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Low cost technologies and traditional ingredients for the production of affordable, nutritionally correct foods improving health in population groups at risk of poverty

Final Report Summary - CHANCE (Low cost technologies and traditional ingredients for the production of affordable, nutritionally correct foods improving health in population groups at risk of poverty)

Executive Summary:
Recent studies have proved that in the European Union more than 80 million people live in households at risk of poverty (ROP). A further 40 million live in households that are not at risk of poverty, but are defined as jobless and/or materially deprived according to the two newly agreed headline indicators, so total is 120 million for the EU-27.
The CHANCE project has found that, with cut-off point at 60% of median equivalised income, Lithuania has the highest ROP rate among the studied countries, with 20.6% of national population, then there is Italy (18.4%), Serbia (17.7%), UK (17.3%), Finland (13.8%) and Denmark (13.1%). Females from all the studied countries are at a higher ROP than males and the population younger than 24 years is evidently at higher ROP rate in all studied countries. From family structure data, single person, one adult older than 65 years, single parent with dependent children and single female separated are the types of household with the highest ROP in all the chosen countries (ROP > 20%).
From literature data (1009 titles and abstracts from Medline and 1430 from WoS), the specific unhealthy dietary habits in ROP population groups have been investigated and have been found to be: i) meal patterns (e.g. missing meals), ii) micronutrient inadequacy (e.g. low iron and high sodium), iii) high intake of high energy density foods (e.g. soft drinks, fast food, fatty meats and fatty meat products, refined grain, added fats), iv) low intake of high nutrient density foods (e.g. fresh fruit and vegetables, lean meats, fish, low fat dairy products, whole grains bread and bakery).
Dietary information (through FFQ and average daily nutrient intakes calculated from multiple pass 24h dietary recalls) have been collected within the CHANCE countries in order to correctly describe the dietary patterns of ROP populations as compared with an affluent population (AFF). To identify the main nutritional criticalities, food consumption patterns and average daily nutrient intakes have been compared with recommendations. Some common trends can be highlighted in all ROP groups: consumption of cereals, meat products, eggs and sugar is higher in the most of ROP groups than in the corresponding AFF; consumption of fruit and vegetables is lower in the ROP groups in all partner countries; ROP people tend to eat less dairy products in almost all countries; the contribution of total fat towards dietary energy is higher than the recommendations in all countries, with no significant differences being found between the two different population groups; average daily intake of protein is adequate in ROP and AFF in all countries; significantly lower intakes of key vitamins has been found in ROP compared with AFF group (particularly folate, vitamin A, vitamin D, and minerals such as iron); in all 5 countries, intake of sodium exceeds the recommendations in both groups.
Although from nutritional data it could be concluded that economic status is not the main determinant of the onset of nutritional criticalities, the purchasing power could be important for the correction of criticalities by nutritionally improved foods, that must be as cheap as possible to impact the nutritional status of ROP. Dietary data on consumption of 39 foods and food groups, reported by ROP groups in the 5 countries participating in CHANCE project, has showed that the most consumed foods are white flour (for bread, sandwich or pizza use), ham, cheese (both soft and hard) and tomato (in the form of paste, pomace, ketchup or juice).
Thus, the CHANCE project has developed low-cost ingredients suitable for the preparation of foods for ROP consumers: cooked ham with 5 % liver; tomato paste; pizza-cheese; fibre-enriched bran. The prototypes of some affordable and tasty CHANCE foods have been developed as well: three different kinds of fibre-rich bread, berry soup, berry jam and berry shot. Moreover, the CHANCE pizza has been developed as RTE meal, containing fibre enriched dough, cooked ham with 7 % liver (rich in iron and vitamin D), tomato paste and pizza-cheese. The involvement of representative SMEs in the CHANCE consortium has allowed the production testing of all CHANCE food prototypes directly in their existing industrial plants, thus confirming that the CHANCE food products so far developed do not require any substantial upgrade of the available structures.
Project Context and Objectives:
CHANCE is a collaborative small-scale research project in the FP7 Cooperation Theme 2 (Food, Agriculture, Fisheries and Biotechnology) whose topic was “Health-value-added food products for populations at risk of poverty”.
Poverty is one of the main causes of food unaffordability, therefore population groups at risk of poverty (ROP) are also at risk of malnutrition or nutritional insufficiency due to food poverty.
Unaffordability can be defined as the inability to obtain healthy food, either because of subjective barriers (e.g. the skills to create healthy meals, cultural barriers) or objective barriers (e.g. lack of availability of healthy foods in local shops).
The general belief is that, as the effect of a complex combination of cultural and economic factors, people on low incomes have the lowest intakes of fruit and vegetables and often the highest intake of energy-rich foods. As a possible consequence, they have higher risk of suffering from diet-related diseases such as cancer, diabetes, obesity and coronary heart disease (CHD). Food poverty is, in effect, also associated with an abuse of “junk” food and a lack of healthy food.
The objectives of CHANCE project have been directed to improve the diet of risk-of-poverty (ROP) population groups through the development of affordable and tasty, nutritionally correct foods. This has been addressed not just by simply focusing on the formulation of new food prototypes possessing a generic correct nutrient profile, but through a more complex strategy, starting from the definition of the specific traits and needs of those population groups at risk of poverty in the EU (varying from country to country), and going on to investigating the factors that, together with affordability, drive their food choices and thus defining the correct food products for all these needs.
On the basis of a consumers’ approach, the project has been addressed to discover ROP-specific barriers, including perceived unaffordability.
Thus, the CHANCE project has considered the dietary habits of the subjects so far recruited for both administration of surveys and for collection of urine sample, in order to improve their preferred food products, keeping similar organoleptic characteristics, while increasing their nutritional value. In this way, the designed foods are not only affordable and nutritionally correct, but also accepted by consumers and with a high probability to be integrated in their usual diet.
CHANCE consortium has been composed by a multidisciplinary team of universities, research institutes, small-and-medium sized enterprises (SMEs) from the food and drink sector as well as two European non-profit organisations in the fields of communication and standardisation. With such a broad composition, the consortium has aimed at developing traditional, nutritionally enhanced food products, by applying low-cost optimized manufacturing technologies in their food processing. The expected and achieved final output has consisted of commonly consumed food prototypes that have taste similar or better than equivalent brand-leading products on the market.
The CHANCE project has been developed along 7 work packages (WP1-7), coordinated by the management WP8, and has been articulated into six logical steps aiming at: 1) understanding the nutrient imbalance and the barriers to healthy nutrition of population at ROP; 2) selecting ingredients or primary foods from a list of local highly consumed products; 3) designing low cost foods and meals improving the nutrients balance of traditional foods; 4) assessing the quality of ingredients and final products; 5) evaluating the nutritional characteristics and consumers’ acceptance of the new food products and meals; 6) testing the production plans of newly developed foods & packages and evaluating their production feasibility.
Socio-economical, nutritional, biochemical, technological and industrial tasks have been sequentially carried out in order to understand and overcome the barriers to healthy nutrition.
The identification of the perceived and actual underlying cultural and economic barriers have been achieved by administrating questionnaires and by organising focus groups; on the basis of the surveys’ output, the dietary habits and the consequent nutrient balance of the identified selected representative population groups at ROP heve been investigated; then, the actual impact of those criticalities on the metabolic profile of the selected subjects has been assessed.
Afterward, the ingredients and the primary foods have been selected from a list of local products which, in relation to their composition and their economic impact and local importance, represented the CHANCE project priorities in the reduction of malnutrition, as observed in the first logical step.
Low cost foods and meals have been developed at a pilot scale by applying formulation design, through predictive mixture methodologies.
Quality assessment and shelf-life studies of ingredients and final products have been performed by using appropriate methodology chosen according to the physico-chemical composition of the prototypes. The assessment of the effects of formulation and processing on digestibility, using human in vitro digestion models, has been also carried out.
Ten CHANCE food prototypes have been selected and produced at the pilot industrial scale by the SMEs partners and by one large food company: cooked ham, pizza-cheese, tomato paste, pizza, three berry-based products and three enriched breads. These foods offer affordable healthier choices to low income consumers, by optimising their intake of micronutrients, fibre, and bioactive compounds, whilst reducing their intake of calories from caloric sweeteners, refined carbohydrates, and fat.
Low cost ingredients and final formulations/products have been analysed for their molecular profiles, in order to establish if processing affects the molecular composition of the starting ingredients in the final formulation. It has been found that CHANCE products kept their higher nutritional values with reference to commercial counterparts. This means that the newly produced foods have the wanted positive characters, together with a low cost and good consumers' acceptance.
The overall feasibility of the designed foods and their production has been economically evaluated, also in terms of the life cycle assessment (LCA).
At the end, the benefit offered by CHANCE project has been twofold: i) consumers at ROP are provided with new healthy foods that could easily substitute those that they are traditionally consuming; ii) SMEs are provided with exploitable models for healthy foods production, targeting new market niches, thus boosting the regional economy and creating new jobs.
In conclusion, the CHANCE project has reached its main goal (i.e. develop foods prototypes addressed to overcome malnutrition in people at ROP) as well as other positive social and economical targets (enhanced industrial competitiveness and jobs creation).
Project Results:
The CHANCE project has been organized by following a work-packages structure also functional to the assignment of specific tasks and responsibilities to all teams and principal investigators.
Seven WPs have been planned aiming at synergistically reaching the expected results. An additional package (WP8) has been also included to supervise and manage all activities.

