Histochemical analysis of chicken meat obtained from birds

Histochemical analysis of chicken meat: M. Korzeniowska and T. Smolinska

XVII th European Symposium on the Quality of Poultry Meat Doorwerth, The Netherlands, 23-26 May 2005

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Histochemical analysis of chicken meat obtained from birds supplemented with plant oils

M. KORZENIOWSKA and T. SMOLINSKA*

Department of Animal Products Technology, Faculty of Food Science, Agricultural University of Wrocław, 50-375 Wroclaw, 25/27 Norwida Str., Poland *[email protected] Key words: chicken, muscle fibre, histology, plant oil supplementation

Abstract The aim of the study was an evaluation of the influence of chicken feed supplementation with plant oils contain high amount of polyunsaturated fatty acids on histochemical characteristics of meat. Chickens Vedetta were divided into three experimental groups and they were fed with feed DKA-SFA; control group and with the addition of rapeseed oil of 2.75% and vitamin E 0.015%, as well as of 8.25% of rapeseed oil and 0.03% vit. E. Two weeks before slaughter levels of added plant oils were adjusted to 5.0% in each experimental blocks. Samples were cut from randomly assigned chicken carcasses. M. pectoralis thoracica and M. pectoralis abdominalis breast, and M. iliotibialis thigh were used for trial. During microstructure analyses the number of muscles fibre, minimal diameter, area of muscles fibre and roundness ratio were measured. Measurements were done on 20 different places of the surface of the muscles with an enlarging ratio of 40x10 on the microscope Kontron Elektronik. Supplementation of 8.25% of plant oils to chicken feed caused significant increase of muscles fibre diameter. It was probably caused by muscles fibre hypertrophy phenomena. No significant differences in muscles fibre diameter were observed for smaller amount of supplemented oils. Countable number of muscle fibres was 1.5 times higher in thigh than in breast muscles. Breast muscles were mainly formed by fibres of 40 – 50 µm diameter. Thigh muscles were characterised by smaller diameter of muscles fibre (30 – 40 µm). Along with increasing of the addition of rapeseed oil to the chicken feed increase of fibres area and fibres diameter were observed. Moreover, connective tissue content was diminished. Histochemical analyses of chicken muscles proved that implementation of higher amount of rapeseed oil to the feed had a negative effect on the structure of muscle tissue, and it can be connected with decreasing of sensory and technological characteristics of chicken meat. Introduction Muscle tissue is formed from the muscle fibres, which differ with length, thickness and appearance. Usually, muscle fibres are thicker when they are situated close to the surface of the muscle than in the centre. Moreover, thick fibres are found in muscles which are out of action. Birds skeletal muscle fibres are not uniform, and they are characterised by different morphological and biochemical aspects. Some of the fibres are intensively red, whereas others are pale. In birds red muscles are situated in those part of the body which are still in motion i.e. legs, wings. White muscles are able to work intensively but for a short time. Chicken breast muscles are qualified as white, due to they are formed with white fibres at least in 60%. Thigh muscles consist of 50% of red fibres and are established as red. Muscle tissue, which is built from 55% of white and 45% of red fibres are called intermediate and their character is not fully recognised. Further research performed by Sosnicki and Wilson (1991) and Wilson et. al. (1990) revealed more intermediate forms of muscle fibres. Shape and metabolism of muscle fibres in poultry tissues are closely related to the breeding system and feeding parameters. The final quality of chicken meat is dependent mainly on the feed constituents, especially cereals and lipids components. Smolinska et. al. (2002) reported that fatty acid composition in chicken meat can be modified by the supplementation of different type and amount of lipids to the birds feed. Lipids were then built into muscle tissues changed their microstructure. There are many papers related to the nutritional and chemical changes caused by plant oil supplementation to the chicken diet (Smolinska et. al., 1999), but muscle tissue microstructure alteration connected with lipids addition to the feed is still waiting to be discover.

