Assignment 2- Research Paper Summary and Critique

Humans Can Taste Glucose Oligomers Independent of the hT1R2/hT1R3 Sweet Taste Receptor

Authors: Trina J. Lapis, Michael H. Penner and Juyun Lim

Summary

Introduction

                The main function of taste is to identify substances in the oral cavity, including food which provides energy or harmful toxins. Currently there are 5 generally excepted tastes; sweet, sour, bitter, salty and umami. Each flavour is known to use its own receptors and transduction pathways. Umami uses G-protein coupled receptors (T1R family) and metabotropic glutamate receptors, bitter uses G protein coupled receptors (T2R family), sour uses cyclic nucleotide-gated channels (HCNs), acid sensing ion channels (ASICS), and transient receptor potential channels (PDK2L1 and PKD2L3), and salty uses epithelial sodium channels (ENaC). The flavour with the most focus in this research is sweet, which utilizes G protein coupled receptors T1R2 and T1R3 (hT1R2/hT1R3 in humans). Recently the idea of the 5 main flavours has been changing with evidence towards flavour detecting of chemicals such as calcium, fat and starch products.

                It would be beneficial to animals and humans if starch had its own flavour and could be detected (due to the nutritional value), but it is unlikely as starch is a very long polymeric molecule. However, it is possible for starch hydrolysis products to be detected in the oral cavity after salivary amylase hydrolyzes it into simple sugars and oligomers. Previous studies have demonstrated the ability of humans to detect maltodextrin (polymer mixture) independent of sweet taste and rats to detect polycose (mixture of glucose oligomers and polymers) and differentiate it from sucrose. However, all previous studies have used mixtures and have not used accurate controlling factors to ensure accurate results.

                This experiment separated maltodextrin into 3 groups based on their degree of polymerization (number of monomers in the chain). Sample 1 (S1) had a DP of 7 (oligomer), sample 2 (S2) had a DP of 14 (oligomer), and sample 3 (S3) had a DP of 44 (polymer). The objectives of the study were to determine the lengths of glucose chains that can be detected, investigate a potential gustatory mechanism, establish glucose oligomer taste qualities, and create a dose-response curve for glucose oligomers. Each objective was the focus of one experiment in a series of 4 completed during this study.

Materials and Methods/Results

Experiment 1A – Taste Discrimination of Glucose Oligomer and Polymer Stimuli as equivalent % w/v solutions

It was required to stop the hydrolysis of the oligomers and polymers by alpha-amylase into simple sugars in order to get accurate results. In order to do this, an alpha-amylase inhibitor (acarbose) was used. Pilot experiments were completed to ensure that acarbose truly inhibited alpha-amylase (in average salivary concentrations, 5mM) when S1, S2, and S3 were present. This test was completed to determine what samples were able to be detected by human gustation, and it was predicted that S1 would have the greatest detection rate and S3 would have the lowest detection rate. 22 subjects (male and females) with an average age of 25 were used. The criteria for entering the experiment were that they were non-smokers, not pregnant, not medication takers, no history of smell or taste loss, no past oral disorders, no oral piercings, no recent dental work, no alcohol consumption in 12h, no food or beverage containing dairy within 4h, no food or beverage at all in 1h, and no products containing menthol within 1h. Solutions of 6% and 8% (w/v) were created, stored at 4-6 degrees Celsius, and returned to room temperature before testing. Each subject was put through 2 sessions (one 6% and one 8%) on different random days. The test consisted of 3 sip-and-spit tests (2 controls and 1 sample), where they sampled all 3 and had to identify the one with different flavours. In order to prevent olfactory input, they wore nose clips as well. This continued 3 times (one for each sample) and they rinsed with water in between, and the orders were counterbalanced and randomized. The results were added by number of correct identifications, which were statistically converted to d' values that accurately represented the stimulus and subtracted background results (noise). The prediction was shown to be accurate as the subjects could distinguish S1 and S2 from blanks, but not S3. d' values were also higher for S1 than S2, and were higher for the 8% samples than the 6% in both cases.

Experiment 1B- Taste discrimination of glucose oligomer and polymer stimuli as equivalent mM solutions

It was thought that the test results in experiment 1A may not have been accurate as the w/v ratios may have skewed the results in favour of S1 and S2 (as each mole of S3 weighs much more, therefore less molecules of S3 were present). This experiment aimed to correct this by testing samples with equal reducing ends (RE). The RE’s signify one oligomer or polymer, therefore it is a direct measurement of the number of molecules and would be more balanced. The problem arose when an S3 solution with the predetermined concentration of 75mM (8% w/v for S1, 17% w/v for S2, and 54% w/v for S3) became notably viscous. A blank was created with tasteless, viscogenic agent, methylcellulose to correct for this. 26 subjects were obtained with the same criteria as experiment 1A. All stimuli were prepared with 5mM of acarbose, and the blanks were created with consistent viscosity. The stimuli were also presented on cotton swabs to reduce further textural clues, although the pattern of experimentation remained unchanged. The results ended up being very similar, with subjects able to discriminate S1 and S2 from blanks, but not S3. Although contrary to prediction, the detection rate of S1 and S2 were actually almost equal, differing from experiment 1A.

 

Experiment 2- Taste Discrimination of sugars and glucose oligomers in the absence and presence of lactisole

It was also important to test if similar mechanisms of detecting simple sugars were used to detect glucose oligomers. In order to do this a chemical called lactisole was used (tasteless, sweet taste blocker that binds to hT1R3). 25 subjects were used with an average age of 25, with the same criteria as previous experiments. S1 and S2 samples were used, along with glucose, maltose and sucralose (artificial sweetener) to test for lactisole effects. Acarbose was added, and all target and blank stimuli were prepared with and without lactisole. 5 stimuli were provided, with and without lactisole (total of 10 tests), and pseudo-randomization was used as it was impractical to do all the possible combinations. The same cotton swab triangle tests were completed exactly as before, and results were analysed into d’ values. All 5 stimuli were equally detected in the absence of lactisole, but when lactisole was present it blocked the taste of glucose, maltose and sucralose. However, lactisole did not compromise the ability to detect the glucose oligomers. 

