• TABLE OF CONTENTS
HIDE
 Title Page
 Acknowledgement
 Table of Contents
 List of Tables
 List of Figures
 Introduction
 Review of literature
 Experimental procedure
 Results and discussion
 Summary and conclusions
 Appendix
 Reference
 Biographical sketch






Title: Physical and chemical changes occurring in beef, post-mortem, as related to tenderness and other quality characteristics /
CITATION PDF VIEWER THUMBNAILS PAGE IMAGE ZOOMABLE
Full Citation
STANDARD VIEW MARC VIEW
Permanent Link: http://ufdc.ufl.edu/UF00098223/00001
 Material Information
Title: Physical and chemical changes occurring in beef, post-mortem, as related to tenderness and other quality characteristics /
Physical Description: xv, 230 leaves : ill. ; 28 cm.
Language: English
Creator: Taki, Ghazi Hussni, 1934-
Publication Date: 1965
Copyright Date: 1965
 Subjects
Subject: Beef   ( lcsh )
Animal Science thesis Ph. D
Dissertations, Academic -- Animal Science -- UF
Genre: bibliography   ( marcgt )
non-fiction   ( marcgt )
 Notes
Thesis: Thesis (Ph. D.)--University of Florida, 1965.
Bibliography: Bibliography: leaves 221-228.
General Note: Typescript.
General Note: Vita.
Statement of Responsibility: by Ghazi Hussni Taki.
 Record Information
Bibliographic ID: UF00098223
Volume ID: VID00001
Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
Resource Identifier: alephbibnum - 000405750
oclc - 24683309
notis - ACF1993

Downloads

This item has the following downloads:

PDF ( 9 MBs ) ( PDF )


Table of Contents
    Title Page
        Title page
        Page i
    Acknowledgement
        Page ii
        Page iii
    Table of Contents
        Page iv
        Page v
        Page vi
        Page vii
    List of Tables
        Page viii
        Page ix
        Page x
        Page xi
        Page xii
        Page xiii
    List of Figures
        Page xiv
        Page xv
    Introduction
        Page 1
        Page 2
        Page 3
        Page 4
        Page 5
        Page 6
    Review of literature
        Page 7
        Page 8
        Page 9
        Page 10
        Page 11
        Page 12
        Page 13
        Page 14
        Page 15
        Page 16
        Page 17
        Page 18
        Page 19
        Page 20
        Page 21
        Page 22
        Page 23
        Page 24
        Page 25
        Page 26
    Experimental procedure
        Page 27
        Page 28
        Page 29
        Page 30
        Page 31
        Page 32
        Page 33
        Page 34
        Page 35
        Page 36
        Page 37
        Page 38
        Page 39
        Page 40
        Page 41
        Page 42
        Page 43
        Page 44
        Page 45
        Page 46
        Page 47
        Page 48
        Page 49
    Results and discussion
        Page 50
        Page 51
        Page 52
        Page 53
        Page 54
        Page 55
        Page 56
        Page 57
        Page 58
        Page 59
        Page 60
        Page 61
        Page 62
        Page 63
        Page 64
        Page 65
        Page 66
        Page 67
        Page 68
        Page 69
        Page 70
        Page 71
        Page 72
        Page 73
        Page 74
        Page 75
        Page 76
        Page 77
        Page 78
        Page 79
        Page 80
        Page 81
        Page 82
        Page 83
        Page 84
        Page 85
        Page 86
        Page 87
        Page 88
        Page 89
        Page 90
        Page 91
        Page 92
        Page 93
        Page 94
        Page 95
        Page 96
        Page 97
        Page 98
        Page 99
        Page 100
        Page 101
        Page 102
        Page 103
        Page 104
        Page 105
        Page 106
        Page 107
        Page 108
        Page 109
        Page 110
        Page 111
        Page 112
        Page 113
        Page 114
        Page 115
        Page 116
        Page 117
        Page 118
        Page 119
        Page 120
        Page 121
        Page 122
        Page 123
        Page 124
        Page 125
        Page 126
        Page 127
        Page 128
        Page 129
        Page 130
        Page 131
        Page 132
        Page 133
        Page 134
        Page 135
        Page 136
        Page 137
        Page 138
        Page 139
        Page 140
        Page 141
        Page 142
        Page 143
        Page 144
        Page 145
        Page 146
        Page 147
        Page 148
        Page 149
        Page 150
        Page 151
        Page 152
        Page 153
        Page 154
    Summary and conclusions
        Page 155
        Page 156
        Page 157
        Page 158
        Page 159
        Page 160
        Page 161
    Appendix
        Page 162
        Page 163
        Page 164
        Page 165
        Page 166
        Page 167
        Page 168
        Page 169
        Page 170
        Page 171
        Page 172
        Page 173
        Page 174
        Page 175
        Page 176
        Page 177
        Page 178
        Page 179
        Page 180
        Page 181
        Page 182
        Page 183
        Page 184
        Page 185
        Page 186
        Page 187
        Page 188
        Page 189
        Page 190
        Page 191
        Page 192
        Page 193
        Page 194
        Page 195
        Page 196
        Page 197
        Page 198
        Page 199
        Page 200
        Page 201
        Page 202
        Page 203
        Page 204
        Page 205
        Page 206
        Page 207
        Page 208
        Page 209
        Page 210
        Page 211
        Page 212
        Page 213
        Page 214
        Page 215
        Page 216
        Page 217
        Page 218
        Page 219
        Page 220
    Reference
        Page 221
        Page 222
        Page 223
        Page 224
        Page 225
        Page 226
        Page 227
        Page 228
    Biographical sketch
        Page 229
        Page 230
        Page 231
        Page 232
Full Text








PHYSICAL AND CHEMICAL CHANGES
OCCURRING IN BEEF, POST-MORTEM,
AS RELATED TO TENDERNESS
AND OTHER QUALITY
CHARACTERISTICS












By
GHAZI HUSSNI TAKI










A DISSERTATION PRESENTED TO THE GRADUATE COUNCIL OF
THE UNIVERSITY OF FLORIDA
IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE
DEGREE OF DOCTOR OF PHILOSOPHY










UNIVERSITY OF FLORIDA
August, 1965























AGRI-
CULTURAL
LIBRARY















UNIVERSITY OF FLORIDA
II 3 1262I II I I 08552III I I5391
3 1262 08552 5391














ACKNOWLEDGMENTS


The author wishes to express his deepest appreciation and thanks to

Dr. A. Z. Palmer, Chairman of his supervisory committee, for encourage-

ment, perceptive counsel and assistance in the completion of this study and in

the preparation of this dissertation.

Special thanks are due Dr. F. W. Knapp, member of the author's ad-

visory committee for his exceptionally kind advice, interest, encouragement

and in the preparation of this dissertation.

The author also wishes to acknowledge the generous and stimulating

counsel which he received from other members of his advisory committee in

planning and conducting his research, Drs. J. W. Carpenter, J. P. Feaster

and T. J. Cunha. The assistance of Drs. F. Martin and M. Koger in the

statistical analysis of the experimental data is sincerely appreciated. Grate-

ful acknowledgment is also extended to Dr. H. L. Chapman, Jr. for making

the animals available for this research.

Recognition is accorded to Mrs. Lynn Fisher for her assistance in the

collection of experimental data throughout the research. Credit also goes to

Mr. J. F. Jeter, Mrs. Bernice Godwin and Mrs. Sherry Bellino for their tech-

nical assistance.

The author is also thankful to the Government of Iraq for its financial

support during his graduate study.








The author is deeply indebted to his wife, Menal, for her encouragement

and patience throughout his graduate work.

The appreciation of the author is extended to Miss Ardeth Heinze for the

accurate and excellent typing of this dissertation.