WP1 - Identification of population groups at risk of poverty and ways to overcome barriers to unhealthy dietary habits along food chain
The activities addressed by WP1 within the CHANCE project, have been firstly focused on:
1) the definition of terms and criteria for the identification of the population at risk of poverty; 2) the analysis of specific data on income and socio-economical well-being from the considered European Countries; 3) the identification of ROP groups in those studied populations according to social and geographical criteria; 4) the analysis of information on socio-economic barriers to correct nutrition in ROP population groups.
In Europe, poverty is officially defined, in relative terms, as the percentage of individuals living in a household whose equivalent income is below the poverty threshold. This threshold is set differently in each country (but coherently defined as equal to 60% of the national median equivalent income), in order to consider the various national income inequalities. Usually, the common threshold applied to at-risk-of-poverty indicators in the EU is that of 60% of median equivalised disposable income after social transfers.
Recent studies have proved that in the European Union more than 80 million people live in households at risk of poverty. A further 40 million live in households that are not at risk of poverty, but are defined as jobless and/or materially deprived according to the two newly agreed headline indicators, so total is 120 million for the EU-27.
Definitions of low income population are often different, depending on the Country or even on the author. The CHANCE project has applied the most updated methods to define “population at risk of poverty”. In effect, the most recent definition of ROP focuses on income, since it is undoubtedly a good proxy of the living standard of an individual or a family; thus, the income insufficiency approach has been employed and proved very effective in guiding policy action and raising public concern for poverty. The method so far adopted to measure the ROP condition combines the EUROSTAT definition and a concept of material deprivation.
Among the studied countries (Italy, Serbia, UK, Denmark, Finland and Lithuania), the results have showed that:
i) Lithuania has the highest risk of poverty rate with 20.6% of national population with cut-off point at 60% of median equivalised income, then there was Italy (18.4%), Serbia (17.7%), UK (17.3%), Finland (13.8%) and Denmark (13.1%); ii) females from all the studied countries are at a higher risk of poverty than males and the population younger than 24 years is evidently at higher risk of poverty rate in all studied countries. Females younger than 24 years of age and female older than 65 are at the highest risk of poverty (ROP) in all six studied countries; iii) from family structure data, single person, one adult older than 65 years, single parent with dependent children and single female separated are the types of household with the highest risk of poverty in all the chosen countries (ROP > 20%). The single person older than 65 is also in high ROP (in 5 countries higher than 25% and in 2 countries higher than 40%). Single parent is in the high ROP (more than 30% for 4 countries); iv) in the UK, ethnic minority groups (Bangladeshi and Pakistani) is within the lowest income population. Unemployment and inactive people are most affected by ROP condition in all 6 countries; v) pre-primary, primary and lower secondary education are clearly identified as characterizing those groups with at highest ROP; vi) material deprivation by gender and age indicates that female is in the highest ROP. However, in Lithuania female older than 65 year and the population from 18-64 year, respectively, are identified as groups in severe material deprivation; vii) people from the lowest income quintile spend most of the money on food and non-alcoholic beverages.
The ROP groups to be recruited in the five chosen countries have been selected using these results and the following criteria : 1) in each country, only groups with ROP percentage equal or higher to EU percentage have been taken into account; 2) adults (25-64 years) have been considered as a whole. This is in agreement with adequacy level of nutrient intake; 3) the evaluation of barriers to healthy nutrition, as well as of dietary habits, in children and adolescents is very difficult because of possible bias such as not independent choice or social programs in schools. Furthermore, the range spanning from 0 to 18 years old considers individuals having very different nutritional requirements. Therefore, it has been decided to recruit adult people only (>18 years old).
In this way, the ROP groups selected for the CHANCE study have been: 1) 25-65 years old: Italy (female), Lithuania and Serbia (both genders); 2) > 65 years old: Finland (both genders), UK (both genders, ethnic minority)
After the choice of population groups to be studied, the barriers to healthy dietary habits have been investigated. Many studies have reported that nutrition can play an important role in the development or prevention of various diseases. This implies that people with better dietary habits will show a healthier status. However, a high quality diet (nutrient rich, low in energy) is often more expensive and thus, it is more abundant in high socioeconomic groups, whereas a high-energy and nutrient-poor diet is generally more consumed by those with limited economic resources. This could partially explain how economic deprivation contributes to nutritional poverty. Inequalities related to food choice are determined by individual preferences, socio-economic and cultural factors. The most important determinants of diet choices so far recognised are social position, income and education.
Nonetheless, the socio-economic position can partially explain the difference in food pattern. Food poverty is defined as ‘the inability to acquire or consume an adequate quality or sufficient quantity of food in socially acceptable ways, or the uncertainty that one will be able to do so’. This definition thus includes many terms that are allied to modern malnutrition: ‘affordability’ related to income, ‘accessibility’ linked to geographic, practical and cultural issues and ‘social acceptability’ connected to social norms.
Accordingly, the specific unhealthy dietary habits in those population groups have been investigated and found to be: 1) low rate of breastfeeding; 2) meal patterns (missing meals, e.g. breakfast); 3) low nutrient density score; 4) high energy density; 5) micronutrient inadequacy; 6) excessive intake of micro/macro nutrients; 7) high intake of soft drinks, fast food, fatty meats and fatty meat products, refined grain, added fats and salt; 8) low intake of fresh fruit and vegetables, lean meats, fish, low fat dairy products, whole grains in bread, vegetable oils; 9) high intake of calories that originate from saturated fat, sugar, salt, sodium; 10) low intake of minerals (iron, calcium, potassium, magnesium); 11) low intake of vitamins such as folate, vitamin D, vitamin C, carotene, vitamin A and vitamin E; 12) low intake of animal protein, dietary fibre, antioxidants, phytochemicals.
Therefore, it has been necessary to identify other determinants of unhealthy dietary habits (beyond the socio-economic position) associated with food choice and nutritional habits in ROP and in low income populations, in order to understand how to overcome these barriers.
Open access and grey literature documents on nutritional health inequalities have been consulted. The resulting determinants found to cause unhealthy dietary patterns in ROP and low-income population groups are: poverty (income distribution by categories); employment; emigration or immigration; education; demographic factors (gender, age, marital status); religion; occupation; urbanisation; food accessibility and preference.
To prove these findings, the chosen population mentioned above have been questioned through interviews and focus groups. The CHANCE study has suggested that ROP consumers perceived different barriers for healthy eating depending on the country and population group.
The investigated subjects most often have agreed with the following barriers for their healthy eating: 1) poor affordability of healthy foods (Lithuania and Serbia); 2) preference for familiar and traditional foods (Finland, Serbia, UK and Lithuania); 3) eating unhealthy delicacies when upset, stressed of tired (Italy, Serbia and Lithuania); 4) different taste preferences of family members (Serbia and Lithuania).
Many barriers are often shared equally by both ROP and non-ROP (AFF) respondents, suggesting that these may be obstacles for healthy eating, but not just for the ROP group. However, a few barriers have been reported significantly more frequently by ROP than by AFF subjects, as indicated at least by 30% of the ROP respondents. These are poor affordability to healthy foods (Lithuania, Serbia) and preference for familiar and traditional foods (Finland, UK).
In addition, in Italy, Serbia and Lithuania, the ROP respondents have declared to perceive less control over their healthy eating than AFF respondents. For some reason, these consumers feel they are not able to follow a healthy diet even if they want to. This feeling may arise from experienced external barriers, such as scarce monetary resources and insufficient knowledge. Supporting this notion, in these same countries, the scores of dietary knowledge have been significantly lower among ROP than AFF respondents.
Results from the focus group studies have showed that there are four main ways to define healthy food: the presence of healthy substances (vitamins, fibre, proteins) or absence of the unhealthy ones (fat, salt, chemicals, pesticides, etc.), the level of processing (most people think the more a food is processed, e.g. frozen food, the more it has lost nutritional value), the needs of our bodies (and many subjects link this to taste, too) and the consequences of eating food (i.e. feeling good).
Another interesting result is that many interviewed subjects started perceiving the connection between well-being and a healthy diet after retirement or a health problem, as they focused the healthy choice in removing one bad habit, usually the one linked with their disease (e.g. sugar for diabetics, salt for people with hypertension). Many have also said that they were more motivated to healthy eating if connected to health care professionals, showing that being well-informed is a strong factor.
'Liking', obviously connected to taste, is the main driving force for food choices, since many respondents believe that healthy food also tasted less good. Affordability is another important factor in the food choice. Lithuanian informants, in particular, have mostly described situations in which to buy healthy food was not possible. Similar information was reported by Serbian and UK subjects. It is important to underline the fact that people wished for more reliable low-priced healthy food, with a low quantity of additives, but they thought this was almost impossible to produce. The barriers found to healthy eating have been demonstrated to be the same ones arising after the individual interviews.
An additional step has been consisting in the investigation on the barriers to the production of this type of food perceived from industries and retailers. A total of 53 respondents (32 food industries and 21 food retailers) have been interviewed, almost equally distributed among the studied countries (with the exception of UK). The food industries interviewed are mostly concentrated in the meat/fish production (38%), while the cereals/bakery category is the less represented (19%). Micro enterprises are the most represented (44%), followed by medium or big companies (31%). Medium size enterprises refer to 22% of the sample.
The 21 retailers interviewed apply to more than one typology of distribution type, the most represented being the traditional ones (48%) followed by large retailing non discounts (38%) and discounts (33%); medium or big retailers are the majority (52%).
People interviewed have had to complete a questionnaire with many topics. The first section in the questionnaire has aimed at obtaining information on the theme of nutritional and healthy food. Most of the respondents has given a definition of healthy food as a product not containing high amount of fat, sodium and sucrose or with high content of fibre and micronutrients. Moreover, the product should be simple, that is, it should be produced with few basic ingredients, of good quality and without additional ingredients for conservation. In many cases, the interviewees have used the term “natural” to define a healthy food, not containing additives or preservatives, containing a high quantity of “raw materials” of good quality to guarantee a safe preservation of the product.
Just like in the case of ROP people interviewed, respondents from retailers and industries mostly have answered that a healthy food must be processed as the least possible, and mainly through “natural” process production systems.
Nutrition is considered to be an interesting training subject for industries/retailers. This interest has been shown much more by food retailers than food industries even if in both cases the percentages were very high (nearly 80% and 90% respectively). This can be due to the different perception of retailers on the attention paid by consumers on these issues, and the major proximity to the consumers with respect to producers. The industries are in particular more affected by production processes, raw materials/main ingredients and prices of materials inputs.
Next step has been the analysis of the nutritional or health claims for foods produced or commercialized by industries or retailers, and the type of production (company brand, other brand, private label).
Foods with low fat content are produced by the 59% of industries and commercialized by 81% of retailers. As second best, products with low/no content of sucrose are produced by 47% of food industries and commercialized by 81% of food retailers. Next, there are low/no sodium products, produced by 44% of food industries and commercialized by 81% of food retailers. Retailers are interested in a bigger range of products than industries, and in particular in nutritional and health food. As a consequence they have put a major attention on the nutritional issue, as seen before. Anyway, the figures have shown that both industries and retailers have a high knowledge of nutritional and healthy food.
The majority of the industries have paid particular attention to the quality of the products, and the processing production system, considering them as main characteristics of their products. The analysis of the produced food has been deepened by investigating the characteristic of production categories, e.g. company label, other company label and private label, by the presence of market leaders in nutritional or healthy food and by using or not nutritional claims. The presence of market leaders is, generally speaking, medium with a low peak with regard to very specific type of nutritional or healthy foods such as those containing bioactive compounds (only 9% with respect to 31% of low/no fat food). Also, first followers are not particularly present and they do not exist when talking about food containing fibre, micronutrients and bioactive compounds. These data have been confirmed by retailers: nearly 70% have declared that only a percentage from 0 to 10% of the annual turnover is represented by healthy food; 19% have declared a percentage between 10% and 20%. Only 10% of retailers have declared a percentage above 50% represented by healthy food.
For what concerns future perspectives, low/no fat food, low saturated fat and low/no sodium food are the type of nutritional or health food that respondents have declared they are planning to invest on. The propensity of investment on healthy food seems to be prevalent in food retailers instead of food industries, with big differences in the investments according to type of nutritional and healthy food. The majority of companies believe that healthy food will be on average of other processed food (43%) or even above average (28%). Again, differences are present between food industries and food retailers, with more food industries than retailers thinking that healthy food in the future will be above the average of other processed food (34% industries vs. 19% retailers).
One of the main barriers to increase the production on healthy food is feasibility. The majority of food industries has considered that producing low cost healthy food is quite difficult (31% when summing extremely difficult and slightly difficult). Around a 19% has declared that it would be slightly or extremely easy. A lower percentage of food retailers think that producing healthy food is difficult; despite nobody considers it extremely easy, 19% of them consider it slightly easy.
At a general level, among food industries, difficulties in producing healthy food are considered strong when industries are micro (10 or less employees) or medium/big, with 250 or more employees. Almost a quarter (23%) of small enterprises so far interviewed has perceived the feasibility of low cost healthy food products slightly easy. Among the industries producing dairy/eggs, 51% of the interviewed have declared not difficult producing low cost healthy foods, from the technological and economical points of view. For meat/fish and cereals/bakery producers, it is possible to distinguish between two groups, with regard to the feasibility in producing low cost healthy food: the interviewed companies distributed almost equally between those expressing easy and difficult feasibility. Among fruit and vegetable industries a wide majority considers in some way technologically and economically difficult (from somewhat difficult to extremely difficult) to produce healthy food.
The last analysed aspect is the knowledge and importance of ROPs consumers for the industry and retailers. The consumers segments that is considered to potentially experience more growth and profit potential are those related to women and men over 50s, people with specific health problems, and also the elderly people. On the contrary the segment related to low-income people at risk of poverty has been considered the one with potential less growth and profit, as well as babies 0-3 years old. Similar trends are also present when looking to retailers answers. However, retailers have a very higher expectation in improving growth and potential profit for all the consumers’ segments, both in term of percentage rate and in term of average rate. Interestingly, it has to be noted that retailers have a significantly higher expectation, with respect to industries, also on the low-income people and ROPs, even if lower than for other segments.
Going into further details on which will be the future trends for consumption among people at ROP, the food products that are considered to have good future trends in the next 3 years are those with good nutritional density commercialized with no nutritional claim, ready-to-eat food and food with health claim/functional food targeting specific diseases. Similar trends are also provided by retailers.
As for the modality to obtain information on consumers’ trends, both food industries and retailers use market data from national/international marketing agencies. In addition, professional seminars/events are considered to be an important source of information, whilst informal sources of information, such as word of mouth, are used less.
Both food industries and retailers think that healthy RTE food can be an effective way to stimulate ROP consumers consumption of healthy food. Food industries think healthy ready-to-eat food can be effective up to extremely effective (66%), with a higher percentage compared to food retailers (57%), but they also presented negative answers: around 9% of industries have considered healthy RTE foods as not effective plus slightly effective against around 15% of negative answers from food retailers.
Food retailers also think that, at present, RTE food is not so compatible with healthy food and diet (more than 57% says slightly and somewhat compatible vs. 31% for industries). Only little percentages of food industries (9%) and retailers (10%) think it is extremely compatible. Industries believe that fresh healthy food could more easily increase ROP consumers consumption over healthy food, followed by healthy snack. On the contrary hot healthy RTE food is the one considered to have a lower possibility to increase consumers consumption. Similar trends are reported also by retailers.
The last objective of these interviews has been to find out the possible barriers to the production, commercialisation and distribution of low cost healthy food. For what industries concern, barriers seem to be mainly connected with the role and the relationship with retailers. As a matter of fact, the option that received more answers is related to the statement that the increasing power of retailers toward what foods will be commercialized impedes food industry to focus interest over low cost healthy food. Immediately after, it comes the option related to the lack of coordination and commercial agreement between industries and retailers, which limits their interest over low cost healthy food. This latter has been considered as a barrier also from retailers, highlighting the importance of the establishment of good relationship for joint working. However, at a general level, it can be said that retailers have a more positive view on solving possible barriers than industries. A better coordination is considered to be the main way to overcome barriers from both industries and retailers. Related to this, also the increased availability of ingredients for low cost healthy food is considered an important way to overcome barriers for both industries and retailers.
Within the second typology of barriers, ROP consumers' perceptions of high price for healthy food are perceived as a barrier both from industries and retailers. This result is also emphasized by the high number of enterprises respondents concerned about the ROP consumers' perception attributing a high price gap between healthy food versus their familiar food. In order to overcome these barriers the affordability of healthy food is considered to be a goal able to stimulate ROP and low income consumers interest towards this kind of food. In this case, industries have expressed a higher number of positive answer.
The second main barrier identified by industry and retailers is related to the high margin performance of other foods in comparison with low cost healthy food. However, industries considered this barrier more crucial than retailers because of the lack of private standards focused on healthy food production, commercialization and distribution, which limits consumers' interest on healthy food.
In order to overcome the identified main barriers, industries and retailers still have a similar approach considering decreasing prices of ingredients for low cost healthy food as a crucial way to increase low cost healthy food commercialization. When looking at market trends, the main barrier has been considered to be the insufficient industry and retailers competition towards healthfulness brand reputation. This is true for both industries and retailers. The other optional barrier – i.e. insufficient industry and retailers positioning strategy focused on low cost healthy food – has also been considered a crucial issue to be tackled.
As for the way to overcome these barriers, while industries focus on an increased positioning strategy of food industry and retailers, retailers consider also the other given options as crucial. For example, strengthening the supporting role of retailers in favour of public health campaign and healthy food consumption has also been considered as a crucial approach.
Within the item of private label, again similar answers can be observed in both industries and retailers even if it can be said that, at a general level, retailers seem to have a more positive view. The main barrier to invest on low cost healthy food is identified by both industries and retailers in the consumers' increasing interest for private label versus commercial brands. With regard to retailers' scarce interest on private label lines for low cost healthy food, differences between industries and retailers can be detected. Retailers consider this aspect as a less problematic barrier than industries. As a consequence, the option of inserting in the market low cost healthy food within private label is considered to be a crucial way to overcome these barriers more by industries than by retailers.
Within public policy and regulations, the lack of adequate public policy intervention over the promotion of healthy food for ROP and low income people is considered to be the main barrier by both industries and retailers, and the latter are the more negative towards this issue. Also, the lack of an official definition of healthy food is considered to be an important barrier and again retailers are those more negative.
However, the improvement of awareness, especially among consumers, is considered a strong way to overcome barriers by retailers. As second option, while retailers consider the funding of public campaigns and the implementation of public policy and public regulations, industries consider more important the identification of financial incentives and disincentives to levering the supply chain in the direction of low cost healthy food.
Finally, the easy access of consumers to unhealthy food is considered an important barrier by industries, whilst retailers have a more positive view. The way to overcome this main barrier is related to the improvement of ROP consumers' access to low cost healthy food, especially for what regards industries.
These interesting results have constituted the baseline for the subsequent work of WP2, aiming at translating this information into actual targets for novel foods, once true nutritional criticalities in the studied population has been assessed. In effect, the combination of the answers from ROP consumers and from industries and retailers has given the chance to translate the needs felt for new healthy food into attractive characteristic for target products.

