*2.6. Warner-Bratzler Shear Force*

Warner-Bratzler shear force were measured according to Peng et al. [11]. Samples were individually placed in a plastic bag in temperature-equilibrated water baths (F38-ME, Julabo, Seelbach, Germany) set at 75 ◦C and cooked to internal temperature of 71 ± 0.5 ◦C. The internal temperature was monitored using T-type thermocouples inserted to the middle of meat samples and the thermocouples were connected to a Grant Squirrel Series 2020 datalogger (Grant Instruments Ltd., Cambridge, UK). After cooking, the samples were chilled in an ice water bath for 30 min and stored at 4 ◦C overnight. Six 4 cm long rectangular strips with 1 cm × 1 cm cross section area were obtained from each sample by cutting parallel to the muscle fibres. Each strip was sheared using a Lloyd Instruments LRX Materials Testing Machine (Lloyd Instruments Ltd., Hampshire, UK) equipped with a 5000 N load cell and a V-shape Warner-Bratzler shear force blade at an extension rate of

300 mm/min. The WBSF (N) of each sample was expressed as the average peak force of measurements from the six strips.

#### *2.7. Texture Profile Analysis (TPA)*

Texture profile analysis was measured using a double bite compression procedure outlined previously Peng et al. [11]. A piece of meat measuring 1 cm in thickness was obtained from each sample. The meat was compressed twice at the same position by a 6.3-mm diameter plunger which was driven 8 mm into the sample at a crosshead speed of 50 mm/min using Lloyd Instruments LRX Materials Testing Machine (Lloyd Instruments Ltd., Hampshire, UK) equipped with a 5000 N load cell. Hardness (N) (first bite compression), cohesiveness, and chewiness (N) were measured. TPA values for each sample were averaged from six measurements.

#### *2.8. Lipid Oxidation*

Lipid oxidation in meat was assessed by 2-thiobarbituric reactive substances (TBARS) procedure as reported by Sorensen and Jorgensen [12] with modifications. For each sample, duplicate (5 g) from each sample were finely chopped and homogenised in 12.5 mL of chilled (4 ◦C) trichloroacetic acid (TCA) solution (20% TCA (*w*/*v*) in 2 M phosphoric acid) at 12,000 rpm for 1.5 min using a Polytron PT 10–35 GT homogeniser (Thermo Fisher Scientific, VIC, Australia). The homogenate was then centrifuged at 1800× *g* using a Rotina 380R Hettich Centrifuge (LabGear, South Melbourne, VIC, Australia) for 10 min at 4 ◦C. The supernatant was filtered using Whatman filter paper no. 1. Equal volumes of the filtrate and 5 μM 2-thiobarbituric acid (TBA) were mixed and incubated in a test tube at 95 ◦C for 60 min. Following incubation, the tube was placed on ice for 15 min. Absorbance at 532 nm was measured for duplicate aliquots from each tube using a Multiskan spectrophotometer (Thermo Fisher Scientific, Scoresby, VIC, Australia). Malondialdehyde (MDA) was quantified against a standard calibration curve with 1,1,3,3-tetraethoxypropane (TEP). Results were expressed as mg MDA·kg−<sup>1</sup> meat.

## *2.9. Total Carbonyl Content*

Carbonyl content of the meat samples was determined according to Lund et al. [13] with modifications. Briefly, 1 g of meat samples were homogenised for 1 min at 15,000 rpm in 15 mL of homogenisation buffer (2.0 mM Na4P2O7, 10 mM Tris-maleate, 2 mM EGTA, 100 mM KCl, pH 7.4) using a Polytron PT 10–35GT homogeniser (Thermo Fisher Scientific, VIC, Australia). Two equal aliquots (0.5 mL) from the homogenate were washed with a HCl:acetone (3:100 *v*/*v*) solution three times followed by washing with 10% (*w*/*v*) TCA twice. Out of the two identical samples, (i) 0.5 mL of DNPH dissolved in 2 M HCl was added to the first sample for carbonyl derivatisation and (ii) 0.5 mL of 2 M HCl was added to the other sample for protein concentration determination. Absorbance of the samples were measured at 280 nm to determine protein concentration against a standard curve with BSA (Sigma-Aldrich, Castle Hill, NSW, Australia); and, at 370 nm to determine total carbonyl content. Carbonyl concentration was determined by using the absorption coefficient at 370 nm for the hydrazones formed (22,000 M−1·cm−1) against the protein concentration and expressed as nmol·mg−<sup>1</sup> protein.
