Liver Modulatory Function And Gastroprotective Effect Of Psidium Guajava (Guava) Leaf Extract On Experimental Rats Model

ABSTRACT

Liver modulatory function and gastro protective effect of aqueous leaf extract of psiduim guajava (guava) on gastrointestinal function of alcohol-induced ulcer in albino Wister rats was studied. Ninety five (95) albino rats weighing 120g- 200g were divided into four different phases. These rats were used to evaluate the liver modulatory function and gastro protective effects of P. guajava leaf.  In phase one, 30 albino rats were used for acute toxicity value of the p. guajava leaf. In phase two, 20 rats were randomly assigned into four groups of 5 rats each. These groups were used for gastric acid, mucus secretions and liver functions parameters. In phase three, 20 rats were also assigned into four groups of 5 rats each and they were used for total acidity, stomach PH and pepsin activity. In phase four, 25 rats were randomly assigned into 5 groups of 5 rats each. Group 1 for control, group 2 untreated controls, group 3 omeprazole treated,   and group 4 extract treated groups. In this phase, all the groups were fed on normal rat chow for 28days and allowed free access to water. Ulcer was induced in the test groups by oral administration of alcohol in a single dose of 20%. Phytochemical screening of the leaf extract was determined. A toxicity study was carried out on the extract, which showed LD50 of more than 6000mg/kg body weight. The extract was administered per oral to the test groups at the dose of 400mg/kg and 800mg/kg body weight respectively. At the end of treatment period, body weight and several tests of gastrointestinal functions were carried out. The body weights of the groups were measured at the beginning and at the end of the treatment period.  The loss in body weight was significant (P<0.05) higher in the test groups than the control. In the liver enzyme group analysis, ALT and ALP were not altered in the test groups when compared with control, they were significantly reduced (P>0.05) when compared with untreated control, AST level was higher in the test groups when compared with the control but was not significantly different .While in the untreated, AST level was lowered (p>0.05). Serum bilirubin and total protein concentrations were not altered when compared with control groups .Mucus secretion was significantly higher in the treated groups than untreated and control groups (p>0.05). The untreated control alcohol induced mean ulcer scores were significantly (P<0.05) higher than ulcer treated and control group. The ulcer index was lower in the treated group when compared with the standard drug omeprazole treated group. Thus the percentage inhibition of ulcer was higher in the extract treated groups than omeprazole treated group. The intestine motility result showed the decrease in the amplitude of concentration of the rabbit jejunum when compared with basal values. The extract like atropine significantly inhibited acetylcholine induced contractions in dose dependent manner. The highest of extract used completely cancelled out Ach induced contraction on the piece of tissue. The microscopic findings in the stomach showed minimal metaplastic and mucosa inflammatory cells in the treated groups when compared with control.  There were regenerating epitheliums noticed in the treated groups than untreated control. The results of   phytochemistry of P.guajava leaf showed that P.guajava leaf contains saponins, tannins and phenolic compounds in higher quantities, while flavonoids, terpenes and glycoside are in a moderate quantities, however, steroids and alkaloid are only present in a small quantities .In conclusion, this findings suggest that alcohols – induced ulcer decreases mucus secretion, increased basal gastric acid secretion, increase ulcer scores and increase liver enzymes in rat. P. guajava treatment ameliorated these derangements and exhibited gastro protective and hepato protective activities.

CHAPTER ONE

 1.0     INTRODUCTION

The liver is a vital organ that plays many important metabolic functions. It is uniquely situated in the right upper abdomen, below the diaphragm. It takes up most of the space under the ribs and some space in the left upper abdomen. The human adult liver weighs between 1.4kg and 1.66kg and contains two robes which are separated by a bound of connective tissue that anchors the liver to the abdominal cavity. (Petridis et al., 1994).

The liver helps to process and distribute dietary nutrients because the venous drainage of the gut and pancreas passes through the hepatic portal vein before entering the general circulation.  Thus after a meal, the liver is bathed in blood containing absorbed nutrients and elevated level of insulin. During the absorptive period, the liver takes up carbohydrate, lipids and most amino acids. These nutrients are then metabolized, stored, or rooted to other tissue (Cheng et al., 2008).

Apart from nutrients metabolism, the liver also functions in the metabolism and excretion of exogenous (drugs, alcohol, copper) and endogenous (bilirubin, hormones) materials. The liver stores vitamins (A, B12 and foliate) and minerals. It also synthesis the following albumin, coagulation factors and inhibitors (except Von willed brand factor) complement factors, transferrin, heptoglobin, caeruloplasmin, protease inhibitors (x1-antitrysin) and fetoprotein. Because of the importance of the liver and its functions, evolution has ensured that it can regenerate rapidly as long as 25 percent of the tissues remain.

Several signal pathways are known to stimulate regeneration in the liver including growth factors, cytokines, coagulation factors and inhibitors, hormones, and nuclear receptors. (Michalopoulos, 2013).

The liver as a complex organ can experience a range of problems, which include; fascioliasis caused by the parasitic invasion of a parasitic worm known as a liver fluke, which can lie dormant in the liver for months or even years. (Blitzer et al., 2006). Cirrhosis of the liver also affect liver cell in a process known as fibrosis, other diseases of the liver include, hepatitis, fatty liver disease, alcoholic liver disease, Gilbert’s syndrome and liver cancer (hepatocellular carcinoma ) (Larson,2016).

The liver has been known as one of the accessory organ of G.I. T that aid in digestion, through its roles in the production of bile and metabolism of nutrients.  Gastrointestinal system has a crucial role in the control of energy homeostasis through its role in the digestion, absorption and assimilation of ingested nutrients. Furthermore, signals from the gastrointestinal tract (GIT) are important regulators of gut motility and satiety. (Duckers, 2007).The protective mechanisms of the gastrointestinal tract mucosa can be overwhelmed by some GIT diseases like peptic ulcer which is associated with several changes in gastrointestinal (GI) motility (Allen et al., 2000).

Ulcer is a break (like a sore) in the lining of the stomach or the upper part of the small intestine, with a diameter of at least 0.5cm penetrating through the muscularis mucosa. (Sung et al., 2009). Peptic ulcer disease (PUD) is generally categorized based on their anatomical origin as gastric or duodenal ulcers. Gastric ulcers are found along the lesser curvatures of the stomach and duodenal ulcer usually occurs in the duodenal bulb, the area’s most exposed to gastric acid. Helicobacter pylori are the main risk factor for 90% duodenal and 80% gastric ulcers. .

