Hanwoo and Chikso steers (24±1 month old) were finished on a grain-based diet with 12% crude protein and 74% total digestible nutrients for 180 days prior to slaughter at Gangwon Provincial Livestock Research Center’s farm. To avoid an effect on aroma volatiles from quality grade or fat content, strip loin cuts (longissimus lumborum) were collected from 11 Hanwoo carcasses and 8 Chikso carcasses with the same quality grade 1. Samples were vacuum packaged and chilled at 2°C±2°C. Meat quality and aroma analyses were performed at day 4 postmortem.

Samples were ground using a food blender at minimum speed for 10 s (HMF-1600PB, Hanil Electronics Co., Ltd., Seongnam, Korea). Proximate composition was determined by AOAC official methods [11]. Moisture content was determined by drying the samples in the oven at 105°C for 24 h. Crude fat content was determined by ether extraction using the Soxhlet system. Nitrogen content was determined using the Kjeltec system (2200 Kjeltec Auto Distillation Unit, Foss, Hillerød, Denmark), and crude protein was calculated as nitrogen content multiplied by 6.25. Crude ash was determined by burning the samples in the muffle furnace at 550°C for 12 h.

An Agilent gas chromatography system (6890N, Agilent Technologies, Santa Clara, CA, USA) with an auto sampler (7683, Agilent Technologies, USA) was used for determining fatty acid composition. The fat was extracted from ground meat samples with a chloroform-methanol (2:1 v/v) solution and prepared in triplicate [12]. Fatty acid methyl esters were prepared prior to injection then dissolved in hexane according to the AOAC method [13]. The sample (1 μL) was injected into the gas chromatograph port by the auto sampler. The injector temperature was set at 250°C with a split ratio of 100:1. Fatty acid methyl esters were separated using a WCOT-fused silica capillary column (100 m×0.25 mm i.d., 0.20 μm film thickness; Varian Inc., Palo Alto, CA, USA) with a 1.0 mL/min helium flow. The oven was programmed as follows: 150°C/1 min, 150°C to 200°C at 7°C/min, 200°C/5 min, 200°C to 250°C at 5°C/min, and 250°C/10 min. The temperature of the detector was set at 275°C. The fatty acid peaks were identified using the retention time of fatty acid standards (47015-U, Sigma-Aldrich Corp., LLC., St. Lois, MO, USA). The peak area of each identified fatty acid was used to calculate the proportion (%) against the total identified peak area.

Samples were sliced into 3×3×2.5 cm pieces and cooked for 4 min using a skillet on a hot plate set at 165°C (Clerin, Zhongsan Tonsun Electric Appliance Co., Ltd., Guangdong, China). During cooking, samples were flipped every minute. Samples were then cooled for 5 min on aluminum foil and ground with a food blender (Hanil Electronics Co., Ltd., Korea), and aroma volatiles and patterns were immediately measured.

Aroma volatiles from cooked samples were separated and identified by gas chromatography-mass spectrometry (GC-MS) using a modified version of the method described in Ba et al [16]. Samples (3 g) were prepared in duplicate and put into 50 mL headspace vials. The vials were heated to 60°C in a drying oven for 10 min, and carboxen/polydimethylsiloxane fiber (Supelco, Sigma-Aldrich Corp., LLC., USA) with a 75 μm diameter was injected into the vial for extraction for another 30 min. Following extraction, the fiber was injected into the GC port, which was set to 250°C, and the volatiles were desorbed for 5 min at a 1:5 split ratio. Separation was performed using a DB5 fused silica column (30 m×0.25 mm inner diameter, 0.25 μm film thickness, J&W Scientific, Folsom, CA, USA) in a gas chromatograph (7890A Agilent Technologies, USA). The GC oven was programmed to operate at an initial temperature of 40°C for 2 min, increasing to 160°C (by 5°C/min), then to 180°C (by 6°C/min, holding time of 5 min), and finally to 200°C (by 10°C/min, holding time of 5 min). The interface and quadruple temperatures were 280°C and 150°C, respectively. Helium was used as the carrier gas with a flow rate of 1 mL/min. Volatile compounds were detected using a mass spectrometer (5975C, Agilent Technologies, USA). The ion source temperature of the MS was set to 280°C with an electron impact of 70 eV. A scanning mass range of 50 to 450 m/z with a scan rate of 1 scan/s was used. Tentative identification was performed by comparing the experimental spectra to the National Institute of Standards and Technology (NIST) Mass Spectral Library. Data are presented as area unit×10⁶/g.

