Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in Paracetamol-Induced Nephrotoxicity and ...

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International Journal of Research in Pharmacy and Biosciences
Volume 6, Issue 3, 2019, PP 1-11
ISSN 2394-5885 (Print) & ISSN 2394-5893 (Online)

     Free Radicals Scavenging Potential of Buccholzia Coriacea
      Extract and Ameliorative Effect in Paracetamol- Induced
             Nephrotoxicity and Hepatotoxicity in Rats
                                       Fakoya, A., Olusola, A. O*
           Department of Biochemistry, Faculty of Sciences, Adekunle Ajasin University,
                               Akungba Akoko, Ondo State. Nigeria
*Corresponding Author: Olusola, A. O, Department of Biochemistry, Faculty of Sciences, Adekunle
Ajasin University, Akungba Akoko, Ondo State. Nigeria, Email: austinolusola@gmail.com

 ABSTRACT
 The present study sought to evaluate the free radical scavenging activities of ethanolic extract of
 Buccholzia coriacea (EBCS) by measuring its ability to scavenge 1,1-diphenyl-2-picrylhydrazyl (DPPH*)
 radical, nitric oxide (NOˉ) radical, 2,2 azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS*), chelate
 Fe2+ radicals, hydroxyl radical (OHˉ) and inhibit lipid peroxidation (LPO). Total flavonoids and phenolics
 were also determined. Antioxidant activity of EBCS was also determined in the plasma of the rats fed with
 the extract by assaying for antiradical activity against DPPH and NOˉ radicals in vitro. In vivo antioxidant
 effect of EBCS was also evaluated in paracetamol treated rats. Fifteen rats were randomly divided into
 three groups for this study. Group 1 received normal feed as control, group 2 received 14.30 mg/kg b.w. of
 paracetamol by gavage, group 3 received 400 mg/kg b.w. of EBCS for 7 days plus paracetamol on the 8 th
 day. Catalase (CAT) and superoxide dismutase (SOD) activity, and malondialdehyde (MDA) levels were
 assayed in the kidney, liver and serum. Histopathological examinations of liver and kidney were also
 carried out. The results showed that EBCS exhibited free radical scavenging ability in dose dependent
 manner towards DPPH*, NOˉ, ABTS*, OHˉ and Fe 2+ radicals as well as inhibition of LPO. The results of
 evaluation of the antioxidant potentials of the extracts in the plasma showed that they were associated with
 free radical scavenging activity in vivo. Paracetamol treatment caused significant (p
Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in
Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

