PROCESSING, PRODUCTS, AND FOOD SAFETY. Microstructure and Thermal Characteristics of Thai Indigenous and Broiler Chicken Muscles

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1 PROCESSING, PRODUCTS, AND FOOD SAFETY Microstructure and Thermal Characteristics of Thai Indigenous and Broiler Chicken Muscles S. Wattanachant,*,1 S. Benjakul,* and D. A. Ledward *Department of Food Technology, Faculty of Agro-Industry, Prince of Songkla University, Hat Yai, Songkhla, Thailand 90112; and School of Food Biosciences, The University of Reading, Whiteknights, Reading, United Kingdom RG6 6AP ABSTRACT The microstructure and thermal characteristics 75.9 to 76.9 C. The fiber diameters of Thai indigenous of Thai indigenous (Gallus domesticus) and broiler chicken muscles were greater (P < 0.05) than those of the chicken (commercial line CP707) biceps femoris and pectoralis muscles were determined. Perimysium thicknesses 28.9 vs µm for pectoralis muscle, respectively. After broiler, 31.7 vs µm for biceps femoris muscle and were 14.2 µm for biceps femoris muscle and 7.10 µm for cooking at 80 C for 10 min, the fiber diameter of indigenous chicken muscles significantly decreased while those pectoralis muscle of indigenous chicken muscles, thicker than those of broiler muscles, which were 9.93 µm for of the broiler significantly increased. The mean of sarcomere lengths of the raw muscles ranged from 1.56 to 1.64 biceps femoris muscle and 3.87 µm for pectoralis muscle (P < 0.05). Five endothermic peaks with peak transition temperatures (T p ) of 54.9, 61.7, 65.4, 70.6, and 76.1 C were µm and decreased to 0.92 to 1.32 µm(p < 0.001) for broiler obtained for broiler pectoralis muscle, whereas only 3 muscles and 1.22 to 1.35 µm (P < 0.001) for indigenous endothermic peaks (T p of 56.6, 62.6, and 74.9 C) were chicken muscles after cooking. The perimysium and endomysium obtained for broiler biceps femoris muscle. Thai indigenous biceps femoris and pectoralis muscles had endothermic peaks with T p ranges of 53.5 to 54.8, 60.7 to 61.9, and of broiler muscles melted after cooking at 80 C, however, only slight disintegration was observed in these tissues in the indigenous chicken muscles. (Key words: microstructure, thermal properties, chicken muscle, indigenous chicken, connective tissue) 2005 Poultry Science 84: INTRODUCTION Muscle structure can be elucidated using microscopy, laser diffraction, transmission electron microscopy, or scanning electron microscopy (Jones et al., 1976; Sundell et al., 1986; Young et al., 1990; Liu et al., 1996; Palka and Daun, 1999). The morphology of muscle structures (original or changed after treatments) can be related to meat tenderness (Jones et al., 1976; 1977). The main factors determining meat texture are myofibrillar proteins, muscle cytoskeleton, and intramuscular connective tissue (Lawrie, 1991; Palka and Daun, 1999). Additionally, thickness of perimysium, fiber diameter or fiber area, shortening sarcomere, and the structural weakening of intramuscular connective tissue can influence the meat texture (Smith and Fletcher, 1988; Smith et al., 1993; Nishimura et al., 1995; Liu et al., 1996; Palka, 1999; Palka and Daun, 1999). The muscle structure varies depending on muscle types, species, and breed of animal, all of which contribute to differences in texture of muscles. Liu et al. (1996) reported that perimysium thickness varied depending on the muscle types of chicken, and had a high correlation with the shear value. A wide range of sarcomere lengths among bovine muscles was observed but had low correlation with the shear value (Torrescano et al., 2003). Furthermore, muscle fiber types and diameter in chickens, geese, and ducks were compared and found to influence their meat quality (Kiessling, 1977; Smith et al., 1993). Thermal characteristics of various muscle proteins are commonly determined using differential scanning calorimetry (DSC). Thermal transitions of breast, thigh, blood, and skin tissues of chicken broilers were related to denaturation of isolated protein fractions (Kijowski and Mast, 1988). Xiong et al. (1987) reported the different thermal denaturation of chicken breast and thigh muscle proteins from different species. Xiong and Brekke (1990) found a difference in thermal denaturation of extracted salt-soluble proteins between pre- and postrigor chicken muscle. Murphy et al. (1998) evaluated the thermal properties of chicken breast patties compared with their isolated pro Poultry Science Association, Inc. Received for publication March 3, Accepted for publication October 10, To whom correspondence should be addressed: swattanachant@ yahoo.com. Abbreviation Key: DSC = differential scanning calorimetry; H = enthalpy of transition; SEM = scanning electron microscope; T o = onset temperature; T p = temperature of peak transition. 328

2 CHARACTERISTICS OF CHICKEN MUSCLES 329 tein fractions. However, no data regarding the thermal characteristics of Thai indigenous chicken muscles has been reported. The Thai indigenous chicken has a unique taste and texture and is very popular among consumers. The indigenous and broiler chickens are consumed at approximately the same commercial live weight. However, the indigenous chicken generally has slower growth than the commercial broiler, which may contribute to differences in properties of their meats. Wattanachant et al. (2004) reported that Thai indigenous chicken muscles possessed firmer texture, particularly after cooking, than those of commercial broilers, and this might be related to the differences in total and soluble collagen contents between their muscles. However, the microstructure and thermal characteristics of the muscles from both breeds should be further studied to fully understand the differences in their texture. Most studies on the structure and composition of the chicken meat have focused on muscles with high commercial value, especially breast and thigh muscles (Smith and Fletcher, 1988; Smith et al., 1993; Ding et al., 1999). In addition, pectoralis breast muscle and biceps femoris thigh muscle represent tender and tough muscles, respectively, as related to their shear values (Liu et al., 1996). The objective of this study was to compare the microstructure and thermal characteristics of pectoralis and biceps femoris muscles in Thai indigenous and broiler chickens to elucidate the differences in muscle texture. MATERIALS AND METHODS Sample Preparation Mixed-sex Thai indigenous chickens (Gallus domesticus) aged 16 wk, and mixed-sex commercial broilers (line CP707) aged 38 d, of similar liveweights (1.5 ± 0.2 kg) were obtained from a local farm, killed by conventional neck cut, bled for 2 min, scalded at 60 C for 2 min, plucked in a rotary-drum picker for 30 s, and eviscerated. Pectoralis major and biceps femoris muscles were dissected from the carcasses after chilling in a cold room at 4 C for 24 h. The skin was removed and the muscles were trimmed of obvious fat and connective tissue. Muscle samples from 4 birds of each breed were used for each analysis: perimysium thickness, muscle structure, and thermal transition (total of 12 birds of each breed). Muscle structure was determined on raw and cooked muscle samples. To prepare cooked muscles, half of each muscle from 4 birds of each breed were cut into approximately cm pieces for pectoralis major analysis, and approximately whole pieces for biceps femoris analysis to weigh approximately 10 ± 0.5 g. The small pieces of each muscle were put into tightly sealed plastic bags and cooked in a waterbath at 80 C for 10 min. The other 2 Olympus BH-2, Olympus Co., Tokyo, Japan. 