Introduction

Humans are continuously exposed to a multitude of pollutants present in the

environment, the food they eat, the air they breathe, and the water they drink (Humblet et al.,
2008). A major challenge in risk assessment is to determine the degree of exposure to multiple
chemicals and the hazards associated with such exposure (Rider et al., 2010). The risk to human
health posed by exposure to these organic pollutants has been one of worldwide concerns.
“POPs” (Persistent Organic Pollutants), a class of toxic chemical pollutants that are widely
present in the environment and that are harmful to human health and to the wildlife. POPs
appear as toxic contaminants in human food in all regions of the world.
The exposure of POPs, in which, dioxin is one example of a complex group of chemicals
is of concern from a human health perspective.
Dioxins are known to be a class of highly toxic, widespread, and persistent
environmental contaminants that include 75 polychlorinated dibenzo-p-dioxins (PCDDs), 135
polychlorinated dibenzofurans (PCDFs), and 12 polychlorinated biphenyls (coplanar PCBs),
which are structurally similar to PCDDs and PCDFs. They received serious concerns from the
general public owing to their potent toxicity, ubiquitous presence in the environment, and their
tendencies to accumulate in humans and wildlife. Dioxin is extremely stable, because of its high
lipophilicity and water insolubility. Dioxin concentrates in sediments and is incorporated into
the fatty tissue of fish, birds, reptiles, and mammals. Much of its presence in plants is due to
atmospheric transport on particles, resulting in settling on the leafy tissues of plants.
Dioxins are generated naturally, through such processes as forest fires and volcanic
eruptions. Human activity has been primarily responsible for the generation of this class of
chemicals over the past two centuries. Historically, industrial activities were the largest sources
of dioxins, but many responsible processes have since been regulated, and today uncontrolled
combustion such as burning of household waste in open containers is the greater concern.
Formed as by-products, these halogenated aromatic compounds are produced during chlorinated
phenoxy herbicide manufacturing, pulp and paper bleaching, metallurgy, and during the
combustion of tobacco as well as wood and fossil fuels (coal, oil and gasoline) and municipal
and hospital incineration (Zook and Rappe, 1994). Dioxin is a global threat, and it requires a
global commitment to combat it. There is an urgent need to ensure that countries can get the
tools and build the base for reducing or eliminating releases of persistent organic pollutants.
The EPA believes that people who eat rich diet containing dioxin may face a 1-in-100
risk of developing some form of cancer, diabetes or immune system disorders.
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 Introduction

India Scenario:
Indians are yet to realize the gravity of dioxin contamination and its related health
effects. The government has not conducted any study to find out the level of dioxin exposure in
the population. Two recent studies, based on human samples, have been found to be in very high
amounts in samples of Indian breast milk, meat and dairy products (International Symposium on
Halogenated Environmental Organic Pollutants and POPs, 2001).
Till date, India has been refusing to acknowledge that dioxin is a problem. In the
recently concluded UNEP Convention on Persistent Organic Pollutants, India was silent on the
dioxin issue treating it as ‘not our problem’.
Liver:
The liver is the largest organ in the body, contributing about 2% of the total body
weight, or about 1.5 kg in the average adult human. It plays an astonishing array of vital
functions in the maintenance, performance and regulating homeostasis of the body. It is the
prime organ concerned with almost all the biochemical pathways to growth, fight against
disease, energy provision, metabolism of nutrients such as carbohydrates, proteins and lipids,
excretion of waste metabolites and detoxification of the exogenous and endogenous challenges
like xenobiotics, drugs, viral infections, chronic alcoholism, environmental pollutants and
chemotherapeutic agents (Ward and Daly, 1999).
Any injury to it or impairment of its function may lead to many implications on one’s
health. Therefore, maintenance of a healthy liver is essential for the overall well being of an
individual. Liver diseases are mainly caused by toxic chemicals (certain antibiotics,
chemotherapeutics, peroxidised oil, aflatoxin, carbon-tetrachloride, chlorinated hydrocarbons,
etc.), excess consumption of alcohol, infections and autoimmune disorder.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD):

Fig.1: Structure of TCDD

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 Introduction

Of the 75 possible congeners, the tetrachloro molecule with chlorine atoms laterally
located at positions 2,3,7 and 8 (TCDD) is the most potent, persistent and ubiquitous
environmental contaminant (van den Berg et al., 2006). TCDD has been most intensively
studied for toxicity, and it has been classified as a human carcinogenic substance by the
International Agency for Research on Cancer (Srogi, 2008) and the health impact of exposure to
TCDD is of great concern to the general public (Turkez et al., 2012a, b and c). TCDD being
approximately 10000 more toxic than potassium cyanide (Sweeney et al., 2000).