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Aim The aim of the study was an evaluation of the influence of chicken feed supplementation with plant oils contain high amount of polyunsaturated fatty acids on histochemical characteristics of meat. Material and methods Chickens line Vedetta were divided into three experimental groups (50 per treatment) and they were fed with feed DKA-SFA for 6 weeks - control group (I) and with the addition of rapeseed oil on the level of 2.75% and vitamin E 0.015% (II), as well as on the level 8.25% of rapeseed oil and 0.03% vitamin E (III). Two weeks before slaughter plant oils addition was adjusted to 5.0% in each experimental block. Samples were cut from randomly assigned chicken carcasses. Muscles Musculus pectoralis thoracica and Musculus pectoralis abdominalis were excised from chicken breast and samples of muscles Musculus iliotibialis were cut from chicken thigh. During microstructure analyses the number of muscles fibre, minimal diameter and area of muscles fibre, as well as roundness ratio were measured. Measurements were done on 20 different places of the surface of the muscles (area 512 x 512 µm) with an enlarging ratio of 40 x 10 on the microscope Axiskop Zeiss with high resolution camera RGB, JVC TK-1070E using Kontron Elektronik KS 100 2.0. programme (Makala, 1995). The results were then statistically analysed using programme Statistica 6.0. Results and discussion Analysis of the structure of the material, especially when analysed meat, is based on the well prepared cross-section of the sample. Cross-section of the muscle can be oval, round or irregular depending on the arrangement of the fibres in muscle tissue. When fibres adhere closely one to another cross-sections of the muscle are multilateral. During describing of the muscle tissue structure terms of short and long axes, as well as maximal and minimal axes are used (Kassner and Kuryszko, 1980). The highest amount of the muscle fibres were calculated (on the same counting area) in breast and thigh meat samples collected from chicken fed with standard feed i.e. without any additives. Within 20 observation areas in breast samples 147 muscle fibres were calculated, whilst in thigh cross-section 235 fibres, which was more than 1.5 times more (Table 1). Average diameter measured for muscle fibre in breast samples was running at a level of 40.0 µm, whereas the same parameter in the case of thigh muscles was at about 29.5 µm (Table 1). According to Kassner and Kuryszko (1980) diameter of muscle fibres can be used as an indicator of the types of the fibre. White muscle fibres have a diameter approximately twice as long as red muscle fibres. In the experiment we calculated about 1.5 times less muscle fibres for breast samples than for chicken thigh cross-section. So, it can be suggested that in chicken breast muscles white fibres are dominated, however there are also intermediate fibres with diverse diameter. That is why, when calculate an average diameter for muscle fibres intermediate fibres has to be put together with white or red fibres. Breast muscles samples obtained from control animals were formed in 80% from the fibres with diameter from 30 µm to 50 µm (Table 2), the rest of fibres were characterised by smaller diameter. It can be concluded, that chicken breast muscles is typical white, but also in a some range heterogenic (Figure 1). Distribution of the fibres in chicken thigh muscle is different than in breast tissue. Muscle fibres with diameter above 40 µm made only 6.0%, whereas almost 50% of the fibres had a diameter in the range from 20 µm to 30 µm (Table 2). This results confirmed that thigh muscles are heterogenic with majority of red fibres (Figure 2). Supplementation of rapeseed oil to the chicken feed effected in lowering number of muscle fibres in breast samples at about 25% (Table 1), when 2.75% of oil addition was used. There were no further changes in countable muscle fibres on the standard area along with increasing the plant oil supplementation. The number of muscle fibres in chicken thigh tissue decreased significantly from 235 to 163 when birds were fed with feed contain 8.25% of rapeseed oil (Table 1). Changes observed in muscle fibres content in muscle tissues were connected with alteration of fibres diameters. However, diameters of breast muscle tissue were changed drastically, the average point calculated for fibres diameter when supplemented with oils, was not significantly different from control group of animals. This was caused by occurrence in muscle tissue some of the fibres with larger diameters and some with smaller. Moreover, together with increasing of rapeseed oil addition to the chicken feed a negative effect in breast muscles expressed by the occurrence of giant fibres was observed (Figure 3). This phenomena is known as pectoral myopathy and it is caused by intensive breeding of chickens. Data collected in this study proofed that rapeseed oil supplementation to the birds feed accelerated the formation of giant fibres. Pathological changes of muscle fibres are expressed also by more loosen tissues caused by alteration of fibres shape observed on their cross-section (Klosowska et. al., 1999).



