mid-term progress report - microcosmos
TRANSCRIPT
Project Title:
To detect damages in liver tissues inducing CCl4 stress and their
recovery after treatment with different plant extracts to establish their
antioxidant and medicinal activities
Sanctioned vide Letter No.:
BT/IN/Indo-US/Foldscope/39/2015 Dated20-03-2018
Funded by:
Department of Biotechnology (DBT), Govt. of India
Submitted by
Prof Manabendra Dutta Choudhury
Principal Investigator, Foldscope Project; Category-B
Coordinator
Assam University Biotech Hub
Assam University, Silchar-788011, Assam, India
MID-TERM PROGRESS REPORT
On
FOLDSCOPE PROJECT
To
Dr Vaishali Punjabi
Scientist D
Block No. 2, 6-8th Floors
Department of Biotechnology
CGO Complex, Lodi Road,
New Delhi - 110 003
Sub: Mid Term Progress Report of Foldscope Project sanctioned vide letter no.
BT/IN/Indo-US/Foldscope/39/2015 dated 20-03-2018
Sir/Madam,
Please find herewith the Mid-Term Progress Report of DBT, Govt of India funded
Foldscope Project for evaluation. UC/ SE of said project is being sent shortly. Necessary action
from your end is highly solicited.
Assam University Biotech Hub Sponsored by Department of Biotechnology (DBT), Govt. of India
Assam University, Silchar – 788011, Assam, India
(Prof. Manabendra Dutta Choudhury)
Principal Investigator, DBT- Foldscope Project, Category -B
Coordinator, Assam Univerrsity Biotech Hub,
Assam University, Silchar - 788011
Date: 15-11-2018
Place: Silchar
E-mail: [email protected]
Phone/ Fax: +91-3842-270920
RESEARCH ACTIVITIES
Objectives of the work:
1. Collection and identification of medicinalplantsfrom Southern Assam, India.
2. Preliminary qualitative phytochemicalscreeningof the crude plant extracts
3. Quantitative phytochemical screening of the crude plant extracts
4. In vitro antioxidant activity evaluation of the crude plant extracts.
5. Foldscope based histopathological studies of the plant extract mediated recovery of
CCl4intoxicated liver in Swiss Albino mice models.
Methodology
Collection of Plant material
Fresh fronds of P. semipinnata (Voucher no. 17602) were collected from Algapur, Hailakandi
district of Southern, Assam, India. The herbarium sheet of the collected specimen have been
submitted to the Assam University Herbarium and identified from Botanical Survey of India (BSI),
Shillong.
Preparation of crude extracts
Collection of the plant materials was followed by air drying at room temperature for some
days. The air dried fronds were ground into powder with the help of a grinder. From this powder
different crude plant extracts were sequentially prepared by Soxhlet’s method of hot extraction
process with various solvents of differential polarity viz., Hexane, Ethyl Acetate, Acetone and
Methanol to furnish different plant extracts of less polarity, medium polarity and high polarity.
Preliminary phytochemical analysis
The preliminary phytochemical analysis of the prepared extracts was carried out using standard
phytochemical methods [1].
Estimation of Total Phenolic Content (TPC)
Folin-Ciocalteau (FC) method was used to determine total phenolic content of the frond extracts.
0.2 mL 10 % v/v Folin-Ciocalteau reagent was added to 0.1 mL of the sample and was constantly
shaken for 5 min, followed by addition of 0.8 mL of sodium carbonate (Na2CO3). This reaction
mixture was incubated for 2 h at room temperature. The absorbance was then measured at 765 nm
of wavelength. The calibration curve was prepared by employing gallic acid at concentrations of 10
to 100 µg/ml of methanol [2].The concentrations of phenolic compounds were calculatedaccording
to the following equation that was obtained from the standard gallic acid graph:
Absorbance = 0.0608 gallic acid (μg) - 0.0081 (R² = 0.9682)
Estimation of Total Flavonoid Content (TFC)
Total flavonoid content was determined using the Dowd method as adapted by [3]. Briefly, 1 ml of
2% aluminium trichloride (AlCl3) in methanol was mixed with the same volume of the methanolic
extracts (2000 μg). Absorption readings at 415 nm were taken after 10 min against a blank sample
consisting of a 1 mL extract solution with 1 ml methanol without AlCl3. The concentrations of
flavonoid compounds were calculated according to the following equationthatwas obtained from
the standard quercetin calibration curve:
Absorbance = 0.0355 quercetin (μg) - 0.2396 (R²: 0.9886)
DPPH radical scavenging activity
The free radical scavenging activities of the plant extracts were measured in terms of hydrogen
donating or radical scavenging ability using the stable radical DPPH [4]. Extract solution (0.1 ml)
in MeOH at different concentrations was added to 3 ml 0.004% MeOH solution of DPPH.
