Total antioxidant capacities of raw and cooked meats Arda Serpena,

a ,

, Vural Gkmena, b, Vincenzo Foglianoc

Food Research Center, Hacettepe University, 06800 Beytepe, Ankara, Turkey Department of Food Engineering, Hacettepe University, 06800 Beytepe, Ankara, Turkey University of Naples Federico II, Department of Food Science, I-80055 Portici, Naples, Italy

Received 15 February 2011; revised 28 May 2011; Accepted 31 May 2011. Available online 7 June 2011. Abstract This study investigated the total antioxidant capacity (TAC) of meats (beef, chicken, pork and fish) and its changes on thermal treatment. The QUENCHER procedure, which is performed directly on the solid material without extraction, was selected and proved to be particularly suitable for meat samples. The ABTS + scavenging capacity of raw meats ranged between 25.9 1.0 and 51.7 1.2 mmol Trolox Eq./kg. Raw chicken had the highest TAC followed by pork, beef and fish. Upon heating at 180 C, TAC of meats increased to an apparent maximum at 5 min followed by sudden decreases until 15 min, while the final stage of heating was characterized by slight increases. The modifications of TAC during cooking can be explained considering factors such as denaturation and exposure of reactive protein sites, degradation of endogenous antioxidants and the formation of Maillard reaction products having antioxidant properties. Highlights We investigate the total antioxidant capacity (TAC) of meats by QUENCHER Procedure. We examine the TAC changes of meats along with thermal treatment. Denaturation and exposure of reactive protein sites primarily affects TAC of meats. Formation of Maillard reaction products upon cooking possibly modify TAC of meats. Balance of endogenous prooxidant and antioxidant factors are crucial in TAC of meats. Keywords: Meat; Antioxidant activity; Quencher procedure; Heating; Denaturation Article Outline
o o

1. Introduction 2. Materials and methods

2.1. Chemicals 2.2. Thermal treatments of meat samples

o o o

2.3. Preparation of meat samples for QUENCHER procedure 2.4. Preparation of ABTS +, DPPH and FRAP radical solutions

o o

2.4.1. ABTS

solution

2.4.2. DPPH solution 2.4.3. FRAP solution

2.5. Measurement of Trolox Equivalent TAC of meats by direct QUENCHER method 2.6. Statistical analysis

3. Results and discussion 4. Conclusions References

1. Introduction The primary importance of meat in human nutrition is related to its high quality proteins that provide essential amino acids upon digestion. However, besides the high protein contents, meat also provides valuable amounts of many essential micronutrients such as some unsaturated fatty acids, vitamins and minerals (Tornberg, 2005). Meat as a food has a complex physical structure and chemical composition that is very susceptible to oxidation (Wood et al., 2008). The oxidative stability of meats depends upon the balance and the interaction between endogenous anti- and pro-oxidant substances and the composition of substrates prone to oxidation including polyunsaturated fatty acids (PUFA), cholesterol, proteins and pigments ( [Bertelsen et al., 2000] and [Decker and Xu, 1998] ). Endogenous antioxidant systems are composed of non-enzymatic hydrophilic and lipophilic compounds such as vitamin E, vitamin C, carotenoids, ubiquinols, polyphenols, cellular thiols, and enzymes like superoxide dismutase, catalase and glutathione peroxidase. Together, enzymatic and non-enzymatic antioxidant systems operate to counteract the action of pro-oxidants in muscle tissues ( [Chan and Decker, 1994] and [Decker et al., 2000] ) both in living animals and also after slaughter. The composition of endogenous antioxidants and pro-oxidant compounds can differ among meat of different species, among animals of a single species ( [Descalzo and Sancho, 2008] and [Pradhan et al., 2000] ) and muscle type. Also the diet of the animal by means of pasture or grain plays a significant role in modifying the concentration of antioxidants, pro-oxidants and fatty acids in the meat. ( [Hernandez et al., 2002] and[Hernandez et al., 2004] ). Depending on the consumers' preferences meat undergoes a wide range of cooking procedures and thermal treatments. Compositional and structural changes during cooking could significantly affect the TAC of meats ( [Palka and Daun, 1999] and [Tornberg, 2005] ).