TABLE OF CONTENTS


Page

ACKNOWLEDGMENTS ..... . . . . . . ii

LIST OF TABLES . . . . . . . . . viii

LIST OF APPENDIX TABLES ................ xii

LIST OF FIGURES ...... ...... .. . . xiv

INTRODUCTION . . . . . . . . . 1

REVIEW OF LITERATURE .. . .. . . . 7
The use of the Longissimus dorsi muscle in beef
tenderness investigations . . ... . . . 7
Post-mortem change in pH of beef muscle as
related to meat quality . . . . . . 9
The relationship between muscle pH and palatability
of pork . . . . . ... .. 11
The relationship between pH and water-holding capacity . 12
The relationship between water-holding capacity and
meat quality . . . . . . . .. 13
Post-mortem changes in water-holding capacity ...... 14
Post-mortem changes in beef tenderness. . . . .. 16
The relationship of muscle proteins to tenderness . . .. 18
Viscosity measurement and muscle protein studies . . .. 25
Ultraviolet absorbance and protein concentration. . . .. 26

EXPERIMENTAL PROCEDURE ............... 27
Animals used . . . . . . . . 27
Treatment, management and feeding of the animals. ... . 27
Slaughter procedure .......... . .... 28
Chilling, carcass data and aging . . . . . . 29
Temperature determinations . . . . . .. 30
pH determinations . . . . . ... 30
Scheme for steaks removed from the Longissimus dorsi
muscle for the different studies conducted . ... . 31
Cooking . . . . . . . . 33
Organoleptic panel and Warner-Bratzler shearing . . . 33
Water-holding capacity determinations . . . . .. 34








TABLE OF CONTENTS Continued


Page

Muscle proteins studies .. . ... .. . ..... 35
Preparation of samples. . . . . . . 35
Extraction and fractionation of buffer soluble,
buffer insoluble and water soluble proteins. ... . 36
Nitrogen analysis . . . . . . . ... 36
Preparation of potassium chloride
Potassium phosphate buffer. . . . . 36
Viscosity determinations . . . ... . . 39
Ultraviolet absorbance measurements . . . ... 41
Starch gel electrophoresis studies . . ... .. . 41
Preparation of muscle protein extracts for starch
gel electrophoresis analysis . . ... . . 42
Preparation of electrolyte buffers. . . ..... 42
Preparation of the starch gel . . . . .. 45
Application of samples and electrophoresis
conditions ........ .......... 45
Staining and washing procedures . . ... . . 46
System of sample analysis and data recording for
the starch gel electrophoresis . . . ... 47
Diagrammatic sketch for the gel .. ... . 47
Statistical analysis . . ... . . ... 49

RESULTS AND DISCUSSION ................. 50
The effect of feeding treatment on rate of gain,
in-transit shrink and slaughter characteristics . . .. 50
The effect of feeding treatment on carcass
characteristics . . . . . . . . 50
Post-mortem changes in the pH of the Longissimus dorsi . 53
The relationship between pH and tenderness. . . . ... 55
Temperature of the Longissimus dorsi during the
chilling period . . . . . . . . 60
The relationship between pH and rate of chilling of
the Longissimus dorsi ................ 62
The relationship between tenderness and rate of
chilling of the Longissimus dorsi post-mortem. ... . 64
Post-mortem changes in bound and free moisture of
the Longissimus dorsi muscle . . . . ... 65
The effect of pH and time post-mortem on free moisture
of the Longissimus dorsi muscle . . ... ..... 70
The effect of pH, initial steak temperature, time
post-mortem, free and bound moisture, cooking
time and cooking method on cooking loss . ..... 73








TABLE OF CONTENTS Continued


Page


Post-mortem changes in juiciness of the
Longissimus dorsi steaks .. .....
The relationship between juiciness and bound moisture,
cooking loss, cooking method and time post-mortem.
Post-mortem changes in flavor of the Longissimus
dorsi steaks . . . . . . . .
The effect of time post-mortem on the flavor of
the broiled and deep fat fried Longissimus
dorsi steaks . . . . . . . .
Post-mortem changes in tenderness of the
Longissimus dorsi steaks . . . . . .
The relationship between 1, 24, 48 and 192 hr
post-mortem tenderness . . . . . .
The relationship between nitrogen concentration in
water soluble proteins and ultraviolet
absorbance . . . . . . . .
The relationship between nitrogen concentration in
buffer soluble proteins and ultraviolet
absorbance . . . . . .
Post-mortem changes in solubility of the water
and buffer soluble and buffer insoluble proteins. .
The relationship between total tissue protein,
water soluble, buffer soluble and buffer
insoluble protein and tenderness . . . .
Post-mortem changes in viscosity of water
and buffer soluble protein extracts . . . .
The relationship between the viscosity of water
and buffer soluble proteins and tenderness . .
The electrophoretic separation of buffer and water
soluble proteins. . . . . . . .
Post-mortem changes in electrophoretic components
of the water and buffer soluble protein extracts .
The relationship between electrophoretic components
of the water and buffer soluble proteins and
tenderness . . . . . . . .


The relative significance of chemical and physical factors
influencing and/or associated with beef tenderness .
The relative significance of factors influencing
and/or associated with tenderness at 1 hr
post-mortem . . . . . . .
The relative significance of factors influencing
and/or associated with tenderness at 24 hr post-mortem


S . 84



S . 89

. . 90

. . 97



. . 108



. . 112

. . 114



. . 117

. . 122

. . 122

. . 125

. 128



. 131


134



134

139








TABLE OF CONTENTS Continued


Page

The relative significance of factors influencing
and/or associated with tenderness at 48 hr
post-mortem .. ... . . . . ... . 144
The relative significance of factors influencing
and/or associated with tenderness at 192 hr
post-mortem . . . ... . . . . . 148

SUMMARY AND CONCLUSIONS . .. . .. . .. . . 155

APPENDIX .......... ........ .. ... .. 162

REFERENCES . .... . . . . . . . 221

BIOGRAPHICAL SKETCH . .... .... ... ... .229














LIST OF TABLES


Table Page

1 Experimental design . . . . . . 27

2 Composition of ration .. . . . . . 28

3 Mean values and significance of difference among the four
treatments in feed lot gain, in-transit shrink and
slaughter characteristics . . . . . . 51

4 Mean values and significance of difference among the four
treatments in carcass qualities and indices of
meatiness . .... ............ 52

5 Simple correlation coefficients between pH values of the
L. dorsi at four post-mortem intervals ...... 55

6 Simple correlation coefficients between tenderness and
the pH of the L. dorsi and the rate of pH change
during post-mortem . . . . . . ... 56

7 Simple correlation coefficients between post-mortem
temperature and pH of the L. dorsi . . . ... 63

8 Simple correlation coefficients between tenderness and
rate of chilling of the L. dorsi post-mortem. ... . 66

9 Simple correlation coefficients between time post-mortem,
pH, free and bound moisture. . . . . ... 71

10 Coefficients for predicting free moisture from pH and
time post-mortem . . . . . . . 72

11 Simple correlation coefficients between cooking loss and
pH,. initial internal steak temperature, time post-
mortem, free and bound moisture, cooking time and
cooking method . ... ......... . 73

12 Coefficients for the most important factors influencing
and/or associated with cooking loss . . . ... 78
viii








LIST OF TABLES Continued


Table Page

13 Mean values of panel juiciness scores of the L. dorsi
steaks . . . . . . . 80

14 Analysis of variance for panel juiciness scores of the
L. dorsi steaks . . . .... .... . 81

15 Simple correlation coefficients between juiciness and
bound moisture, cooking loss, cooking method
and time post-mortem . . . . . ... 83

16 Coefficients for the most important factors influ-
encing and/or associated with juiciness of the
L. dorsi steaks . . . . . . ... 85

17 Mean values of panel flavor scores of the L. dorsi
steaks . . . . . . ... . 86

18 Analysis of variance for panel flavor scores of the
L. dorsi steaks . . . . ... . . . 87

19 Coefficients for predicting the flavor of broiled and
deep fat fried L. dorsi steaks from time post-
mortem . . . . . . . . 90

20 Mean values of W-B shear force values of the L. dorsi
steaks ... . . . . . . 91

21 Mean values of panel tenderness scores of the L. dorsi
steaks . . . . . . . . 92

22 Analysis of variance for W-B shear force values of
the L. dorsi steaks . . . . . ... 93

23 Analysis of variance for panel tenderness scores of
the L. dorsi steaks . . . . . . 94

24 Simple correlation coefficients between tenderness
values obtained by taste panel and W-B shear at
four post-mortem intervals . . . . ... 96

25 Simple correlation coefficients between 1, 24, 48 and
192 hr post-mortem tenderness . . . ... . 98








LIST OF TABLES Continued


Table Page

26 Coefficients in predicting tenderness value at 192 hr
post-mortem from tenderness values at 48, 24
and 1 hr . . . . . . . . 106

27 Simple correlation coefficients between nitrogen con-
centration in water soluble proteins and ultraviolet
absorbance at 260 and 280 mp .. . . . . 109

28 Coefficients in predicting nitrogen concentration in
water soluble proteins and ultraviolet absorbance
at 260 and 280 m ............... ....... 111

