The document summarizes a study that investigated the phytochemical screening, antioxidant, and anti-inflammatory activities of different extracts from the leaf, stem, and bark of Tectona grandis. Qualitative phytochemical analysis revealed the presence of compounds like phenols, saponins, steroids, and terpenoids in the leaf and stem extracts. The leaf, bark, and stem extracts showed antioxidant activity in Ferric Reducing Antioxidant Power and DPPH radical assays. The extracts also demonstrated anti-inflammatory effects by inhibiting albumin denaturation and stabilizing red blood cell membranes in heat-induced hemolysis tests. The study thus supports the traditional use of T. grandis in treating inflammation

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Available Online at: www.ijrpp.com Print ISSN: 2278 - 2648
_________________________________
* Corresponding author:
*1Shruthi D.P
Department of Biotechnology,
Shridevi Institute of Engineering & Technology,
Sira Road, Tumkur-572 106, Karnataka, INDIA.
E-mail: shruthidavasam@gmail.com
Online ISSN: 2278 - 2656
International Journal of
Research in Pharmacology and
Pharmacotherapeutics
(Research article)
PHYTOCHEMICAL SCREENING, ANTIOXIDANT AND ANTI-INFLAMMATORY
ACTIVITY OF DIFFERENT EXTRACTS
FROM LEAF, STEM AND BARK OF TECTONA GRANDIS
*1Shruthi D.P, 2Sunith K.E, 3Haritha Kumari E, 4Govindappa M, 5Siddalingeshwara K.G
1Department of Biotechnology, Shridevi Institute of Engineering & Technology, Sira Road,
Tumkur-572 106, Karnataka, India.
5Department of Biochemistry, Tumkur University, Tumkur, Karnataka, India.
_________________________________________________________________________
ABSTRACT
Interest in natural products as a source for innovation in drug discovery and agrochemicals is still growing
worldwide. Natural products, whose immense diversity has been appreciated for many years, may become in a
rich source of novel chemical structures. Our country is a rich source of both biological and chemical diversity,
which may be useful as a source of novel chemical structures. The timber value of Tectona grandis has been
well known from decades. Teak is the major exotic species found in tropical regions. The present study was
meant to characterize pharmacological potential of different extracts from leaf, bark and stem of teak. The aim
of present study to investigate the preliminary phytochemical screening of Tectona grandis. This work
highlights on qualitative phytochemical investigation on leaf, bark and stem. Of the Tectona grandis. A
comparative phytochemical analysis was carried to prove that the amount of phyto constituents varied with the
fresh and dry stages of plants contributing to the activity of the extract. Total phenolic and saponnins content of
both the extracts was estimated and was found to be more in the leaf, bark and stem giving positive results and
the presence of steroids and Terpenoids also showed in leaf and stem. Antioxidant activity of extracts was
carried out using ferric ion reducing antioxidant power (FRAP) and 2, 2-diphenyl-1- picrylhydrazyl (DPPH)
assay and anti-inflammatory activity of the Tectona grandis extract were also made.
KEY WORDS: Antioxidant, anti-inflammatory activity, Tectona grandis, phytochemical screening.
INTRODUCTION
Plants are the basis for traditional medicine systems
and used for thousands of year in countries such as
India and China. Numbers of medicinal plants and
its derived extracts are used in the treatment of
various disorders by Ayurveda, Unani and Siddha
systems in India. Scientifically, few of
pharmacological properties have been studied to
support its traditional use 1. Throughout the world,
many plant species are used for the treatment of
inflammation and other diseases. Despite the
availability of anti-inflammatory and analgesic
agents searching of newer therapeutic agent in this
segment from the natural plants is still progressing
due to presence of diverse chemical substances that
have a better alternative and safer effect on
inflammation without or lesser side effects. The
World Health Organisation (WHO) estimates that
80% of the world’s inhabitants continue to rely on
traditional medicines systems and its products 2.
Tectona grandis belonging to the family
Verbinaceae is commonly called as teak. The
various parts of the plants are reported to posses

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various activities. Some of the activities include its
action as a cooling agent, laxative and sedative,
bronchitis, as diuretic and in the treatment of
urinary discharge, in the treatment of the common
cold and headache, hair problems and in scabies 3, 4.
The various phyto constituents isolated from
Tectona grandis are Juglone, which has been
reported to posses anti-microbial activity 5, Betulin
aldehyde is reported to posses anti tumor activity 6,
Lapchol shows anti ulcerogenic activity 7. The
present work was designed to give a detailed
picture of antioxidant and anti inflammatory
potentiality of extracts from different parts of
Tectona grandis and also qualitative studies of
photochemistry of the plants were studied.