In African, the incidence of peptic ulcer continues to increase by four fold especially the duodenal ulcer. Gastric ulcer proximately arise in subject particular occur between the ages of 20-50 years and associated symptoms such as heart burn, abdominal pain, and constipation.(Sung et al.,2009).

The prevalence of ulcer and other gastrointestinal tract disorders over the world is rapidly assuming an epidemic proportion.

The search for a common, most effective, safe, convenient   and affordable treatment for GIT disease is unending at the moment. The use of herbal medicinal plant in the treatment of human disease conditions is currently of immense interest to scientists and orthodox medical researchers, as the efficiency of some herbal medicinal plant is no more in doubt.

During the last century, the practice of herbalist has become mainstream throughout the world. Moreover, plants now form an integral part in the great advances made in modern medicine. This is due in part to the recognition of the value of traditional medicinal systems, and the identification of medicinal plants from indigenous pharmacopoeias, which have significant healing power. (Hotlez et al., 2001).

Medicinal plants are distributed worldwide but they are most abundant in tropical countries. Among all families of the plants kingdom, members of the myrtaceae have been used for centuries in folk medicine. Psidium guajava (myrtaceae), commonly known as guava has been used naturally in the treatment of different diseases like diarrhea stomach ache (Hassimotto et al., 2005). Thus, nature has not only provided us with a very beautiful environment to live, it has also been a dependable resource for solution to the number of health problem playing man. P. guajava is believed to originate from Mexico and extents to South America and Brazil. It is also found in West Africa and Asia hot climates zones. In Nigeria, it grows widely as a seasonal fruit. The wide usage of P. guajava is as a result of its vague of biological importance, ranging from traditional, nutritional and medicinal value.

1.1     BACKGROUND OF THE STUDY

The botanical name Psidium guajava (P.guajava) is derived from the Greek language psidion due to a fancied resemblance between the two fruits. It is one of the species from the genus psidium. It is commonly called Guava (Mark, et al., 2011). It is classified as follows:

Kingdom – planate

Division – Angiosperms / eudicots

Class – Rosids

Order – Myrtles

Family – Myrtaceae

Genus – Psidium

Species – P. guajava.

Psidium is a genus of about 150 species of evergreen trees and stubs in the family myrtaceae. P. guajava commonly called guava is most widely distributed in tropical and subtropical areas of the world adapts to different climatic conditions but prefers dry climates. P. guajava is an evergreen tree growing up to 10m high, branching from the base and often producing suckers. The leave, usually 3-10mm long 4-6cm wide. the fruit an ovoid or pear shaped is usually 4-12cm  long weighing up to 500g , skin yellow when ripe , sometimes flushed with red pulp juicy creamy white or creamy yellow to pink or red , the soft enveloping numerous , cream brown , kidney – shaped or  flattened seeds. The exterior of the fruit is fleshy and the centre consists of a seedy pulp (stone, 2009).

There are two most common varieties of P. guajava. The red psiduim guajava (pomifera) and the white P. guajava (pyrifera), (Lorenz et al; 2012). All parts of this tree, including fruits, leaves, bark and root have been used for treating stomach ache and diarrhea in many countries. Leave, pulp and   seed are used to treat respiratory and gastrointestinal disorder, and as an antispasmodic, anti- inflammatory, as a cough sedative, in the management of hypertension, obesity and in the control of diabetes mellitus. It also possesses anticancer properties .The seeds are used as antimicrobial, anti- allergic and anti carcinogenic activity (Hassimotto et al., 2005).

PHOTOCHEMISTRY OF PSIDIUM GUAJAVA

Photochemical are naturally occurring biologically active chemical compounds in plants and act as natural defense system for host plants and provide color, aroma and flavor. Photochemical test on the plant p.guajava revealed important constituents as tannins, alkaloids, saponins, flavonoids mainly quercetin derivative, and phonetic compounds. (Zachariah et al., 1994). The psidium guajava leaves contain essential oil with the main components being alpha- pinene, limonene, menthol, terpenyl acetate, and isopropyl alcohol. It also contains Gallic acid, catechins, epicatechins and kaempferol.

Also ascorbic acid, cinnamyl alcohol, ethyl benzoate, and alpha bisabolene have been identified. The antioxidant properties of the guava pulp can be related to anticancer effects. Catechins contained in the leaves of p.guajava are important as a preventive treatment for diabetes type 2 and obesity. Quercetin has been associated to decreased mortality from heart disease and decreased incidence of stroke. The antispasmodic action of thymol contained in the leaves of p. guajava could be associated with reduce intestinal motility. (Zachariah et al., 1994).

This action in conjunction with anti bacterial property of the plant offers potential for controlling epidemic of cholera. Begum et al., 2000 also reported that p.guajava leaves have a broad spectrum of antimicrobial action that could be associated in controlling diarrhea due to a wide range of pathogens. This antimicrobial activity can be linked to the presence of flavonoids contained in the leave.

Tannins act on proteins and also form a protective layer on the skin and mucus membrane. Thus, they can bind the tissue of the gut and reduce diarrhea or internal bleeding externally. They are useful in the treatment of burns, for healing wounds and reduce infection in the eye, mouth, vagina and cervix (Willard, 2002).

Alkaloids are the most potent group of plant constituent that act upon the human body and mind. Their actions improve the state of the liver, nerve, lungs and digestive system. (Kom and keviles, 1996).

Sponins can be used in the synthesis of cortisone, a strong anti-inflammatory drug and also in the synthesis of sex hormones. The sponins found in herbs is used by the body as raw materials to build up chemicals. They are expectorant in action through stimulation of reflex in the upper digestive tract. (Kom and keviles, 1996).

Phenols have antioxidant properties which carry out their protective activity on cells either by preventing the productions of free radicals or by scavenging free radicals produced in the body or chelating the transition metals. It has been reported that the total phenol content of two species of psidium guajava was seventy –two with the highest amount of phenol compounds in pyrifera varieties than that of pomifera verities (Baby et al., 2007, Mello et al., 2010).