capparaceae, locally known as ‘Uke’ (Ibo),            Biochemical In-Vitro Assays
‘Uworo’ (Yoruba), Owi (Edo), (Ezekiel, 2009)
                                                      Dpph Radical Scavenging Assay
is traditionally macerated in water or local gin
as a cure for diabetes. The kola is also              The in-vitro antioxidant activity of the sample
recommended in the treatment of migraine. The         was determined according to the method
leaves and seed have been reported to have anti-      described by Mensor et al. (2001). To 1ml of
helminthic activity (Karmeswararao, 2003) as          plant extract, 1ml of ethanolic solution of 2, 2-
well as anti-microbial properties, (Nweze, 2006)      diphenyl-1-picrylhydrazyl (DPPH) (0.3 mM)
an attempt has been made to investigate the           was added. The mixture was incubated in the
blood glucose lowering activity of methanolic         dark for 30 min. The assay was standardized
extract of seeds of B. coriacea on type 2             with Tannic acid. The absorbance of the yellow
diabetes model to ascertain the folkloric claims      colour solution was read at 517 nm on a
of local healers.                                     spectrophotometer using methanol as blank.
In the present study, an attempt has been made        DPPH scavenged (%) = (A control – A sample)
to evaluate the in-vitro antioxidant activities,      /A control x 100.
and in vivo antioxidant of ethanolic extract of       Where A control = the absorbance of the
B,coriacea in paracetamol induced toxicity in         ethanol, A sample = the absorbance of the
order to justify the folkoric claims of traditional   reaction mixture.
healers..
                                                      Nitric Oxide Radical Scavenging Assay
MATERIALS AND METHODS                                 The in-vitro nitric oxide scavenging activity was
Reagents and Chemicals                                estimated according to the method of Marcocci
                                                      et al., 1994). To 1ml sample, 1ml of sodium
The 2, 2-diphenyl-1-picrylhydraxyl (DPPH) and         nitroprusside (10 mM, aqueous) and 1 ml buffer
Hydrogen peroxide, Potassium ferricyanide,            (sodium phosphate buffer, 0.2 M) were added.
sodium carbonate, butanol, methanol, acetic           The mixture was incubated at room temperature
acid, thiobarbituric acid, sodium dodecyl             for 150 min. this is followed by the addition of
acrylamide sulphate, Iron ii tetraoxosulphate         0.1 ml Griess reagent. The absorbance of the
(vi) used were a product of Sigma- Aldrich,           pink colour solution was read at 540 nm on a
USA. 1, 10-phnanthroline used were products of        spectrophotometer.       The     reaction     was
Merck, Germany.                                       standardized with ascorbic acid. The pink
Collection of Plant Materials                         chromophore generated during diazotization of
                                                      nitrite ions with sulphanilamide and subsequent
The seeds of B. coriacea were gotten from Oja         coupling with N-naphthyl ethylene diamine
Oba Market at Ikare, Ondo State. The plant was        dihydrochloride      (NED)      was     measured
identified in the Department of Plant Science         spectrophotometrically at 540 nm. The in-vitro
and      Biotechnology,   Adekunle      Ajasin        NOˉ scavenging activity of the sample was
University, Akungba Akoko, Ondo State,                calculated by using the following formula:
Nigeria.
                                                      Nitric oxide scavenging activity (%) = (A
Extraction Procedure                                  control - A sample)/ A control x 100.
Cold extraction method was employed. 500 g of         Where A control = the absorbance of the
the clean, air dried and pulverized plants            reaction mixture in the absence of sample.
samples were weighed differently into
extraction jars respectively and 1400 ml of           A sample = the absorbance of the reaction
analytical grade ethanol was added to the jars        mixture.
containing B. coriacea. The extraction mixture        ABTS* Radical Scavenging Activity Assay
was given constant agitation and left for 72
hours. The supernatant was decanted separately        The in-vitro ABTS* scavenging activity of the
and concentrated using a rotary evaporator at 40      polyphenolics was determined according to the
0
  C and the extract was freeze dried. The extract     method of Re et al., 1999. The stock solutions
was packed inside an airtight sample bottle and       were of 8mM ABTS and 3mM Potassium
kept at 4 0C inside refrigerator until required for   persulphate. The working solution was then
various in-vitro and in vivo antioxidant              prepared by mixing the two stock solutions in
assessments.                                          equal quantity and allowing them to react for 12
                                                      hours at room temperature in the dark. To 1ml

2                        International Journal of Research in Pharmacy and Biosciences V6 ● I3 ● 2019
Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in
Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