3 JSM-5800LV, JEOL Scanning microscope, Jeol Ltd., Tokyo. muscle samples were analyzed raw. The raw and cooked muscles from each breed and muscle type were stored at 4 C and their microstructures were determined within 24 h. Thickness of Perimysium Perimysium thickness was determined using the Picro- Sirius Red polarization method as described by Liu et al. (1996). Raw muscle samples ( cm) were excised from each muscle type of each breed and frozen quickly in liquid nitrogen. Transverse sections (4 µm thick) of the frozen muscle were cut in a cryostat at 20 C. The sections were placed in acetone for 60 min and then fixed in picro-formalin fixative containing 5% formalin, 90% ethanol, and saturated picric acid for 10 min at room temperature. The sections were rinsed in 90% ethanol and under gently running water for 1 min and 10 min, respectively. The sections were then stained with Picro- Sirius Red solution containing 0.1% Sirius Red F3BA and saturated picric acid for 60 min, transferred into 0.01 M HCl for 5 min, and rinsed in distilled water for 1 min. They were dehydrated in 3 changes of absolute ethanol, cleared in 2 changes of xylene, mounted in Perma-Fluor Permanent Aqueous mounting medium, and examined under a light microscope. 2 The thickness of the perimysium was measured at 5-mm intervals along single perimysia on 2 micrographs obtained from each of 4 birds, for both muscle types and breeds. The mean thickness was estimated from the measured values (n = 40). Muscle Structure Muscle structures of samples were determined using scanning electron microscopy (SEM) according to the procedure of Palka and Daun (1999) with slightly modifications. Pieces ( cm) were excised from raw and cooked muscle samples and fixed in 2.5% glutaraldehyde in 0.1 M phosphate buffer ph 7.3 for 2 h at room temperature. The specimens were rinsed with distilled water and dehydrated in graded ethanol solutions of 25, 50, 70, 95%, and absolute ethanol (twice), with dehydration conducted for 1 h in each solution. The samples were cut in liquid nitrogen. Fragments of dried specimens were mounted on aluminum stubs and coated with gold. The specimens were examined and photographed in an SEM, 3 using an accelerating voltage of 5 or 10 kv. The micrographs and videoprints were taken at magnification of 500( (5 kv) for transverse sections and 10,000 (10 kv) for longitudinal sections. Muscle fiber area and sarcomere length were measured on videoprints, using a special morphometric facility. Two videoprints from each of 4 birds of each breed and muscle type were taken. Ten measurements of fiber area and 5 measurements of sarcomere length were made on each (n = 80 and n = 40, respectively). Fiber diameter was calculated from fiber area. The micrographs and videoprints for transverse sections of endomysium and perimysium from raw and cooked muscle specimens

3 330 WATTANACHANT ET AL. TABLE 1. Thickness of perimysium of chicken muscles from broiler and Thai indigenous chickens Thickness of Breed Muscle perimysium (µm) Broiler Bicep femoris m ± 2.28 b pectoralis m ± 1.32 d Thai indigenous Bicep femoris m ± 3.45 a pectoralis m ± 2.56 c a d Means with differing superscripts in the same column are significantly different (P < 0.05). Data are presented as mean ± standard deviation, n = 40. of each breed and muscle types were taken at magnification of 1,000 (5 kv). Thermal Transition Analysis Raw chicken muscles from each breed and muscle type were ground and stored at 4 C for 3 d before thermal analysis. The average moisture content of muscle samples before analysis ranged from 75.2 to 75.9%. For thermal analysis, samples were scanned in a differential scanning calorimeter (DSC), 4 calibrated with indium for heat flow, temperature, and enthalpy. Each muscle sample was weighed (approximately 20 mg) into an aluminum pan and sealed hermetically. The heating rate of the DSC scans was 5 C/min over a range of 25 to 120 C. Empty aluminum pans were used as the reference and for baseline corrections. The onset temperature (T o ), temperature of peak transition (T p ), and enthalpy of transition ( H) were determined from typical thermograms. The determination was done in duplicate for each of 4 birds of each breed and muscle type, n=8. Statistical Analysis Data were evaluated statistically using the SPSS version 10.0 computer program. 