29 Simple correlation coefficients between nitrogen con-
centration in buffer soluble proteins and ultraviolet
absorbance at 260 and 280 my ........ . 113

30 Mean values of protein distribution of the L. dorsi of
20 animals ... ..... . . .. . . 115

31 Simple correlation coefficients between total, water
soluble, buffer soluble and buffer insoluble
protein and tenderness . . . .... . . 118

32 Mean values of viscosity of water and buffer soluble
protein extracts at four post-mortem intervals
(20 carcasses) ........... .. . .. .123

33 Simple correlation coefficients between the viscosity
of water and buffer soluble proteins and tenderness . 124

34 Mean values of number of electrophoretic bands
obtained from water and buffer soluble proteins
after starch gel electrophoresis (four post-mortem
intervals of 20 carcasses) . . . . . .129

35 Simple correlation coefficients between the number of
some electrophoretic bands of water and buffer
soluble proteins and tenderness . . . ... .132

36 The relationship between tenderness of the broiled
L. dorsi steak at 1 hr post-mortem and selected
chemical and physical measurements as shown by
simple correlations and step-wise regression .... .136








LIST OF TABLES Continued


Table Page

37 The relationship between tenderness of the deep fat fried
L. dorsi steak at 1 hr post-mortem and selected
chemical and physical measurements as shown by
simple correlations and step-wise regression .... .137

38 The relationship between tenderness of the broiled
L. dorsi steak at 24 hr post-mortem and selected
chemical and physical measurements as shown by
simple correlations and step-wise regression .. ... .140

39 The relationship between tenderness of the deep fat
fried L. dorsi steak at 24 hr post-mortem and
selected chemical and physical measurements
as shown by simple correlations and step-wise
regression ......... ....... 142

40 The relationship between tenderness of the broiled
L. dorsi steak at 48 hr post-mortem and selected
chemical and physical measurements as shown
by simple correlations and step-wise regression . . 145

41 The relationship between tenderness of the deep fat
fried L. dorsi steak at 48 hr post-mortem and
selected chemical and physical measurements as
shown by simple correlations and step-wise
regression ... .. .. ... . . 146

42 The relationship between tenderness of the broiled
L. dorsi steak at 192 hr post-mortem and
selected chemical and physical measurements as
shown by simple correlations and step-wise
regression .. . . .... . . . . 149

43 The relationship between tenderness of the deep fat fried
L. dorsi steak at 192 hr post-mortem and selected
chemical and physical measurements as shown by
simple correlations and step-wise regression . .. 151













LIST OF APPENDIX TABLES


Table Page

44 Description of scales used in evaluating carcass
characteristics. ........ . . . . 163

45 Description of scales used in evaluating palatability
characteristics .. . . . . . . 165

46 Individual feed lot performance, in-transit shrink
and carcass characteristics . ... . . . 166

47 Individual carcass grade data . . . . . .. 168

48 Post-mortem changes in temperature of L. dorsi
muscle of 36 animals . . . . . ... .170

49 Post-mortem changes in pH of L. dorsi muscle
of 36 animals ..... .. .. .... 176

50 Percentage of total, bound and free moisture in
raw steaks from L. dorsi muscle of 36
animals relative to post-mortem time . . ... 182

51 Cooking characteristics of L. dorsi steaks (broiled)
of 36 animals relative to post-mortem time . . .. .184

52 Cooking characteristics of L. dorsi steaks
(deep fat fried) of 36 animals relative to
post-mortem time. . . . . . 188

53 Values for W-B shear and palatability characteristics
of L. dorsi steaks (broiled) of 36 animals
relative to post-mortem time . .. .. 192

54 Values for W-B shear and palatability characteristics
of L. dorsi steaks (deep fat fried) of 36 animals
relative to post-mortem time .. . . .. .196








LIST OF APPENDIX TABLES Continued


Table

55 Tenderness rank of broiled L. dorsi steaks as
evaluated by W-B shear at 1 hr post-mortem

56 Tenderness rank of broiled L. dorsi steaks as
evaluated by W-B shear at 24 hr post-mortem

57 Tenderness rank of broiled L. dorsi steaks as
evaluated by W-B shear at 48 hr post-mortem

58 Tenderness rank of broiled L. dorsi steaks as
evaluated by W-B shear at 192 hr post-mortem

59 Nitrogen concentration and ultraviolet absorbance of
water soluble and buffer soluble protein ex-
tracts . . . . . . .

60 Nitrogen concentration in water soluble, buffer
soluble and buffer insoluble fractions of the
L. dorsi of 20 animals . . . . .

61 Viscosity of water and buffer soluble protein
extracts of the L. dorsi muscle of 20 animals

62 Number of electrophoretic bands obtained by starch
gel electrophoresis of water and buffer soluble
protein extracts of the L. dorsi of 20 animals


Page


200


. . 202


. . 204


. . 206




. 208




. . 212


. . 216


xiii














LIST OF FIGURES


Figure Page

1 Schedule of removing L. dorsi steaks for studies
conducted.................. ... 32

2 Scheme for the extraction of buffer soluble and
buffer insoluble proteins and other quantita-
tive and qualitative analysis conducted ...... 37

3 Scheme for the extraction of water soluble proteins
and other quantitative and qualitative analysis
conducted ............. . . . . 338

4 Scheme for the extraction of buffer soluble proteins
for starch gel electrophoresis analysis ...... 43

5 Scheme for the extraction of water soluble proteins
for starch gel electrophoresis analysis. . . .. 44

6 Starch gel electrophoresis data sheet . . .... . 48

7 Post-mortem changes in pH of L. dorsi muscle as
presented by average values with standard
deviations. .................. 54

8 Post-mortem changes in temperature of L. dorsi muscle
as presented by average values with standard
deviations .................. .. 61

9 Post-mortem changes in total, bound and free moisture
of raw steaks from L. dorsi muscle. . . . 68

10 Post-mortem changes in cooking losses and pH . . .. 74

11 Post-mortem changes in cooking losses and free
moisture . . . . . . .. 75

12 Post-mortem changes in cooking losses and cooking
tim e . . . . . . . . 76








LIST OF FIGURES Continued


Page


Post-mortem changes in juiciness of L. dorsi steaks
cooked by broiling and deep fat frying . .


14 Post-mortem changes in flavor of L. dorsi steaks
cooked by broiling and deep fat frying . . . . 88

15 Post-mortem changes in tenderness of L. Dorsi
steaks cooked by broiling and deep fat frying ..... 95

16 Average tenderness changes for the "tough" and
"tender" groups based on 1 hr shear ranking . .101

17 Average tenderness changes for the "tough" and
"tender" groups based on 24 hr shear ranking .... .102

18 Average tenderness changes for the "tough" and
"tender" groups based on 48 hr shear ranking .... .103

19 Average tenderness changes for the "tough" and
"tender" groups based on 192 hr shear ranking .... .104

20 Starch-gel electrophoretic patterns of buffer
soluble proteins. . . . . . . 126

21 Starch-gel electrophoretic patterns of water
soluble proteins. . . . . . . . 127


Figure

13














INTRODUCTION


Tenderness is the most important palatability characteristic of beef.

Consumers more frequently express disappointment in lack of tenderness than

either lack of flavor or lack of juiciness. In general, beef produced today may

be considered highly palatable as seen in the decided trend over the past decade

of increasing per capital consumption. Beef producers and processors, obvi-

ously, are providing beef that meets the consumers' demand. Yet, problems

of unexpected toughness in some of the higher priced cuts and the variability in

tenderness between identical cuts from different carcasses of the same grade

still exist. Differences in tenderness between the different muscles of the beef

carcass are real, as established scientifically, and these differences are of

importance. In practice, tender muscles go into steaks and less tender muscles

go into roasts, stew meat and ground meats. A knowledge of why muscles with-

in a carcass vary in tenderness and why the same muscles of different carcasses

differ in tenderness could lead, through further research, to an even greater

acceptability of beef. Perhaps beef could be produced more economically, for

now most of our beef is fed out in the feed lot, a rather expensive part of the over-

all production, with the assumption that increased grade will substantially improve

over-all palatability. This broad assumption has brought about the wide-spread

practice of finishing cattle by grain feeding. It is obvious that before such an

assumption should have had such an influence over production practices, the








validity of that assumption should first have been well established by scientific

research. There is a striking lack of research work reported in the literature

demonstrating that beef palatability is appreciably enhanced by feeding. The

literature that is available, however, does indicate that palatability is some-

what improved by the feeding of concentrate rations prior to the slaughter.