Therefore, a reliable estimation of the TAC value of meat can be useful to describe the capacity of muscle to resist oxidation processes and to determine modifications in TAC values during thermal processing. Due to its complex structure, various extraction methodologies involving different solvents (water, aqueous buffers, alcohols, chloroform, etc.) have been applied to assess the TAC of meats ( [Descalzo et al., 2007]and [Sacchetti et al., 2008] ). Unfortunately, extraction-based methodologies can only measure the antioxidant capacities of the soluble and extractable hydrophilic or/and lipophilic fractions of meat. On the other hand, independent of the cooking process, meat contains a significant insoluble fraction. Moreover, various factors like the type and the pH of the extraction solvents, molecular interactions, aggregation phenomena, denaturation and oxidation can affect the extraction capacities and lead to an underestimation in TAC values of meat. A robust extraction-independent procedure to measure the TAC of solid foods, based on the interaction occurring at the interface between the solid matrix and a liquid colored radical probe has been developed (Serpen, Capuano, Fogliano, & Gkmen, 2007). The method, so called QUENCHER has been applied to various food matrices including cereals, bakery products and nuts and seeds ( [Aar et al., 2009] , [Gkmen et al., 2009] and [Serpen et al., 2008] ). The QUENCHER procedure in principle can be used for all food matrices and adapted to various assays commonly used to measure antioxidant activity (Amigo-Benavent, del Castillo, & Fogliano, 2010). In this work, the QUENCHER procedure was used to measure the TAC of four types of meat (beef, chicken, pork and fish) and to evaluate changes in TAC by thermal treatment. 2. Materials and methods 2.1. Chemicals All chemicals and solvents used were analytical grade. Water, potassium persulfate (di-potassium peroxdisulfate), acetic acid (glacial), sodium acetate and ferric chloride were purchased from Merck (Darmstadt, Germany). Cellulose (powder from spruce) and 2,2-azinobis(3-ethylbenzothiazoline-6sulfonic acid) (ABTS) were purchased from Fluka Chemie AG (Buchs, Switzerland) and ethanol from Carlo Erba (Italy). 2,4,6-tris-2,4,6-tripyridyl-2-triazine (TPTZ), 1,1diphenyl-2-picryl-hydrazyl (DPPH) and 6hydroxy-2,5,7,8-tetramethylchroman-2carboxylic acid (Trolox) were purchased from Sigma-Aldrich (Steinheim, Germany). 2.2. Thermal treatments of meat samples Fresh samples from chicken breast, pork tenderloin, beef fillet and sea bream (Sparus aurata) fillets were purchased from a local market. Proximate composition of the meats determined by AOAC (1996) official procedure is reported in Table 1. Minced meat samples were shaped into cylinders, having a diameter of 5 cm and thickness of 1 cm, by spreading them into aluminum caps. Using this procedure, any variation in the rate of heat transfer during cooking, which would further affect the rate of antioxidant/pro-oxidant degradation/formation, was limited.

Table 1. Proximate composition of the raw meat samples. Meat Chicken (breast) Pork (tenderloin) Beef (tenderloin) Water Protein Lipid Ash

73.3 0.47 23.1 0.37 2.4 0.21 1.2 0.09 73.2 0.21 22.1 0.39 3.6 0.33 1.1 0.11 73.1 0.28 20.0 0.33 5.9 0.52 1.0 0.12

Fish (Sea bream, fillet) 67.4 1.12 19.7 0.25 10.6 0.51 1.4 0.06 Meat samples were put into a preheated and air-ventilated oven (REX, Italy) at 180 C and cooked for 5, 10, 15 and 20 min. After cooking, samples were immediately cooled in an ice bath and stored at 20 C before freeze drying. Freeze dried meat samples were ground to powder form using a mortar and were consecutively passed through a sieve (Endecotts Test Sieve, London, U.K.) having 50 mesh size (297 m). All raw and cooked meat powders were stored at 20 C prior to analysis and all analyses were performed within 3 days. 2.3. Preparation of meat samples for QUENCHER procedure Freeze dried meat samples prepared as described above could be directly used for the QUENCHER procedure. Due to the high antioxidant activity (the values of absorbance were below the linear response range of the radical discoloration/color formation) a preliminary dilution was necessary. Dilution was performed by mixing the freeze dried meat samples with cellulose at a ratio between 1:1 to 1:10 (w:w) in a tube depending on the TAC values, following by rigorously shaking or squeezing in a mortar to achieve better homogeneity. Preliminary tests showed that a 1:5 (w:w) solid-state dilution with cellulose was suitable for both ABTS and DPPH assays, but not for the FRAP assay where no dilution was necessary. After dilution with cellulose 10 mg per sample was used, an amount ensuring good reproducibility for high antioxidant materials. Cellulose was found to be inert towards the various colored probes (Serpen et al., 2007): mixing cellulose powder (10.0 mg) with each radical probe solution no discoloration (i.e. decrease in absorbance) was observed over the 180 min of the assay. 2.4. Preparation of ABTS +, DPPH and FRAP radical solutions 2.4.1. ABTS
+