Absorbance at 517nm was determined after 30 min. The decreasing absorbance of the DPPH
solution indicated an increase in the DPPH radical scavenging activity. Quercetin(50 μg/ml) was
used as a positive control. DPPH radical scavenging activity (%) was calculated by using the
following formula:
DPPH radical scavenging activity (%) = [(A control– A sample)/ Acontrol]x 100
where, Acontrol is the absorbance of the control and Asampleis the absorbance of the test.
Reducing power assay
The reducing power of the prepared frond extracts was determined according to the method of
Oyaizu [5]. Each extract (0.2–1.0 mg/ml) in methanol (2.5 ml) was mixed with 2.5 ml of 200 mM
sodium phosphate buffer (pH 6.6) and 2.5 ml of 1% potassium ferricynideandthe mixture was
incubated at 500C for 20 minutes. Then, 2.5 ml of 10% trichloroacetic acid was added, and the
mixture was centrifuged at 200g for 10 min. The upper layer (2.5 ml) was mixed with 2.5 ml of
deionized water and 0.5 ml of 0.1% ferric chloride. Finally the absorbance was measured at 700 nm
against a blank. The standard antioxidant i.e., ascorbic acid was used as a control for this
experiment.
Hydroxyl radical scavenging Activity
Hydroxyl radical-scavenging activity was measured according to Smirnoff’s work [6]. 0.5 mL
FeSO4(1.5 mM) was mixed with 0.35 mL H2O2 (6 mM), 0.15 mL sodium salicylate (20 mM) and 1
mL sample (0.2-1.0 mg/mL), then incubated for 1 h at 370C. The absorbance of the hydroxylated
salicylate complex was measured at 562 nm. Ascorbic acid was used as the positive control. The
antioxidant activity was calculated with the following equation:
scavenging effect (%) = 1- (Asample- Ablank) / AcontrolX 100
whereAsample was the absorbance of the test (sample or ascorbic acid), Acontrol was the absorbance of
the solvent control, and Ablank was the absorbance of the reagent blank without sodium salicylate.
Superoxide radical scavenging Activity
Superoxide radicals were generated by pyrogallic acid method [7] and the method was innovated
slightly. The system contained 2.5 mL of PBS buffer (0.1 M, pH 8.2), 4 mL of sample solution, 2.5
mL of pyrogallic acid (6.0 mM), and0.5 mL of thick hydrochloric acid for termination the reaction.
The solution was incubated at 250C and determined at 299 nm. Ascorbic acid was used as a
reference material. All tests were performed in triplicate. The scavenging activity was calculated as
follows:
scavenging activity (%) =A0 - (As - Ac)/ A0 X 100
where As, with the presence of pyrogallic acid and test extracts;A0, with the presence of pyrogallic
acid but without test Text extracts; and Ac, with the presence of test extracts but without pyrogallic
acid.
ABTS+
cation scavenging activity
The ABTS+ radical scavenging activity was determined by spectrophotometric analysis [8]. The
ABTS+
cation radical was produced by the reaction between 7 mM ABTS in water and 2.45 mM
potassium persulfate, stored in the dark at room temperature for 12 h. Before usage, the ABTS+
solution was diluted to get an absorbance of 0.700± 0.025 at 734 nm with phosphate buffer (0.1 M,
pH 7.4). Then, 1 ml of ABTS+solution was added 3 ml of extract solution in methanol at different
concentrations (8–40 lg/ml). After 30 minutes, the percentage inhibition at 734 nm was calculated
for each concentration relative to a blank absorbance (methanol). The scavenging capability of
ABTS+radical was calculated using the following equation:
ABTS+ scavenging effect (%) = [(Acontrol - Asample)/ Acontrol]X 100
where Acontrol is the initial concentration of the ABTS+ and Asample is the absorbance ofthe remaining
concentration of ABTS+ in the presence of extract.