Tenderness of beef has been attributed to the breeding of the animal,

feeding, age at time of slaughter and sex. Carcass grade or grade character-

istics such as maturity, marbling, color and texture have been related to

tenderness. The quantity and character of connective tissue as well as muscle

fiber diameter has been associated with tenderness. Method of freezing and

method of cooking have been shown to influence tenderness. The study of all

of these factors that influence or are associated with tenderness has been help-

ful in the production and marketing of beef possessing a higher degree of consumer

acceptability. Yet research of this nature has failed to show clearly the funda-

mental nature of tenderness or toughness in beef. Applied type of research has

opened several promising avenues of research that might lead to a better under-

standing of many complexities of tenderness. There is a lack of a fundamental

knowledge to explain why beef may be tender or tough. We know how to obtain

tender beef by different production and process practices, but we do not fully

understand why it is tender.

It has not been possible until the last decade to explore the ultramolecular

structure of the muscle cell. Now electron micrographs may be used to locate

the myosin and actin filaments on the sarcomers thus allowing identification and

definition of some of the many physical and chemical changes that take place in








the muscle during the periods of contraction and relaxation. Many studies of

the histological and biophysicochemical properties of muscle have been con-

ducted in the past; most of these studies, however, were conducted on laboratory

animals with emphasis mainly toward muscular contraction mechanisms.

Tenderness studies have been confined for the most part to the period

following the completion of rigor mortis. Studies of the change in the physical

and chemical properties of meat during the pre-rigor, rigor and immediately

post-rigor periods have been somewhat lacking.

Muscle tissue contains a large number of proteins that differ in structure,

chemical and physical properties and function. There is a present lack of entire-

ly acceptable standard techniques for the isolation, purification and characteri-

zation of these proteins. It is not surprising, therefore, that the exact nature

of muscle protein changes post-mortem is not clear.

Wierbicki et al. (1954) suggested that initial toughness of meat after

slaughter was due to the formation of actomyosin and that the subsequent tender-

ization was the result of actomyosin dissociation. Wierbicki et al. (1956) later

reported data that failed to support their first suggestion about post-mortem.

actomyosin dissociation, although the amount of actomyosin seemed to be related

to tenderness. Partmann (1963) reported that the actomyosin complex formed

during rigor development became dissociated or, at least, may become disso-

ciated easily in aged meat, and that tenderness changes in the aging period were

correlated to this process. Furthermore, the same author demonstrated the close

correlation in the interactions between adenosine triphosphate (ATP), actin

and myosin that took place in living muscle during contraction and extension








and during post-mortem muscle changes. The Partmann (1963) report and the

new molecular theory of muscle contraction developed by Davies (1963) who

described the anatomical process of contraction by the movement of the thin

filaments of actin along the channels between the thick filaments of myosin

without overall changes in length allows theorization of why meat should be

tough during rigor mortis.

The recent research on extraction and fractionation of muscle proteins

according to solubility behavior has been promising. Relationship between the

solubility of some intracellular protein components and tenderness and water-

holding capacity has been demonstrated by Hill (1962) and Hegarty etal. (1963).

Also, the alterations of protein solubility by post-mortem time and conditions

have been illustrated by Sayre and Briskey (1963), Kronman and Winterbottom

(1960), Weinberg and Rose (1960), Goll et al. (1964) and Khan and Van den Berg

(1964).

Starch gel electrophoresis was developed by Smithies (1955) for serum

protein studies. This technique showed a greater resolution power than any of

the previously used electrophoretic methods, and serum separation was quite

successful with this technique. The use of this technique in muscle protein

studies has been limited, even though the muscle protein patterns obtained on

the starch gel by Hartshorne and Perry (1962), Scopes and Lawrie (1963) and

Neelin and Rose (1964) were greater in number and clearer in appearance com-

pared to patterns obtained by previously used methods.

Considering the above factors and others, the present author concluded

that a comprehensive study on tenderness, protein, water-holding capacity,








temperature and pH changes during post-mortem might yield information on

the nature of post-mortem tenderness change and, at the same time, might

give some insight as to why individual carcasses differ in tenderness.

The primary objectives of this study were, therefore, as follows:

1. To study post-mortem changes in tenderness and other organolep-

tic characteristics, using the Longissimus dorsi muscle of beef carcasses.

2. To study post-mortem changes in solubility of some of the struc-

tural and functional muscle protein components and to correlate the quantities

of these components to tenderness.

3. To study qualitatively, by using starch gel electrophoresis, the

nature of the different muscle protein extracts, to detect any post-mortem

changes, and to relate these changes to changes in tenderness and protein

solubility.

4. To study post-mortem changes in the water-holding capacity of

muscle and to relate such change to palatability characteristics and to pH.

5. To study post-mortem changes in temperature as an index of rate

and extent of rigor development and to correlate temperature changes with

tenderness.

6. To study post-mortem changes in pH as an index of rate and extent

of glycolysis, ATP decomposition and rigor development. Also, to correlate

pH readings at different post-mortem intervals and rate of pH drop with tender-

ness and other factors studied.

7. To study post-mortem changes in cooking loss and to correlate such

change to muscle tenderness, juiciness, pH and water-holding capacity.




6



8. To compare the effect of two methods of cooking (broiling and

deep fat frying) on tenderness, flavor, juiciness and other factors studied.













REVIEW OF LITERATURE


The use of the Longissimus dorsi muscle in beef tenderness investigations

The muscles of the beef carcass vary widely in tenderness and in many

physical and chemical characteristics. For this reason in a fundamental study

it is most desirable to make comparisons between protein components of

muscle and between animals on a within-muscle basis.

The L. dorsi is the largest muscle of the beef carcass. It is the major

muscle found in the high-priced retail cuts of the loin, rib and chuck. Because

of its size in the beef carcass, the L. dorsi can provide a relatively large quan-

tity of muscle for use in experimental work. Data obtained on the L. dorsi should

be most meaningful in a practical sense by having been obtained on the largest

and one of the most expensive muscles of the carcass.

Satorius and Child (1938) determined the tenderness variability of three

different rib cuts from 13 beef carcasses that had been aged 12 days at 2 to 30C.

Their results showed no significant difference in tenderness between the 7 8th,

9 10th and 11 12th rib roasts when only the physical properties of tenderness

were measured. Ramsbottom et al. (1945) reported that the L. dorsi muscle was

fairly uniform in tenderness, except at the extremities, when compared with

other beef muscles. Blakeslee and Miller (1948) used 18 short loin roasts to

determine the tenderness of several different grades of beef. Their data demon-

strated that the short loins were less tender at the rib end than at the porterhouse

end.








Hiner and Hankins (1950) studied the tenderness of nine muscles,

including the L. dorsi, from 52 beef animals varying widely in age and sex.

Differences in tenderness between samples taken from the 8th rib, shortloin

and loin end areas were of no statistical significance from one of the groups

studied that consisted of 25 steers averaging 900 pounds in live weight.

Ginger (1957) reported that the L. dorsi muscle presented fewer prob-

lems of design for tenderness studies due to its length and width when compared

to the Semimembranosus muscle. In later work, Ginger and Weir (1958) studied

tenderness variability of the Semimembranosus, Semitendinosus and Biceps

femoris using the taste panel and on the Biceps femoris and Semimembranosus

using the Warner-Bratzler (W-B) shear measurement. All muscles varied

significantly in tenderness throughout their length.

Two muscles, the L. dorsi and the Semitendinosus, from the left side

of 12 Hereford heifer carcasses were studied by Mjoseth (1962). Variations

in tenderness, gross chemical composition, pH and cooking loss due to carcass

and position effect were investigated. The investigation revealed that the effect

of position variation in the Semitendinosus muscle was much greater for impor-

tant variables such as shear force, per cent fat and per cent moisture than was

the case in the L. dorsi muscle. In the above study, the amount of variation

accounted for by carcass differences in the L. dorsi muscle was greater for all

variables except cooking loss than that accounted for by steak position.

The histological study of 50 of the principal beef muscles conducted by

Strandine et al. (1949) showed that the cross sectional area of muscles taken any-

where in the muscle except at the extreme ends (origin and insertion) were rather







uniform and presented a regular pattern or arrangement of muscle bundles

and connective tissue. Hiner et al. (1953) found no significant difference

among fiber size at the third rib, eighth rib, and short loin areas of the L.

dorsi. Swanson et al. (1965), however, found a significant difference in cross-

sectional area of muscle fibers along the L. dorsi muscle. These researchers

reported that the smallest fibers were found over the 12th rib, and the fibers

increased in size both anteriorly and posteriorly to this region. The largest

fibers were found between the 13th rib and first lumbar vertebra in the short

loin region.