solution

ABTS solution was prepared by adding 5 mL of deionized water to 38.41 mg of ABTS. Potassium persulfate solution was prepared by mixing 5 mL of deionized water with 6.615 mg potassium persulfate. A total of 10 mL of stock solution of ABTS + was prepared by reacting 5 mL of each solution described above which resulted in final concentrations of 7 mmol/L ABTS and 2.45 mmol/L potassium persulfate. The stock solution of ABTS + was allowed to stand in the dark at room temperature for 12

16 h before use (Re et al., 1999). The working solution of ABTS + having absorbance of 0.750.80 at 734 nm was prepared daily by diluting the 10 mL of stock ABTS + solution with approximately 800 mL of a water/ethanol (50:50, v/v) mixture. 2.4.2. DPPH solution Stock solution of DPPH was prepared daily by dissolving 40 mg of DPPH in 100 mL of ethanol. Then the ethanolic solution of DPPH was further diluted with 100 mL of deionized water to obtain a DPPH stock solution in ethanol/water mixture (50:50, v/v). The working solution of DPPH having an absorbance value of 0.750.80 at 525 nm was prepared by diluting 200 mL of stock DPPH solution with approximately 800 mL of water/ethanol (50:50, v/v) mixture (Brand-Williams, Cuvelier, & Berset, 1995). 2.4.3. FRAP solution FRAP solution was prepared by diluting an aqueous solution of 10 mM TPTZ and 20 mM ferric chloride in 300 mM sodium acetate buffer (pH 3.6) at a ratio of 1:1:10 (v:v:v) as described by Benzie and Strain (1996). 2.5. Measurement of Trolox Equivalent TAC of meats by direct QUENCHER method Ten ( 1.0) mg of powdered meat sample (diluted at a ratio of 1:5 (w:w) with cellulose for the ABTS and DPPH assays) was weighed into a centrifuge tube. The reaction was started by adding 10 mL of ABTS +, DPPH or FRAP working solution. The tube was shaken rigorously for 1 min and placed on an orbital shaker in the dark. The mixture was shaken at 300400 rpm at room temperature on the orbital shaker until centrifugation to facilitate the surface reaction between the solid meat particles and the ABTS +, DPPH or FRAP solution. 15, 30, 60, 120 and 180 min after introduction of the radical solution to the solid meat samples, centrifugation was performed at 9200 g for 2 min. The clear supernatant (2 mL) was transferred into a cuvette and the absorbance measured at 734 nm (for ABTS assay), 525 nm (for DPPH assay) or 593 nm (for FRAP assay) at room temperature. Measurements at five different times (15, 30, 60, 120, 180 min) were needed to estimate the time required to reach the plateau (i.e. the end point of the reaction). The inhibition percentage of the ABTS + or DPPH radicals was calculated using the following equation (Eq. (1)):(1)Inhibitionsample(%)=(AbsblankAbssample)/Absblank100where, Absblank is the initial absorbance of the ABTS + or DPPH radical solution without sample and Abssample is the absorbance of the ABTS + or DPPH radical solution having sample after t min (15, 30, 60, 120, 180 min) of incubation. Trolox was used as a standard reference to convert the inhibition capability of each sample to the trolox equivalent antioxidant capacity (TEAC). First, standard trolox solutions in methanol were prepared at a concentration range between 0 and 600 g/mL. A 0.1 mL of each trolox solution was added to 9.9 ml of ABTS + or DPPH radical solution (024 M trolox in radical solution). After 30 min of incubation at room temperature, (enough time to reach a stable value) 2 mL of the radical solution was transferred into a cuvette and the absorbance measured at 734 nm (for ABTS assay) or 525 nm (for DPPH assay).

Standard calibration curves were constructed by plotting percentage inhibition (Eq. (2)) against concentration of trolox at 734 nm and 525 nm for the ABTS and DPPH assays, respectively (Fig. 1a and b).(2)Inhibitiontrolox(%)=(AbsblankAbstrolox)/Absblank100where, Absblank is the absorbance of the ABTS + or DPPH radical solution without trolox (only methanol) and Abstrolox is the absorbance of the ABTS + or DPPH radical solution with trolox after 30 min of incubation.