History:
In 1947, X-disease, a hyperkeratotic condition akin to chloracne, which could culminate
to death, was first reported in American cattle, and was later, demonstrated to arise from
exposure to dioxin-like compounds (Bell, 1953). In 1949, an explosion in a Monsanto chemical
plant in Nitro, West Virginia resulted in the exposure of workers to the dioxin-contaminated
herbicide, 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), and persistent chloracne was observed in
the exposed individuals (Suskind et al., 1984). US EPA issued its first health assessment of the
most toxic dioxin molecule, TCDD, in 1985. Shortly thereafter, in 1991, the National Institute
of Occupational Health and Safety (NIOHS) reported its cancer mortality study of US workers
exposed to TCDD, and found that mortality was increased from all cancers combined,
particularly among those with the longest occupational exposure and the greatest latency from
exposure (Fingerhut et al., 1991).
TCDD gained notoriety and the public's interest as a result of environmental disasters at
places such as Love Canal, New York; Times Beach, Missouri; Pensicola Florida; and the
herbicide, Agent Orange, used in Operation Ranch Hand during the Vietnam conflict.
Half-life:
Much of the concern with exposure to TCDD is due to their environmental and
biological persistence, which may result in the bioconcentration and bioaccumulation of the
chemicals up to the food chain. The half-life of TCDD is approximately 10 to 14 days in rats
and mice, while in humans the half-life has been estimated at 7 to10 years (Miniero et al.,
2001). This bio-persistence is thought to be an important factor in dioxin’s toxic effects.

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 Introduction

Route of exposure:
The main route of human exposure to dioxin is through the diet, primarily through the
consumption of contaminated meat and fish. Once ingested, dioxin preferentially accumulates in
lipid rich tissues due to its hydrophobic nature.

Fish Human Cattle Plants

Planktons
Air Soil
Water
DIOXIN
Automobile Exhaust
Incinerator
PVC Production Disposal

Backyard Burning
Building Fires
Waste Pesticide

Fig.2: Anthropogenic sources of dioxins

Mechanism of Action of TCDD:

TCDD is a ligand of the aryl hydrocarbon receptor (AhR) and all major toxic effects of
TCDD and related halogenated aromatic hydrocarbons appear to be mediated by a soluble
protein, the aryl hydrocarbon receptor (AhR) (Bunger et al., 2003 and Walisser et al., 2004).The
binding of TCDD to the AhR results in translocation of the receptor complex to the nucleus,
where it dimerizes with another basic helix-loop-helix PAS protein known as AhR nuclear
translocator (ARNT).
This heterodimer then binds specific DNA elements, termed dioxin response elements
(DREs), in the regulatory regions of target genes leading to changes in gene expression.
Through such binding, AhR up-regulates the expression of several downstream genes including
those encoding xenobiotic metabolizing enzymes such as Phase I (e.g., cytochrome
P450monooxygenases) and Phase II (e.g., glutathione S-transferases, sulfotrans-ferases)
biotransformation enzymes (Hankinson, 1995). TCDD induces the transcription of Cyp1A1,
Cyp1B1 and NQO1, which are all significantly up-regulated and this may lead to the generation

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 Introduction

of reactive oxygen species (ROS; includes superoxide anion and hydrogen peroxide) and
thereby oxidative stress/DNA damage (Knerr et al., 2006). Polymorphism in the Ah locus,
which has been shown to be the structural gene for the cytosolic receptor, seems to determine
the sensitivity of genetically different strains of mice to TCDD and congeners.
TCDD toxicity involves many different types of symptoms, which vary from species to
species and from tissue to tissue, both quantitatively and qualitatively. Age and sex-related
differences in sensitivity have also been reported.