Because the short loin is generally considered a more tender cut,

Swanson's et al. (1965) work provided support for the observation by Tuma et

al. (1962) that no relationship between shear force and fiber diameter existed

when the effect of animal age was removed.


Post-mortem change in pH of beef muscle as related to meat quality

The hydrogen ion concentration of meat has a direct or indirect relation-

ship with the commercial value of beef carcasses by influencing color of lean,

bacterial growth, organoleptic properties and processing characteristics.

Post-mortem changes in pH were studied by Ramsbottom and Strandine

(1949) who found that the muscle pH of six utility grade cattle dropped from about

6.4 at 2 hr after the dressing operation to about 5.5 one day later. Post-mortem

changes in pH of Semitendinosus and Biceps femoris steaks removed from two

prime, two good and two commercial carcasses were studied by Paul et al. (1952).

The authors found that the pH of the raw meat decreased with storage, rapidly for

the first 24 hr and more slowly thereafter. The pH values for the six carcasses







were quite close and the averages were: zero hr, pH 6.68; 5 hr, pH 6.50;

12 hr, pH 6.23; 24 hr, pH 5.90; 48 53 hr, pH 5.55; and 144 149 hr, pH

5.48. Wierbicki et al. (1954) reported that the pH of muscle dropped from

7. 3 7.4 in the live animal to 5.4 5.6 in the carcass within 48 hr after

slaughter due to muscle metabolism changing from an aerobic to an anaerobic

state. The authors found that the drop in pH was concurrent with the disap-

pearance of ATP and the appearance of lactic acid and inorganic phosphate.

Although no direct relationship was shown, the authors concluded that pH and

increasing tenderness may be indirectly related. The authors doubted if pH

was the primary factor causing increases in tenderness with post-mortem

aging. This observation was in agreement with the work of Husaini et al.

(1950 b), who conducted a study using 20 animals and failed to show a significant

correlation between pH and tenderness of broiled short loin steaks, which were

aged at 3.50C for 14 days.

Hedrick et al. (1959) reported little or no change in tenderness of beef

after ante-mortem treatments that raised the pH sufficiently to produce dark

cutting beef. High ultimate pH, taken after 24 hr of chilling, and dark meat

were the results of continuous excitement over a period of 24 hr by periodic

prodding with an electric "hot shot."

In a continuation of tests begun by Paul et al. (1952), Paul and Bratzler

(1955) used the L. dorsi muscle of the previously mentioned carcasses plus

L. dorsi muscles from two commercial grade cows. The pH was determined

on every fourth steak of each muscle. Animal differences in pH values were

the only significant values obtained. Average pH values for the six groups








ranged from a high of 5.80 to a low of 5.22. The pH differences due to storage,

handling and position on the steak were not significant. The authors felt that

two days of cold storage were adequate to complete the initial drop in pH nor-

mally observed in beef after slaughter, while nine days were not long enough to

cause the slight rise in pH due to increased storage periods observed by

Wierbicki et al. (1954).

Briskey (1959) indicated that the initial and ultimate pH values of muscle

were critical in determining the time course of rigor; therefore, the factors

which predetermine these pH values are of great importance. The author stated

that the initial pH was a result of the severity of the death struggle, while the

ultimate pH was an indicator of the animal's state of fatigue and level of feeding.

He also indicated that the initial pH was not correlated with the ultimate pH.

This observation was in line with the finding of Marsh (1954) who reported no

relationship between initial and ultimate pH values. The author found that even

those muscles with an initial pH of 6.6 6.8 apparently contained sufficient

glycogen to attain low ultimate pH values. Kronman and Winterbottom (1960)

stated that the ultimate pH value of muscle was determined chiefly by the rate of

lactic acid production, the buffering capacity of the muscle and the rate of inac-

tivation of the significant glycolytic enzymes.


The relationship between muscle pH and palatability of pork

The relationship between muscle pH, organoleptic and processing

characteristics of pork has been investigated.

Judge et al. (1960) reported on the chemical analyses and sensory

scoring data obtained from 54 pork loins. Dark, firm muscle was higher in








pH and lower in free water than was light, soft muscle. Water-holding capacity

and pH were positively related. A highly significant correlation coefficient of

,66 was found between pH and tenderness.

Kauffman et al. (1961) used 439 pigs of varying sex, weight and chrono-

logical age to investigate the relationships of chilled, aged and cooked pork mus-

cle acidity with palatability and economical traits. Results indicated that

increased muscle acidity was significantly indicative of a higher percentage of

expressible juice and was characteristic of pale, soft tissue. Darker, drier,

firmer muscles exhibited relatively high pH values, shrank less (P <.01) during

curing and cooking and were more juicy and tender (P<. 01) when compared to

pale, soft muscle.

Lewis et al. (1963) utilized the L. dorsi, Psoas major and Quadriceps

femoris muscles of 12 hogs to study the relationships of certain chemical and

physical measurements to organoleptic characteristics of pork. The authors

found that lactic acid concentration was negatively correlated with tenderness,

but not as highly correlated with tenderness as was pH or expressible water.

They also found a significant correlation between pH and expressible water.


The relationship between pH and water-holding capacity

The relationship between pH and water-holding capacity has been demon-

strated by several investigators. Howard and Lawrie (1956) and Bouton et al.

(1957) found that the water-holding capacity or the susceptibility to "drip" after

freezing and thawing, as well as flavor and tenderness of beef muscle were all

pH-related.







Eight muscles from two cows, a bull, a heifer and a steer were util-

ized by Swift and Berman (1959). Highly significant correlations of-. 89 and

.93 between pH and protein content and pH and water retention, respectively,

were obtained. The authors demonstrated that the higher the ultimate pH

attained by the muscle, the more water the muscle proteins held or "bound."

Penny et al. (1963) demonstrated that injection of adrenaline in beef and

rabbits before slaughter improved rehydration, juiciness and tenderness of the

rehydrated products. The effect of adrenaline was ascribed to a higher ultimate

pH in the meat as a result of glycogen depletion prior to slaughter.

The influence of the pH of fresh meat before heating on the change in

water-holding capacity after cooking has been reported by Hamm and Deatherage

(1960). Raw L. dorsi muscle from utility cows was adjusted to various pH

values and heated to 800C. The authors found that the pH of meat had a marked

influence on water-holding capacity (measured by the press method) before and

also after heating. The lowest water-holding capacity of cooked meat was that

of meat adjusted to pH 5.0 before heating.


The relationship between water-holding capacity and meat quality

The biochemistry of meat hydration and the relationship of the water-

holding capacity to the different characteristics of meat quality has been reviewed

by Hamm (1960).

Early work by Satorius and Child (1938) and Hall et al. (1944) indicated

no relation between subjective scores for meat palatability and the amount of

expressible juice. Gaddis et al. (1950), however, found a low but significant

correlation between the amount of press fluid and the juiciness scores in 97 beef








ribs; in 115 lambs and sheep, the authors found no significant correlation. The

results obtained by Urbin et al. (1962) indicated a positive relationship between

free moisture values and tenderness of the loin-eye in pork.

The effect of heating or cooking on the eating quality of beef has been

discussed by Hamm (1960), who suggested that the amount of bound water rather

than the amount of expressible juice may be related to juiciness of meat. Ritchey

and Hostetler (1964), however, reported that correlations between subjective

scores for juiciness and either free or bound water were low and varied irreg-

ularly with the different final internal temperatures used in cooking the L. dorsi

and Biceps femoris steaks of beef animals. Yet Tannor et al. (1943) and Hardy

and Noble (1945) reported a significant correlation between subjective juiciness

scores and objective tests for expressible fluid or juice. This finding was in

contrast to the report of Ritchey (1965) who failed to find any significant corre-

lations between subjective scores for eating quality in two beef muscles and

either bound or free water.


Post-mortem changes in water-holding capacity

Post-mortem change in water-holding capacity is a problem of practical

importance in meat processing operations; consequently, the literature provides

several interesting reports on the subject.

Wierbicki and Deatherage (1958), Hashimoto et al. (1959), and Hamm

(1963) reported that meat hydration dropped very markedly within a few hours

after slaughter, reached a minimum in 24 to 48 hr, and increased slowly there-

after. However, the hydration of the aged meat was never as high as that found

1 to 3 hr after slaughtering.








