SEX VARIATION ON THE EFFECTS OF CARICA PAPAYA SEED EXTRACT ON RED BLOOD CELL, PACKED CELL VOLUME AND HAEMOGLOBIN IN MALE AND FEMALE WISTAR RATS
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SEX
VARIATION ON THE EFFECTS OF CARICA PAPAYA SEED EXTRACT ON RED BLOOD CELL,
PACKED CELL VOLUME AND HAEMOGLOBIN IN MALE AND FEMALE WISTAR RATS
TABLE OF
CONTENT
Title
page----------i
Declaration----------ii
Certification----------iii
Dedication----------iv
Acknowledgement---------v
Table of
Contents---------vi
Lists of
Tables --------- vii
Lists of
graphs --------viii
Abstract----------ix
CHAPTER ONE
INTRODUCTION--------1
CHAPTER TWO
MATERIALS
AND METHODS-------18
CHAPTER
THREE
RESULTS----------21
CHAPTER FOUR
DISCUSSION--------27
CONCLUSION---------30
REFERENCES----------31
vi
LISTS OF
TABLES
Table 1:
Haematological parameter in male and female wistar rats………………21
LISTS OF
GRAPHS
Fig 1Effect
of Carica Papaya on RBC--------23
fig
2:----------24
fig
3:----------25
ABSTRACT
The effects
of oral administration of different doses ofCaricapapayaseed extracts on
haematological parameters specifically red blood cells (RBC), packed cell
volume (PCV) and haemoglobin concentration in male and female wistar rats was
examined. Previous studies have shown that Carica papaya has antisickling
effects (Sofoworaet al., 1975), antifertility effects (Chinoyet al., 1997),
membrane stabilization effects (Priyangaet al., 2012), nephroprotective effects
(Olagunjuet al., 2009) and these effects tend to vary with sex. Other possible
variations on the effects of Carica papaya seed extract due to sex is being
investigated in this study.
Blood
samples were taken by cardiac puncture from twenty (20) male and twenty (20)
female wistar rats which were divided into four groups (n=5) each and graded
dosages of Carica papaya seed extracts administered for twenty-one days (three
weeks). The red blood cell mass, haemoglobin concentration and packed cell
volume of the rats was determined by routine methods.
Two-way
analysis of variance followed by Student’s T-test were performed on data and
differences were considered statistically significant for p<0.05. There was
an increase in parameters studied with response being higher in males than in
females. Significant difference wereobserved between males and females only in
red blood cell count though there were slight differences in the other parameters.
This study
showed that Carica papaya seed extract has haemato-stimulatory effects on both
males and females with higher response in males which is via another mechanism
apart from its effects on sex hormones.
CHAPTER ONE
INTRODUCTION
BACKGROUND
STUDY
Blood is a
special type of connective tissue composed of formed elements in a fluid
matrix. The formed elements include red blood cells (erythrocytes), white blood
cells (leukocytes) and platelets (Bacha and Bacha, 2000). It is a bodily fluid
in animals that delivers necessary substances such as nutrients and oxygen to
the cells and transports metabolic waste products away from those same cells.
Assessment
of haematological parameters can be quite informative about the health status
of the individual and it is among the methods which may not be apparent during
physical examination but may affect the fitness of the animals (Kronfeld et
al., 1969). The red blood cell count, white blood cell counts, packed cell
volume and haemoglobin concentration are regularly measured haematological
parameters but this study is concerned with red blood cells, packed cell volume
and haemoglobin.
Most of the
haematological parameters are influenced by factors like sex, age, breed,
seasonal variations, lactation, health and nutrition status (Aengwanich, 2002;
Al-shami, 2007; Mohammed et al., 2007). Since it is a well-documented fact that
there are variations due to age, sex, attitude etc. (Hawkins et al., 1954,
Viteri et al.,1972, De Grunch et al.,1989), it is acknowledged that for
comparisons between individuals and with reference data in a clinical
diagnostic situation, it is necessary to consider normal variations due to sex,
age and breed in order to increase diagnostic precision (Satue et al., 2009).