sample (1mg/ml), 1ml of ABTS was added. The           Hydroxyl radical scavenging activity (%) =
absorbance was read at 734nm on a                    [ Acontrol-Asamole/Acontro] x 100.
spectrophotometer. Trolox was used as standard.
                                                     Where Acontrol= Absorbance of the control.
The in-vitro ABTS* radical scavenging activity
of the sample was calculated by using the            Asample = Absorbance of plant extract sample.
following formula:                                   Determination of Total Phenolic Content
ABTS* scavenging activity (%) = (A control -
                                                     Folin Ciocalteu reagent was used for analysis of
A sample)/ A control x 100.
                                                     total phenolics content (TPC) according to Chun
Where A control = the absorbance of the              et al. (2003). In a 10 ml volumetric flask, a 0.2
reaction mixture in the absence of sample.           ml aliquot of the extract in ethanol (1.0 mg/ml)
A sample = the absorbance of the reaction            was mixed with 0.4 ml of Folin-Ciocalteu
mixture.                                             reagent. The solution was allowed to stand at
                                                     25oC for 5-8 min before adding 0.2 ml of 4.0 ml
Inhibition of Lipid Peroxidation Assay               of sodium carbonate solution 7.0 % and made to
In-vitro inhibition of lipid peroxidation was        10 .0 ml with distilled water. The mixture was
estimated according to the method of Ruberto         allowed to stand for 2 h before its absorbance
and Baratta, 2000. In this assay, egg yolk           was measured at 725 nm. Tannic acid was used
homogenate served as lipid rich medium, and          as standard for the calibration curve. TPC was
FeS04 acts as initiator of lipid peroxidation.       expressed as mg tannic acid equivalents (TAE)
Briefly, 40 μl of plant extract was mixed with       per gram of sample (mg/g).
0.25 ml 10% egg yolk. This was followed by the       Determination of Total Flavonoid Content
addition of 10 ul FeS04 (0.07 M, aqueous). The
                                                     The total flavonoid contents were measured by a
mixture was incubated at room temperature for
                                                     colorimetric assay (Zhishen et al. 1999; Zou et
30 min. This was followed by the addition of
                                                     al., 2004). A-100.0 μl aliquot of extract in
0.75 ml of glacial acetic acid (5%,v/v aqueous)
                                                     ethanol was added to a 10 ml volumetric flask
and 0.75 ml of thiobarbituric acid 0.6% in 0.2 M
                                                     containing 4 ml of distilled water. At zero time,
NaOH .The mixture was incubated in a boiling
                                                     0.3 ml 5% sodium nitrite was added to the flask.
water bath (90oC) for 20 min, cooled and
                                                     After 5 min, 0.3 ml of 10% aluminium chloride
centrifuged at 3000 rpm. One milliliter (1 ml) of
                                                     was added. At 6 min, 2 ml of 1 M sodium
the pink colour supernatant was read at 532 nm
                                                     hydroxide was added to the mixture.
on a spectrophotometer. The assay was
                                                     Immediately, the mixture was diluted to volume
standardized using quercetin.
                                                     with the addition of 2.4 ml distilled water and
In-vitro inhibition of lipid peroxidation = (A       thoroughly mixed. Absorbance of the mixture,
contol – A sample)/A control X 100. Where A          pink in color, was determined at 510 nm versus
control = absorbance of the control, A sample is     a blank containing all reagents except sample of
the absorbance of the sample.                        extract. Rutin was used as standard for the
                                                     calibration curve. Total flavonoid content of the
Determination of Hydroxyl Radical Scavenging
                                                     extract was expressed as mg rutin equivalents
Activity
                                                     (RE) per gram of sample (mg/g).
The hydroxyl radical scavenging activity of the
                                                     Determination of Fe2+ Chelation Activity
extract was determined spectrophotometrically
according to the method describe by Wyu et al.,      The metal chelation activity of ethanolic extract
(2004). 0.09 ml of 1,10 phenanthroline was           of Buccholzia coriacea was carried out
added to 1.5 ml of varying concentrations of         according to the method described by Mnnoti
manitol standard (25-400 mg/ml) and the extract      and Aust, 1958, modified by Pentel et al., 2005.
(25-400 mg/ml) in their respective test tubes        The assay was standardised using EDTA. 900 µl
arranged in triplicate. This was followed by 0.06    of 500 µM FeSO4 was added into 150 µl of
ml of FeSO4, 0.015 ml of H2O2 and 2.4 ml of 0.2      varying concentrations (25 mg/ml – 400 mg/ml)
M phosphate buffer. Allowed to incubate for 5        of the extract. 78 µl of 0.25% w/v 1, 10-
minutes, absorbance readings were taken at 540       phenanthroline was added. Incubate at room
nm against the methanol blank. The percentage        temperature for 5 minutes. EDTA was used as
hydroxyl radical scavenging effect of plant          standard. The absorbance was read against the
extract was calculated as follows.                   blank at 510 nm.