Chelating effects on ferrous ions
The chelating effect was determined according to the method of Dinis et al. [9]. Briefly, 2 ml of
various concentrations (0.05–0.25 mg/ml) of the extractsin methanol was added to a solution of 2
mM FeCl2 (0.05 ml). The reaction was initiatedby the addition of 5 mM ferrozine (0.2 ml). Then,
the mixture was shaken vigorously and left at room temperature for 10 min. Absorbance of the
solution was measured spectrophotometrically at 562 nm. The inhibition percentage of ferrozine–
Fe2+ complex formation was calculated by using the formula given below:
Metal chelating effect (%) = [(Acontrol – Asample) / Acontrol]× 100
where Acontrol is the absorbance of control (the control contains FeCl2 and ferrozine, complex
formation molecules) and Asample is the absorbance of the test compound. EDTA was used as a
control.
Foldscope based histopathological studies of the plant extract mediated recovery of CCl4
intoxicated liver in Swiss Albino mice models.
Test Animals:
Swiss albino mice weighing between 25g and 30g, used for the study were obtained from
the College of Veterinary Science and Animal Husbandry, Khanapara, Guwahati. The animals were
housed in cages under standard environmental conditions of temperature (24º ± 2º C) and humidity
(60 ± 5%)withfood and water ad libitum. An acclimatization time of 10 days was given prior to
start of the experiment.
The experiments were performed in accordance with the guidelines in the care and use of
laboratory animals and were approved by the Ethical Committee(AUS/IAEC/2017/PC/06, dated
25/8/2017) of Assam University, Silchar.
Acute toxicity study:
Swiss albino mice were selected by a random sampling technique for acute toxicity study.
The animals were fasted overnight prior to the experiment and maintained under standard
laboratory conditions. The methanolic extract of P. semipinnata was orally administered in
increasing dose up to 2000mg/kg, but no mortality was observed.
Animal treatment and preparation of test:
Mice were randomly divided into four groups (n=6).Group 1(normal control) animals were
administered a single oral dose of water (25ml/kg) daily for 6 days and received olive oil (10ml/kg,
i.p) on day 1 and day 2. Group 2 (CCl4) received water (25ml/kg) once daily for 6 days and
received 0.2% CCl4 in olive oil (10ml/kg, i.p.)on day 1 and day 2. Group 3 received standard drug
Silymarin (100mg/kg) orally once daily for 4 days from day 3 to day 6and received 0.2% CCl4in
olive oil (10ml/kg, i.p) on day 1 and day 2.Group 4 received orally a dose of 250mg/kg of
methanolic extract of P. semipinnata was for 4 days from day 3 to day 6 and received 0.2% CCl4in
olive oil (10ml/kg, i.p) on day 1 and day 2. On day 7, the mice (n = 6 per group) were anaesthetised
by chloral hydrate (350 mg/kg b.w.; I.P.), and then mice were perfused transcardially with 50mL
each of ice-cold 0.1M phosphate buffered saline (PBS; pH 7.4) and 4% w/vparaformaldehyde (in
PBS). After perfusion, liver of the mice were dissectedout, and stored in 4% paraformaldehyde,
cryoprotected overnight in 30% w/v sucrose solution, 5 μm thick liver sections weretaken on poly-
L-lysine coated slides and Haematoxylin-Eosin staining was performed. (Table.1)
Table.1: Animal treatment and experimental set up
Hepatic histological parameters:
To analysis the result of induced hepatic toxicity and their recoveryatthe histological level, hepatic
histological studies were performed. Haematoxylin-Eosin routine staining procedure was performed
to see possible hepatic damage in the hepatic tissues. The mice were transcardially perfused with50
mL each of 0.1M phosphate buffered saline (PBS; pH 7.4) and 4% w/v paraformaldehyde (in PBS).
