## 2 Materials and Methods  
The experimental materials comprised _longissimus lumborum_ (LL, n = 8) and _semimembranosus_ (SM, n = 8) muscles obtained from the carcasses of Polish Holstein-Friesian bulls (\(19.2\pm 0.6\) months of age, body weight of \(598\pm 18.8\) kg). The same animals were used in our previous study which investigated the influence of different feed additives on beef quality [29], and they constituted the control group. In that study, blooming time was determined in all experimental groups fed different feed additives. The rearing conditions were described in detail in our previous study [29]. The muscles were removed from the left half-carcass of each animal 24 h post-mortem, and were transported to a laboratory in a cool box at refrigerated temperature. The muscles were kept at refrigerated temperature (4 \(\pm\) 1 \({}^{\circ}\)C) overnight. Subsequently, each muscle was divided into two cuts, and after color measurement (day 1), each cut was packaged individually in a vacuum pouch (PA/PE, thickness 70 \(\upmu\)m, Inter Arma Ltd., Rudawa, Poland). Vacuum-packaged meat was stored until 9 and 14 days post-mortem [30] at 4 \(\pm\) 1 \({}^{\circ}\)C (Memmert GmbH, Schwabach, Germany). Beef color was determined on days 1, 9, and 14 of ageing, on freshly cut muscles (2.5 cm thick steaks, approx. 230 g) during 60 min blooming (4 \({}^{\circ}\)C), according to the standard conditions of the CIE LAB system using, (MiniScan XE Plus, HunterLab, Reston, Virginia 20190 USA) with standard illuminant D65, a 10\({}^{\circ}\) standard observer angle and a 2.54 cm diameter aperture. Before measurements, the device was calibrated using white and black tiles. The values of lightness (L *), redness (a *), and yellownes (b *) were measured at 5 min intervals in three different locations on the muscle's cross-section. The values of hue (H * = tan\({}^{-1}\)(b */a *)) and chroma (C * = (a *2 + b *2)\({}^{1/2}\)) were calculated [12]. The total color change (\(\Delta\)E) over the entire blooming period (between 0 and 60 min measurements) was then calculated for each sample, using Equation (1):  
\[\Delta\mathrm{E}=[(\Delta\mathrm{L}^{*})^{2}+(\Delta\mathrm{a}^{*})^{2}+( \Delta\mathrm{b}^{*})^{2}]^{0.5} \tag{1}\]  
where \(\Delta\)L *, \(\Delta\)a *, and \(\Delta\)b * are the derivatives of corresponding parameters, respectively. \(\Delta\)E\({}_{0}\) was calculated to determine the time after which the color stabilizes (reflected in the absence of significant differences between mean values). \(\Delta\)E\({}_{0}\) denotes the difference in color measured on freshly cut beef (time 0) and during blooming. \(\Delta\)E\({}_{\mathrm{xy}}\), which denotes the difference in color between two consecutive measurements made on a sample during blooming, was calculated to determine the intensity of color changes taking place at 5 min intervals. Due to the fact that the data in each experimental group met the assumptions of normal distribution (Shapiro-Wilk test) and the homogeneity of variance ([PERSON]'s test), an analysis of variance (ANOVA) was conducted, followed by [PERSON]'s test for comparisons of mean values (at \(p<0.05\)). Two-way ANOVA (Statistica 12, StatSoft Inc., Tulsa, OK, USA) was performed to determine the effects of ageing time (three levels: 1, 9, and 14 days) and muscle type (two levels: LL and SM) on the color parameters of beef. Differences in the values of color parameters L *, a *, b *, C *, and H * over time were analyzed using repeated measures ANOVA with 13 sampling dates, separately for each color attribute. The same procedure, with 12 sampling dates, was used when analyzing the values of \(\Delta\)E\({}_{0}\) and \(\Delta\)E\({}_{\mathrm{xy}}\). The values obtained on different sampling dates were treated as dependent variables, whereas time was a repeated measures factor. The significance of differences between the values obtained on different sampling dates was determined with the Bonferroni test.