Effect of rigor temperature, ageing and display time on the meat quality
and lipid oxidative stability of hot boned beef Semimembranosus muscle
Tanyaradzwa E. Mungure
a,⁎
, Alaa El-Din A. Bekhit
a, E. John Birch
a, Ian Stewart
ba
Department of Food Science, University of Otago, PO Box 56, Dunedin, New Zealand
bDepartment of Chemistry, University of Otago, PO Box 56, Dunedin, New Zealand
a b s t r a c t
a r t i c l e i n f o
Article history: Received 10 August 2015
Received in revised form 16 December 2015 Accepted 19 December 2015
Available online 23 December 2015
The effects of rigor temperature (5, 15, 20 and 25 °C), ageing (3, 7, 14, and 21 days) and display time on meat quality and lipid oxidative stability of hot boned beef M. Semimembranosus (SM) muscle were investigated. Ulti-mate pH (pHu) was rapidly attained at higher rigor temperatures. Electrical conductivity increased with rigor
temperature (pb 0.001). Tenderness, purge and cooking losses were not affected by rigor temperature; however purge loss and tenderness increased with ageing (pb 0.01). Lightness (L*) and redness (a*) of the SM increased as rigor temperature increased (pb 0.01). Lipid oxidation was assessed using1H NMR where changes in aliphatic to
olefinic (Rao) and diallylmethylene (Rad) proton ratios can be rapidly monitored. Rad, Rao, PUFA and TBARS were
not affected by rigor temperature, however ageing and display increased lipid oxidation (pb 0.05). This study shows that rigor temperature manipulation of hot boned beef SM muscle does not have adverse effects on lipid oxidation.
© 2016 Elsevier Ltd. All rights reserved.
Keywords: Ageing Meat Oxidation rigor temperature 1. Introduction
Several studies reported significant effects of rigor temperature on meat quality attributes (Bekhit, Farouk, Cassidy, & Gilbert, 2007; Farouk & Swan, 1998; Geesink, Bekhit, & Bickerstaffe, 2000). As rigor temperature increases, the rate of glycolysis in muscle also increases (Hertzman, Olsson, & Tornberg, 1993). This leads to a rapid decline in pH while the muscle has relatively high temperature providing protein denaturing conditions. This process results in pale, soft, and exudative (PSE)-like meat characteristics (Thompson et al., 2005; Warner, Dunshea, Gutzke, Lau & Kearney, 2014).
High rigor temperatures (N25 °C) have been reported to have an ad-verse impact on the most important quality attribute, meat tenderness, due to heat shortening (Hertzman et al., 1993; Rees, Trout, & Warner, 2002). While the pH andμ-calpain activity rapidly decline with increas-ing rigor temperature (Geesink et al., 2000), lightness (L*) of meat have been reported to increase with increased rigor temperatures (Farouk & Swan, 1998; Warner, Kerr, Kim & Geesink, 2014). Most of the reported studies have examined the effects of rigor temperature on longissimus et lumborum muscle as it is the most important commercial muscle and little attention was given to metabolically different muscles.
Furthermore, the effects of rigor temperature, ageing and display time on lipid oxidative stability of fresh meat are rarely reported. Lipid oxida-tion is a major mechanism of meat quality deterioraoxida-tion manifesting
changes inflavour and colour. Lipid oxidation can also cause formation of toxic compounds such as aldehydes and ketones; this leads to re-duced shelf life (Gray, Gomaa, & Buckley, 1996). The present study in-vestigated the effect of rigor temperature on the quality of hot boned beef semimembranosus (SM) muscle at various ageing and display times. Lipid oxidation was also assessed using thiobarbituric acid reac-tive substances (TBARS) method.1H NMR was used as a novel tech-nique to monitor the change in oxidation rate by analysis of the relative change in aliphatic to olefinic proton ratio (Rao) and the aliphat-ic to diallylmethylene proton ratio (Rad). Lipid oxidation was also analysed using changes in polyunsaturated fatty acids (PUFAs) as assessed by gas chromatographyflame ionisation detection (GC-FID). 2. Materials and methods
2.1. Meat
Topside (SM) muscles were obtained from 6 heifers (n = 6) raised on pasture with an average hot-carcass weight of 278.9 ± 28.4 kg. The animals were slaughtered in an export licensed meat-processing facility (Alliance Group, Pukeuri plant, Oamaru, New Zealand), electrically stimulated [Square mono wave, 80 V, 25 s0], hot boned at 45 min post-mortem and processed within 2 h. Muscles from right and left top sides of each animal were divided into 4 samples from each animal. Samples were randomised to 5, 15, 20 and 25 °C rigor temperature treatment groups. Samples measured for their pH decline every hour and then removed from the treatment incubator once the samples reached their ultimate pH thus attaining rigor. They were vacuum
⁎ Corresponding author.