Hamm (1960) reported that the decrease of water-holding capacity after

slaughtering was due partly to the drop of pH caused by glycolytic formation of

lactic acid. The author reported an experiment in which the press method was

used to measure the water-holding capacity of muscle homogenate adjusted to

different pH values by adding acid or base; minimum hydration occurred around

pH 5.0. Marsh (1952 a,b), however, emphasized the effect of ATP cleavage

post-mortem on the decrease in water-holding capacity of muscle. The author's

emphasis on ATP water-holding capacity theory was based on observations with

rabbit and whale muscle, in which the drop of pH alone was not sufficient explan-

ation for the post-mortem drop in water-holding capacity. This observation was

in agreement with Hamm (1963) who reported that the high water-holding capacity

of meat immediately after slaughter was largely the result of the presence of

ATP. The latter author also stated that two-thirds of the fall in water-holding

capacity in beef post-mortem was caused by the breakdown of ATP and one-third

caused by the drop in pH as a result of lactic acid formation.

The phenomenon of increased water-holding capacity after "aging" was

discussed by Hamm (1960) who reported that the slight rise in pH after "aging"

was not the entire explanation. The author reported that only 23 33% of the

total increase of water-holding capacity of beef muscle after 10 days of aging

was due to the increased pH. The author, hence, concluded that other biochem-

ical changes were responsible for the hydration effect of aging. These biochemical

changes were explained by the author to be the result of increased net charge of

protein due to cleavage of stable (nonelectrostatic) cross linkages or due to

proteolytic influences (Hamm, 1963).








Post-mortem changes in beef tenderness

The progressive improvement of tenderness during aging is well estab-

lished in the literature but there is little information available on the initial

tenderness of beef at the time of slaughter and the change in tenderness that occurs

during rigor mortis.

Ramsbottom and Strandine (1949) used the L. dorsi muscle from three

good carcasses and three utility carcasses to study tenderness changes at the

following post-mortem intervals: 2, 5, 8, 11 and 14 hr and also at 1, 2, 3, 6,

9 and 12 days. At the above mentioned intervals, steaks one inch in thickness

were removed from the carcass, cooked and evaluated subjectively and objectively

for tenderness. The authors found that the L. dorsi steaks were more tender at

two hr after slaughter than at any time during the next two days. Steaks removed

from the L. dorsi muscle of two carcasses at 3, 6, 9 and 12 days post-mortem

were more tender than steaks cut from the loins of two other unchilled carcasses,

then held for comparable time post-mortem. Tenderness differences between

muscle left intact in the carcass and muscle removed from the carcass immediately

post-mortem lessened with aging time and after 12 days post-mortem, muscles

left attached to the skeleton were more tender than muscles removed and then

held in the cooler.

Recently, in an effort to relate changes in protein solubility to differences

in tenderness during post-mortem, Goll et al. (1964) used the Semitendinosus

muscle from 15 steers which ranged from 16 to 19 months in age. Nine different

sire groups were represented, and animals from the same sire group were

sampled at different times post-mortem. The first sample was removed after








15 20 min post-mortem, and others were removed at 6, 12, 24, 72 and 312 hr

post-mortem. The sampling technique used by the authors was as follows: The

Semitendinosus muscle was taken from the left side of each carcass and stored

at 4C until further sampling. At an appropriate post-mortem time for each

animal, the Semitendinosus muscle was excised from the right side of the carcass.

Thus, the left Semitendinosus muscle of each animal had two steaks removed for

W-B shear tests of tenderness and two samples taken for measurements of protein

solubility. The corresponding muscle from the right side was excised from the

carcass at a certain time post-mortem and had one steak removed for a W-B shear

test of tenderness and one sample taken for measurements of protein solubility.

Here, the authors found that muscles left attached to the skeleton were least ten-

der during the first 12 hr of post-mortem aging and then gradually increased in

tenderness. However, even after 312 hr aging, muscles excised from the skeleton

immediately post-mortem were still less tender than muscles that remained with

the carcass. This observation was in agreement with that reported by Paul et al.

(1952), who found that Semitendinosus steaks decreased in tenderness during the

first 24 hr post-mortem and then returned approximately to their original tender-

ness after 144 149 hr. The muscles used in Paul's study had been removed from

the skeleton approximately 1 hr post-mortem.

Change in tenderness during aging of beef has been observed by Deatherage

and Harsham (1947), who determined tenderness in the loin of 14 beef carcasses at

3, 6, 10, 17, 24, 31, 38 and 41 days post-mortem. The authors found that not all

the carcasses increased in tenderness with age throughout the period of observation;

i.e. tenderness did not increase smoothly with age.








Husaini et al. (1950 a) found that the meat of 20 carcasses was more

tender at 15 days post-mortem than at 3 days. The correlation between initial

and final tenderness values was statistically significant but lacked magnitude.


The relationship of muscle proteins to tenderness

Recent interest in the relationship of certain muscle proteins to tender-

ness has been based on work reported during the late forties and middle fifties.

Post-mortem changes in tenderness were observed by Deatherage and Harsham

(1947), Ramsbottom and Strandine (1949) and Paul et al. (1952). Husaini et al.

(1950 a,b) presented evidence against the previously held view that increased

tenderness with aging was due to autolysis by the group of enzymes collectively

known as kathepsin. Husaini et al. (1950 a,b) failed to find any increase in the

non-protein or proteose-peptone nitrogen fractions during aging of beef.

Wierbicki et al. (1954) observed that increased tenderness with aging was not com-

bined with significant changes in the nature of connective tissue as measured by

per cent of alkali-insoluble protein and per cent of hydroxyproline. The authors

stated that if change in connective tissue was responsible for the increased tender-

ness during the aging period, then, one should expect connective tissue to break

down during the aging period and at least some solubility changes in connective

tissues should take place. This observation led the authors to suggest two lines of

thought about tenderness changes: 1) initial toughness of meat at slaughter was

due to the formation of actomyosin complex from actin and myosin and that subse-

quent tenderization was the results of actomyosin dissociation and 2) subsequent

tenderization involved, rather than extensive dissociation of actomyosin, only a








slight dissociation; this was coupled with, or was brought about by, a redistribu-

tion of the ions within the muscle, thus causing increased hydration and tenderness.

Later, Wierbicki et al. (1956) reported again that toughness of meat during the on-

set of rigor mortis was due to the formation of actomyosin; however, actomyosin

was not dissociated during post-mortem aging, although the amount of actomyosin

seemed to be related to tenderness. The authors therefore suggested that post-

mortem tenderization was due to certain ion-protein or protein-protein interactions

rather than dissociation of actomyosin.

The insignificant effect of proteolysis in improving tenderness during

aging of beef for 16 days was demonstrated by Locker (1960 b). The author found a

slight decline in both non-protein nitrogen and free amino acid values during rigor,

followed by a slow rise to values above the original (at 1 hr post-mortem) during

aging. The extremely small rise in values after aging led the author to conclude

that proteolysis was most unlikely to be of any importance in the normal tender-

ization period. This report, however, was in contrast to the finding of Van den Berg

et al. (1963) with chicken meat aged at 0 C. Here, the authors observed an appre-

ciable increase in the amount of amino acid nitrogen, as determined by the

ninhydrin method, during aging. This report was in agreement with the study of

Ginger et al. (1954), who reported that aging beef rib cuts for two weeks at 35F

caused a slight increase in the amount of free arginine, leucine and tyrosine con-

tent, as well as in the non-protein nitrogen fraction of the aged meat. The increase

noted was interpreted by the authors to be due to proteolytic enzyme activity.

Wang et al. (1957) studied the mode of action of twelve enzyme prepara-

tions on the structural organization of beef and the effect of the enzymes on








tenderness. The authors found that enzymic action on the muscle fibers began

with disintegration of the sarcolema and nuclei and ended in complete disappear-

ance of the cross-striations. The authors confirmed the early suggestion of

Szent-Gyorgyi (1951) that morphological changes in the myofibrils were the result

of chemical modification of the actomyosin molecule.

The more recent approach to a better understanding of the nature of meat

tenderness has been in the fractionation of muscle cell proteins and the investi-

gation of the quantitative and/or qualitative relationship between the different

fractions and tenderness.