It has however been observed that the modifications correlated with sex, age
and taxonomic position are best shown by the erythrocytic counts (Duguy 1970).
SEX
VARIATION IN RED BLOOD CELLS, PACKED CELL VOLUME AND HAEMOGLOBIN CONCENTRATION
Red blood
cells are the most common type of blood cells and the vertebrates’ organism’s
principal means of delivering oxygen to the body tissues via blood flow through
circulatory system (Pierige et al., 2008). They are also called erythrocytes.
They are biconcave and anucleated and there are about 5.2 million red cells per
cubic millimeter of blood in the adult human (4.7 to 6.1 million in males and
4.2 to 5.4 million for females). They take up oxygen and release it while
squeezing through the body’s capillaries. They are formed in the bone marrow
and the average life cycle of a red blood cell is 120 days. Erythropoietin and
oxygen levels are factors that regulate the production of red blood cells.
Haemoglobin
is an iron-containing biomolecule found in red blood cell cytoplasm for
transporting oxygen and carbon (iv) oxide. Iron can bind to oxygen and heme is
responsible for the blood’s red colour. It also carries some of the waste
products back and from the tissue. It is concerned with blood’s ability to
carry oxygen and iron. Low iron could lead to morbidity and mortality. On the
average, haemoglobin concentration for males is about 14 – 18g/dl and 12
-16g/dl.
The
haematocrit is also known as packed cell volume or erythrocyte volume fraction.
It is the volume percentage (%) of red blood cells in blood.it was coined in
the year 1903. It comes from the Greek words ‘hema’ meaning blood and ‘krites’
meaning gage or judge. It is normally higher in males than in females and that
is about 45% in men and 40% for women (Purveset al., 2004). It can be determined
by centrifuging heparinized blood in a capillary tube at 10,000 RPM for five
(5) minutes. The volume of packed red blood cells divided by the total volume
of blood sample gives packed cell volume. It can also be calculated from
automated analyzer.
Mean red
cell mass and hematocrit levels are higher in men than women but the reason for
this is yet to be discovered. It was reported that testosterone and other
androgens have an erythropoietic stimulating effect that can cause
polycythemia, which manifests as an increase in hemoglobin, hematocrit, or red
blood cell count (Siddique et al., 2004, Dobs et al., 1999).Some studies
however do not agree with this theory. It was reported that the increase of
hemoglobin depending on testosterone is not along with the increase of
erythropoietin. Hence another mechanism was proposed which stated that
androgens increased the response of immature bone marrow cells to the effects
of erythropoietin which in turn stimulates the production and regulation of red
blood cells. However, other studies have shown that greater increase of
hematocrit and hemoglobin observed in older males during testosteronetherapy is
due to changes in erythropoietin rates (Alexanian, 1969). There is an important
difference between male and female genders in testosterone and hematocrit rates
but there is no important difference in their erythropoietin rates (Molinari,
1982). Then, androgens stimulate haematogenesis by a passage other than
erythropoietin. Also the studies on patients with renal disease demonstrated
that erythropoietin does not react to androgentherapy (Dobs et al., 1999). In
contrast, studies have shown that estrogen can inhibit the production of red
blood cells (Watanuki et al.,2002). Hence it can be seen that the sex of an individual
is a determining factor on the haematological factors.
CARICA
PAPAYA
In the long
history of the world, plants have been used medicinally. A large and increasing
number of patients use medicinal herbs or seek the advice of their physician
regarding their use (‘O’ Hara et al., 1998). It has been estimated roughly,
that presently more than half of the total population of the world use herbal
drugs (Chang, 1987). Increasing interest in medicinal herbs has increased
scientific scrutiny of their therapeutic potentials and safety thereby
providing physicians with data to help patients make wise decisions about their
use (‘O’ Hara et al., 1998).
