 and TA100 tester strains of Salmonella typhimurium.

Treatment Dose TA 100 (without S9) TA 98 (without S9) TA 100 (with S9) TA 98 (with S9)
/0.1 ml/plate % Inhibition % Inhibition % Inhibition % Inhibition

Co- 12.5 g 16.65 2.12 34.70 2.08 78.59 1.95 74.88 1.23
incubation 25.0 g 25.16 2.85 41.00 0.33 84.79 1.44 79.27 0.96
50.0 g 47.05 2.35 43.16 1.56 93.44 1.64 85.57 0.73
100 g 55.96 1.92 49.86 1.55 98.19 0.65 89.73 0.29

Pre- 12.5 g 19.58 3.43 41.81 1.09 81.98 1.22 86.81 0.52
incubation 25.0 g 29.10 1.30 46.54 0.75 89.42 2.39 90.31 0.63
50.0 g 52.39 0.65 50.45 0.78 94.18 0.33 92.94 0.28
100 g 74.51 1.17 52.75 1.07 99.45 0.75 94.84 0.57
588 Robin et al. / Industrial Crops and Products 74 (2015) 585591

TA100 (without S9) TA100 (with S9)

100 100

Inhibion (%)
Inhibion (%)
80 80
60 60
40 40
20 20
0 0
0 20 40 60 80 100 0 20 40 60 80 100
Conc. (g/0.1 ml) Conc. (g/0.1 ml)

TA98 (without S9) TA98 (with S9)

100
100

Inhibion (%)
80
Inhibion (%)

80
60 60
40 40
20 20
0 0
0 20 40 60 80 100 0 20 40 60 80 100
Conc. (g/0.1 ml) Conc. (g/0.1 ml)

Fig. 2. Antimutagenic effect of DCM extract of B. rapa var. rapa L. seeds in Ames assay.

Table 2
Antimutagenic effect of DCM extract of B. rapa var. rapa L. seeds in Ames assay.

Experimentation mode Regression equation (R2 ) IC50 (g/0.1 ml/plate)

TA100 (without S9) co-incubation y = 20.17ln(x) 35.71 2

R = 0.965 70.0620
TA100 (without S9) pre-incubation y = 27.13ln(x) 52.84 R2 = 0.972 44.2846
TA100 (with S9) co-incubation y = 8.247ln(x) + 61.85 R2 = 0.989 0.6595
TA100 (with S9) pre-incubation y = 9.731ln(x) + 54.05 R2 = 0.988 0.2377
TA98 (without S9) co-incubation y = 6.872ln(x) + 17.67 R2 = 0.967 110.4539
TA98 (without S9) pre-incubation y = 5.298ln(x) + 28.99 R2 = 0.978 52.7544
TA98 (with S9) co-incubation y = 7.335ln(x) + 56.20 R2 = 0.993 0.4294
TA98 (with S9) pre-incubation y = 3.855ln(x) + 77.48 R2 = 0.982 0.0008

Fig. 3. 3-D graphical representation of antimutagenic effect of DCM extract of B.

rapa var. rapa L. seeds between co- and pre-incubation for TA98 and TA100 strain
of S. typhimurium in Ames assay with S9.
The comparison among all experiments in Ames assay, with
and without S9, has been shown in Figs. 3 and 4, respectively.
It was observed that there is less variation in inhibition in Ames
assay performed with S9, as compared to without S9, between
all the concentrations of DCM extract. The comparison enables
the conspicuous activity of DCM extract with S9, even at lower
concentrations, as compared to without S9 experiments. So, the
whole experimentation setup facilitates the noticeable activity of
DCM extract, with S9 mix in pre-incubation mode. The assay was
also examined statistically by two-way ANOVA for the statisti-
cal difference and interaction as shown in Table 3. The analysis
revealed statistically signicant difference among co-incubation
Fig. 4. 3-D graphical representation of antimutagenic effect of DCM extract of B.
rapa var. rapa L. seeds between co- and pre-incubation for TA98 and TA100 strain
of S. typhimurium in Ames assay without S9.
and pre-incubation mode of experimentations, as well as
between different concentrations applied in the Ames assay at
5% level of signicance. Also, the experimentation with TA100
(without S9) and TA98 (with S9) displayed statistically signi-
cant interaction among experimentation mode (i.e., co-incubation
and pre-incubation) and DCM extract doses (12.5, 25, 50 and
100 g/0.1 ml/plate) on the antimutagenic effect at 5% level of sig-
nicance. Therefore, conclusively the DCM extract of B. rapa var.
rapa L. showed, pronounced antimutagenicity activity in Salmonella
histidine point mutation assay.
 Robin et al. / Industrial Crops and Products 74 (2015) 585591 589