International Journal of Research in Pharmacy and Biosciences V6 ● I3 ● 2019                         3
Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in
Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

The metal chelation activity (%) = Ao – As/Ao        6M H2SO4 was added to the sample, blank and
x 100.                                               standard tubes while 0.05 M phosphate buffer
Where, Ao = absorbance of control; As =              pH 7.4 (for the standard tube) was added and
absorbance of the sample.                            thoroughly mixed by inversion and thereafter
                                                     7.0 ml 0.01 M KMNO4 was added to all the
Estimation of Malondialdehyde (MDA) Status
                                                     tubes. Finally, the content of each tube was
The assay method of Buege and Aust, 1998 was         thoroughly mixed by inversion and the
adopted.                                             absorbance read at 480nm after 30-60 seconds
Principle                                            against distilled water as blank. Catalase activity
                                                     was calculated.
MDA which is formed from the breakdown of
polyunsaturated fatty acid serves as a               Estimation of Superoxide Dismutase (SOD)
convenient index for the determination of the        Activity
extent of lipid peroxidation (LPO). MDA reacts       This was determined according to the method of
with thiobarbituric acid to give a red product       Misra and Fridovich (1972).
absorbing at 535nm.
                                                     Principle
Procedure
                                                     Adrenaline auto-oxidizes rapidly in aqueous
To 1 ml sample, 2 ml of TBA-TCA-HCL                  solution to aderenochrome, whose concentration
Reagent (ml) was added, and 3 ml of the reagent      can be determined at 420nm.The auto-oxidation
was added to the blank test tube. The contents of    of adrenaline depends on the presence of
each test-tube was heated for 15 minutes in
                                                     superoxide anions. Superoxide Dismutase
boiling water. After cooling, each tube was
                                                     catalyzes the breakdown of superoxide anions
centrifuged at 4000rpm for 10 minutes to
                                                     thus inhibiting the auto-oxidation of adrenaline.
remove flocculent precipitates. Absorbance of
                                                     The degree of inhibition is thus a reflection of
each supernatant was read at 535nm against the
                                                     the activity of SOD and is determined as one
blank.
                                                     unit of the enzyme activity.
Estimation of Catalase (CAT) Activity
                                                     Procedure
This was determined according to the method of
Cohen et al., 1970.                                  3 ml of the sample tubes contained 0.2 ml
                                                     sample, 2.5 ml of 0.05 M carbonate buffer
Principle                                            pH10.2 and 0.3 ml of 0.03 mM adrenaline. The
Catalase catalyses the breakdown of hydrogen         blank tubes contained 3.0 ml of distilled water
peroxide introduced in the presence of               while the standard tubes contained 0.2 ml
phosphate buffer to water and oxygen.                distilled water, 2.5 ml of 0.05M carbonate
                                                     buffer pH10.2 and 0.3 ml of 0.03 mM
       2H2O2                      2H2O + O2          adrenaline. The content of each tube was mixed,
The reaction is thereafter quenched (stopped) by     and absorbance read at 420 nm. SOD activity
the addition of H2SO4. The amount of H2O2            was calculated.
remaining in the reaction mixture after few
                                                     Histological Test Preparations
minutes of catalase action will be determined by
titration   with     potassium     permanganate      The liver and kidney were carefully removed
(KMNO4), a very strong oxidizing reagent. The        and piece of the tissues were cut and kept in
amount of substrate remaining in the mixture is      10% formalin in carefully labeled universal
inversely proportional to the activity of the        containers for preservation prior to processing.
enzyme.                                              The slides were then stained using the
Procedure                                            Haematoxylin-Eosin staining technique through
                                                     increasing grade of alcohol and xylene before
To 0.5 ml of the sample, 5 ml of 30 mM H2O2          being mounted with a cover strip dried before
was added while the blank tube contains 0.5 ml       viewing microscopically for possible malignant
distilled water. The contents of the tubes were
                                                     changes.
mixed and left standing for three minutes,1.0 ml

