EFFECTS OF TOOTHPASTE ON ORAL BACTERIAL LOAD

ABSTRACT

The effects of toothpaste on oral bacterial load were tested  using different brands of tooth paste, Colgate, Oral B pro-health, Red Close Up, Close Up Herbal, Darbur Herbal, Sensodyne, Aquafresh, Macleans, Holdent and Pepsodent. The Total Oral Bacterial Count of toothpaste (Pepsodent) has the highest oral bacterial count immediately after brushing of the teeth at the serial dilution factor of 10-6 giving 9.8x10-8cfu/ml, while the bacterial count of (Holdent) has the lowest oral bacterial count of 1.0x10-7cfu/ml immediately after brushing of teeth at the serial dilution of 10-4, because the viable count of bacterial load reduces with the time exposure. The finding should be taken into account of further research concerning the prolong use of toothpaste containing fluoride, and some toothpastes that increase bacterial load of the oral cavity.

TABLE OF CONTENTS

Title page                                                                                                                                i

Certification                                                                                                                            ii

Dedication                                                                                                                              iii

Acknowledgment                                                                                                                   iv

Table of contents                                                                                                                    v

List of tables                                                                                                                           vii

Abstract                                                                                                                                  viii

CHAPTER ONE

1.0       Introduction                                                                                                                1

1.1      Aims and objectives                                                                                                     4

1.2       Literature review                                                                                                         4

1.2.1    Definition of toothpaste                                                                                             4

1.2.2    History of toothpaste                                                                                                  5

1.2.3    Ingredients of toothpaste                                                                                            7

1.2.4    Types of toothpaste                                                                                                    9

1.3       Oral micro-flora that effects bacterial load of toothpaste                                          10

1.4       Composition of tooth paste                                                                                        11

CHAPTER TWO

2.0       Materials and method                                                                                                 16

2.1       Toothpastes and sources                                                                                             16

2.2       Collection of sample                                                                                                   16

2.3       Sterility of materials                                                                                                    16

2.4       Preparation of culture media                                                                                       16

CHAPTER THREE

3.0       Results                                                                                                                        18

CHAPTER FOUR

4.0       Discussion                                                                                                                   32

4.1       Conclusion                                                                                                                  33

REFERENCES

LIST OF TABLES

CHAPTER ONE

            1.0              INTRODUCTION

Bacteria form an important group of microorganisms found in both healthy and diseased mouth (ADA, 2010).There are 300 types of bacteria found in the mouth. One might think or suggest that the oral carvity is an easy environment for bacteria to colonize (Williams and Cummins, 2003). However, relatively few of the oral bacteria are found further along the gastro-intestinal tract. This illustrates the unique ecology of the oral carvities(Becks and WainWaright, 1993). Moreover, a bacterial accumulation on oral surfaces is a major factor in the development of most of the common dental diseases. Streptococcus mutans a bacterium in the mouth is the chief bacterium that causes plaque and may cause dental caries. Plaque is a complex biofilm found on the development of dental caries. The accumulation and development of plaques depends upon the outcome of the interactions between the adhesiveness of plaque to the tooth surfaces and physical shear forces which serve to dislodge and remove the plaque. Toothpastes and brushes are among the physical forces that removes plaque. Flouride containing toothpaste has significant effects on the initiation and progression of caries. Within these flourides containing toothpastes is a cationic agent called Chlorhexidine. Chlorhexidine has been documented for its ability to inhibit the formation of dental plaque. However, the use of chlorhexidine has a few draw backs. The first draws back is the staining of the teeth and on tongue. Another drawback is unpeasant taste (Brailsford etal., 2005).

When eating or drinking something after the use of chlorhexidine, one can experience a change in taste perception.

Triclosan, a compound commonly used for disinfection is another broad spectrum antibacterial agent manufactured specially for use in oral care (Williams and Cummins, 2003). It has been shown in vitro to be active against many of the organisms associated with plaque, gingivitis and bad breath.

Triclosan works by disrupting the bacterial cytoplasm membrane resulting in the leakage of cellular content and the death of the bacteria (Bobby and Vankesten, 1998).

It is used in most oral care ingredients and has a long history of use in consumer products. Fluoride in drinking water and toothpaste do not appear to have demonstrated effects on the compositor on dental plaque, fluid pH change and reduces lactate production following consumption of sugar (Black and Bora, 2000).