 

Experiment 3- Determination of taste quality of glucose oligomer through a focus group discussion

                

This experiment was aimed at subjectively describing the taste quality of the glucose oligomers, which had already been established as using different mechanisms of detection in experiment 2. 7 subjects who had already participated in one of the studies were used. Different amounts of equally intense aqueous solutions of sucrose, sucralose and S2 were provided using the cotton swab technique. In a focus group the subjects tasted the unknown stimuli and were asked to describe the flavour and, as a group, come up with a one word descriptor for the taste they were experiencing. It was seen that sucrose was “sweet” like sugar water, sucralose was “sweet” like artificial sweetener, and the glucose oligomer was “starchy” like a root vegetable, corn, bread, or pasta. Sucrose and sucralose were also identified as being much more similar than either sucrose or sucralose to the S2 oligomer.

Experiment 4- Establishing dose-response curves for sugars and glucose oligomers

                 

Once the new flavour was characterized, it was of interest to create a dose-response curve which could be compared to natural sweeteners. 20 subjects (mean age of 25) were recruited again with the same criteria. Sucrose, glucose and S2 were provided at 3 concentrations (45, 100, and 224mM) with acarbose present in all. The cotton swab technique was used again, and the subjects were asked to rate the intensity of a general version of the Labeled Magnitude Scale (gLMS) which they were previously trained to use. The data was plotted on a dose response curve based on molar and %w/v concentrations. These results showed that glucose and the glucose oligomer had almost indistinguishable curves based on moles, however, when using %w/v ratios, it was seen that glucose oligomers were shifted to the right of the glucose curve. 

Discussion

                Based on the results from experiment 1, it was seen that humans can discriminate glucose oligomers from water, but not glucose polymers from water. It is important to note that taste is not the only sense responsible for detection of glucose oligomers, although the other sense were controlled in these experiments (nose clips to reduce olfactory clues and similar textures to reduce somatosensory clues). It is also important to note that the breakdown of starch and glucose oligomers to simple sugars can contribute to the recognition of glucose oligomers in vivo, but this was constantly controlled for in every experiment using acarbose. A conclusion can be made that, in the absence of confounding effects, glucose oligomers can be sensed by the human gustatory system. Although these results conflict with previous studies which stated that rats can taste glucose polymers, the previous studies did not control for glucose polymer hydrolysis.

                Based on the results from experiment 2, it was seen that subjects lost the ability to taste glucose, maltose, and sucralose, in the presence of lactisole but did not lose the ability to taste glucose oligomers. As lactisole (which blocks T1R3 sweet receptors) did not stop the sensation of glucose polymers, it is clear that glucose oligomers use a different taste receptor. These results are consistent with previous finding where T1R2/T1R3 knockout mice could still taste glucose oligomers.

                These findings were backed up by evidence of experiment 3, where subjects rated the flavour of oligomers as “starchy”. In the past, other receptors have been thought to be used for detection of sugars and glucose oligomers, such as T1R-independent pathways. However, the findings in this study also disprove this theory, and it is now thought that there is a completely novel receptor used for starch hydrolysis products. The main function of this receptor is likely to identify incoming starch which has begun being broken down by amylase while entering the digestive system.

Personal Critique

This paper did an excellent job at covering all of the bases when determining the detection of starch hydrolysis products on the tongue. It reviewed many pieces of previous work done on this topic, and improved upon them by adding more controlling factors and doing multiple experiments to back up their theory. The use of acarbose as an amylase blocker was what differentiated this paper from the rest and seemed to produce more accurate effects and results. Experiment 1 was also repeated using different measurements in order to fully ensure that the results they obtain were accurate. Using step by step experiments (determining if humans could taste starts, and then determining if they could taste them without sweet receptors) was a very intelligent thing to do. If they relied on previous work, which stated that rats could taste glucose polymers, it may have skewed the results when they blocked the sweet receptors with lactisole. The experimenters also made it very clear that they completed a pilot study (ensures the chemicals were functioning accurately) whenever it was required. Although the results of this experiment were not specific (they did not find a receptor responsible for glucose oligomer detection), they did their best to lay down the groundwork for further research on this topic and ruled out some possible paths of research.  

In my opinion, experiments 1 and 2 were enough information to produce a paper on their own. The research completed in experiments 3 and 4 seemed unnecessary and generally not important to the main idea of the paper. Experiment 3 produced subjective flavour characteristics of the glucose oligomers which were deemed “starchy”, however this idea had already been demonstrated in the first 2 experiments. Experiment 4 created a dose-response curve for the concentrations needed to detect glucose oligomers, which could have been done in a separate paper, especially because the results were not even discussed in the discussion section. One important thing to note about the first 2 experiments are the small sample sizes that were used (about 25 people). Even though the data was statistically significant, it could have increased the validity of the research if more subject were used in each experiment. Other than these factors, I think the paper was well written and organized in a way that made sense. The results they obtained could open up a door into more research about flavour detection of not only glucose oligomers, but many other potential chemicals.

Reference

Lapis, T. J., Penner, M. H., & Lim, J. (2016). Humans Can Taste Glucose Oligomers Independent of the hT1R2/hT1R3 Sweet Taste Receptor. Chemical Senses, 41(9), 755-762. doi:10.1093/chemse/bjw088 

Note: All Figures and Images in this section were retrieved from this paper which can be found here.