4                       International Journal of Research in Pharmacy and Biosciences V6 ● I3 ● 2019
Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in
Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

RESULTS
Table1. Antioxidant constituents of B. coriacea
                Antioxidant                                                 Values
 Total phenol                                   18.0 mg/g of GAE
 Total flavonoid                                37.5 mg/g QE
 Vitamin C                                      300 mg/100 g of Ascorbic acid
Table2. DPPH free radical scavenging effect of ethanolic extract of B. coriacea
        Drug                Dose (µg/ml)          DPPH (measured at 518 nm)                  % Inhibition
 Control               -                        0.763±0.009                            -
 B.coreacea            25                       0.476±0.004                            37.78
                       50                       0.404±0.006                            47.23
                       100                      0.314±0.003                            58.91
                       200                      0.224±0.008                            70.72
                       400                      0.063±0.004                            91.76
 Tannic acid           25                       0.571±0.003                            25.32
                       50                       0.488±0.015                            36.25
                       100                      0.345±0.011                            54.95
                       200                      0.120±0.009                            84.27
                       400                      0.046±0.005                            93.99
Table3. Nitric oxide scavenging effect of ethanolic extract of B. coreacea.
        Drug               Dose (µg/ml)              Nitric oxide scavenging                 % Inhibition
 Control               -                       0.547±0.021
 B. coriacea           25                      0.087±0.013                             84.14
                       50                      0.079±0.021                             85.71
                       100                     0.072±0.015                             86.98
                       200                     0.083±0.008                             84.98
                       400                     0.066±0.005                             87.93
 Vitamin C             25                      0.302±0.002                             44.79
                       50                      0.250±0.001                             54.59
                       100                     0.106±0.005                             80.80
                       200                     0.077±0.004                             85.95
                       400                     0.035±0.005                             93.58
Table4. ABTSˉ free radical scavenging effect of ethanolic extract of B. coriacea
        Drug               Dose (µg/ml)            ABTSˉ (measured at 734nm)                  % Inhibition
 Control               -                      0.403±0.002                                 -
 B.coriacea            25                     0.395±0.003                                 02.07
                       50                     0.383±0.008                                 04.88
                       100                    0.377±0.004                                 06.37
                       200                    0.351±0.007                                 12.16
                       400                    0.311±0.006                                 22.91
 Trolox                25                     0.224±0.004                                 44.49
                       50                     0.086±0.004                                 78.66
                       100                    0.034±0.004                                 91.65
                       200                    0.008±0.001                                 98.09
                       400                    0.001±0.000                                 99.75
Table5. Inhibition of lipid peroxide (LPOˉ) formation by the ethanolic extract of B. coriacea
        Drug               Dose (µg/ml)               Lipid peroxide formation                % Inhibition
 Control               -                      0.577±0.004                                 -
 B. coriacea           25                     0.253±0.004                                 56.15
                       50                     0.226±0.008                                 60.83
                       100                    0.154±0.005                                 73.37
                       200                    0.110±0.001                                 80.94
                       400                    0.082±0.002                                 85.73
 Quercetin             25                     0.357±0.006                                 38.19
                       50                     0.315±0.003                                 45.35
                       100                    0.254±0.015                                 55.92
                       200                    0.142±0.003                                 75.33
                       400                    0.101±0.001                                 82.50

International Journal of Research in Pharmacy and Biosciences V6 ● I3 ● 2019                                 5
Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in
Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