5 One-way ANOVA was used to analyze the effects of each muscle type from each breed on thickness of perimysium, fiber diameter, sarcomere length, transformation temperature, and enthalpy of denaturation. A significant difference between treatment means was analyzed by Duncan s Multiple Range Test, and the paired sample t-test was applied to analyze the significant difference of means between raw and cooked conditions (Steel and Torrie, 1980). The interaction between breed and condition was not determined. RESULTS AND DISCUSSION Perimysium Thickness The thicknesses of perimysium of Thai indigenous and broiler chicken pectoralis and biceps femoris muscles are shown in Table 1. The perimysia of indigenous chicken 4 Perkin Elmer DSC7, Perkin Elmer Corp., Norwalk, CT. 5 SPSS Inc., 1999, Chicago, IL. muscles were thicker than those of broiler muscles (P < 0.05). The perimysia of pectoralis muscle were thinner than those of the biceps femoris muscle (P < 0.05). Thick perimysium in indigenous chicken muscles was coincidental with higher collagen content and shear value of this breed muscle, as previously reported by Wattanachant et al. (2004). Liu et al. (1996) reported a high correlation (r 2 = 0.95) between the thickness of perimysium and shear value. In this study, perimysium thickness of the muscles correlated well with the magnitude of the shear values of the muscles, indigenous biceps femoris (5.20 kg), broiler biceps femoris (2.89 kg), indigenous pectoralis (1.78 kg), and broiler pectoralis (1.20 kg) (Wattanachant et al., 2004). The differences in the thickness of perimysium and the collagen contents between breeds may be attributed to differences in age (Dawson et al., 1991). However, total collagen content of muscle does not increase with increasing animal maturity (Dransfield, 1994) and does not correlate significantly with the tenderness of meat (Nakamura et al., 1975). Therefore, the difference between the breeds might be due to genetic factors influencing perimysium thickness and collagen content, and resulting in the differences in textural properties. Thermal Denaturation The onset (T o ) and peak (T p ) temperatures of protein denaturation were determined for the chicken muscles from both breeds and muscle types (Table 2). The enthalpies of protein denaturation of chicken muscles from both breeds are presented in Table 3. Five endothermic peaks at 54.9, 61.7, 65.4, 70.6, and 76.1 C were obtained for broiler pectoralis muscle, whereas only 3 endothermic peaks were found for broiler biceps femoris muscle and Thai indigenous biceps femoris and pectoralis muscles, at temperature ranges of 53.5 to 56.6, 60.7 to 62.6, and 74.9 to 76.9 C. Chicken breast muscle from 7-wk-old chicken broilers has been reported to have 5 endothermic peaks at 57, 62, 67, 72, and 78 C, whereas their thigh muscle had 3 transitions at 59.6, 65.6, and 75.8 C (Kijowski and Mast, 1988). Murphy et al. (1998) found 3 transitions at 52 to 57, 67 to 72, and 76 to 83 C for ground and formed chicken breast patties. The difference in numbers of peaks could be due to the differences in source of raw material, type, age, sex, and storage of chicken meat (Xiong and Brekke, 1989; Murphy et al., 1998). Stabursvik and Martens (1980) concluded that greater differences were found between red and white muscle than between muscles from different species. However, the differences between the breast muscle thermoprofile, with a higher proportion of white muscle fibers, and the thigh muscle thermoprofile, with more red fibers, was not clear (Kijowski and Mast, 1988). Myofibrillar proteins have been reported to exhibit 2 transitions at T p of and C, with H of 0.44 and 0.33 J/g, respectively (Murphy et al., 1998). In this study, we found only one peak corresponding to myofibrillar protein in chicken muscles from both breeds (peak no.1, Tables 2 and 3). Myofibrillar proteins in broiler chicken muscles showed higher T o,t p (peak no.1, Table