Days
Group
Day 1 Day 2 Day 3 Day 4 Day 5 Day 6 Day 7
Control
Water orally
+
Olive oil i.p
Water orally
+
Olive oil i.p
Water
orally
Water
orally
Water
orally
Water orally
Sacrifice
CCl4
Water orally
+
0.2% CCl4 in
olive oil i.p.
Water orally
+
0.2% CCl4 in
olive oil i.p.
Water
orally
Water
orally
Water
orally
Water orally
CCl4
+
Silymarin
Water orally
+
Olive oil i.p
Water orally
+
Olive oil i.p
Silymarin
(100mg/kg)
Silymarin
(100mg/kg)
Silymarin
(100mg/kg)
Silymarin
(100mg/kg)
CCl4
+
Methanol
extract of
P.semipinnata
Water orally
+
0.2% CCl4 in
olive oil i.p.
Water orally
+
0.2% CCl4 in
olive oil i.p.
MeOH of P.
Semipinnata
fronds
250mg/kg
(orally)
MeOH of P.
Semipinnata
fronds
250mg/kg
(orally)
MeOH of P.
Semipinnata
fronds
250mg/kg
(orally)
MeOH of P.
Semipinnata
fronds
250mg/kg
(orally)
Liver were dissected out and stored in the same fixative for 48 hours at 40 C, and then cryoprotected
in 30% w/v sucrose solution for pending histological studies.
Haematoxylin-Eosin staining:
From the paraformaldehyde fixed liver of all groups of mice, mid-longitudinal, 5μm thick sections
were taken on poly-L-lysine coated slides and stained using Haematoxylin-Eosin procedure.
Briefly, the sections were cleared in xylene,hydrated in decreasing alcoholic grades (100%, 90%,
70%, 50%, 30% and distilledwater) for 2-5 min each, stained with Haematoxylin for 20 min,
washed in runningtap water, dehydrated in increasing alcoholic gradient (50%, 70%, 90% and
absolute) for 2-5 min each, counterstained with Eosin (1% Eosin in absolutealcohol), washed in
absolute alcohol, cleared in xylene, mounted in DPX andphotographed using a mobile phone
camera attached to foldscope microscope under bright-field illumination and also with Olympus
DSLR under Olympus CX41 bright field microscope at nosepiece set at10x and 40x with eyepiece
fixed at 5x.
Results:
Preliminary phytochemical analysis
The preliminary phytochemical screening results implicated the presence of phenolic and flavonoid
compounds in all the prepared extracts, which indicates the probability of antioxidative behavior of
the frond extracts of the plant. Alkaloids were found to be present in the ethylacetate, acetone and
methanol extract of the fronds, whereas, saponin was found to be present in the acetone and
methanol extract of the fronds.
Estimation of TPC and TFC
As all the extracts showed the presence of phenols and flavonoids, hence, the TPC and TFC of all
the extracts were quantified in terms of gallic acid equivalents and quercetin equivalents
respectively. The methanolic extract of the fronds showed maximum phenolic content (370.5±2.1
µgGAEs/ mg of extract) as well as flavonoid content (324.3±1.1 µgQEs/ mg of extract) in
comparison to the other extracts. (Table.2)
DPPH radical scavenging activity
The free radical scavenging activities of the plant extracts were measured in terms of hydrogen
donating or radical scavenging ability using the stable radical DPPH. The IC50 values, i.e., 50%
DPPH radical - inhibitory concentration values of the extracts were found to be the lowest in case
of methanolic frond extract again in comparison to the other extracts (Table.3). However, the IC50
value of the quercetin standard was still a bit lower than the methanol extract. The DPPH radical-
scavenging activity was found to be in the order of Quercetin>Methanol>Acetone > Ethyl Acetate
> Hexane. This indicates that the compounds with strongest radical-scavenging activity in the
fronds of the plant are of high polarity. This radical-scavenging activity of extracts/ fractions could
be related to the nature of phenolics, thus contributing to their electron transfer/hydrogen donating
ability.