WP2: Conversion of nutritional criticality data into targets for new foods
The results of WP1 have evidenced that the diets of low-income groups are likely to be inadequate. In the absence of any literature study addressing diet-related health inequalities within ROP groups, WP2 has been focused on this specific goal. Thus, in order to address the right nutrition-targeted action, aiming at improving the nutritional status and decrease the risk of diet-related disease in low-income populations, a critical assessment of available information on the main nutritional criticalities has been carried out. This task has been accomplished by systematic literature review and by qualitative analysis of the relevant studies from Medline and Web of Science (WoS) databases (1009 titles and abstracts from Medline and 1430 from WoS). Reports on dietary intake of food groups/subgroups/micronutrients in low-income groups compared to high-income groups, or to reference values, in adults in Europe have been analyzed. In addition, other sources were accessed, e.g. grey literature, EU projects research and papers, and national food and nutritional surveys.
Only a few countries have conducted country-specific dietary assessment of low-income people. These studies differ in methodologies and this hinders cross-country comparisons. In addition, suitable dietary intake methods are required to assess the diets of low-income groups. A recent publication on best practice dietary methods by the FP6 EURRECA project (www.eurreca.org) suggested that many of the methods used in low-income and immigrant populations have not been subjected to validation, and consequently may not demonstrate sensitivity and/or specificity when used in these groups. Therefore, in order to correctly improve the diets of low-income population groups, an overview of their nutritional criticalities has been prepared.
Thus, WP2 has focused the research activities on the study of existing information about unhealthy dietary patterns in low-income groups, including: 1) the consumption of food groups and nutrients in low-income populations in comparison to higher-income groups; 2) the identified micronutrient deficiencies; 3) the extent to which nutritional recommendations are met and 4) the guidelines for an optimal diet.
Nutritional criticalities, as emerging from reported food consumption patterns of low-income populations, have been identified by considering the low-income definition and the food groups and macro- and micronutrients which are prioritised for nutritional assessment.
A qualitative approach has been applied for the systematic analysis, by extracting the data on food groups or macro- and micronutrients intakes in low-income groups compared with high- income groups, and in comparison with dietary recommendations.
Food Consumption patterns of low-income groups across different European countries have been shown to present slight variations from one country to another, and the trends for consumption are consistent for most food groups.
For what the CHANCE countries concerns, the main criticalities found were: lower intake of wholegrain products, fruit and vegetables than in high socio- economic groups (Denmark); lower consumption of fresh vegetables and lower intakes of vitamin C and carotenoids (Finland); lower consumption of fruit and vegetables and lower intake of some nutrients (Italy); insufficient consumption of vegetables, fruits, cereal products, fish and fish products, and overconsumption of fats and confectionary products (Lithuania, from data on the general population because lacking on the low-income population groups); lower intake of vitamin D (Serbia, adults 30-75y); lower consumption of wholegrain bread, fruit and vegetables, lower intake of vitamin C and carotenoids, higher intake of Na and K (UK).
To summarize, in a general picture, it has been assessed that the main characteristics of food consumption and nutritional criticalities for low-income populations in Europe are: 1) low intake of fruit and vegetables, assessed either by comparing low-income groups to high-income groups or by meeting 5-a-day recommendation; 2) low intake of wholegrain food products (high intake of starchy food, potatoes and low intake of fibre); 3) slight-to-moderate increased likelihood of high fat intake (vegetable-based margarine on bread and oil for cooking) - where data are available, there is a consensus that higher fat contribution to total daily energy intake is found in low-income populations when compared to affluent groups, except in Poland; 4) low consumption of vitamin C, iron (in women), vitamin A, vitamin B12, and high sodium intake; 5) high intake of meat (in men) and meat products; 6) variable consumption of type and amounts of milk and dairy products; 7) less-frequent consumption of breakfast; 8) high intake of table sugar and sweets; 9) high intakes amongst women of crisps and snacks; 10) increased likelihood of developing obesity.
The CHANCE study has recruited the specific population groups emerging from the activity of WP1. Female adults (Italy), male and female adults (Serbia), elderly males and females (aged 65 yrs and above, Finland) and adult males and females from ethnic minority groups (UK) have been directly surveyed for malnutrition or nutrients imbalance. To identify the main nutritional criticalities, food consumption patterns and average daily nutrient intakes have been compared with recommendations. Additional data on physical activity and anthropometric measurements have been collected and analysed in relation to both the risk of being overweight/obese and to overall lifestyle patterns.
It has been found that overweight still remains one of the main nutritional criticality in EU. A high prevalence of overweight and obesity has been detected among both ROP and AFF groups, without significant differences among the two groups. Interestingly, the rates of overweight and obesity were lower than previously reported in Italy. Further data analysis is still in progress to evidence which changes in the dietary pattern could be at the basis of this trend inversion.
Data on dietary patterns collected through FFQ and average daily nutrient intakes calculated from multiple pass 24h dietary recalls have showed variations among the different population groups investigated in the five reported countries (Italy, UK, Serbia, Finland and Lithuania). However, some common trends can be highlighted in all ROP groups and most of these confirm the previous results from literature analysis: consumption of cereals, meat products, eggs and sugar is higher in the most of ROP groups than in the corresponding AFF; consumption of fruit and vegetables is lower in the ROP groups in all partner countries; ROP people tend to eat less dairy products in almost all countries; the contribution of total fat towards dietary energy is higher than the recommendations in all countries, with no significant differences between the two different population groups; the intake of saturated fats is higher than recommendation in several ROP and AFF groups; average daily intake of protein was adequate in ROP and AFF in all countries; the consumption of sugar is higher among the AFF groups, with the exception of Italy; significantly lower intakes of key vitamins is found in ROP compared with AFF group, although the intakes of many vitamins, particularly folate, vitamin A, vitamin D, and minerals is below recommended levels in both ROP and AFF people; in all 5 countries, intake of sodium exceeded the recommendations in both groups.
In conclusion, the multi-centre nutritional survey performed within the CHANCE project has found main nutritional criticalities not only in the ROP populations, but also in the AFF groups used for comparison. It is worth noting that main nutritional criticalities found in ROP people are the same highlighted in participants with a better economic status.
The data obtained from the dietary assessment in the five countries have evidenced the main nutrients imbalance for ROP populations, and the output has been considered during the formulation of new foods within the CHANCE project. The food design has also considered the cultural differences, the popularity of foods and the age groups studied. For this reason, only the most popular, readily-accepted/liked foods across Europe have been chosen for foods development using novel, traditional and ethnic ingredients.
In the light of the reported nutritional data, it is conceivable that foods formulated to correct the malnutrition in ROP could be also consumed by the rest of the population. The main trait of the CHANCHE foods has been therefore linked to their cost. In fact, although the economic status is not the main determinant of the onset of nutritional criticalities, the purchasing power could be important for the correction of nutrients imbalance in the whole population, provided that the improved foods must be as cheap as possible to positively impact the nutritional status of ROP.
Therefore, the targets that the new CHANCE foods have had to reach are:
1. reduction of energy density;
2. decrease of the total fat, in particular saturated fat, while increasing the polyunsaturated fat content;
3. increase of dietary fibre through the employment of wholegrain and fruit and vegetables;
4. reduction of the contents of sodium and sugars;
5. conservation and improvement of the contents of the majority of essential micronutrients, in particular iron, copper, selenium, vitamins A, D, C, B1, B2, B6 and folate;
6. optimisation of the bioavailability of micronutrients
7. improvement of sensory quality;
8. improvement of the nutritional quality of commonly-consumed traditional and ethnic ingredients
9. increased affordability
Therefore, the subsequent step has been the generation of the so-called “CHANCE score” for foods that were identified as most commonly consumed foods among ROP.
Using the information obtained from the analysis of dietary habits of ROP population groups, both from literature review and from dietary surveys in CHANCE countries, two lists have been compiled: 1) a list of 39 simple and raw foods most commonly used in ROP groups; and 2) a list of 34 imbalanced nutrients among ROP.
From these list the following steps have been executed:
1. analysis of the content of 34 nutrients in the 39 raw foods listed
2. assessment of the percentage contribution of 100 g of the 39 selected food to the 34 nutrients, considering the total daily intake
3. assignment of nutritional scores to each nutrient within each food. A score from 1 to 5 has been assigned to nutrients of which daily consumption in ROP should be increased: this means that a higher score is for nutrients that had to prioritised. Scores from 1 to 5 have been given to vitamins, minerals (except sodium), proteins, monounsaturated fats, polyunsaturated fats, and total fibres, depending on their content in the most 39 consumed foods (0-20% →1, 21%-40%→2, 41%-60%→3, 61%-80%→4, > 80% → 5). The same idea, but with opposite scoring, has been applied to nutrients that should be preferably decreased in CHANCE meals (like total fats, SFA, sugars, cholesterol and sodium), for example -1 means that the specific negative nutrient contributes in a food between 0 and 20%, whilst -5 that the negative nutrient is more than 80% of 100 g of that food.
4. multiplication of these scores with the coefficients of importance, to set nutrients imbalance and target nutrients for development of CHANCE foods/meals final scoring system
5. sum of all the nutrient scores for each food and inclusion of a polyphenols score (0, 5 or 10, added to account for the beneficial effect of polyphenols) to obtain the total nutritional score for that food
6. assignment of cost score to each food based on prices and daily consumption. Cost coefficients are assigned to cost of amount of daily consumption of specific foods. These coefficients range from 0.5 to 1 (lower price = lower score) and they are used as a denominator to the nutritional score. All food groups are scored as follows: 0–0.20€ = 0.50 0.21–0.40€ = 0.67 0.41–0.60€ = 0.75 0.61–0.80€ = 0.8 >0.80€ =1.
7. integration of the total nutritional score and of the cost score to obtain total CHANCE score.
Food with the highest total score in each food group has been defined as CHANCE food suitable for improvement and/or development of CHANCE recipes. However, the overall approach for development of CHANCE meal has been complex, as it also have took into consideration the appropriateness of CHANCE target foods for technological modification/development, estimated health benefit and cost-effectiveness. Foods with the highest CHANCE score have been suggested to the teams working in WP4 for technological development. According to the data on foods consumption, collected among the ROP groups in the 5 countries participating in the CHANCE project, the foods so far selected as the most consumed have been: white flour (for bread, sandwich or pizza use), cooked ham, cheese (both soft and hard) and tomato (in the form of paste, pomace, ketchup or juice). Such foods have been considered either as matrix for nutrient fortification, or to be replaced with others, selected from the same category, with higher nutrient or higher total CHANCE score. The choice of nutrients to be added to foods and/or foods to be substituted has accounted for: i) identified nutritional criticalities, ii) the applicability of the foods to be used as a matrix for fortification and iii) the price of technological development of CHANCE foods/meals. Overall, the choice of CHANCE foods/meals did not aim to change the dietary habits of ROP.
Accounting for the aforementioned factors, the following have been recommended:
1. Change of recipes for bread, sandwich or pizza dough: white wheat flour should be substituted with CHANCE flour which consists of 50% white wheat flour and mixture of whole wheat flours.
2. Enrichment of cooked ham with fat soluble vitamins A, E and D, and iron and creation of the so called CHANCE ham which consists of 95% pork and 5% liver, with reduced sodium content.
These changes have been estimated to be simple and easily applied, i.e. cost-effective in CHANCE companies.
After these information have been passed for the production of CHANCE foods and meals, WP2 have had the role of evaluating the nutritional and overall quality of the corresponding food prototypes produced by the enterprises participating in the project.
CHANCE project has developed the following low-cost ingredients, aiming at correcting some of the imbalanced nutrients previously identified:
- Bio-modified bran, rich in dietary fibre and very low in fat
- YASO (germinated soya based additive), very rich in dietary fibre, protein, omega 3 and omega 6 fatty acids, minerals, but low in carbohydrate content
- Special aleuron fraction of wheat) which is high in protein and ash content and extremely high in dietary fibre content which provide very positive nutritional characteristics (95% fibres)
- Berry press cake, rich in dietary fibre (soluble and insoluble) and proteins
- Milk protein – Casein, rich in calcium, amino acids, especially in lysine
- Pork liver, rich in protein, vitamins A, D and E, vitamin B complex and iron
By combining the above mentioned ingredients, CHANCE project has developed the following low-cost simple foods or RTE meals suitable for ROP consumers:
1. CHANCE cooked ham with 5 % liver
2. CHANCE tomato paste
3. CHANCE pizza-cheese (mozzarella-like)
4. Pizza – with enriched pizza dough and cooked ham with 7 % liver, CHANCE tomato paste and CHANCE pizza-cheese
5. Fibre-enriched bread
6. Berry soup
7. Berry yoghurt
8. Berry shot
These foods have been thus evaluated in relation to three different key points:
I. reformulation effect on increasing/decreasing nutrients level in each food, as the amount of some nutrients can be unintentionally changed, with positive or negative impacts on the CHANCE score.
II. comparison of CHANCE score for foods before and after reformulation (including the cost score) using an expanded score range (positive scale from 1 to 20, negative scale from -1 to -20) in order to be more sensitive to changes.
III. evaluation of CHANCE meal with respect to EAR (Estimated Average Requirement) and population goals
The results so far obtained have been the followings:
1. CHANCE cooked ham with 5% of pork liver: the primary goal of increasing iron content has been achieved, as this nutrient has been more than doubled in CHANCE ham. Calcium, copper and zinc contents have been slightly increased and sodium content has been reduced by 5.7%. Vitamin D has been analysed in the forms of D2 and D3, because of the higher concentration of D3 in pork liver; it has been found that this vitamin was 0.5 µg in commercial cooked ham, whilst 20.9 µg in the CHANCE version. Vitamin A has been also greatly increased in CHANCE cooked ham: vitamin A content has met more than the 90% DRI (daily-recommended intake). Other vitamins increased in the novel products have been niacin, vitamin B6 an vitamin B12. Total fats have been reduced by 46% by means of a reduction of 46% of saturated fats and of 54% of mono-saturated fats.
2. CHANCE tomato sauce: the fibre content has been increased by 46.7%, meeting the desired targets. In this way, also the carbohydrate content has been increased (10-fold). The iron content has increased by 25%, though vitamin C unexpectedly decreased, probably due to technological procedures.
3. CHANCE pizza-cheese: the aimed reduction of fats has been reached, decreasing their level by 82%. This reduction has proportionally been the same for each type of fatty acid, though the reduction of SFAs was particularly important, as this type of cheese usually has a high level of these fatty acids. The reduction of PUFAs, although of 83.3%, has been low because the commercial product already contains negligible amounts of these FAs. Vitamin D3 has increased by 47.3% in the CHANCE cheese, thus the combination of this product with the CHANCE cooked ham in a meal could, will generate a significant improvement in the intake of this vitamin. In addition, iron, calcium, magnesium and potassium have been increased in CHANCE cheese, whilst the sodium content has been decreased.
4. CHANCE bread enriched with YASO: has reached a 3-fold increase in fibre content. Total fats also has been increased, particularly, the healthy MUFA and PUFA. Cholesterol has been decreased by 53% and vitamin E has increased from 0.5 to 3.6 mg/100 g, allowing 100 g of this product to provide 30% of the DRI for this micronutrient. Calcium, copper and potassium have also been increased, whilst, again, sodium has been decreased as suggested by nutritionists.
5. CHANCE bread enriched with wheat bran: fibre content has been more than doubled, going from 0.7 g/100 g measured in commercial bread to 1.8 g/100 g of CHANCE bread. Also Vitamin D content has been increased, together with the levels of iron, magnesium, phosphorous, zinc and potassium.
6. CHANCE yogurt with jam (enriched with berry press cake): it has been the only dairy food product developed by the project (pizza-cheese is considered as an ingredient), with higher fibre content (increased by 50%), as the consequence of the addition of the berry press cake in the jam. Vitamin E has also increased from 0.12 mg/100 g to 1.28 mg/100 g and folate by 73%.
7. CHANCE berry shot: the addition of the berry press cake has determined an increase in fibre content (by 44.5%). Vitamin C has increased by 13% and folate by 53%. In addition, all minerals, except for sodium, have increased.
8. CHANCE berry soup: the fibre content has 5-fold increased thanks to the addition of berry press cake. All the other nutrients are linearly higher since this type of berry soup is more concentrated than the commercial one.
9. CHANCE pizza: this product has been designed with the combination of different CHANCE ingredients (tomato sauce, cooked ham, pizza-cheese and pizza dough with bio-modified bran fibres and YASO soy). It has gained a significant improvement in nutrient contents with respect to the analogue commercial pizza. Total fats have been found in the same amount, although CHANCE pizza has a lower level of SFA (by 46%) thanks to their decrease in both ham and cheese. Together with that, MUFA and PUFA are higher in CHANCE pizza, again from ham and cheese, but mostly from YASO soy in the pizza dough. Fibres are increased by 83% in CHANCE pizza thanks to the modified dough and tomato sauce. Vitamins have showed increased levels for what concerned beta-carotene, thiamine, vitamins A, C, D2, D3 and E. Iron, magnesium, phosphorous and potassium are also increased in CHANCE pizza, whilst sodium content is decreased.
Using the CHANCE score to evaluate these products it has been found that the greatest difference is between CHANCE and commercial pizza, the latter having 118 as nutritional score and 1 as cost score, whilst the former has 177 as nutritional score and 0.67 as cost score. This means that commercial pizza has got 118 as CHANCE score, whilst CHANCE pizza has reached 318 for CHANCE score. This reflects the fact that the combined use of CHANCE ingredient can generate great positive changes in a food product.
The last performed analysis has been focused on the evaluation of the so-called CHANCE menu with respect to target population goals and EAR. The CHANCE menu chosen as representative has been made of 200g of CHANCE pizza (containing 5 reformulated foods) and 125 g of CHANCE yogurt. This menu has got an almost perfect distribution of energy: 57% from carbohydrates, 20% from proteins and 23% from fats, thus satisfying the nutritional recommendations. The CHANCE menu is very rich in fibres, satisfying 50-70% of DRI, and is nutritionally correct with respect to vitamins and minerals. In effect, one CHANCE menu a day would provide over 50% of the EAR for riboflavin, thiamin, vitamins A, C and E, phosphorous and zinc. Vitamin D is also very high, satisfying more than double the EAR, which is good considering the widely spread deficiency and tolerable upper intake level for individuals older than 9 years. In addition, sodium is lower than the upper limit, as recommended by nutritionists.
From all these results, it is evident that the widespread use of the novel CHANCE food prototypes could potentially improve the intake of many critical nutrients among the ROP population. Fibre is the critical food component mostly affected by CHANCE reformulations. Such food component has been identified as a critical nutrient among both population, ROP and AFF, in all countries. Finalizing the CHANCE project, 7 foods with increased fibre content have been developed. A wide usage of these products could potentially improve fibre intake among users.
CHANCE pizza contains 5 newly developed CHANCE ingredients, which are mostly improved for higher content of fibres, and vitamin D and lower level of fats. The whole idea of the project and application of the food prototypes was based on changing the food formula and improving the nutritive values but not dietary habits of ROP people. CHANCE pizza was proved to be the solution, perfectly matching the project goal. A wide consumption of this pizza among the ROP population could contribute to the improvement of their nutrient intake and nutritional status.

