Food Analytical Methods

, Volume 12, Issue 10, pp 2150–2160 | Cite as

Measuring Primary Lipid Oxidation in Food Products Enriched with Colored Microalgae

  • Lore GheysenEmail author
  • Céline Dejonghe
  • Tom Bernaerts
  • Ann Van Loey
  • Luc De Cooman
  • Imogen Foubert


Microalgae are a valuable alternative source of n-3 LC-PUFA and have already proven their potential in different food products. However, enrichment of food products with n-3 LC-PUFA implies an increased sensitivity to lipid oxidation. Numerous analytical techniques have already been developed to determine and follow lipid oxidation. Photoautotrophic microalgae often contain, besides n-3 LC-PUFA, other compounds like carotenoids and chlorophylls. These colored compounds may interfere with the standard analytical techniques. This study contributes to optimize a simple and low-cost method to measure the degree of primary lipid oxidation in food products enriched with photoautotrophic microalgae. The standard iodometric titration and three spectrophotometric methods (FOX, IDF, and CD & CT) were investigated. The FOX method was selected as the preferable method, although interference due to the presence of metal ions could occur. This could partially be solved by a supplemental step with TPP addition. However, as this additional step did not change the trend in oxidative values during storage, it was suggested to normalize the values of the FOX method to week zero to investigate the trend of oxidation during storage of products enriched with photoautotrophic microalgae.


Photoautotrophic microalgae Colored food product Peroxide value Lipids Hydroperoxide determination 



Conjugated dienes & conjugated trienes


Docosahexaenoic acid


Eicosapentaenoic acid

FOX method

Ferrous oxidation–xylenol orange method

IDF method

Ferric thiocyanate-based International Dairy Federation method


Omega-3 long chain polyunsaturated fatty acids





We want to thank K. Raes (Ghent University Campus Kortrijk, Belgium) for the support with the ICP-OES analysis.


The research presented in this paper was financially supported by the Research Foundation - Flanders (FWO SB PhD fellowship Lore Gheysen 1S 151287 16N and Tom Bernaerts 1S 099 16N) and Industrial Research Fund KULeuven (IOF-KP Vegetalgae).

Compliance with Ethical Standards

Conflict of Interest

Lore Gheysen declares that she has no conflict of interest. Céline Dejonghe declares that she has no conflict of interest. Tom Bernaerts declares that he has no conflict of interest. Ann Van Loey declares that she has no conflict of interest. Luc De Cooman declares that he has no conflict of interest. Imogen Foubert declares that she has no conflict of interest.

Ethical Approval

No conflicts, informed consent, human or animal rights applicable.

Informed Consent

Not applicable.