Table6. Hydroxyl (OHˉ) free radical scavenging effect of ethanolic extract of B. coriacea
          Drug                   Dose                          OHˉ                          % Inhibition
                                (µg/ml)                 (measured at 510nm)
    Control            -                        1.024±0.020                              -
    B.coriacea         25                       0.038±0.006                              96.26
                       50                       0.038±0.003                              96.26
                       100                      0.043±0.0005                             95.77
                       200                      0.041±0.010                              95.99
                       400                      0.049±0.004                              95.25
    Mannitol           25                       0.894±0.004                              12.66
                       50                       0.788±0.003                              23.01
                       100                      0.605±0.005                              40.89
                       200                      0.430±0.002                              57.98
                       400                      0.306±0.004                              70.38
Table7. Metal chelaing effect of ethanolic extract of B. coriacea
         Drug                    Dose                         ABTSˉ                         % Inhibition
                                (µg/ml)                 (measured at 734nm)
 Control               -                        0.784±0.012                              -
 B.coriacea            25                       0.739±0.008                              09.90
                       50                       0.722±0.004                              11.86
                       100                      0.476±0.004                              41.88
                       200                      0.475±0.002                              41.96
                       400                      0.417±0.007                              49.03
 EDTA                  25                       0.629±0.006                              23.21
                       50                       0.585±0.004                              28.57
                       100                      0.420±0.009                              48.75
                       200                      0.368±0.003                              55.09
                       400                      0.204±0.004                              75.07
Table8. DPPH free radical scavenging activity of plasma from rats treated with ethanolic extracts of B.
coriacea
         Drug                   Dose                          DPPH                          % Inhibition
                               (mg/ml)                  (measured at 518nm)
 Control               -                        1.649±0.064a                            -
 Normal                -                        0.836±0.070b                            49.30
 B.coriacea            400                      0.349±0.004c                            78.84
Values with different superscript are significant different at p≤0.05, Mean ± SD; n=3
Table9. Nitric oxide free radical scavenging activity in the plasma of rats treated with ethanolic extract of B.
coriacea.
       Drug                  Dose (mg/ml)          Nitric oxide scavenging activity         % Inhibition
 Control               -                        1.660±0.038a                            -
 Normal                -                        0.549±0.049b                            66.93
 B.coriacea            400                      0.317±0.003c                            80.90
Values with different superscript are significant different at p≤0.05, Mean ± SD; n=3
Table10. Effect of ethanolic extract of Buccholzia coriacea on changes in SOD activity induced by
Paracetamol.
    SOD/Treatments            SOD in Liver                    SOD in Kidney                SOD in Serum
                            (Units/mg tissue)                (Units/mg tissue)          (Units/ml) (n=5)x103
    Control            28.802±2.919bc                  30.400±0.894c                    05.000±0.000a
    Paracetamol only   22.668±3.653a                   22.668±3.653a                    05.000±0.000a
    B.C +              31.656±1.775c                   26.670±0.000b                    05.000±0.000a
    Paracetamol
Values with different superscript are significant different at p≤0.05, Mean±SD; n=5
Key: B.C = Buccholzia coriacea

6                             International Journal of Research in Pharmacy and Biosciences V6 ● I3 ● 2019
Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in
Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

Table11. Effect of ethanolic extract of Buccholzia coriacea on changes in MDA status induced by Paracetamol
                        MDA in liver (Units/mg        MDA in kidney (Units/mg         MDA in serum (Units/ml
    Treatments         tissue) Mean±SD (n)x10-5       tissue) Mean±SD (n)x10-5       serum) Mean±SD (n)x10-5
Control               2.400±1.226ab                  1.755±0.219a                    0.481±0.258a
Paracetamol only      3.650±0.435b                   5.183±0.640c                    0.853±0.447b
B.C + Paracetamol     2.885±1.933ab                  1.794±0.657a                    0.506±0.185a
Values with different upper case are significant different at p≤0.05, Mean±SD; n=5.
Key: B.C = Buccholzia coriacea
Table12. Effect of ethanolic extracts of Buccholzia coriacea, Psidium guajava and Morinda lucida on changes
in CAT Activity induced by Paracetamol.
   CAT Treatments         CAT in Liver (Units in mg tissue)         CAT in kidney (Units in mg tissue) (n)x
                                 (n)x 10-1 K/minute                               10-1 K/minute
 Control                 0.433±0.012b                              0.442±0.013 ab