Reducing power assay
The reducing power or activity assay was based on the reduction of Fe3+
/ferricyanide complex to
the ferrous form in presence of reductants (antioxidants) in the tested samples. The Fe2+
was then
monitored by measuring the formation of Perl’s Prussian blue coloured solution at 700 nm(Oyaizu,
1986). The reducing power of all the extracts were observed to get increased with the extract
concentration (Table.4). At all the subsequently increasing concentrations of the plant extracts, the
reducing power of the methanolic frond extracts was found to be the highest as compared with that
of the ascorbic acid control as well as other extracts. Therefore, as a result of this experiment the
methanolic extract was found to be a potential free radical reducer for this system.
Hydroxyl radical scavenging activity
The hydroxylradical scavenging activity of all the extracts has been shown in Table.5. The ascorbic
acid standard was found to exhibit the highest hydroxylradical scavenging activity with an IC50
value of 0.290± 0.007 mg/ml followed by the methanol extract of P. semipinnata having IC50 value
of 0.410±0.011 mg/ml. The hydroxyl radical scavenging activity was found to be the lowest in
hexane extract of P. semipinnata.
Superoxide radical scavenging activity
The superoxide radical scavenging activity of the plant extracts/ standard is shown in Table.6. The
methanolic frond extract of P. semipinnata revealed the highest superoxide radical scavenging
activity with an IC50 value of 0.280 ± 0.004 mg/ml which was slightly less than that of the ascorbic
acid standard having IC50 value of 0.230 ± 0.006 mg/ml.
ABTS+
cation scavenging activity
Table.7 illustrates the ABTS+
cation scavenging activity of the plant extracts. As a result of this
assay the highest activity was observed in the methanolic extract of P. semipinnata (84.57±1.26%)
at the concentration of 40 µg/ml followed by the other extracts of decreasing solvent polarity.
Chelating effect on ferrous ions
The percentage metal chelating effect (Table.8) of the standard chelating ligand i.e., EDTA was
found to be the highest (97.24 ± 5.6%) at the concentration of 0.250 mg/ml followed by the
methanol extract of P. semipinnata fronds (95.46±1.4%). The metal chelating effect of the extracts
was also found to be decreasing with respect to the decreasing solvent polarity.
Hematoxylin-Eosine staining:
The CCl4 treated animals (group 2) developed hepatocyte damage. Severe necrosis of the hepatic
cells has been observed in the H-E staining of the liver sections (Fig. 2). However, no changes were
observed in the liver morphology (Fig. 1) of the control animals (Group 1). Liver morphology (Fig.
3) of CCl4 treated along with silymarin (positive control, Group 3) is showing recovery of liver
tissue damaged due to CCl4 administration. And the liver morphology (Fig. 4) ofCCl4 treated along
with MeOH extract of P. semipinnata was (Group 4) interestingly showing comparable recovery of
liver tissue damaged due to CCl4 administration.
Table. 2: Total Phenolic Content (TPC) & Total Flavonoid Content (TFC) of the prepared
frond extracts
Extracts TPC
(μg eq. of Gallic Acids/mg of
extract)
Mean ± SD
TFC
(μg eq. of Quercetin/mg of
extract)
(Mean±SD)
Hexane 49.6±2.7 21.6±1.2
Ethyl Acetate 162.8±2.2 87.2±4.2
Acetone 243.4±3.4 167.3±3.1
Methanol 370.5±2.1 324.3±1.1
Table. 3: DPPH Free Radical Scavenging Activities of the P. semipinnata frond extracts