  1. Abuzaytoun R, Shahidi F (2006) Oxidative stability of algal oils as affected by their minor components. J Agric Food Chem 54:8253–8260CrossRefGoogle Scholar
  2. Antolovich M, Prenzler PD, Patsalides E, McDonald S, Robards K (2002) Methods for testing antioxidant activity. Analyst 127:183–198CrossRefGoogle Scholar
  3. AOCS (2003) Official methods and recommended practices of the AOCS, 5th edn. American Oil Chemists’ Society, ChampaignGoogle Scholar
  4. Ashoka S, Peake B, Bremner G, Hageman K, Reid M (2009) Comparison of digestion methods for ICP-MS determination of trace elements in fish tissues. Anal Chim Acta 653(2):191–199CrossRefGoogle Scholar
  5. Barriuso B, Astiasarán I, Ansorena D (2013) A review of analytical methods measuring lipid oxidation status in foods: a challenging task. Eur Food Res Technol 236:1–15CrossRefGoogle Scholar
  6. Belleza OJV, Villaraza AJL (2014) Ion charge density governs selectivity in the formation of metal-xylenol orange (M-XO) complexes. Inorg Chem Commun 47:87–92CrossRefGoogle Scholar
  7. Bou R, Codony R, Tres A, Decker EA, Guardiola F (2008) Determination of hydroperoxides in foods and biological samples by the ferrous oxidation-xylenol orange method: a review of the factors that influence the method’s performance. Anal Biochem 377:1–15CrossRefGoogle Scholar
  8. Brajter K, Olbrych-Śleszyńska E (1983) Application of xylenol orange to the separation of metal ions on amberlyst A-26 macroreticular anion-exchange resin. Talanta 30:355–358CrossRefGoogle Scholar
  9. Colston BJ, Robinson VJ (1997) Use of metal ion indicators to determine complex stability constants: the method of competitive equilibration. Analyst 122:1451–1455CrossRefGoogle Scholar
  10. DeLong JM, Prange RK, Hodges DM, Forney CF, Bishop MC, Quilliam M (2002) Using a modified ferrous oxidation-xylenol orange (FOX) assay for detection of lipid hydroperoxides in plant tissue. J Agric Food Chem 50:248–254CrossRefGoogle Scholar
  11. Farhoosh R, Moosavi SMR (2009) Evaluating the performance of peroxide and conjugated diene values in monitoring quality of used frying oils. J Agric Sci Technol 11:173–179Google Scholar
  12. Frankel EN (2005) Lipid oxidation, 2nd edn. Oily Press, BridgewaterCrossRefGoogle Scholar
  13. Gay C, Collins J, Gebicki JM (1999) Determination of iron in solutions with the ferric – xylenol orange complex. 148:143–148Google Scholar
  14. Gheysen L, Matton V, Foubert I (2018a) Microalgae as a source of omega-3 polyunsaturated fatty acids. In: Catala A (ed) Polyunsaturated fatty acids (PUFAs): food sources, health effects and significance in biochemistry. Nova Science Publishers, New YorkGoogle Scholar
  15. Gheysen L, Bernaerts T, Bruneel C, Goiris K, Van Durme J, Van Loey A, De Cooman L, Foubert I (2018b) Impact of processing on n-3 LC-PUFA in model systems enriched with microalgae. Food Chem 268:441–450CrossRefGoogle Scholar
  16. Giri A, Osako K, Ohshima T (2010) Identification and characterisation of headspace volatiles of fish miso, a Japanese fish meat based fermented paste, with special emphasis on effect of fish species and meat washing. Food Chem 120:621–631CrossRefGoogle Scholar
  17. Gotoh N, Miyake S, Takei H, Sasaki K, Okuda S, Ishinaga M, Wada S (2011) Simple method for measuring the peroxide value in a colored lipid. Food Anal Methods 4:525–530CrossRefGoogle Scholar
  18. Hornero-Méndez D, Pérez-Gálvez A, Mínguez-Mosquera MI (2001) A rapid spectrophotometric method for the determination of peroxide value in food lipids with high carotenoid content. JAOCS J Am Oil Chem Soc 78:1151–1155CrossRefGoogle Scholar
  19. Jacobsen C, Sørensen ADM, Nielsen NS (2013) Stabilization of omega-3 oils and enriched foods using antioxidants, food enrichment with omega-3 fatty acids. Woodhead Publishing LimitedGoogle Scholar
  20. Johnson DR, Decker EA (2015) The role of oxygen in lipid oxidation reactions: a review. Annu Rev Food Sci Technol 6:171–190CrossRefGoogle Scholar