The exact mechanism underlying this inhibition is not known but fluoride has been shown to inhibit a variety of bacterial processes that are mediated (Brailsford et al., 2005). On occasion, many people experience stale or unpleasant breath upon waking in the morning. However, some people, about 25% of the population, experience halitosis or bad breath on a regular basis. The problem results from the anaerobic breakdown of protein from food and salivary debris by gram (-) negative bacteria, which generates amino acids such as cysteine  and methionine. Many different products are currently marketed that promised to provide consumers with fresh breath. It is estimated that more than one billion dollars are spent. Annually world-wide on lozenges, chewing gum, mouth rinse, toothpaste and dentrifices in an effort to resolve this condition (Black and Bora, 2000). The active agents that are incorporated into treatment forms includes surfactants, antibacterials agent, baking soda, perioxide; metal sacks herbal and natural extracts and chlorine dioxide. Contaminated toothbrushes which are regularly become contaminated with microorganisms that colonize the teeth and oral carvity. Under he usual conditions of storage, a toothbrush can therefore serve as a vector for the reintroduction potential pathogens into the oral carvity and also for the introduction of  other microbial species originating from the bathroom environment. It has reported that toothbrushes could be a source of repeated oral infection (Bowen, 1997).Significant bacteria on toothbrushes have been reported after tooth brushing, especially in patients with severe periodontitis. This can be caused by simply leaving ones toothbrushes kept in a moist environment, like that of a bathroom, retain up to 50% of herpes simplex virus types 1 after one week. An in-vitro study involving, patients who had oral inflammatory disease found that 34% required no additional therapy after they changed their toothbrush contamination (Bowen, 1997).

Some studies suggest that the general population replace toothbrushes every month or after any illness(Caldwell and Stallard, 1999).

The use of disposable toothbrushes is also a good option to reduce oral bacteria flora. Another idea to prevent the contamination of brushes is to coat the brush and the chlorhexine after each use. One study found that soaking tooth brushes for 20mins in a mouth rinse contain essential oils killed 100% of bacteria (Warren et al., 2000).Bacteria in the mouth are an issue everyone has to deal with some of the bacteria are harmful and cause plaque and bad breath. There are tooth paste and other remedies that help to kill an d prevent bacteria in peoples mouth. Calcium carbonated and silicic acid ensure through removal of plaque, which is mainly responsible for caries, paradentosis and tartar. With sea salt and minerals, in addition to extracts of amine, myrrh and yarrow toothpastes ensures a healthy bacteria flora in the mouth with food water and air.; the presence of mucosa folds interdental species gums and other places where food¸ designated epithelium and saliva are easily trapped creating favorable conditions for the reproduction of most microorganisms. Dentalcaries is localized and transmissible pathological I nfections process that results in the destruction of hard enamel tissues (Brailorsky, 1995).

Streptococcus mutans, an acidogenic and acid uric microorganisms colonizing the oral carvity is considered to be main cause of dental caries. Fluoride therapy has been the cornerstone of caries preventive strategies since the infuoridation scheme over five decade ago. Toothpaste is by far the most widespread form of fluoride usage (Black and Bora, 2000).

            1.1              AIMS AND OBJECTIVES

i.                    To isolate bacterial resident in human oral cavity

ii.                  To determine the effect (reduction in microbial load) of some commonly used toothpaste on the bacterial flora of the human oral cavity.

1.2       LITERATURE REVIEW

1.2.1    Definition of toothpaste

Toothpaste is a paste or gel dentrifice used with toothbrush as an accessory to clean and maintaining the aesthetics and health of teeth.  Tooth paste is used to promote orsl hygiene. It serves as an abrasive that aids in removing the dental plaque and food from the teeth and assists in suppressing Halitosis and deliver active ingredients (most common fluoride to help prevent tooth and gum disease (gingivitis) (Carlson et al., 1998).

Most the cleaning is achieved by mechanical action of the toothbrush and not by the toothpaste. Salt and sodium bicarbonate (Baking soda) are among materials that can be substituted for commercial toothpaste (Cheesebrough, 2000).

Toothpaste is untended to be swallowed due to the fluoride content, but is generally not veyharmful if accidentally swallowed in small amounts, however, one should seek medical attention after swallowing abnormally large amount (Davis, 1995).