WP3 -Nutri-metabonomics to assess the metabolic consequences of critical diets for individuals at ROP
The team involved in Work Package 3 has been committed to acquire the metabolic fingerprinting of populations with different economic income and different diets. Nutri-metabonomics is a particularly innovative and important approach since it can give a direct idea about if and how diet (and maybe income) can affect the metabolic status, and thus the well-being, of people. Unravelling tiny changes in metabolic profiles among ROP and AFF populations has been expected to be a serious challenge, and therefore all state-of-the-art pattern recognition methods as well as up-to-date algorithms recently developed by the project partners have been exploited.
In order to collect good and robust quantitative metabolomics data, it is of paramount importance that human urine samples are collected under similar conditions, following a detailed protocol. Similarly, NMR spectrometers working in different laboratories have to be run using the same protocols. For these reasons SOPs for each analytical step have been compiled and followed by the recruiting centres in the different CHANCE countries. In this way, the data obtained have been comparable and are suitable to give useful information on the possible differences and similarities among samples from different origins.
The preparation of the sample is very simple, a characteristic which makes NMR spectroscopy a prominent tool for metabolomics. For each urine sample, a one-dimensional NMR spectrum was acquired applying a water signal suppression pulse sequence. To make sure that there would be an unbiased sample identification and to make the subsequent data analysis easier, a data reorganization step was included.
The CHANCE spectral dataset of urine samples collects a total of 2732 NMR spectra made up of 128 thousands variables covering a chemical shift range broad enough to allocate the signals generated by all metabolites. The samples have been collected in 5 different recruiting centres: 523 from Finland, 616 from Lithuania, 581 from Italy, 602 from Serbia and 410 from United Kingdom. Urine samples have been shipped under dry ice from the recruiting centres to the two analytical centres (CIRMMP – Florence, Italy - and UCPH – Copenhagen, Denmark -) and analysed using the same NMR spectrometers that have been previously utilized for the ring test. In particular, CIRMMP has analysed the samples from Italy (UNIBO), Serbia (IMR) and United Kingdom (UL), whereas UCPH has analysed those from Finland (VTT) and Lithuania (VU). Every analytical step (e.g. urine collection, shipment, sample preparation and analysis) has been executed by following the predefined aforementioned SOPs, so to achieve the best standardization and to reduce the sources of unwanted variance. Spectral data have been kept in both their original format (Bruker Topspin) and in more accessible formats for multivariate data analysis such as Matlab (full resolution) and Excel (reduced resolution).
After the application of the acceptance criteria, required to assess the spectral data quality, 25 spectra has been removed from the data matrix, which was then reduced to 2707 spectra × 131072 variables. After the pre-statistic processing, the number of variables in the dataset was reduced from 131072 to 58000. This reduced dataset, sized 2707 × 58000, is the one successively used for further data analysis. Additionally, before multivariate analysis, a comprehensive alignment of all NMR spectra and a normalisation step have been applied. At last, the variable dimensionality has been reduce by means of window averaging (binning), resulting in a 10-fold reduction in the number of variables.
A preliminary thorough literature review of nutri-metabolomics biomarkers has been carried out, in order to aid analysis and interpretation of the spectral datasets that were collected during the CHANCE project.
The first step of multivariate analysis has been consisting of outlier detection and removal. In effect, this process is one of the key points in order to have data ready to undergo a thorough data mining analysis and to get useful information from the spectra. Outlier detection has revealed that diabetic subjects have to be omitted in the global analysis in order to avoid overlooking smaller sources of variations in the data.
After this step, unsupervised analytical techniques has been firstly applied to the data matrix in order to check if the two urine samples collected from each subject were systematically different. The results has excluded the bias introduced by the time of collection. For these reasons, the two spectra for the same subject have been subsequently averaged considering the mean spectrum to be a better representative of the actual subjective metabolic status than just a one-time point sample. The second explorative data analysis has been therefore aimed at highlighting any differences in the urinary metabolome amongst the populations represented by the 5 different recruiting centres.
From the PCA scores plot, some weak tendencies to differentiate among recruitment centres has been observed. This is the particular case when inspecting the PC2 scores, where most of the samples from UK subjects have positive values, whereas those from Finland are mainly shifted towards negative ones. Inspection of the PCA loadings have pointed towards the aromatic region where the signals mostly responsible for the discrimination are located. Indeed, the hippurate content for the UK people (south Asian ethnic minority) has been found sensibly lower than that of Finnish ones, maybe for genetic reasons. Such outcome is indeed expected since an objective diversity exists between the two populations both for the ethnicity (Scandinavian vs South Asian) and for the nutritional habits. It is important to exclude that the difference is due to different age groups, since the recruited subjects have been, in both countries, over 65.
In order to focus on pattern differences originated from gender, the data analysis has been limited only to one recruiting centre, since no evidence has appeared pointing towards male and female separation when PCA analysis was applied to the entire data set. The Finnish cohort was analysed apart, and a quite good separation between the genders can be retrieved by means of PCA. As already shown in literature the spectral regions containing hippurate, creatinine, TMAO and citrate signals are those leading the gender differentiation.
It has been relevant to investigate, using a non-supervised approach, on possible metabolic differences between the two socio-economic different populations, namely affluent (AFF) and at-risk-of-poverty (ROP) people, since this was one of the main targets of the metabonomics investigation of WP3. The results of such an investigation, based on unsupervised PCA, has shown not a clear tendency towards a socio-economic differentiation when the whole data set was analysed. At last, the unsupervised analysis has been carried out on the only samples belonging to the same gender collected in Finland. Also in this specific case, in which the gender and country variances are eliminated, no difference could be found between the two socio-economic conditions.
The result has made evident either that an unsupervised approach is not powerful enough for retrieving any possible subtle metabonomic difference caused by the subjects’ socio-economic condition, or that such a difference indeed did not exist.
In order to investigate the latter hypothesis, finally a supervised approach has been carried out by means of more dedicated and tailored chemometric tools. To expand other possible separation among recruiting centres, the Extended Canonical Variates Analysis tool (ECVA) has been applied, and the results has showed again that the UK and Finnish samples are the most extreme (especially the UK ones), whereas the other three origins are more similar (less separated). Apparently, metabolic phenotypes (metabotypes) of Italians, Serbian and Lithuanians are very similar and a clear differentiation cannot be pointed out.
Another supervised technique, namely PLS-DA, has been applied to amplify, if actually existent, even weak tendency to group ROP separately from AFF. In this case the information about the economic status was provided to the chemometric algorithm, to seek for possible combination of latent variables that could maximize such a difference. When analysing the whole dataset by PLS-DA, an encouraging result towards a differentiation of the two groups has been obtained giving a model with a tendency for separation between the two economic groups, although not clear as requested. However, the PLS-DA method is prone to overfitting, and indeed after validation such result has been found to be overfitted, as many predicted subjects fall in the wrong category. Still, it is plausible that the little variance caused by the different income status can be shadowed by more significant factors of variance such as gender and origin. Therefore, in order to limit, as much as possible, the sources of other variance a further analysis trying to separate ROP and AFF has been attempted on the Italian cohort, the most numerous group constituted only by subjects of female gender. The result has shown an even better separation compared to the one obtained on the global dataset, indicating that a more homogeneous group is a better choice for studying the metabonomic differences caused by a different economic status.
The expected difference based on the income is based on the assumption that ROP and AFF have different quality of the food intake which affects the metabolic fingerprint. The influence of food intake, however, could be accounted for by directly inspecting the nutritional habits of the populations.
For this reason, a parallel study has been conducted on nutritional data derived from 24 hour recall questionnaires. Only those nutrients categorized as CHANCE Nutrients have been considered and analysed with a multivariate analysis approach, since many nutrients differed among countries and the CHANCE nutrients represented a consistent core of data available for every recruiting centre. Along with the nutritional values also the anthropometric meta-data, as well as the information about gender, age and economic status have been included.
The nutrient data of the same subject have been averaged out between the two days of diary recall and used for the successive multivariate data analysis. Nutritional tables have been analysed to investigate their ability to discriminate between ROP and AFF people, and by applying the supervised PLS-DA approach. For each subject, a total of 43 variables has been gathered, consisting of 7 physical metadata (gender, age, anthropometric) and 36 CHANCE nutrients divided into 12 macronutrients, 13 micronutrients and 11 minerals. Since the units of these meta-data differed among a few order of magnitude, the data were autoscaled prior to any chemometric analysis in order to equalize their importance and variance.
The main results obtained from the multivariate analysis on these data matrices have been:
1. dietary (24 HR) and anthropometric data has given indications about some trends differentiating ROP from AFF dietary habits, nevertheless they are unable to provide a robust model for prediction of ROP.
2. data fusion and preliminary correlation analysis between the spectral and the nutritional data have provided some encouraging results that require a more dedicated studies to be fully disclosed.
Some more promising predicting models have been obtained from the Italian data set, for which other techniques have been also tested. The results have been the following:
1. fish-based foods (and nutrients) seemed to help discriminating for AFF vs. ROP classification for the Italian population.
2. hierarchical clustering (HCA) of metabolic profiles has selected a particular group of individuals (named Group_5), mostly AFF subjects, that could have peculiar features in terms of dietary habits. Evaluation of metabolites content in Group_5 urines has shown some statistically significant difference from the other subjects: phenylacetilglycine and mHHPA are higher in the investigated group, while creatinine plus creatine, valine, valerate, isoleucine, leucine and vitamin-C are lower.
3. alternative statistical tools applied to dietary data has revealed that nutrients patterns are indicative about dietary habits: fast-food products consumption seem higher in ROP, the same holds for snacks. Consumption of some meat such as beef, veal and rabbit seems higher in AFF; the same has been observed for vegetable foods like artichokes, cucumbers, broccoli and mushrooms.
4. for the Italian population, urinary metabolomics profiles have been correlated to nutritional data by means of canonical correlation pointing out that the urinary metabolites hippurate and creatinine could be associated with pattern of nutrients rich in dietary fibres and their derivatives, as well as alcohol, riboflavin and phytic acid.
5. correlation between PCA scores of nutritional H24 and spectral data of Italian populations didn’t show any strong correlation: maximum correlation value (0.27) is obtained between two uninformative PCs and the minimum correlation value (-0.33) although not significant, shows a link between nutrient PC3 and spectral PC2. The inspection of the loadings of these two PCs pointed out that oligoelements and fatty acids, for nutrients, and creatinine, TMAO and isocitrate, for urine metabolites, are negatively correlated each others.
In conclusion, some important results have been obtained, starting from the validation of the CHANCE protocols for the metabonomic analysis. It has been shown, as well, the evidence that even for observational studies conducted on freely fed subjects, different population groups (e.g. based on country or gender) show distinct metabolic patterns. However, belonging to the ROP or AFF groups has effects on metabolome that are hidden under all the other stronger effects (genetic, gender or age), and needs more time to be investigated carefully in parallel with the nutritional data. A conclusive feature could be drawn for the metabolome of people at ROP: it appears much heterogeneous in comparison with the one associated to the AFF population. It could be argued that, assuming a healthy status in the centre of the metabolomic space occupied by all subjects, there are more ROP than AFF people more distant from this centre.