The papaya
/pa’paia/ (Spanish name) also known as papaw or pawpaw is a fruit of the plant
Carica papaya, the sole species in the genus carica of the plant family
Caricaceae. The name papaya is widely recognized but it has been corrupted to
kapaya, lapaya or tapaya in Southern Asia and the East indies. In French, it is
papaya (the fruit) and papayer (the plant) or sometimes figuier des iles.
Spanish speaking people employ the names melon zapote, payaya (fruit) and
papayero (plant). Name in Brazil is mamao. It is known as pawpaw or papaw in
both Africa and United Kingdom (Delbridgeet al., 1988, Watson 1997). It was
nicknamed tree melon in Europe. It is a small, frost-tender, succulent broadleaf
evergreen tree which is native to South America. It is best grown in topical or
semi-tropical climates like Malaysia, the West Indies and Africa (Soforowa,
1996) and it is planted in well drained loams in full sun. It is properly
called a giant herb because it never produces true woody tissue. Papaya is not
a tree but an herbaceous succulent plant that possess self-supporting stems
(Gross, 2003). Its fruits are large, fleshy and melon like. It is an erect fast
growing tree measuring 7 – 8m tall, with copious latex and trunk of about 20cm
in diameter (Duke, 1984). Its leaves are spirally arranged and confined to the
top of the trunk.
The papaya
has a rather complicated means of reproduction. The plants are male,
hermaphrodite, or female (Bruce and Peter, 2008).The male trees are uncommon,
but sometimes occur when homeowners collect their own seeds. Hermaphrodite
trees (flowers with male and female parts) are the commercial standard,
producing a pear shaped fruit. These plants are self - pollinated (Jari, 2009).
Female flowers give way to smooth-skinned green pea-sized black seeds. Fruits
and seeds are edible. The interior flesh of the fruit goes through colour
changes from green (immature) to yellow (ripe and when it is overripe) (McGrath
and Karahadian, 1994). It has been found t be available all through the year
(Banerjee et al., 2006).
Phytochemical
Components of Carica papaya
Proteolytic
enzymes
Papaya
contains several unique protein-digesting proteolytic enzymes including papain
and chymopapain (Aravind et al.,2013). Papain is similar to pepsin and it is
also used to treat arthritis. Aside from its value as a remedy in kindred
ailments, it has been utilized for the clarification of beer. The juice has
been in use on dyspepsia meat to make it tender, (Wilson, 1994). Chymopapainis
a more abundant proteolytic enzyme found in Carica papaya but papain is twice
as potent (Morton 1987). Both papain and chymopapain can help lower
inflammation and improve healing from burns.
Carpaine
This
alkaloid occurs in all the green parts of the Carica papaya plant and in the
seeds (Everette,1971). It slows the heart rate in humans and thus reduces blood
pressure. Its action is similar to the drug prescribed for heart patients,
digitalis. The alkaloid is reported to be able to kill worms and amoebas.
Lycopene
Lycopene is
a bright red carotene and carotenoid pigment and phytochemical found in
tomatoes and other red fruits and vegetables including papayas. Papaya has an
abundance of cancer fighting lycopene. It is a key intermediate in the
biosynthesis of many important carotenoids, such as beta-carotene and
xanthophylls.
Fibrin
Another
useful compound not readily found in the plant kingdom is Fibrin. It reduces
the risk of blood clots and improves the quality of blood cells, optimizing the
ability of blood to flow through the circulatory system. Fibrin is
alsoimportant in preventing stroke.
Nutritional
value of Carica papaya
The fruit of
Carica papaya is not just delicious and healthy but whole plant parts, fruit,
roots, bark peel, seeds and pulp are also known to have medicinal properties.
