INTRAOCULAR PRESSURE MARKERS IN MALARIAL INFECTED MICE RECEIVING PHYLLANTHUS  AMARUS  TREATMENT

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ABSTRACT

Phyllanthus amarus has been consistently reported as a rich herb having medicinal value and ethnomedical importance. It has been used to eliminate gallstone, malaria and some other aliments, but the effect of its use on changes in ocular glucose, proteins and lipids — analytes that influence intraocular pressure, has not been fully documented. So, in this study, the effect of ethanolic leaf extract of Phyllanthus amarus on the levels of ocular glucose, proteins and lipids was investigated using homogenized ocular tissue of experimental mice. Forty five (45) adult mice weighing between 22-27g were randomly divided into 9 groups  and used for the study. Group 1: normal control (uninfected and untreated mice) Group 2: malaria control (mice infected with plasmodium berghei and untreated) Group 3: parasitized (infected with P. Berghei treated with 100mg/kg P. amarus) Group 4: parasitized (infected with P. Berghei treated with 200mg/kg P. amarus) Group 5: parasitized (infected with P. Berghei treated with 300mg/kg P. amarus)

Group 6: parasitized (infected with P. Berghei treated with 5mg/kg chloroquine)

Group 7: uninfected but treated with 100mg/kg P .amarus Group 8: uninfected but treated with 200mg/kg P. amarus Group 9: uninfected but treated with 300mg/kg P. amarus .Each group was treated for

7days and on the 8th day the animals were scarificed under chloroform anaesthesia after an overnight fast. The mice eyes were carefully excised, rinsed in cold normal saline and prepared for the biochemical analysis of glucose, proteins and lipids using standard methods. Results show that Phyllanthus amarus administration (irrespective of dose) did not significantly (p>0.05) alter ocular glucose and protein levels, but increased (p<0.05) lipids (cholesterol and triglycerides) concentrations when compared with control values. The altered ocular lipid homeostasis may have some biochemical implications and clinical significance. This should be validated in further studies.

 

LIST OF TABLES

Table 1: glucose in the serum of p. berghei infected and uninfected mice treated with phyllanthus amarus leaf extract.    ———————————————————        25

Table 2: changes in blood cholesterol and triglyceride induced by plasmodium berghei infected mice    ——————————————————————————–       26

Table 3:  changes in blood albumin and total protein induced by p. berghei infected mice    —————————————————————————————————        27

Cover         ——————————————————————–      i

Title            ——————————————————————–    ii

Certification   —————————————————————-     iii

Dedication      ————————————————————–       iv

Acknowledgement   ——————————————————         v

Abstract         —————————————————————       vi

List of Tables   ————————————————————-     vii

Table of contents   ——————————————————-       viii

CHAPTER ONE

• INTRODUCTION ———————————————————————–      1

1.1.1 BACKGROUND OF STUDY    ——————————————————-      1

1.2 STATEMENT OF PROBLEM    ——————————————————-        3

1.3 OBJECTIVE OF STUDY    ————————————————————-        4

1.4 SIGNIFICANCE OF STUDY    ——————————————————–        4

1.5 HYPOTHESIS    ————————————————————————–     5

1.6 BRIEF REVIEW OF RELEVANT LITERATURE    ———————————     5

1.7 BIOMAKERS   —————————————————————————-    5

1.8 RETINOPATHY    ————————————————————————   6

1.9 MUSCULAR ODEMA   —————————————————————–    6

1.10 WHAT IS MALARIA?    ———————————————————    7

1.11 CAUSES OF MALARIA       ———————————————————–     8

1.12 EPIDEMIOLOGY OF MALARIA         ———————————————-     8

1.13 GLOBAL AND GEOGRAPHICAL DISTRIBUTION OF MALARIA..   ——        9

1.14TRANSMISSION AND LIFE CYCLE OF PLASMODIUM PARASITE   —–      11

1.15 LIFE CIRCLE OF PLASMODIUM PARASITE   ———————————–    12

CHAPTER TWO   ——————————————————————————    14

2. 0MATERIALS AND METHOD ——————————————————– 14

2.1 MATERIALS    —————————————————————————–    14

2.2 METHODS    ——————————————————————————–   14

2.2.1 ANIMALS CARE AND HANDLING    ———————————————    14

2.2.2 ANIMAL GROUPING AND INOCULATION WITH PLASMODIUM BERGHEI    —————————————————————————————————–     15

2.2.3 ANIMAL SACRIFICE AND COLLECTION OF SPECIMEN    —————-     16

2.2.4 ANALYSIS OF SPECIMEN   ———————————————————-   16

 

2.3 GLUCOSE ESTIMATION    ———————————————————–    16

2.3.1 REACTION PRINCIPLE    ———————————————————-     16

2.3.2 PROCEDURE    ———————————————————————–     17

 

2.4 TOTAL CHOLESTEROL ESTIMATION    ——————————————   17

2.4.1 PRINCIPLE    ————————————————————————–     17

2.4.2 EQUATION    ————————————————————————–    17

2.4.3 PROCEDURE    ————————————————————————    18

2.4.4 CALCULATION    ———————————————————————   19

 

