“CHARACTERIZATION OF Hibiscus rosa-sinensis Linn LEAVES AQUEOUS EXTRACT ON THE

ANTIPYRETIC ACTIVITY”

SAIFUL IRWAN BIN ZUBAIRI

RESEARCH OFFICER (QA41)CHEMICAL ENGINEERING PILOT PLANT

(CEPP)

UNIVERSITI TEKNOLOGI MALAYSIA

UTM SKUDAI, JOHOR e-mail: saiful-z@utm.my

Hibiscus rosa-sinensis leaves

Skim Latihan Akademik Bumiputera (SLAB): KURSUS PENYELIDIKAN 2006Bilik Mesyuarat Fakulti, FKKKSA, 07 July 2006

INTRODUCTION

Fever or pyrexia is an adaptive mechanism of the body to react against pathogens The core body temperature is elevated to levels which denatures, weaken or kill the pathogen Unfortunately, there are circumstances when fever accelerates to temperature where the body is at risk of devastating its own cells and should be brought under control with suppressive medication

TYPES OF FEVER

Pyrexia can be classed as low grade (38 °C to 39 °C), moderate (39 °C to 40 °C) or high grade (more than 40 °C) depending on how much the body temperature has deviated from normal

FEVER BENEFICIAL

Fever may be of benefit and is part of the body's response to a disease However, if the fever goes above 42 °C (107.6 °F), then it may cause significant tissue damage and

would most certainly be harmful. This is termed as hyperpyrexia

NORMAL TREATMENT Drugs that reduce fever are known as antipyretics Common antipyretics are acetaminophen, also called paracetamol and Non Steroidal Anti- inflammatory Drug (NSAIDs) such as ibuprofen A popular household remedy is soaking a cloth in cold water and placing it on the patient's forehead

Nowadays, the concern is on the modern antipyretic drugs which associated with the hepatotoxicity problems

For that reason, Hibiscus rosa-sinensis Linn leaves is used traditionally as an antipyretic remedy (Edward, 1999)

This is because, the aqueous extract of Hibiscus rosa-sinensis Linn leaves are comprised with -sitosterol, taraxeryl acetate, flavonoids compound and high in calcium and phosphorus content (Duke and Ayensu, 1995)

These bio-active compounds, minerals and vitamins that available in the extracts are assumed to lower down the temperature and acts as a coolant to the fever patient

Since there is not scientific research on this particular plant species as regards to the antipyretic activity, this research could be a new BREAKTHROUGH or ALTERNATIVE REMEDY to the chemically synthesized ANTIPYRETIC DRUG in which posse major side effects especially associated with hepatic organ

Hence, the synergistic of these compounds could be contributes to the antipyretic activity

WHY CHOOSE THE CONVENTIONAL WAY TO THREAT FEVER?

WHAT IS -SITOSTEROL?

-sitosterol is a major bio-active compound of phytosterol group in the higher plants

-sitosterol, with an empirical of C29H50O, is a sterol compounds with a molecular weight of 414 g/mol

-sitosterol is not synthesized endogenously in mammals with minimal intestinal absorption

-sitosterol has been shown to have anti-inflammatory and antipyretic properties and a study on animal model have found that antipyretic effects of -sitosterol was comparable to that of aspirin (Gupta et al., 1980)

Moreover, there have been numerous clinical and animal studies that have established the benefits of -sitosterol such as immune system support (especially during stress)-relieves allergies-reduces cancer risk (prostate, breast and colon)-analgesic properties-relieves symptoms of enlarge prostate

Molecular structure of -sitosterol

POSTULATE

An aqueous extract of Hibiscus rosa-sinensis is used as an acute dermal and acute oral to treat the elevation of body temperature

above normal temperature (i.e.: 98.4 °F or 37 °C)

PROBLEM STATEMENT/RESEARCH PROBLEM/QUESTION

HOW does -sitosterol, flavonoids compound, minerals and vitamins in the aqueous extract of Hibiscus rosa-sinensis affect the body immune system simultaneously alleviate the elevation of body temperature?

HYPOTHESIS

-sitosterol, flavonoids compound, minerals and vitamins in the aqueous extract of Hibiscus rosa-sinensis acts as an antipyretic, antioxidant and

enhancer of body immune system by producing a SUFFICIENT amount of white blood cells (leukocytes: WBC count/mm3) to fight the infections of

pyrogens simultaneously alleviate the elevation of body temperature

Literature review on the current development of conventional (ethno-botanical studies) and un-conventional antipyretic drugs

To identify and analyze the major bio-active constituents, trace minerals and vitamins in the extract of Hibiscus rosa-sinensis

To study the effect of major bio-active constituents in the extract of Hibiscus rosa-sinensis on the antioxidant and antimicrobial activity

To study the effect of major bio-active constituents in the extract of Hibiscus rosa-sinensis on the white blood cells count (WBC count/mm3) and antipyretic activity

