## 3 Results and Discussion
### Case Study Review: Analytical Advantages Offered by \({}^{1}\)H NMR Analysis of LOPs in CFOs
#### 3.6.2 Spectrophotometric TBARS Test  
The authors of the current paper also continue to express much concern that the spectrophotometrically-based thiobarbituric acid (TBA)-reactive substances (TBARS) test is still neglectfully very frequently employed for determinations of its malondialdehydedehyde (MDA) target analyte, in edible oils and other foodstuffs [68], despite the availability of many reports which clearly demonstrate and confirm the extremely limited value of any results arising therefrom. Indeed, such results are completely unreliable and serve little or no value for the estimation of these species. We are, of course, already aware of the induction of the lipid peroxidation process at standard frying temperatures, and since the TBARS test requires the heating of oil or food extracts with TBA at ca. 96 \({}^{\circ}\)C for periods of 15 min or more, this temperature is certainly more than sufficient to induce the peroxidation of PUFAs located therein. Hence, in general it appears that all results derivable from this heat-dependent test system represent nothing more than artefactual data. Indeed, our \({}^{1}\)H NMR experiments have clearly proven the thermally-induced oxidation of PUFAs in commercial oil formulations to CHPDs and aldehyde LOPs at both Pasteurisation and ambient room temperatures (72 and ca. 25 \({}^{\circ}\)C, respectively) [12]. Moreover, further caution should always be employed, since it is known that a series of additional agents present in foods in general and not CFOs, especially reducing and non-reducing sugars, also react with TBA to form chromophores which also absorb strongly at a wavelength of 532 nm [69; 70]. Hence, if it was not already producing artefactual results from the thermal stressing of MUFAs and PUFAs, this test's specificity would be less than zero. Attempts to improve the selectivity and specificity of this aberrant test system by the separation of the 2:1 TBA:MDA chromophore from others generated during the heating phase of the assay are of little or no value, however. Indeed, this 'improvement' is, of course, also of negligible value if the MDA is already artefactually generated at the elevated temperatures involved in chromophore development. If, however, this method involves a prolonged equilibration at ambient temperature, and the 2:1 TBA:MDA chromophore arising (\(\lambda_{\text{max}}\) 532 nm, \(\varepsilon=1.55\)\(\times\) 10\({}^{5}\) M\({}^{-1}\) cm\({}^{-1}\)) is then allowed to develop slowly, perhaps for 24 h or more, then this test system does have the potential to monitor aldehyde LOPs in unheated or heated oils, but not exclusively MDA, since a wide range of other aldehydes (including \(\alpha,\beta\)-unsaturated ones) also react with TBA to generate the same or similar chromophores, or similar interfering adducts. Notwithstanding, we have previously shown that the careful extraction of water-soluble MDA from thermally-stressed or unheated CFOs into an aqueous medium prior to the above heating stage of the assay serves to overcome this problem [19]. Indeed, visible region electronic absorption spectra acquired from such aqueous extracts demonstrate that the characteristic 2:1 TBA:MDA chromophore absorption bands are clearly distinguished from those of other coloured products present (Figure 6). Moreover, this colourimetric method is also suitable for producing an impacting visual display of aldehyde formation during the heating of UFA-containing CFOs (Figure 7); the intensity of the red colouration formed is, of course, critically dependent on the unsaturation status of the oil tested (with PUFA-rich corn and sunflower oils generating much higher intensities than MUFA-rich olive oil, for example), and the heating exposure period at standard frying temperature (generally, but not exclusively at 180 \({}^{\circ}\)C).