WP4 -Development of methodologies for food production and packaging
The fourth work package has been focused on the development of the most effective methods for the production of CHANCE food and their packaging.
The ingredients to be used for these products have been chosen among those evidenced by WP2 on the basis of national and regional differences, their technological and expected physiological functionality as well as the production costs.
Processing techniques, instead, have been optimized for the conversion of chosen raw and low-cost materials into ingredients which are going to be used for the design of low cost foods.
Each selected ingredient is evaluated in terms of its compositional and nutritional profiles and a set of quality parameters have been assessed as well.
This activities have lead to the obtainment of the following low cost ingredients:
- Bio-fermented modified bran
- YASO (germinated soy based additive)
- Special aleuron fraction of wheat
- Tomato pomace (Tomato-processing by-product)
- Berry press cake
- Milk protein (casein)
- Pork liver
Each selected ingredient has been evaluated in terms of:
1. Safety (chemical, microbiological and allergenic risks);
2. Technological feasibility (estimated cost, nutritional value, shelf life, chemical, physical and sensory properties);
3. Nutritional quality
For what food safety concerns, each food ingredient has been assessed with regards to the key aspects of microbiological and chemical food safety, considering European legislation.
Additional issues regarding food labelling have been evaluated in the light of the recent implements on food information regulations that came in to force during the lifetime of the CHANCE project and are especially relevant with regards to nutrition and allergen labelling.
All food ingredients have been manufactured according to the principals of Good Manufacturing Practice and all food safety issues have been managed using food safety management systems and the application of hazard analysis and critical control points (HACCP). In addition to the General Food Law Regulation 178/2002, specific legislation that has been addressed is as follows:
- microbiological food safety (Commission Regulation (EC) No 2073/2005).
- chemical food safety (Commission Regulation (EC) No 882/2004, 669/2009, 187/2011).
- food information for consumers regulation (FIR; Commission Regulation (EC) No 1169/2011).
In order to ensure the durability of the approaches and ingredients identified by CHANCE, they have been assessed with regards to this legislation and its requirements concerning food allergen and nutrition labelling.
A questionnaire has been submitted to food ingredient producers, to ensure the principals of GMP and HACCP have been followed, and identify if particular microbiological, chemical or food allergen issues needed to be specifically assessed or addressed.
Each ingredient has been tested to have an associated certification to ensure compliance.
One of the proposed ingredients has been the special wheat aleuron fraction. Whole grain cereals are recommended by food and health authorities because of evidence of a reduction of the risk of several chronic disease due to their consumption. The possibility to exploit bioprocessed bran fractions with different granulometry is a useful opportunity to obtain new functional ingredients suitable for the preparation of several foods. The special wheat aleuron fraction (BKL) has nutritional benefits. High level of minor bioactive components like tocols, alkylresorcinols, phenolics, sterols, lignans etc. have direct health promoting effects (anti-inflammatory, anticarcinogenic, antioxidative etc.) while the enhanced level of dietary fibre components like arabinoxylans and β-glucans improves the health status of digestive tract. The special aleuron fraction of wheat can be used as substituting component of normal wheat flour or can even be applied (in 100% level) instead of wheat flour in different food systems. There is no difference between normal flour and special aleuron fraction flour concerning manufacturing cost, so this special additive can be highly recommended in low price food development.
Another ingredient evaluated by the CHANCE team has been YASO, a germinated soybean product having a meat-analogue character, concerning gross chemical composition, but additional high level of functional minor constituents and bioactives. Typical applications of YASO could be implemented where the increasing of protein and/or dietary fibre level and/or fatty acid composition is targeted. Beside the functional benefits of YASO, it is easy to handle from a technological point of view and even because of its neutral taste and smell can be flexibly applied in different food systems.
Berry press cake has been also tested. This is a by-product of juice process which is normally considered as waste and not used further, but it has been found that it contains a very high amount of natural fibre (65%), mostly insoluble. This product is not usable as such, because the large sized particles give a bad mouth feel. It is thus necessary to mill the berry cake in order to include it in the final product. So far it has been almost impossible to mill berry press cake to become an ingredient of good quality, but during the CHANCE project, a new colloidal mill has been used to get very smooth milled berry press cake, that can be further used in different type of food products.
Milk proteins have been other possible CHANCE ingredients. These proteins are rich in amino acids, especially essential lysine. Also, dairy casein contains high amounts of calcium needed for bone health. Several health and wellbeing benefits have been shown with the consumption of dairy proteins. Dairy proteins can be used in wet form or dried to powders. They can be processed to obtain food products by using different processing methods, to produce for example yogurt, cheeses and beverages. Both proteins fractions (casein and whey) have very different properties to heat and acid.
Finally, pork liver has been demonstrated to be a good ingredient for the final product cooked ham. At levels of 5 and 7% (of total weight) in the cooked ham, liver increases iron content without significantly affecting sensory properties. Taste was one of the main concerns in CHANCE food development, because some consumers could reject extraneous aroma and taste, like those of pure liver. At these levels in ham, pork liver has demonstrated to be a nutritionally good and low-cost ingredient. For these reasons it could be used as an ingredient in cooked ham and some other meat products.
The next step has been the development of CHANCE test food and their evaluation through different types of analysis: chemical, rheological, sensory, etc, together with innovative approaches like metabonomics and in vitro digestion to assess the bioavailability of their ingredients.
The tested foods have been:
- CHANCE cooked ham
- CHANCE tomato paste
- CHANCE pizza-cheese
- CHANCE pizza
- CHANCE bread
- CHANCE berry products (blueberry jam, blueberry soup and a berry and vegetable shot)
The results from chemical and sensory analysis for CHANCE ham with 5% and 7% of pork liver (compared with standard commercial cooked ham) have proved that a production of ham with 7% of liver is preferable, since it will bring more iron and vitamins into the product and, at the same time, it will not significantly affect taste and other sensory properties on the final product compared to commercial ham already on the market and ham with 5% of added pork liver.
Tomato paste and ketchup like products developed by WP4 team have increased cellulose and dietary fibre content in comparison to conventional products. Additionally, they also have lower costs of ingredients in comparison to the conventional products which they are supposed to substitute. Final products could be adjusted to consumers preferences in different regions (example of tomato paste and ketchup-like products have been presented to sensorial panels). The basic developed product is convenient for further nutritional improvement by reduction of salt content, addition of vitamin, mineral, probiotics, prebiotics or other supplements
The produced pizza-cheese has been evaluated too and has shown good hot melting properties.
The CHANCE pizza, with improved nutritional quality, has shown satisfactory results related to both technological performances and quality and sensorial characteristics, comparable to the control one.
Selected novel additives have been used to produce different breads. A 10% addition of bran or 30-50 % addition of YASO or even up to 100 % addition of special aleuron fraction flour (BKL) could be properly used in production of bread. These levels of additions do not have any negative effects on physical or sensory characteristics of bread products. The applied additives improve significantly the chemical composition (protein, dietary fibre, bioactives, etc.) and nutritive values of bread products. Wheat bread containing bio-processed bran was shown to effectively improve nutritional quality with good sensory and textural properties.
Berry press cake is now possible with the new colloidal mill technology. The optimal particle size of milled press cake is around 150 μm. With this ingredient different food items have been produced and tested: blueberry jam, blueberry soup and a berry and vegetable shot. The taste and mouth feel is pleasant in all those product with milled blueberry press cake. It is therefore safe to say that in reference recipes, blueberries can be substituted with milled blueberry press cake thus giving cost reduction. The production can be claimed source of fibre (yogurt with blueberry jam containing 2 % of fibre) and rich in fibre (blueberry soup with 1.3 % of fibre) and rich in fibre (shot with 2 % of fibre) and all fibre will be from berries.
After these first analyses, bio-accessibility studies of some of the CHANCE food products using a combination of in vitro digestion models and a suite of profiling methods have been carried out.
The following foods have been tested:
1. CHANCE cheese
2. CHANCE ham
3. CHANCE ham (5% liver)
4. CHANCE pizza base
5. CHANCE tomato paste
6. CHANCE bread
The in vitro digestion model has been applied to mimic the real human process, thus comprising three main stages: oral, gastric and duodenal. The models used in the study of CHANCE foods have been developed through national funding (The UK Biological and Biotechnological Sciences Research Council, BBSRC DRINC) and in collaboration with researchers at the University of Birmingham. There are two main types of digestion models: batch models, which do not employ extensive modelling of the physical characteristics of the human gut, and ‘dynamic models’ which, to varying degrees, model physical food and fluid behaviour. The model employed for the CHANCE project foods has been a batch model, since previous research undertaken in the BBRC DRINC project has demonstrated that this type of digestion model can provide an excellent, cost-effective way of modelling digestion and allowed CHANCE research to focus on profiling methodologies.
The batch digestion models are also called ‘test tube’ or ‘static’ models. These models do not mimic the complex processes of dilution, pH titration and mixing behaviour that actually takes place in vivo nor do they model absorption. Instead, in batch models, foods are rigorously mixed to a homogeneous state and digestion products are not removed throughout the digestion time course. This aids practical work in sampling and reproducibility. A number of different batch model modes have been employed which differ in parameters such as mixing, pH, volume, time, fluid composition and volume, as well as amount and activity of digestive enzymes used. One of the advantages of batch models is that such parameters can be varied systematically to answer specific questions. However, the values are not always transparent in digestion study publications. In addition to this problem, even when conditions are stated, the reasoning behind them and the model limitations are not always clear. In spite of this, studies using batch model systems are useful in giving a snapshot of the in vivo process and due to their low cost they are often favoured over other systems.
The model employed for the CHANCE food digestion studies has been planned as follows:
- oral phase, using simulated salivary amylase and mechanical break-up of the food,
- gastric phase
- intestinal phase.
A simplified sampling strategy has been developed based on the BBSRC DRINC project approaches, in order to allow effective nutrimetabolomic and proteomic analysis for the mapping of nutrient bioaccessibility and bioavailability in CHANCE foods and their comparators.
To do this, CHANCE foods have been analysed using several complementary techniques which allowed the elucidation of the constituents with a focus on nutrients.
Analysis has been performed on both the food as provided and on samples taken during model digestion. The approach to the analysis includes :
1. An assessment of nutrient release from the CHANCE food products, including ‘small molecule’ analysis of amino acid, sugar and fatty acid release throughout the simulated digestion model. This has allowed the modelling of both the digestibility of the CHANCE food products and bio-accessibility of identified nutrients upon release from the food matrix. This data has been then compared to commercially available comparator food products in order to place the results into context.
2. Proteomic analysis of food digests, allowing the determination of digestion profiles of identified protein constituents, such as myosin and beta-casein. This analysis has been also complementary to the aforementioned analysis of amino acid release from the CHANCE food products following simulated digestion.
The chosen analytical methodologies have been SDS-PAGE, 1H 1D NMR and Orbitrap MS.
1H NMR has been chosen over GC-MS since it doesn't require prior knowledge of the nutrients present within the sample, allowing the simultaneous observation of all the relevant nutrients, which upon examination where within both LOD (limit of detection) and LOQ (limit of quantification). This technique, even if less sensitive than GC-MS, also requires only one individual standard for nutrient quantification (TSP), and this is less time-consuming than the selection of standards suitable to the system required for establishing LOD and LOQ in GC-MS.
These analyses have shown that the tested foods are at least comparable with comparator foods or commercial counterparts with regards to the bioaccessiblity and bioavailability of nutrients. NMR analysis identified the release of 31 individual ‘nutrients’ during simulated gastroduodenal digestion including various sugars, amino acids, fatty acids and various biological precursor/intermediate products.
The combination of the ham, cheese and pizza-base has been highly synergistic with regards amino acid availability. The NMR nutrimetabolomic approach applied has showed that, collectively, these CHANCE foods provided 13 bio-accessible amino acids including 6 essential amino acids. Many of the detected nutrients showed specific bio-accessibility profiles as a function of the simulated digestion period. The rate of release and extent of amino acid bio-accessibility from the cheese and pizza base has been superior to the ham and this might be due to the possible presence of exopeptidases in these foods, possibly as a consequence of the fermentation processes used in the production. The release of the essential amino acids L-leucine, L-phenylalanine, L-tryptophan and L-valine during simulated gastro-duodenal digestion of CHANCE cheese has been complementary to that from the pizza base. In addition, the release of the essential amino acid L-lysine could be assigned within the CHANCE cheese product, since it has been not observed in either ham or pizza base products. L-tryptophan in particular is shown to be released following duodenal digestion only, and this result is consistent across both pizza base and cheese, whilst it has been unassigned in ham. In particular, amino-acid release has been found to vary between in vitro gastric and duodenal digestion. Variation in the amino acid release profiles of cheese and pizza base products is most likely due to the difference in their respective protein profiles, matrix structure and the types of fermentation involved in their production.
All three analysed CHANCE foods contained bio-accessible sugars such as sucrose, D-galactose, D-glucose (cheese), D-lactose and D-maltose (pizza base) and sucrose (ham and cheese). CHANCE ham also provided other important nutrients in a bioaccessible form, including choline, creatine, carnosine, cholesterol, triacylglyceride and fatty acids (saturated and unsaturated). Some of these are important nutrients: in effect, carnosine has been proposed as an anti-aging nutrient for human and biomarker for meat intake, choline must be consumed in the diet to maintain health and creatine can help to preserve the muscle strength.
Incorporation of multivariate statistical analysis such as principal component analysis (PCA) to the analysis of the bio-accessibility of nutrients following simulated digestion allowed a rapid method of identifying which nutrients are undergoing changes in concentration (as reflected in their respective 1H resonance signal intensities). This also provided a method for the comparison of the 1H spectral profiles of the newly developed products against commercially equivalent ones, as well as allowing the rapid identification of which specific nutrients result in the variability between the products. When applied to the analysis of the CHANCE cheese product against commercially available mozzarella cheese, PCA analysis revealed that the two products are similar with respect to their nutrient profiles, with the exception of the presence of lactate within the commercially available product. The occurrence of lactate within the commercially available product is assumed to be a consequence of the addition of sodium lactate during the manufacture process, but this needs confirmation. These data support the conclusion that the CHANCE cheese product is equivalent, or has superior, digestibility characteristics to the commercially equivalent product. A similar conclusion can be drawn by the comparison of the other two analysed CHANCE products (pizza and ham) with their commercial counterparts.
Concluding, it has been proved by the activity of WP2 an WP4 how the developed CHANCE products are good solutions for the improvement of diet both in ROP and AFF populations. These novel foods showed to have the targeted nutritional characteristics both in the native ingredients and in the final matrix, even after digestion.

