INTRODUCTION:

The Liver is the leading structure in the human body and the place for a powerful breakdown. Liver diseases remain one of the serious health problems¹. The customary method of a remedy, particularly Ayurveda has put onward an integer of therapeutic plants and their preparations for the above situations. In this current age, it is very vital to deliver technical evidence to defend the abundant remedial usages of basils. Herbal drugs are assumed extensively even when their organically active constituents are unfamiliar because of their competence, fewer side effects and comparatively within budget². However, we are not cognizant of a pleasing therapy for severe liver illnesses and hunt for present and harmless drugs for liver ailments endures to be an area of interest. Ficus benghalensis (Moraceae) is a deciduous tree, which grows in hot and subtropical districts of Asia and is normally known as the banyan tree³.

In outmoded medication the aerial parts are used in the management of antioxidant consequence due to its polyphenolic components⁴, bark extract showed blood glucose dropping stuff⁵. The plant fruits and latex are used in a toothache⁶, diarrhoea⁷, menorrhagia⁸, nervous disorders⁹ and astringent¹⁰. The broad literature also proved that F. benghalensis plant studied for its Pharmacognosy¹¹, phytochemistry¹², antioxidant¹³, antiatherogenic¹⁴, antitumor¹⁵, anti-worm, anti-swelling¹⁶, analgesic¹⁷, an antidepressant¹⁸, antipyretic¹⁹, anti-histaminic²⁰, immunity increasing²¹ and wound curative²² properties. Personal information from users tinted that it is existence castoff in dismissing jaundice. Former investigations exposed the chemical arrangement of this F. benghalensis plant to have ketones (20-tetratriacontene-2-one, pentatriacontan-5-one and 6-heptatriacontene-10-one) in bark²³, flavonoids (quercetin-3-galactoside and rutin) in bark²⁴, 5, 7 Dimethyl ether of leucoperalgonidin-3-0-α-L-rhamnoside, 5, 3 dimethyl ether of leucocyanidin 3-O-β-Dgalactosyl cellobioside and 5, 7, 3 trimethoxy leucodelphinidin 3-O-α-L-Rhamnoside in bark, terpenoids (β-sitosterol friedelin, 3-friedelanol, 20-traxasten-3-ol, Betulinate and β-amyrin) in leaves²⁵, coumarins (Bergapten) in bark, esters (Keto-n-cosanyl stearate, phenyl tetradecanyl oleiate and hydroxypentacosanyl palmitate in bark and enzyme (Serine protease) in latex.

Previously it was reported the excerpts of different of F. benghalensis plant proved significant liver shielding in rats²⁶ with bark, with leaves. But no efforts have been made on F. benghalensis fruits for Liver shielding activity. F. benghalensis fruits are confirmed to be safe as they are edible by ancestral people. Because of the reported liver shielding of Ficus species and traditional claims, the fruits of F. benghalensis were evaluated in contrast to perchloromethane persuaded hepatic mutilation in rats with an ambition of evolving a natural Liver shielding drug.

Induction of liver toxicity in rats by perchloromethane is widely used for evaluating Liver shielding drugs/formulations. The Liver plays a vital part in the biotransformation of perchloromethane, which depends on cytochrome P-450 enzyme²⁷. The process of lipid peroxidation is originated by free radicals, which causes inhibition of enzyme activity. Trichloromethyl and its peroxy radicals are produced after biotransformation, which binds to H⁺from unsaturated fat, originating fat peroxidation and liver mutilation²⁸.

MATERIALS AND METHODS:

Collection and Identification of Plant Material:

The branches and fruits of Ficus benghalensis were unruffled from plants growing around Anantapur, AP, India. F. benghalensis was recognized and confirmed genuineness in the Dept. of Botany, SK University, Anantapur, AP, India. The voucher specimen (number: SKUB 0084/18) was obtained and kept at the Herbarium of SK University, Anantapur.

Materials:

Ether, trichloro acetic acid, sodium hydroxide, acetone, chloroform and Ethanol (95%) were procured from Fisher Scientific, Secunderabad, Telangana, India. Enzyme kit was from Span Diagnostic Ltd. Deionized water was used in the experiment whenever desirable.

Preparation of Extract:

Fresh fruits were cut, seeds were detached, wash with water and drifted with ether (60±5^oC) and further distinctly extracted with ethanol and water by soxhlet extractor for 72h. The solvent was detached underneath reduced pressure and semisolid mass obtained dried in vacuum to a solid residue yield (7.4 %w/w and 5.9% w/w respectively). The chemicals of the extracts²⁹ were recognized by qualitative examination tracked by their authorization by thin-layer chromatography.