Post-mortem changes in the water soluble proteins during aging and

freezing were studied by Kronman and Winterbottom (1960). Eight muscles from

four animals (one cow and three steers) were used. It was found that aging and

freezing of beef muscle for 7 days and 35 days, respectively, rendered protein

less soluble in water compared to that of 3 hr post-mortem sample. They reported

that the 10 to 30% decrease in protein solubility observed during aging and freezing

was due to protein denaturation. The authors also analyzed the fresh, aged and

frozen protein extracts by boundary electrophoresis and ultracentrifuge and re-

ported that the patterns obtained by those methods of analysis indicated that some

protein components were lost during aging and freezing. Hamm and Deatherage

(1960), however, reported that quick freezing and thawing resulted in no consider-

able denaturation of muscle protein.

Weinberg and Rose (1960) investigated post-mortem changes in protein

extractability of chicken breast muscle. Pectoralis muscle of chickens were

extracted with phosphate buffer (pH 7.5) in 0.4M KC1 (Total ionic strength M= 0.55),







and the extracts were fractionated by dilution to lower ionic concentrations. It

was reported that the amount of nitrogen extracted as "actomyosin" (protein pre-

cipitated during dilution of the extracts to ionic strength of 0. 225) was increased

post-rigor (24 hr post-mortem) compared to pre-rigor extracts (within 30 min

after death). However, the amount of nitrogen extracted as myosinn" (protein

precipitated during dilution of the extracts from an ionic strength of 0.225 to

0. 05) was less post-rigor than in pre-rigor extracts. Although actin was not ex-

tracted as such in the author's experiment, the data obtained indicated that more

actin was extracted from post-rigor meat and that actin was combined with

myosinn" in the extract. This observation led the authors to suggest that tender-

ization was not merely random autolysis but resulted from a specific cleavage of

an actin association responsible for the maintenance of the muscle matrix.

Hill (1962) examined the distribution of nitrogen within characteristically

tough (Semitendinosus and tender (L. dorsi) muscles of different species (cattle,

lambs and pigs). The components examined quantitatively for nitrogen content

were sarcoplasmic, myofibril, stroma (by difference) and non-protein soluble

nitrogen. The author found that the stroma nitrogen and myofibril nitrogen

(expressed as per cent of fat-free total tissue nitrogen) were highest in beef mus-

cles and lowest in pig and lamb muscles. Also, on a stroma nitrogen-free basis,

beef Semitendinosus was higher in myofibril nitrogen and lower in sarcoplasmic

nitrogen content compared to the L. dorsi muscle. These observations led the

author to suggest that the amounts of these protein fractions were associated with

tenderness.

In an investigation of protein solubility as influenced by physiological








and W-B shear values at the five different post-mortem times studied for muscle

removed immediately after death, muscle left attached to the skeleton and muscle

excised from the skeleton. Goll et al. (1964) found no pattern of relationship

between protein solubility and tenderness. However, the amount of sarcoplasmic

protein extracted at any of the post-mortem times was less than the amount ex-

tracted immediately after death, and this was alos true of the myofibrillar protein

for muscles left attached to the skeleton.

The solubility of different protein extracts from breast and leg muscle of

chicken was examined at 30 min and at 2, 4, 24 and 48 hr post-mortem by Khan

and Van den Berg (1964). In this study, total extracted nitrogen (soluble in KC1-

borate or KC1-phosphate buffer, ionic strength = 1 and pH 7.4) was fractionated

into myofibrillar proteins, soluble at P = 0.5 and insoluble at p = 0.08, and

sarcoplasmic proteins, soluble at = 0.08. The authors found that the buffer-

extracted nitrogen rapidly decreased during the onset of rigor (2 and 4 hr after

death) and gradually increased to a maximum value during post-rigor aging (48 hr).

The authors stated that these changes in nitrogen extractability were mainly a re-

sult of changes in the solubility of myofibrillar proteins which showed the same

patterns of post-mortem change. Quantitative changes in stroma and sarcoplasmic

fractions were small. The authors concluded that post-rigor tenderization was the

result of the weakening or the breakdown of some bonds which bind myofibrils to

the matrix of the muscle.

Electrophoretic separation of rabbit muscle proteins has been conducted by

Bate-Smith (1940), Jacob (1947) and Amberson et al.(1949). The identification of

new fibrous protein in skeletal muscle by the use of electrophoresis was also







condition in the muscle, Sayre and Briskey (1963) used the L. dorsi from 15

market-weight pigs. The authors found that muscle protein solubility was grossly

altered by the conditions of both temperature and pH which existed at the onset of

rigor mortis or during the first few hours after death. Sarcoplasmic protein solu-

bility at 24 hr was decreased to 55% of that found at 0 hr in muscle groups exhibiting

high temperature and low pH (pH 5.3 5.6, temperature >350C) at the onset of

rigor mortis. Conversely, only a 17% reduction of sarcoplasmic protein solu-

bility was noted in groups with high pH (pH 6.04) at onset of rigor mortis.

Myofibrillar protein solubility ranged from no reduction during the first 24 hr after

death, when pH remained high at onset, to 75% reduction in solubility in muscle

with low pH and high temperature at the onset of rigor mortis.

The relationship of intracellular protein characteristics to beef muscle

tenderness was reported by Hegarty et al. (1963). In this study the carcasses of

20 yearling bulls were aged 7 days, at which time steaks from the L. dorsi were

removed; some were used for tenderness evaluation, and others were held in the

frozen state for a period of approximately one month for protein fractionation.

The authors reported that the ratio of sarcoplasmic nitrogen to total fibrillar nitro-

gen was correlated with tenderness ( r =-. 43 for shear and r = 41 for panel). A

higher correlation coefficient between soluble fibrillar nitrogen/total fibrillar

nitrogen ratio and tenderness ( r =-.69 for shear and r = .59 for panel) was reported.

Also, a correlation of 49, significant at the 5% level, between water-holding

capacity and tenderness as measured by the W-B shear technique was found. This

finding was in contradiction with the results of Goll et al. (1964) who found insignif-

icant correlations between protein solubility (sarcoplasmic and myofibrillar)








Chromatography also has been used recently for muscle protein investi-

gations. Fujimaki and Deatherage (1964) found that the sarcoplasmic proteins

(proteins extracted with water) of beef muscle showed at least 14 fractions

immediately after slaughter (about 1 hr post-mortem) when fractionated chro-

matographically on ion-exchange cellulose. The numbers and levels of eluted

peaks in the effluent diagram decreased with aging (7 days at 1 30C) of muscle

and freeze-drying of sarcoplasma. The authors concluded that these decreases

were due to denaturation. When two beef animals and/or two muscles (L. dorsi

and Semimembranosus) were compared, the authors found that quantitative differ-

ences appeared even though the qualitative similarities were quite clear in the

chromatograms.


Viscosity measurement and muscle protein studies

Brey (1958) defined viscosity of a fluid as a measure of the resistance of

the fluid to flow. The methodology, techniques and apparatus used are discussed

by the above author and by other, Joslyn (1950).

Perry (1951) demonstrated, by means of viscosity measurements, the

effect of adding trypsin to a myofibril solution isolated from rabbit muscle. The

author observed that short incubation of a myofibril solution with trypsin caused

a considerable decrease in the viscosity of the solution. The author found that

trypsin degraded the myofibril and destroyed the actomyosin-forming ability of

extracted myosin, yet at the same time had little effect on the ATPase activity.

Viscosity measurements were also a valuable technique in understanding the re-

lationship between actomyosin and ATP. Barany et al. (1952) found that ATP

lowered the viscosity of actomyosin solutions prepared from beef hearts. By this

method, the authors were able to demonstrate the marked activating effect of Mg







demonstrated by White et al. (1957). The electrophoretic properties of myosin

and actin was illustrated by Ziff and Moore (1944) and Spicer and Gergeley (1951).