WP5 - Processing scale up and technology optimization
The activities included in WP5 have identified the best raw material sources, processing conditions and packaging solutions and have put them together into flow charts to show the most feasible production schemes.
It has been estimated that no less than 6 processes have to be analysed. Analysed processes have been selected based on the potential to fulfil nutritional targets with minimum cost structure.
In this way, industrially feasible production designs for these new food products, based on the food concepts developed in WP4 have been established. The different scenarios for ingredients and food production, as well as food packaging, have be designed in strong collaboration between RTD and industrial partners. These scenarios have been analysed for availability of processing equipment to typical food processing companies, complexity of processing protocol and estimation of production scenario to deliver eligible nutritional benefit. Each possible scheme was scored and the best one was thus chosen.
The results showed that industrial formulation and processing steps could need slight modifications, in order to allow product processability and optimize product quality and shelf-life in real industrial conditions. This is particularly needed in more complex ingredients and products (e.g. ham and pizza) with a high number of processing variables and parameter combinations.
It was also found that the nutritional characterization of the final industrial products was necessary, particularly for the different thermal impact of the industrial heat processes.
After the choice of scenarios, preliminary economics for selected combinations of ingredients and raw materials (low cost foodstuffs or RTE meal suitable for consumers at risk of poverty) developed in the project have been analysed. Incremental approach was used, so to take into account the additional costs and benefits from the new production system compared to the corresponding traditional ingredients and raw materials. Moreover, it was assumed that the new production systems are integrated into existing ingredient and raw material production systems. The production systems considered include, as mentioned before:
- enriched low cost breads,
- cooked ham with liver,
- tomato paste rich in fiber,
- “mozzarella” like pizza-cheese,
- pizza with ingredients designed in CHANCE Project,
- berry press cake based jam usable in soup, yogurt or vegetable shot.
Preliminary techno-economic evaluation of the production systems has been conducted using, as said, incremental production cost estimate of the new products as the economic performance measure. The production systems have been defined by the development work packages. The information collection was conducted using partner survey requesting following information:
1. Production process block-flow diagrams (original vs. CHANCE food)
2. Material and energy balances per single operational unit (e.g. t/d and kWh/d, dry matter content, temperature, pressure)
3. Price estimates for all the process feeds and end products (€/kg tai €/kWh)
4. Investment cost estimates for single operational units and infrastructure (€, for e.g. containers, mill, fermenter)
5. Labour requirements
6. Market potential in European core markets
7. Logistics costs (raw material and product)
The block-flow diagram of the production process, the recipe for the new product, and main process conditions (e.g. temperature, pressure, residence time) have been obtained through the survey for some of the CHANCE products. Based on this information obtained from the partners, the difference in production costs between the original product and the CHANCE product has been estimated (incremental cost analysis).
Keeping in mind that the results are in many cases very uncertain, the following are the main findings from this analysis. The cost analysis has shown that in the case of fibre-enriched breads competitive cost structure can be obtained if low cost fibre ingredient, such as bran, is added without any modifications. In the case of cooked ham with liver, savings obtained from lower pork need cannot offset the additional costs due to the new process costs. In the case of tomato paste rich in fibre the cost seems to be very competitive if the replaced products are sold at about 700 - 1500€/t. The new developed cheese seems cost efficient compared to traditional cheese. The preliminary cost analysis has shown that the costs of the CHANCE pizza are less than 1/3 of the costs of a traditional pizza. This results mainly from a large difference in the prices of a) ham, b) wheat flour, c) cheese, and d) ketchup. The ham, ketchup and cheese price/cost comparison results from the fact that in reference case the ingredient is purchased with market value and in CHANCE case with the production cost of the ingredient. The variable costs of different berry press cake products have been also evaluated but no comparison to corresponding berry products has been possible.
All in all, it was found how CHANCE products could be competitive or even cheaper than their commercial counterparts, therefore providing healthy nutritious meals at a low-price.