E-mail address:[email protected](T.E. Mungure).
http://dx.doi.org/10.1016/j.meatsci.2015.12.015
0309-1740/© 2016 Elsevier Ltd. All rights reserved.
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Meat Science
packed and aged for 3, 7, 14, and 21 days at 4 °C. At the designated ageing time, the pH, purge loss and conductivity of samples were measured then subsampled for colour, tenderness and lipid oxida-tion. Samples for colour were processed as described below and sam-ples for tenderness measurements and lipid analysis were stored at −80 °C until analysis.
2.2. pH analysis
The pH of the meat samples was measured using a puncture pH elec-trode (InLab 427, Mettler-Toledo Process Analytical Inc., Wilmington, MA) attached to a pH metre Hanna HI 9025 (Hanna Instruments, Woon-socket, RI) every hour during the incubation time until pHuwas reached. The pH was also measured directly after the ageing period.
2.3. Electrical conductivity
The electrical conductivity (mS·cm−1) of the samples was mea-sured after the ageing period using a hand held electrical conductivity metre (LF-Star, Matthäus, Pӧttmes, Germany).
2.4. Purge loss
Sample weight was determined prior to and following the ageing process. Following ageing, the samples were blotted dry with paper towels before reweighing. Purge loss was calculated as a percentage using the formula below:
Purge lossð Þ ¼% ðweight before storage weight after storage timeÞ weight before storage time
X 100
2.5. Cooking loss
The samples were thawed at 2 °C overnight and cooked individually in plastic bags immersed in a water bath at 80 °C until the internal tem-perature reached 75 °C as measured by temtem-perature probes. The cooked meat was cooled on ice, blotted dry with paper towels and weighed. The cooking loss was expressed as a percentage and calculated by the for-mula below:
Cooking lossð Þ ¼% ðweight before cookingweight before cooking weight after cookingÞ
X 100
2.6. Shear force
Shear force was determined using a MIRINZ tenderometer as de-scribed byChrystall, Devine, Graafhuis, and Muir (1993). The cooked samples were sliced parallel to the musclefibre axis to produce 6 sub-samples with a 10 mm × 10 mm cross section. The subsub-samples were each sheared on a MIRINZ tenderometer with a wedged shaped tooth and the peak shear force was recorded. The values were converted to Newtons using the following formula;
Shear force Nð Þ ¼ 0:2035 shear force=kPA½ð Þ−2:2945 9:8
2.7. Colour analysis
Objective colour measurements were obtained as described byBekhit et al. (2007). After ageing, the beef SM muscle samples were placed in polystyrene trays and wrapped with oxygen permeable polyvinylchloride film (O2permeabilityN2000 mL·m−2·atm−1·24 h−1at 25 °C) (AEP FilmPac Ltd., Auckland, New Zealand). Samples were displayed under coolfluorescent light (1076 lux) at 4 °C. Colour measurements were
taken at 1, 3, 5 and 7 days of display time using a HunterLab colorimeter (model 45/0-L, Hunter Associates Laboratory Inc., Reston, VA). The color-imeter was calibrated with a black and a white standard tile C2-36852. CIE L*, a*, b* values were taken using illuminant C and a 10° observer with an aperture size 2.5 cm and the reflectance values in the wavelength range of 400 to 700 nm. The Chroma (C = [a*2+b*2][1/2]) and hue angle (HA = tan−1b*/a*) were calculated. The browning index parameter 630 nm/580 nm ratio was determined.