1.2.2    History of toothpaste                                                                     

Early toothpaste

According to ( Davis,1995), he stated that the Greeks and then the Romans improved the recipes for toothpaste by adding abrasives such as crushed and oysters shells. In the 9^th century, the Arab musician and fashion designer Ziryab invented a types of toothpaste, which he popularized thought Islamic Spain. The exact ingredients of this toothpaste are known but it was reported to have both functional pleasant to taste. It is not known whether those early toothpaste where used aloe or were to be rubbed into the teeth with rags or were to be used with early  toothbrushes such as neem- a tree twig and miswak.

Toothpaste powder

Toothpaste powder for use with toothbrushes came into general use in the 19^th century in Britain. Most were homemade with chalk, pulverized brick or salts as ingredients (Gunsolley et al., 2006).An 1866 home encyclopedia recommended  pulverized charcoal and cautioned that many potent toothpowder that were commercially marketed did more harm than good.

Modern toothpaste

An 18^th century American and British toothpastes recipe called for burnt bread. Another formula fund this time called for dragons blood (La resin), cinnamon and burnt (ADA,  2010).In 1900,a paste made of hydrogen perioxide (N₂O₂) and baking soda as recommended for use with toothbrushes.

Pre-mixed toothpaste were first marketed in the 19^th century,  but not surpass the popularity of tooth powder until world war 1. In 1892, Doctor Washington Sheffield of New London manufactured toothpaste into a collapsible tube, Dr Sheffield Bremem dentifrice.

The original collapsible toothpaste tube was made to lead.Fluoride was first added to toothpaste in the year 1890 ‘tanagra’ containing calcium fluoride as the active ingredients , was sold by Karl GF Toellner company of Bremen, Germany based upon the early work of chemist Albert Deninger.Fluoride toothpaste developed in the 1950’s and receive the American dental approval.

Procter and gamble started a research program in the early 1940’s.procter and gamble developed a joint research project team headed by Dr Joseph Muhler at Indiana university to study new toothpaste with fluoride (Konen, 2000).

Proter and Gamble crest launched its first clinically proven fluoride  containing toothpaste. On august 1, 1969, the American dental association reported that crest has been shown to be an effective anticarvity (decay peventative) dentrifice that can be of significant value when used in a conscientiously applied program of oral hygiene and regular professional care.

Amount of fluoride in toothpaste varies from country: toothpaste in UK and Greece typically contains more fluoride than US toothpaste. In 2006, Bio-repairappeared in Europe with the first toothpaste contain synthetic hydroxylapatite as an alternative to fluoride for the demineralization and reparation of tooth enamel (MacCarty, 1995).

1.2.3                Ingredients of Toothpaste

According to Onysi et al., (1990) in addition to 20-42% water, toothpastes are derived from a variety of compounds, the three main ones are abrasives, fluoride and detergents

Abrasives

Abrasives constitute at least 50% of a typical tooth [paste. These insoluble particles help to remove plaque from the teeth. The removal of plaque and calculus helps minimize cavities and periodontal diseases . Abrasives includes aluminumhydroxide, (AL(COH)₃), hydrogen phosphates, various silica’s and zeolites and hydroxyapatite (Ca5(PO4)3OH). Abrasives helps in the removal of stains from tooth surfaces, but has not been shown to improve dental health over above the effects of the removal of plaque and calculus,

Fluoride

Fluoride in various forms is the most popular active ingredients in toothpaste to prevent  cavities.Fluoride occurs in small amount in plants and animals, some natural water sources (Perlich et al., 1995). The additionalfluoride in toothpaste has beneficial effects on the formation of dental enamel and bones, sodium fluoride (Naf) is the most common source of fluoride,  but stannout fluoride (SnF₂), Olafur(an organic salt of fluoride), and sodium monoflurophospahte (Na₂PO₃F) are also used. Most toothpaste sold in united states has the  1000 to 1100 parts per million fluoride .

Surfactants

Many although not all, toothpaste contain sodium  lauryl sulfate found in many other personal care products , as well as shampoo and is mainly a foaming agent, which enables  uniform distribution of toothpaste, improving its cleansing power or ability.

Other components include

Antibacterial agents

Triclosan, an antibacterial agent is a common toothpaste ingredient in the United Kingdom. Triclsoan or zinc chloride prevents gingivitis and according to American dental association helps reduce tartar and bad breath.

A 2006 review of clinical research concluded that there was evidence for reducing plaque and gingivitis (Perlich et al, 1995).

Flavorants

Toothpaste comes in a varietyof odours and flavour intended to encourage use of the product. Three most common flavorants are peppermint spear mint and winter green.