4 CHARACTERISTICS OF CHICKEN MUSCLES 331 TABLE 2. Transformation temperature 1 of chicken muscles from broiler and Thai indigenous chickens Peak 1 Peak 2 Peak 3 Peak 4 Peak 5 Breed Muscle T o ( C) T p ( C) T o ( C) T p ( C) T o ( C) T p ( C) T o ( C) T p ( C) T o ( C) T p ( C) Broiler Bicep femoris m a a a a b c (± 0.29) (± 0.29) (± 0.36) (± 0.34) (± 0.86) (± 0.29) pectoralis m bc b b b a b (± 0.82) (± 0.38) (± 0.31) (± 0.36) (± 0.28) (± 0.15) (± 0.15) (± 0.22) (± 0.66) (± 0.11) Indigenous Bicep femoris m b b c b a a (± 0.73) (± 0.86) (± 0.22) (± 0.84) (± 0.77) (± 0.52) pectoralis m c c d c a b (± 0.36) (± 0.30) (± 0.24) (± 0.40) (± 0.58) (± 0.43) a d Means with differing superscripts in the same column are significantly different (P < 0.05). 1 Data are presented as mean ± standard deviation, n=8.t o = onset temperature; T p = temperature of peak transition. TABLE 3. Enthalpy of denaturation ( H) of chicken muscles from broiler and Thai indigenous chickens Peak 1 Peak 2 Peak 3 Peak 4 Peak 5 Breed Muscle H (J/g) H (J/g) H (J/g) H (J/g) H (J/g) Broiler Bicep femoris m ± a ± a ± b pectoralis m ± b ± c ± ± ± c Indigenous Bicep femoris m ± d ± a ± a pectoralis ± c ± b ± bc a d Means with differing superscripts in the same column are significantly different (P < 0.05). Data are presented as mean ± standard deviation, n = 8. 2), and greater enthalpy of denaturation compared with those in Thai indigenous chicken muscles (P < 0.05) (Table 3). For the stromal proteins in broiler and Thai indigenous chicken muscles, one transition (peak no. 2) was observed at peak temperature range of 60.7 to 62.6 C, which was lower than previous reports at 63 C by Xiong et al. (1987), 65.5 C by Kijowski and Mast (1988), and 64.2 C by Murphy et al. (1998). For sarcoplasmic proteins of chicken breast muscle, 3 transitions have been observed at peak temperature of 62, 67, and 72 C (Wang and Smith, 1994), and 64.2, 71.9, and 78.4 C (Murphy et al., 1998). For Thai indigenous chicken muscles, only one peak (peak no. 5) was observed with T p of 75.9 and 76.9 C for biceps femoris and pectoralis muscles, respectively. Three transitions (peaks 3, 4, and 5) were obtained only in broiler pectoralis muscle with T p of 63.8, 65.4, and 76.1 C. These peaks most likely represented sarcoplasmic proteins. For the same muscle tested, Thai indigenous chicken muscle exhibited significantly lower enthalpies for peak no. 1 but higher enthalpies for peak no. 2, compared with broiler muscles. The results suggested that stroma in Thai indigenous chicken muscle was less susceptible to thermal denaturation than that in broilers. This might be associated with the firmer structure of indigenous chicken stroma, resulting in the tougher texture of meat from this breed. Microstructure The results of quantitative structural measurements of raw and cooked Thai indigenous and broiler chicken pectoralis and biceps femoris muscles are shown in Table 4. In raw muscles, the fiber diameter of Thai indigenous chicken muscles was larger than that of broiler muscle (P < 0.05). The biceps femoris muscle of broiler chickens had a smaller fiber diameter than the pectoralis muscle (P < 0.05), whereas the opposite results were observed in the indigenous chicken muscles. The average diameter of chicken white fibers has been variously reported as 68.2 µm (Kiessling, 1977), 38 to 46 µm (Smith and Fletcher, 1988), and 32.6 µm (Smith et al., 1993). These differences TABLE 4. Fiber diameter and sarcomere length of chicken muscles from broiler and Thai indigenous chickens 1 Fiber diameter (µm) Sarcomere length (µm) Significance 2 Significance Breed Muscle Raw Cooked (t-test) Raw Cooked (t-test) Broiler Bicep femoris m ± 2.40 d 26.2 ± 4.36 c *** 1.60 ± 0.14 ab 0.92 ± 0.09 c *** pectoralis m ± 6.43 c 29.9 ± 6.46 a ** 1.64 ± 0.12 a 1.32 ± 0.12 a *** Thai indigenous Bicep femoris m ± 7.21 a 27.3 ± 3.26 bc *** 1.56 ± 0.15 c 1.22 ± 0.15 b *** pectoralis m ± 5.95 b 28.8 ± 5.14 ab NS 1.61 ± 0.16 ab 1.35 ± 0.14 a *** a d Means with differing superscripts in the same column are significantly different (P < 0.05). 1 Data are presented as mean ± standard deviation, n=80formeasurement of fiber diameter, and n = 40 for measurement of sarcomere length. 2 Significant differences between raw and cooked samples were determined by t-test: **P < 0.01; ***P < 0.001; NS = nonsignificant difference.