2.8. Lipid oxidation analysis using thiobarbituric acid reactive substance (TBARS) Assay
TBARS assay was used to determine lipid oxidation as described by
Du, Nam, and Ahn (2001). TBARS were measured at each ageing time and after 7 days of display time. Meat samples from the surface (3 g) were placed in 50 mL tubes and homogenised with 9 mL of deionised distilled water (DDH2O) using a Polytron homogeniser at 14,000 rpm (PT-MR 2100, Kinematica AG, Switzerland) for 30 s. Meat homogenate (1 mL) was transferred to a 10 mL Falcon tube and 50μL butylated hy-droxytoluene (BHT, 7.2% w/v in ethanol) and 2 mL thiobarbituric acid (TBA)/trichloroacetic acid (TCA) (20 mM TBA in 15% TCA, (w/v)) solu-tion were then added and the mixture vortexed for 30 s. The samples were incubated in a water bath at 90 °C for 30 min for colour develop-ment, and then cooled in cold slurry of ice water for 10 min. The sam-ples were vortexed (30 s) and centrifuged at 2520 × g (Beckman Coulter, Inc. California, USA) for 15 min at 5 °C. The absorbance of the resulting supernatant was read at 531 nm against a blank prepared with 1 mL DDH2O and 2 mL TBA/TCA solution. The amount of malondialdehyde (mg·kg−1) in the sample was determined by using the following equation;
Mg MDA=kg meat ¼ A@531 E 72:063=100 x dilution factor Where E is the molar extinction coefficient of MDA¼ 156; 000M−1cm−1
Molar mass of malondialdehyde¼ 72:063gmol−1
2.9. Lipid extraction for1H NMR lipid oxidation analysis and GC-FID Lipid extraction was performed as described byFolch, Lees, and Sloane-Stanley (1957). Meat samples (100 g), from rigor temperatures 5 and 25 °C only due tofinancial constraints, aged for 7 and 14 days, and subsequent post display time were cut into small pieces and ex-tracted in an Omnimixer with 200 mL chloroform/methanol (2:1 v/v) in a 1.9 L Mason jar. After dispersion, the whole mixture was agitated for 20 min in an orbital shaker at room temperature. The homogenate was thenfiltered through a funnel with Celite 545 with a folded filter paper. The solvent was washed with a 0.9% NaCl solution. The samples were vortexed for 30 s and the mixture was centrifuged at 2000 rpm for 5 min to separate the two phases. After centrifugation and siphoning of the upper phase, the lower chloroform phase containing lipids was evaporated under vacuum in a rotary evaporator and under nitrogen gas.
2.9.1.1H NMR analysis
1H NMR spectra were acquired using a Bruker Avance 400 spectrom-eter operating at 400 MHz. Each lipid sample (30 mg) was mixed with 650μL of deuterated chloroform (purity 99.8%) and 6.5 μL 1, 4-dioxane as an internal reference in a 5 mm diameter tube. The acquisi-tion parameters were: spectral width 5000 Hz, relaxaacquisi-tion time 3 s. Spec-tra were recorded at room temperature with 65,536 data points where the number of scans was 128, acquisition time 1.6 s and pulse width 80° with a total acquisition time of 21 min 27 s. Exponential line broadening 0.30 Hz, automatic phase correction and baseline correction were ap-plied to each spectrum. The assignment of the signals integrated and used for lipid oxidation analysis was made as described in previous studies (Guillén & Ruiz, 2004) and is given inTable 1. The ratios for
Radand Raowere obtained as follows;
Rad¼Integrated area for diallylmethylene proton signalIntegrated area for aliphatic proton signal
Rao¼Integrated area for aliphatic proton signalIntegrated area for olefinic proton signal
2.9.2. Fatty acid methyl ester (FAME) preparation and GC-FID analysis A modified method ofVan Wijngaarden (1967)was used to prepare fatty acid methyl esters. Each lipid sample (15 mg) was dissolved in 10 mL of hexane, and 2 mL was transferred into a glass tube. A 2 mL ali-quot of 0.5 N methanolic KOH (prepared by adding 5.6 g of KOH into 200 mL methanol with 15 min stirring) was added into the glass tube and heated for 20 min at 80 °C. Diethyl ether (2 mL) and water (5 mL) were added to the glass tube and the two phases were separated. The diethyl ether layer (upper phase) was discarded. A few drops of HCI were added until the solution was acidic, as assessed by litmus paper. A 2 mL aliquot of diethyl ether was subsequently added into the glass tube. Two layers of separation were formed and the top diethyl ether layer was collected. Boron trifluoride (BF3; 1 mL, 14% in methanol) was added into the new glass tube and heated for 20 min at 80 °C. Saturated NaCl (5 mL) was added to the cooled solution and vortexed for 1 min. The two phases were separated and the FAME phase (upper phase) was col-lected into a vial for GC analysis (Van Wijngaarden, 1967). The FAMEs were separated using a BPX-70 capillary column (100 m × 0.22 mm i.d; 0.25μm film thickness). The gas chromatographic system consisted of a 6890 N GC equipped with an autosampler (HP7673) and ChemStation integration. The column oven was held at an initial tem-perature of 165 °C for 52 min, and then increased at a rate of 5 °C·min−1to afinal temperature of 210 °C for 59 min (total run time 120 min). Both the injector andflame ionisation detector ports were at 250 °C. Carrier gasflow (Helium) was maintained at 1.0 mL·min−1 (linear gas velocity was 0.2 m·s−1) throughout the temperature pro-gramme with an inlet split ratio of 30:1. Fatty acid peaks were identified by matching retention time with authentic standards. A composite stan-dard was made from commercially available methyl esters (NuCheck Prep, Elysian, Minnesota and Sigma, St. Louis, Missouri).