Fig. 3: A schematic model of the action of dioxin. Binding of the ligand (dioxin) to the aryl
hydrocarbon receptor (AhR) results in the release of Hsp90 and translocation to the nucleus
followed by dimerization to the aryl hydrocarbon receptor nuclear translocator (ARNT). The
AhR-ARNT heterodimer binds with specific cis-acting enhancer, called dioxin responsive
element (DRE), that promote the activation of several genes. Dioxin can also exert its effects in
the cytosol through AhR-associated protein kinase, c-Src, to alter the function of many proteins
through a cascade of protein phospholylations ( Overmeire et al., 2001).

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 Introduction

Health effects:
Effects on other animals:
In the Great Lakes area, which has been extensively polluted with dioxin and dioxin-like
compounds, multiple species of birds, fish, and reptiles have exhibited developmental toxicity,
reproductive impairment, compromised immunologic function, and other adverse effects
correlated with these exposures. Specific observations correlated with dioxin or dioxin-like
compound levels in multiple vertebrate species included hyperplasia of the thyroid and adrenal
glands, porphyria, suppressed T-cell-mediated immunity, mammary and ovarian pathologies,
reduced viability of offspring, congenital malformations, growth retardation, and an edematous
syndrome among the offspring of fish-eating birds comparable to chick edema disease (Fox,
2001).
In laboratory animals, TCDD brings about a wide spectrum of toxic or adaptive
biochemical and morphological changes, including acute lethality associated with a wasting
syndrome; tissue-dependent induction or inhibition of a variety of enzyme activities, endocrine
perturbations; immune suppression coupled with thymic atrophy; lipid peroxidation; alterations
in growth factors and cytokines and in their receptors; disturbed synthesis and degradation of
heme; dysregulation of glucose and lipid metabolism, liver damage, neurobehavioral disorders,
to reproductive toxicity, terato- and carcinogenicity (Pohjanvirta and Tuomisto, 1994).

Effects on mammals:
Dioxins have proven to be developmentally toxic, immunotoxic, neurotoxic, and
hepatotoxic to mammals. Adverse health effects of dioxin exposure in humans may include
cardiovascular disease, diabetes, cancer, porphyria, endometriosis, early menopause, reduced
testosterone and thyroid hormones, altered immunologic response, skin, tooth, and nail
abnormalities, altered growth factor signalling, and altered metabolism (US EPA, 2003). TCDD
at high doses (lethal or near lethal) causes a starvation-like effect or wasting syndrome (El-
Sabeawya et al., 2001).TCDD also affects the cholinergic receptors causing long-term
behavioural dysfunctions and significant impairment in discrimination-reversal learning
activities (Rice, 1997).
TCDD is also typical endocrine disruptors toxicity mimics endocrine imbalance,
interfering with the estrus cycle in combination with some steroid-like activities of TCDD,
impair thyroid functions (Van Birgelen et al., 1994). TCDD also causes impairment of

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 Introduction

reproduction, prenatal mortality, significant decreased fertility, litter size and number of pups
alive at birth (Li et al., 2006).
TCDD acts as a complete carcinogen in several species and induces neoplasms in the
lung, oral/nasal cavities, thyroid and adrenal glands, and liver (Huff et al., 1991). TCDD also
adversely affects the multilineage hematopoiesis in the hematopoietic stem cells of the bone
marrow, mediated through AhR in the target tissue.
TCDD and Hepatotoxicity:
The highest amounts of TCDD are found in the liver and adipose tissue. The liver has
been also identified as the primary and most important tissue target for TCDD toxicity and the
prominent hepatotoxic effect is progressive centrilobular necrosis (Karami et al., 2001). TCDD
leads to hypertrophy of hepatocytes, glycogen depletion and lipidosis of the liver (Zodrowa et
al., 2004). TCDD elicits hepatomegaly and liver pathologies that include hepatocellular
neoplasms, inflammation, necrosis, steatosis, and the differential expression of lipid metabolism
and transport genes in mice ((Zodrowa et al., 2004; Kopec et al., 2008, 2010a, 2010b).
TCDD treatment in animals resulted in a significant decline in the activities of
superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase with
concomitant increased levels of hydrogen peroxide and lipid peroxidation.
Medicinal plants:
Since time immemorial, mankind has made the use of plants in the treatment of various
ailments. Phytochemicals, which modulate the host's defense, are found in several leaves, roots,
fruits and vegetables.
Hippokrates (460-370 B.C.) was also well aware of the pharmacological effects of food
plants, as evident from his statement, “Let your ailments be your medicament and your
medicament be your food”.
The Indian Traditional Medicine like Ayurveda, Siddha and Unani are predominantly
based on the use of plant materials. These practices incorporated ancient beliefs and were passed
on from one generation to another by oral tradition and/or guarded literature.
Herbal drugs have gained importance and popularity in recent years because of their
safety, efficacy and cost effectiveness. Today, it is estimated that about 80% of the world
population relies on botanical preparations as medicine to meet their health needs (Ogbera et al.,
2010). A large number of medicinal plants have been tested and found to contain active
principles with curative properties against a variety of diseases. Plants contain a variety of
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 Introduction