Toothpaste flavoured in peppermint-arise oil is popularly in the Mediterranean region. These flavours are provided by the respective oils e.g pepper mints oil. More exorticflavours include anethole arise, apricot, bubble gum, cinnamon, fennel, lavender, neem, vanilla. Lemon, orange and pine, more unusual flavours have been used e.g peanut butter, iced tea and even whisky. Unflavoured toothpaste exists.

1.2.4                Types of toothpaste

Whitening toothpaste

Many types of toothpaste make whitening contains, some of these toothpaste removes the stains not the peroxide (Quirynen et al., 2003).

Whitening toothpaste cannot alter the natural colours of the teeth or reverse discolouration by penetrating surface stains or decay. To remove surface stains, whitening toothpaste may include abrasives to gentle polish the teeth  to break down or dissolve stains.

It reduces the strength of ones teeth (the process scrapes away a protective outer layer of enamel).

Herbal and natural toothpastes

According to (Roberts, 2005) many consumers have started to switch over to natural tooth pastes to avid synthetic and artificial  flavours that are commonly found in regular toothpastes, because of the increased demand of natural  products, most of the toothpaste manufactures now produce herbal tooth paste.

This types of toothpaste does not contain dye or artificial flavours. Many herbal toothpastes do not contains fluoride or sodium lauryl sulphate. The ingredients found on natural toothpaste vary widely but often include  baking soda, aloe, eucalyptus oil, myrrh, plant extracts (strawberryextract) and essential oils. In addition of the commercially available products, it is possible to make ones’ own toothpaste using similar ingredients.

           1.3              ORAL MICRO-FLORA THAT EFFECTS BACTERIAL LOAD OF TOOTHPASTE

The oral microflora is divided into two groups and is as follows. They are permanent microflora of the oral cavity whose presences is necessary for normal functioning of the dental system, as well as the entire body.

Saprophytic micro-flora: They affect the conditions of local immune system , prevents the development of pathological conditions and support bacterial equilibrium.

Pathogenic microflora: They affect the organs of the tissue of the mouth. The microflora ideally should not or perhaps be present in the very limited quantities that do not substantially affects the oral carvities of the body. The species composition of permanent oral; microflora is normally quite stable and includes representatives of various micro-organism (bacteria). Predominant are anaerobic bacteria (Lactobacilli), bacteriods, fuzi forms, prevotelly, vellonella, spirochets and actinomycetes. Personal hygiene is at a low level or is missing altogether, the qualitative composition of bacterial flora changes. pathogenic micro-flora prevails its own quantity increases by the tens or hundred of times within a very short time (Warren et al., 2000).

1.4              COMPOSITION OF TOOTH PASTE

Humectants and binders

Humectants are used in dentifrices to prevent loss of water and subsequent hardening of the paste when it is exposed to air. The most commonly used humectants are glycerol and sorbitol. Binders are hydrophilic colloids which disperse or swell in the presence of water and are used to stabilise toothpaste formulations by preventing the separation of the solid and liquid phases (Orward, 1991). Examples of binding agents used in toothpaste include the natural gums (arabic, karaya and tragacanth), the seaweed colloids (alginates, Irish moss extract and gum carrageenan) and synthetic celluloses (carboxymethyl cellulose, hydroxyethyl cellulose), with the latter now being used increasingly for economic reasons. These substances are used routinely in the food and pharmaceutical industries and should pose a minimal health risk when used in toothpaste. However, sorbitol may cause diarrhoea in large doses as it acts as an osmotic laxative. The FAO/WHO Expert Committee on Food Additives recommends that the intake of sorbitol be limited to 150 mg/kg/day. Therefore, the use of 60-70% sorbitol gel toothpastes by small children should be supervised by parents (Sullivan, 1990).

Detergents, flavours, preservatives and colourings

Detergents lower the surface tension and therefore help loosen plaque deposits and emulsify or suspend the debris removed from the tooth surface during cleaning. Detergents also contribute to the foaming property of dentifrices, an effect which appeals to consumers. The commonly used detergent in toothpaste is sodium lauryl sulphate. Flavours constitute only a minor part of a dentifrice, but are important components for consumer acceptance. The flavour of a toothpaste is usually a blend of several components. The principal flavours used are peppermint, spearmint and wintergreen modified with other essential oils of anise, clove, caraway, pimento, eucalyptus, citrus, menthol, nutmeg, thyme or cinnamon. The humectants and some of the binders in toothpastes can act as nutrients for various micro-organisms. Microbial contamination of dentifrices is restricted by a low water activity and by the inclusion of preservatives such as benzoates. Colouring agents are also added to dentifrices. These include titanium dioxide for white pastes and various food dyes for coloured pastes and gels (Chess et al., 1950).