1.2     USES OF P. GUAJAVA

1.1.2.1 NUTRITIONAL VALUE OF P.GUAJAVA

The plant psidium guajava contain protein (3.8g), water (197g), ash (2g), and total calories 167. Calories from carbohydrate are 13g. (Mello et al., 2010). The total carbohydrate found in P.guajava is 38g, dietary fiber 13g and sugar 22g. Total fat found 1.9g, saturated fat 542mg, monounsaturated fat 173mg, polyunsaturated fat 798mg, omega 3 fatty acids 223mg, omega, 6 fatty acid 573mg.

The P.guajava has also been reported to have many vitamins include vitamin A, 87/iu, vitamin B, 268mcg, foliate 120mcg, pantothenic acid 1.1mg. The plant also contain major minerals like calcium 48mg, iron 586mcg, magnesium 48mg, phosphorus 82mg, potassium 865mg, sodium 48mg, zinc 561mcg, copper 56mg, manganese 366mcg and selenium. (Baby et al., 2007).

1.1.2.2 MEDICINAL USES OF PSIDIUM GUAJAVA

The whole plant of P .gaujava have many uses in   traditional medicine, especially in Mexico and other central American countries including the Caribbean, Africa and Asia.

Throughout the world, P. guajava is used as an anti inflammatory, used in the treatment of gastroenteritis, anti-diarrhea, dysentery and anti hypertension (Heinrich, et al., 1998).

Oils from the leaves have been reported to possess an interesting spectrum of anti microbial properties, antibacterial and antioxidant properties (Nicholson, et al., 2000) and catechins found in the leaves are important as a preventive treatment for diabetes type 2 and obesity. Rustin in the leaves is also effective in the inhibition of triglyceride accumulation in adipocytes. Ghosh et al 2009, isolated two terpenoids from the leaf extract of P. guajava (betulinic and Lupeol) and reported their potential antimicrobial and phytotoxic activities and these can be used in the treatment of cardiovascular disease, obesity and atherosclerosis.  The leaf also has high potential anti plague agent by inhibiting the growth of the streptococcus mutas. (Shaom et al., 2010).

Clinical trials in creams formulated against dermatological disease have yielded favorable result (Felice et al., 2012). In coastal areas of Nigeria, the plant is used in the treatment of diarrhea, reducing pain and reducing fever. (Felice et al., 2012).

1.1.2.3   NON MEDICINAL USES OF P. GUAJAVA

In Asia, the leaves are employed to give a black color to cotton. In Indonesia, the leaves also serve to dye malting. (Argueta et al., 1994).

The wood is used in the construction of houses in Nigeria (Lucas et al., 2006). In Malaya, the wood and leaves are used by carpentry and turnkey to make a black dye for silk, while in Mexico, the tree is seeded to give shade to the coffee and it may be parasitized by the mistletoe, calyculatus on producing the rosette like malformations called wood flowers, which are sold as ornamental curiosities (Argueta et al., 1994).

1.1.3 STUDY OF THE GASTROINTESINAL SYSTEM AND FUNCTIONS

Gastro-intestinal system says “I pulverize the food and supply the same for energy generation and body growth. But greedy eaters abuse me only to become obese and Sick (Reddy et al., 2008).

Human beings and other animals must obtain their basic organic molecules from food. The main process by which this is achieved is through the gastro intestinal system. This process involves ingestion of food, digestion, absorption, and elimination of waste products. The gastrointestinal system consist of the alimentary canal which extent the full length of the truck, open at both ends (mouth and anus) and its associated structures and accessory organs that assist the tract by secreting enzymes  to help break down food into its component nutrients. Thus the salivary glands, liver, pancreas and gall bladder have important functions in the gastrointestinal system. (Reddy et al., 2008).

The functions include:

Motility:  This refers to the movement of food through the digestive tract through the processes  of :  Firstly;  food must  be  ingested  into the mouth to be  mechanically processed and moistened .

Secondly, Deglutition, which means swallowing of food.

Thirdly, Digestion, which refer to the breakdown of food into smaller subunits which can be absorbed.

Fourth, Absorption, the passage of smaller molecules across the epithelium of the smaller intestine and subsequently enter the blood or lymph.

Fifty, Defecation .when undigested material and secreted waste products are expelled from the body [passing of feces].

Finally, Excretion, certain harmful substances like heavy metals, metabolic end products like bilirubin are excreted. Also included are, production of some vitamins. The colonic bacteria are capable of producing vitamin K and B complex vitamins. Body   defense, the tonsils of oral cavity and pharynx and the lymphoid tissue of acid gastric juice kill the pathogens that enter via food. Acid base balance, the gastrointestinal system secretions are acidic at some location and alkaline at other sides. This type of secretion with varied participates in acid base balance.

1.1.3.1: COMPONENTS OF THE GASTRO INTESTINAL SYSTEM AND                  ASSOCIATED ORGANS

Oral cavity or Mouth. This is responsible for the intake of food. It is lined by a stratified squamous oral mucosa with keratin covering those areas subjected to significant abrasion, such as the tongue, hard palate and roof of the mouth.

Salivary glands

Three parts of salivary glands communicate with the oral cavity. Each is a complex gland with numerous acini lined by secretary epithelium. The acini secrete their contents into specialized ducts. Each gland is divided into smaller segments called lobes. They glands include

• Parotids glands. They are large, irregular shaped glands located under the skin on the side of the face, and secrete 25% of the saliva. It secretes serous secretions.
• Submandibular gland, secrete 70% of the saliva in the mouth and are found in the floor of the mouth, in a groove along the inner surface of the mandible. Its secretions are mucus and serous in nature.
• Sublingual glands are the smallest salivary glands covered by thin layer of tissues at the floor of the mouth. They produce 5% of the saliva and their secretions are very sticky due to the large concentration of mucin.

 Esophagus:

The esophagus is a muscular tube of approximately 25cm in length and 2cm in diameter it extends from the pharynx to the stomach after passing through the an opening in the diaphragm. The wall of the esophagus is made up of inner circular and outer longitudinal layers of muscle that are supplied by the esophageal nerve plexus. This nerve plexus surrounds the lower portion of the esophagus. (Osim et al., 1991).

Other components include stomach, small intestine, liver, gall bladder and pancreas.  The mucosal layer mainly consist of three different structures namely, lamina propria, connective tissues layer, muscularis mucosal smooth muscles layer, and the innermost epithelial cell layer resting on a basement membrane. This is thrown into columnar epithelial cells. The villa, give rise to hair like processes called microvillus. These arrangements increase the surface area for absorption of digested foods and secretions (Osim, 2002).