chemical constituents like phenols, coumarins, lignans, essential oil, carotinoids, glycosides,
flavanoids, organic acids, lipids, and alkaloids which showed hepatoprotective activity. A
number of traditional remedies from plant origin are evaluated for their possible antioxidant and
hepatoprotective effects against different chemicals (e.g., CCl4, paracetamol, alcohol,
antitubercular drug, nitrobenzene, bromobenzene, lead etc.) induced liver damage in
experimental animal’s. Most of the hepatotoxic chemicals damage liver cells mainly by
inducing lipid peroxidation and other oxidative damages. Natural antioxidants from plants are
reported to provide substantial protection that slows down the process of oxidative damage
caused by reactive oxygen species (ROs) (Hamid et al., 2010). Therefore, a large number of
plants and formulations have been claimed to have hepatoprotective activity. Nearly 160
phytoconstituents from 101 plants have been claimed to possess liver protecting activity. In
India, more than 87 plants are used in proprietary multi ingredient plant formulations (Vishnu et
al., 2010).
India is richly endowed with diverse flora with good potential to be developed
into various useful natural products. Our country is known to have a large amount of plant
species and a number of them have been traditionally used to treat many diseases and
ailments.
Among the promising medicinal antioxidative plants, we have focused on Curcuma
longa and Eclipta alba, which are amazing herbs with rich historical and medicinal background.

Curcuma longa (L.)

Curcuma longa L. (turmeric), “the golden spice of life” is one of the oldest and most
important and valuable herb spice to humankind. Its use dates back nearly 4000 years, to the
Vedic culture in India where it was used as a culinary spice and had some religious significance
(Bhowmik et al., 2009). Curcuma is also known as Indian saffron, Haridra (Sanskrit,
Ayurvedic), Jianghuang (yellow ginger in Chinese) and Kyoo (Japanese). It is grown and
harvested commercially in India, China, and many regions of tropical south Asia and needs
temperature between 20°C and 30°C and a considerable amount of annual rainfall to thrive. It
has oblong, pointed leaves and funnel-shaped yellow flowers. The rhizomes are specialized
underground stems that are root-like in structure, have a brown surface and bright orange or
yellow interior flesh and after cutting, it is in curved cylindrical or oblong tubers 2 or 3 inches in
length, and an inch in diameter, pointed or tapering at one end, yellowish externally, with

8
 Introduction

transverse, parallel rings internally deep orange or reddish brown, marked with shining points,
dense, solid, short, granular fracture, forming a lemon yellow powder (Kumar and Sakhya,
2013).
Turmeric was described as C. longa Linn. by Linnaeus and its taxonomic position. It is a
perennial herb, which belongs to Kingdom- Plantae, Class-Liliopsida, Family- Zingiberaceae,
Genus - Curcuma, Species - longa, Linn.
Chemical compositions:
The major constituent of turmeric is curcumin (diferuloylmethane) which constitutes up
to 90% of total curcuminoid content, with demethoxycurcumin and bis-demethoxycurcumin
comprising the remainder (Aggarwal and Shishodia, 2004). The active constituents of turmeric
are the flavonoid curcumin (diferuloylmethane) and various volatile oils, including tumerone,
atlantone, zingiberone, sugars, proteins, resins, proteins, vitamins, and minerals (including iron
and potassium) (Akarm et al., 2010).

