With the increasing awareness of health, people are becoming more and more cautious about the choice of food additives.Tagatose is a kind of natural rare sugar, which has similar sweetness to sucrose, but only one-third of the calories of sucrose. It is especially suitable for weight-loss people and diabetic patients.The low glycemic index of tagatose helps stabilize blood sugar levels and, as a prebiotic, improves the gut flora and makes it healthier.In addition, tagatose does not cause caries and is more friendly to teeth.Tagatose offers a less burded-down sweet option than regular sugar, meeting the modern consumer's need for a healthy diet. However, the yield of tagaose limits its wider application, so our project is dedicated to increasing its yield through scientific research.
In order to better complete our work and achieve our goals, we carried out a series of human practice activities. We started with a brainstorming session to discuss project stakeholders, implementation plans and approaches. Later, we consulted large enterprises and researchers, conducted extensive social research , obtained user feedback to understand what society brings to us, how we can improve the program, and how our program affects society.
The whole human practice part is cohesive, closely connected to society, and responsible to the world.
To make sure our project is accountable to the world, we held a brainstorming session where we discussed a range of issues. The discussion results and summary are as follows.
Figure 1 Pictures from when we were brainstorming
Which resources or communities should we consult to ensure we are prioritizing appropriate values in the context of our project?
1.Masses
2.Researchers in Synthetic Biology
3.Industry professionals
4.Government regulations
Figure 2 The communities we consulted and their priorities
What evidence do we have to show that our project is responsible and good for the world?
1.Communication with experts
2.Questionnaires
3.Feedback form users
4.Public acceptance
Figure 3 Evidence that our project is responsible and good to the world
How do we envision others using our project?
1.Health and Nutrition
2.Individual Consumers
3.Food and Beverage Industry
4.Pharmaceutical Industry
5.Research Institutions and Universities
6.Education
7.Investors
Figure 4 The way others use our project
How would we implement our project in the real world?
Step1.Conduct market research to assess market demand;
Step2.Review current research progress through literature research;
Step3.Design appropriate experimental methods;
Step4.Communicate with relevant individuals and teams to refine experimental methods;
Step5.Seek partnerships, invest in production, and establish collaborations with other companies in the industrial chain;
Step6.Apply for relevant food safety certifications;
Step7.Boost promotional efforts to increase visibility.
What impact will our project have?
1.Advance research in biomanufacturing and synthetic biology research;
2.Improve human health levels;
3.Develop the field of education;
4.Facilitate the tagatose supply chain, reduce costs, and increase efficiency;
5.Expand the market size for tagatose, reducing market prices;
6.Provide consumers with new sugar substitute options and improve their quality of life.
What values-environmental, social, moral, scientific, or other-inspired our project?
1.Health Orientation: According to the International Diabetes Federation (IDF), the number of diabetes patients in China reached 140 million in 2021, accounting for 26.2% of the total number of patients worldwide, of which 96% were type 2 diabetes patients. The World Health Organization reports that more than one billion people worldwide suffer from obesity. People's health problems are becoming more and more serious, and the main reason is insulin resistance and intestinal microbial imbalance caused by high-sugar diets.
2.Social responsibility: With the acceleration of the pace of life, people increasingly rely on fast food to meet their nutritional needs, despite its high content of fats, salt, and sugar.It is difficult for us to change the current fast pace of life, but we can start from changing the food we consume to minimize the health risks associated with fast food.
3.Environmental sustainability: Currently, due to cost considerations, the main production method of tagatose is chemical synthesis. While biosynthesis is more environmentally friendly, it is significantly more expensive—about ten times more, according to interviews with relevant companies. To reduce the cost of biosynthesis, the main method is to choose cheaper substrates or improving the conversion rates. Our project focuses on improving the conversion rates.
4.Sustainable development: Minimize harm and pollution to the environment while developing new approaches.
Which communities will be most interested in or most affected by our project?
1.Diabetic patients:The development of sugar substitute products is closely related to their health. The development of high-yield and low-priced sugar substitutes will be of great benefit to their lives and health;
2.Fitness and weight loss individuals: While many sugar substitutes are developed, their low production volume and high prices remain a burden for those wanting to manage their sugar intake. Reducing costs and increasing efficiency would be beneficial for them.