 Paracetamol only        0.382±0.033a                              0.465±0.015b
 B.C + Paracetamol       0.418±0.015b                              0.452±0.023b
Values with different upper case are significant different at p≤0.05, Mean±SD; n=5
Key: B.C = Buccholzia coriacea,
Effect of Ethanolic Extracts of Buccholzia Coriacea on Liver and Kidney Ultrastucture in
Paracetamol Induced Oxidative Stress
Histological Readings Gotten from the Rat’s Liver Examinations
Results on Liver
         Treatments                                              Observations
 Untreated control                No lesions seen
 Paracetamol only                 There was focal hepatocyte necrosis with heavy infiltrates of polymorphs.
 Paracetamol + B. coriacea        There was reduction in the area of necrosis and inflammation to a
                                  considerable degree.
Histological Readings Gotten from the Rat’s Kidney Examinations
Results on Kidney
         Treatments                                               Observations
 Untreated control                No lesions seen
 Paracetamol only                 There was an induction of in the patchy areas of acute tubular necrosis.
 Paracetamol + B. coriacea        Reduction in the tubular necrosis to focal area.
DISCUSSION                                                 al.,1997). The toxicity of NO increases greatly
B. coriacea has considerable amounts of phenols            when it reacts with superoxide radical, forming
and flavonoids as seen in the table 1 above.               the highly reactive peroxynitrite anion (ONOO-)
                                                           (Hue and Padjama, 1993). The nitric oxide
Nitric oxide is an essential bioregulatory
                                                           generated by sodium nitroprusside reacts with
molecule required for some physiological
                                                           oxygen to form nitrite. The extracts inhibit
processes like neural signal transmissions,
                                                           nitrite formation by directly competing with
immune response, control vasodilation and
                                                           oxygen in the reaction with nitric oxide. Table 1
control of blood pressure (Palmer et al., 1987;
                                                           illustrates the % inhibition of nitric oxide
Rees et al., 1989; Bredt and Synder, 1990; Gold
                                                           generation by the extract: B. coriacea and
et al., 1990). Nitric oxide has an important role
                                                           ascorbic acid in concentrations 25, 50, 100, 200,
in various inflammatory processes. Excessive
                                                           and 400 µg/ml which significantly scavenged in
production of this radical is directly toxic to
                                                           dose dependent manner with an IC50 of 21
tissues and contribute to the vascular collapse
                                                           µg/ml by B. coriace and 45.8 µg/ml by ascorbic
associated with septic shock, whereas chronic
                                                           acid. B. coriacea exhibited the better inhibition
expression of nitric oxide radical is associated
                                                           of nitric oxide formation in vitro when
with various carcinomas and inflammatory
                                                           compared with the ascorbic acid standard
conditions including juvenile diabetis, multiple
                                                           p
Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in
Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