The serosal layer is the outermost covering of the alimentary tract; it carries nerves, blood and lymphatic vessels. It forms the smooth moist membrane which reduces friction between contacting surfaces. The muscular layer is made up of smooth muscles (visceral type). There are two types of layers, the inner circular and outer longitudinal; in between the two layers are the presents of mesenteric nerve plexus. In the stomach, the oblique layer is also present. In oral cavity and the upper 1/3^rd of esophagus, striated muscle is present.

The submucosal layer is loose areola connective tissue which contains large blood and lymphatic vessels, nerves and glands etc. (Osim, 2003). Between the circular muscle layer and the mucosa, the sub mucosal nerve plexus.

1.1.3.2 REGULATION OF THE GIT

The digestive system has a complex system of motility and secretion regulation which is vital for proper function. This task is accomplished via a system of long reflexes from the central nervous system (CNS), short reflexes from the enteric nervous system (ENS) and reflexes from GI peptides working in harmony with each other. (Towers et al., 2014).

LONG REFLEXES

Long reflexes to the digestive system involve a sensory neuron sending information to the brain, which integrates the signal and then sends messages to the digestive system. In some situations, the sensory information comes from the GI tract itself, information is received from sources other than the GI when the information comes from GI tract; these reflexes are called feed forward reflexes. This type of reflex includes reactions to food or danger triggering effects in the GI tract. Emotional responses can also trigger GI response such as the butterflies in the stomach feeling when nervous. The feed forward and emotional reflexes of the gastrointestinal tract are considered cephalic reflexes (Silver thorn, 2006).

SHORT REFLEXES

Control of the digestive system is also maintained by enteric nervous system, (ENS) which can be thought of as a digestive brain that can help to regulate motility, secretion and growth. Sensory information from the digestive system can be received, integrated and acted upon by the enteric system alone. When this occurs; the reflex is called a short reflex. Although this may be the case in several situations, the ENS can also work in conjunction with the CNS, vagal afferents from the viscera are received by the medulla, and efferent are affected by the vague nerve. When this occurs, the reflex is called vagovagal reflex. The myenteric and sub mucosal plexus are both located in the gut wall and receive sensory signals from the lumen of the gut or the CNS.

 GASTRO INTESTINAL PEPTIDES.

GI peptides are signal molecules that are released into the blood by the GI cells themselves. They act on a variety of tissues including the brain, digestive accessory organs, and GI tract. The effects range from excitatory or inhibitory effect on motility and secretion to feeling of satiety or hunger when acting on the brain. These hormones fall into three major categories, the gastrin and secretin families, with the third composed of all the other hormones unlike those in the other two families. These hormones include glucagon – like peptide which stimulates insulin release and inhibits glucagon release, motilin which stimulates migrating motor complex etc. (Kom et al., 1996).

1.1.3.3   GIT SECRETIONS

GIT secretion, include salivary secretion, gastric secretion, pancreatic secretion, small intestinal secretions (succus entericus) and large intestinal secretions. These are regulated by neural, mechanical and hormonal mechanisms. At any time during digestion all the three mechanisms interact to bring about optimum secretion. (Farrell, 1998.)

 1.1.3.4   SALIVARY SECRETION

The first secretion into the GI tract takes place in the oral cavity. There are three main types of salivary glands contributing to the production of saliva. These are the parotid, submandibular and sublingual glands, but also many small buccal glands contribute to the overall secretion. All salivary glands are comprised of multiple acini, which empty in collecting ducts before being secreted to the buccal cavity. Secretions differ for each type of salivary gland. Saliva contains a variety of protein compounds for different purposes and these changes are reflected in the secretion of each gland. Saliva also contains large amount of fluids, ions, and as well as bicarbonate (HCo3) for buffering purposes (Farrell, 1998).

The main component is a serous type of secretion which is based on an enzyme for the digestion of starches called ptyalin. This enzyme is an alpha-amylase. The other component of saliva is a mucus like secretion that contains mucin which a lubricant. This facilities swallowing and protects buccal surfaces. The parotid glands secrete only the enzyme containing type of saliva (serous). The submandibular and sublingual glands secrete a combination of serous and mucus type of saliva.

Finally, the buccal glands only produce mucus to protect the oral cavity. The saliva maintains a PH for the maximal activity of the alpha-amylase (Fox, 2004).

 1.1.3.5. GASTRIC SECRETION

In the stomach there are many mucus cells covering surfaces and pits. These cells produce thick mucus, alkaline in nature, which protects the tissue from the acid content. Within each pit there are tubular glands which can be oxyntic, also known as gastric, or pyloric. Oxyntic glands consist of parietal cells that secrete HCL and chief or (zymogenic) cells that secrete pepsinogen.  Enterochromaffin, like cells, found in the stomach and intestine, secrete histamine and 5-hydroxyptamine (serotonin) as paracrine regulators of the GIT (Fox, 2004).

Gastrin (G) cells secrete gastrin. D cells secrete somatostatin. In addition to these products, the gastric mucosa (probably the parietal cells) secretes a polypeptide called intrinsic factor, which is required for the intestinal absorption of VitaminB12 (Fox, 2004). Also, the stomach has recently been shown to secrete a hormone called gremlin. Its level rises before meals and falls after meals, this may serve as a signal from the stomach to the brain that helps regulate hunger. The secretion of HCL is stimulated by Vagal stimulation, histamine and gastrin. Vagal stimulation increases HCL secretion by a direct pathway and an indirect pathway (Linda, 1998). In the direct pathway, histamine is released from mast cells in the gastric mucosa and diffuses to the nearby parietal cells to stimulate Hydrogen ions secretion by activating H2 receptor on the cell membrane. The second messenger for histamine is CAMP. H2 receptor, blocking drugs, e.g. Cimetidine inhibit Hydrogen ions secretion by blocking the stimulatory effect of histamine. The 2^nd messenger for gastrin on the parietal cell has not been identified but is clearly different from those for ach and histamine because their actions are additive with those of gastrin (Feldman et al., 1986).

Negative feedback mechanisms inhibit the secretion of Hydrogen ions by the parietal cells. Low Ph in the stomach inhibits gastric secretion and thereby inhibits Hydrogen ions secretion. Chyme in the duodenum inhibits Hydrogen ions secretion both directly and via hormonal mediators (Feldman et al., 1986) .The hormonal mediators are GIP (released by fatty acids in the duodenum) and secretin.