2.5. TRIGLYCERIDE ESTIMATION    —————————————————    19

2.5.1 PRINCIPLE    ————————————————————————–     19

2.5.2 EQUATION    —————————————————————————   20

2.5.3 PROCEDURE    ———————————————————————–     20

2.5.4 CALCULATION    ——————————————————————–    21

 

2.6 TOTAL PROTEIN ———————————————————————-     21

2.6.1 PRINCIPLE    ————————————————————————–     21

2.6.2 PROCEDURE    ———————————————————————–     21

2.6.3 CALCULATION    ——————————————————————–     22

 

2.7 ALBUMIN ———————————————————————————–22

2.7.1 PRINCIPLE    ————————————————————————–      22

2.7.2 PROCEDURE    ———————————————————————–      23

2.7.3 CALCULATION    ——————————————————————–     23

2.7.4 STATISTICAL ANALYSIS ————————————————————– 24

CHAPTER THREE    —————————————————————————   25

3.0 RESULTS    ———————————————————————————-   25

CHAPTER FOUR    —————————————————————————–   28

4.0 DICUSSION    ——————————————————————————    28

4.1 CONCLUSION    —————————————————————————– 29

REFERENCES   —————————————————————————30

 

CHAPTER ONE

1.1     INTRODUCTION

1.1.1. BACKGROUND OF STUDY

Nature can be considered as the ultimate chemist, about 80% of the world inhabitants still depend on natural products that have inspired chemists and physicians for years because of their rich structural diversity and complexity considerable advances have been obtained for the understanding of natural product biosynthesis in the recent decades.

Malaria, a mosquito borne infectious disease is endemic in the tropical and sub tropical regions of the world (Ahimanah et al., 2000). It is a deadly disease which lowers life expectancy and a major cause of infant mortality in highly endemic areas (WHO, 2011).

Malaria infection in humans and animals is caused by the Plasmodium. Several species of Plasmodium have the ability to cause malaria in animals, including rodents (mice). These parasites are not direct practical concern to man or his domestic animals. The interest of these parasites is that they are practical model organism in the laboratory for the experimental study of human malaria.

Plasmodium berghei is considered a comparable genetic model to human There is a high degree of genetic conservation this up to 99% (pennachio, 2003) and it is well established that mice also exhibit natural differences in susceptibility to malaria infection (Greenberg et al., 1954). P. berghei is transmitted by Anopheles mosquito and it infects the liver after being injected into the blood stream by the bite of the infected female mosquito. After a few days of development and multiplication, these parasites leave the liver and invade erythrocyte (red blood cells). The multiplication causes Anemia and damage essential organs in the body. P. berghei infection also affects the brain and can cause cerebral complications in laboratory mice.

The plant, phyllanthus schumach (Euphorbiaceac) is commonly known as bhuirali Usually occurs in Asain, Maharashitra, Burma, Nicobar, Islands malesia and America. Phyllanthus amarus schumach is a native to Americans (van Holthoon, 1999). Phyllanthus amarus is small, erect, annual herb that grows 30-40cm in height which is indigenous to the rain forests of the Amazon and other tropical areas throughout the world including America, India and Nigeria. The Spanish name of the plant, chanca piedra, means stone breaker, wind breaker, gulf leaf flower or gala of wind (Celia, A.  et al., 2006).

Phyllanthus amarus is an Ayurvedic system of medicine which is used in the problems of stomach, genitourinary system, liver, kidney, and spleen, it plays an important role in Ayurvedia, an Indian system of medicine and it is used to treat jaundice, gastropathy, diarrhea, dysentery, fevers, scabies, gential, infections, ulcers, and wounds (Patel et al., 2011).

The different plant parts are ethnobotanically used in various diseases and disorders. For example, the leaves are used as expectorant and diaphoretic and the fruits as carminative, laxative, astringent. Diuretic and tonic to the liver. The juice or extract of its thinner roots and young leaves are taken internally to stimulate the kidney. Heyne recorded its uses in the Dutch  Indies (Indonesia) for stomach, aches, gonorrhoea and children cough (Karuna,  et al., 2009).

Research have shown that the plant has demonstrated anti-viral property against hepatitis B virus (Boeira  et al., 2 011), hepatoprotective (Amin,  et al., 2013), anticarcinogenic (Rajeshkumar,  et al., 2002), antimutagenics, anti-nociceptive and anti-inflammatory (Obidike,  et al., 2010), anti-diabetics (Okoli, et al., 2011) and antilipidermic (Khanna,  et al.,  2002) activities.

1.2 STATEMENT OF PROBLEM

Since malaria has been speculated to alter biochemical functions of organs of the body such as brain, liver, heart, and spleen, this research is designed to know if such biochemical alternations also include changes in the intraocular pressure markers in experimental mice. Phyllanthus amarus is a medicinal herb used for the treatment of several diseases including malarial infection. In southern Nigeria the utilization of Phyllanthus amarus is popular but whether it improves or disturbs