Advanced Topics in Biomedical Science

Table of Contents

Introduction.

Background.

Aims and rationale.

Materials and Methods.

Selection of subjects.

Data collection and experimental protocol

Results.

Discussion and Literature review..

Recommendation.

Conclusion.

Reflection.

Reference list

Introduction to Consumption of Chocolate

The consumption of chocolate has recently increased at a drastic rate throughout the world. In the last few years, many independent research groups have published works that suggested that the consumption of chocolate containing products is likely associated with improved memory and cognitive function of the brain. Most of them performed correlation studies with a large number of participants to support their observations. One such longitudinal study published in 2016 found that memory and abstract thinking improved in those reporting more chocolate consumption. This was a follow-up study where the members filled questionnaires and undertook physical assessments at different periods so that the researchers could assert changes related to aging, the development of coronary diseases, and brain performance. The analysts compared among the individuals who never or once in a while consumed chocolate from those individuals who consumed chocolate once every week. Individuals enlisted in the study were given different brain function tests – including recalling where things were (spatial memory), theoretical thinking, reasoning, working memory, and attention. The association between chocolate consumption and the execution of cognitive tasks were then systematically analyzed. The members revealed that they ate chocolate more than once in a week performed better in the vast majority of the subjective tests; especially on their "visual-spatial memory and association" score (Afoakwa, 2016). No relationship was found with working memory.

Background of Chocolate

Chocolate is regularly disparaged because of the high sugar and fat substance in chocolate bars and its relationship with "indulgence". In any case, cocoa (or chocolate in its crude structure) contains many compounds that may affect the functioning of the brain. Specifically, cocoa contains flavonoids, which are found in organic products, for example, grapes and apples. Cocoa likewise contains caffeine and theobromine, which are found in tea and coffee. These organic compounds are thought to improve readiness and hence can also render a positive effect on the functioning of the brain. In any case, the amounts of these synthetic compounds rely upon the concentration of cocoa in chocolate. Milk chocolate contains significantly less amount of these organic chemicals in it than dark chocolate. A review published in the year 2017 analyzed the proof to date that flavanols found in cocoa may profit the human brain function.

The review gave a glimpse of some of the major findings published in different research articles (Afoakwa, 2016). The review pointed out that short-term consumption of chocolate may be useful for increasing memory and brain function. The benefits were visible only in the case of dark chocolate and not white chocolate. Another study pointed out that long-term consumption for at least 3 months may be helpful for the enhancement of brain performances. Several other studies provided increasing evidence that consumption of cocoa-based products rapidly increases the electrical activity in the brain. These changes do not share any positive correlation with improved cognition and hence it is difficult to associate these results with improved brain function.

These researches highlighted the fact that all chocolate is not the same (Bakoyiannis et al., 2019). Dark chocolate has high levels of flavanol, while white chocolate has much lower levels. Besides, many different types of chocolate are high in fats and sugar.

Chocolate contains many compounds like methylxanthines which have been associated with increased brain activity and cognition. Some investigators pointed out in their study that the amount of methylxanthine present in a bar of chocolate largely affects the mood and cognitive behavior of individuals. They showed that the cognitive function improves greatly with the increasing amount of cocoa in the chocolate. In their study white chocolate showed a similar kind of effect as placebo or water, with no significant change in the cognitive function.

During the last decade, a few investigations have shown that cacao-containing beverages or food items, for example, chocolate and cocoa may categorically and positively affect human wellbeing. The most imperative constituents of cacao laden foods are flavonoids (Catechin and epicatechin) and methylxanthines (Caffeine and theobromine) (Bakoyiannis et al., 2019). The flavonoids play a major role as an anti-cancer agent or as focal sensory energizers of the brain whereas the methylxanthine is expected to improve the cognitive and cardiovascular activity.