WP6 – Technology transfer and test of production
The task of WP6 activity has been to produce, in industrial scale, 7 different products and analyse their production thoroughly.
The analysed products have been:
- bioprocessed (fermented) wheat bran
- bread
- ham
- tomato paste and ketchup
- pizza
- pizza-cheese
- berry-based products (yogurt with berry jam, berry shot and blueberry soup)
The results found have been as follows:
WHEAT BRAN: This product, obtained from the outer layers of grain separated in milling processes, is considered a good source of dietary fibre, proteins and nutritional molecules. For this reasons, it has been studied as a possible CHANCE ingredient, even if it is usually limitedly used in food because it can alter negatively food quality. In this case, though, the application of microbes and enzymes in its bio-processing has been proved useful to obtain good sensory and textural properties in a bread containing this ingredient. The specific wheat bran used in CHANCE project is a sourdough containing 80% of water and 20% of wheat bran, with an average particle size of 400 µm, bio-processed with yeasts (Kazahstania exigua) and lactic acid bacteria (Lb. Brevis) and commercial carbohydrases mixtures (Depol 740L and Grindamyl).
During the industrial scale production test, bran has been bio-processed at VTT (Espoo, Finland). The employed ingredients for bran production have been 3.4 kg of wheat bran, 13.4 kg of water and appropriate concentrations of starter cultures and enzymes. This has been all mixed and incubated at 20°C for 8 h, then cooled down to 4°C and transferred to Hungary where bread was produced in an industrial bakery (Lipoti Bakery, Hungary) within 24h, keeping the temperature below 4°C.
The issues found have been the following:
- CHANCE studies has employed pure starter cultures prepared by VTT, but in commercial industrial production more easily available starters or previous sourdough should be employed. The latter, though is not applicable for all microbial strains.
- The stability of the fermented bran is limited: it can be stored only up to one day by cooling the slurry to about 4°C. The maximum storage time needs to be tested.
The quality control of the product has been done by analysing the level of yeasts, lactic acid bacteria, total heterotrophic aerobic bacteria and spore-forming bacteria and all those showed the expected growth pattern, proving a good quality of the product.

CHANCE BREAD: Four different CHANCE bread products have been prepared: with addition of native wheat bran (14%), with fermented bran-sourdough (14%), with special milling fraction (25%) or with the addition of YASO-germinated soybean (30%). These new products have been designed to be nutritionally improved, high-fibre bakery products.
Product manufacturing has been carried out in the industrial facilities of the Lipóti Bakery (Lipót, Hungary). There has not been change or modification in the original technology, the only difference has been in the leavening step, and temperature in the case of sourdough addition where the added material (in slurry form) has been cooled. The length of baking step has been optimized in each case to avoid the formation of unwanted products from an accelerated Maillard reaction.
The compositional improvement of bread products has been different depending on the types of additive applied. The application of wheat bran and fermented wheat bran (sourdough) enhanced the level of dietary fibre by 5 % with respect to control, while the protein content has been elevated by 0.3 %. The level of minerals (specifically Cu, Fe, Mg, Mn, Zn and P) has been improved by approximately 15%. These additives have been rich in vitamin B components and their sterol, alkylresocinol and arabinoxylan components have been significantly higher compared to normal wheat flour.
The addition of the special milling fraction (aleuron), instead, resulted in a 5% increase in dietary fibre, 1,2% in protein content and 10% in minerals. The level of bioactive components has been also significantly higher compared to bran.
The germinated soybean addition improved the dietary fibre content by 5%, the level of protein content has been enhanced by 0.5% (with a balanced amino acid composition) and a high level of fitoestrogens has been found in bread products with this ingredient.
Products have been then evaluated for their sensory characteristics and it has been found that they had similar or poorer organoleptic quality in comparison to control products.
Other tests have been for volume, density, water absorption and shelf life and all of those showed to have a higher variability compared to the commercial counterparts.
From these results it has been proved how the four additives can be used to produced bread at their highest level, without great negative organoleptic effects. Nutritional effects vary depending on the additive used: they all increase highly the level of dietary fibre, but bran and aleuron fraction can enhance protein level, too, and influence protein functionality and water absorption capacities. All the ingredients, also, generated a great amount of bioactive and positive components in the final product.
It has been also shown how there is no need for modifications in the industrial processing steps. The newly formulated breads will only need a longer leavening time and probably a higher temperature when the fermented bran is used. In any case, however, baking time must be optimised according to the product wanted.
The quality test finally performed on the products have been done using standard ICC and ISO methodologies in order to measure compositional, physical and functional characteristics. To test sensory quality and product stability, instead, the twenty points weighted test method is advised. It is thus evident that the existent tests are already sufficient for the characterisation of these new products.
It is also to add how these results prove that the new ingredients can be add in other bakery products, though prototypes need to be tested to check for the compatibility with existing technologies.
CHANCE HAM: it has been prepared with the addition of 7% (maximum sensorially accepted level) of pork liver to increase the levels of iron and vitamin A.
Quality and safety parameters for the enriched product have been similar to the commercial cooked ham products but with added nutritional value.
CHANCE ham is composed by the typical ingredients for cooked ham, with the exception of liver:
- pork first category - leg;
- pork first category – shoulder;
- pork second category;
- water;
- ice;
- soy isolate;
- modified starch;
- nitrite salt;
- soybean fibres;
- polyphosphate;
- sugar;
- color stabilizer;
- coloring agents.
Three kinds of cooked ham have been subjected to chemical analyis: the commercially avalaible product (standard cooked ham, used a control) and two types of CHANCE ham with 5 or 7% of added pork liver.
During the production processes, only existing technologies have been applied. The only additional operation has been the preparation of liver as an ingredient, while the rest of the production process has been the same as for the production of regular cooked ham.
These minimal changes resulted in no special technological issues and this made production of the new cooked ham products suitable with existing production processes and technologies.
Chemical analysis on the final products revealed that in the newly developed ham iron and vitamin A levels have been increased. In addition, the levels of salt (sodium) have been decreased, enhancing again the nutritional profile of this product.
Microbiological testing of all samples has been also performed, using ISO methods, and it showed that Salmonella spp., coagulase positive Staphylococcus, sulphite reducing Clostridia, Proteus sp.and Escherichia coli have not been found in any of the samples. Therefore the product has been considered safe.
For sensory analysis, samples of raw and baked ham (standard and modified 5% and 7% hams) have been used. Since the results showed no statistically significant differences between the standard and ham with 5% and 7% of added liver for baked ham, the 7% ham has been chosen as a final product, since it increases more the level of beneficial nutrients. In addition, this product could be easily produced on existing equipment and by the same technology as standard ham products.
To control the quality of this new product, standard ISO methods for meat products have been applied in order to assess the chemical, microbiological and sensory properties. Therefore, no additional method of analysis needs to be developed.
This new product can be produced on the large scale, just as its commercial counterpart. Additionally, the price of the added ingredients is very low and the one of the final product is very affordable for consumers, providing therefore a good nutritional value at a low price. Good labelling and advertising could carve a good market share for this type of product.
CHANCE TOMATO-BASED PRODUCTS: Two nutritionally correct foods have been developed at lab scale in FINS and then produced at industrial scale by SME “Zdravo” (Selenča, Serbia):
1) Tomato paste like product based on tomato press cake
2) Ketchup like product based on tomato press cake
These two products contain tomato press cake, a by-product of tomato juice production with a high fibre content. The benefits of the use of tomato press cake is that it's free and its use contributes to a decrease in waste disposition costs for companies. The other ingredients used for the two CHANCE tomato-based products have been: tomato juice, sugar, salt, vinegar and starch syrup and they are all low-cost and widely available ingredients. Additional ingredients are xanthan and guar gum, used to increase the products' thickness.
For both products, the same existing production line in Zdravo, has been used. The only differences between the two products are:
1) the ratio between the employed quantities of tomato press cake and tomato juice
2) the quantity of additives used to obtain the product
3) the grade of concentration of product by water evaporation

The technological process consists of the following steps:
1) preparation of the mixture of water, additives, salt and sugar in order to achieve the swelling of the additives.
2) addition of assumed quantities of tomato press cake and tomato juice to the mixture
3) heating and stirring of the mixture in order to extract soluble fiber from tomato press cake
4) separation of large peel particles and seeds from the mixture in order to obtain smooth product
5) evaporation of water from the mixture under vacuum until the desired dry matter content and consistency are achieved
6) packaging of the products in glass units
7) pasteurization of packed products
8) labelling and forming of commercial packages in polymer foil

The line for tomato paste and ketchup like products in Zdravo consists of separate equipment units that are not interconnected in a way which would enable continual industrial production. However, with a minor investment, the production line could be completely connected with initial capacity of approximately 600 to 650 kg/h of tomato paste or ketchup like products. Similar equipment can be found in almost any plant involved in fruit and vegetable processing with production lines for juices and for cooked products like spreads, chutneys, jams or jellies.
The applied methods for process control and final control of the product are the usual methods used in fruit and vegetable processing industry: total soluble solids (brix), acidity (pH) and Bostwick consistency during the production and in the final product.
The final specification of developed products are the following:

Quality parameters Tomato paste like product Tomato ketchup like product
Total soluble dolids (Brix) 20±1 20±1
pH 4,0±0,1 4,0±0,1
Bostwick consitency, cm 8±1 20±1