3.Companies producing tagatose: Our experiments have optimized the production pathway for tagatose, improving production efficiency and reducing costs;
4.Industries producing other sugar substitutes and traditional sugars: With the decrease in tagatose costs, other sugars will need to lower their prices or find alternative methods to improve production efficiency to maintain market share.
5.Primary and secondary schools: Students can receive cutting-edge scientific and technological knowledge at an age when they are most receptive to knowledge. Additionally, fostering healthy living habits early can help reduce health issues later in life;
Which communities will be left out or negatively impacted if our project succeeds?
1.Some traditional sugar industries;
2.Other sweetener industries;
3.Individuals who cannot afford high-priced sweeteners;
4.People with concerns about synthetic biology synthesis methods.
How can we use our Human Practices work to inform our team's ethical, technical, safety and/or communication decisions?
1.Understand the needs and values of different groups, ensuring those factors are incorporated into the team's decision-making process.
2.Emphasize ethical consideration, ensuring that research and production processes meet ethical standards and respect the rights and dignity of all participants and audiences.
3.Consider safety in technical decisions, ensuring that all operations meet safety regulations to reduce risks to people and the environment.
4.Demonstrate transparency and integrity in communication strategies, sharing project information in a timely and accurate manner to build public trust.
Who are the stakeholders of our project?
Upstream:
R&D teams(With the advantage of pioneering, making it difficult to be surpassed by other developers in a short period of time, gaining fame and fortune);
Universities(Develop new methods, potentially involve commercial transfers of technology and patents);
Raw material suppliers(Change the original expensive raw materials to make the raw material manufacturers of the newly used cheap substrates profitable);
Equipment suppliers(Provide equipment to make profits during the production process);
Midstream:
Manufacturers (Reduce costs, save time, and enhance profitability);
Enterprises (Receive reputation, get popularity benefits);
Investors (Receive dividend benefits from their investments.);
Downstream:
Consumers including diabetics (Get health benefits);
Retailers (Pocket the difference);
Media (Promote new technologies and new products to gain traffic benefits);
All people(Benefit from environmentally friendly outcomes);
Figure 5 Stakeholders in our project
How can we optimize the entire process in future production?
Figure 6 A loop of future use and optimization
Who are the end users of our project?
1.People who have diabetes or need to control their blood sugar
2.Individuals with corresponding medical needs
3.Fitness enthusiasts concerned with health and sugar control
4.Adolescents and children who love to eat sugar and want to avoid dental and intestinal problems
As a new low-calorie sweetener, tagatose has potential health and environmental advantages, which are consistent with global health initiatives and sustainable development goals. Therefore, many countries have introduced relevant policies to support the development of tagatose in the food industry.
In 2001, the Joint FAO/WHO Expert Committee on Food Additives (JECFA) recognized tagatose as a safe sweetener. Subsequently, tagatose was approved as a food additive by the U.S. Food and Drug Administration (FDA) and passed the "Generally Recognized as Safe (GRAS)" certification, which considered tagatose to be generally recognized as safe and allowed to be added to food. In 2014, China’s Health and Family Planning Commission issued an announcement approving tagatose as one of six new food ingredients, which could be used in foods other than infant foods. Tagatose officially entered the Chinese food industry. In addition, on June 24, 2019, the State Council issued the Opinions of Healthy China Action, and the “Healthy China Program (2019-2030)” at the national level,which explicitly set forth the strategic goal of "sugar reduction". Among them , food producers were clearly advocated to replace sucrose with natural sweeteners and sweeteners approved by food safety standards and scientifically reduced the sucrose content in processed foods. This national-level support was intended to foster the development and growth of the tagatose and alternative sweetener industries.
Through the recognition and approval of governments and international organizations, the safety and health benefits of tagatose have been widely verified and promoted. This provides a solid policy foundation for our project, relying on which we can promote the application of tagatose in the food industry more effectively to achieve broader social and health benefits.
Figure 7 JECFA Certification
Figure 8 GRAS Certification
Figure 9 Announcement by the National Health and Family Planning Commission of the People's Republic of China
Figure 10 Healthy China Program (2019-2030)
In the early stage of the project, we designed a questionnaire to investigate public usage of sugar substitutes and distributed it through social media such as WeChat. The questionnaire consisted of 17 questions with logic jump, and we eventually collected 690 valid responses. By analyzing these results, we were able to draw some conclusions and gain preliminary insights into the public’s use of sugar substitutes.