Refinement of the Mucilage:

F. benghalensis mucilage was homogenized using icy 5 % dilute trichloro acetic acid. Later rotated at 3,500rpm for 20 min, nullified with NaOH by dropwise mixing and finally made it to dialyze for 30 h likened to distilled water. The mucilage was chelated with 3 times the quantity with ethanol and washed uninterruptedly with ethanol (90%), propanone and ether³⁰.

Selection and maintenance of animals:

New Zealand albino rats (male) with 200-250g weight were employed for the investigation (procured from National Institute of Nutrition, Hyderabad, India). The New Zealand albino rats were upheld at a temperature of 25±2^oC and RH of 50±5% (standard laboratory situations) with dark and light photo stages. Marketable pellet diet (Varna Enterprises, Bangalore) and water were given when anticipated. The investigational protocol has been putative by the IAEC and by the Regulatory Body of the Govt. (Reg. no. 878/ac/05/CPCSEA/004/2015).

Acute toxicity studies:

The serious oral poisonousness findings were conceded out as per the strategies of the Organization for Economic Co-operation and Development (OECD), draft guidelines 423 acknowledged from CPCSEA, Ministry of Social Justice and Empowerment, Govt. of India. Acute toxicity studies on ethanolic and aqueous extracts of F. benghalensis fruits (EEFG and AEFG correspondingly) are approved out at a dose range of 50 mg to 3,000mg/kg b.w (oral) and the numeral of rats that croaked in a 7-days next a single dose was chronicled. The rats were also carefully inspected for ciphers of inebriation, lethargy, behavioural alteration and indisposition³¹.

Liver shielding consequence counter to perchloromethane- tempted toxic hepatitis in rats:

The detailed depiction of groups and integer of rats used were clarified in Table 1.

Table 1: The full representation of groups and numeral rats used

Constraint	Explanation
Animals used	New Zealand albino rats.
Number of groups	5
Rats per group	6
Drug administered	Perchloromethane(1ml/kg) to all rats by S.C. route.
Group-I	Normal saline (10 ml/kg) by i.p route (control)
Group-II	Liv-52 (25mg/kg) by i.p route (drug reference)
Group-III	Perchloromethane (1ml/kg i.p).
Group-IV	aqueous extract of F. benghalensis (300mg/kg) (test)
Group-V	Ethanolic extract of F. benghalensis (300mg/kg) (test)
Testing duration	15 days

The different steps involved in sampling and detections as shown in figure 1.

Figure 1: Steps involved in sampling and estimation

Histopathological studies:

The histopathological readings in the present investigation were illustrated in figure 2.

Figure 2: Process of histopathological studies of rat’s liver

Statistical Analysis:

The results are communicated as average ±S.D. The variance amongst experimental groups was treated by one-way ANOVA (toxic control vs. treatment), Bonferrioni’s method (utilizing Sigma stat statistical software-4.0) and was measured statistically considerable when p< 0.05.

RESULTS AND DISCUSSION:

Liver shielding actions of F. benghalensis fruits:

Both alcoholic (AEFB) and ethanolic extracts (EEFB) of fruits of F. benghalensis were found to be practically non-toxic since no mortality was observed even at the dose of 3,000mg/kg body weight. Hence, the biological dose was fixed 300mg/kg for EEFB and AEFB. Rats treated with perchloromethane developed noteworthy liver mutilation and showed elevated serum concentration of hepato-specific enzymes as well as an unadorned amendment in other biochemical strictures (P<0.001). The values of the biochemical constraints were perceived to be increased in the perchloromethane intoxicated rats. Treatment with EEFB and AEFB decreased the perchloromethanebrought alterations in AST, ALT, alkaline phosphatase and total bilirubin³². It is found that both the extracts offer protection in contradiction of the toxin as shown by a remarkable lessening in all biochemical stricture. Histopathological readings established that perchloromethane causes focal necrosis, portal infiltration, fatty change, kupffer cell hyperplasia and hydropic change. In the conserved groups, necrosis which is an extra severe form of grievance is markedly banned; a milder form of injury like fatty change and abridged necrosis continued in both the extracts.

The EEFB fruits, administered prophylactically, unveiled important protection alongside perchloromethane persuaded liver injury. High lipid peroxidation values indicate excessive free radical convinced peroxidation. Pre-treatment of animals with EEFB and Liv-52 prevented the perchloromethaneencouraged a rise in serum level of transaminases and total serum bilirubin, confirming the protective properties of EEFB fruits counter to perchloromethaneprompted liver mutilation. The liver shielding of F. benghalensis (500mg/kg) was comparable with the activity of standard Liv-52 (100mg/kg). However, there was no result on the increase in serum alkaline phosphatase concentrations by the test extract and Liv-52. A relative histopathological study of the livers from dissimilar groups further validated the Liver shielding potential. In animals treated with ethanolic extract and Liv-52, the rise in lipid peroxides in liver tissue homogenate was disallowed expressively. The diminution in lipid peroxides owing to the antioxidant concern of the extract. A likely mechanism of the F. benghalensis extract as Liver shielding e may be owing to its antioxidant upshot or reticence of cytochrome P450s which damage the bioactivation of perchloromethane into their consistent reactive species. The preliminary phytochemical studies designated the occurrence of steroids/ triterpenoids and their glycosides and coumarins in the EEFB. Since coumarins have liver shielding activity, it may be speculated that these constituents of Ficus are been accountable for the detected protective possessions. However, the role of steroids/triterpenoids cannot be ruled out. The result of F. benghalensis concentration of liver components in the serum of rats (table 2) was exemplified in figure 3.