Contact sensitivity or mucosal irritation by dentifrices is relatively rare. Occasionally, the flavours, colourings or preservatives can cause allergic reactions in some individuals.4 These may include desquamation and oedema of the lips and tongue, perioral dermatitis, angular cheilitis, gingivitis and intra-oral ulceration. Some toothpastes can cause mild irritation of the oral mucosa which disappears after use and is usually attributable to the detergent or essential oil flavours. This again is quite rare as the majority of people prefer the more flavoured dentifrices as the tingling sensation makes the mouth feel fresh and clean, albeit for only a few minutes (Machackova and Smid, 1991).

Therapeutic agents

The use of fluoride dentifrices is beneficial in the prevention of dental caries. Most dentifrices today contain 0.1%(1000 ppm) fluoride, usually in the form of sodium monofluorophosphate (MFP); 100 g of toothpaste containing 0.76 g MFP (equivalent to 0.1 g fluoride). The concentration of fluoride in toothpaste is limited in Australia to a maximum of 1000 ppm by a recommendation of the Standard for Uniform Scheduling of Drugs and Poisons. The suggested toxic dose of fluoride ion is 5 mg fluoride per kg body weight.6 For a 10 kg child, this corresponds to approximately half the contents of a 90 g tube of toothpaste. Therefore, young children should not be allowed unsupervised access to fluoride dentifrices (Rolla, 1991).

A review of fluoride benefits and risks by the U.S. Public Health Service concluded from more than 50 human studies that no evidence existed showing an association between fluoride and cancer. However, there is evidence of an increase in the prevalence of mild dental fluorosis. The consumption of excessive amounts of fluoride during enamel formation may result in dental fluorosis. This is a continuum of changes in the enamel varying from fine white lines in mild cases to very chalky, opaque enamel which breaks apart soon after tooth eruption. Since fluoride appears to affect the activity of the ameloblast, especially during the late secretion or early maturation of enamel, excessive fluoride intake is of concern during the first 7 years of life. Toothpastes have been identified as a significant source of fluoride for the young child. The NHMRC has reported in its review of the effectiveness of fluoridation that fluoride from toothpaste accounts for up to 53% of the total fluoride intake of children aged two years (Whitford, 1987).

For the purpose of enhancing the safe use of fluoride dentifrices by children, several measures should be taken to minimize the risk of developing dental fluorosis. Parents should be advised to supervise tooth cleaning closely using only small (pea-size) quantities of toothpaste. Manufacturers should be encouraged to market a low fluoride dentifrice (e.g.400-500 ppm fluoride) for infant use. This level of fluoride in toothpaste, given that all other sources are constant and low, should result in a total fluoride intake which does not exceed the recommended upper limit of 0.07 mg/kg of body weight for a child between 2 and 7 years of age. A low fluoride, sorbitol-based toothpaste designed specifically for children is available (Colgate Junior Toothpaste) and contains 0.304% MFP (400 ppm fluoride). Data from several independent studies indicate that, although a dose-response relationship does exist for fluoride levels in toothpaste and caries, use of a 400 ppm fluoride-containing paste by children under 7 years of age instead of the standard 1000 ppm fluoride paste should not increase their caries risk. There is currently no glycerol-based, 400-500 ppm fluoride toothpaste available in Australia (Machackov 1991).

In addition to claims of the anticaries activity of fluoride, new therapeutic dentifrices are being promoted to the public and the dental profession for the control of dental plaque and gingivitis. Triclosan (2,4,4'-trichloro-2'-hydroxy diphenyl ether), an antimicrobial agent used extensively in deodorants, soaps and other dermatological preparations, is the active agent in these new dentifrices. In various clinical studies, brushing with a 0.3% triclosan-containing dentifrice when compared with a control paste resulted in significant reductions in dental plaque formation and gingival inflammation.11 From animal toxicity, mutagenicity, teratogenicity and carcinogenicity studies, as well as pharmacokinetic studies in man, it has been proposed by the major toothpaste companies that 0.3% triclosan-containing toothpastes are safe for human use (Machackova, 1991). However, the U.S.A. Food and Drug Administration has stated that new drug approval will be required for a toothpaste containing triclosan because the ingredient has not been used in any intra-oral drug or cosmetic preparation previously.

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