Aims and Rationale: Stars Project Concept

In the past few years, a lot of evidence has been put forward supporting the role of chocolate and other cocoa derived products in the enhancement of executive neurocognitive functions including memory and attention. In a recent cross-sectional study involving adult participants from a particular community, it was found that consumption of a high amount of dark chocolate was largely associated with a much better cognitive function (Cavarretta et al., 2018). Short-term studies among healthy older adults or middle-aged individuals revealed the cognitive benefits of flavonol consumption in cocoa drinks or supplements. The trials that have conducted enlisted very few individuals and the cognitive brain functioning have also been measured in very heterogenic ways.

In contrast to all the aforementioned and discussed researches, some studies led by other groups using a much more chronic administration of cocoa flavanols or in particular dark chocolates have found different results. In this case, with a consumption period of 5 to 42 days, the investigators failed to draw any positive correlation between chocolate intake and improved cognition (Cavarretta et al., 2018). The possible variability in the flavanol content in different varieties of chocolate may vary largely according to the type of bean used, country of origin, and processes of extraction and manufacture. This may generally contribute to the variable outcomes that have been observed in different studies to date on the effects of cocoa flavanols on brain activity and cognition. The objective of this study was quite subtle; it was to evaluate the effect of administration of dark and white chocolate on the working ability of the brain in students belonging to two different campuses. In clearer words, this study intended to assess the effects of chocolate consumption on brain functioning in healthy young subjects (Cicvaric et al., 2018). We hypothesized that the consumption of chocolate especially those with high cocoa content would result in enhanced brain function. We also tried to assess whether these improvements significantly differ from those observed in those consuming white chocolate.

Materials and Methods: Stars Project Concept 

Selection of Subjects

This is a randomized single-blind study performed with young healthy university students. The hypothesis was to determine the extent of improvement in brain function upon consumption of chocolate both dark and white. Students were allowed to participate freely as per their choice, feeling questionnaires' and recording all individual data. The inclusions of the participants were based on the following criteria: they should be non-smokers, not under any medication, without chronic clinical problems like diabetes, hypertension, renal complications, cardiovascular complications, gastrointestinal complications, and neurological problems. The participants were not provided with any information about the objective of the study and no examination data was provided to them. All the researches and data analysts were kept blinded to the groupings and sub-groupings of the participants. All the participants were instructed to maintain a healthy diet and avoid intake of any caffeinated products including beverages and chocolate bars from any outside source during the study. All participants were restricted to participate in any form of intense exercise. The procedures and potential risk factors for the study were carefully explained to all the participants and informed consent was collected from each of them.

Data Collection and Experimental Protocol

The following data were collected for each subject: Gender, Age (years), Body Mass Index (BMI), Number of hours of sleep per day: Heart Rate (per day), Self-assessment survey of general health, general mood, energy levels once at the beginning and once at the end and memory on a day to day basis (Cova et al., 2019). The brain function was assessed based on the scores obtained on the Lumosity Brain Performance Test. This test assesses the three main aspects defining a functioning brain i.e. flexibility, attention, and memory (Decroix et al., 2016).

The total population of students was divided into two experimental groups, one with those consuming dark chocolate (Lindt-85% cocoa) and the other with those consuming white chocolate (Lindt Extra Silky). University students belonging to the Burwood campus were selected to consume only dark chocolate, whereas those belonging to the Geelong campus were selected to consume only white chocolate (Crichton, Elias & Alkerwi, 2016). Now students from each campus were further divided into 4 separate subgroups based on the consumption of chocolate and participation in the lumosity brain performance test. The subgroups are as follows: Sub-group A1- The subjects did not consume any chocolate but played the lumosity brain training game 2 times per week; Sub-group A2- The subjects consumed 20 gram of chocolate per day for a period of 20days and played the lumosity brain training game 2 times per week; Sub-group A3- The subjects did not consume any chocolate but played the lumosity brain training game every day; Sub-group A4- The subjects consumed 20 gram of chocolate per day for 20 days and played the lumosity brain training game every day. 

