The thiobarbituric acid test

One molecule of MDA reacts stoichiometrically with two molecules of 2-thiobarbituric acid (Sinnhurber et al., 1958 Yu et al., 1986) the reaction occurs at a pH of 2-3 but excess acid (pH 2) inhibits the colour development. 

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Nair, V. and Turner, G.A. (1984). The thiobarbituric acid test for lipid peroxidation structure of the adduct with malondi-aldehyde. Lipids 19, 804—805. 

Exposure to periodate yields 3-formylpyruvic acid, which reacts with thiobarbituric acid, giving rise to red condensation products having an absorption maximum at 545-550 nm. Certain workers have used the thiobarbituric acid test without previous oxidation of the unsaturated oligo-D-galactosiduronates with periodic acid.3-29,239,260 Even here, the products of reaction with thiobarbituric acid have an absorption maximum at 550 nm. 

Haase, G. and Dunkley, W. L. 1969A. Ascorbic acid and copper in linoleate oxidation. I. Measurement of oxidation by ultraviolet spectrophotometry and the thiobarbituric acid test. J. Lipid Res. 10, 555-560. 

The thiobarbituric acid test (AOCS method Cd 19-90), developed initially for measurement of malondialdehyde, is used to measure the oxidative degradation product of polyunsaturated fatty acids (47, 48). However, it lacks sensitivity (49). 

Brett R, Rumsby MG. Susceptibility of myelin gjycerophospholipids and sphingolipids to oxidative attack by hydroxyl free radicals as measured by the thiobarbituric acid test. Neurochem Int 1994 24 241-51. 

Bieri (1959) was able to demonstrate by the thiobarbituric acid test that peroxidation occurred in homogenates of chicken brain incubated in air at 37 °C for 1 hour. The degree of peroxidation in brain homogenates was much higher than in homogenates of liver and muscle moreover, the peroxidation in brain homogenate was independent of whether the chicks had received vitamin E or not. These results indicate that tocopherol does not easily pass the blood-brain barrier. 

Patton, S. Malonaldehyde, lipid oxidation, and the thiobarbituric acid test. J. Am. Oil Chem. Soc. 51, 114(1974). 

The thiobarbituric acid test (TBA) is a preferred method for detecting lipid peroxidation in biological systems. However, the reaction is nonspecific since a number of primary and secondary products of lipid peroxidation form malonaldehyde which in turn reacts in the TBA test. In food containing oleic and linoleic acids, the TBA-test is not as sensitive as the Fe +-test outlined above. 

The periodate-thiobarbituric acid test developed by Waravdekar and Saslaw269 is used in the modification of Weissbach and Hur-witz229 for identification of the products of /3-eliminative cleavage. 

Other indices measure a secondary stage of oxidation, such as the anisidine value (ANV), pointing to formation of carbonyl compounds, capable of undergoing condensation reactions with p-anisidine, and the thiobarbituric acid reactive substance (TBARS) pointing to the presence of malondialdehyde (MDA) in particular. In biological systems, TBARS is of widespread use as a measure for the extent of oxidation damage. Another test for stability of oils to oxidation is based on the development of acidity as secondary product, for example, standards using the Rancimat equipment or a similar setup. 

The periodic acid-thiobarbituric acid test has some disadvantages that may cause severe errors in the determination of sialic acid. 

Hoyland, D.V. and Taylor, A.J. 1991. A review of the methodology of the 2-thiobarbituric acid test. Food Chem. 40 271-291. 

Rhee, K.S. 1978. Minimization of further lipid peroxidation in the distillation 2-thiobarbituric acid test of fish and meat. J. Food Sci. 43 1776-1778, 1781. 

Tarladgis, B.G., Pearson, A.M., and Dugan, L.R. Jr. 1964. Chemistry of the 2-thiobarbituric acid test for determination of oxidative rancidity in foods. II.-Formation of the TBA-malonaldehyde complex without acid-heat treatment. J. Sci. Food Agric. 15 602-607. 

