ABSTRACT
In vitro assessment of chemical and medicinal properties of some snail slime and honey samples from different vegetation zones of Nigeria was performed. The study was carried out in four vegetation zones, namely: Mangrove (Eleme-Rivers State), Rainforest (Umudike, Abia State), Derived Savanna Zone (Ezzamgbo-Ebonyi State) and Central Guinea Savanna (Jos North Plateau State) during rainy season. Honey samples were collected by purchase directly from modern bee-keeping farmers in each location. Slime from four snail species sourced from the rainforest zone was employed namely Achachatina maginata, Achatina achatina, Limicolaria flamulata and Limicolaria martensis. The honey and snail slime were both analysed for their physico-chemical properties. They were also combined together at different ratios of honey to slime and then applied to some bacteria and fungi samples and the zones of inhibition were observed. Chromatographic analysis of the honey was also done. Results showed that honey from Central Guinea Savanna Vegetation Zone yielded the highest total soluble solids (62.81mg/l), while that of derived savanna was the least (68.87mg/l) which in many cases is a measure of the sugar content of the honey. The honey sample with the highest proportion of vitamins was from the rainforest zone (6.18mg/l) and this came from vitamin C, while the lowest vitamins (0.02mg/l) and it came from Vitamin B2. In the physico-chemical analysis of the snail slimes, it was observed that A. achatina slime had the highest in flavonoid content (0.17), L. martensis, the highest in phenol content (0.005), while A. maginata has the highest alkaloid content (0.028). Heavy metal content was highest in A. achatina and lowest in L. martensis for all the tested heavy metals. A. maginata also had the least acidity (3.86), highest in L. flamulata (3.64). In honey and snail slime combination at different ratios against fungi and bacteria samples zones of inhibition was observed. It was also observed in the control experiment (antibiotics was added). In the results obtained honey and snail slime were found to have great antimicrobial effect. Based on the results of the study, it was concluded that honey from central Guinea Savana zone yielded the best efficacy in terms of physico-chemical and medicinal properties. Farmers, cosmeticians, pharmaceutical industries and traditional healers are therefore advised to patronize honey from this ecological zone.
TABLE OF CONTENTS
Title Page i
Declaration ii
Certification iii
Dedication iv
Acknowledgement v
Table of Contents vi
List of Tables viii
List of Figures ix
List of Plates x
Abstract xi
CHAPTER 1: INTRODUCTION
1.1 Background
of the Study 1
1.2 Statement
of Problem 4
1.3 Aim
of the Study 4
1.4 Objectives
of the Study 4
1.5 Justification
of the Study 5
CHAPTER 2: LITERATURE REVIEW
2.1 Healing
Properties of Honey 6
2.2 Use
of Honey as Wound Dressing 7
2.3 Active
Healing Properties in honey 8
2.4 Nitric
Acid 9
2.5 Prostaglandins 10
2.6 Antibody
Production 10
2.7 Economic
Importance of Honey 11
2.8 Healing
properties of Snail Slime 11
2.9 Active
properties in Snail Slime 12
2.9.1 Cosmetic benefit of snail slime 13
2.9.2 Beneficial effect of snail slime on the skin 14
CHAPTER 3: MATERIALS AND METHODS
3.1 Study
Area 15
3.1.1 Rainforest
vegetation 15
3.1.2 Derived
savanna (Dr. S. Egwu’s Farm in Ohaukwu LGA, Ebonyi State) 18
3.1.3 Central
guinea savanna zone (Jos-North, Plateau State) 20
3.1.4 Mangroove
vegetation zone (Eleme LGA Ogoni-Rivers State) 24
3.2 Sample
Collections 25
3.2.1 Honey
collection 25
3.2.2 Snail
slime collection 25
3.3 Laboratory
Analysis 25
