The Hypoglycemic Activity of Native Carica papaya leaves extract on alloxan-induced Mus musculus domesticus

A Research Project Presented to the Department of Education (DepEd) National Capital Region (NCR) and Division of City Schools, Manila Ramon Magsaysay High School Espana Boulevard, City of Manila Morales, Yvette B. Ramos, Jonah Leigh E. Ventanilla, Ila M. September 2011

Chapter 1 Introduction

Diabetes is a chronic disease which is caused by high levels of sugar in the blood. It cannot be treated once diagnosed, but different medicines established were made to prolong life by decreasing the blood sugar level and decrease the incidence of related complications such as blindness, heart disease, kidney failure, and amputation of limbs.

Diabetes is currently the 7th leading cause of death in the Philippines. The common perception for this problem is that Filipinos are in the habit of eating sweets, and also considering the fact that the countrys staple food is rice, a starchy food item that makes diet a primary risk factor to diabetes. Mathematical modeling on projection yields that over 380 million people are expected to develop diabetes by 2025 based on International Diabetes Federation World Health Organization, to which a good percentage will be coming from the Southeast Asian countries, including the Philippines (J. P. Simbulan 2008).

On the other hand, previous studies reported the potential of dried Carica papaya leaves in decreasing glucose level. Its leaf components were found out to help produce insulin, resulting to the decrease of glucose level. Carica papaya leaves may aid possibly on diabetes mellitus (DM) type 1 where production of insulin is needed.

Background of the Study

Diabetes type 1 is caused by lack of insulin in the body. Although it is not very common, it is still a high threat to everyone because the exact cause is unknown. This type of diabetes is diagnosed in childhood.

Diabetes is not only for older people. Hospital records show cases where patients are aged 20 years old or lower. Cases being filed involve operations wherein some parts of the body are needed to be removed such as the legs; heart attack may occur; dialysis; blindness; and several others. Filipinos are not prone to such a disorder being that rice is the staple food of the country, and therefore risks to diabetes could not be avoided.

On the other hand, Carica papaya leaves have been associated in studies related to the lowering of glucose level and production of insulin. Its components were found to help produce insulin which may be helpful to DM type 1.

This brings the proponents of the project to study on the possible relation of the extract of papaya leaves to helping decrease troubles with diabetes.

Purpose of the Study

The scope of this study is to test for the feasibility of dried Carica papaya leaves extract to work on decreasing the bodys sugar level. The proponents are aiming to come up with results positively relating the leaves of papaya to diabetes.

Significance of the Study

This study would be a big benefit to those with DM type 1. Carica papaya leaves were tested to be more effective than the ordinary glucose analogues. This project may also be helpful to the country since the disorder had become common among Filipinos. Moreover, this may contribute to the field of science in terms of new discoveries.

Problem Statement

Diabetes is a metabolic disorder that needs a maintained treatment all throughout a patients years of living. The disorder is now ordinary that Filipinos are able to manage well, however the recent declaration that it is among the top ten leading causes of death in the Philippines makes it clear that diabetes is still a nuisance despite the fine management of diabetic Filipinos.

Scope and Limitation

This project is limiting its study to testing only for the potential of dried Carica papaya leaves in lowering the glucose level, as well as to proving the ability of Alloxan monohydrate to work as a glucose analogue.

Chapter 2 Theoretical Framework

Native Papaya (Carica papaya Linneus) leaves have been linked to treatments of different diseases. One of these was diabetes. Although there have been no studies conducted to prove that papaya can prevent diabetes, it was said to lower the glucose level.

Carica papaya has been known by its nutritional values. Great progress has been achieved regarding the biological activity and medicinal application of papaya. (Zunjar et al)

Tea extract from papaya leaves was reported to treat cancer, based on anecdotes of patients with advanced cancers. Significant growth inhibition on tumor cell lines was observed when the mice were treated with extract of papaya leaves. Cytotoxicity was also enhanced during the addition of the extract. Immunomodulatory genes which are classified by gene ontology analysis were also enhanced by the addition of Carica papaya extract. Active components which inhibit tumor cell growth and stimulate antitumor effects were identified. (Otsuki et al.)