The Statistical analyses were performed in Excel using descriptive measures, mean and standard deviation calculated as per the formula. An unpaired t-test was performed in excel. Statistical significance was considered at values < 0.05 (Fox et al., 2019).

Results: Stars Project Concept 

Of the different variables taken into account like general health, end energy, general mood, and memory measured as a single value on a fixed scale, no significant change was observed in the mean values of the parameters in case of all the four sub-groups, fed with white chocolate. All of these values were ascribed specific values one at the beginning and one at the end of the 20 days study (Crichton et al., 2016). The BMI in case of the Geelong group was found to be increased in case of both A2 and A4 sub-groups where there was an intake of chocolate (25.6 and 22.4 respectively) compared to the sub-groups A1 and A3 where there was no chocolate consumption (20.08 and 19.5 respectively) (Cicvaric et al., 2018). The rest of the parameters were more or less similar between the sub-groups in the case of the Geelong group. The other parameters that were measured include Sleep day, BPI, Speed, Memory, Attention, Flexibility, and problem solving for each participant (Decroix et al., 2016).

Values were measured every single day for 20 days with a break-up after 10 days. When unpaired t-test was performed, it was found that only the mean value of attention among the participants (Baseline to 10 days), Flexibility (Baseline to 10 days), Problem-solving ability (Baseline to 10 days) and BPI (11 to 20 days) varies significantly among the sub-groups A3 and A4. The P-values are as follows: 0.034, 0.036, 0.016, and 0.001 respectively. There were no significant differences in the mean values of any of the parameters between sub-groups A1 and A2. Among all of these parameters taken into account, values for sleep day and speed remained almost the same in the case of all the sub-groups (Crichton, Elias & Alkerwi, 2016). 

In the case of the Burwood group, the same parameters were tested for the Geelong group. General health, end energy, general mood and memory measured as a single value on a fixed scale, when tested for the individuals belonging to the Burwood group i.e. those who were fed dark chocolate, no significant change was observed in the mean values of the parameters in case of all the four sub-groups. BMI was significantly high in the case of the A4 subgroup (Mean- 29) compared to the other three (Fernell, Swinton & Lukowiak, 2016). The BMI for the A2 group (25.7) was higher compared to both A1 (23.5) and A3 (23.9). The other parameters that were measured include Sleep day, BPI, Speed, Memory, Attention, Flexibility, and problem solving for each participant (Socci et al., 2017). Values were measured every single day for 20 days with a break-up after 10 days. When an unpaired t-test was performed, it was found that only the mean value of Flexibility among the participants (Baseline to 10 days), Flexibility (11 to 20 days), and Memory (11 to 20 days) varies significantly among the sub-groups A3 and A4. The P-values are as follows: 0.017, 0.001, and 0.025 respectively (Fernell, Swinton & Lukowiak, 2016). There were no significant differences in any of the parameters among sub-group A1 and A2.

Discussion and Literature Review on Stars Project Concept 

In the last few years, scientists from all over the world have suggested having noticed increased cognitive memory performance and improved information processing ability of healthy individuals after consumption of cocoa-based flavanols following a dedicated regime (Socci et al., 2017). They hypothesized that the cocoa flavonols quite significantly increase the normal cognitive functioning in healthy working people but only with serious and dedicated training activities (Fox et al., 2019). The study also demonstrated that the flavonols also have the neuroprotective ability. In some recent reports, it has been shown that the chocolate-based flavonoids do increase cognitive functions in healthy people and also in those who are suffering from any sought of cognitive impairment (Crichton, Elias & Alkerwi, 2016). Many commercial companies have synthetically modified the normal flavonoid content in chocolate bars, thereby increasing their amounts along with decreasing the calorific content. Recently, an experimental cross-sectional study has reported that chocolate consumption readily increases attention and improves working memory in students (Cova et al., 2019).