Table 2: Data for the result of F. benghalensis fruit extracts on perchloromethaneprompted toxic hepatitis in rats

Group	Treatment (p.o)	Biochemical Parameters
Group	Treatment (p.o)	AST (U/I)	ALT (U/I)	ALP (IU/L)	ACP (U/L)	Total Bilirubin (mg/dl of blood)
I	Normal saline (control)	45.8±0.51	29.5±1.28	18.5±1.11	9.2±0.02	0.29±0.01
II	perchloromethane extract	141.5±2.07	151.5±1.67	97.5±2.25	35.1±1.01	0.94±0.02
III	Liv-52 (standard)	89.3**±2.52	49.8**±1.34	23.9**±0.46	10.2*±0.05	0.26*±0.01
IV	Ethanolic extract	91.5**±1.20	59.1**±1.07	28.4**±0.13	11.5*±0.05	0.39*±0.02
V	Aqueous extract	104.1**±3.65	78.8**±1.07	32.8**±1.01	15.1*±0.02	0.49*±0.01

Values in mean ± S.E.M; Number of trials (n) =6; *P<0.01 Vs. Control; **P<0.001 Vs. Control

Table 3: The histological comments in rats of different groups

Part of the liver	Group
Part of the liver	I	II	III	IV	V
Liver tissue	Typical lobules	Typical lobules	Necrosis	Typical lobules	Typical lobules
Nuclei	vesicular	vesicular	degenerated	vesicular	vesicular
Individual lobules	consist of hepatocytes arranged as cords radiating around centrally placed terminal hepatic veins	consist of hepatocytes arranged as strings scorching nearby centrally located terminal hepatic veins	Injured, degenerated and necrosed	Liver cells organized as threads searing around centrally sited terminal hepatic veins	Hepatocytes arranged as leads baking around centrally sited terminal hepatic veins
Hepatocytes	Uniform in size, polyhedral in shape, with centrally located large nuclei	showed variable size There is a mild increase in fibrous connective tissues	Hepatocytes show ballooning degeneration and steatotic changes	Even and polyhedral in shape, with a clear nuclei	Adaptable size with a little hike in fibrous connective tissues
cytoplasm	Strongly eosinophilic with a fine basophilic granules	Strongly eosinophilic with basophilic granules	Eosinophilic with inadequate cytoplasm	Eosinophilic with an adequate basophilic granules	Moderately eosinophilic with basophilic granules
Portal tracts	Terminal twigs of the hepatic portal vein and hepatic artery (PV andHA) at the periphery in a fibrous stroma	Liver PV and HA at the boundary in a fibrous stroma	Some amount of fibrosis seen in portal tracts.	Terminal divisions of the PV and HA at the periphery in a fibrous stroma	Terminal divisions of the PV and HA at the fringe in a fibrous stroma
Impression	Normal	Recovering cells	Mild necrosis and fibrotic changes	Redeveloped cells	Regenerated cells

Figure 3: The concentration of the liver component in serum in various groups

Histological Observation:

The histological observations in rats of various groups were illustrated in table 3 and shown in figure 4.

Figure 4: Microphotograph of rat liver section with various treatments at 200x A) Normal B) perchloromethane C) Liv-52 D) Ethanolic extract E) Aqueous extract

CONCLUSION:

The present work was revealed liver protecting potential of Ficus benghalensis fruit extracts in perchloromethane induced toxic hepatitis in New Zealand albino rats. The ethanolic extract of F. benghalensis fruits were evaluated for Liver shielding potential in New Zealand Albino rats. F. benghalensis fruit extract showed liver protection by reducing serum concentrations of alanine aminotransferase, aspartate aminotransferase, total serum bilirubin and malondialdehyde. This study concludes the potential liver protecting activity of F. benghalensis fruit extracts.

ACKNOWLEDGEMENT:

The authors are grateful to the RERDS-CPR of RIPER for providing facilities for this work.

CONFLICT OF INTEREST:

The authors declare no conflict of interest.

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Received on 02.12.2019 Modified on 11.03.2020

Research J. Pharm. and Tech. 2021; 14(7):3739-3743.

DOI: 10.52711/0974-360X.2021.00647