MEASURING THE OXIDATIVE STABILITY OF FISH OIL BY THE RANCIMAT TEST P.J. Garca-Moreno, M.M. Muo, R. Prez-Glvez, A.

Guadix, E.M. Guadix Department of Chemical Engineering, University of Granada, Spain.

Introduction Composition of fish oil is different from that of other oils due to its unique content of Omega3 polyunsaturated fatty acids. Among these PUFAs, particularly important are the eicosapentaenoic acid (C20:5 n-3, commonly called EPA) and the docosahexaneoic acid (C22:6 n-3, commonly called DHA) which have been reported to promote several benefits on human health (Lees and Karel, 1990; Uauy and Valenzuela, 2000). As consequence of its high degree of unsaturation, fish oil is very susceptible to oxidation. The oxidation process causes the development of rancid off-flavours which deteriorate the overall quality of the oil and reduce its shelf life. Different accelerated methods have been developed to assess the resistance of oils and fats to oxidation. Among these techniques, the Rancimat Test is the most commonly used due to its ease of use and reproducibility (Mndez et al., 1996). Moreover, this method neither requires periodic analytical nor organic solvents (Hasenhuettl and Wan, 1992). The Rancimat Test is based on the conductivity measurements of water which collects the volatile organic acids produced in an accelerated oxidation process. The aim of this work is to evaluate the influence of the operational parameters of the Rancimat Test, including temperature, air flow rate and sample weight, on the determination of the oxidative stability index (OSI) of a commercial cod liver oil.

Materials and methods The partially refined cod liver oil employed was purchased from Acofarma (Barcelona, Spain). This oil had a peroxide value of 3.12, an acid value of 0.16 and a composition of 8.18 % (w) in EPA and 11.02 % (w) in DHA. The oil was kept in a bottle covered with aluminium foil at 4 C under nitrogen atmosphere. Antioxidants were not added. A Metrohm Rancimat model 743 (Methrom Instruments, Herisau, Switzerland) was utilised. A stream of filtered, cleaned and dried air at different rates (10-15-20-25 L h-1) is bubbled into oil samples (3-5-7-9 g) contained in reaction vessels. These vessels are placed in an electric heating block which is set at different temperatures (60-70-80-90 C). Effluent air 1

containing volatile organic acids from the oil sample are collected in a measuring vessel with 60 mL of distilled water. The conductivity of the water is continuously recorded and the OSI is automatically determined by the apparatus. Analysis of variance (ANOVA) was performed with the statistical package Statgraphics ver. 5.1. Differences were considered significant at p<0.05.

Results and discussion The OSI results for the factorial experimental design carried out are presented in Table 1. As expected, the OSI values significantly decreased with temperature. However, the influence of the air flow rate and sample weight is not so obvious. In consonance with other studies (Farhoosh, 2007), the natural logarithms of the OSIs were fitted to a linear model, log(OSI) = b0 + b1M + b2Q + b3T, and the regression coefficients were calculated, Table 2. The correlation between experimental and predicted values for the natural logarithms of the OSIs is shown in Figure 1. As can be observed, most of the experimental points present a deviation of 10% between the predicted and measured variable, that is, they are placed in the region delimited by the dotted lines. This fact means that the linear model assayed can adequately predict the influence of the experimental factors studied upon the response variable, R2 = 0.9609. Furthermore, the statistical significance of the three parameters evaluated was studied by means of an analysis of variance, ANOVA. The p-values presented in Table 2 state that just the temperature is statistically significant and, therefore, it is the only operational parameter of the Rancimat Test which has significant influence on the OSI determination by this method. Also, this result was obtained by other authors (Mndez et al., 1996; Farhoosh, 2007).

Conclusions Firstly, the lineal model assayed, log(IP) = 3.1072 0.0012M 0.0018Q 0.0307T, having a high correlation coefficient R2 = 0.9609, predicts satisfactorily the influence of the experimental factors studied. Secondly, neither the air flow rate nor the sample weight have major influence on the OSI determination by the Rancimat Test. Temperature, therefore, is the only operational parameter of the Rancimat method statistically significant.

Acknowledgements This work was funded by the project CTQ2008-02978 from the Spanish Ministerio de Ciencia e Innovacin.

References Farhoosh, R, (2007). The effect of operational parameters of the Rancimat method on the determination of the oxidative stability measures and shelf-life prediction of soybean oil. Journal of the American Oil Chemists Society 84: 205-209. Hasenhuettl GL, Wan PJ, (1992). Temperature effects on the determination of oxidative stability with the Methrom RAncimat. Journal of the American Oil Chemists Society 69: 525-527. Lees RS, Karel M, (1990). Omega-3 fatty acids in health and disease. Marcel Dekker, New York. Mendez E, Sanhueza J, Speisky H, Valenzuela A, (1996). Validation of the rancimat test for the assessment of the relative stability of fish oils. Journal of the American Oil Chemists Society 73, 1033-1037. Uauy R, Valenzuela A, (2000). Marine Oils: the health benefits of n-3 fatty acids. Nutrition 16: 680-684.

Figures and Tables

1.4 1.2

Predicted log(OSI)

1 0.8 0.6 0.4 0.2 0 0 0.2 0.4 0.6 0.8 1 1.2 1.4 y = 0.9609x + 0.0298 R2 = 0.9609

Measured log(OSI)

Figure 1.Correlation between predicted and measured values of log(OSI)

Table 1.Oxidative Stability Index (OSI) for Cod Liver Oil assessed by the Rancimat Test

Exp. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32

T, C 60 60 60 60 60 60 60 60 60 60 60 60 60 60 60 60 70 70 70 70 70 70 70 70 70 70 70 70 70 70 70 70

Q, L/h 10 10 10 10 15 15 15 15 20 20 20 20 25 25 25 25 10 10 10 10 15 15 15 15 20 20 20 20 25 25 25 25

M, g 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9

OSI, h 18.03 17.91 17.76 17.98 17.71 17.51 17.55 17.72 17.67 17.56 17.34 17.63 18.12 17.35 17.38 17.46 8.12 8.03 8.09 8.15 8.10 8.01 7.90 8.08 8.01 7.97 7.85 7.87 8.21 8.04 7.78 8.03

Exp. 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64

T, C 80 80 80 80 80 80 80 80 80 80 80 80 80 80 80 80 90 90 90 90 90 90 90 90 90 90 90 90 90 90 90 90

Q, L/h 10 10 10 10 15 15 15 15 20 20 20 20 25 25 25 25 10 10 10 10 15 15 15 15 20 20 20 20 25 25 25 25

M, g 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9 3 5 7 9

OSI,h 4.22 3.95 3.97 4.06 3.85 3.76 3.81 3.89 3.47 3.38 3.33 3.42 3.31 3.42 3.11 3.26 1.97 2.00 2.04 2.08 1.93 1.93 1.93 1.96 3.47 3.38 3.33 3.42 1.78 1.74 1.69 1.52

Table 2. Statistical analysis.

Term b0 b1 b2 b3

Coefficient 3.1073 -0.0012 -0.0019 -0.0307

p-value 0.7587 0.2497 0.0000