INTRODUCTION:

Liver is a vital organ that plays a role in controlling critical biochemical and physiological activities including homeostasis, growth, energy and nutrient supply, detoxification of Drugsand otherxenobiotics, and also combating infections^[1,2].

Therefore, it is very susceptible to being damaged by hepatotoxic agents^[3]. The Liver injuries induced by various hepatotoxins have been recognized as a major toxicological problem for years. Liver Cirrhosis is one of the liver injury occurs due to the necrosis of liver cells by fibrosis and nodule formation. Impairment in function and structureof liver leads to impaired liver blood flow and function.^[4]

Cirrhosis is defined as a diffuse hepatic process characterized by fibrosis and the conversion of normal liver architecture into structurally abnormal nodules.

Liver cirrhosis has become one of the major causes of morbidity and mortality. Plants are a rich source used for centuries to cure various diseasesand disorders from natural products.

Hence, it is necessary to find out new formulation from natural sources that have less expensive, least side effects and on a long term therapy which can provide better safety and efficacy However, there are a number of herbal formulations available on liver disorders in ayurvedicmedicine.^[5] Various formulations like Liv-52,^[6]Kamilari,^[7] APCL-A AmalkadiGhrita,^[8]and Panchagvya Ghrita, Himoliv,^[9]are well known for their Hepatoprotective effects. Polyherbal combination contain different herbal plant, they have as different pharmacological activities, the plant like Terminalia chebula, Terminaliabelerica, Emblica officinalis, azadiracta aindica, swertiachirata, Picrorhiza kurroo, adhatodavasica, tinosporacardifolia.

MATERIAL AND METHODS:

Material:

The various crude drugs Terminalia chebula, Terminalia bellerica, Emblica officinalis, Azadiracta indica, Swertia chirata, Picrorhoza kurroa, Adhatoda vasica, Tinospora cardifolia were used for livodac formulation.

Methods:

1] Preparation of Polyherbal formulation:

All crude drug powders were collected and mix by using doubling up method and the tablets were prepared by direct compression method.

2] In vivo model:

A] Acute oral toxicity study:

Acute toxicity test was done for the livodac tablet following OECD guidelines 425. Female Swiss albino mice used for the study. The limit test at single dose administration of 2000mg/kg and 5000 mg/kg of the livodac tablet was conducted, it was observed up to fourteen days and no any sign of toxicity and mortality was observed in 2000mg/kg dose of livodac tablet.

B] Procedure:

36 either sex Wistarrats weighing 200-250 g were selected for the study. All these rats were divided into six groups. In that the six groups, Group I animals served as normal control, animals belonging to Group II served as positive control, animals belonging to Group III served as standard and animals belonging the group IV, V, VI are the groups received different doses low dose (100mg/kg), mid dose (200mg/kg), and high dose (400mg/kg) of test formulation. The group first animal received CMC 1%. Group second give Paracetamol (1% CMC), group third animals received Silyamarin tablet (54mg/kg) with Paracetamol (2gm/kg) and group fourth, fifth and six give Paracetamol (2gm/kg) and livodac formulation in doses low dose (100mg/kg), mid dose (200mg/kg), and high dose (400mg/kg). All the animals were weighed before (day1), during (day5), and after (day10) the drug treatment. After completion of the treatment, animals were anaesthetized under light chloroform anesthesia. Blood was collected by retro-orbital plexus puncture. Biochemical estimations of various parameters such as total proteins, total Bilirubin, serum glutamic-oxaloacetictransaminase (SGOT), serum glutamic pyruvic transaminase (SGPT), and alkaline phosphatase (ALP) using serum are assessed. Animals are sacrificed, and liver was removed, weighed and perfused in 10%formalin solution for histopathology studies.

Group. No.	Treatment	No. of animal
I	Normal control: CMC1%	6
II	Positive control: Paracetamol (2gm/kg)	6
III	Standard control: Silyamarin (54mg/kg body weight) + Paracetamol (2gm/kg body weight)	6
IV	Super therapeutic: Testdrug (400mg/kgbody weight) +Paracetamol (2gm/kg body weight)	6
V	Therapeutic: Test drug (200mg/kg body weight) + Paracetamol (2gm/kg body weight)	6
VI	Sub therapeutic: Test drug(100mg/kg body weight)+ Paracetamol (2gm/kg body weight)	6

Evaluation Parameter:

The rats were sacrificed on the eleventh day by using chloroform and blood was collected in plain tubes. The various blood tests SGOT, SGPT, Total Bilirubin, Total Alkaline Phosphate, Total protein were done.

Statistical Analysis:

The data were statistically analyzed and allvalues were expressed as Mean ±SEM. The datawere also analyzed by One Way ANOVA using Graph pad prism 8 versionSoftware followed by Dunnets multiple comparisons tests.

Table No. 1 Effect of livodac tablet on Bilirubin, alkalinephosphate, total protein, SGOT, SGPT on 11^th day.

Groups	Bilirubin	ALP	SGOT	Total Protein	SGPT
NC	0.403±0.01***	112.8±17.08***	32±1.00**	6.82±0.21*	32.76±1.16***
DC	0.82±0.10	291.5±10.25	39.26±2.45	8.07±0.50	44.35±0.98
STD	0.331±0.02***	127.3±13.79***	31.2±0.61***	6.30±0.32**	31.33±0.67***
T100	0.283±0.02***	226.6±8.56**	23.98±1.28***	5.91±0.11***	25.68±1.21***
T200	0.518±0.02***	211.5±15.79***	24.76±0.55***	6.04±0.30***	28.08±1.12***
T400	0.396±0.02***	188.31±6.20***	26.35±0.65***	6.50±0.33**	29.03±1.71***

Values are expressed as mean ± S.E.M. (n=6). *P<0.05 **P<0.01 ***P<0.001 when compared with disease control (ANOVA followed by Dunnets test).

