Methods for Measuring Lipid Oxidation

Other accelerated methods for measuring lipid oxidation include the oxygen bomb test and the Schaal oven test. These methods are described by Wan (1995). 

Pikul, J., Leszczynski, D.E., and Kummerow, F.A. 1989. Evaluation of three modified TBA methods for measuring lipid oxidation in chicken meat. J. Agric. Food Chem. 37 1309-1313. 

Salih, A.M., Smith, D.M., Price, J.F., and Dawson, L.E. 1987. Modified extraction 2-thiobarbituric acid method for measuring lipid oxidation in poultry. Poultry Sci. 66 1483-1488. 

As oxidation normally proceeds very slowly at the initial stage, the time to reach a sudden increase in oxidation rate is referred to as the induction period (6). Lipid hydroperoxides have been identified as primary products of autoxidation decomposition of hydroperoxides yields aldehydes, ketones, alcohols, hydrocarbons, volatile organic acids, and epoxy compounds, known as secondary oxidation products. These compounds, together with free radicals, constitute the bases for measurement of oxidative deterioration of food lipids. This chapter aims to explore current methods for measuring lipid oxidation in food lipids. 

There are many other methods for measuring lipid oxidation and quality by chemical means. Among the best-known procedures are the thiobarbituric acid (TEA) test, carbonyl value, and headspace oxygen analysis. These methods have been reviewed and discussed elsewhere (287, 307). 

A simple method for assessing lipid oxidation is measuring the headspace concentration of hexanal by capillary GLC. Also, the total volatiles appearing in the chromatogram up to hexanal can be taken as oxidation index. The method was applied to determine the amounts of lipid peroxides present in rat liver cells. Enhancement of the hexanal concentration can be achieved on adding ascorbic acid (22), that reduces Fe(ni) present in the matrix to Fe(II), which catalyzes decomposition of hydroperoxides to aldehydes. Significant correlations are found between hexanal concentrations and various oxidation indices, such as TBARS (Section IV.D.2)" . ... 

Lillard, D. A. and Day, E. A. 1961. Autoxidation of milk lipids. II. The relationship of sensory to chemical methods for measuring the oxidized flavor of milk fats. J. Dairy Sci. 44, 623-632. 

Measurement of hydroperoxides is the classical method for quantifying lipid oxidation and a variety of assay procedures are available. The oxidation of ferrous to ferric iron by hydroperoxides in the presence of ammonium thiocyanate to produce ferric thiocyanate, which can be quantified spectrophotometrically at 505 nm, has been used extensively to study lipid oxidation in milk (Loftus-Hills and Thiel, 1946). Newstead and Head-ifen (1981) recommend that extraction of fat from whole milk powder be carried out in the dark when using this procedure to avoid artefactually high... 

For these reasons, and because they can be measured in urine and plasma, PGF are generally considered to offer a noninvasive, sensitive, specific direct method for measuring lipid peroxidation in vivo (Y1). The main drawback is that the methods are very complicated, tedious, and not usually available in clinical laboratories. Like other lipid oxidation products, PGF can be generated ex vivo. For this reason, fluids should be preserved with butylated hydroxy toluene (BHT) and EDTA to prevent further oxidation and measured immediately or stored at —70°C (M8). An ELISA has been developed for urine for 8-Iso-PGF2a that is commercially available (02), but the method is tedious since it requires a column separation prior to ELISA (Bl). Also, since PGF are mainly indicators of arachidonic acid oxidation, they do not reflect oxidation of the major PUFA comprising lipoproteins. 

Methods to measure lipid oxidation have generally been unspecific and not sufficiently sensitive to measure flavor and low levels of oxidative deterioration of food lipids. Several specific complementary methods are needed to determine the contribution of lipid oxidation products formed at different stages of the multi-step process of oxidative deterioration in foods. Since polyunsaturated hydroperoxides decompose more readily at higher temperatures to form aldehydes causing rancidity, it is important to measure both aldehydes and hydroperoxides to monitor lipid oxidation. For the reliable prediction of shelf life of foods containing polyunsaturated lipids, it is essential to use more than one specific method to determine oxidation at different levels and to store samples at several temperatures, preferably between 40 and 60°C. 

The TBA test is perhaps the most widely used method for determining lipid peroxidation. The representative adduct of lipid peroxidation, malondialdehyde, forms a 1 2 adduct with TBA that can be measured by spectroscopy or fluorometry. The general procedure, of which there are numerous variations, simply involves heating a small quantity of the test substance for a defined period of time in an aqueous acidic solution of TBA, and then measuring the absorbance (535 nm) of the red color which is produced in the TBA reaction. It should be considered as an index of oxidative stress that represents primarily lipid peroxidation. ... 

Vogt, G., Aursand, M., Aaby, K., and Nilsson, A. 2001. Evaluation of conventional methods for measuring oxidation in fish oil, Lipidforum, 21 stNordic Lipid Symposium, Bergen, June 5-8, 2001. 

To evaluate oxidative stability, different methods are used to measure lipid oxidation after the sample is oxidized under standardized conditions to a suitable end-point. In Table 7.1, different lipid oxidation tests are ranked in decreasing order of usefulness in predicting Ae stability or shelf life of a food product. Methods for sensory evaluations, conjugated diene, gas chromatography of volatiles, peroxide values and thiobarbituric acid-reacting substances were discussed in Chapter 5. 

There is increasing evidence that degradation of proteins is associated with the development of the so-called warmed-over flavor . Selective methods to measure protein oxidation may be useful as a complementary approach in evaluating oxidative deterioration of meat products. The determination of protein carbonyls (as dinitrophenyl hydrazine derivatives) is now used for this purpose. More reliable methods are needed, however, to determine specific products of lipid oxidation and their interaction products acting as precursors of flavor compounds to establish the relative contribution of heme and nonheme iron to the development of rancidity in various meat products. 

Decomposition of the primary products of lipid oxidation generates a complex mixture including saturated and unsaturated aldehydes such as hexanal. Hexanal is the most commonly measured end product of lipid oxidation, and both sensory and physicochemical methods are used for its determination. Where other antioxidant activity tests may be nonspecific, physicochemical measurement of hexanal offers the advantage of analyzing a single, well-defined end product. 

A procedure for determination of lipid hydroperoxides in human plasma is based on kinetic measurement of the CL of luminol (124) with hemin (75a) catalysis . CLD of microperoxidase-catalyzed oxidation of luminol (124) or isoluminol (190) was applied to detection and determination of amino acid hydroperoxides after exposure to UV and y-irradiation A method for determination of hydroperoxides in the phospholipids of cultured cells uses isoluminol (190) and microperoxidase as catalyst " . Simultaneous determination of phosphatidylcholine hydroperoxides and cholesteryl ester hydroperoxides in human serum is carried out by quantitative extraction of the lipids, HPLC separation by column switching and CLD using isoluminol (190) with microperoxidase catalysis . ... 

---