The aldehydic products from the autoxidation of PUFA have always been of interest, since the thiobarbituric acid (TBA) test detects malonaldehyde (or malonaldehyde precursors) with considerable sensitivity (76-78) and since malonaldehyde itself is mutagenic (H). More recently, Esterbauer and his coworkers have shown that 4-hydroxy-2-alkenals, and particularly 4-hydroxy-2-nonenal, are extremely cytotoxic (76f77,7Q). 

The thiobarbituric acid (TBA) test was proposed over 40 years ago and is now one of the most extensively used methods to detect oxidative deterioration of fat-containing foods (41). During lipid oxidation, malonaldehyde (MA), a minor component of fatty acids with 3 or more double bonds, is formed as a result of the degradation of polyunsaturated fatty acids. It is usually used as an indicator of the lipid oxidation process, both for the early appearance as oxidation occurs and for the sensitivity of the analytical method (42). In this assay, the MA is reacted with thiobarbituric acid (TBA) to form a pink MA-TBA complex that is measured spectrophotometrically at its absorption maximum at 530-535 nm (Figure 2) (9,43,44). The extent of oxidation is reported as the TBA value and is expressed as milligrams... 

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). 

The thiobarbituric acid (TBA) value test is a popular way of measuring rancidity in certain foods and oxidation products in biological systems. It is based on the formation of a colored complex between two molecules of TBA and one molecule of malonaldehyde resulting from thermal decomposition of polyunsaturated peroxides. This reaction is not specific due to the presence of many TBA-reactive substances (TBARS), such as browning reaction products, protein and sugar decomposition products, amino acids, nucleic acids and nitrite. 

The thiobarbituric acid (TEA) test is one of the most frequently used methods to assess lipid peroxidation, basically based on the determination of malonaldehyde which is assumed to be an important lipid oxidation product in food and biological systans. ... 

According to Zalkin and Tappel (1960) the in vivo peroxidation occurring in vitamin E deficiency damages the mitochondria in the tissues of rats, rabbits, and chicks as measured by oxygen absorption, thiobarbituric acid test, and enzyme inactivation. 

Wilbur, K. M., Bernheim, F., and Shapiro, O. W. 1949. The thiobarbituric acid reagent as a test for the oxidation of unsaturated fatty acids by various agents. Arch. Biochem. 24, 305. 

There are many studies relating the benefits of vitamin E supplementation to improved lipid stability of muscle foods. The majority of studies concerned with vitamin E supplementation and its effect(s) on improving lipid stability have often utilized the thiobarbituric acid (TEA) test to monitor rancidity development. A high TEA value represents high lipid oxidation, while a lower TEA value indicates less rancidity development. The specific treatments, and subsequent benefits of the various studies previously cited, cannot be individually addressed in the restricted space of this chapter. However, the general trends of the studies are similar in that supplementation generally results in improved lipid stability. Noteworthy aspects of some specific studies are highlighted as follows. 

The extent of prior oxidative activity in a fat sample may be estimated by the following peroxide value (PV), thiobarbituric acid test (TEA) (which measures the presence of malonaldehyde), anisidine value—Totox, Kreis test, oxirane test, total and volatile carbonyl... 

The simple spectrophotometric thiobarbituric acid (TBA) test has been frequently used for many years as an indicator of the peroxidation of PUFAs present in biological matrices. This test involves the reaction of aldehydes in the sample with TBA at c. 100°C under acidic conditions (Equation 1.13) to produce a pink-coloured chromogen, which absorbs light strongly at a wavelength of 532 nm (Nair and Turner, 1984). 

The products formed during lipid peroxidation include unsaturated aldehydes, such as 4-hydroxynonenal. Their quantification is of great interest because of their extremely reactive and cytotoxic properties. This extreme reactivity and metabolic conversion, however, may make them unsuitable as test analytes for in vivo antioxidant activity studies except at high levels of oxidative stress. Furthermore, simple chemical tests such as the TBARS (thiobarbituric acid reactive substances) and LPO-586 (colorimetric... 

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