MATERIALS AND METHODS
Plant materials collection and identification
Plant material was collected from the campus of
Shridevi Institute of Engineering and Technology,
Tumkur, Karnataka, India. The collected plant was
authenticated from the Department of Botany,
Manasa Gangotri, University of Mysore, Mysore,
Karnataka, India and Government Ayurvedic
College, Mysore, and herbarium was prepared.
Extraction of plant material
The fresh and dry T. grandis plant material were
powdered and extracted with distilled water at room
temperature for 24 h. and filtered to the help of
Whatman No. 1 and fresh and dry plant materials
were also used for extraction by 90% methanol
using soxhlet apparatus for 72 h. All extracts were
concentrated under reduced pressure using rotary
evaporator and stored at 2 - 8 0C until the
completion of qualitative phytochemical studies,
antioxidant and ant-inflammatory activity.
Antioxidant activity
Antioxidant activity of extracts was carried out
using ferric ion reducing antioxidant power
(FRAP) and 2, 2-diphenyl-1- picrylhydrazyl
(DPPH) assay.
FRAP assay:
FRAP reagents was freshly prepared by mixing 25
ml acetate buffer (300 mM, pH 3.6), 0.5 mL 2,4,6-
tris (2-pyridyl)-S-triazine (TPTZ) solution (10 mM
TPTZ in 40 mM/l HCl) and 2.5mL FeCl3 (20 mM)
water solution. Each sample (150 μl) (0.5 mg/ml)
dissolved in methanol was added to 4.5 mL of
freshly prepared FRAP reagent and stirred. After 5
min, absorbance was measured at 593 nm, using
FRAP working solution as blank 8,9. A calibration
curve of ferrous sulfate (100 to1000 μmol/l) was
used and results were expressed in μmol Fe2+/mg
dry weight extract. The relative activity of the
samples was compared to L-ascorbic acid.
DPPH radical assay:
The effect of plant extracts on DPPH radical was
estimated using the method of Liyana-Pathirana
and Shahidi 10. DPPH solution was freshly prepared
by dissolving 24 mg DPPH in 100 ml methanol,
stored at -20°C before use. 10 μl of the sample was
added to 140 μl distilled water and allowed to react
with 2850 μl of DPPH reagent (190 μl reagent +
2660 μl distilled water) for 24 h in the dark
condition. Absorbance was measured at 515 nm. A
linear standard curve between, 25 to 800 μM
ascorbic acid, was obtained and expressed in μm
AA/g fresh mass. Additional dilution will be
needed if the DPPH value measured is over the
linear range of the standard curve; mix 10 ml of
stock solution in a solution of 45 ml of methanol, to
obtain an absorbance of 1.1 ± 0.02 units at 517 nm
using spectrophotometer 11. All determinations
were performed in triplicate. The percentage
inhibition of DPPH radical by the samples was
calculated according to formula of Yen and Duh 12,
% inhibition= [{Abs control - Abs sample}/Abs
control] x 100,
Where Abs control is the absorbance of the DPPH
radical + ethyl acetate, Abs sample is the
absorbance of DPPH radical+ sample
extract/standard.
Anti-inflammatory activity
Inhibition of albumin denaturation
Methods of Mizushima and Kobayashi 13 and Sakat
et al. 14 followed with minor modifications. The
reaction mixture was consisting of test extracts and
1% aqueous solution of bovine albumin fraction,
pH of the reaction mixture was adjusted using
small amount at 37 0C. The extracts incubated
(20min) and then heated to 51 0C the samples the
turbidity was measured spectrophotometrically at
660nm. The experiment was performed in
triplicate. Percent inhibition of protein denaturation
was calculated as fallows.
% inhibition= [{Abs control- Abs sample}/Abs
control] x 100,
Where Abs control is the absorbance without
sample, Abs sample is the absorbance of sample
extract/standard.
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Membrane stabilization test
Preparation of red blood cells (RBCs)
suspension
Fresh whole human blood (10ml) was collected and
transferred to the centrifuge tubes 14. The tubes
were centrifuged at 3000 rpm for 10min and were
washed three times with equal volume of normal
saline. The volume of blood was measured and re
constituted as 10% v/v suspension with normal
saline.