5 332 WATTANACHANT ET AL. FIGURE 1. Scanning electron micrographs of longitudinal sections of chicken muscles; biceps femoris muscle of broiler, raw (A) and cooked (B), pectoralis muscle of broiler, raw (C) and cooked (D), biceps femoris muscle of Thai indigenous chicken, raw (E) and cooked (F), pectoralis muscle of Thai indigenous chicken, raw (G) and cooked (H). Scale bar indicates 1 µm, magnification = 10,000. in muscle fiber diameter may have been due to the differences in age, rate of rigor onset, and degree of sarcomere shortening (Smith and Fletcher, 1988). After cooking, however, the equivalent muscles from both breeds did not differ in fiber diameter. The fiber diameter of cooked broiler muscles increased by 28.4% for biceps femoris muscle (P < 0.001) and 12.4% for pectoralis muscle (P < 0.01) compared with their raw counterparts. Conversely, the fiber diameter of the Thai indigenous chicken biceps femoris decreased 13.9% (P < 0.001) after cooking, whereas that of pectoralis muscle did not change (P > 0.05) after cooking. The mean sarcomere lengths of the raw muscles from both breeds were similar, although those of the indigenous biceps femoris muscle were slightly shorter than those of the others (P < 0.05). The sarcomere lengths of both muscle types from both breeds were in the range reported by Young et al. (1990). The sarcomere length of all muscles decreased after cooking (P < 0.05), as shown in Table 4. The sarcomere lengths of the cooked muscles decreased by 42.5% for broiler biceps femoris muscle, 19.5% for broiler pectoralis muscle, 21.8% for indigenous chicken biceps femoris muscle, and 16.1% for indigenous chicken pectoralis muscle. These results indicated that the shrinkage found in cooked broiler mus-

6 CHARACTERISTICS OF CHICKEN MUSCLES 333 FIGURE 2. Scanning electron micrographs of transverse sections of chicken muscles; biceps femoris muscle of broiler, raw (A) and cooked (B), pectoralis muscle of broiler, raw (C) and cooked (D), biceps femoris muscle of Thai indigenous chicken, raw (E) and cooked (F), pectoralis muscle of Thai indigenous chicken, raw (G) and cooked (H). Scale bar indicates 50 µm, magnification = 500. cles was more parallel to the fiber direction than the transverse direction compared with shrinkage of the indigenous chicken muscles. This was probably caused by the difference in content and thermal property of myofibrillar protein and collagen between the breeds as evidenced by different thermal transition between them (Tables 2 and 3). As reported by Lepetit et al. (2000), the extent of muscle fiber deformation during cooking depends on the compression stress applied by collagen fibers and resistance of the muscle fibers to compression. The compression forces applied by collagen networks on muscle fiber bundles depends on the amount of collagen present and its thermal solubility, because the greater the thermal solubility of collagen, the less its force of thermal contraction (Lepetit et al., 2000). Broiler muscles contained higher myofibrillar protein content with higher denaturation temperatures and enthalpies (Tables 2 and 3), but lower collagen content with high thermal solubility (Wattanachant et al., 2004). This resulted in greater resistance to compression and less force of thermal contraction from collagen fibers, which, in turn, resulted in the broiler muscle shrinking in parallel and expanding transversely

7 334 WATTANACHANT ET AL. FIGURE 3. Scanning electron micrographs of intramuscular connective tissue of chicken muscles; biceps femoris muscle of broiler, raw (A) and cooked (B), pectoralis muscle of broiler, raw (C) and cooked (D), biceps femoris muscle of Thai indigenous chicken, raw (E) and cooked (F), pectoralis muscle of Thai indigenous chicken, raw (G) and cooked (H). Scale bar indicates 10 µm, magnification = 1,000. to fiber direction. Thai indigenous chicken muscles contained lower myofibrillar protein content, with lower denaturation temperatures but contained higher collagen content with less thermal solubility than broiler muscle, leading to more force of thermal contraction in both directions. The qualitative changes in the microstructure of raw and cooked biceps femoris and pectoralis muscles from both breeds are presented in Figures 1 and 2. On longitudinal sections (Figure 1), shrinkage of the sarcomeres was observed for cooked biceps femoris muscles of broiler and indigenous chickens. Cooking caused weakening and loss of structure of the Z-disks in cooked broiler pectoralis (Figure 1D) and biceps femoris muscles (Figure 1B). The sarcomeres of the cooked indigenous chicken muscles (Figure 1F and 1H) were more compact with little loss of the Z-disks compared with the broiler muscles. This difference was probably because the Z-disk structure of