Full-size image (16K) Fig. 1. Calibration curve for (a) ABTS, (b) DPPH and (c) FRAP assay probes as a function of standard trolox concentration. The ratio between % inhibition of the sample and the slope of the trolox calibration curve was defined as the TEAC, which was used to indicate the scavenging free radical capability of the samples on a dry basis by the following equation (Eq. (3)).(3) where, s represents the slope of trolox calibration curve for ABTS (sABTS = 4.0895) and for DPPH (sDPPH = 3.0069) and m is the sample amount in mg dry basis, 10 is the conversion factor to obtain TEAC values in mmol Trolox Eq./kg of meat (d.w). Since color formation is monitored in the FRAP assay, no inhibition percentage was calculated as in the DPPH or ABTS assays. A calibration curve was constructed at room temperature by plotting concentration of trolox (020 M in radical solution) against the absorbance at 593 nm (Fig. 1c). TEAP values of the samples were calculated by (Eq. (4))(4) where, Abs593nm represents the absorbance of the FRAP solution with the sample at 593 nm after t min (15, 30 60, 120, 180 min) of incubation. n and s represent the intercept (0.04) and the slope (0.0444) of the trolox calibration curve of the FRAP assay, respectively and m indicates the sample amount in mg (dry basis), 10 is the conversion factor to obtain TEAC values in mmol Trolox Eq./kg of meat (d.w). 2.6. Statistical analysis

The analytical data are reported as mean standard deviation of triplicate independent measurements and were subjected to ANOVA, the significance of mean differences was determined by Duncans' posthoc test and t test using SPSS version 14.0 (2005). 3. Results and discussion Most natural antioxidant or neo-formed antioxidants upon processing are multifunctional, and in complex heterogeneous foods such as meat and meat products, their activity cannot be evaluated by a single method (Perez-Jimnez & Saura-Calixto, 2005). Two or more radical scavenging capacity assays are required to investigate heterogeneous samples since each assay involves different chemical mechanism(s) and may reflect different aspect(s) of their antioxidant properties. Here three common radical probes were used, namely ABTS, DPPH and Fe+ 3 (FRAP) to assess in-vitro antioxidant activity/power of meat in the QUENCHER procedure. Scavenging of DPPH radical allows evaluation of the hydrogen-donating potency of antioxidative compounds (Brand-Williams et al., 1995) while the ABTS radical determines the single electron-transfer capabilities of these (Re et al., 1999). Fe+ 3 probe in FRAP assay reflects the reductive antioxidant power (Benzie & Strain, 1996). To determine the appropriate end point for the evaluation of the TAC of meat, the reaction with the different probes was followed for different times. In Fig. 2 representative time-antioxidant capacity curves obtained with the three radical solutions (ABTS, DPPH, FRAP) for the pork samples cooked at 180 C for 15 min are reported. Similar time-antioxidant capacity curves were obtained for all raw or cooked meat samples studied. As shown in Fig. 2, the reaction between the probes and the meat samples is completed within 60 min for the ABTS and FRAP assays and within 120 min for the DPPH assay. Therefore, 60 min for the ABTS and FRAP assays and 120 min for the DPPH assay were considered satisfactory end-points to determine the TAC of meat by the direct QUENCHER procedure.

Full-size image (18K) Fig. 2. Representative time antioxidant capacity/power curve of pork cooked for 15 min at 180 C using ABTS, DPPH and Fe+ 3 (FRAP) as antioxidant assay probes.

TAC of the samples determined by the ABTS + and DPPH radical probes are presented in Fig. 3 and expressed in mmoles of Trolox per kg of dry weight. The total ABTS + scavenging capacity of the raw meat samples was in the range 25.9 1.0 mmol Trolox Eq./kg d.w. to 51.7 1.2 mmol Trolox Eq./kg (d.w). Comparing the ABTS + results of the raw meat samples, the TAC of chicken was significantly higher than that of pork, beef and fish (p < 0.05), while no significant differences were observed when the DPPH probe was used. Considering per fresh weight values (the average content of water in meat is 70%), the TAC for raw meats is around 10 mmol Trolox per kg.