Fig. 4: The chemical structures of the major constituents obtained from Curcuma longa Dohare et al., 2008).

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 Introduction

Ethno medicinal uses of C. longa:

Turmeric (Curcuma longa rhizomes), commonly used as a spice, is well documented for
its medicinal properties in Indian and Chinese systems of medicine. In current traditional Indian
medicine, it is used for the treatment of biliary disorders, anorexia, cough, diabetic, hepatic
disorders, rheumatism and sinusitis (Jain and DeFillips, 1991). It has been also used topically
on the skin for wounds, blistering diseases, for parasitic skin infections, and for acne. It
has been used via oral administration for the common cold, liver diseases, urinary tract
diseases, and as a blood purifier (Kumar and Sakhya, 2013). For the last few decades,
widespread work has been done to establish the biological activities and pharmological actions
of C. longa rhizome and its extract, such as anti-inflammatory, antioxidant, anticarcinogenic,
anticoagulant, antidiabetic, antiamoebic, antifungal, antiviral and anti HIV activities (Mortellini
et al., 2000). The active constituents of turmeric are yellowish orange volatile oils called
curcuminoids known as curcumin, which has demonstrated antioxidant, antineoplastic, antiviral,
anti-inflammatory, antibacterial, antifungal, antidiabetic, anticoagulant, antifertility,
cardiovascular protective, hepatoprotective, and immunostimulant activity in animals (Shishodia
et al., 2005). There are over 1,500 citations in Medline relating to the biologic effect of
curcumin.

Antioxidative Hepatoprotective Anticarcinogenic

Effects
Antimicrobial
Antidiabetic effect:

Nephroprotective
Wound healing Curcuma longa
Muscle Gastrointestinal
regeneration Effects

Anti-inflammatory Antihyperlipedemic Cardiovascular Effects

Fig.5: Health effects of C. longa

Eclipta alba (L.)

Eclipta alba (L.) (syn. Eclipta prostrate L., Eclipta erecta, Verbesina alba), is an annual
herbaceous plant, commonly known as False Daisy, yerba de tago, and bhringraj. In India, the

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 Introduction

plant is known as bhangra, "bhringaraj" or bhringraja. It is an erect or prostrate, much branched,

roughly hairy, annual, rooting at the nodes; the leaves are opposite, sessile and lanceolate. Floral
heads 6-8 mm in diameter, solitary, white, achene compressed and narrowly winged. It grows
commonly in moist places as a weed all over the world. It is widely distributed throughout
India, China, Thailand, and Brazil.
It belongs to Kingdom - Plantae, Class - Magnoliopsida, Family - Asteraceae, Genus -
Eclipta, Species - alba, Linn.
Chemical compositions:
The herb E. alba contains mainly coumestans i.e., wedelolactone (I) and demethyl
wedelolactone (II), polypeptides, polyacetylenes, thiophene-derivatives, steroids, triterpenes,
flavonoids and nicotine (Thorat et al., 2010). The plant also contains other chemicals such as
wedelic acid, apigenin, luteolin, b-amyrin, oleanane & taraxastaneglycosides etc. (Anonymous,
1998). The polypeptides isolated from the plant yield cystine, glutamic acid, phenyl alanine,
tyrosine and methionine on hydrolysis.

Fig.6: The chemical structures of the major constituents obtained from E. alba (Prakash et al., 2011)
Ethnomedicinal uses of E. alba:
In Ayurveda, the plant is considered a rasayana for longevity and rejuvenation. Charaka
advises taking the juice of E. alba with honey to prevent the onset of senility. It is an active
ingredient of many herbal formulations prescribed for liver ailments and shows effect on liver
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 Introduction

cell generation. It is widely used in India as a chologuague and deobstruent in hepatic

enlargement, for jaundice and other ailments of the liver and gall bladder (Treadway, 1998). E.
alba is reported to possess antiseptic, analgesic, antipyretic, antispasmodic, antimicrobial and
antiviral properties (Bakht et al., 2011). It is also useful in inflammations, hernia, eye diseases,
bronchitis, asthma, leucoderma, anemia, heart and skin diseases, night blindness, syphilis etc.
The alcoholic extract of this plant has shown antiviral activity (Dalal et al., 2010). The
fresh juice of leaf is used for increasing appetite, improving digestion and as a mild bowel
regulator (Lans, 2007). The plant has a reputation as an anti-ageing agent in Ayurveda. E. alba
also has traditional external uses, like athlete foot, eczema and dermatitis, on the scalp to
address hair loss, inflammation, minor cuts and burns (Singh et al., 2010).