8 CHARACTERISTICS OF CHICKEN MUSCLES 335 the indigenous chicken muscles was more heat stable than that of the broiler muscles and may contribute to the firmer structure of the cooked indigenous chicken muscles. These observations might partly explain the previous observation that the shear values of the broiler muscles decreased after cooking whereas the shear values of the indigenous chicken muscles increased after processing under the same conditions (Wattanachant et al., 2004). However, differences in the nature of the collagen structures were probably of more importance, as suggested previously. For the transverse section (Figure 2), the fibers in the raw broiler muscles had smaller diameters and were more compact than those of the indigenous chicken muscles. After cooking, denaturation and melting of the endomysium together with the denatured myofibrils resulted in swelling of the fibers of the broiler chicken muscles (Figure 2B and 2D). For the cooked indigenous chicken muscles, however, the muscle fibers were separated from the sheaths of the endomysium (Figure 2F and 2H). This difference may have been caused by differences in crosslinked collagen content between the breeds (Wattanachant et al., 2004). For the indigenous chicken muscles, more highly crosslinked collagens remained insoluble and shrank during heat denaturation, effectively squeezing the heat-denatured myofibrils, and resulting in loss of moisture and decreased fiber diameter. These microstructural changes could result in increasing toughness in the cooked indigenous chicken muscles. For the broiler muscles, heating might cause a softening of connective tissue caused by conversion of collagen to gelatin (Larick and Turner, 1992). Lan et al. (1995) suggested that collagen was the predominant stromal protein in chicken breast muscle. Therefore, the DSC result of peak 2 indicated that the collagen began to shrink in which an endothermic transition was observed at a temperature range of 57.8 to 62.6 C for chicken muscles from both breeds (Table 2), and then the shrunken collagen was converted to gelatin at 80 C as reported by Larick and Turner (1992). The changes in the structure of the intramuscular connective tissues after cooking are shown in Figure 3. The perimysium of broiler biceps femoris and pectoralis muscles lost their wavy sheet structures and melted (denatured) to produce a soft, compact texture after cooking at 80 C (Figure 3A-D). However, the structure of the perimysium and endomysium of the indigenous chicken muscles changed only slightly after cooking under the same conditions (Figure 3E-H). The wavy sheets of perimysium and the sheaths of the endomysium were retained with only slight disintegration. The indigenous chicken muscles had low content of myofibrillar protein with less resistance to thermal denaturation but had thicker perimysium and more highly crosslinked collagen, which were associated with heat stability. Therefore, the thermal characteristics and microstructural results indicate that the thickness of the perimysium directly contributes to the texture of raw chicken muscles, whereas the crosslinked collagen content primarily influences the texture of cooked muscles. The broiler muscle fibers shrank more in parallel than transverse to the fiber axis and expanded in transverse dimensions after cooking, resulting in increasing tenderness of muscles. The indigenous chicken muscle fibers shrank slightly in both dimensions and resulted in a toughening of muscle fibers caused by heat coagulation and shrinkage of myofibrils and shrinkage of intramuscular connective tissue after cooking. Consequently, muscles from both breeds differed in textural characteristics. 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