PHYSICOCHEMICAL AND PHYTOCHEMICAL ANALYSIS OF HONEY AND SHEA BUTTER SAMPLES AND THEIR ANTIBACTERIAL EFFECT ON STAPHYLOCOCCUS AUREUS AND KLEBSIELLA PNEUMONIAE

ABSTRACT

This research work was carried out to examine the physicochemical and phytochemical constituents of honey and Shea butter samples respectively and their antibacterial effect on Staphylococcus aureus and Klebsiella pneumoniae. The physicochemical screening of honey showed that the honey samples used have low water activity, low moisture content, low pH value below 4.0 and Ash while the phytochemical analysis of Shea butter revealed the presence of Saponnins, Tannins, Alkaloids, Steroid and Phenol. The organisms Staphylococcus aureus and Klebsiella pneumoniae were isolated and identified by standard microbiological procedures. The antibacterial screening protocol was carried out by disc diffusion method. The result obtained showed that the honey alone was able to give an inhibition zone that is up to 29mm and the Shea butter gave up to 26mm. The synergistic effect of honey and Shea butter showed higher inhibitory effect of 42mm at 100% concentration. The standard antibiotics; Ciprofloxacin and Streptomycin had the highest mean of 36mm. The synergism is hereby recommended for its high antibacterial properties for therapeutic purposes and can be used as an alternative medicine against Staphylococcus aureus and Klebsiella pneumoniae infections. It may be even more conventional to use this natural means as it has very low antibiotic resistance compared to synthetic antibiotic.

TABLE OF CONTENT

Title page                                                                                                                                i

Certification                                                                                                                            ii

Dedication                                                                                                                               iii

Acknowledgement                                                                                                                  iv

Table of Content                                                                                                                     v

List of tables                                                                                                                           vi

Abstract                                                                                                                                   x

CHAPTER ONE                                                                                                                   1

1.0 INTRODUCTION                                                                                                            1

1.1   Aims and Objectives                                                                                                 2

CHAPTER TWO                                                                                                                  3

2.0 LITERATURE REVIEW                                                                                                 3

2.1 Honey                                                                                                                               4

2.1.1 Extraction                                                                                                                      6

2.1.2 Local test for real honeys                                                                                              6

2.1.3 Classification of honey                                                                                                  7

2.1.3.1 Crystallized honey                                                                                                      7

2.1.3.2 Pasteurized Honey                                                                                                      7

2.1.3.3 Raw honey                                                                                                                  7

2.1.3.4 Ultrasonicated honey                                                                                                  7

2.1.4 Mode of action of some of the antibacterial substances in honey.                              7

2.1.4.1 High osmotic pressure                                                                                                8

2.1.4.2 Low water activity                                                                                                      8

2.1.4.3 Glucose oxidase enzyme                                                                                            8

2.1.4.4 Low pH/ acidic environment                                                                                      8

2.2 Medical Uses of Honey                                                                                                    8

2.2.1 Remedy for Diarrhea                                                                                                     8

2.2.2 Gastritis/gastric Ulcers                                                                                                  9

2.2.3 Wound healing                                                                                                              9

2.2.4 Treatment for arthritis:                                                                                                  9

2.2.5 For post-operative wound infection:                                                                             9

2.2.6 Adverse reaction of honey                                                                                             10

2.3 Shea Butter Tree                                                                                                               10

2.3.1 Extraction of Essential Oil from Shea Kernel                                                               11

2.3.2 Composition of Shea butter                                                                                                                                           12

2.3.3 Shea Butter Phenolics                                                                                                    12

2.3.4 Therapeutic uses of Shea Butter                                                                                    12

2.3.5 General uses of shea butter                                                                                            13       

2.4 Antimicrobial Agent                                                                                                         13

2.5 Antimicrobial synergy                                                                                                      14

2.6 Antimicrobial Susceptibility and Resistance                                                                   15

 2.7 An overview of the test organisms                                                                                  15

2.7.1 Staphylococcus aureus                                                                                                  15

2.7.2 Klebsiellapneumoniae                                                                                                   16

CHAPTER THREE

3.0 MATERIALS AND METHODS                                                                                                             17

3.1 Collection of test samples                                                                                                                        17

3.1.1 Collection of clinical samples                                                                                                               17

3.2 Physicochemical Analysis of Honey                                                                                                        17

3.2.1 Colour determination                                                                                                                            17

3.2.2 PH determination                                                                                                                                  17

3.2.3 Moisture content                                                                                                                                    18

3.2.4 Ash content                                                                                                                                            18

