ANÁLISIS E INTERPRETACIÓN DE LA OBSERVACIÓN INICIAL A LOS

Cl1 Cl2 Cl3 Cl4 Cl5 Cl6 Cl7 Cl8 Cl9 Cl10 Cl11 Cl12 Cl13 Cl14 BEF vs PLA 1,00 0,482 0,920 0,227 0,494 0,000 0,178 0,878 0,959 0,854 0,052 0,006 0,000 0,036 BEF Cl1 Cl2 Cl3 Cl4 Cl5 Cl6 Cl7 Cl8 Cl9 Cl10 Cl11 Cl12 Cl13 Cl14 T0 vs T20 0,757 0,632 0,313 0,951 0,966 0,187 0,633 0,259 0,533 0,243 0,398 0,348 0,517 0,609 T0 vs T40 0,008 0,382 0,141 0,733 0,425 0,008 0,796 0,893 0,931 0,063 0,946 0,146 0,452 0,931 T0 vs T60 0,823 0,203 0,866 0,574 0,513 0,456 0,007 0,309 0,723 0,243 0,057 0,919 0,929 0,894 T20 vs T40 0,216 0,706 0,189 0,145 0,205 0,379 0,933 0,777 0,587 0,994 0,103 0,218 0,012 0,380 T20 vs T60 0,897 0,693 0,034 0,213 0,668 0,807 0,027 0,796 0,980 0,693 0,016 0,706 0,029 0,116 T40 vs T60 0,069 0,620 0,361 0,396 0,485 0,450 0,193 0,840 0,929 0,697 0,281 0,524 0,109 0,002 PLA Cl1 Cl2 Cl3 Cl4 Cl5 Cl6 Cl7 Cl8 Cl9 Cl10 Cl11 Cl12 Cl13 Cl14 T0 vs T20 0,839 0,901 0,960 0,003 0,128 0,000 0,179 0,525 0,241 0,231 0,498 0,206 0,133 0,723 T0 vs T40 0,827 0,722 0,099 0,134 0,418 0,008 0,576 0,168 0,097 0,199 0,215 0,032 0,048 0,758 T0 vs T60 0,349 0,499 0,588 0,434 0,944 0,927 0,094 0,172 0,982 0,563 0,661 0,669 0,613 0,856 T20 vs T40 0,025 0,364 0,294 0,005 0,870 0,013 0,899 0,461 0,192 0,371 0,441 0,694 0,000 0,267 T20 vs T60 0,932 0,249 0,430 0,825 0,479 0,078 0,347 0,792 0,705 0,592 0,601 0,613 0,217 0,891 T40 vs T60 0,042 0,574 0,567 0,228 0,108 0,163 0,582 0,817 0,371 0,814 0,298 0,710 0,009 0,068 BEF vs PLA – Time points Cl1 Cl2 Cl3 Cl4 Cl5 Cl6 Cl7 Cl8 Cl9 Cl10 Cl11 Cl12 Cl13 Cl14 T0 BEF vs PLA 0,923 0,098 0,872 0,486 0,009 0,007 0,369 0,532 0,494 0,474 0,138 0,489 0,133 0,295 T20 BEF vs PLA 0,296 0,949 0,069 0,513 0,850 0,001 0,432 0,634 0,454 0,701 0,500 0,341 0,043 0,602 T40 BEF vs PLA 0,352 0,860 0,376 0,641 0,969 0,042 0,449 0,415 0,242 0,526 0,694 0,536 0,000 0,064 T60 BEF vs PLA 0,078 0,800 0,271 0,223 0,074 0,096 0,614 0,560 0,425 0,798 0,067 0,661 0,085 0,051

BEF and PLA samples had differences in their profiles, though these were not that evident along the storage, proving that the enrichment did not effectively affect the shelf-life of the product. The trend observed for some signals, where the area decreases at T20 to later increase again, might be due to the reorganization of carbohydrates and sugar molecules during storage (i.e. staling processes, starch retrogradation, etc.), though no literature is present on 1H-NMR study of this specific behaviour.

Conclusion

The proposed method for the evaluation of the food matrix stability is based on the molecular profile of its aqueous extract, acquired through NMR spectroscopy. In this study two different

165

types of pancakes, enriched with two bioactive ingredients or controls, were evaluated during their shelf-lives. The method employed consisted in the acquisition of NMR spectra of samples and the evaluation of their profile variations during storage. It was seen that PLA and BEF pancakes had different molecular profiles, though the effect of storage was masked by differences in the production of the pancake batches. Further studies are needed for the evaluation of the shelf-life of the products, though no major impact on the matrix was seen in the selected conditions and period of storage, proving that the enrichment did not cause any major stress to the food matrix. The study proposes an effective analytical pipeline for this kind of evaluations and can be employed in further stability assessments.