The many benefits of papaya is due mainly to high content of vitamins A,B and C
(Peterson et al., 1982). Proteolytic enzymes like papain and chymopapain which
have antiviral, antifungal and antibacterial properties. The extracts of unripe
Carica papaya contain terpenoids, alkaloids, flavonoids, carbohydrates,
glycosides, saponins and steroids (Aravindet al., 2013).
Table 1:
Papaya raw Nutritional value per 100g
Energy 163KJ
Sodium 3mg
Potassium
257mg
Phosphorus
5mg
Magnesium
10mg
Iron 0.10mg
Calcium 24mg
Vitamin C
61.8mg
Folate (vit
B9) 38mg
Vitamin B6
0.1mg
Niacin (vit.
B3) 0.338mg
Riboflavin
(vit. B2) 0.05mg
Thiamine
(vit. B1) 0.04mg 0.04mg
Vitamin A
328mg
Protein
0.61mg
Fat 0.14mg
Dietary
fibre 1.8mg
Sugars 5.9mg
Carbohydrates
9.81mg
(Source:
Aravindet al., 2013)
Actions of
Carica papaya
A.
Antisickling Effect of Carica papaya
Sickle cell
disease is a genetic disease affecting the haemoglobin causing it to assume a
sickle shape that cannot effectively carry oxygen to tissues and one of the
many symptoms is anaemia (Koch et al., 2000). Some tropical plants like Carica
papaya have been found to have antisickling properties and are employed in the
management of sickle cell disease (Sofowora et al., 1975, Elekwa et al., 2003;
Imaga et al., 2009). The antisickling potential of tropical plants is due to
their peculiar content of a wide variety of biologically active substances and
amino acids capable of reversing sickling (Sofowora 1975, Ekeke and Shode,
1990). Antisickling activity of Carica papaya aqueous extract has been
established and the antisickling agent was found to reside in the ethylacetate
fraction of the methanol extract (Thomas et al., 1987). P-hydroxybenzoic acid
and 3,4-dihydroxybenzoic acid (protocatechuic acid,) and
4-hydroxy-3-methoxybenzoic acid (vanillic acid) are chemical constituents
responsible for the antisickling activity of these plant and has also been used
as a positive control in antisickling assays (Khadem et al., 2010). It has also
been reported that Carica papaya oil extract is effective in enhancing sickle
cell (Hbss) polymerization in only females but positively modulates peroxidase
activity in both sickle cell sexes. It can hence be seen that the effectiveness
of Carica papaya in managing sickle cell disease is sex dependent and this
should be considered when administering the plant extract (Afolabiet al.,2012).
B. Membrane
Stabilization Properties of Carica papaya
Extracts of
papaya leaves have showed significant reduction in heat-induced hemolysis
(Priyanga et al., 2012). This shows that Carica papaya has a therapeutic
potential on disease processes causing destabilization of biological membranes.
Carica papaya leaf extracts have also demonstrated a positive effect on
increasing platelets counts in healthy mice (Sathasivam et al., 2009). Any
compound or drug having a stabilization effect on the plasma membrane may
effectively enhance survival of platelets with a potential morbidity and
mortality benefits in patients with dengue viral infections. Mechanisms for
this effect of Carica papaya is not yet known but a number of studies have
revealed that flavonoids (David, 2007) and a host of other plant compounds
(Jorge et al., 2004) exhibit analgesic and anti-inflammatory effects as a
result of their membrane stabilizing ability in various experimental models.
Antiferility
Effects of Carica papaya
Infertility
is one of the adverse reactions from the consumption of Carica papaya. Aqueous
extracts and benzene extracts given orally to female rats caused infertility
and irregular oestrous cycles while in male rats given ethanol seed extracts
orally or intramuscularly had decreased sperm motility. The oral doses also
decreased testis mass and sperm count. The fertility of the male and female
rats returned to normal within sixty (60) days after the treatments were
discontinued (Harsha and Chinoy, 1996; Chinoy et al., 1997). Carica papaya seed
extract and antifertility activities have also been discovered in langur
monkeys and male humans (Lohiya et al., 2002). A study was carried out which
revealed that alkaloid extracts of Carica papaya inhibited the serum level of
androgen- testosterone which might result in infertility and reduce the effect
of androgens to increase erythropoiesis (Udoh et al., 2009).