Heat induced hemolytic
The reaction mixture (2ml) consisted of 1 ml of test
sample solution and 1 ml of 10% RBCs suspension,
instead of test sample only saline was added to the
control test tube. Aspirin was used as a standard
drug. All the centrifuge tubes containing reaction
mixture were incubated in water bath at 56⁰C for
30min. At the end of the incubation the tubes were
cooled under running tap water. The reaction
mixture was centrifuged at 2500 rpm for 5 min and
the absorbance of the supernatants was taken at 560
nm 14. The experiment was performed in triplicates
for all the test samples. Percent membrane
stabilization activity was calculated by the formula
mentioned below.
Protein inhibitory action
The test was performed according to the modified
method of Oyedepo et al.15and Sakat et al.14. The
reaction mixture (2ml) was containing 0.06mg
trypsin, 1ml of 20mM Tris HCl buffer (pH7.4) and
1ml test sample of different concentrations of
different solvents. The reaction mixture was
incubated at 37⁰C for 5min and then 1ml of 0.8%
(W/V) casein was added. The mixture was
inhibited for an additional 20 min, 2ml of 70%
perchloric acid was added to terminate the reaction.
Cloudy suspension was centrifuged, and the
absorbance of the supernatant was read at 210nm
against buffer as blank. The experiment was
performed in triplicate. The percentage of
inhibition of proteinase inhibitory activity was
calculated.
Xanthine oxidase assay
Xanthine oxidase activity was assayed
spectrophotometrically at 300 nm as described by
Yamamoto et al. 16. Briefly, the reaction mixture
consisting of 500 μl of solution A (0.1Mphosphate
buffer containing 0.4mM xanthine and 0.24 mM
NBT), 500 μl of solution B (0.1 M phosphate
buffer containing 0.0449 units/ml xanthine
oxidase) and 50 μl of a 10% of each solvent
extracts were incubated in a cuvette at 37 0C for 20
min. The enzyme activity was expressed as the
increment in absorption at 300 nm per unit time.
RESULTS AND DISCUSSION
Phytochemical screening
Phytochemical analysis of methanol and water
solvent extract for saponins, phenolic compounds,
anthraquinones, steroids, terpenoids, tannins and
alkaloids are done. Plant extracts have all the
phytochemical except anthraquinones and
alkaloids. The presence of saponnins and phenolic
compounds was observed in higher amount than
compared to steroids, tannins and terpenoids. The
phytochemical studies were presented in
Table 1 and 2 with solvent extract.
Table 1: Phytochemical analysis of different plant parts (20ml distilled water)
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Tests
Leaf Stem Bark
Fresh dry Fresh dry Fresh dry
Saponins +ve +ve +ve +ve +ve -ve
Phenols +ve +ve +ve +ve +ve -ve
Anthaquinones -ve -ve -ve -ve -ve -ve
Steroids +ve -ve +ve -ve -ve -ve
Tannins +ve +ve -ve +ve -ve -ve
Terpenoids +ve +ve -ve +ve -ve -ve
Alkaloids -ve -ve -ve -ve -ve -ve

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Table 2: Phytochemical analysis of different plant parts (99% methanol
Antioxidant activity of Tectona grandis
extract
Antioxidant status of all extracts was checked by
DPPH and FRAP free radical. Its ability to
scavenge those free radicals at different
concentrations was analysed. Plant at a
concentration of 0.1mg/ml, the scavenging activity
of the endophytes reached at high concentration,
shows the dose response curve of DPPH radical
scavenging activity of ethyl acetate of plant extract
(leaf, stem and flower). The study on antioxidant
activity of Tectona grandis with its crude ethanol
extracts by H2O2 scavenging activity, DPPH and
FRAP proved its potential. The fig 1 and Table 3
will show the DPPH and FRAP antioxidant
activity.
Figure 1. DPPH scavenging activities
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Tests
Leaf Stem Bark
Fresh dry Fresh dry Fresh dry
Saponins +ve +ve +ve +ve +ve -ve
Phenols +ve +ve +ve +ve +ve +ve
Anthaquinones -ve -ve -ve -ve -ve -ve
Steroids +ve +ve +ve -ve -ve -ve
Tannins +ve +ve -ve +ve -ve -ve
Terpenoids +ve -ve -ve +ve -ve -ve
Alkaloids -ve -ve -ve -ve -ve -ve
% of inhibition
Concentration in μg/ml

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Table 3.Total antioxidant (FRAP) activities
Extracts FRAP
Leaf Fresh 1623.21±0.06a
Dry 1432.64+0.08d
Stem Fresh 810.14±1.2f
Dry 773.32+1.4g
Flower Fresh 1611.26+0.06b
Dry 1368.57±0.09e
Ascorbic acid 1648.52±0.06c
BHT 64.84±1.5h
The total phenolic content of the metabolites of
host plant extract are shown and they had the
highest total phenolic content (TPC). The Phenolic
compounds in the plant extract may have
contributed significantly to their antioxidant
activity.