2.10. Statistical analysis
Data for pH, conductivity, purge loss; cooking loss, shear force, and lipid oxidation analysis by proton NMR was tabulated in Microsoft excel spreadsheets. All the samples run for lipid oxidation analysis using GC and NMR were carried out in duplicate. Analysis of variance (ANOVA) was carried out using Minitab (version 16.2.4). When the main effect or interactions were significant, mean separation was attained by least squares F-test procedures. Significant differences among mean values were determined at a 5% significance level. The pH decline results werefitted using Sigmaplot (Systat Software Inc. (SSI), San Jose, California, USA).
3. Results and discussion 3.1. pH
The rate of pH decline decreased with the decrease in the pre-rigor temperature showing that it was temperature-dependent. Both 20 °C and 25 °C treated SM muscles reached pHuafter 9 h of incubation whereas the samples incubated at 5 °C reached pHuat 24 h postmortem (Fig. 1). At 5 °C the glycolysis process is slower, reflecting the time re-quired for the glycolysis reaction to run to completion (Jeacocke, 1977). The rapid decline at 20 °C and 25 °C reflects an accelerated rate of glycolysis. These results were in agreement with rate of pH decline in other studies investigating the effect of rigor temperature on beef muscles (Farouk & Lovatt, 2000; Hertzman et al., 1993; Jeacocke, 1977). This trend has been reported in venison longissimus dorsi (LD) muscle (Bekhit et al., 2007) and lamb (Geesink et al., 2000).
The effects of post-mortem time and rigor temperature on the sam-ples pH are shown inTable 2. The 25 °C and 20 °C (pH 5.50 and 5.52, re-spectively) incubated beef SM muscle samples had lower pHuvalues (pb 0.05) compared to 15 °C and 5 °C treatments (5.59 and 5.69 respec-tively). This is in agreement with results on incubation of other beef muscles (Farouk & Swan, 1998; Marsh, 1954). The difference in pHu may again be due to higher glycolysis rates at high temperatures (Bendall, 1972; Jeacocke, 1977). Glycolysis by-products may also hinder enzymatic activity at lower rigor temperatures (Bendall, 1972).
Ageing time also affected pH (pb 0.01). At 3 days post-mortem, the pH values were lower than at rigor values for all treatments (pb 0.05). This trend was observed in venison LD muscle under similar treatments (Bekhit et al., 2007). The 3 and 7 day aged samples did not statistically differ. The pH increased at days 14 (5, 15 and 25 °C treatments) and 21 (all treatments) of ageing compared to 3 days post-mortem time (Table 2). The increase in pH could be attributed to the proteolysis of muscle proteins resulting in increased free amino acids and dipeptides like carnosine which are well known for their buffering abilities (Braggins, Frost, Agnew, & Podmore, 1999; Braggins et al., 2004).
Braggins et al. (1999)observed similar increases in pH with ageing time in mutton and concluded that the change was due to an increase in basic amino acids relative to the acidic free amino acids. The same trend was reported in beef (Farouk & Wieliczko, 2003) and chicken (Craig, Fletcher, & Papinaho, 1999).
3.2. Electrical conductivity
Conductivity describes the ease at which ionic species move in the cellular matrix. This ionic species movement is normally inhibited by
Table 1
The chemical shifts for functional groups integrated to monitor lipid oxidation in beef SM muscle using1H NMR sources.
Signal Functional group 1.45–1.20 ppm –(CH2)n-(acyl group)
2.84–2.70 ppm HC–CH2–CH(acyl group)
5.40–5.26 ppm –CHCH-(acyl group) Source:Guillén & Ruiz, 2006
Fig. 1. Effect of pre-rigor temperature (5, 15, 20 and 25 °C) incubation of hot boned beef SM muscle on the pH decline during post-mortem time.
an intact cellular membrane (Lebovka, Bazhal, & Vorobiev, 2002). Elec-trical conductivity at 3 days post-mortem increased with an increase in rigor temperatures (Table 2, pb 0.001). The relationship between rigor temperature and ageing time together was found to impact the conduc-tivity (pb 0.05). The most noticeable significant increase was found in the 5 °C treatment group between days 3 and 21(pb 0.001), with a 3.83 mS rise in conductivity (Table 2). This increase is likely due to the effect of endogenous processes in the muscle encouraging cellular damage during the ageing process that promoted increasedflow of elec-trolytes within the muscle due to an increase in cell membrane perme-ability (Byrne, Troy, & Buckley, 2000). The lack of difference over ageing time at higher rigor temperatures (20 °C and 25 °C) may indicate that a maximum permeability was achieved as a result of rigor temperature incubation.