3.2.5 Water activity                                                                                                                                        18

3.3 Phytochemical Analysis of  Shea Butter                                                                  18

3.3.1 Detection of alkaloids                                                                                                                            18

3.3.2 Detection of flavonoids                                                                                                                         19

3.3.3 Detection of saponins                                                                                                                            19

3.3.4 Detection of tannins                                                                                                                               19

3.3.5 Detection of phenol and polyphenol                                                                                                     19

3.3.6 Detection of steroids                                                                        19

3.4 Preparation of media                                                                         20

3.5 Isolation and identification of bacteria from upper respiratory tract                                                     20

3.5.1 Gram staining                                                                                                                                        20

3.5.2 Indole Test                                                                                                                     21

3.5.3 Catalase Test                                                                                                                  21

3.5.4 Citrate test                                                                                     21

3.5.5 Oxidase test                                                                                                                   21

3.5.6 Coagulase test               

3.6 Antibacterial activity testing                                                                                                                    22

3.6.1 Preparation of test samples                                                                                                                    22

3.6.2 Sensitivity test                                                                                                                                       23

3.7 Synergistic antimicrobial assays                                                                                                              24

3.8 Determination of Minimum Inhibition Concentration (MIC)                                          25

3.9 Determination of Minimum Bactericidal Concentration                                                                         25

CHAPTER FOUR

4.0  RESULTS                                                                                                                   26

CHAPTER FIVE

5.0 DISCUSION, CONCLUSION AND RECOMMENDATION                                                                35

5.1 Discussion                                                                                                                                                 35

5.2 Conclusion                                                                                                                        37

5.3 Recommendation                                                                                                              37

REFERENCES                                                                                                                       38

LIST OF TABLES

CHAPTER ONE

1.0 INTRODUCTION

Antimicrobial agents are essentially important in reducing the global burden of infectious diseases. However, as resistant pathogens develop and spread, the effectiveness of antibiotics is diminished. This type of bacterial resistance to the antimicrobial agents possess a very serious threat to public health, and for all kinds of antibiotics, including the major last resort drugs, the frequencies of resistance are increasing worldwide (Levy and Marshal, 2004). Therefore, alternative antimicrobial strategies are urgently needed, and thus this situation has led to a re-evaluation of the therapeutic use of ancient remedies, such as plants based products including honey and shea butter (Mandal et al., 2010).

The use of traditional medicine to treat infections has been practiced since the origin of mankind and honey produced by Apis mellifera and shea butter produced from Vitellaria paradox are one of the old traditional medicine considered to be important in the treatment of several human ailments (Onyeagba et al., 2004). Currently, many researchers have reported the antibacterial activity of honey and found that natural unheated honey has some broad spectrum antibacterial activity  when tested against pathogenic bacteria, oral bacteria as well as food spoilage bacterial (Lusby et al., 2005). Today, traditional structure of medicine is being practiced on many accounts. Medicinal plants products such as shea butter and other antibacterial products are having great impact in the field of curing diseases and as a source of medicines for a wide variety of human ailments (Onyeagba et al., 2004). Also shea butter and honey contain some bioactive components such as phenolic, flavonoids, hydrogen peroxide, etc, these constitute their phyto components responsible for their antibacterial activities. Several up to date research work and practical experience have shown that using medicinal plants is better than allopathic drugs by being safer besides have Synergistic effect (Ifra and Ahmed, 2009).

Upper respiratory tract organisms have posed a lot of problems and discomfort on patients. The most common symptoms being cough, catarrh, nausea, etc, give a major concern to many nursing mothers and their children as it leads to nocturnal cough coupled with sleepless night.

Over the years, there have been reports on the production of more potent antibiotics e.g. third and fourth generation of cephalosporin by pharmaceutical companies, which are not readily available and expensive.

Problems of various antibiotics include, but are not limited to, low efficacy and side effect, hence the need to investigate into natural and potent antibiotics.

The invasion of pathogenic organisms has been on the rise; as a result efforts are being made to develop antimicrobial agents from natural sources for better therapeutic effects (Gills, 1992).

1.2  Aims and Objectives.

1. To ascertain the physicochemical and phytochemical components of honey and shea butter.

2. To isolate upper respiratory tract organisms; Staphylococcus aureus and Klebsiella pneumoniae from sputum, nose swab, and sneezes of patients.

3. To determine antibacterial potential of honey, shea butter, and their synergistic effects on the bacterial isolates.

4. To determine the Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) of honey and shea butter.

5. To compare the antibacterial effect of honey and shea butter and standard antibiotics.

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