Full-size image (14K) Fig. 3. TAC of raw meats determined using ABTS and DPPH as antioxidant probes. Different components are responsible of this TAC. It has been demonstrated that proteins and peptides have an important antioxidant action in meats due to their ability to scavenge free radicals and chelate prooxidative metals ( [Diaz and Decker, 2004] , [Elias et al., 2008] and [Elias et al., 2007] ). Chan and Decker (1994) reported that chicken meat is rich in histidine-containing dipeptides such as carnosine and anserine, which have high antioxidant activities. The presence of these antioxidant peptides in chicken could account for its higher ABTS + scavenging activity. Sacchetti et al. (2008) investigated the ABTS + scavenging antioxidant capacities of hydrophilic and lipophilic extracts of chicken meat by classical extraction dependent procedures. They reported the average ABTS +scavenging antioxidant capacities of chicken meat (breast) as 10.3 mmol Trolox Eq./kg (d.w.) and 5.3 mmol Trolox Eq./kg (d.w.) for the hydrophilic and lipophilic extracts, respectively. This led to a total of 15.6 mmol Trolox Eq./kg (d.w.) which is almost 70% lower than the value obtained in present study (51.7 mmol Trolox Eq./kg d.w.). This result, which was previously found comparing QUENCHER with conventional extraction based procedures on other matrixes, is likely due to the fact that the QUENCHER procedure includes the contribution of compounds which are not solubilized with the traditional extraction procedure. The total DPPH scavenging capacity of the raw meat samples in this were between 19.1 1.8 mmol Trolox Eq./kg (d.w.) and 31 0.9 mmol Trolox Eq./kg (d.w.). The DPPH scavenging capacity of chicken, pork and beef was similar (p > 0.05) while that of fish was significantly lower (p < 0.05).

Interestingly, the TAC value measured with the two total radical scavenging antioxidant capacity assays was significantly different for chicken and fish meats but not for raw pork and beef (p < 0.05) which could be ascribed to the different affinities of the radicals to scavenge the various antioxidant groups present in different samples. Dean, Yamamoto, and Niki (1991) demonstrated that hydrophobic radicals have less ability to attack macromolecules such as proteins in solution. Therefore, it could be one of the reasons that DPPH , a relatively hydrophobic radical, would have less interaction with polar macromolecular antioxidant compounds than more hydrophilic probes like ABTS. In addition, DPPH is likely more selective than ABTS +in the reaction with H-donors (Roginsky & Lissi, 2005) and this could explain the low TAC values obtained by the DPPH compared with the ABTS assay. On the other hand, the TAC values of fish samples were the lowest in both assays. Beside a lower content of blood in the fish sample the loss of endogenous antioxidant compounds that may inhibit unsaturated lipid oxidation could be one of the reasons for the low total radical scavenging antioxidant capacity of fish compared with beef, pork and chicken. FRAP of raw meat samples is presented in Fig. 4 expressed as mmoles of Trolox Eq. per kg of meat (d.w.). Among the raw meat samples, the highest FRAP value was in beef (4.9 0.2 mmol Trolox Eq./kg d.w.) whereas the lowest level of 3.0 0.1 mmol Trolox Eq./kg (d.w.) was that of the fish sample. The differences in FRAP values between the raw beef and chicken were not statistically significant (p > 0.05). Similarly, there were no significant differences between the FRAP values of raw fish and pork. In general, reductive antioxidant power of the raw samples was lower as compared to the ABTS + and DPPH scavenging capacities. The FRAP method is based on the reduction of the Fe+ 3TPTZ complex to the ferrous form at low pH. The low values of FRAP could be due to the poor ability of meat antioxidants to reduce ferric ion to its ferrous form.

Full-size image (12K) Fig. 4. FRAP of raw meats. The influence of thermal treatment on the TAC of the meat samples was measured using the ABTS + and DPPH radical probes; Fig. 4 and Fig. 5, respectively. The TAC of most meat samples increased on heating at the beginning of the cooking period peaking after 510 min using both assays. The maximum

TAC values were clearly visible in all the meat samples but not the fish. On increasing the heating time, TAC levels started to decrease, although at the longest heating time a further increase of TAC is seen, particularly using the ABTS probe.

Full-size image (7K) Fig. 5. Effects of time at 180 C on TAC values of meats measured using ABTS as the antioxidant probe. The behavior on heating of proteins, which are the main component of meat, helps explain this result. Thermal treatments can alter the secondary and tertiary structure of proteins, resulting in modifications of their physical properties ( [Sante-Lhoutellier et al., 2007] and [Tironi et al., 2002] ). Unfolding of the proteins (i.e. denaturation) can increase their ability to scavenge radicals by increasing the solvent exposure of antioxidant amino acids that would normally be located in the core of the native protein structure (Elias et al., 2007). Therefore, it is possible that mild heat treatments could increase the antioxidant activity of some proteins due to changes in their quaternary and tertiary structures. On the other hand, it is well known that more severe thermal treatments can cause thermoxidation of different components of muscle foods resulting in consumption of antioxidant substances and a decrease of TAC. The trend of TAC in the first minutes of heating (Fig. 5 and Fig. 6) can be explained because in the initial period, the consumption of antioxidant substances is less marked compared to the exposure of hidden antioxidant amino acids due to protein denaturation. Moreover some antioxidant compounds (for example GSH) can be released as a consequence of cell membrane destruction, thus being more reactive towards the radical probes.