3.3. Purge and cooking loss
Rigor temperature did not have an effect on the purge loss (pN 0.05) (Table 3). However, other researchers have found increased purge loss with an increase in rigor temperature.Farouk and Swan (1998)and
Fernandez and Tornberg (1994)reported that beef semitendinosus (ST) and pork LD muscles, respectively, exhibited significant increases in purge loss with increases in rigor temperature. However, other stud-ies on pork longissimus thoracis et lumborum muscle showed a decrease in purge loss with an increase in rigor temperature (Rees et al., 2002). The results of the present research are in agreement with those reported byBekhit et al. (2007). In venison LD muscle no differences in purge loss were observed within the rigor temperature range of 0–35 °C. It is im-portant to register that the effect of rigor temperature on purge loss is possibly affected by the animal species and muscle type (Bekhit et al., 2007).
Ageing time did have an effect on purge loss (pb 0.001). There was gradual increase in purge loss, but only samples stored for 3 days (1.93 ± 0.34%) were significantly different from those aged for 21 days (6.90 ± 0.34%) (Table 3). There were no significant differences between day 3, 7 and 14 days of ageing time for all samples. The purge loss in samples incubated at 25 °C and aged for 21 days was numerically higher than the other treatments but there were no significant differ-ences among any of the treatments. The increase in purge loss with age-ing time can be explained by the denaturation of muscle proteins and reduced water holding capacity (WHC) that consequently increases the purge loss (Wiklund, Stevenson-Barry, Duncan, & Littlejohn, 2001).
Analysis of rigor temperature and the interaction of rigor tempera-ture and ageing time showed no effect on cooking loss (Table 3). This is in agreement with previous work done on several animal species (Bekhit et al., 2007; Devine et al., 2002; Geesink et al., 2000; Molette, Rémignon, & Babilé, 2003).Farouk and Swan (1998), however, found that cooking loss decreased in beef ST samples with an increase in rigor temperature. An effect of rigor temperature on cooking loss may be related to the muscle type, but further investigation is required.
Ageing time increased cooking loss (pb 0.01). The increased cooking loss during ageing is possibly due to the myofibrillar protein degrada-tion by endogenous proteolytic enzymes. This disrupts the cellular ma-trix system thereby reducing the muscles' ability to retain moisture resulting in increasing cooking loss (Han, Sung, Kim, Kim, & Ahn, 1996; Kim, Warner, & Rosenvold, 2014).
The relatively high cooking loss percentage in all samples can be at-tributed to the low intramuscular fat (IMF) found in the meat samples. Beef SM muscles used in this study were lean (1.59%, determined from lipid extraction). This is supported by results reported byHan et al. (1996)that showed lower cooking losses with high IMF content. Muscle type is an important factor in determining cooking loss (Bekhit et al., 2007). For instance, LD muscle from Korean native beef is reported to have high IMF and lower cooking loss compared to ST muscle with lower IMF and higher cooking losses (Ba, Park, Dashmaa, & Hwang, 2014).
3.4. Shear force
Shear force values have a direct relationship with meat tenderness (Geesink et al., 2000). An increase in shear force indicates a decrease in the tenderness of meat. The present study showed that rigor temper-ature had no effect on the shear force of beef SM (p = 0.24) (Table 3.). It is interesting, however, to note that beef SM muscles incubated at 15 °C attained relatively low numerical shear force values. It has been report-ed that 15 °C is the optimal temperature for activation ofμ-calpain, deg-radation of calpastatin and increased proteolysis (Zamora, Chaïb, & Dransfield, 1998). Increased proteolysis leads to reduced muscle short-ening and therefore increased meat tenderness (Devine et al., 2002; Geesink et al., 2000). Holding carcasses for up to 10 h at 15–16 °C have been reported to decrease the shear force values thus increase meat tenderness by up to 40% compared to other incubation tempera-tures (Bouton, Harris, Shorthose, & Baxter, 1973).
Table 2
The predicted means for pH and electrical conductivity for hot boned SM muscle aged for 3, 7, 14 and 21 days.
Rigor temperature (°C) pH Electrical conductivity (mS)
3 days 7 days 14 days 21 days 3 days 7 days 14 days 21 days 5 5.58bcd 5.65abc 5.78a 5.81a 6.90d 9.17cd 9.62bcd 10.73abc
15 5.54cd 5.61bc 5.69b 5.75ab 9.95abc 11.33abc 10.22abc 11.90abc
20 5.52cd 5.57bcd 5.62bc 5.71ab 12.32ab 12.62a 12.25ab 12.20ab 25 5.51d 5.54cd 5.61bc 5.68b 12.17ab 12.67a 11.82abc 12.32ab SEM 0.28 0.28 0.28 0.28 0.57 0.57 0.57 0.57
a–dValues with different superscript letters within a column or row and main effect are significantly different at p b 0.05.