Table 3
Two-way ANOVA analysis of DCM extract of B. rapa var. rapa L. seeds in Ames assay.

Two-way ANOVA analysis Degree of F-ratio for TA100 F-ratio for TA100 F-ratio for TA98 F-ratio for
freedom (df) (without S9) (with S9) (without S9) TA98 (with S9)

Difference between co-incubation and pre-incubation experimentation 1 25.50* 06.89* 40.78* 310.68*
Difference between DCM extract doses or concentrations applied 1 197.79* 71.59* 37.59* 99.66*
Interaction among experimentation mode and doses applied 4 5.76* 00.91 01.30 10.03*

Error df = 16.
*
p 0.05.

Fig. 5. GCMS chromatographic prole of volatile components of DCM extract of B. rapa var. rapa L.

3.3. Gas chromatography-mass spectrometry (GCMS) analysis The present study emphasizes the DNA protective effect of DCM
extract of B. rapa var. rapa L. seeds. The antimutagenic effect of seed
The volatile components in DCM extract of B. rapa var. rapa extract has been observed against mutagenesis induced by direct
L. seeds were determined by GCMS analysis as shown in Fig. 5. (sodium azide and 4-nitro-o-phenylenediamine) and indirect (2-
Total sixteen compounds were detected with phenylethylbrassinin aminouorene) acting mutagens. Two strains of S. typhimurium
(phenylethyl isothiocyanate) and 3-phenylpropionitrile as major i.e., TA100 and TA98 were utilized to recognize antimutagenic
compounds, which account for the presence of phytoanticipins in nature of extract against base pair substitution and frameshift
the extract (Table 4). The compounds are hydrolytic products of glu- mutations, respectively. The study reveals that the desmutagenic
cosinolates and might be accountable for notable effective activity effect prevails conspicuously among all applied concentrations of
of the seed oil extract of B. rapa var. rapa L. in inhibition of DNA DCM extract against both mutations. This enables the effective
oxidative damage and antimutagenicity. inhibition by extract against mutagenesis, before the mutation
on genes occurred, by direct inactivation of the mutagen. How-
Table 4
ever, more prominent inhibition (74.5%) was observed with TA100
Volatile compounds in the DCM extract B. rapa var. rapa L. detected by GCMS. strain having base pair substitution mutation in the assay without
S9 mix at 100 g/0.1 ml/plate concentration. Also, in the presence
Peak R. Time Area (%) Name
of cytochrome P450 the promutagen (2-aminouorene) activation
1. 5.872 0.53 Dimethyl trisulphide was prevented in experimentations with S9 mix in both strains
2. 6.135 0.47 1-Decene
of S. typhimurium. But overall the Ames assay with TA98 (with
3. 9.347 4.73 4,5-Dimethylthiazole
4. 10.175 2.28 Adacene 12 S9) in pre-incubation mode of experimentation showed lowest
5. 10.660 0.79 2,9-Dithiadecane IC50 value enabling its highest antimutagenic ability. The inhibi-
6. 10.906 0.52 Dimethyl tetrasulde tion by DCM extract might be due to either by preventing the
7. 11.404 12.10 3-Phenylpropionitrile conversion to mutagen or by inactivating the mutagenic form
8. 11.551 7.90 1H-Cyclopenta[c]thiophene, hexahydro-, cis-
9. 12.569 1.59 3-Methyl-2-cyclopentenone
by binding with it. The observations in the present study are in
10. 13.420 7.45 1-Tetradecene agreement with the earlier reports of Rampal et al. (2010). The
11. 14.817 12.91 Phenylethylbrassinin study observed the antimutagenic effect of seed extracts of Bras-
12. 15.519 5.51 N.D. sica oleracea L. var. italica Plenck (Broccoli) against direct acting
13. 15.877 1.00 N.D.
mutagens (sodium azide and 4-nitro-o-phenylenediamine) and
14. 16.095 0.74 1-Tetradecene
15. 16.169 11.84 1-Hexadecene indirect acting mutagen (2-aminouorene) with TA100 and TA98