To study the correlation between the antipyretic activity with the major bio-active constituents in the extract of Hibiscus rosa-sinensis and white blood cells count (WBC count/mm3)

Thesis write-up and publications

To investigate the efficacy of bio-active constituents, trace minerals and vitamins in the aqueous extract of Hibiscus rosa-sinensis Linn leaves on the antipyretic activity

RESEARCH SCOPES

RESEARCH OBJECTIVE

Research scope/semester S1 S2 S3 S4 S5 S6

1. Literature review (conventional and unconventional antipyretic drugs)

2. To identify and analyze the major bio-active constituents, trace minerals and vitamins

3. To study the effect of major bio-active constituents on the antioxidant and antibacterial activity

4. To study the effect of major bio-active constituents on the white blood cells count (WBC count/mm3) and antipyretic activity

5. To study the correlation between the antipyretic activity with the major bio-active constituents and white blood cells count (WBC count/mm3)

6. Thesis write-up and publications

RESEARCH PLANNING AND EXECUTION

Publications: International Scientific Journal

(A) Pretreatment of the Hibiscus fresh leaves:Forced with air oven at 30 0C (3 hours) and ground into fine powder (particles size of 0.5 mm )

(B) Extraction of the Hibiscus ground dried leaves:

Microscale batch solid-liquid Pyrex extractor: = 5.0 mlg-1, t = 40 0C for 24 hours

(C) Physico-chemical characterization:Relative density (20d20) and refractive index (20nD)

(D) Nutritional composition and proximate analysis: (LIQUID EXTRACT)a) Identification of trace minerals: Induced Coupled Plasma-mass spectrometry (ICP-MS)

b) Proximate analysis (Ash, Fat, Carbohydrate and Vitamin A, B, C: AOAC Method)

(E) Antioxidant test (LIQUID EXTRACT):Radical scavenging activity using 1,1-diphenyl-2-picrylhydrazyl (DPPH) method

(F) Microbiology test - Agar dilution method (SOLID EXTRACT):

Fungal colony of Cladosporium sphaerospermum

(G) Antipyretic test (LIQUID AND SOLID EXTRACT): (1) An ED50 value calculated as the dose of Hibiscus extract solution required to decrease pyresis by 50 %

(2) Acute Dermal Heat Transfer Assessment (Q = Joules) - Newtonian method/Cooling method

(H) Blood cells count (count/mm3): Complete blood count (RBC + WBC) and leukocytes content

PROCESS FLOW AND RESEARCH METHODOLOGY

Lyophilization process

HPLC: Bio-active compounds and flavonoids content

EXPECTED RESULTS Mechanism of -sitosterol, flavonoids compound, trace minerals and vitamins in the aqueous

extract of Hibiscus rosa-sinensis Linn leaves on the antibody immunization system, antioxidant, antimicrobial and antipyretic activity will be observed significantly due to its well known TRADITIONAL USAGE to treat fever and as a food supplementary

-sitosterol is expected to be the major contribution of all constituents in the extract due to the fact that plant sterol appears to modulate the immune function by controlling the production of inflammatory cytokines that relieve the pain activity

This phytosterol compound is also expected to produce high inhibitory on the targeted microorganisms given that phytochemicals which consist with antioxidant and anti-inflammatory properties will also produced a significant effect on the antimicrobial activity

Whilst flavonoids compound (e.g.: kaempferol, rutin, catechin, luteolin and quercetin) are expected to yield significantly in the aqueous extract in which these compounds will scavenged the pre-radical molecules (oxidation agent) that usually found abundantly in the body system during the demotion of body immunization system

The macro mineral (e.g.: calcium, potassium, phosphorus and sodium) are expected to increase the hormones activity to the hypothalamus system function by regulating the high body temperature to normal temperature (negative feed back mechanism)

As for the pyrexia effect of the Hibiscus extract, it is expected to produce high effective dose (ED50) values to decrease the pyrexia level of rat induced fever (↑ ED50; ↓ pyrexia level)

The higher effective dose (ED50) indicates the higher of changes in mean dorsal temperature lowering produced by the Hibiscus extracts treatment versus the non treatment animal of an induced fever

CONCLUSION AND RECOMMENDATION

From an extensive literature reviews study, understanding the mechanism of plant sterol, flavonoids compound, minerals and vitamins that existed abundantly in the particular plants or herbs as a potential antipyretic drug and body immunization system enhancer is an important innovative breakthrough and alternative to allopathic remedy (e.g.: tylenol, aspirin, acetaminophen or paracetamol) in the future to come

The study can be a viable references and the configuration of the research can be a useful tools for developing on other potential plant that posses a specific therapeutic action

**THE END**

THANK YOU FOR YOUR KIND ATTENTION

QUESTION AND ANSWER SESSION