If the normal protective barrier of the stomach is damaged, the presence of Hydrogen ions and pepsin secretions may injure the gastric mucosa, causing duodenal or gastric ulcer. Experiments demonstrate that the plasma membranes of the parietal and chief cells of the gastric mucosa are highly impermeable to the acid in gastric lumen (Reddy et al., 1998). Other protective mechanisms include a layer of alkaline mucus containing bicarbonate, covering the gastric mucosa, tight junctions between adjacent epithelial cells, preventing acid from leaking into the submucosa, a rapid rate of cell division, allowing damaged cells to be replaced and several protective effects provided by prostaglandins produced, by the gastric mucosa. A major causative factor in the development of peptic ulcer is helicobacter pylori infection. Helicobacter pylori have a high urea’s activity and converts urea to NH4, which damages the gastric mucosa.

Duodenal ulcers are more common than gastric ulcers. (Song et al., 2009]. A common cause of gastric ulcers is also believed to be use of no steroidal anti-inflammatory drugs (NSAIDS). These classes of drugs, including aspirin and ibuprofen, act to inhibit the production of prostaglandins. Experimentally peptic ulcers could be induced with 20% alcohol. When the gastric barriers to auto-digestion are broken down, acid can leak through the mucosa to the submucosa, causing direct damage and stimulating inflammation. The histamine released from mast cells may stimulate further acid secretion and result in further damage to the mucosa (Moore et al., 1993).

 1.1.3.6   PANCREATIC SECRETION

The pancreas is the next organ contributing to the digestive process. This organ has a similar glandular design to the salivary gland, but the secretions are different .Pancreatic secretions are derived from two functionally difference portions, the endocrine and exocrine pancreas.  The endocrine part produces insulin, gremlin, glucagon, pancreatic polypeptide (PP) and somatostatin                                                                                              . Exocrine part produces enzymes, HCO3 and water essential for digestion of protein, fat and carbohydrate. Pancreatic secretion is stimulated by secretin, CCK, and Ach (via vagovagal reflexes). Secretion is secreted by the tubular cells of the duodenum in response to the Hydrogen ions and, acts on the pancreatic ducal cells to increase HCO3 secretion using CAMP as 2^nd messenger (Heller et al., 1995). CCK is secreted by the cells of the duodenum in response to small peptides, amino acids, and fatty acids in the duodenal lumen and acts on then pancreatic acinar cells to increase enzyme secretion (amylase, lipases, proteases) ,using ip3 and increase intracellular (Ca2+) as 2^nd messenger. CCK potentiates the effect of secretion on ductal cells to stimulate HCO3 secretion. The potentiating effects of CCK on secretion are explained by the two GI hormones (i.e. CAMP for secretin and Ca2+ for CCK. (Walsh, 1994).

Disorders associated with pancreatic secretion include: steotorhea, cystic, fibrosis of pancreas and acute pancreatitis. Steotorhea increase lipid excretion into stools due to pancreatic exocrine or bile insufficiency. Fibrosis of pancreatic parenchyma causes a deficiency in exocrine secretions. This is due to defect in the secretion of chloride ions, which is a genetic defect. The defective gene is present on the long arm of chromosome 7. This may again cause mal absorption steotorhea. It appears that lack of trysin inhibitor of bile into the pancreatic duct causes activation of pancreatic enzymes within the pancreas leading to auto digestion of pancreas. This results in acid pancreatitis (Edwards et al., 1995).

 1.1.3.7 BILE SECRETION AND GALLBLADDER FUNCTION

The secretary function of the liver is limited to the production of bile. The bile is secreted by the liver at a rate of 30-60ml/hr. Total secretion is 500- 1000ml/24hrs. It is golden yellow in color and is stored in the gallbladder, till it is evacuated (Reddy et al., 2008). Bile contains water, bile salts, bilirubin, cholesterol, fatty acid, lecithin as well as Na+, k+, Ca+, CL- and HCo3 Most of these materials are continuously recycled. It is estimated that 94% of the secreted bile salts reabsorbed by the small intestine. Half of it re- enters the mucosa of the upper small intestine by diffusion and the rest is actively transported by the distal ileum. Bile is concentrated in the gallbladder as a result of isometric absorption of solutes and water. The serve as the emulsification and solubilization of fat into micelles for digestion and absorption in the small intestine as carrier for circulatory waste, which once it is secreted into the small intestine, is not reabsorbed and it is eventually discarded with the faces (Reddy et al., 2008).

Abrams et al., 1988 also reported that CCK is released from duodenal mucosa in response to peptides and fatty acids, and causes contraction of the gallbladder and relaxation of sphincter of Odd.

 1.1.3.8   INTESTINAL SECRETION

The GIT secretes electrolytes from blood to lumen. The secretary mechanisms are located in the crypts of Lieberkuhn. These crypts contain some goblet cells, which produce mucus to protect the villas surface and the enterocytes. The enterocytes within the crypts secrete large volumes of water and electrolytes. Water secreted follows Nacl to maintain is- osmotic conditions. Chloride is the primary ion secreted into the intestinal lumen. It is transported through CL channels in the luminal membrane that are regulated by CAMP (Linda, 2008). Na+ is secreted in the lumen by passively following CL channels.  A potassium ion is actively secreted in the colon by a mechanism similar to that for Potassium ions secretion in the renal distal tubule. As in the distal tubule secretion in the colon is stimulated by an aldosterone. (Linda, 2008).

 1.1.3.9 DIGESTION AND ABSORPTION

Carbohydrates, proteins and lipids are digested and absorbed in the small intestine. The surface area of absorption in the small intestine is greatly increased by the presence of the brush border.

 CARBOHYDRATES

Carbohydrates must be digested to glucose, galatose, and fructose for absorption to proceed, as only monosaccharides are absorbed. Alpha amylases (Pancreatic and salivary) hydrolyses 1, 4 glycoside linkage in starch yielding maltose, maltotriose and alpha limit dextrin’s (Southgate, 1995).

The disaccharides produced alpha-dextrin; maltase and sucrose are all converted to glucose by brush border enzymes. (Levin, 1994).Glucose and galatose are absorbed across the apical membrane by secondary active transport along with Na ions through the sodium glucose co-transporter (SGLTI). This is facilitated diffusion. The sugar is transported “uphill” and Na ions is transported “downhill” (Kellet et al., 1984 ).