Firstly, most respondents were familiar with sugar substitutes, with 68.26% reporting regular use in their daily lives, mainly stevia, xylitol and erythritol, accounting for 61.15%, 57.96% and 52.44% respectively. These sugar substitutes were primarily used in baking. The main reason for choosing sugar substitutes is to control blood sugar, and the taste of sugar substitutes also met their expectations. Among those using sugar substitutes, most people used sugar substitutes every day, accounting for 73.89%, and only a small percentage used sugar substitutes occasionally. This indicated a certain level of demand for sugar substitutes among the public.
The survey also revealed that 60% of the public believed that sugar substitutes were beneficial for health and supported their promotion and development. At the same time, they also expressed a desire for more information about sugar substitutes, especially their health benefits, usage methods and potential side effects. In addition, we gathered their expectations and suggestions on sugar substitute products, mainly focusing on lower product prices, higher safety and wider availability.
Furthermore, 69.28% of respondents had heard of tagatose in real life or through social media, recognizing its main advantages as a low glycemic index and taste close to sucrose. Notably, 62.9% of respondents clearly expressed their willingness to try tagatose, which shows that the promotion of tagatose has achieved certain results among the public. However, concerns remained regarding the safety, price, and taste of tagatose. Moving forward, we should prioritize publicizing the safety of tagatose to help alleviate doubts and misconceptions.
Overall, the public has a wide range of knowledge and use of sugar substitutes, and there is a positive attitude towards their health benefits. While tagatose, as a new sugar substitute, has garnered significant attention, further promotion is still needed. By addressing these concerns, we hope to promote the wider application of sugar substitutes and tagatose in the market to meet consumer demand.
After we released the trial recruitment for tagatose through online platforms, we received a large number of enthusiastic responses from users. Once the tagatose samples were sent to the participants, we collected detailed feedback from them, which helped us better understand the practical application of tagatose in daily life.
Many users expressed great satisfaction with the taste of tagatose. They found that the sweetness of tagatose is similar to regular sugar but without the cloying sensation that sometimes comes with it. Some users specifically mentioned that the sweetness of tagatose felt more natural, especially when used in beverages. For instance, several people who used tagatose in coffee or tea reported that the original flavors were preserved without the strange aftertaste often associated with other sugar substitutes. Additionally, some baking enthusiasts shared that tagatose worked well as a substitute for traditional sugar in their recipes, resulting in a cleaner and more natural sweetness in their baked goods.
The feedback regarding health benefits was also very positive. Many participants, especially those who need to manage their blood sugar levels, found that tagatose had minimal impact on their blood sugar. Some diabetic users mentioned that after replacing regular sugar with tagatose in their daily diet, they experienced significantly less fluctuation in their post-meal blood sugar levels. This allowed them to enjoy sweet flavors without worrying about potential health risks. Fitness enthusiasts and those focused on healthy eating also appreciated that tagatose enabled them to maintain a low-sugar diet while still enjoying sweet flavors, greatly enhancing their overall dietary experience.
However, some users did offer suggestions. A few people noticed that when used in high-temperature baking, the sweetness of tagatose might slightly diminish. Although this was not a major issue, it could require some adjustments in specific cooking scenarios. Nonetheless, the vast majority of users reported that tagatose performed excellently in their everyday use.
Overall, user feedback on tagatose has been very positive, especially in terms of taste and health benefits. This trial activity verified the practical application potential of tagatose and proved that our project was meaningful.
Figure 11 Feedback from our testers
Introduction:Chunming Zhou Nanjing Haiming Microbial Technology Co.
After interviewing Chunming Zhou, the head of a leading sugar manufacturing enterprise, to understand the advantages and disadvantages of biological manufacturing and chemical manufacturing to produce sugar substitutes, we determined to use biological methods to produce tagatose.
Figure 12 Interview process
Figure 13 Junping Zhou
Introduction: Junping Zhou, male, PhD, Distinguished Associate Researcher, Master's supervisor. She is mainly engaged in the topics of rational design and targeted modification of enzymes, biological cascade reaction and metabolic engineering, etc. She has published papers in SCI journals such as ACS catalysis, Advanced Materials, Current Opinion in Chemical Biology, Biotechnology Journal, etc. with total impact factor and co-authorship. He has published papers in SCI journals with impact factor greater than 50, and has been granted several patents.