Enhancing one’s general cognitive abilities has in recent times become a tremendously fascinating challenge for the scientific community. This interest is owing to the demand among the public for either prevention of age-related cognitive decline or increasing the cognitive output or performance to match the demand of the work environment. In the last few years, cocoa-based consumable products which are a rich source of flavonoids have been clearly shown to exert neuroprotective as well as modulatory functions (Grassi, Ferri & Desideri, 2016). Various studies have also reported the positive effects of chronic or in cases acute administration of cocoa-based flavonoids on memory, attention, and certain other cognitive functions. Currently, new-age research is speculating that through different biological actions both direct and indirect, cocoa and cocoa-derived food are suggested to efficiently counteract the decline in general cognitive ability as well as sustain the positive cognitive abilities, particularly among those patients who are at risk.

Although this whole research idea and experimental work is still at a very preliminary stage, researchers investigating the relations between cocoa and cognition have found a dose-dependent improvement in general attention, speed of processing information, cognition, and memory (Grassi, Ferri& Desideri, 2016). Administration of the flavanols found in cocoa has been shown to enhance the normal functioning of the cognitive process and do exert a significantly protects the cognitive performance. The consumption of flavonoid-rich foods like dark chocolate has been associated with many health benefits, and a thorough literature search indicates that cocoa flavonoids are associated with high benefits to cognitive function. White chocolate is most likely a matched alternative for many consumers because it shares almost the same sensory properties like dark chocolate but without the presence of any flavonols (Cicvaric et al., 2018).

We planned for a similar outcome based on the effect of chocolate in brain function improvement. After carefully analyzing the data we found that there was not much of any difference involved in brain function among the participants based on the consumption of chocolate only (Afoakwa, 2016). Our results suggested that white chocolate might cause a significant increase in attention, flexibility, and problem-solving ability, starting immediately after the first day of consumption. But the improvement was only visible up to 10 days from the start and for only those who participated in the brain training game every day (Grassi et al., 2016). The observations we found are not clear indicators of whether only chocolate consumption is causing an improvement in these factors, there was no direct measure to check this out. We did not find any significant changes in these parameters for sub-group A2, in which case the participants participated in the brain training game only twice a week when compared to subgroup A1 (Bakoyiannis et al., 2019).

Thus our results indicate that taking 20gm of white chocolate along with brain training exercises every day might help to improve some cognitive parameters like attention, flexibility, and problem solving of brain functioning. The effects were visible starting from the first day of consumption and up to 10 days thereafter. These results were based on a standard observer's scales which are supposed to be error-prone (Kord-Varkaneh et al., 2018). A more authenticated procedure involving acute effect analysis, to measure the brain electrical activity after chocolate intake, in the tested and control group might yield a much-advanced result. The parameters like speed, sleep a day, and memory was in no way influenced by the consumption of white chocolate. General health or mood was also not influenced by the consumption of white chocolate (Cavarretta et al., 2018).

The effect on attention or flexibility which only remains for a limited time like 10 days since the beginning of consumption might suggest that the effect is not very prominent and the results were more likely due to the daily brain training exercises. More frequent white chocolate consumption and regular brain training exercises were significantly associated with better performance on the lumosity brain test score (Kord-Varkaneh et al., 2018). The brain training games are vital factors for brain parameter improvements as it was observed that in case of sub-groups A1 and A2, where the training was performed only twice a week, even after consuming 20 gms of chocolate there was no significant changes in the brain cognitive parameters concerning the control sub-group. There was no direct evidence to support our hypothesis that white chocolate might increase the cognitive functioning of the brain but the associated observations might help to claim that white chocolate consumption with regular brain training exercises surely helps to improve the brain functioning (Moreira et al., 2016).