The aroma pattern was observed using an electronic nose (FOX3000, Alpha MOS, Toulouse, France). A total of 2 g of cooked sample was placed into a 10 mL headspace vial and was prepared in duplicate. The vials were then sealed with rubber septa (Supelco 29176-U, Sigma-Aldrich Corp., LLC., USA). The samples were heated at 60°C for 600 s with an agitation speed of 500 rpm. The 2.5 mL of gas in the headspace of the samples was extracted with an automatic sampler syringe (HS 100, Alpha MOS, France) at 65°C, flow-injected into the carrier gas (synthetic air with a purity quotient >99.99%, pressure set to 0.5 bar) flow (150 mL/min) of the electronic nose and detected using a six metal oxide sensors (MOS) array system consisting of PA2, T70/2, P40/1, P10/2, P10/1, and T30/1 sensors.

The statistical significance of the differences between means from different breeds was determined using one-way analysis of variance. Analyses were performed using R-version 3.2.0 with the “Agricolae” library (The R Foundation for Statistical Computing, Vienna, Austria). For the aroma pattern determined by electronic nose, the maximum resistance ratio was considered as the value for one measurement. Two-dimensional principal component analysis was used for data processing (Alpha Soft package version 8.01, Alpha MOS, France).

As samples were taken from carcasses of the same quality grade 1, no significant differences were found in proximate composition of strip loin between Hanwoo and Chikso beef, as shown in Table 1. The results of the present study were similar to those of a previous study, in which the fat content in quality grade 1 Hanwoo strip loin ranged from 10% to 16% [17].

Hanwoo beef has a unique fatty acid composition with abundant oleic acid (C18:1n9). The proportion of oleic acid increases as IMF content increases [17–19]. A comparison study revealed that the proportion of oleic acid in Hanwoo beef with quality grade 1++ is significantly higher than that in American crossbreed beef with choice grade [20]. However, no significant differences were found in fatty acid composition of strip loin fat between Chikso and Hanwoo beef in this study, as shown in Table 2. Overall, the fatty acid composition agreed with previous work [18]. Monounsaturated fatty acids (MUFA) were predominant in both breeds, contributing approximately 55% of total fatty acids. MUFA were followed by saturated fatty acids (SFA) and polyunsaturated fatty acids (PUFA), which contributed approximately 42% and 2.7%, respectively. Among MUFA and SFA, oleic acid had the highest contribution of up to 49% of total fatty acids, followed by palmitic (C16:0, 27%), stearic (C18:0, 10%), palmitoleic (C16:1n7, 5.8%), and myristic acid (C14:0, 3.3%), respectively. Linoleic acid (C18:2n6) was the predominant PUFA, contributing approximately 1.9% of total fatty acids. It was followed by arachidonic (C20:4n6, 0.30%), alpha-linolenic (C18:3n3, 0.10%), gamma-linolenic (C18:3n6, 0.09%), adrenic (C22:4n6, 0.08%), eicosapentaenoic (C20:5n3, 0.06%), and docosahexaenoic acid (C22:6n3, 0.04%), respectively. These results suggest that fatty acid composition is similar in Hanwoo and Chikso meat.

Differences in meat physical quality traits were not observed between Hanwoo and Chikso in this study, as shown in Table 3. The surface lightness (L*) values are in line with previous study [17]. However, the redness (a*) values of our findings are slightly higher than previously reported [17]. The longer blooming time used in this study (60 min) might have contributed to these differences. Wulf and Wise [21] reported that the L* value of beef would be stable after approximately 30 min of blooming, while a* and b* values would be stable after 78 min of blooming. In general, moisture, fat and meat pH affect water-holding capacity, cooking loss and tenderness [22]. Meat physical quality traits observed in this study (e.g., instrumental surface color, water holding capacity, cooking loss, and shear force values) were not different among breeds as no differences were found in proximate composition and meat pH.