In our efforts to enhance the activity of tagatose-4 epimerase, we encountered difficulties in selecting appropriate mutation sites. For this reason, we consulted Professor Junping Zhou. He suggested that when conducting rational design of enzymes, appropriate bioinformatics tools such as AlphaFold can be selected to predict the three-dimensional structure of the enzyme, and molecular docking can be used to simulate the interaction between the enzyme and the tagatose substrate.He emphasized that key residues in the active site need to be precisely modeled and optimized to ensure that the mutated enzyme can bind more effectively with the substrate after the modifications.
Figure 14 Rational Design and Optimization
Figure 15 Zheng Xu
Introduction:Zheng Xu, male, PhD, MSc supervisor. In his scientific research, he studies food biotechnology, including biosynthesis methods of functional monosaccharides and oligosaccharides, development of industrial enzymes and industrialisation technology of microbial cell factories; he has published nearly 40 academic papers and filed more than 10 invention patents.
After reviewing a large amount of literature, we discovered that naturally occurring tagatose-4 epimerase has a problem of low catalytic efficiency, so we consulted Professor Zheng Xu. He believed that mining new enzymes is an important approach to improving the efficiency of tagatose conversion. Professor Zheng Xu pointed out that the production of tagatose relies on the efficient catalysis of specific enzymes. Through enzyme mining, it is possible to find new enzymes with stronger catalytic abilities or better adaptability, especially those that can operate stably under industrial conditions.
In discussing how to more effectively mine new enzymes, Professor Zheng Xu emphasized the role of phylogenetic trees. By analyzing the evolutionary relationships of different enzymes, we can predict which enzymes might have functions related to tagatose conversion and further optimize their activity through directed evolution. Additionally, phylogenetic trees can help us identify enzymes that adapt to extreme environments, which is particularly important for enhancing the tolerance of enzymes in tagatose production.
Professor Zheng Xu suggested that we could combine phylogenetic trees with enzyme mining techniques to search for novel enzymes related to tagatose conversion and apply them in subsequent directed evolution experiments to improve conversion efficiency.
Figure 16 Enzyme Mining
Figure 17 Xu Li
Introduction:As an associate professor and master's tutor in the College of Food Science of Shihezi University, Li is committed to the cause of education, practicing the purpose of ‘educating people for the Party, educating talents for the country’, and has made remarkable achievements in teaching, scientific research, and social service, and has been highly evaluated by the teachers and students.
During the expression of tagatose enzymes, we encountered the issue of inclusion bodies, which caused improper folding of the enzymes. This created difficulties for subsequent studies on the enzymatic properties. To address the problem, we consulted Professor Xu Li.
Professor Xu Li analyzed several major causes of inclusion body formation, such as excessive protein hydrophobicity leading to aggregation and misfolding caused by rapid protein synthesis rate. He suggested that we could optimize codon preference to reduce rate-limiting steps during translation, thereby minimizing the formation of inclusion bodies. Additionally, he mentioned that using molecular chaperones and fusion tags could assist in proper enzyme folding and improve its solubility.
In response to our suggestion on how to further improve the solubility of the enzyme, Professor Xu Li recommended employing a secretion expression system to screen out mutants that are easier to fold correctly, thus improving the soluble expression of tagatose-4 epimerase.
Introduction:Yang Liu, Nanjing Haihe Biotechnology Research Institute Co.
In our experiments, we performed iterative mutations on tagatose-4-epimerase ,but encountered some challenges regarding how to enhance enzyme activity. For this reason, we consulted Professor Yang Liu. He explained that by means of point mutations, one can change the active site of the enzyme, the polarity of the active pocket, the size of the pocket channel, or enhance the hydrogen bonding interaction between the substrate and the enzyme to improve the conversion efficiency.
We also faced issues with fronting and tailing peaks in our HPLC analysis. Professor Yang Liu analyzed these problems, pointing out that fronting peaks might result from sample overloading, while tailing peaks could be related to the condition of the chromatography column. To resolve these issues, he suggested reducing the sample amount and attempting to backflush or replace the column.
In the future, we hope to further enhance enzyme activity through directed evolution. However, directed evolution involves complex screening processes. Therefore, Professor Yang Liu suggested that we can integrate high-throughput screening techniques to identify mutants that can significantly improve the activity of tagatose-4-epimerase.
Figure 18 Mutation