Nephroprotective
properties of Carica papaya
In previous
studies, it has been reported that pretreatment with Carica papaya seed extract
prevented chloroform tetrachloride toxicity on the kidneys in both a time- and
dose- dependent manner, an effect which could be mediated by any of the
phytocomponents present in it via either antioxidant and/or free radical
scavenging mechanism(s) (Olagunju et al., 2009). Another study also revealed
that pretreatment with aqueous extract of Carica papaya L. significantly
attenuated the physiological and histopathological alterations induced by
gentamicin in the kidney and liver (Nale et al., 2012).The biochemical and
histological findings obtained from these studies were suggestive of the fact
that the extract may be mediating its protective effects by acting as a chain-breaking
antioxidant for scavenging free radicals or by some other mechanisms depending
on the condition of the kidney. In an earlier study, the presence of alkaloids,
flavonoids, saponin, tannin, anthraquinones, and anthacyanosides in Carica
papaya seed extract was reported (Adeneye and Olagunju, 2009).
Other uses
of Carica papaya include the following
Antioxidant
Property of Carica papaya
Pawpaw
fruits have a juicy taste rich in antioxidant nutrients like carotene, vitamin
C, vitamin B, flavonoids, folate, panthotenic acids and minerals such as
potassium and magnesium. The antiradical and antioxidant activities of Carica
papaya leaf extract has been correlated with the polyphenolic components
(Srikanth et al., 2010; Aiyegoro and Okon, 2009) hence it helps to prevent some
forms of cancer.
Gastroprotective
Properties of Carica Papaya
Papain
belongs to a family of related proteins with a wide variety of activities,
including endopeptidases, aminopeptidases, dipeptidyl peptidases and enzymes
with both exo- and endopeptidase activity (Rawlings et al., 1994). Green or
unripe papaya contains more papain than the fully ripened fruit hence green
papaya is normally prescribed for acid indigestion and stomach ulcers.
Antihypertensive
Actions of Carica Papaya
Hypertension
is the most common cardiovascular disease and constitutes a major factor for
several cardiovascular pathologies including atherosclerosis, coronary artery
disease, myocardium infarction (oparil et al., 1999). Root extract of the
Carica papaya plant gave similar effect of excretion of urinary electrolytes to
that of the standard drugs. The determination of the osmolality of urine and
electrolyte per unit time suggests that the extracts of Carica papaya exhibit
such effect due to its high salt content (Sripanidhulchai et al., 1994). It has
been reported that oral administration of papaya plant has anhypotensive effect
(Arya et al., 2005). Proper standardization and clinical trials is needed to
exploit therapeutic utility to combat cardiovascular diseases (Thakur et al.,
2012).
Antidiabetic
Effect of Carica Papaya
Some
tropical herbs like Carica papaya offers both benefits as it has been recently
informed in experimental models (Andrade-Cetto et al., 2005, Oliveira et al.,
2009, Runnie et al., 2004). It has been observed that the aqueous extract of
Carica papaya exerted a hypoglycemic effect. It also improved the lipid profile
in diabetic rats (Isela et al., 2012).
Antibacterial
and Antiparasitic Action of Carica papaya
Carica
papaya fruit and seed extracts have been reported of possessing pronounced
bactericidal activity against Staphylococcus aureus, Bacillus cereus,
Escherischia coli, Pseudomonasaeruginosa and Shigellaflexnori (Emeruwa 1982).
The pulverized seeds are also documented to possess anti-parasitic activities
against Entamoebahystolica and Dirofilariaimmitis infections (Suhaila et al.,
1994; Tona et al., 1998).The seed is used for intestinal worms when chewed.