Extracts/Standard IC 50 Values
(mg/ml)
(Mean±SD)
Hexane 1.900±0.020
Ethyl Acetate 1.430±0.011
Acetone 0.920±0.009
Methanol 0.870±0.009
Quercetin 0.510±0.012
Table. 4: Reducing Power Assay of the frond extracts
Extracts/
Standard
Absorbance at 700 nm (OD)
(Sample Concentrations in mg/ml)
(Mean±SD)
0.2 0.4 0.6 0.8 1.0
Hexane 0.046±0.010 0.067±0.003 0.085±0.002 0.097±0.006 0.152±0.001
Ethyl Acetate 0.196±0.007 0.212±0.001 0.234±0.008 0.278±0.020 0.312±0.002
Acetone 0.387 ± 0.001 0.491 ± 0.004 0.598 ± 0.002 0.786 ± 0.001 0.856 ± 0.031
Methanol 0.621 ± 0.004 0.647 ± 0.002 0.761 ± 0.005 0.889 ± 0.009 1.124 ± 0.010
Ascorbic Acid 0.215 ± 0.004 0.298 ± 0.008 0.387 ± 0.002 0.452 ± 0.015 0.561 ± 0.003
Table. 5: IC50 Values of the plant extracts for eliminating hydroxyl radicals
Extracts/Standard IC 50 Values
(mg/ml)
(Mean±SD)
Hexane 0.940±0.006
Ethyl Acetate 0.610±0.001
Acetone 0.490±0.003
Methanol 0.410±0.011
Ascorbic Acid 0.290±0.007
Table. 6: IC50 values of the plant extracts for eliminating superoxide radicals
Extracts/Standard IC 50 Values
(mg/ml)
(Mean±SD)
Hexane 0.730±0.011
Ethyl Acetate 0.480±0.003
Acetone 0.320±0.007
Methanol 0.280±0.004
Ascorbic Acid 0.230±0.006
Table. 7: Scavenging (%) effect of the extracts on the stable ABTS+
Extracts/Standard % Scavenging effect
(Mean±SD)
Sample Concentrations in µg/ml
8 20 40
Hexane 35.11±2.01 41.25±1.34 47.94±2.55
Ethyl Acetate 44.09±1.25 56.31±0.21 69.78±1.38
Acetone 49.72±1.41 62.47±0.58 81.21±2.21
Methanol 69.32±1.33 78.15±2.24 84.57±1.26
Table. 8: Metal chelating effect of the extracts on Fe2+
ions
Extracts/Standard % Chelating effect
(Mean±SD)
Sample Concentrations in mg/ml
0.050 0.150 0.250
Hexane 34.20±0.7 50.26±1.1 61.31±2.4
Ethyl Acetate 57.89±2.1 64.91±2.3 81.98±1.4
Acetone 71.26±1.2 76.76±2.9 88.32±2.2
Methanol 84.22±1.6 89.99±2.1 95.46±1.4
EDTA 89.29±3.6 92.83±2.4 97.24±5.6
Fig. 1 Fig. 2
Fig. 3 Fig. 4
Fig.: Representative photographs of Haematoxylin-Eosin stained sections of Liver of control
(Fig. 1), CCl4 (Fig.2), CCl4 + Silymarin (Fig.3) and CCl4 + MeOH extract of P. semipinnata
(Fig.4)
Twinning collaboration developed with:
1. Dr Arati Prabhu & Dr Munira Momin, SVKM’s Dr Bhanuben Nanavati College of
Pharmacy, Mumbai, Maharashtra
2. Dr Arindam Biswas, Indian Institute of Engineering Science and Technology, Shibpur,
Kolkata, West Bengal
Work still to be done:
1. Computational analysis of the foldscope based photographs of prepared histopathological
slides.
2. Same experimentations to be performed for histopathological evaluation of hepato-
protective potential of some more medicinal plant extracts.
3. Comparative analysis of the foldscope based inferences with that of electron microscopic
studies of the same samples.
OUTREACH/ AWARENESS
ACTIVITIES
Outreach Workshops and Awareness Programmes on Foldscope Microscopy
1. Organized lecture cum hands on demonstration on Foldscope microscope at Assam
University Biotech Hub for the visiting school students of Sonapur M.E. School, Silchar,
Assam on 20th
August, 2018.
2. Organized lecture series cum hands on demonstration on Foldscope Microscope for the
teachers and high school students of Vivekananda Kendra Vidyalaya, Borjhalenga,
Silchar in collaboration with other associated PIs of Foldscope Projects sanctioned at
Dept. of Microbiology, Assam University,Silchar as well as VijnanaBharati on 24th
August, 2018.