The volatiles released from roasted Hanwoo and Chikso beef are shown in Table 4. The total peak area of all identified volatiles was higher (p<0.02) in Chikso beef than in Hanwoo beef. Although the fat amount of Hanwoo beef was slightly higher than that of Chikso beef, the aroma intensity did not exhibit the same trend. Chikso beef released more toluene, heptanal, octanal and nonanal than Hanwoo beef. According to total peak area, toluene, heptanal, octanal and nonanal contributed 2.46%, 3.28%, 3.63%, and 3.72% of total identified volatiles, respectively, for Chikso beef. Lower proportions were observed in Hanwoo beef, in which toluene, heptanal, octanal and nonanal contributed 1.23%, 2.50% 2.19%, and 2.60% of total identified volatiles, respectively. Overall, aldehydes were predominant, in addition to other volatile groups. Hexanal and butyraldehyde were two predominant aldehydes, contributing 23.4% and 19.6% of total identified volatiles, respectively, for Hanwoo beef and 27.5% and 22.8% of total identified volatiles, respectively, for Chikso beef. Previous studies also demonstrated aldehydes to be the major volatiles in beef and showed that volatile compounds are mostly affected by a diet that alters the meat fatty acid composition [19,23,24]. Most aldehydes are the products of thermal oxidation of unsaturated fatty acids and exhibit some aroma notes such as fatty, buttery and green [4,8,25]. Although no significant differences were found in fatty acid composition, a slight difference in fat content affects the amount of each fatty acid. Higher fat content was found in Hanwoo beef, suggesting that Hanwoo beef also contained higher unsaturated fatty acids than Chikso beef. However, the estimated proportion of fat from IMF (neutral lipid) and muscle cell membrane (phospholipid bilayer or polar lipid) was not measured in this study. Wood et al [26] mentioned that the deposition of PUFA into muscle phospholipids is higher in cattle than in pig and sheep. The amount of muscle cells, which contain phospholipids, also declines when the IMF is deposited more because muscle cells and adipocytes interplay during growth [27]. As Hanwoo beef had a slightly higher fat content than Chikso beef, the tendency of bigger or wider marbling areas might be present in Hanwoo beef. In this study, a whole slice was used for pan-roasting the steak instead of a ground patty. The marbling pattern could affect the aroma development on the surface of the steak. The polar lipids in the muscle cell membrane, which contains mainly PUFA, are more prone to thermal oxidation than the neutral lipids in IMF [28]. Therefore, Chikso beef released more lipid oxidation-related aldehydes (heptanal, octanal, and nonanal) than Hanwoo beef. Further, as aldehydes content were higher than the content of other volatile groups, the area unit for total identified volatiles was higher in Chikso beef than in Hanwoo beef.

Other identified volatiles found in this study were pyrazines (2-methyl and 2,5-dimethyl), butanoic acid and 2(5H)-furanone. Pyrazines contributed 8.2% in Hanwoo and 3.9% in Chikso, butanoic acid contributed 7.6% in Hanwoo and 5.6% in Chikso, while 2(5H)-furanone contributed 6.0% in Hanwoo and 5.2% in Chikso. Meaty, roast and nutty aromas are influenced by Maillard-derived pyrazines, and the sweaty note and odor tenacity are positively correlated with the amount of butanoic acid, whereas 2(5H)-furanone is associated with sweet or caramel notes [25,29].

The aroma pattern of roasted Chikso and Hanwoo beef was well-discriminated by electronic nose (Figure 1A). The sensor resistance ratio, which was computed using a radar fingerprint chart of six MOS, is shown in Figure 1B. Figure 1A shows that Chikso beef has more distance from blank (air) than Hanwoo beef, as Chikso beef had higher resistance ratios of PA2, T70/2, P40/1, P10/2, P10/1, and T30/1 than Hanwoo beef. The total contribution rate of component 1 (C1) and component 2 (C2) was 99.82% and 0.11%, respectively. These results suggest that the aroma patterns of Hanwoo and Chikso beef differed in terms of concentration and proportion of volatiles. These findings agree with our gas chromatography results, in which Chikso beef released more intense aroma than Hanwoo beef.