In the case of the group consuming dark chocolate, a slight increase and improvement in some of the cognitive parameters like flexibility and memory were observed for those who consumed 20gm of dark chocolate daily and performed the brain training exercises regularly. We did not find any significant changes in these parameters for sub-group A2, in which case the participants consumed the same amount of chocolate daily but participated in the brain training game only twice a week when compared to subgroup A1. Thus our results indicate that taking 20gm of dark chocolate along with brain training exercises every day might help to improve some cognitive parameters like flexibility and memory related to the advancement in brain functioning(Moreira et al., 2016). The effects were visible only after 10 days since the start of the test process and continued till the end. Owing to its caloric value daily dark chocolate consumption caused an increase in the BMI of the participants.

The results though not very subtle give a hint towards the fact that dark chocolate consumption does change some of the cognitive parameters associated with brain function (Sumiyoshi et al., 2019). The other parameters like speed, sleep a day, and attention was in no way influenced by the daily consumption of dark chocolate. General health or mood was also not influenced by the daily consumption of dark chocolate (Oberrauter et al., 2018). The effect on flexibility and memory which was only observed after 10 days from the start of initial consumption might suggest that the effect is not very prominent and the results were more likely due to the daily brain training exercises. More frequent dark chocolate consumption and regular brain training exercises were significantly associated with better performance on the lumosity brain test score but not with an immediate effect like that with white chocolate.

The change happened slowly and took at least 10 days to be visible (Scaramuzza & Zuccotti, 2015). The brain training games are vital factors for brain parameter improvements as it was observed that in case of sub-groups A1 and A2, where the training was performed only twice a week, even after consuming 20 gms of chocolate there was no significant changes in the brain cognitive parameters concerning the control sub-group (Pereira et al., 2019). There was no direct evidence to support that dark chocolate consumption might increase the cognitive functioning of the brain but the associated observations might help to claim that eating dark chocolate daily with regular brain training exercises surely helps to improve the brain functioning to some extent with the effects being prominent only after a few days (Pereira et al., 2019).

Recommendation on Stars Project Concept

Modern experimental research depicting the synergistic effects of chocolate on the cognitive ability of the human brain is at this moment at its beginning or preliminary stage. Over the years many pieces of evidence have built up that suggests that consumption of cocoa flavonoids might be very effective in supporting the performance of active cognition which might simultaneously improve the measures of normal cognitive ability, speed of processing information, memory and attention (Pereira et al., 2019). If proved then this idea can be converted to point out and present the idea of using cocoa as an interesting and new tool having nutraceutical properties that might help to protect active brain functions, neuromodulation, cognition. This might also help to prevent different forms of cognitive decline in affected patients. Further investigations are required to establish this theory.

Future research on this field should be focused relatively on the proper identification of advanced experimental processes that are capable of detecting even minute changes induced by the flavonols in cognitive performance. It is also very important to characterize the exact dose, effective dosage time, as well as the proper type of flavanols required for the study to generate the particularly desired effects in the case of different populations. The inclusion of control groups is also important to assert the effect of the intervention in the tested group (Santiago-Rodríguez, Estrada-Zaldívar& Zaldívar-Uribe, 2018). To understand whether cocoa exhibits any long-term as well as acute/ immediate properties causing neuro-modulation more scientific investigations should be specifically designed in the future to including different advanced techniques associating with the different associated forms of cognitive and physiological measures.

Conclusion on Stars Project Concept

It can be concluded that the results generated from this study support the fact that chocolate consumption might have a link in increasing some of the cognitive abilities of the brain including attention, flexibility, problem-solving ability, and memory. White chocolate consumption with a daily regime of brain training exercises increases the flexibility and attention of the brain to a significant proportion, and the change was immediate. In the case of dark chocolate flexibility and memory, both increased for participants taking part in the daily routine of game training. The improvement, in this case, was not immediate but only observed after 10 days of consumption. This might suggest that the increased flavonoid content might provide a more long-lasting solution to the cognitive abilities of the brain. In the end, we can say that our hypothesis has been tested and it seems to hold. Chocolate consumption might be linked to increased brain activity. 