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Shape of the fibres is more round what causes the formation of bigger free spaces between fibres. In our experiment there were not significant differences in average muscle fibres shape indicators between control samples and samples obtained from animals fed with oil supplemented feed (Table 2, 3, 4). Smolinska et. al. (1999) suggested that consumer preferred meat with many small fibres instead of meat formed with large muscle fibres, explained this phenomena by a negative correlation between diameters of the fibres and meat tenderness. Data collected in this study showed that supplementation of rapeseed oil to chicken feed effected in presence of 1.5 times more muscle fibres in thigh muscles than in breast tissue (Table 1). However, summary areas counted for 100 fibres in breast tissue were significantly bigger than in thigh muscles, moreover along with increasing of the oil addition to the feed countable areas of 100 fibres increased (Table 1). Those observation confirmed that supplementation of rapeseed oil to the chicken feed diminished the quality of breast muscles due to occurrence of giant fibres. According to Klosowska et. al. (1999) background of hypertrophy phenomena lies in thickening of muscle fibres or increasing of connective tissue content. In the performed experiment any significant changes in connective tissue content between treatments were observed (Table 1). So, it can be concluded that structural changes, especially in breast muscles, were caused by hypertrophy phenomena. Results collected in this work showed that rapeseed oil supplementation on the level 8.25% effected in increasing of the number of lipid droplets in connective and muscle tissues, which resulted in diminishing of the muscle tissue integrity (Figure 4). The results were comparable with those reported by Smolinska et. al. (2002) and Sosnicki and Wilson (1991). Conclusion

1. Muscle fibre diameter was raised along with increasing of rapeseed oil addition to the chicken feed.

2. Supplementation of rapeseed oil to the chicken feed in amount of 8.25% caused increasing the surface of muscles fibre of about 11% and decreasing of connective tissue content of about 23% in comparison to the control group.

3. Rapeseed oil addition to the chicken feed increased at about 1.5% the amount of muscle fibres in thigh muscles. Thigh muscles tissue was mainly formed with fibres from 20 µm to 40 µm, whilst breast muscles were built with longer fibres (from 40 µm to 50 µm).

4. Plant oil supplementation to the chicken feed changed the integrity of breast and thigh muscles tissue, muscle fibres were less coherent due to loosen connective tissue structure.

References KASSNER J., KURYSZKO J. (1980) Some problems of the morphometric analysis of the mouse

oocytes. Zoologica Poloniae, 27, 10. KLOSOWSKA D., LEWANDOWSKA M., PUCHAJDA H. (1999) Zmiany histopatologiczne w miesniu

piersiowym powierzchniowym (M. pectoralis superficialis) i w miesniu dwugłowym uda (M. biceps femoris) indyczek z roznych grup genetycznych. Zeszyty Naukowe PTZ Przeglad Hodowlany. 45, 15-19.

MAKALA H. (1995) Komputerowa analiza obrazu w technologii żywności ze szczególnym uwzglednienim technologii miesa. Gospodarka Miesna, 9, 12-14.

SMOLINSKA T., MALCZYK E., KRZOWSKI R. (2002) Analiza histochemiczna lipidow miesni jasnych i ciemnych kurczat żywionych pasza wzbogacana w oleje roślinne i α-tokoferol. Żywność, Nauka, Technologia, Jakosc, 30, 116-131.

SMOLINSKA T., POPIEL A., MALCZYK E. (1999) Influence of oil and vitamin E (alfa-tocopherol) supplementation on lipid oxidation and flavour of poultry meat. Proceed. 14th Europ. Symp. Quality Poultry Meat, Bologna, Italy, September 1999, 167-172.

SOSNICKI A., WILSON B.W. (1991) Structure and development of meat animals and poultry. Food Structure, 10.