3.3.1 Total soluble solid (TSS) Determination 25
3.3.2 Refractive index (RI) 26
3.3.3 Density 26
3.3.4
pH determination 26
3.3.5 Determination
of vitamins 27
3.3.5.1 Vitamin A 27
3.3.5.2 Vitamin B1 27
3.3.5.3 Vitamin B2 28
3.3.5.4 Vitamin B3 29
3.3.5.5 Vitamin C
29
3.3.6 Heavy
metals 30
3.3.7 Phytochemicals 30
3.3.7.1 Tannins
30
3.3.7.2 Alkaloids 31
3.3.7.3 Saponin 31
3.3.8 Microbial
analysis 32
3.3.8.1 Source of test organisms 32
3.3.8.2 Sample preparations and extraction 32
3.3.8.3 Standardization of bacteria cell suspension 33
3.3.8.4 Antibacterial susceptibility assay 33
3.3.8.4.1 Agar diffusion method 33
3.3.8.4.2 Disc diffusion
method 33
3.3.9 Minerals 33
3.3.9.1 Digestion and analysis for minerals 33
3.3.9.2 Preparation of standards for analysis of
minerals in samples 34
3.3.9.3 Macro nutrients 34
3.3.9.3.1 Calcium 34
3.3.9.3.2
Phosphorous 35
3.3.9.3.3
Potassium 36
3.4 Statistical
Analysis 36
CHAPTER 4: RESULTS AND DISCUSSION
4.1 Physiochemical Analysis of Honey Samples 42
4.1.1 The physiochemical analysis of honey samples
from four vegetation
zones 42
4.1.2 Vitamins 45
4.1.3 Phytochemical screening of honey samples 47
4.1.4 Mineral contents of screened honey samples 49
4.2 Physico-chemical properties of snail
slime 51
4.2.1 Phytochemical properties of snail slime
samples assessed 51
4.2.2 Heavy metal content of snail slimes 53
4.2.3 Physical properties of sampled snail slimes 55
4.3 Antifungal and Antibacterial properties
of snail 57
CHAPTER 5: CONCLUSION AND RECOMMENDATIONS
5.1 Conclusion 70
5.2 Recommendations 70
REFERENCES 72
APPENDICES 82
LIST OF TABLES
4.1: Physiochemical parameters of
honey samples collected 44
4.2: Vitamin contents of honey
samples 46
4.3:
Totals of phytochemical
contents 48
4.4:
Mineral contents of honey
samples 50
4.5: Phytochemical contents of
snail slime accessed 52
4.6: Heavy metal content of snail
slime assessed 54
4.7: Physical properties of snail
slimes samples 56
4.8: Zone of inhibition test with
100% slime of A. marginata (fungal) 59
4.9: Zone of inhibition
test with 100% slime of A. achatina (fungal) 59
4.10: Zone of inhibition
test with 100% slime of L. flammulata
(fungal) 60
4.11: Zone of inhibition
test with 100% slime of L. martensis (fungal) 60
4.12: Zone of inhibition
test with 100% honey from mangrove
Vegetation zone (Fungal) 62
4.13: Zone of inhibition test with 100%
honey from rain forest vegetation
zone (fungal) 62
4.14: Zone of inhibition test with 100%
honey from derived savanna vegetation
zone (Fungal) 63
4.15: Zone of inhibition test with 100%
honey from central guinea
savanna
vegetation zone (fungal) 63
4.16: Zone of inhibition test with 100%
honey from mangrove
vegetation zone (Bacteria) 65
4.17: Zone of inhibition test with 100%
honey from rainforest
vegetation
zone (Bacteria) 65
4.18: Zone of inhibition test with 100%
honey from derived savanna vegetation
zone (Bacteria) 66
4.19: Zone of inhibition test with 100%
honey from central guinea
savanna
vegetation zone (Bacteria) 66
4.20: Zone of inhibition test with 100%
slime from A. maginata
(Bacteria) 68
4.21: Zone of inhibition test with 100%
slime from A. achatina
(Bacteria) 68
4.22: Zone of inhibition test with 100%
slime from L. flammulata (Bacteria) 69
4.23: Zone of inhibition
test with 100% slime from L. martensis
(Bacteria) 69
LIST
OF FIGURES
1: Map
showing rainforest vegetation (Umudike in Ikwuano
LGA,
Abia State) 17
2: Map showing the study
location in Ebonyi State (Ezzamgbo in Ohaukuwu
LGA, Ebonyi State) 19
3: Map showing study area
(Jos-North LGA, Plateau State) 22
4: Map
of Rivers State showing the study area (Eleme) 22
LIST OF PLATES
1: Some snail species used
for the study 37
2: Nutrient medium