There happen to be some limitations in our study like sample size, which was essentially not very large, especially in the control groups; discrepancies in sample data collection with many leaving the study in between. The brain functioning was based only on the lumosity online test parameters and not on any other advanced techniques. The major findings for this project were: (1) White chocolate does have an effect on the flexibility, attention, and problem-solving ability of the participants but does not influence speed or memory in the Lumosity test (2) Dark chocolate improved the flexibility and memory parameter in the Lumosity tests but does not influence any other parameter.

Reflection on Stars Project Concept 

The STARS project is a unique concept that aims to build a more constructive view of research in the minds of young students. The idea was to create a platform through which research ideas could easily be formulated and reciprocated in the form of hand-on experience (Santiago-Rodríguez, Estrada-Zaldívar& Zaldívar-Uribe, 2018). This project brought about a new experience in my life as it helped me explore the dimensions of the field of research. When the project started, I was a bit hesitant but as the work progressed it gradually turned into a more comfortable one. The whole project was quite well organized and enabled us to slowly learn all the logical and experimental things. The idea of hypothesis testing involving a wide range of healthy subjects is the most distinct feature of this project. A large data set with a huge number of participants always generates a statistically significant output (Sumiyoshi et al., 2019). The most challenging aspect was to collect all the necessary data from all the individual participants. The process for data collection was practically very troublesome. Most of the data collected was categorically measured at a fixed scale.

The values ascertained like general health, memory, thinking, speed, etc. are all measured based on observed criteria which are subjected to human error. This has created statistical irregularities in the data set. A lot of participants left mid-study hence this also made it very difficult to construct a statistically significant data set (Sumiyoshi et al., 2019). The STARS project helped me to channelize the direction of my logical thinking and generate a scientific output by modulation of the associated environment. This is a very promising opportunity for me as this experience will help extensively in the development of skills related to project handling and data generation for my future endeavors. Skills like this will put me forward in the line for a potential job position. This project helped me gain a better insight into the nature of research projects and how to design them.

It has also helped me to improve my public relations by interacting with different participants. The whole research project was a demonstration based longitudinal study with a thorough objective and simple experimentation techniques that have helped me gather a prospective outlook of the whole scenario (Scaramuzza & Zuccotti, 2015). The most important aspect was the field-based data collection and assessment of individual performance. The hypothesis testing helped me to build a huge technical stance to reciprocate my theoretical knowledge. Collecting and analyzing the data set, got me face to face with the statistical discrepancies one could face in the field of study. This has prepared me in a much better way for the future, which might turn out to be important for different career opportunities. In the end, it can be said that this whole STARS project was a very entertaining and knowledgeable experience that has surely changed by scientific outlook to a great extent (Oberrauter et al., 2018).

Reference List for Stars Project Concept 

Afoakwa, E. O. (2016). World cocoa production, processing, and chocolate consumption pattern. Chocolate Science And Technology, 17-48.

Bakoyiannis, I., Daskalopoulou, A., Pergialiotis, V., & Perrea, D. (2019). Phytochemicals and cognitive health: Are flavonoids doing the trick?. Biomedicine & Pharmacotherapy109, 1488-1497.

Cavarretta, E., Peruzzi, M., Del Vescovo, R., Di Pilla, F., Gobbi, G., Serdoz, A., ... & De Falco, E. (2018). Dark Chocolate Intake Positively Modulates Redox Status and Markers of Muscular Damage in Elite Football Athletes: A Randomized Controlled Study. Oxidative Medicine And Cellular Longevity2018.

Cicvaric, A., Bulat, T., Bormann, D., Yang, J., Auer, B., Milenkovic, I., ... & Monje, F. J. (2018). Sustained consumption of cocoa-based dark chocolate enhances seizure-like events in the mouse hippocampus. Food & Function9(3), 1532-1544.

Cova, I., Leta, V., Mariani, C., Pantoni, L., & Pomati, S. (2019). Exploring cocoa properties: is theobromine a cognitive modulator?. Psychopharmacology236(2), 561-572.