WILSON B.W., NIEBERG P.S., BUHR R.J. (1990) Turkey muscle growth and focal myopathy. Poultry Science, 69, 1553-1562.



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Table 1 The influence of rapeseed oil supplementation to the chicken feed on the number and average diameter of muscle fibres, summary areas of 100 muscles fibres and connective tissue content in chicken breast and thigh muscles.

Breast muscles Thigh muscles Experimental group Control

group 2.75% oil addition

8.25% oil addition

Control group

2.75% oil addition

8.25% oil addition

Number of muscle fibres

147 109 109 235 225 163

Average diameter of muscle fibres

40.04 42.09 42.76 24.49 30.30 32.92

Summary areas of 100 muscle fibres

171452.7 194527.3 196085.3 88926.7 95132.0 117253.3

% content of connective tissue

34.51 34.48 28.05 31.98 27.53 24.70

Table 2 Distribution of muscle fibres number and average shape indicator in breast and thigh muscles excised from chicken fed with standard feed (control group) in relation to range of muscle fibres diameter.

Breast muscles Thigh muscles Range of muscle fibres

diameter μm Number of

fibres % of

muscles fibres

average shape

indicator

Number of fibres

% of muscles

fibres

average shape

indicator 0-10 0 0.0 0.00 0 0.0 0.00 10-20 2 1.3 0.79 14 9.3 0.85 20-30 16 10.7 0.84 73 48.7 0.83 30-40 54 36.0 0.81 54 36.0 0.84 40-50 66 44.0 0.83 8 5.3 0.89 50-60 11 7.3 0.85 1 0.7 0.92 60-70 0 0.0 0.00 0 0.0 0.0 70-80 1 0.7 0.79 0 0.0 0.0 80-90 0 0.0 0.00 0 0.0 0.0

90-100 0 0.0 0.00 0 0.0 0.0 Table 3 Distribution of muscle fibres number and average shape indicator in breast and thigh muscles excised from chicken fed with feed supplemented with 2.75% of rapeseed oil in relation to range of muscle fibres diameter.



fibres % of

muscles fibres

average shape

indicator

Number of fibres

% of muscles

fibres

average shape

indicator 0-10 0 0.0 0.00 0 0.0 0.00 10-20 3 2.0 0.89 12 8.0 0.84 20-30 23 15.3 0.83 59 39.3 0.83 30-40 33 22.0 0.83 73 48.7 0.82 40-50 57 38.0 0.82 6 4.0 0.84 50-60 27 18.0 0.87 0 0.0 0.0 60-70 5 3.3 0.89 0 0.0 0.0 70-80 2 1.3 0.89 0 0.0 0.0 80-90 0 0.0 0.00 0 0.0 0.0

90-100 0 0.0 0.00 0 0.0 0.0



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Table 4 Distribution of muscle fibres number and average shape indicator in breast and thigh muscles excised from chicken fed with feed supplemented with 8.25% of rapeseed oil in relation to range of muscle fibres diameter.



fibres % of

muscles fibres

average shape

indicator

Number of fibres

% of muscles

fibres

average shape

indicator 0-10 0 0.0 0.00 1 0.7 0.81 10-20 1 0.7 0.82 11 7.3 0.86 20-30 16 10.7 0.85 43 28.7 0.81 30-40 49 32.7 0.80 61 40.7 0.81 40-50 47 31.3 0.82 30 20.0 0.80 50-60 33 22.0 0.85 3 2.0 0.76 60-70 4 2.7 0.89 1 0.7 0.71 70-80 0 0.0 0.00 0 0.0 0.0 80-90 0 0.0 0.00 0 0.0 0.0

90-100 0 0.0 0.00 0 0.0 0.0



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Figure 1 Cross-section of chicken breast muscle (control group)

Figure 2 Cross-selection of chicken thigh musle (control group)

Figure 3 Cross-selection of chicken breast muscle obtained from birds fed with fodder supplemented with 2.75% of rapeseed oil.

Figure 4 Cross-section of chicken thigh muscle obtained from birds fed with feed supplemented with 8.25% of rapeseed oil.

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Histochemical analysis of chicken meat obtained from birds