Lowering of glucose level by papaya leaf extract was not affected by higher dosage. The extract reduces the hypoglycaemic activity of glimeperide and increases the hypoglycaemic activity of metformine. (Fakeye et al)

Dry seeds of mature, unripe fruit of papaya manage hepatic and renal diseases effectively It includes drug-related liver and kidney diseases. The extract of the dried seeds of mature papaya possesses protective effect against CCl4. Dosage and time affects the activity of the extract. (Adeneye et al)

Active constituents of Carica papaya, including chymopapain and papain are used in treating arthritis and disorders in the digestive system. Diabetic patients are said to take leaf extracts together with hypoglycaemic prescription drugs. (Fakeye et al)

Rats pretreated with Carica papaya extract showed significant reduction in the gastric ulcer index, as compared with alcohol treated controls. Carica papaya extract induced in rats showed protection with reduction in plasma lipid peroxidation level. Acute alcohol damage was reflected in the blood, according to biochemical analysis. Glutathione peroxidase activity was also increase. It could be concluded that Carica papaya leaves can be a good therapeutic agent against gastric ulcer and oxidative stress.

Finding out the effects of given with to Mus monohydrate extract can lower hypoglycaemic activity of mice the extract alloxan musculus domesticus artitioning and testing of the different fractions of the extract on Mus musculus domesticus Research Paradigm

Statement of the Hypotheses

This study aims to prove that Carica papaya extract can lower the glucose level of Mus musculus domesticus pretreated with alloxan monohydrate.

H1: Carica papaya leaf extract lowers the glucose level of Mus musculus domesticus since it contains components that do that function.

H2: The effectiveness of each fraction varies from one another because each is composed of different constituents.

H3: The extract causes side effects to Mus musculus domesticus because some of the chemicals used are found to be quite harmful.

Gathering papaya leaves for Testing of extract to Mus musculus domesticus extraction Observation and data gathering Extraction and partitioning of papaya leaf extract Conceptual Framework

Definition of Terms

Hypoglycemic Activity takes place when an agent, such as a food, herb or medication, causes the insulin in the blood to quickly drop

Alloxan Monohydrate - (2,4,5,6-tetraoxypyrimidine; 2,4,5,6-pyrimidinetetrone) is an oxygenated pyrimidine derivative; induced in rodents to achieve a high glucose level

Glucometer - a medical device for determining the approximate concentration of glucose in the blood

Chapter 3 Methodology

Extraction Five kilograms of native Carica papaya Linn was purchased from the National Museum of the Philippines. It was air dried for one week and cut into small pieces. Afterwards, it was percolated with methanol as the solvent for 72 hours. It was then filtered and brought to Adamson University for rotary evaporation. The extract underwent Kupchan Partitioning. Using a separatory funnel, the crude extract was mixed with chloroform to obtain two fractions of the extract: chloroform extract and the methanolic extract. Only the methanolic extract was gathered in the partitioning. After the process, there are two fractions of the extract, namely: the crude extract and the methanolic extract.

Testing There are three groups of Mus musculus domesticus, with three Mus musculus domesticus in each group. These mice has undergone acclimation period, which lasted for one week. On the first day, the baseline of the glucose level of Mus musculus domesticus was taken using a glucometer. For the next three days, all of them were given with alloxan monohydrate to obtain a high glucose level. Afterwards, the extract was given twice a day through oral gavage to determine if it can lower the glucose level. The first group, group (a) was given the crude extract. Group (b) was given the methanolic extract. Lastly, group (c), the control group, was given metformin. Observation and data gathering was made during the experiment.

Disposal The chemicals used was packed and kept secured by covering it with old newspapers. The apparatuses used were thoroughly washed, to prevent the spread of diseases.

Experimental Gathering of leaves and chemicals Flowchart

Extraction (percolation, filtration, rotary evaporation and partitioning)

Testing to Mus musculus domesticus (three mice per group)

Group A Crude extract

Group B Methanolic extract

Group C (control) Water only

Observation and Data Gathering

Disposal

Chapter 4 Results and Discussion

Extraction The papaya leaves were cut into smaller pieces, air-dried for a week, and then soaked into methanol for three days. Afterwards, it was then filtered to obtain the extract. It was then bought to Adamson University for rotary evaporation. The collected extract possesses a deep, dark green color, and smells like the leaves itself. The collected extract was then divided into two for partitioning. The first half was the crude extract itself. The second half of the extract was mixed with chloroform in order to obtain the chloroform and the methanolic extract. However, only the methanolic extract was used in the experiment. After the partitioning, there are already two kinds of extract: the crude and methanolic extract.

Testing The mice underwent acclimation period for one week. After the acclimation period, the baseline of each mice was then determined through the use of a glucometer. For three days, all of them were given alloxan monohydrate to increase the glucose level. Afterwards, the mice were given the extract to determine if it can lower the glucose level. The mice were divided into three groups: group (a) was given the crude extract. Group (b) was given the methanolic extract. Lastly, group (c), the control group, was given metformin. The extract was given twice a day, for three days. Some mice died during the period. The table shows the relative glucose level for each mice.