  21. Marmesat S, Morales A, Velasco J, Ruiz-Méndez MV, Dobarganes MC (2009) Relationship between changes in peroxide value and conjugated dienes during oxidation of sunflower oils with different degree of unsaturation. Grasas Aceites 60:155–160CrossRefGoogle Scholar
  22. Mizuguchi H, Yotsuyanagi T (2001) Visual threshold detection of trace metal ions using a bi-functional metallochromic reagent. Anal Sci 17:1687–1689Google Scholar
  23. Morales-Sánchez D, Martinez-Rodriguez OA, Kyndt J, Martinez A (2015) Heterotrophic growth of microalgae: metabolic aspects. World J Microbiol Biotechnol 31:1–9CrossRefGoogle Scholar
  24. Nielsen NS, Timm-Heinrich M, Jacobsen C (2003) Comparison of wet-chemical methods for determination of lipid hydroperoxides. J Food Lipids 10:35–50CrossRefGoogle Scholar
  25. Nourooz-Zadeh J, Tajaddini-Sarmadi J, Wolff SP (1994) Measurement of plasma hydroperoxide concentrations by the ferrous oxidation-xylenol orange assay in conjunction with triphenylphosphine. Anal BiochemGoogle Scholar
  26. Orefice I, Gerecht A, D’Ippolito G, Fontana A, Ianora A, Romano G (2015) Determination of lipid hydroperoxides in marine diatoms by the FOX2 assay. Mar Drugs 13:5767–5783CrossRefGoogle Scholar
  27. Rawat I, Ranjith Kumar R, Mutanda T, Bux F (2013) Biodiesel from microalgae: a critical evaluation from laboratory to large scale production. Appl Energy 103:444–467CrossRefGoogle Scholar
  28. Ryckebosch E, Bruneel C, Muylaert K, Foubert I (2012a) Microalgae as an alternative source of omega-3 long chain polyunsaturated fatty acids. Lipid Technol 24:128–130CrossRefGoogle Scholar
  29. Ryckebosch E, Muylaert K, Foubert I (2012b) Optimization of an analytical procedure for extraction of lipids from microalgae. JAOCS J Am Oil Chem Soc 89:189–198CrossRefGoogle Scholar
  30. Ryckebosch E, Bruneel C, Termote-Verhalle R, Lemahieu C, Muylaert K, Van Durme J, Goiris K, Foubert I (2013) Stability of omega-3 LC-PUFA-rich photoautotrophic microalgal oils compared to commercially available omega-3 LC-PUFA oils. J Agric Food Chem 61:10145–10155CrossRefGoogle Scholar
  31. Schaich KM (2016) Chapter 1 – Analysis of lipid and protein oxidation in fats, oils, and foods, oxidative stability and shelf life of foods containing oils and fats. Elsevier IncGoogle Scholar
  32. Schaich KM, Shahidi F, Zhong Y, Eskin NAM (2013) Lipid oxidation, Third Edit. ed, Biochemistry of Foods. ElsevierGoogle Scholar
  33. Shahidi F, Zhong Y (2005) Lipid oxidation: measurement methods. Bailey’s Ind. Oil fat prodGoogle Scholar
  34. Shantha NC, Decker EA (1994) Rapid, sensitive, iron-based spectrophotometric methods for determination of peroxides values of food lipids. J AOAC Int 77:421–424Google Scholar
  35. Södergren E, Nourooz-Zadeh J, Berglund L, Vessby B (1998) Re-evaluation of the ferrous oxidation in xylenol orange assay for the measurement of plasma lipid hydroperoxides. J Biochem Biophys Methods 37:137–146CrossRefGoogle Scholar
  36. Vieira SA, Zhang G, Decker EA (2017) Biological implications of lipid oxidation products. J Am Oil Chem Soc 94:339–351CrossRefGoogle Scholar
  37. Wang N, Ma T, Yu X, Xu L, Zhang R (2016) Determination of peroxide values of edible oils by ultraviolet spectrometric method. Food anal. Methods 9:1412–1417Google Scholar
  38. Wrolstad RE, Acree TE, Decker EA, Penner MH, Reid DS, Schwartz SJ, Shoemaker CF, Smith DM, Sporns P (2005) Handbook of food analytical chemistry, volume 1 : water, protein, enzymes, lipids, and carbohydrates. Handb Food Anal ChemGoogle Scholar

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© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Research Unit Food & LipidsKU Leuven KulakKortrijkBelgium
  2. 2.Leuven Food Science and Nutrition Research Centre (LFoRCe)KU LeuvenLeuvenBelgium
  3. 3.Laboratory of Food TechnologyKU LeuvenLeuvenBelgium
  4. 4.Laboratory of Enzyme, Fermentation and Brewing TechnologyKU Leuven Technology Campus GhentGhentBelgium

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