PHYTOCHEMICAL ANALYSIS AND ANTIOXIDANT POTENTIAL OF FICUS BENGHALENSIS L

Genus Ficus belongs to family Moraceae having 40 genera and over 1000 species worldwide. Different methods have been used for phytochemical screening of medicinal plants like total phenolics content (TPC) and total flavonoids content (TFC) assays to quantify phenolics and flavonoids. The phytochemical analysis exhibited highest total phenolics content in M extract of stem and total flavonoids content in ethyl acetate (EA) extract of leaves i.e. 61.2±1.3 μg GAE/mg extract and 25.1±0.9 μg QE/mg extract respectively. Total reduction power and total antioxidant capacity were maximum in the M extract of stem i.e. 243.89±1.6 μg AAE/mg extract and 127.08±2.7 μg AAE/mg extract respectively.


INTRODUCTION
Genus Ficus belongs to family Moraceae having 40 genera and over 1000 species worldwide.It is also known as fig which plays an important role in ecosystem by providing fruit throughout the year to the insects, birds and animals (Chaudhary et al., 2012).F. benghalensis leaves contain leucopelargonin, bengalenoside, rutin, β amyrin along with psoralen, β sisterol, bergapten, quercetin-3-galactoside20, leucodelphinidin derivative, A glucoside and leucocynidin derivatives (Joseph and Raj, 2010b).

Phytochemicals
are chemically diversified group of compounds produced by plant as a part of defense against pathogens, predators and competitors.They have specific chemical properties that are useful for humans in treatment of diseases such as antioxidant, antimicrobial, interference with DNA replication and inhibition or stimulation of enzymes (Halliwell, 2007).Phytochemicals responsible for broad spectrum of bioactivities include alkaloids, glycosides, tannins, polyphenols, flavonoids and many more (Joseph and Raj, 2010a).Flavonoids are bioactive leads with the ability to chelate and reduce metal ions (Nazli et al., 2018).Different methods have been used for phytochemical screening of medicinal plants like total phenolics content (TPC) and total flavonoids content (TFC) assays to quantify phenolics and flavonoids (Angelova et al., 2008).
Antioxidants in optimal range inhibit the process of oxidation to retain normal physiology of body.Plants possess numerous antioxidants which play an essential role as radical scavengers and transform free radicals into less reactive species.(Shahwar et al., 2012).Antioxidant potential is determined by a set of different assays because one technique is not sufficient to predict exact antioxidant potential.These techniques involve different mechanisms e.g.prevention of chain initiation, scavenging of radicals and disintegration of peroxides (Aliyu et al., 2012).

Phytochemical analysis a) Assessment of TPC
A well stated protocol recounted by Fatima et al. (2015) was adopted for the estimation of total phenolics content in test extracts.
The results were expressed as µg gallic acid equivalent (GAE)/mg extract of the plant.

Procedure
A well stated procedure described by Khan et al. (2015) was performed for the estimation of total flavonoids content in test extracts (4 mg/ml DMSO).The results were expressed in µg quercetin equivalent (QE)/mg extract of plant.

Procedure
A well-defined procedure described by Ahmed et al. (2017) was used.Samples that showed a scavenging potential of more than 50% at initial concentrations were further analyzed for their IC50 values using threefold serial dilution methodology.

Procedure
A well stated procedure described by Fatima et al. (2015) was adopted for the calculation of TAC of test extracts.The results were stated as µg ascorbic acid equivalent (AAE)/mg extract.

Procedure
A well-defined procedure elaborated by Zahra et al. (2017) was followed for the estimation.1).

Antioxidant assays a) DPPH assay
A Free radical scavenging assay was performed and change in color from purple to yellow was measured.Maximum reducing activity in fruit was shown by M extract with an IC50 value of 3.18 µg/ml.In case of adventitious root, highest potential was observed in DW extract with an IC50 value of 4.10 µg/ml, while the M extract of leaves showed maximum potential with an IC50 value of 10.02 µg/ml.Highest activity in bark was shown by M extract with an IC50 value of 13.41 µg/ml, while DW extract of stem showed maximum potential with an IC50 value of 16.67 µg/ml.In case of root, M extract showed highest activity with an IC50 value of 20.98 µg/ml.

Phytochemical Analysis
Medicinal plants naturally synthesize aromatic secondary metabolites which are responsible for pharmacological response in humans.
Therapeutically active subcategories include phenolics, tannins, alkaloids, flavonoids etc. (Angelova et al., 2008).Second most abundant secondary metabolite in plants are hydroxylated phenolic substances called flavonoids.They are responsible for antioxidant properties due to presence of hydroxyl group.Antioxidant properties play important role in reduction of oxidative stress, cytotoxicity and strengthen oxidative defense by scavenging free radicals (Chang et al., 2002).Maximum TFC was shown by EA extract of leaves.The possible reason is the solubility of flavonoids in medium polarity solvents.Other species of Ficus such as F. religiosa also show maximum flavonoids in EA extract (Sultana et al., 2009).

Antioxidant assays
Oxidation in biological systems is a natural phenomenon which results in formation of highly reactive peroxyl and hydroxyl radicals.These radicals ultimately cause damage to DNA, protein and polyunsaturated fatty acid residues of cell membrane and lead to pathological effects such as cancer and vascular diseases.Immune system inactivates the reactive species but the overburden of radicals needs exogenous supply of antioxidants.Antioxidants are integral part of plants as secondary metabolites which play important role as free radical scavengers and by converting highly reactive free radicals into less reactive species (Moon and Shibamoto, 2009).Different studies verify the significant role of antioxidants in reduction of oxidative stress, which provoke us to determine the antioxidant potential of various extract of F. benghalensis.The antioxidant capability of samples cannot be assessed by a single assay.Therefore, DPPH free radical scavenging, total reduction power and TAC assays were performed to verify the antioxidant potential.
DPPH is the stable free radical and antioxidant potential of crude extract was determined on the basis of scavenging of free radical i.e.DPPH.The principle of the assay is based on the conversion of purple color of the free radical to the yellow color molecule by accepting a hydrogen electron from donor antioxidants present in samples (Floegel et al., 2011).In the current study, high percentage free radical scavenging activity was shown by M extract of fruit.Various studies expressed linear relationship between TPC and reducing activity (Roy et al., 2010).Significant TPC was also detected in M extracts, which might be responsible for free radical scavenging activity.
TAC of various samples of F. benghalensis was determined by phosphomolybdenum based calorimetric assay.The principle of the assay is based on reduction of Mo (VI) to Mo (V) by natural antioxidant and formation of green color complex which show maximum absorption at 695 nm (Pellegrini et al., 2003).The highest TAC was expressed by M extract of stem.Previous studies also indicated that M extract showed maximum antioxidant activity due to presence of polyphenols in polar solvents.Studies indicate positive correlation between polyphenols and TAC (Kumaran and Karunakaran, 2007).
Total reducing power is based on the presence of reductones which are associated with antioxidant activity by breaking the radical chain by donating a hydrogen atom (Wong et al., 2006).Maximum reducing power was shown by the M extract of stem.Studies performed on certain plant extracts proved that polyphenols and reducing power has direct correlation so results were in agreement with the previous findings (Ou et al., 2002).

CONCLUSION
The phytochemical analysis exhibited highest TPC in M extract of stem and TFC in EA extract of leaves i.e. 61.2±1.3 µg GAE/mg extract and 25.1±0.9µg QE/mg extract respectively.

Figure 1 .
Figure 1.TPC and TFC of samples.Values presented are expressed as a mean of triplicate ± standard deviation.