In addition, the characterization and confirmation of nutritive value of developed products include the determination of total and soluble dietary fiber, NaCl and sugar content and texture analysis, which can be determined in any accredited testing laboratory for food. Therefore no new test is necessary for these products.
The newly developed tomato paste and ketchup will thus be an affordable and more nutritive option for consumers.
PIZZA: CHANCE Pizza product is a new type of pizza obtained with low cost raw materials and ingredients able to improve the overall nutritional quality of traditional pizza. Its preparation is performed with the addition in the base of germinated soybean-YASO paste (20%), wheat fibre (3%) and native wheat bran (3%), partially replacing wheat flour. Cooked ham with liver and tomato paste obtained by tomato by-products have been used for the topping in order to enrich the product in vitamin D, iron and fibre. The original recipe has been developed in lab-scale and then optimised for the industrial production.
The industrial production of the CHANCE enriched pizza has been carried out in the Italian company “La Pizza + 1” (Piacenza, Italy), equipped with industrial production lines for pizza products. The unit operations and related technological process parameters applied have been the same used for the standard pizza production: mixing (13 min); resting (20 min); lamination; topping with tomato paste; leavening (1.30 h at 35 °C and 85% RH); baking (6.53 min at 250 °C); topping with ham and mozzarella cheese; cooling (30 min at 16 °C plus 60 min at 0-2 °C); packaging in high barrier box and multilayer film in modified atmosphere (60 % CO2; 40 % N2); refrigerated storage (at 4 °C). The fixed expiry date has been 20 days.
The ingredients ratio has been slightly modified in order to optimize the visco-elastic properties of the dough that can strongly influence the technological performances in the industrial production. Following some formulation improvement, the pizza dough has been suitable for processability at industrial level especially for what concerned the main critical steps such as mixing, lamination and leavening.
There have been no technological issues during the industrial production of CHANCE pizza. The only remark is regarding a shorter leavening time necessary for a complete fermentation of CHANCE pizza dough compared to the standard one.
The CHANCE pizza has been then evaluated for other quality characteristics. Regarding the measured rheological parameters, CHANCE cooked pizza had a slightly consistent texture and a lower volume increase compared to the control sample. Its crumb structure had homogeneous pore size and distribution. From the sensory analysis the product texture and appearance have been considered acceptable. However, for smell and flavour CHANCE pizza has been judged not as good as the standard control, at least for Italian pizza consumers, mainly due to the odour and taste of the topping ingredients.
In addition, the variation of the main quality indices (sensory, microbiological, physico-chemical properties) has been followed as a function of time to evaluate the quality degradation of the product. After 30 days of storage at the required storage temperature of 4°C (ten days more of the fixed expiry date) no modifications in terms of sensorial odour, flavour and texture have been perceived. Measured physico-chemical properties (texture, moisture, water activity, pH, peroxide number) did not change significantly. Acceptable microbial counts have been maintained until the end of storage, and no product spoilage has been evidenced. In addition, pathogenic bacteria (Salmonella spp and Listeria), moulds and yeasts have been absent or under the detection limit. This means that performed technological choices (ingredients, formulation, packaging and storage conditions) have been able to maintain product quality.
Quality characteristics of pizza samples have been assessed after production and during shelf life by means of: main nutrients analysis (by means of Official Methods), gas measurement in the head space of packed pizza; moisture; water activity; textural measurements; microbiological analysis (mould, yeasts and total bacterial count); peroxide number of extracted fat; sensory analysis.
The used test methods applied are sufficient to characterise the overall quality of pizza after production and during shelf-life, thus no new test needs to be developed for CHANCE pizza.
Concluding, the new formulation developed has been suitable to produce CHANCE pizza at industrial level. The main general issue observed is related to the not well-accepted characteristics of the topping ingredients, in particular, smell and flavour of cooked ham and tomato sauce. Moreover, the cheese used did not have the requested physical properties, being in particular not able to properly melt during the final cooking step. This aspect has been probably emphasised since the panel has been constituted by Italian consumers, that are used to eat pizza with in the topping ingredients having very different organoleptic characteristics.
CHANCE PIZZA-CHEESE: This new product has been made using casein protein from milk, alginate and butter. In this case, the product has been made in a laboratory pilot scale with cooking kettle with mixing properties has been developed. In this way a cheese prototype with required melting characteristics has been produced.
Anyway, more studies are needed to obtain casein material from milk in industrial scale need and the processing of cheese in industrial scale with appropriate equipment needs to be further evaluated, together with storage stability (structure, microbiological, and sensory).

CHANCE BERRY-BASED PRODUCTS: Three different products have been chosen to be made using blueberry press cake: yogurt berry with jam, berry shot and blueberry soup. Berry press cake is a by-product from juice manufacturing and, up to now, it has not been used and available as an ingredient, even if it's rich in fibre and nutrients.
The main problem is therefore the obtaining of good berry press cake and in the right yearly amount for the production of these new foods, whilst for their processing no additional technology or process is needed.
Therefore, more studies are necessary to solve the problems concerning the industrial manufacturing of the new ingredient from blueberry press cake.
Potential Impact:
In the actual market, the size of low-price foods is in the order of billion of euro. The majority of low-cost food products also have a low-nutritional value, being energy-rich and nutrient-poor, with a high content of ingredients such as preservatives and flavours, which are poorly viewed by consumers. As it is evident, those products are not useful for the proper satisfaction of consumers' nutritional needs. A high consumption of this type of foods, especially possible in the low-income population groups, can cause malnutrition and chronic diseases. This could have a great social reflection, often underestimated, but driving in the long run to sensible social participation reduction. Because of the recent global economic crisis and with the increase of migratory fluxes from other Countries, the number of inhabitants within European borders that can be considered as at risk of poverty has increased and it is probably going to grow further. Therefore, addressing the problem of providing nutritionally viable food with a low cost is coming to be an important challenge to be faced at EU level.
The challenge here was to keep in mind that population at risk of poverty are anyway seeking tasteful foods, therefore to address the production to healthy, safe and low-cost products which were also tasteful and appreciated by consumers. The increasing flux of EU and extra EU migrants had also to be considered, since nutritional habits represent an important aspect of the cultural identity, and food choices are often focused on traditional foods. Another important point is the increase in the use of ready to eat food due to the reduction of time available for cooking.
One of the objective of the CHANCE programme of work was thus at firstly to increase the knowledge of the main nutritional criticalities and of the barriers that are dividing population at risk of poverty from an healthy nutrition.
The determination of the problematics behind the nutrition of such layers of populations found out through this projects could allow EU to act on several levels to obtain:
- A reduction of health problems of a significant proportion of the EU population (considering both the “already-poor” and the “at risk of poverty” EU citizens and migrants), which contributes disproportionately to the burden of associated chronic diseases across EU. The consequential reduction of health care costs connected to diet-related diseases will be a benefit as well as allowing this group of citizens to contribute more effectively to income generation.
- The possibility to develop new products or implement traditional foods with healthier traits that can integrate and hence improve diets of population at ROP, triggering the industrial interest into this new market sector. Optimization of the use of traditional and low-cost ingredients and raw materials into different food types will lead to the development of new food products. The tools and approaches developed by CHANCE will reduce the barriers to development of low-cost nutritious foods, especially targeted to SME food producers which prepare so much of European food. This will contribute to maintaining and increase the competitiveness of the European food industry.

Since the main thematic areas of the FP7 were Food, Agriculture and Fisheries, and Biotechnology Priority, CHANCE was addressing the challenge to meet the demand for safer, healthier, higher quality food with the affordable prices by bringing together excellent research centres and academia, industry and technology transfer centres.

The impacts of this project on the European Food Quality and Safety regulations and standards were made by cross-linking strategic research priorities such as:
- impact of food on health, by providing new formulations of healthy foods deriving from the optimization of traditional low cost resources, raw materials and ingredients;
- Safer and environmentally friendly production methods of foods and of healthier foodstuffs, by the development of improved/integrated food production processes (in production, handling and packaging) relying on the valorisation of raw materials of different origins;

The main strategy applied was the following:
- Delivering accessible and affordable health-value added foods targeted for specific social groups;
- Optimisation of the use of resources by the involvement of academia and industry (SMEs especially) sectors;
- Delivering scientifically sound data to policy makers in order to collaborate for the definition of an European policy on the regards of the thematic of poverty and nutrition.

As a consequence, the following impacts were reached:
1) Since CHANCE project helped in gaining a better understanding of existing barriers to healthy nutrition of populations at risk of poverty along the whole food chain, it was possible to define more effective ways to overcome unhealthy nutrition habits involving all chain actors and, therefore, allowing better access and supply of healthy food and meals to populations at risk of poverty
2) The project collected social and nutritional data on feeding habits of European ROP populations, based on metabonomic analysis and surveys realized in several European areas. In this way it is possible to address communitarian policies on nutrition, health and social issues.
3) CHANCE developed laboratory-scale experimental protocols, based on optimization of current processes turning raw materials into newly affordable formulated foods. Therefore, specifications on quality and safety were identified and gathered in protocols describing the foods and employed food processing strategies/technologies and the role of factors that might affect their economical and environmental impacts and legislative follow up.
4) Due to the integration of a robust scaffold of excellent and highly complementary European food expertise among the CHANCE project partners, it would be possible to improve career development opportunities within the food sector for young scientists, thus establishing the basis for the effective cooperation on the long-term in the field of the food technologies.
5) The interdisciplinary planned CHANCE protocols aimed at the assessment of quality, chemical and microbial safety of the new foods and feeds generated. Thus, synergic knowledge-based trans-national innovation for the food industry was generated.

CHANCE project impacts are generated on: public health, the European food industry, consumers, science and economics. As far as the impacts on Public Health concern, a reduction of low-quality food intake would guarantee a balanced assumption of nutrients and decrease of insurgency of diet-related health problems, thus improving the health status of low-income population. Moreover, the scientific data and tools so far collected will support policies and priorities in European public health.
As far as the impacts on the European Food Industry concerns, CHANCE model would increase the competitiveness of European food industry suggesting new formulations, and by providing some principles driving to the development of new market sectors previously not evaluated. The benefits to SMEs will come by production of foods which do not require important technological conversion of standard machinery and supplying, keeping the cost low and maximising the related benefits. The CHANCE protocol provides hints for further optimization of product quality and consumer acceptability, thus increased utility in the future, bringing higher level of innovation on the food products and companies. The increased competiveness of industry will generate new job opportunities
As far as the impacts on consumers concern, ROP people will have the possibility to buy foods with the quality, taste and convenience demanded, without being pushed towards consensus on food products that are less accepted. The possibility to preserve their cultural identity in food habits, while ameliorating their nutritional balance, is also taken into account.
In the food technology field there was the establishment of a technological methodology to produce specific meals for ROP consumers, the assessment of the use of low-cost formulation in term of final food quality and acceptability, determining physico-chemical, rheological and stability properties of new meals. New information on stability of quality parameters and on the objective determination of shelf-life of designed foods through kinetic studies during storage were obtained.
At last, the economic impacts include: i) long term benefits of nutritionally correct diets and consequent less expenses on Health Services; ii) benefits to the food industry, to both SMEs and large companies in the whole food production chain due to higher competitiveness; iii) new job opportunities generated.
List of Websites:
The homepage of the CHANCE project: http://www.chancefood.eu/

Coordinator: Prof. Francesco Capozzi UNIBO (francesco.capozzi@unibo.it)
Co-coordinator: Prof. Alessandra Bordoni UNIBO (alessandra.bordoni@unibo.it)
WP1 Leader: Prof. Vaiva Hendrixson VU (vaiva_hendrixson@yahoo.com)
WP2 leader: Dr. Maria Glibetic MRI (mglibetic@gmail.com)
WP3 leader: Prof. Søren Balling Engelsen UCPH (se@food.ku.dk)
WP4 leader: Dr. Anamarija Mandic FINS (anamarija.mandic@fins.uns.ac.rs)
WP5 leader: Dr Kaisa Poutanen VTT (Kaisa.Poutanen@vtt.fi)
WP6 leader: Prof. Andras Salgo BME (salgo@mail.bme.hu)
WP7 leader: Dr. Laura Fernandez EUFIC (laura.fernandez@eufic.org)
Project Manager. Dr. Alessandro Zamboni UNIBO (a.zamboni@unibo.it)