Table 3
The predicted means for cooking loss, purge loss and shear force for hot boned SM muscle aged for 3,7, 14 and 21 days.
Rigor temperature (°C) Purge loss (%) Cooking loss (%) Shear force (N)
3 days 7 days 14 days 21 days 3 days 7 days 14 days 21 days 3 days 7 days 14 days 21 days
5 2.05e 2.53de 3.71cde 5.59abcd 31.81cd 39.81a 38.83a 37.51abc 107.56a 95.09ab 81.55abc 59.86bc 15 1.98e 2.98cde 4.13bcde 7.17ab 31.85cd 39.06a 37.35abc 37.36abc 92.60ab 78.72abc 76.07abc 50.14c 20 1.99e 2.71de 4.40bcde 6.33abc 30.53d 38.74a 39.33a 38.28ab 87.45abc 91.11abc 82.88abc 62.19bc 25 1.71e 3.55cde 3.76bcde 8.52a 32.20bcd 37.82abc 34.65abcd 38.28ab 104.90a 84.46abc 73.16abc 57.37bc SEM 0.68 0.68 0.68 0.68 0.61 0.61 0.61 0.61 4.18 4.18 4.18 4.18
Ageing time had an effect on shear force (pb 0.01). As shown in
Table 3, mean shear force decreased with the increase in ageing time (21 days). The substantial decrease in mean shear force with ageing was possibly due to the myofibrillar protein degradation by endogenous enzymes (Koohmaraie, 1996; Lepetit, 2008). Ageing of meat is a useful, common practice in the meat industry as it increases the tenderness (Silva, Patarata, & Martins, 1999).
3.5. Colour analysis
The results for the colour analysis are shown in supplementary data. Lightness (L*) was affected by rigor temperature (p = 0.012), ageing (pb 0.01), and display time (p b 0.01). L*-value was significantly in-creased between 15 and 25 °C treatment groups (pb 0.012). This could be explained by the shrinkage of myofibrils as the rigor tempera-ture increased and moistempera-ture was released to the sample surface in turn leading to increased light scattering (Farouk & Swan, 1998; MacDougall, 1982; Offer et al., 1989).
L*-value was increased with the ageing (pb 0.01). Samples aged for 14 and 21 days were significantly lighter than samples aged for 3 and 7 days (pb 0.05; see supplementary 1). This can be attributed to protein structure modification resulting in higher light reflection due to reduced WHC (Joseph & Connolly, 1977).The low purge loss and higher WHC at earlier ageing times will have contributed to the low light scattering thus lowering L* values. The ageing and display process may increase muscle protein denaturation leading to more purge loss and ultimately higher light scattering accounting for some L* increases (Brewer & Harbers, 1991; Farouk & Swan, 1997).
Redness (a*) increased (p = 0.027) with an increase in rigor tem-perature. This can be attributed to decreased oxygen consumption rates in muscle at higher rigor temperatures (Ledward, 1985). Ageing time and display time both had an effect on a*-value (pb 0.01). The in-teraction of ageing time and rigor temperature also had an effect on a*-value (pb 0.05). The meat samples aged for 3 and 7 days had higher mean a*-values compared to 14 and 21 days. This could be due to the re-duced mitochondrial respiration and oxygen consumption after 7 days of vacuum package storage (Bendall, 1972; DeVore & Solberg, 1974).
The a*-values of all treatments were decreased with display time (pb 0.01). At the start of display time, samples with the highest a*-values were those aged for 3 days, with rigor temperatures of 20 and 25 °C. Among samples aged 7 days, there was little variation between the rigor temperature treatments. The same trend of decreasing a*-values with display time was observed. This decrease can be attributed to increased lipid oxidation due to the exposure of the samples to aero-bic conditions (Bekhit et al., 2007). The large decrease in a*-values of 14 and 21 day aged samples during display (pb 0.001; see supplementary 2) can be explained by the highly reduced metmyoglobin reducing en-zyme activity; this would lead to the accumulation of metmyoglobin and hence lower a* (Farouk & Swan, 1998; Ledward, 1992).
Rigor temperature did not have an effect on yellowness (b*-value) (p = 0.141). The b*-values decreased with both ageing and display times (pb 0.001). Chroma, C was affected by ageing and display time (pb 0.001); however rigor temperature did not affect C. C values de-creased with both ageing and display time.
Browning index (colour difference) was affected by rigor tempera-ture, ageing time, and display time (pb 0.001). There was a significant effect on the ageing and display time interaction (p = 0.014).