Crichton, G. E., Elias, M. F., & Alkerwi, A. A. (2016). Chocolate intake is associated with better cognitive function: The Maine-Syracuse Longitudinal Study. Appetite100, 126-132.

Crichton, G. E., Elias, M. F., Stranges, S., & Abhayaratna, W. P. (2016). Relation of habitual chocolate consumption to arterial stiffness in a community-based sample: preliminary findings. Pulse4(1), 28-37.

Decroix, L., Tonoli, C., Soares, D. D., Tagougui, S., Heyman, E., & Meeusen, R. (2016). Acute cocoa flavanol improves cerebral oxygenation without enhancing executive function at rest or after exercise. Applied Physiology, Nutrition, and Metabolism41(12), 1225-1232.

Fernell, M., Swinton, C., & Lukowiak, K. (2016). Epicatechin, a component of dark chocolate, enhances memory formation if applied during the memory consolidation period. Communicative & Integrative Biology9(4), 816-23.

Fox, M., Meyer-Gerspach, A. C., Wendebourg, M. J., Gruber, M., Heinrich, H., Sauter, M., ... & Juengling, F. (2019). Effect of cocoa on the brain and gut in healthy subjects: A randomised controlled trial. British Journal of Nutrition121(6), 654-661.

Grassi, D., Ferri, C., & Desideri, G. (2016). Brain protection and cognitive function: cocoa flavonoids as nutraceuticals. Current Pharmaceutical Design22(2), 145-151.

Grassi, D., Socci, V., Tempesta, D., Ferri, C., De Gennaro, L., Desideri, G., & Ferrara, M. (2016). Flavanol-rich chocolate acutely improves arterial function and working memory performance counteracting the effects of sleep deprivation in healthy individuals. Journal Of Hypertension34(7), 1298-1308.

Kord-Varkaneh, H., Ghaedi, E., Nazary-Vanani, A., Mohammadi, H., & Shab-Bidar, S. (2018). Does cocoa/dark chocolate supplementation have favorable effect on body weight, body mass index, and waist circumference? A systematic review, meta-analysis, and dose-response of randomized clinical trials. Critical reviews in food science and nutrition.

Moreira, A., Diógenes, M. J., de Mendonça, A., Lunet, N., & Barros, H. (2016). Chocolate consumption is associated with a lower risk of cognitive decline. Journal of Alzheimer's Disease53(1), 85-93.

Oberrauter, L. M., Januszewska, R., Schlich, P., & Majchrzak, D. (2018). Sensory evaluation of dark origin and non-origin chocolates applying Temporal Dominance of Sensations (TDS). Food Research International111, 39-49.

Pereira, T., Bergqvist, J., Vieira, C., Sveälv, B. G., Castanheira, J., & Conde, J. (2019). Randomized study of the effects of cocoa-rich chocolate on the ventricle–arterial coupling and vascular function of young, healthy adults. Nutrition63, 175-183.

Santiago-Rodríguez, E., Estrada-Zaldívar, B., & Zaldívar-Uribe, E. (2018). Effects of Dark Chocolate Intake on Brain Electrical Oscillations in Healthy People. Foods7(11), 187.

Scaramuzza, A. E., & Zuccotti, G. V. (2015). Dark chocolate consumption and lower HbA1c in children with diabetes: Direct cause or pure happiness?. Clinical Nutrition34(2), 333-334.

Socci, V., Tempesta, D., Desideri, G., De Gennaro, L., & Ferrara, M. (2017). Enhancing human cognition with cocoa flavonoids. Frontiers In Nutrition4, 19.

Sumiyoshi, E., Matsuzaki, K., Sugimoto, N., Tanabe, Y., Hara, T., Katakura, M., ... & Shido, O. (2019). Sub-Chronic Consumption of Dark Chocolate Enhances Cognitive Function and Releases Nerve Growth Factors: A Parallel-Group Randomized Trial. Nutrients11(11), 2800.

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