containing ethanol extract of honey and snail
slime in different
combinations. Ready to be analyzed for
antimicrobial activity 38
3: Checking the cultured
nutrient media for zones of inhibitions 39
4: Appearance of the nutrient media contanining ethanol extract
of
honey and snail slime in various combinations after
culture 40
5: Getting the snail slime samples ready for physico-chemical
analysis 41
CHAPTER 1
INTRODUCTION
1.1 BACKGROUND
OF THE STUDY
Honey is a natural sweetener and flavourful
product ingested for its elevated healthful value and for its consequences on
human health with inhibitive, bacteriostatic, anti-inflammatory and
anti-microbial elements, as well as lesion and sunburn curing results
(Alvarez-Suareaz, et al., 2013). The Codex
Alimentarious Commission (1989) explained honey as the unadulterated sweet
substance produced by honeybees from the necter of flower. The dominant constituents
of honey include sugars such as fructose, glucose, sucrose, maltose as well as trisaccharide
sugars. Besides sugars, honey contains an array of chemical nutrients namely
protein, vitamins, fats, enzymes, minerals, amino acids, volatile aromatic
substances etc (Ahmad et al., 2003).
Simple sugars like glucose (31%) and fructose
(38%), are the vital constituents in honeys, and proteins, organic acids, minerals,
enzymes, vitamins, aromatic compounds. phenolic compounds, free amino acids and
carotenoids form the minor components (Alvarez-Suaez et al., 2010; The National Honey Board, 2017; Bogdanov et al., 2008).
Honey has been reviewed to contain more than
500 active components that are useful and mostly adopted as herbal medicines.
These constituents bind to the anti-bacterial, anti-oxidant, anti-inflammatory,
anti-browning, anti-allergic, anti-parasitory, anti-ulcer, anti-tumour and
anti-viral actions (Bogdanou et al.,
2008; Vinda-Martos et al., 2008).
Vitamins such as phyllochinon (F), thiamin (B1) riboflavin (B2),
niacin (B3), panthothetic acid (B5), pyridoxine (B6),
Folic acid (B9), ascorbic acid (O) and α-tocopherol (E) are present
in small amounts in honey and their contribution to the permissible day-to-day
intake is marginal (National Honey Board, 2017; Bogdanou et al., 2008).
Honey has remarkably increased height of
monosaccharaides fructose and glucose and it accommodates about 70 to 80
percent sugar which provides its sweetness, it also has sterile and
antibacterial elements. Contemporary medical inquiry has co-ordinated to elicit
usefulness of honey in incessant wound management and contending diseases (Juli
et al., 2015).
Honey is regularly consumed as an energy
food, it has simple sugars that are assimilated into blood stream in the
absence of digestion. Honey combines effectively as a sweetener in heated and chilly
drinks, honey accompanies nearly all edible materials, the liquid assimilating potentials
of honey facilitates cakes, breads, cookies as well as candies remain fresh and
longer (Caron et al., 2004).
Natural medicinal commodities have been useful
in the remedy of arrays of ailments for millennia (Baker et al., 2007). Although many have been surprised by common medicinal
steps, there is contemporarily, revival in concern in the usefulness of honey
and honey commodities by the broad public (Manyi-Loh et al., 2011)
Honey’s greatest medical capacities is its utilization
as contemporary agent in healing of wounds and dermis contaminations (Carter et al., 2010). Honey has
anti-inflammatory, antioxidant and immune supportive property, enough of the curative
features of honey are as a result to the increased sugar aggregation and the
resulting osmotic impact (Osato et al.,
1999).