Full-size image (6K) Fig. 6. Effects of time at 180 C on TAC values of meats measured using DPPH as the antioxidant probe. Prolongation of the heating process reduced the TAC of meats in both radical scavenging assays (Fig. 5 andFig. 6). This can be due to the accumulation of oxidized proteins and the loss in functionality of active meat peptides. Moreover, degradation of endogenous antioxidative factors such as vitamin E, vitamin C, carotenoids, ubiquinols, polyphenols, and cellular thiols could be promoted by heating. In this stage, the prooxidantantioxidant balance was dominated by the prooxidant properties. In the last stage of the heating process, the TAC of some meat samples showed a marked increase in both assays. This could be due to reactions between reducing sugars and free amino acids or free amino groups in proteins, the Maillard reaction. There have been several reports on the antioxidative properties of MRPs ( [Bailey and Um, 1992] , [Borrelli et al., 2002] and [Serpen et al., 2007] ). Since the Maillard reaction is favored at low water activity a significant increase in the MRPs concentration could be achieved only at the end of the cooking time. The highest increases in TAC levels in the final cooking period were observed in chicken, beef and fish samples by the ABTS assay while the net increase was only observed in pork samples by the DPPH assay. Low reactivity of the hydrophobic DPPH radicals against hydrophilic MRPs could explain the differences between the two assays. The effects of heating on the FRAP values of meats are reported in Fig. 7. The FRAP assay gave different results showing a sharp increase of activity for all samples, particularly beef. All the samples produced MRPs upon thermal treatment, but with different properties depending on their composition of proteins and carbonyl groups.The rapid increase of FRAP values in beef in the last heating stage might be due to the accumulation of MRPs with higher reducing properties than those of fish, chicken and pork.

Full-size image (6K) Fig. 7. Effects of time at 180 C on FRAP values of the meats. In order to detect a possible release of free iron from myoglobin to the matrix that would account for the increase in FRAP value during heating, the FRAP measurements were performed without adding ferric iron to the reaction mixture, this resulted in no color development indicating that there was no detectable free ferrous iron formed from the reduction of iron released from the heme group. Also there were no detectable agents in the heated meat samples that could react directly with TPTZ to form the blue chromogen. These results agree with previous reports regarding the relatively high stability of the heme group to thermal treatments and the binding of released iron to proteins ( [Berisha et al., 2003] and [Kristensen and Andersen, 1997] ). As a result it is concluded that the increase of FRAP values with heating is mainly related to the development of MRPs and does not reflect the loss of endogenous antioxidative factors during the cooking of meat. 4. Conclusions Traditional extraction-based methodologies for TAC evaluation are only partially useful as they take into account only the soluble and extractable hydrophilic or/and lipophilic fractions of meat. The QUENCHER procedure which measures the antioxidant capacity of both soluble and insoluble parts together, gives a reliable measure of the TAC value for raw and processed meat. The QUENCHER approach revealed that the effect of cooking on TAC is dependent on meat type and severity of heating. The balance between phenomena occurring during cooking such as i) denaturation and exposure of reactive sites of proteins; ii) thermoxidation and degradation of endogenous antioxidants; iii) formation of antioxidant MRPs might be responsible for the overall trend of TAC observed for different cooked meats. Using the direct QUENCHER procedure it will be possible to build a database and compare results for the TAC of meat samples, particularly after very different cooking procedures. References Aar et al., 2009 O.C. Aar, V. Gkmen, N. Pellegrini and V. Fogliano, Direct evaluation of the total antioxidant capacity of raw and roasted pulses, nuts and seeds. European Food Research and Technology, 229 (2009), pp. 961969.