Anti inflammatory properties
Inhibition of albumin denaturation
Denaturation of proteins is a well documented
cause of inflammation. As part of the investigation
on the mechanism of the anti-inflammation
activity, ability of different solvent plant extract
protein denaturation was studied. It was effective in
inhibiting heat induced albumin denaturation.
Maximum inhibition 89.61% was observed from
methanol extract followed by ethanol 86.81% and
water 51.14%. All the solvent extracts inhibited the
albumin denaturation, the methanol extract stood
first compared to ethanol and water extracts.
Aspirin, a standard anti-inflammation drug showed
the maximum inhibition 75.89% at the
concentration of 200 μg/ml. Repeated the
experiments three times for each replicates and
results were presented in Table. 4. According to
Duncan’s Multiple Range Test (DMRT), values
followed by different subscripts are significantly
different at P<0.05, SE-standard error of the mean.
Repeated the experiments three times for each
replicates, According to Duncan’s Multiple Range.
Test (DMRT), values followed by different
subscripts are significantly different at P<0.05, SE-Standard
error of the mean.
Statistical analysis
Analysis of variance (ANOVA) was used to
determine the significance of difference between
treatment groups (p < 0.05). Means between
treatment groups were compared for significance
using Duncan’s new Multiple Range post test.
Proteinase inhibitory activity
The T.grandis different solvent extracts exhibited
significant antiproteinase activity represented in
Table 4. The maximum inhibition was observed
from methanol extract (83.91%) in decreasing
order was ethanol (81.17%) and water (61.73%).
The methanol and ethanol extract have showed
highest proteinase inhibitory activity compared to
water extract. The standard drug aspirin have
showed the maximum proteinase inhibitor activity
is 92.83%
Table 4. Effect of water extracts on albumin denaturation, membrane stabilization and proteinase
inhibitory activity Xanthane oxidase percentage inhibition
Test sample Albumin
denaturation
Membrane
stabilization
Proteinase
inhibition
Xanthane oxidase
Fresh extract
Leaf 89.61±0.06a 78.82±0.04b 83.91±0.03b 41.13±0.07f
Bark 51.14±0.08e 52.31±0.06d 64.84±0.06d 36.92±0.08f
Stem 86.81±0.06b 76.65±0.05b 81.17±0.03c 38.97±0.08f
Dry extract
Leaf 86.26±0.06b 74.63±0.06c 79.33±0.06c 40.16±0.07f
Bark 48.22±0.07f 50.72±0.06d 64.84±0.06d 34.86±0.08f
Stem 82.11±0.06c 72.46±0.04d 77.55±0.05c 36.75±0.08f
Aspirin (200μg/ml) 75.89±0.06d 85.92±0.02a 92.83±0.03a 95.96±0.03f

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Xanthine oxidase assay
The maximum inhibition of xanthine oxidase was
observed from methanol extract (41.13%) followed
by ethanol (38.97%) and water (36.72%).
Maximum inhibition was noticed in methanol and
ethanol extracts compared to the water extract. The
standard drug aspirin have showed the maximum
proteinase inhibitor activity is 95.96% (Table 4).
CONCLUSION
In this study results indicate that the different
solvent extracts of T.grandis possess antioxidant
and anti-inflammatory properties. These activities
may be due to the strong occurrence of
polyphenolic compounds such as tannins, steroids,
phenols, terpenoids and Saponins. The extract
fractions serve as free radical inhibitors or
scavenger or acting possibly as primary oxidants
and inhibited the heat induced albumin
denaturation and proteinase activity and stabilized
the Red Blood Cells membrane. The solvent
fractions exhibited a moderate xanthine oxidase
(XO) inhibitory activity and therefore may due to
presence of bioactive constituents and these can
useful in the treatment of xanthine oxidase induced
diseases. This paper proposing its potential
application as a lead compounds for designing
potent anti-inflammatory activity and they can be
used for treatment of various diseases like
bronchitis, biliousness, hyperacidity, dysentery,
diabetes, leprocy, and inflammatory.
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