The aim of this study is to determine how different ingredients which are found in food and other consumer products affect the ability of saliva to form surface films.

Saliva forms a protective film on the surfaces inside the mouth. This film affects the way we consume and digest food and protects our teeth and other surfaces inside the mouth from disease and decay. It is not well known how this film forms, or which components in everyday life affect formation of the film, and therefore its protective properties.

Samples of saliva will be collected from apparently healthy volunteers. The properties of the film (thickness, density, strength, protein composition) will be studied in the laboratory under controlled conditions. The effects of common ingredients in foods and other products such as proteins, emulsifiers, preservatives and surfactants on the formation and properties of the film will be determined.

The results of this study will help us to understand how different ingredients can change the properties of saliva that may affect the way that we sense and digest food, and the ability of saliva to protect against decay and infection. This could, in the future lead to new ideas for developing:healthy foods (e.g. lower fat) that taste better, or more effective dental care products.

The aim of this study is to determine the physical and biochemical mechanisms responsible for the formation and structure development (in vitro) of the salivary mucous film and the impact of common ingredients found in food or oral hygiene products.

Saliva forms a film or pellicle on oral surfaces which serves as a protective barrier and lubricates the oral tissues during food consumption. However, it also provides the primary site for bacterial attachment and the build up of plaque. Therefore the physical and molecular structure of the salivary film is important for how we sense and taste food, how food is broken down in the mouth, and its effectiveness as a protective barrier. Much is known about the composition of saliva, but very little is known about the composition and structure of the salivary film or pellicle. We believe that the film formation and its physical properties, such as viscosity and strength are the result of complex interactions between different proteins within saliva, and with other components such as calcium and other salts, food components such as proteins and emulsifiers, and active compounds found in oral hygiene products such as preservatives and surfactants. This study will determine how the salivary film forms, its molecular composition and which other critical components affect its ability to form a stable, protective barrier. This knowledge will hopefully, in the future contribute to strategies to design healthier foods and more effective oral hygiene treatments.

The physical properties of saliva are not stable as the proteins tend to aggregate following secretion, which affects the length of time the film formation can be studied (ex vivo). Therefore we need to use fresh human saliva samples collected from healthy volunteers using a defined protocol to ensure sample quality, and assess them within four hours of collection. Saliva will be collected (either whole mouth or parotid), taken to the laboratory and allowed to form films on model surfaces, representative of those found in the mouth. The rate of formation, strength, thickness and mass of the films will be studied using a range of physical methods. The protein content and composition will also be determined to identify the components responsible for various stages of film formation. Then, the influence of environmental conditions such as pH, calcium and salt will be determined. Finally, we will determine how the films interact with external components found in food and oral hygiene products such as proteins, emulsifiers, preservatives and surfactants. Although we are not studying the effects of dietary intervention on saliva properties, we want to determine the underlying mechanisms responsible for film formation and break down. Paired T-tests will be used to compare parotid with whole mouth saliva from the same volunteers. Two sample T-tests will be used for comparison of similar measurements on samples from different volunteers. Diagnostic checks on results will be performed to check whether transformations are required to meet normality assumptions.

Two types of saliva samples will be collected: Whole Mouth Saliva (WMS) and Parotid Saliva (PS). The saliva will be stored at 4ºC (WMS may also be centrifuged at 2700xg for 20 minutes at 5°C to remove aggregates) and used within 4 hours of collection, as protein aggregation begins immediately after secretion, affecting the film formation properties, as determined in previous work. A range of physical techniques will be employed to determine the rate and extent of film formation, as well as the thickness, mass and rheological properties, under the different solution conditions and in the presence and absence of added ingredients as described above. Protein concentration and composition will also be determined. This will enable improved experimental design (e.g. normalising protein concentration) to improve the quality of the results. The results will be used to inform the design of further studies with other centres of expertise in order to understand the impact on in vivo behaviour of the salivary film and, for example the sensory perception of foods and the role of saliva on digestion of food structures.

Inclusion Criteria:

• Age: 18 - 65 years old
• Gender:Male or Female
• nonsmokers

Exclusion Criteria:

• diabetics
• medical history of Tuberculosis or Hepatitis
• depressed or elevated blood pressure measurements (<90/50 or <95/50 if symptomatic or >160/100).
• take any medication which may affect saliva production
• are related to someone in the study team i.e. spouse, partner or immediate family member
• are pregnant, planning to become pregnant, or are currently breastfeeding.
• have results of the clinical screening which indicate, or are judged by the Human Nutrition Unit (HNU) Medical Advisor to be indicative of, a health problem which compromise the wellbeing of the volunteer if they participated, which would affect the study data.

In addition, prior to each sample donation, the following conditions will be checked, and volunteers will only be allowed to donate saliva if none of the following apply:

• They have any current dental or mouth problems (bleeding gums, sores on mouth, dry mouth syndrome)
• They have had a viral illness or chest infection within last 7 days
• They are not able to produce sufficient saliva
• Volunteers will be required to avoid eating and drinking (except water) for 1 hour prior to saliva collection.

Apparently normal healthy volunteers from the local community able to donate saliva samples.

Volunteers were required to donate saliva for ex-vivo study, as described in protocol. A pool of 12 volunteers was maintained. 3 further volunteers had to be recruited following 3 losses due to personal reasons, i.e. not connected with study.

No adverse events were reported. The number of volunteers required gave acceptable statistics as anticipated. The study was completed on time