The       ABTS-        (2,    2’-azinobis-3-ethyl-    acid in joint is an important feature of disease.
benzothiozoline-6-sulphonic       Acid)    radical    This plant was able to scavenge OH- radicals at
reactions involve electrons transfer and the          lower doses better than the standard mannitol.
process take place at faster rate when compared
                                                      Ferrozine can quantitatively form complexes
to DPPH- radicals. Re et al., 1999 reported that
                                                      with Fe2+. The complex formation can be
the decolouration of the ABTS-+ radicals also
                                                      disrupted by the presence of other complexing
reflects the capacity of an antioxidant species to
                                                      agents which cause a decrease in the red colour
donate electrons or hydrogen atoms to inactivate
this radical species. In the ABTS- radical cation     intensity of complexes. Substances or samples
scavenging activity, the extracts showed              that can reduce its colour intensity can be
concentration dependent scavenging activity.          considered as antioxidant through the
The percentage inhibition was observed to be          mechanism of inhibition of heavy metal. It was
concentration dependent. The IC50 of the extract      reported that chelating agents, which form σ-
is >400 µg/ml B. coriacea and 28.10 µg/ml             bonds with a metal, are effective as secondary
Quercetin standard. Buccholzia coriacea               antioxidants because they reduce the redox
exhibited a weak inhibition of ABTS* radical          potential, thereby stabilizing the oxidized form
when compared with the standard quercetin.            of the metal ion (Kumaran and Karunakaran,
                                                      2006). Our results showed that the extract is not
1,1-diphenyl-2-picrylhydrazyl (DPPH) has been         as potent as the standard EDTA, hence low or
used extensively as a free radical to evaluate        weak metal chelating activity.
reducing substances. In this model it was
observed that the tested extracts showed              It is known that paracetamol (PCM) induces
scavenging ability of DPPH* radicals in dose          liver injury through the action of its toxic
dependent manner with IC50 values of 52.80            metabolite,          N-acetyl-benzoquinoneimine,
µg/ml B. coriacea and 90.99 µg/ml Tannic acid         produced by the action of Cytochrome P-450.
as standard. The extract showed better                This metabolite reacts with reduced glutathione
scavenging activity when compared with the            (GSH) to yield non-toxic 3-GS-ylparacetamol.
standard p
Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in
Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

The decrease values of MDA formed in group           enzymes activities. Similar results on PCM
treated with extract plus paracetamol when           induced collapse of the antioxidant defense had
compared with paracetamol treated only group         earlier been reported (Uma et. al., 2010; Sabina
showed that the extract was able to offer            et. al., 2013; Dash et. al., 2007; Sowemimo et.
protection against cytotoxicity presented by the     al., 2007). This effect of PCM was well
paracetamol. In the serum, kidney and liver          tolerated by experimental animals receiving
tissues of rats treated with paracetamol only,       extract of M. lucida hence, preventing the
there were profound injuries to the tissues as       collapse of the antioxidant enzymes SOD and
seen from the histopathology results. However,       CAT (Fogha et. al., 2015). The observed
the damage was ameliorated in the group treated      increase of SOD activity suggests that the
with extract plus paracetamol. This implied that     aqueous extract of M. lucida stem bark have an
the extract was able to show protective effect       efficient protective mechanism in response to
possibly by the process of antioxidant due to the    oxidative stress and may be associated with
presence of phenolic compounds in them.              decreased oxidative stress and free radical-
                                                     mediated tissue damage (Fogha et. al., 2015).
Catalase is a phase II enzyme involved in the
conversion of hydrogen peroxide to water and         Super oxide dismutase (SOD) is associated with
oxygen. In this study, the level of catalase         increase in ROS which arises from adverse
activity was low in the liver of rats treated with   metabolic activities such as lipid peroxidation.
paracetamol only when compared with the              Activity of SOD was significantly low both in
extract treated group p
Free Radicals Scavenging Potential of Buccholzia Coriacea Extract and Ameliorative Effect in
Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

potentials in-vitro, and in-vivo against the                  Morinda lucida stembark protects paracetamol
oxidative stress induced by PCM by lowering                   induced liver damage. Int. J. Pharm. Sci. Rev.
the elevated levels of MDA, and increasing the                Res. 31: 198 – 204.
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compounded with PCM drug.                              [13]   Grice, H. C (1986): Safety Evaluation of
                                                              Butylated Hydroxytoluene (BHT) in the Liver,
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Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats

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 Citation: Fakoya, A., Olusola, A. O " Free Radicals Scavenging Potential of Buccholzia Coriacea Extract
 and Ameliorative Effect in Paracetamol- Induced Nephrotoxicity and Hepatotoxicity in Rats", International
 Journal of Research in Pharmacy and Biosciences, vol. 6, no. 3, pp. 1-11, 2019.
 Copyright: © 2019 Olusola, A. O. This is an open-access article distributed under the terms of the
 Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in
 any medium, provided the original author and source are credited.

International Journal of Research in Pharmacy and Biosciences V6 ● I3 ● 2019                                  11
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