Histopathology of liver:

Fig 1.9 Group: NC Pathological change: NAD

Fig 1.10 Group: DC Pathological change: Moderate (+++)

Fig 1.11Group: STD Pathological change: Mild (++)

Fig 1.12 Group: Test (sub therapeutic) Pathological Change: Moderate (+++)

Fig 1.13test (super therapeutic) Pathological change: Mild (++)

Histopathology of kidney

Fig1.14Group: Test (Therapeutic)Group: Pathological change: Moderate (+++)

Fig1.15Group: NC Pathological change: NAD Pathological change: NAD

Fig1.16 Group: DC Pathological change: Moderate (+++)

Fig1.17 Group: STD Pathological change: Mild (++)

Fig1.18 Group: Test (sub therapeutic) Pathological Change: Mild (++)

Fig 1.19 Group: Test (Therapeutic) Test Pathological pathological change

Fig1.20 Group: Mild (++) change: Moderate (+++)

The severity of various features of liver and kidney was evaluated based on following scoring scheme

NAD	No abnormality detected
+	Minimal changes in organ histopathology
++	Medium changes in organ histopathology
+++	Moderate changes in organ histopathology
++++	Severe changes in organ histopathology

DISCUSSION:

Paracetamol, an over the counter drug, is a commonly used antipyretic and analgesic which can lead to liver damage if taken in over dose.^{[10] [11]}

The hepatotoxins are associated with changes at cellular levels that may lead to deterioration of organ functions. Therefore, any improvement in the treatment of hepatic function could be of potentially a great importance. The possiblemechanism of herbal preparation as Hepatoprotective agent against Paracetamol could be by substantially decreasing lipid per- oxidation through the elevation of MDA level in liver homogenate. Generally it is known that most of the Paracetamol is excreted by conjugating with glucuronate and sulphate, while metabolized by cytochrome p-450 system to produce a highly toxic N acetyl- p- benzoquinone- imine (NAPQI) which is readily detoxified by enzymatic conjugation with hepatic glutathione (GSH). But, when the detoxification process is disturbed, an active agent NAPQI is produced which in turn binds covalently to tissue macromolecules thereby causing severe hepatic damage.^[12,13]

The overall body weight of rats in livodac tablet received groups was significantly decreased as compared with Paracetamol induced hepatotoxicity models in rats. This indicates the prevention of hepatotoxicity related fatty changes in liver and improvement in liver function which leads to overall enhancement in metabolism and health by the entire livodac tablet received groups. When the liver tissues from all the groups were subjected to histopathological examination, it was observed that liver tissues of normal control group showed normal cellular architecture with distinct hepatic cells, sinusoidal spaces, and central vein.

Whereas Paracetamol treated liver tissues showed signs of toxicity as necrotic cells, inflammation, hemorrhage, fibrosis, and even fatty changes. In standard control these signs of toxicity were not demonstrated; the tissues showed normal architecture, but focal enlargement was noted in Standard group.

When the kidney tissue from all the groups were subjected to histopathological examination, it was observed that liver tissues of normal control group showed normal cellular architecture with distinct kidney cells, Whereas Paracetamol treated kidney tissues showed signs of toxicity were abnormal changes like tubular swelling, as well as cellular infiltration and degeneration of nodular tissue, and the Paracetamol containing group changes in cellular. In all the groups of livodac tablet, i.e. low(100mg/kg), medium(200mg/kg) and high dose(400mg/kg), successfully preserved the normal cellular architecture of liver and prevented the cellular damage caused by Paracetamol. Therefore, from the results, we could conclude that all the formulations have been successful in preventing the pathological changes generated by Paracetamol.

All the above findings suggest that high (400mg/kg), medium (200mg/kg), and low-dose (100mg/kg) formulation groups show significant Hepatoprotective activity but in medium dose (200mg/kg) was more Hepatoprotective effect in terms of reduction in serum levels of liver enzymes such as SGOT, SGPT, and ALP that are usually released in response to the damage to hepatic parenchyma. Lowering of total Bilirubin indicates improvement in liver function.

Oral administration of livodac tablet was standardized and has a significant Hepatoprotective activity in preventive treatments against hepatotoxins induced hepatic damage.

CONCLUSION:

The livodac tablet was Hepatoprotective and which could be safe with no interactions and beneficial in Hepatoprotection. The biochemical studies revealed a dose dependent significant fall in the levels of SGOT, SGPT, ALP, total Bilirubin, total protein a decrease in the body weight of livodac tablet treated animals against Paracetamol induced hepatotoxic animals an increases the body weight.

Histopathological studies supplemented the findings by showing mild hepatic degeneration with absence of necrosis in comparison with the model control. Thus indicating the prominent significance of livodac tablet in hepatoprotection against Paracetamol induced hepatotoxicity.

REFERENCES:

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Received on 09.06.2019 Modified on 01.07.2019

Res. J. Pharmacology and Pharmacodynamics.2019; 11(3): 83-88.

DOI: 10.5958/2321-5836.2019.00014.4

Groups	NC	DC	STD	T100	T200	T400
Day1	244.16±2.71	229.14±5.68	234.16±6.50	234.16±1.57	238.16±1.57	248±0.85
Day5	229.5±7.13***	274.16±7.12	248.5±1.99**	250.5±1.47**	253.16±1.53**	250±2.23**
Day11	229.16±6.37***	283.33±4.77	247.3±3.81***	246.6±6.00***	237.5±4.95***	253.3±3.57***