Browning index increased with an increase in rigor temperature (pb 0.001), indicating less browning. At higher rigor temperatures there is rapid fall in pH and a lower pHucompared to lower rigor tem-peratures (Section 3.1above). Myoglobin oxidation decreases with an increasing pH (Abril et al., 2001). In the current study, this is likely the reason why lower rigor temperatures showed lower colour differences. Browning index decreased with ageing time (pb 0.001) and display time. This can be attributed to an increased browning in meat due to myoglobin oxidation (Abril et al., 2001). Day 21 samples showed the
fastest rate of decline in colour difference; in other words, day 21 sam-ples had the least colour stability once exposed to aerobic conditions (supplementary 3).
3.6. Lipid oxidation: TBARS
Lipid oxidation was not affected by rigor temperature (pN 0.05) (Fig. 2). Lipid oxidation was, however, affected by ageing time, display time, interaction between ageing time and display time, interaction between ageing time and temperature, and interactions between ageing time∗ display ∗ temperature (all p b 0.01).
Lipid oxidation of the meat samples aged in vacuum packaging for 3 and 7 days was not statistically different (pN 0.05) (Fig. 2). Lipid oxida-tion in samples aged for 14 and 21 days ageing time showed a small in-crease in TBARS value (pb 0.001). The lipid oxidation rapidly increased after 7 days of display time (pb 0.001). The increase in TBARS value with ageing and display time can be explained by the depletion of en-dogenous antioxidants with ageing and exposure to oxygen/aerobic conditions that cause the meat to rapidly oxidise on display (Bekhit et al., 2005; Cheng & Ockerman, 2004; Scanga et al., 2000). Samples aged for 14 and 21 days recorded the highest TBARS value after 7 days of display time. This was possibly because of the changes in the cellular and tissue structure during ageing time. Muscle membrane was no lon-ger an efficient barrier to enzymes and substrates with cellular lipids ox-idation being the net outcome (Morrissey, Sheehy, Galvin, Kerry, & Buckley, 1998). During the ageing process there is release of iron from high molecular sources like haemoglobin, myoglobin and ferritin. This iron is availed to amino acids (made readily available due to cell struc-ture disruption with increased ageing) to form chelates which are active lipid oxidation catalysts (Decker & Crum, 1993; Monahan, 2000). As the aged beef SM meat was exposed to aerobic conditions during display time, it is likely that lipid oxidation became a rapid reaction.
There was also a consistent increase in TBARS values on all meat sam-ples during display time; this is likely attributed to free radical formation initiating further lipid oxidation (autoxidation and photosensitised oxi-dation) due to exposure to aerobic conditions and light. The ageing re-sults were consistent with the rere-sults reported by (Ismail, Lee, Ko, & Ahn, 2008) on ageing of beef SM muscle up to 21 days.
3.7. Lipid oxidation: Rao,Radratios and fatty acid content
The aliphatic to diallylmethylene proton ratio (Rad) was not affected by rigor temperature (5 °C and 25 °C) and ageing time (day 7 and 14) (pN 0.05) (Fig. 3). Display time, however, did have an effect on Rad (pb 0.05). The comparable Radratio between day 7 and day 14 of stor-age suggest minimal loss of PUFAs thus implying low oxidation in
Fig. 2. Predicted means of lipid oxidation (μg/g) of hot boned beef SM muscle at various rigor temperatures aged for 3, 7, and 14 days.(abc
Values with a different letter are significantly different at p b 0.05.)
vacuum packaging and dark storage. Radincreased across the display time for both 5 °C and 25 °C aged samples (Fig. 3A). This can again be explained by the exposure of meat to aerobic conditions and light that encourage formation of potent free radicals like hydroxyl radicals that attack the vulnerable PUFAs to form more fatty acyl radicals (Halliwell & Chirico, 1993; Halliwell & Gutteridge, 1986). With more exposure to oxygen and light, the fatty acyl radicals would react to form hydroper-oxides, fundamental primary oxidation products of lipid degradation. The samples aged for 14 days also had a higher Radratio after the display time. This is possibly due to the depletion of natural antioxidants in the meat and the degraded muscle membrane system from the ageing peri-od; these two factors make the accessibility of chelates that are respon-sible for lipid oxidation catalysis much easier (Decker & Crum, 1993).
The aliphatic to olefinic proton ratio was not affected by rigor temper-ature and ageing time (pN 0.05). Display time did have an effect on Rao (pb 0.05). There was an increase in ratio from the end of ageing time
(14 days) to the end of display time for 25 °C rigor temperature samples as illustrated inFig. 3B. The rest of the samples showed a numerical in-crease in Raoratio from pre-display to post-display but statistically not sig-nificant. The increase in the ratio was low compared to Rad; this is because the olefinic protons are attached directly to the double bonded carbons contributed by the oleic acyl group (monounsaturated fatty acid, MUFA) which are readily oxidised in comparison to the diallylmethylene group in PUFAs (Yang, Grey, Archer, & Bruce, 1998).