Amigo-Benavent et al., 2010 M. Amigo-Benavent, M.D. del Castillo and V. Fogliano, Are the major antioxidants derived from soy protein and fructo-oligosaccharides model systems colored aqueous soluble or insoluble compounds. European Food Research and Technology, 231 (2010), pp. 545 553. AOAC, 1996 AOAC, K. Hilrich, Editor, Official methods of analysis, Association of Analytical Communities, Arlington, VA (1996). Bailey and Um, 1992 M.E. Bailey and K.W. Um, Maillard reaction products and lipid oxidation. ACS Symposium Series, 500 (1992), pp. 122139. Benzie and Strain, 1996 I.F.F. Benzie and J.J. Strain, The ferric reducing ability of plasma (FRAP) as a measure of antioxidant power: The FRAP assay. Analytical Biochemistry, 1996 239 (1996), pp. 70 76. Berisha et al., 2003 A. Berisha, E. Yasushi and K. Fujimoto, The effect of heat-induced structural changes on the prooxidant activity of myoglobin. Italian Journal of Food Science, 15 (2003), pp. 8594. Bertelsen et al., 2000 G. Bertelsen, M. Jakobsen, D. Juncher, J. Moller, M. Kroger-Ohlsen and C. Weber, et al.Oxidation, shelf-life and stability of meat and meat products, Proceedings of the 46th international congress of meat science and technology (2000), pp. 516524 Buenos Aires, Argentina. Borrelli et al., 2002 R.C. Borrelli, A. Visconti, C. Mennella, M. Anese and V. Fogliano, Chemical characterization and antioxidant properties of coffee melanoidins. Journal of Agricultural and Food Chemistry, 50 (2002), pp. 65276533. Brand-Williams et al., 1995 W. Brand-Williams, M.E. Cuvelier and C. Berset, Use of a free radical method to evaluate antioxidant activity. Lebensmittel-Wissenschaft und Technologie, 28 (1995), pp. 2530. Chan and Decker, 1994 K.M. Chan and E.A. Decker, Endogenous skeletal muscle antioxidants. Critical Reviews in Food Science and Nutrition, 34 (1994), pp. 403426. Dean et al., 1991 R.T. Dean, Y. Yamamoto and E. Niki, Free-radical damage to proteins The influence of the relative localizaton of radical generation, antoxidants, and target proteins. Free Radical Biology and Medicine, 11 (1991), pp. 161168. Decker et al., 2000 E.A. Decker, S.A. Livisay and S. Zhou, Mechanisms of endogenous skeletal muscle antioxidants: Chemical and physical aspects, E. Decker, C. Faustman, C.J. Lopez-Bote, Editors ,Antioxidants in muscle foods, John Wiley & Sons Inc., New York (2000), pp. 3947. Decker and Xu, 1998 E.A. Decker and Z. Xu, Minimizing rancidity in muscle foods. Food Technology, 52 (1998), pp. 5459.

Descalzo et al., 2007 A.M. Descalzo, L. Rossetti, G. Grigioni, M. Irurueta, A.M. Sancho and J. Carrete, et al.Antioxidant status and odour profile in fresh beef from pasture or grain-fed cattle. Meat Science, 75 (2007), pp. 299307. Descalzo and Sancho, 2008 A.M. Descalzo and A.M. Sancho, A review of natural antioxidants and their effects on oxidative status, odor and quality of fresh beef produced in Argentina. Meat Science, 79 (2008), pp. 423436. Diaz and Decker, 2004 M. Diaz and E.A. Decker, Antioxidant mechanisms of caseinophosphopeptides and casein hydrolysates and their application in ground beef. Journal of Agricultural and Food Chemistry, 52 (2004), pp. 82088213. | View Record in Scopus | | Cited By in Scopus (42) Elias et al., 2008 R.J. Elias, S.S. Kellerby and E.A. Decker, Antioxidant activity of proteins and peptides.Critical Reviews in Food Science and Nutrition, 48 (2008), pp. 113. Elias et al., 2007 R.J. Elias, D.J. McClements and E.A. Decker, Impact of thermal processing on the antioxidant mechanisms of continuous phase beta-lactoglobulin in oil-in-water emulsions. Food Chemistry, 104 (2007), pp. 14021409. Article | in Scopus (13) PDF (523 K) | | View Record in Scopus | | Cited By

Gkmen et al., 2009 V. Gkmen, A. Serpen and V. Fogliano, Direct measurement of the total antioxidant capacity of foods: The QUENCHER approach. Trends in Food Science and Technology, 20 (2009), pp. 278288. Article | PDF (343 K) | | View Record in Scopus | | Cited By in Scopus (18)

Hernandez et al., 2002 P. Hernandez, D.K. Park and K.S. Rhee, Chloride salt type/ionic strength, muscle site and refrigeration effects on antioxidant enzymes and lipid oxidation in pork. Meat Science, 61 (2002), pp. 405410. Article | Scopus (13) PDF (129 K) | | View Record in Scopus | | Cited By in