5.3 Final conclusions

The present studies on various enriched food products represent the first step towards the devlopment of a specific analytical method employing NMR for the investigation of the stability of a food matrix. These studies have shown the capability of NMR spectroscopy to capture a lot of information on what occurs in a food items in relation to many concurrent factors (enrichment, bacth production, storage) and the consequences of these factors and their interaction on the molecular profile of the food item analyzed. In effect, the non-target approach of NMR analysis allows to define a more complex metabolite profile for the food product and observe even unforeseen and otherwise not visible changes in its matrix. This is particularly important in enriched products which have to undergo a great variety of testing before being ready for the market. It is in fact fundamental that the bioactive molecules added in the product are stable during shelf-life but can easily become available when the product is consumed and the capability of this method to picture the interaction between the food matrxi and the added molecule is of great help in this kind of assay.

166

REFERENCES CHAPTER 5

Augustin, Mary Ann, and Luz Sanguansri. "Challenges and Solutions to Incorporation of Nutraceuticals in Foods. “Annual review of food science and technology” 6 (2015): 463-477.

Ciampa et al., 2012 “Changes in the Amino Acid Composition of Bogue (Boops boops) Fish during Storage at Different

Temperatures by 1_{H-NMR Spectroscopy” – Nutrients (2012), 4(6), 542-553.}

Dave N. et al., 2014, "Antioxidant Activity, Textural Properties and Storage Stability of Honey Fortified Biscuits." (2014). Dieterle, F., Ross, A., Schlotterbeck, G., & Senn, H. (2006). Probabilistic quotient normalization as robust method to account for dilution of complex biological mixtures. Application in 1H NMR metabonomics. Analytical chemistry,78(13), 4281-4290.

Edel A. L. et al., 2015, "Stability of bioactives in flaxseed and flaxseed-fortified foods." Food Research International. Fundo J. F. et al., 2015, “Molecular Dynamics and Structure in Physical Properties and Stability of Food Systems”. Ganesh V. et al., A Review: Supplementation of Foods with Essential Fatty Acids – Can It Turn a Breeze without Further Ado?, 2015, Critical Reviews in Food Science and Nutrition.

Heertje I., Structure and function of food products: A review, 2014, Food Structure, 1, 3-23.

Kilcast, D., & Subramaniam, P. (2000) “The Stability and Shelf-Life of Food” Elsevier, ISBN: 978-1-85573-500-2

Laghi et al., 2014 - Laghi, Luca, Gianfranco Picone, and Francesco Capozzi "Nuclear magnetic resonance for foodomics beyond food analysis." TrAC Trends in Analytical Chemistry 59 (2014): 93-102.

Li-Chan, Eunice CY. "Bioactive peptides and protein hydrolysates: research trends and challenges for application as nutraceuticals and functional food ingredients. “Current Opinion in Food Science” 1 (2015): 28-37.

López-Córdoba et al., Yerba mate antioxidant powders obtained by co-crystallization: Stability during storage, Journal of Food Engineering 124 (2014) 158–165.

Mc Clements D. J., 2015, “Enhancing nutraceutical bioavailability through food marix design”, Current Opinion in Food Science, 4:1-6.

Moro A. et al., 2015, “Dairy matrix effect on the transference of rosemary (Rosmarinus officinalis) essential oil compounds during cheese making”, J. Sci. Food Agric., 95: 1507-1513.

Norton J. et al., 2014, “Designing Food Structures for Nutrition and Health Benefits”, Annu. Rev. Food Sci. Technol., 5:177-95.

Picone et al. (2011) "Metabolomics as a Powerful Tool for Molecular Quality Assessment of the Fish Sparus aurata." Nutrients 3.2.

Salcedo-Sandoval L. et al., Shelf-life of n-3 PUFA enriched frankfurters formulated with a konjac-based oil bulking agent, LWT – Food Science and Technology 62 (2015) 711-717.

Samuelsen A. et al., Effects of orally administered yeast-derived beta-glucans: A review, Mol. Nutr. Food Res. 2014, 58, 183–193.

Sinn N. et al., Effects of n-3 fatty acids, EPA v. DHA, on depressive symptoms, quality of life, memory and executive function in older adults with mild cognitive impairment: a 6-month randomised controlled trial, British Journal of Nutrition (2012), 107, 1682–1693.

167

Turgeon S. L. et al., 2011, “Food matrix impact on macronutrients nutritional properties”, Food hydrocolloids, 25 1915- 1924.

Yang M. et al., 2011, “Food Matrix Affecting Anthocyanin Bioavailability: Review”, Current Medicinal Chemistry, 18:291- 300.

Yousouf et al., Health Benefits of Anthocyanins and Their Encapsulation for Potential Use in Food Systems: A Review, 2015, Critical Reviews in Food Science and Nutrition.

168

CHAPTER 6: FOODOMICS TO INVESTIGATE THE DIGESTION