Decreased pH and acidity (Gethin et al., 2008), are as a result of hydrogen
peroxide produced from the aerobic alteration of blood sugar to gluconic acid
by glucose oxidase upon dilution (Krell, 1996). Studies have pointed out that
hydrogen peroxide gets the largest credit for the medicinal advantage of honey
(White and Suber, 1964).
Snails secrete viso-elastic mucus with a sticking
and greasing potentials and facilitates the organisms to attach to, entirely different
kinds of exterior inclusive of uneven or promising antagonistic terrain. In
addition, mucus facilitates in prevention of creatures from dehydrating, contributes
to them having fairly disgusting characteristics as nutriment for prey (Anon,
2013) and is also perceived to facilitate disease healing.
Snails have been adopted sporadically as dermis
therapy since days of the ancient Greeks; Hipporates informatively suggested
the application of smashed snails to alleviate incited dermis and two decades
later, the efficacy of snail gelled waste (slime) was recognized by Chilean
snail cultivators who observed that dermis abrasions healed speedily without
traces of blister, when they handled snails for the French foods market
(Reporter, 2019). This observation resulted in the production of ‘Elicina’ a Chilean snail slime based
product. In 2010, Missha then launched super-Aqua cell renew snail cream,
claiming that its 70% snail extracts, soothes, regenerates and heals skin
(Missah, 2010).
Snail slime dependent commodities are affirmed
to be the trending mystery face fixer in U.S where they are useful in curing of
acne, lessen colouration and scurrying and combat wrinkles (Reporter, 2019).
Snail products may even be useful in orthopedics researchers at Herriot-Wart
University found that the slime of Giant African Land Snails (Archatina archatina) contains usual
crystals of calcite in the presence of harsh situations the mollusc will pull
back into its covering and manufacture outstanding aggregate of this sludge which
dehydrate and speedily solidify to establish the animal’s epiphragm – a shielding
housing created across the orifice of the shell-case when the molluscs enters
into periods of passionate relaxation (Aitken, 2010).
Snail slime possess outstanding hidden
characteristics, abundant of them have been disclosed even in ancient antiquity
and in current years, experimental scientists have shown that sludge derived medicine
can be useful in a substantial array of cures. For example, it is used in
creams to ease skin abrasions and scars, to cure respiratory diseases and
heartburn (Chiare and Joshua, 2013).
In 1959 the researcher A.P Williams investigated the collagen captured from the
physique wall of the garden snail Helix
aspersa and explored that it consists
of huge number of mucopolysaccharides elements fortified with glycerine,
glutamic acid and proline (The chemical composition of snail gelatin, 1960).
1.2 STATEMENT OF PROBLEM
Chemical composition and
medicinal value of honey and snail slime combination at different ratios have
not been ascertained. Although honey and snail slime are used in traditional medicine.
1.3 AIM OF THE STUDY
The aim of this study was to ascertain the
chemical composition and medicinal properties of snail slime and honey samples
from different vegetation zones of Nigeria.
1.4 OBJECTIVES
OF THE STUDY
The specific objectivs of
the study were:
i)
to determine the physicochemical properties of honey
and snail slime from the different vegetation zones of Nigeria.
ii)
to determine the physicochemical properties of snail
slime
iii)
to access the efficacy of the mixture of snail slime
and honey at different ratios on some pathogenic organisms.
1.5 JUSTIFICATION OF THE STUDY
Honey is both referred to as nature’s wonder,
it has been an important source of food and medicine (Marquis et al., 2016). Snail slime also has numerous benefits
especially in cell regeneration, there has been limited scientific report on curative
benefits of honey and snail slime (Carter et
al., 2010).
Therefore, this study will provide more
information on the physico-chemical and medicinal properties of honey from the
four zones: Mangrove
(Eleme-Rivers State), Rainforest (Umudike, Abia State), Derived Savanna Zone
(Ezzamgbo-Ebonyi State) and Guinea Savanna (Jos North Plateau State) when compared, as well as provide more data
to prove the effectiveness of slime from four (4) snail species in Umuahia
(Umudike).
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