Antihepatotoxic Antioxidative Immunomodulatory

Antimicrobial
Antiaggressive
effect:

Antidiabetic
Eclipta alba Neuropharmacologic
al
Analgesic
Anticancer

Anti-inflammatory Hair growth & Alopecia

Antihyperlipedemic

Fig.7: Health effects of E. alba

Assessment of Toxicity:
Toxicity is the degree to which a substance is able to damage an exposed organism. The
study of the adverse effects of chemicals on living organisms; the study of symptoms,
mechanisms and treatments is called toxicology. Toxicity can refer to the effect on a whole
organism, such as an animal, bacterium or plant, as well as the effect on a substructure of the
organism, such as a cell (cytotoxicity) or liver (hepatotoxicity).
Toxicity may be acute, sub-acute/sub-chronic or chronic.

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 Introduction

Acute Toxicity:
Acute toxicity is the toxicity that manifests itself immediately or within 14 days
following exposure of animal or humans to single doses of chemicals (xenobiotics) by oral,
dermal, inhalation or through the skin. In acute toxicity, there is a single exposure to a toxicant
which may result in severe biological harm or death; acute exposures are usually characterized
as lasting no longer than twenty four (24) hours.
The LD50 value is currently the starting point for toxicological classification of
chemicals. Laboratory mice and rats are the species typically selected for a classic LD50 study
and widely used for the assessments of the degree of potential harm to the environment caused
by industrial processes (Barlow et al., 2002).

Sub-Acute toxicity:
These tests express the overall side effects observed after repeated administration of the
test substance (2-6 weeks) and enable to determination of the principal behavioral changes as
well as anatomical, physiochemical and biochemical manifestations of the tissue damages
provoked by the substances (WHO, 1993).
Chronic Toxicity:
Chronic exposure to a toxin is over an extended period of time, often measured in
months or years, and can cause irreversible side effects. The aim of this test is to determine the
concentration of a test material/substance that produces adverse effects on a group of test
organism during long term exposure under controlled conditions. Unlike acute toxicity study,
chronic toxicity studies not only mortality, but investigate endpoints such as individual
growth/growth rate, developments, hatching time and success, reproduction, behaviour,
physiological parameters and histology (Barlow et al., 2002).

Target organ toxicity:

The extent to which an organ is susceptible to toxicity varies from organ to organ. For
example the liver is more highly vascularised making them more susceptible to toxicity than the
bone tissues.
Blood Toxicity:
Blood which forms the main medium of transport in the body is a very important tissue.
It serves to transport many drugs and xenobiotic. Since all foreign compounds are distributed

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 Introduction

via the bloodstream, the various components, cellular and non-cellular, are initially exposed to
significant concentrations of toxic compounds (Timbrel, 2009).
The assessment of blood is relevant to the evaluation of risks since the haematological
system carries a higher predictive value for toxicity in humans (Olson et al., 2000).
Haematological characteristics have also been widely used in the diagnosis of variety of
diseases and pathologies induced by industrial compounds, drugs, dyes, heavy metals, pesticides
and several others (Mansour & Mossa, 2005).