16. 18.597 11.45 1-Heptadecene strains. The results showed overall better antimutagenic activity of
17. 20.841 8.81 1-Nonadecene 0.1 mol/L HCl extract against frameshift mutation with TA98 strain
18. 23.724 5.26 1-Tricosene at 1000 g/0.1 ml/plate concentration. The study indicated that
19. 28.374 3.04 N.D.
20. 36.551 1.08 N.D.
nitriles formed at lower pH predominates the antimutagenic activ-
ity against mutagen 4-nitro-o-phenylenediamine. Earlier, Hecht
N.D. = Not detected.
590 Robin et al. / Industrial Crops and Products 74 (2015) 585591
et al. (1999) reported on clinical trial of smokers that phenylethyl
ITC rich diet inhibited the cytochrome P450-mediated bioactivation
of procarcinogen. The study concluded that watercress consump-
tion of 170 g/d by smokers reduced the activation of procarcinogen
in Tobacco. The current study enables to demonstrate the reticence
of oxidative DNA damage against OH radicals. The study revealed
the protective nature of extract by inhibiting the nicking of plasmid
DNA. The extract effectively conserved the supercoiled form of DNA
at 1000 g/ml concentration. Manesh and Kuttan (2003) reported
the hydroxyl radical scavenging effect of allyl and phenyl ITCs in
deoxyribose degradation assay. At 25 g/ml concentration allyl and
phenyl ICTs exhibited 61.9% and 69.7% inhibition of hydroxyl radical
production. The observations in the present study are in accor-
dance with previous reports on hydroxyl radical scavenging activity
of extract. The GCMS analysis has showed phenylethyl isothio-
cyanate and 3-phenylpropionitrile as major compounds in the seed
extract. The phenylethyl isothiocyanate along with the combina-
tion of other constituents in the extract may help in the inhibition
of hydroxyl radicals. Gluconasturtiin (phenethyl glucosinolate) is
the precursor of both the compounds. The presence of both com-
pounds might be responsible for the DNA protective nature of seed
extract. The phenylethyl ITC has been widely studied for its chemo-
prevention against carcinogenesis (Wu et al., 2009). Phenylethyl
ITC and their cysteine conjugates inhibit the in vitro growth and
induce apoptosis of human leukemia and myeloblastic leukemia-1
cells (Wu et al., 2009). Study suggests that phenylethyl ITC exhib-
ited less toxicity to surrounding cells and cytotoxicity was more
selective towards leukemia cells. Presence of both nitrile and isoth-
iocyanate in the extract may render the promising antimutagenic
and DNA protective nature of seed extract. The results facilitate the
possible therapeutic effect of the DCM extract and further studies
will establish the mechanistic pathways to these effects.
4. Conclusion
In present study, it has been found that the oil extract of B.
rapa var. rapa L. seeds exhibits strong inhibition of DNA oxidative
damage from OH radicals and signicant antimutagenicity against
frameshift and base pair mutations. Conclusively, the study support
therapeutic effect of the B. rapa var. rapa L. and further investiga-
tions are required to unravel its chemoprotective effects against
carcinogenesis.
Conicts of interest
The authors declare no conict of interest.
Acknowledgment
The authors gratefully acknowledge University Grants Com-
mission (UGC) for nancial support and Dr. Bikram Singh, CSIR-
Institute of Himalayan Bioresource Technology (IHBT), Palampur
(H.P.), India, for the GCMS analysis of the extract. Also, the authors
would like to acknowledge Prof. A. K. Thukral, Department of
Botanical and Environmental Sciences, Guru Nanak Dev Univer-
sity, Amritsar (Punjab), India, for the development of 2-way ANOVA
program for statistical analysis.
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