The PUFAs numerically decreased with ageing and significantly de-creased (Pb 0.05) with display time as shown inTable 4. There was a higher drop in PUFAs with samples aged 14 days to post-display (end). The high drop supports the increase in Radobserved with1H NMR suggesting that the loss of PUFAs is due to lipid oxidation. Over the display period samples aged for 7 days with rigor temperature 25 °C showed a decline in PUFAs from 6.8 to 5.0% (Table 4) of total FAME (pb 0.05). Samples aged 14 days with rigor temperature 5 °C
Fig. 3. A & B. Aliphatic to diallylmethylene (Rad) (A) and aliphatic to olefinic (Rao) (B) proton ratios for beef SM lipid for rigor temperatures 5 °C and 25 °C, 7 and 14 day aged samples plus
(initial) 7 day display time (end). (abc
Values with different letters are significantly different at p b 0.05.)
Table 4
The distribution of fatty acids in hot boned beef SM muscle exposed to rigor temperatures of 5 °C and 25 °C, aged for 7 and 14 days (initial) and displayed for 7 days (end). 7 days ageing time 14 days ageing time
Pre-display (initial) Post-display (end) Pre-display (initial) Post-display (end) Rigor Temperature 5 °C 25 °C 5 °C 25 °C 5 °C 25 °C 5 °C 25 °C SFA (%) 52.2 ± 0.4ab 52.2 ± 0.5ab 55.6 ± 1.0b 54.9 ± 0.9bc 53.4 ± 1.9ab 53.3 ± 1.2ab 56.6 ± 1.9c 55.9 ± 2.1bc MUFA (%) 40.3 ± 0.4a 41.0 ± 0.8a 38.8 ± 0.6ab 40.1 ± 1.0b 39.9 ± 0.9a 41.0 ± 1.2a 39.4 ± 0.9ab 40.5 ± 1.3b PUFA (%) 7.5 ± 0.2c 6.8 ± 0.6c 5.5 ± 0.9b 5.0 ± 0.9ab 6.7 ± 0.8cd 5.7 ± 0.7cd 4.1 ± 0.6ab 3.7 ± 0.9a PUFA/SFA (%) 0.14cd 0.13cd 0.09b 0.09ab 0.12bc 0.11c 0.07a 0.06a a–dValues with different superscript letters within a row are significantly different at p b 0.05
showed a 6.7 to 4.1% decline (pb 0.05); this supports changes observed in the Radratios. This also suggests increased primary oxidation prod-ucts and ultimately higher secondary oxidation prodprod-ucts as confirmed by TBARS values. The PUFA/SFA ratio showed a decline with ageing and display time. This decrease further illustrates the loss of the unmodified polyunsaturates with ageing and display time under aero-bic conditions.
MUFA decline with ageing and display time was not significant; for instance day 7/5 °C samples showed no significant change (p N 0.05) over the display period. This is probably due to the low propensity of the monounsaturates to oxidise compared to PUFAs. This also goes on to support thefindings with Raoratios showing a low decrease in the ratio with ageing and display time (Yang et al., 1998).
4. Conclusion
This study shows that rigor temperature can be manipulated to hasten the glycolysis process in hot boned beef SM muscle without overly compromising some of the meat quality characteristics such as tenderness, colour and cooking loss. Ageing time also improved the ten-derness of meat and colour L*-values. These qualities, tenten-derness and colour L* values are the most important meat attributes to consumers. Lipid oxidation was not affected by rigor temperature manipulations. Changes in PUFAs over ageing and mainly display time were observed by GC–FID and Radratios. The GC-FID data and Raoalso provided concise information on MUFAs oxidative stability, which were not affected by the ageing times.
1
H NMR was a useful non-destructive technique to monitor oxida-tion in extracted lipid from meat during the ageing and display storage times in hot boned SM beef. Extracted lipid used for1H NMR analysis was readily recovered for further use in different analytical work. The process was rapid and did not require much sample manipulation allowing assessment of quality deterioration in meat during processing and storage.
Acknowledgements
The assistance of the management and staff from Alliance Group and Pukeuri Plant is appreciated. Assistance from Kristina Mungure, Ayodeji Agbowuro, and Siddarth Sai is also appreciated.
Appendix A. Supplementary data
Supplementary data to this article can be found online athttp://dx. doi.org/10.1016/j.meatsci.2015.12.015.
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