Carica
papaya L. leaf tea or extract has a reputation as a tumour-destroying agent
(Walter Last, 2008). It appears to boost the production of key signaling
molecules called Th1-type cytokines, which help regulate the immune system.
Papaya
leaves are made into tea as a treatment for malaria (Titanjiet al., 2008).
Antimalarial and antiplasmodial activity has been noted in some preparations of
the plant, but the mechanism is not understood and not scientifically proven.
Rubbing the
white pulp of raw papaya improves pimples as well as wrinkles. Papaya works as
a good bleaching agent. It is an important ingredient in bath soaps,
astringents, detergent bars and hand washes.
Protein
enzymes including papain and chymopapain and antioxidant nutrients found in
papaya; including vitamin C, vitamins E, and beta-carotene, reduce the severity
of the conditions such as asthma, osteoarthritis, and rheumatoid arthritis
(Elizabeth, 1994; Owoyele, 2008).
Also home
applications of leaf and bark papaya extract is used to deal with mouth gums
and toothaches which are being effectively practiced in many cultures around
the world.
The black
seeds of the papaya are edible and have a sharp, spicy taste. They are
sometimes ground and used as a substitute for black pepper.
Carica
papaya is also known to have nephroprotective (Olagunju, 2009) and
neuroprotective function
SEX
VARIATIONS AND THE EFFECTS OF CARICA PAPAYA
Evidently,
it can be seen that Carica papaya mediates its effects on blood parameters
directly or indirectly. Carica papaya high iron and vitamin B contents are
contributing factors for stimulating erythropoiesis. It has however been
observed that Carica papaya effects tend to vary with sex as reported by
Afolabi (2013) in its antisickling and Lohiyaet al (2002) in its antifertility
effects.
These
variations of Carica papaya extract due to sex could be due to the direct
effect of Carica papaya one sex hormones. It has been previously reported that
the major cause of the difference in blood parameters in males and females is
due to the stimulating effects by androgens in males (Siddique et al., 2004,
Dobset al., 1999) and inhibitory effects of estrogen (Watanukiet al., 2002).
However, there could be some other mechanisms that could cause sex variation in
the effects of Carica papaya on haematological parameters.
AIMS
To study the
effects of Carica papaya seed extract on red blood cell, packed cell volume and
haemoglobin.
To investigate
the sex variation of the effects of Carica papaya seed extract on red blood
cell, packed cell volume and haemoglobin.
CHAPTER ONE
INTRODUCTION
BACKGROUND
STUDY
Blood is a
special type of connective tissue composed of formed elements in a fluid matrix.
The formed elements include red blood cells (erythrocytes), white blood cells
(leukocytes) and platelets (Bacha and Bacha, 2000). It is a bodily fluid in
animals that delivers necessary substances such as nutrients and oxygen to the
cells and transports metabolic waste products away from those same cells.
Assessment
of haematological parameters can be quite informative about the health status
of the individual and it is among the methods which may not be apparent during
physical examination but may affect the fitness of the animals (Kronfeld et
al., 1969). The red blood cell count, white blood cell counts, packed cell
volume and haemoglobin concentration are regularly measured haematological
parameters but this study is concerned with red blood cells, packed cell volume
and haemoglobin.
Most of the
haematological parameters are influenced by factors like sex, age, breed,
seasonal variations, lactation, health and nutrition status (Aengwanich, 2002;
Al-shami, 2007; Mohammed et al., 2007). Since it is a well-documented fact that
there are variations due to age, sex, attitude etc. (Hawkins et al., 1954,
Viteri et al.,1972, De Grunch et al.,1989), it is acknowledged that for
comparisons between individuals and with reference data in a clinical
diagnostic situation, it is necessary to consider normal variations due to sex,
age and breed in order to increase diagnostic precision (Satue et al., 2009).
It has however been observed that the modifications correlated with sex, age
and taxonomic position are best shown by the erythrocytic counts (Duguy 1970).
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