Na+K+ pump in the bilateral membrane keeps the intracellular Na+ concentration low, thus maintaining the Na+ gradient across the luminal membrane poisoning the Na+ gradient . Fructose is transported exclusively by facilitated diffusion; therefore it cannot be absorbed against concentration gradient.

 1.1.3.10  PROTEINS

Endopeptidases degrade proteins by hydrolyzing interior peptide bonds, while exoptidase hydrolyze one amino acid at a time from the terminus of proteins and peptides (Freeman et al., 1979). Pepsin is not essential for protein digestion when the PH is less than 5. At this PH, pepsin is denatured. Thus, in the intestine as HCo3 is secreted in pancreatic juice, duodenal PH increases and pepsin is inactivated. Pancreatic proteases, trysin, chymotrypsin elastase, carboxypetidase A and B are activated consequentially by trysin, after enterokinase (a burse border enzyme) has activated trysinogen to trypsin. The process of digestion is completed in the small intestine. These enzymes split the Oligopeptides into amino acids, dipeptides and tripeptides which is absorbed. (Freeman et al., 1979).

Digestive products of protein can be absorbed via N+ dependent amino acid co-transport which occurs into luminal membrane that acid transport of amino acid from cell to blood by facilitated diffusion. There are four separate Corners for neutral, acidic, basic and the amino acids, proline and hydroxyproline respectively. Dipeptides and tripeptide are absorbed faster than free amino acids. H+ dependent co-transport of dipeptides and tripe- tides also occurs in the luminal membrane after the dipeptides and tripeptides are transported into the intestinal cells, cytoplasm peptides hydrolyze them to amino acids, the amino acids are then transported from cell to blood by facilitated diffusion.(Slazenger  et al ., 2009)

1.1.3.11   LIPIDS

Digestion of lipid began with the action of lingual lipases which digest some of the ingested triglycerides to monoglycerides and fatty acids (Carey et al., 1983). However, most of the ingested lipids are digested in the intestine, by pancreatic lipases. Bile acids emulsify lipids in the small intestine, increasing the surface area for digestion. Pancreatic lipases hydrolyze lipids to fatty acids, monoglycerides, cholesterol, and phospholipase A2 (Carey et al., 1983).The hydrophobic product of lipid digestion is solubilized in micelles by bile acids. Micelles bring the products of lipid digestion into contact with the absorptive surface of the intestinal cells. Then, fatty acids, monoglycerides, and cholesterol diffuse across the luminal membrane into the cell (Linda, 1998). Glycerol is hydrophilic and is not contained in the micelles. In the intestinal cells, the products of lipid digestion are re-esterifies to triglycerides, cholesterol esters, and phospholipids and with lipoproteins, form chylomicrons (Glaze et al., 2002).

The later are transported out of the intestinal cells by exocytosis. Because chylomicrons are too large to enter the capillaries, they are transferred to lymph vessels and are added to the blood stream via the thoracic duct (Reddy et al., 2008).

1.1.3.12   ABSORPTION OF WATER AND ELECTROLYTES.

Water and electrolytes may cross intestinal epithelial cells by either cellular or par cellular routes. Tight junctions attach the epithelial cells to one another at the luminal membrane .The permeability of the tight junctions varies with the type of epithelium. ‘A tight’ (impermeable) epithelium are the small intestine and gallbladder (Linda, 1998).

Na+ moves into the intestinal cells, across the luminal membrane, and down its electrochemical gradient by passive diffusion through Na+ channel, Na+ –glucose or Na+- amino acid co-transport system.

Na+ is pumped out of the cell against its electrochemical gradient by the Na+ K+ channel pump in the basolateral membrane. CL- absorption accompanies Na+ absorption throughout the GIT, by Na+-CL- Co-transport and CL- HCo3 exchange (Reddy et al., 2008).

Absorption of water is secondary to solute absorption and iso-osmotic in the small intestine and gallbladder. In the colon, water permeability is much lower than in the small intestine, and faces may be hypertonic (Linda 1998).

 1.1.4 GIT MOTILITY

Contractile tissue of the GIT is almost exclusively unitary smooth muscle, with the exception of the pharynx, upper one third of the esophagus, and external anal sphincter, all of which are striated muscle .GIT motility is meant for the passage of ingested food from oral cavity towards the anus. Only in the first part of GIT i.e. oral cavity, the movements are initiated voluntary and are purely under the control of somatic nerves. The rest of the GIT exhibits different types of spontaneous movements, depending on the region and are, modified by autonomic nerve fibers in association with the intramural nerve plexus and local hormone thus neural and hormonal mechanisms influence GIT motility. (Fox et al., 1998).

1.1.4.1   GASTRIC MOTILITY

The movements that occur in the stomach include periodic movements of empty stomach, filling of stomach digestive peristalsis and emptying of stomach. Sinusoidal basal electric rhythm (BER) arising from the stomach or small intestines causes the periodic movements of the empty stomach, this is referred to as “migrating myo-electric complex (MMC) (Linda, 1998). Motilin initiates MMC, substance p’ somatostalin and neurotensin influence MMC.

MMC is considered to be the “house keeper” of the stomach and the small intestine and clears off any food debris that remains after the previous meal. The frequency of the BER in the stomach decides the rate of gastric peristalsis and the spike potentials decide the intensity of the peristaltic wave.

Linda, 1998 reported that the BER co-ordinates gastric peristalsis. The rate of gastric emptying is fastest when the stomach contents are isotonic. If the stomach contents are hypertonic or hypotonic, gastric emptying is slowed (Linda, 1998). Fats inhibits gastric emptying i.e., increases gastric emptying time by stimulating the release of CCK. H+ in the duodenum inhibits gastric emptying via direct neural reflexes. It receptors in the duodenum relay information to the gastric smooth muscles via inter neurons in the gastro intestinal plexuses (Linda, 1998).

1.1.4.2    SMALL AND LARGE INTESTINAL MOTILITY.

As in the stomach, slow waves set the BER, which occurs at a frequency of 12waves/min. Action potentials occur on top of the slow waves and lead to contractions. Current evidence suggests that the slow waves are produced by unique cells, often associated with autonomic nerve ending. However, these pace maker cells are neither neurons nor smooth muscles cells, they are the cells identified histological as the intestinal cells of cajal. (Long et al., 2004).

