Genetics of Bitter and Fat Taste

Demographic data Participants will complete a questionnaire asking about their demographics provided in Appendix. The answers to these questions will be used to control for potential confounding effects in statistical analysis.

Genetic Analysis Saliva samples (2 ml) will be collected for DNA analyses (Genefix, Isohelix, Kent, UK) and stored in the fridge at 4 °C in M201 lab at St Mary's University. DNA extraction will be carried out through use of a PSP® SalivaGene 17 DNA Kit 1011 following the standard manufacturer protocol. DNA quantification and quality control will be assessed with spectroscopy. Following this, genotyping will be carried out using prepared TaqMan® SNP genotyping assays for genes coding for bitter and fat taste receptors and a StepOnePlus thermocycler. All samples will be analysed in duplicate in accordance with the manufacturer's protocol. All genetic analyses will be carried out at St Mary's University, Twickenham.

Anthropometrics Height (m), weight (kg) and waist circumference (cm) will be recorded by the researcher team. BMI will be calculated using the equation: weight (kg)/ height (m2) ± standard deviation and waist circumference and distribution will be compared to the UK standards.

Fat and bitter taste sensitivity The Oral Fatty Acid Threshold Assessment and Ascending Forced Choice Triangle Procedure will be used to determine each participant's oleic acid (C18:1) detection threshold (fat taste sensitivity). Briefly, each participant will be presented with three cups containing 30 ml vehicles in a random arrangement, two controls and the third containing oleic acid (0.02, 0.06, 1, 1.4, 2, 2.8, 3.8, 5, 6.4, 8, 9.8, 12, 20 mM). Participants will have to identify the oleic acid solution by tasting but not ingesting the solutions. A participant will be required to choose the oleic acid solution correctly three times at the same concentration to define their threshold. If participant is incorrect at any point, further three cups will be presented, one containing the higher concentration. There will be thirteen concentrations in total. The bitter taste compound used to classify participants into bitter taste hypo-, normal- and hypertasters will be PTC. Participants will place a strip on the tongue and use the general Labeled Magnitude Scale (gLMS) to report the intensity of the taste compound.

Food preference Food preference will be measured by asking participants to taste food usually perceived as bitter (dark chocolate and broccoli) as well as food that are defined as sweet (milk chocolate and carrots).To minimise variability, the vegetables will be presented raw. Overall preference will be tested using a nine-point Likert scale. Each participants will rinse their mouth with water between each food and a minimum of one minute will be left between each test. Eating behaviour will be determined by use of the validated Adult Eating Behaviour Questionnaire. Fat preference will be determined by a UK adaption of the validated Fat Preference Questionnaire.

Dietary Intake The validated EPIC Norfolk Food Frequency Questionnaire (FFQ) will be used to measure dietary intake over the previous year.