Hernandez et al., 2004 P. Hernandez, L. Zomeno, B. Arino and A. Blasco, Antioxidant, lipolytic and proteolytic enzyme activities in pork meat from different genotypes. Meat Science, 66 (2004), pp. 525 529. Article | PDF (162 K) | | View Record in Scopus | | Cited By in Scopus (17)

Kristensen and Andersen, 1997 L. Kristensen and H.J. Andersen, Effect of heat denaturation on the prooxidative activity of metmyoglobin in linoleic acid emulsions. Journal of Agricultural and Food Chemistry, 45 (1997), pp. 713. | View Record in Scopus | | Cited By in Scopus (21) Palka and Daun, 1999 K. Palka and H. Daun, Changes in texture, cooking losses, and myofibrillar structure of bovine M. semitendinosus during heating. Meat Science, 51 (1999), pp. 237 243. Article | PDF (1212 K) | | View Record in Scopus | | Cited By in Scopus (71)

Perez-Jimnez and Saura-Calixto, 2005 J. Perez-Jimnez and F. Saura-Calixto, Literature data may underestimate the actual antioxidant capacity of cereals. Journal of Agricultural and Food Chemistry, 53 (2005), pp. 50365040. | View Record in Scopus | | Cited By in Scopus (65)

Pradhan et al., 2000 A.A. Pradhan, K.S. Rhee and P. Hernandez, Stability of catalase and its potential role in lipid oxidation in meat. Meat Science, 54 (2000), pp. 385390. Article | Record in Scopus | | Cited By in Scopus (25) PDF (122 K) | | View

Re et al., 1999 R. Re, N. Pellegrini, A. Proteggente, A. Pannala, M. Yang and C. Rice-Evans, Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radical Biology and Medicine, 26 (1999), pp. 12311237. Article | Scopus (2404) PDF (158 K) | | View Record in Scopus | | Cited By in

Roginsky and Lissi, 2005 V. Roginsky and E.A. Lissi, Review of methods to determine chain-breaking antioxidant activity in food. Food Chemistry, 92 (2005), pp. 235254. Article | Record in Scopus | | Cited By in Scopus (170) PDF (485 K) | | View

Sacchetti et al., 2008 G. Sacchetti, C. Di Mattia, P. Pittia and G. Martino, Application of a radical scavenging activity test to measure the total antioxidant activity of poultry meat. Meat Science, 80 (2008), pp. 10811085.Article | Scopus (5) PDF (365 K) | | View Record in Scopus | | Cited By in

Sante-Lhoutellier et al., 2007 V. Sante-Lhoutellier, L. Aubry and P. Gatellier, Effect of oxidation on in vitro digestibility of skeletal muscle myofibrillar proteins. Journal of Agricultural and Food Chemistry, 55 (2007), pp. 53435348. | View Record in Scopus | | Cited By in Scopus (18) Serpen et al., 2007 A. Serpen, E. Capuano, V. Fogliano and V. Gkmen, A new procedure to measure the antioxidant activity of insoluble food components. Journal of Agricultural and Food Chemistry, 55 (2007), pp. 76767681. | View Record in Scopus | | Cited By in Scopus (43) Serpen et al., 2008 A. Serpen, V. Gkmen, N. Pellegrini and V. Fogliano, Direct measurement of the total antioxidant capacity of cereal products. Journal of Cereal Science, 48 (2008), pp. 816820. Article |

Penelitian ini menyelidiki kapasitas antioksidan total (TAC) daging (sapi, ayam, babi dan ikan) dan perubahan perlakuan termal. Prosedur memuaskan, dilakukan langsung pada bahan solid tanpa ekstraksi, dipilih dan terbukti sangat cocok untuk sampel daging. Para ABTS + pemulungan kapasitas daging mentah berkisar antara 25,9 1,0 dan 51,7 1,2 mmol Trolox EQ / kg. Ayam mentah memiliki TAC tertinggi diikuti oleh daging babi, daging sapi dan ikan. Ketika dipanaskan sampai 180 C, peningkatan daging maksimum TAC jelas pada 5 menit, diikuti oleh tetes mendadak sampai 15 menit, sedangkan pemanasan tahap akhir ditandai oleh peningkatan sedikit. Modifikasi TAC saat memasak dapat dijelaskan dengan mempertimbangkan faktor-faktor seperti denaturasi dan eksposur situs reaktif protein, degradasi antioksidan endogen dan pembentukan produk reaksi Maillard memiliki sifat antioksidan.