Both segmentation and Peristaltic contractions occurs. Ideally, peristalsis occurs after digestion and absorption have taken place. The peristaltic reflex is highly co-ordinate by the enteric nervous system.

Ordog, 2008 reported that gastro ileac reflex is mediated by the extrinsic ANS and possibly by gastrin. The presence of food in the stomach triggers increase peristalsis in the ileum and relaxation of the ileocecal sphincter. As a result, the intestinal contents are delivered to the large intestine. Haustra or sac like, segments appear after contractions of the large intestine. Fecal material moves from the cecum to the colon to the rectal (by mass movement) and then to the anal canal. The presence of food in the stomach increases the motility of the colon/gastric colic reflex and increases the frequency of mass movements.

1.1.5 THE LIVER AND ENERGY METABOLISM.

The liver is one of the largest organs in the body. It is uniquely situated to process and distribute dietary nutrients because the venous drainage of the gut and pancreas passes through the hepatic vein before entering to the general circulation. It has many metabolic functions. It converts the nutrients in our diets into substances, and supplies cells with them when needed. It also takes up toxic substances and converts them into harmless substances or make sure they are release from the body. During the absorptive period, the liver takes up carbohydrates, lipids and most amino acids. These nutrients are then metabolized, stored, or rooted to other tissues. (Cheng et al., 2008).

Hepatic metabolism of glucose is increased by the following mechanisms, increased phosphorylation of glucose, increased glucose synthesis, Increased activity of the hexose monophosphate pathway (HMP), increased glycolysis and decreased gluconeogenesis (Pamela et al.,2008).

The liver response to high blood glucose levels by increasing the phosphorylation of glucose by glucokinase, which has a high Km for glucose. Glucose uptake by the hepatocytes is not rate limiting because of the abundance of insulin-independent GLUT-2 glucose transporters. The liver is the primary tissue for de novo synthesis of fatty acids. This process is favored by substrates (acetyl-co A and NADPH derived from the metabolism of glucose) availability, and by the activation of acetyl-coA carboxylase, both by dephosphorylation and by the presence of its allosteric activator, and citrate. There is also increased synthesis of tri acylglyceriols (TAG). The liver package TAGs into very-low-density lipoprotein (VLDL) particles that are secreted into the blood for use by extra hepatic tissues particularly adipose and muscle tissue (Ketbamma et al., 1990).

The body cannot store protein in the same way that it maintains glycogen or TAG reserve. However, a transient increase in the synthesis of hepatic proteins does occur in the absorptive state. The surplus amino acids are not stored but are either released into the blood for all tissues to use in protein synthesis or are deaminated, with the resulting carbon skeletons being degraded by the liver to pyruvate, acetyl/COA, or TAG cycle intermediates (Panel et al., 2008).

Apart from nutrient metabolism the liver also functions in the metabolism and excretion of exogenous (drugs, alcohol, copper) and endogenous (bilirubin, hormone) materials.

The liver stores vitamins (A, B12, K and Foliate) and minerals (iron). With the help of vitamin K, the liver also produces proteins that are important in blood clotting. It also synthesizes the following: albumin, coagulation factors and inhibitors (except von wile brand factor), complement factors, transferrin, heptoglobin, caeruloplasmin,protease inhibitors (XI-antitrypsin) & fetoprotein. The liver is a vital organ that plays a major rule in human metabolism. Because of its peculiar anatomy and physiology, it is extremely susceptible to the influence of a wide variety of injurious agents including infections and toxins. Liver damage may also result from immunologic and chemical processes that characterize various systemic disorders.

Liver disease has a worldwide distribution (WHO, 2008). Very often, affected individuals are asymptomatic for a long period of time, making it very difficult to generate accurate incidence and prevalence data in the general population. The pattern of liver disease varies in different geographical location. In the united states between 1988 and 2008 except non-alcoholic fatty liver disease, which increased steady and is expected to continue to contribute substantially to the burden of chronic liver disease (DLD) because of increasing rates of obesity (Bojuwoye et al.,2010). Viral hepatitis remains a leading cause of morbidity and mortality affecting millions of individuals worldwide including Nigeria. According to the world health organization (WHO), 2 billion people have been infected with the hepatitis B virus (HBV). In addition, it has been estimated that up to 3% of the world’s population have been infected with hepatitis C virus (HCV) of which 170 million people are chronically infected (WHO, 2008).

Cancer of the liver (Hepatocellular carcinoma (HCC) is a major cause of death by cancer worldwide, accounting for over half a million deaths per year. In men HCC is the fifth most frequently diagnosed cancer worldwide, and also is the second leading cause of death in theworld. In most of Africa, and indeed Nigeria, liver disease is an important cause of morbidity and mortality. Cirrhosis and primary liver cancer account for two-thirds of liver disease related death in Africa including Nigeria. The major risk factors are hepatics B virus infection, alcohol consumption, use of herbs and root, cigarette smoking, family history of liver diseases and hepatitis C virus infection. (Robert et al., 2010).

 1.1.6 LIVER FUNCTION PARAMETERS.

Hepatocytes contain many enzymes which maybe released into the blood during pathological processes. Measurement of their activity in the blood may give evidence of hepatobiliary disease. In practice, maximal information is obtained by measuring the activity of relatively few enzymes. None of the enzymes is specific to the liver and alternative origins should be considered particularly where abnormalities have been found incidentally in patient with no clinical evidence of liver disease (Finlayson et al., 1998).

Alanine amino transferase (ALT) a cytoplasm enzyme and aspartate amino transferase (AST) present both in cytoplasm and mitochondria are the two important amino transferases.

Normal plasma contains low activities of both enzymes, the source of which is unknown.

Neither enzyme is specific to the liver, but ALT occurs in much higher concentration in the liver than elsewhere and consequently increased serum ALT activity reflects hepatic damage more specifically. ALT and AST are liberated into the blood whenever liver cells are damaged and increased plasma enzymes activity is a sensitive index of hepatic damage.

Alkaline phosphatase (ALP) is situated principally in the canalicular and sinusoidal membranes of liver cells. Blood normally contains alkaline phoshatase derived mainly from bone and liver, and in pregnancy additional activity of placenta origin is found. (Finlayson et al., 1998).