Liver Toxicity:
Liver is the vital organ of metabolism and excretion thus it is continuously and widely
exposed to toxins and chemotherapeutic agents that lead to impairment of its functions. All
types of insults/injuries to the liver (e.g. circulatory, traumatic, toxic or microbiological) lead to
damage of hepatocytes which results in its mal-functioning and cause hepatic disease. Hepatic
or liver disease is a term that affects the cells, tissues, structures, or functions of the liver. About
20,000 deaths are found every year due to hepatic diseases (Sharma and Sharma, 2009).
It is also a unique organ in the sense that a significant loss of liver cells due to drug
toxicity or other insults can be overcome by regeneration (Mehendale, 2005). Though, it
possesses a large capacity for regeneration, slight damage to the liver by toxins or infective
agents can hamper its functioning. Thus, the normal functioning of liver can be disturbed by
various infections, infiltrations or toxic agents, drugs and environmental factors. Liver injuries
induced by various hepatotoxins have been recognized as a major toxicological problem for
years (Azer et al., 1997). Chemical induced liver injury is known as toxic hepatopathy and
is one of the most common and important environmental health and clinical problem
worldwide.
Histopathology of liver:
Hepatocytes, with their high degree of metabolic activities, are readily disturbed by
toxins and may demonstrate the histological cell responses known as cloudy swelling, fatty
changes, and necrosis. The fatty changes due to metabolic injury to hepatocytes are manifested
by large cytoplasmic vacuoles within some hepatocytes, which usually displace the nucleus to
one side. The dead cells stain a bright pink (eosinophilia) and standout from the other cells. This
is due to degeneration of structural proteins which form a compact homogenous mass, whereas

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living cell’s nucleus is smaller, condensed, and intensely stained with hematoxylin (basophilia)
(Burkitt et al., 1996).
Acute inflammation of the liver parenchyma is usually marked by focal accumulations
of inflammatory cells in the site of necrotic hepatocytes (Ishak, 2000). Liver sections may
appear portal tracts enlargement with inflammatory cellular infiltration mainly plasma cells,
vacuolated hepatocytes, dilated sinusoids, and increased number of Kupffer cells in
histopathological study (Ebaid et al., 2007). Cirrhosis is the end stage of many types of liver
damage. It is characterized by destruction of normal liver architecture, which is replaced by
regenerative nodules of hepatocytes separated by bands of fibrous tissue (Hubscher, 2006).
In chronic hepatitis, the portal triads are infiltrated and usually expanded by
lymphocytes. In some cases, the metabolic disruption may be irrecoverable and some
hepatocytes undergo necrosis (Burkitt et al., 1996).
Biomarkers of liver cell injury:
Assessment of biochemical parameters is an important tool in assessing the health status
of the individuals and is valuable in predicting clinical and prognostic outcomes (Krishna and
Ramachandran, 2009). The hepatoxicity is associated with alteration in the levels and activities
of certain biochemical parameters such as ALT, AST, ALP, total protein, albumin, bilirubin etc.
The noticed increase in the levels of aminotransferase (ALT and AST) and the level of ALP as
well as the decrease in the levels of total protein and albumin in the serum, are the major
diagnostic symptoms of liver diseases (Chatterjee and Shinde, 2005).
Thus, the present work entitled “Biochemical and Histopathological assessment of
Effects of Dioxin on liver of mice and Hepatoprotective action of Curcuma longa & Eclipta
alba” has been undertaken to investigate TCDD (Tetrachloro-dibenzo-p-dioxin) induced effects
on the physical parameters, biochemical profile and histology of liver of Swiss albino mice and
to evaluate the hepatoprotective effects of aqueous extracts of rhizome of C. longa and leaf of E.
alba.

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 Introduction

Objectives

 To study the morphological features and phytochemical analysis of the herbal plants C.
longa (rhizome) and E. alba (leaf).
 To find the LD50 and therapeutic dose of aqueous rhizome extract of C. longa and
aqueous leaf extract of E. alba in Swiss albino mice.
 To assess the physical, biochemical (LFTs), haematological and histological changes in
liver tissues at acute, sub- chronic and chronic doses of the aqueous rhizome extract of
C. longa and aqueous leaf extract of E. alba in Swiss albino mice.
 To find out the LD50 of TCDD (Tetrachloro-dibenzo-p-dioxin) in Swiss albino mice.
 To observe the acute, sub-chronic and chronic toxicity of TCDD on the physical,
biochemical (LFTs) and histological parameters of the liver in Swiss albino mice.
 To evaluate the protective effects of aqueous rhizome extract of C. longa and aqueous
leaf extract of E. alba in TCDD intoxicated mice on the basis of body weight, liver
weight, biochemical parameters, haematological parameters and histopathological
changes in the liver tissue of mice.
 To study and compare the efficacy of aqueous rhizome extract of C. longa and aqueous
leaf extract of E. alba against TCDD induced haepatotoxicity.

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