3. Organized Awareness Programme on Foldscope Microscopy for senior secondary science
students at Karimganj College, Karimganj, Assam on 16th
September, 2018 in
collaboration with Institutional Biotech Hub of Karimganj College, Karimganj.
4. Organized Awareness Programme on Foldscope based Diagnosis of Malaria for school students
and public at Deodhar LP School, Debodwar Village, Sonebeel, Karimganj on 19th
September,
2018 in collaboration with Dr Neelanjana Dutta Roy, Co PI of Foldscope Project sanctioned at
Indian Institute of Engineering, Science & Technology, Shibpur, Kolkata and other associated
PIs of Foldscope Projects sanctioned at Dept. of Microbiology, Assam University, Silchar.
5. Organized Awareness Programme on Foldscope Microscopy for Higher Secondary and
Degree students of various colleges of Hailakandi district at Hailakandi Womens’
College Auditorium, Hailakandi, Assam on 23rd September, 2018 in collaboration
with Prescientia Coaching Centre, Kalibari Road, Hailakandi and Hailakandi Women’s
College.
6. Delivered invited awareness talk on Foldscope Microscopy and demonstrated Foldscope
Microscope to the senior secondary and degree students at Workshop cum Inauguration
programme of Foldscope Project organized by and held at Dept. of Botany, Pandit
Deendayal Upadhyaya Adarsha Mahavidyalaya, Eraligool, Karimganj on 4th
October, 2018.
Acknowledgements
We acknowledge Department of Biotechnology (DBT), Govt. of India, for providing financial
assistance in the form of Foldscope project (Sanction letter no. BT/IN/Indo-US/Foldscope/39/2015
dated 20-03-2018) in Assam University Biotech Hub, Assam University, Silchar -78801. We also
acknowledgeDepartment of Biotechnology (DBT), Govt. of India, for providing instrumental facility in
the form of Institutional Biotech Hub (sanction letter no. BT/04/NE/2009 Dated 21/09/2010) and free
access of Delcon’s e-Journal library in the form of Bioinformatics Infrastructure Facility (BIF)
(sanction letter no. BT/BI/12/042/2007 Dated 11/02/2008) in Assam University, Silchar-788011.
REFERENCES
[1] A. A. Siddiqui and M. Ali,“Practical Pharmaceutical Chemistry,” 1st Edn.CBS Publishers and
distributors, New Delhi, 1997, pp. 126―131.
[2] V. L. Singleton and J.A. Rossi,“Colorimetry of total phenolics with phosphomolybdic-
phosphotungstic acid reagents,”Amer.J.Enol.Viticul., 1965, vol.16, pp.144―158.
[3] A. Arvouet-Grand, B. Vennat and A. Pourrat, “Standardization of propolis extract and identification
of principal constituents., 1994, vol.49, pp.462―468.
[4] L. L. Mensor, “Screening of Brazilian plantextracts for antioxidant activity by the use of DPPH free
radical method,” Phytother. Res., 2001, vol.15, pp.127―130.
[5] M. Oyaizu, “Studies on products of browning reactions: antioxidative activities of browning
reaction prepared from glucosamine,” Jap. J. Nutr.,1986, vol.44, pp.307―315.
[6] N. Smirnoff and Q.J. Cumbes, “Hydroxyl radical scavenging activity of compatible solutes,”
Phytochem., vol. 28, 1989, pp.1057―1060.
[7] B. Jiang,“Extraction of water-soluble polysaccharide and the antioxidant activity from Ginkgo
biloba leaves,” Med. Chem. Res., 2010, vol.19, pp.262―270.
[8] R. Re, N. Pellegrini, A. Proteggente, A. Pannala, M. Yang and C. Rice-Evans,“Antioxidant activity
applying an improved ABTS+ radical cation decolorization assay,” Free Rad. Biol. Med., 1999, vol. 26,
pp.1231―1237.
[9] T. C. P. Dinis, V.M.C. MadeiraandL.M. Almeida, “Action of phenolic derivatives (acetaminophen,
salicylate, and 5-aminosalicylate) as inhibitors of membrane lipid-peroxidation and as peroxyl radical
scavengers,” Arch. Biochem.Bioph., 1994, vol. 315, pp.161―169.