About a fifth of people have plasma alkaline phosphatase of intestinal origin and its activity may increase after meals. When hepatocytes are damaged, little alkaline phosphatase is liberated into the blood, most probably coming from cell which are killed. Consequently, plasma alkaline phosphatase activity does not usually raise more than two fold in acute or chronic hepatocellular disease (Edwards et al., 1995).

In billiary tract obstruction at any level new alkaline phosphatase activity is therefore the main indicator of billiary obstruction though it provides no information about the site of that obstruction. Gamma glutamyl transpeptidase (GGT) is also a microsomal enzyme found in hepatocytes, biliary epithelial cells, renal tubules, pancreas, and intestines .It helps in glutathione metabolism by transporting peptides across the cell membrane. In acute viral hepatitis, the GGT level can peak at 2^nd and 3^rd week of illness, and remained, elevated in 30% of the hepatitis C patient. When GGT level is elevated, the triglyceride level is elevated also.  GGT is also elevated in 50% of the patients with alcoholism and non-alcoholism liver diseases (Linda, 1998).

Albumin is a protein made specifically by the liver, and can be decreased in chronic liver disease, such as cirrhosis of the liver. It is also decreased in nephritic syndrome, where it is lost through the urine. The consequence of low albumin can be edema since the intravascular oncotic pressure becomes lower than the extra vascular space (Edwards et al., 1995).

1.2     STATEMENT OF RESEARCH PROBLEMS

Recent epidemiological studies have established that there is high prevalence of gastrointestinal diseases including ulcer and diarrhea globally. These have been associated with impaired quality of life, especially poverty. Many ulcer disease patients suffer from upper and lower gastrointestinal (Gl) symptoms (Bauer et al., 2002).The complications of ulcer diseases affect almost all organ systems of the body. The gastrointestinal system is of special concern, since it is known to play a crucial role in the pathogenesis of many of the complications of ulcer diseases. Thus, any treatment or prevention modality, design, imperatively should assess its effect on the gastro intestinal system. Current treatments and managements drugs for these diseases have either been ineffective or too expensive to buy (Bauer et al., 2002).Not much is known about the use of P.guajava leaf in management of ulcer and other GIT disorders.  Hence, the need for this study.

 1.3 AIM OF THIS STUDY

The aim of this research work is to evaluate the liver modulatory function and gastro protective effects of P. guajava leaf extract in experimental rat models.

 1.4     OBJECTIVES OF THIS STUDY

The main objectives of this study are to evaluate the following;

1. Acute toxicity effect of P.guajava leaf
2. To assess liver enzymes of the psidium guajava leaf treated qualify rats
3. To assess the anti motility effect of P.guajava leaf, both invitro and in vivo.
4. To assess the gastro protective effect of p. guajava leaf on gastric acid secretion.
5. To assess the effect of P. guajava leaf on extractible mucus.
6. To assess the gastro protective effect of P. guajava leaf (ulcer score)
7. To assess the total pepsin activity and total acidity of P. guajava.
8. Histology of the isolated rats stomach.

1.5     JUSTIFICATION OF THE STUDY

The near pandemic proportion of the ulcer incidence has called for a concerted effort to explore and formulate the commonest treatment and preventive principle. More so, the current treatment modalities of ulcer and other GIT disorders still leaves much to be desired, in terms of efficacy, side effects, convenient, accessibility, affordability and even availability.

The reduction in the quality of life as a sequence of GIT diseases is very costly in terms of social-economic implications. The morbidity and mortality associated with complications of GIT diseases is on the increase (Bauer et al., 2002).

The gastrointestinal system plays a crucial role in the metabolism and regulation of body fuel. Since the problems with GIT disorders usually disarrange the fuel metabolism, any claim on thetherapeutic provinces of any agent cannot be complete without assessing its effect on the gastro-intestinal system. . The ability of the extract of p. guajava leaf to ameliorate ulcer induced GI function derangement will be a further proof of its efficacy as an anti ulcer agent.

This justified the imperative need for this study on psidium guajava (P. guajava) leaf.

1.6     SCOPE OF THE STUDY

This study will involves an assay on liver modulatory functions and gastro-protective effects of normal Wister rats administered with the aqueous leaf extract of P. guajava for three weeks and comparing the results with the control group of five Wister rats feeds only on normal rats feed and distilled water for the same period of time .Also, comparing the Omeprazole treated rats and P.guajava leaf extracts treated rats after induction of ulcer with 20% of alcohol

 1.7     SIGINFICANT OF THE STUDY

The finding of this study will be to the benefit of the society, considering that the use of herbal medicine plays an important role in the treatment of diseases today.  The greater demand for the use of herbal medicine in the society today justifies the need for more effective life changing ulcers treatment approaches.    Thus, hospital that applies the recommended approach derived from the result of this study will be able to treat their ulcer patients better. This study will also help the researcher to uncover critical areas in the treatment process that many researchers were not able to explore .Hence; a new method of ulcer treatment may be arrived at

1.7 APPORACH TO STUDY

Gastrointestinal disorder relies on the existence of acid and peptic activity in gastric juice with breakdown of the mucosal defense system.  Its prevalence being attributed due to a variance between the offensive (acid, pepsin (helicobacter pylori). The current drug remedial modalities available for peptic ulcer (Pylori) and defensive factors (prostaglandins, nitricoxide) is employed with proton pump inhibitors (Omeprazole, bicarbonate secretion, gastric mucus, rabeprazole, etc, and histamine receptor blockers mucosal cell factors).

The common factors responsible for causing peptic ulcer are misoprostol, antacids, helicobacter pylori (a gram negative rod, accountable for trisilicate combination), and antibiotics (Clarithromycin). Potentially beneficial but many of them have B. Cell lymphoma. (Konturek et al., 2003).

Incessant efforts for searching of new novel anti ulcer or duodenal ulceration manifested with erosions, hemorrhage moiety are on progress intent to have fewer side effects with perforation and psychological stress. Hence, this leads to the search of alternative drugs probably of plant origin which can inhibit the production of acid in the stomach, thereby gaining activity in the treatment of peptic ulcer diseases.

1.9    RESEARCH HYPOTHESIS

It is hypothesized that, over the past decades P. Guajava has become increasingly accepted as one of the medicinal herbs used in the treatment of many gastro –intestinal diseases, including ulcer and diarrhea.

Pages:  130

Category: Project

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Chapters: 1-5                                          

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Material contains Table of Content, Abstract and References.