Smithies' (1955) starch gel electrophoresis technique has been used in re-

cent work for the separation of muscle proteins. Scopes and Lawrie (1963) used

vertical starch gel electrophoresis to compare, and detect changes in, sarcoplas-

mic proteins of beef L. dorsi. Sarcoplasmic proteins were extracted with water

from muscle pre-rigor and from muscles held either at 0C for 20 hr or at 370C

for 4 hr. The authors found that several components were "removed completely

or very much diminished by the post-mortem glycolysis" which occurred at 37C

compared either with pre-rigor muscle or with muscle in which the post-mortem

pH fall had been relatively slow (at 0C). The instability of some components was

explained by the authors as due to either denaturation or isoelectric precipitation

caused by the post-mortem pH fall in muscle. Many of the protein constituents

were completely stable, however, and showed no diminution in the starch-gel

patterns under the conditions utilized. The authors also identified one of the major

components as creatine and phosphoryltransferase. The above method was used

by Neelin and Rose (1964) for examining protein extracted from chicken muscle

during post-mortem aging. The authors found no detectable, consistent change in

the "myofibrillar" proteins during the two-day aging period. Some of the zone

appearing on the starch gel were tentatively identified by the authors as myosin and

actin. "Sarcoplasmic" extracts of chicken breast muscle, however, revealed sig-

nificant changes during the tenderization period. These differences were described

by the authors in terms of the number of zones appearing or to the intensity of the

components.








ions and a smaller inhibitory effect of Ca++ions in the absence of Mg+" on a com-

bination between actomyosin and ATP. In the presence of Mg+ions, however,

low Ca" concentrations were without effect, but higher Ca++ concentrations inhib-

ited the reaction. Szent-Gyorgyi (1960) stated that when solutions of actin and myo-

sin were brought together, a complex, actomyosin, was formed. The viscosity of

this complex as reported by the author was higher than that of the sum of the com-

ponent proteins.


Ultraviolet absorbance and protein concentration

Haurowitz (1963) reported that all proteins strongly absorbed ultraviolet

light and that the maximum absorbance was found near 280 mui. The absorbance

at 280 mil was due to the aromatic residues of tryptophan, tyrosin and phenylala-

nine; their absorbance maxima, as reported by the author, were found at 280, 275

and 258 mu, respectively.

Fischer (1963) approximated the protein concentration in his water and

salt soluble protein extracts by using the absorbance value obtained at 280 mu,

adjusted with a correction factor obtained from the absorbance value at 260 mp.

Even though the author realized that this determination was not an exact one, he

concluded that it was possible to compare the extracts obtained at various times

from different chickens, since they were prepared and analyzed under similar

conditions.














EXPERIMENTAL PROCEDURE


Animals used

The 36 steers used in this study, of Brahman, Hereford and Angus

breeding, were raised at the Everglades Experiment Station. They were

approximately 30 months of age at the beginning of the experiment.


Treatment, management and feeding of the animals

In the summer of 1964, the steers were randomly allotted (within

weight, grade and breed group) to one of four groups to be treated as shown

in Table 1.


Table 1. Experimental design


Lot Number No. of Animals Vitamin Treatments


1 9 Control

2 9 A; 25,000 I. U./animal/day

3 9 E; 50 I. U./animal/day

4 9 Combination of 2 and 3



Vitamins were injected at the beginning of the experiment and on 28-day

intervals throughout the total feeding period of 120 days.

At the beginning of the study, each steer received 113.4 g of phenothiazine,

and all steers were implanted with 24 mg of diethylstilbestrol. The steers were








weighed at the beginning and end of the trial and at every four week interval

during the experiment.

The steers were fed on Roselawn St. Augustinegrass pasture. The

basal ration was composed as shown in Table 2.



Table 2. Composition of ration




Ingredients Kg


Ground snapped corn 352

Dried citrus pulp 363

Cottonseed meal (41% c.p.) 182

Mineral mixture- 11



1/
SMineral mixture of 40% defluorinated phosphate, 22.5% steamed bonemeal,
24.2% salt iodizedd and trace mineralized), 2.5% ferrous sulfate, 3.2% copper
sulfate, 0.15% cobalt sulfate and 7.45% black-strap molasses.


The steers were hand fed once a day an average of 3.6 kg of the basal

ration per day. Fresh water was available at all time.


Slaughter procedure

At the completion of the feeding trial, and for a period of three consecutive

weeks, three steers were randomly selected from each of the four different vita-

min treatments, weighed ( shipping wt. ) and transported by truck 440 kilometers

to the University of Florida Meat Laboratory at Gainesville. Upon arrival, the

twelve steers were weighed (Gainesville wt.). Four steers, one from each lot,








were randomly selected, placed in separate pens, fasted overnight with access

to fresh water and slaughtered the following day. The other eight steers were

provided with hay and fresh water until the night before slaughtering; the above

stated procedure was followed in slaughtering the remaining eight steers. For

the first four steers, the Gainesville weight was used for the slaughter weight;

for the remaining eight steers, the slaughter weight was determined the night

before slaughtering. The twelve steers were slaughtered on three consecutive

days.

The animals were slaughtered using routine slaughter procedures, and

the hot carcass weights were recorded. The hot right side of each carcass was

weighed separately. The left side of each carcass was used in obtaining pH,

temperature and other data, while the right side was used chilled and intact

for carcass grading and rib eye area measurement.

A strict slaughter time schedule was designed for the three successive

slaughter days in order to facilitate data collection and to permit the removal

of the muscle samples at rather exact post-mortem times. In this time sched-

ule, periods of one and a half to three hours were provided between slaughterings.

To minimize the influence of temperature on rigor development, pH and temper-

ature drop, an exact interval of 70 min was maintained between the time of

slaughter and the time each carcass was rolled into the chill cooler.


Chilling, carcass data and aging

All carcasses were chilled at -10C to 2C for 48 hr before being graded

by a Federal meat grader. At the same time, rib eye tracings at the twelfth rib

were obtained, and the chilled right side of each carcass was weighed. Dressing








percentage was calculated using the following formula:


Chilled right side wt. + estimated chilled left side wt.
Slaughte wtx 100 = dressing percentage
Slaughter wt.


After 48 hr of chilling, the left side was rolled into the aging room for

holding for a period of 192 hr (8 days). The temperature of the aging room was

maintained between 4.40C and 5.60C.


Temperature determinations

Temperature readings were obtained with general laboratory mercury

thermometers which ranged from -100C to 1100C with 10C divisions.

Immediately after sticking, a small slit in the hide was made in the right

side of the animal body approximately at the location of the 4th lumbar vertebrae.

The thermometer stem was inserted into the L. dorsi muscle perpendicular to

the back bone to a depth of eight cm. Two min later, the first temperature was

recorded and the thermometer removed to allow further slaughtering procedures.

At 55 min post-mortem, the thermometer was inserted in the left side at approx-

imately the same position as stated for the right side. The thermometer remained

in this position until temperature readings were completed.

After the first temperature reading, temperature measurements were taken

each successive hour on the hour for 24 hr. After the 24th hr reading, observa-

tions were recorded every 4 hr for an additional 24 hr.


pH determinations

For pH measurements, a Beckman expanded scale pH meter (Model 76)

was used. The pH meter was equipped with a flat bulb combination electrode,








sensitive for surface pH measurements, and a thermocompensator that provided

temperature correction for pH measurements.

Starting at the 9th rib and continuing toward the 8th, a small piece of the

L. dorsi was removed, and surface pH readings were made on the fresh cut sur-

face immediately. In the same way, pH measurements of the L. dorsi muscle

were made on the 36 animals at one-hr intervals; reading began one hr after

slaughter and continued through the 24th hr. Readings were then taken every 4

hr for the next 24 hr. The final pH reading was taken at 192 hr post-mortem.


Scheme for steaks removed from the Longissimus dorsi muscle for the different
studies conducted

Exactly one hr after slaughtering, the left side of the beef carcass was

ribbed by cutting between the 9th and 10th ribs perpendicular to the long axis of

the L. dorsi muscle and following the contour of the 9th rib. Cutting toward the

posterior end of the animal, four adjacent 2.54 cm thick boneless steaks were

removed. Another four boneless steaks of the same thickness were removed

at 24, 48 and 192 hr post-mortem.

The steaks were numbered consecutively 1 through 16 starting with the

most anterior steak and proceeding posteriorally as shown in Fig. 1.

Steaks numbered 1, 8, 12 and 16 were wrapped immediately in freezer

paper and were identified with the following data: slaughter number, animal

number and post-mortem interval. These steaks were then frozen and stored at

-240C until used for the protein studies. The remaining twelve steaks were used

immediately after removal from the muscle for the determinations below.

Steaks numbered 2, 7, 11 and 15 were used fresh and uncooked for the












































S1, r p O 11 CST ''I OR TE .E


'92 ^







~I


.9

A I, C 9



3 6
2,7 F N


- S E3 % 51 c

CHAR aC'( 9 15
A R

A', 4 C
S Sei"~


AC I rV


r ScZ edule of reS ev no TT 7 orsi steaks for itud es conducted.


S a,







water-holding capacity determination. The other eight steaks were used for

cooking time, cooking loss, taste testing by trained panel and W-B shear deter-

minations. Steaks numbered 4, 5, 9, 13 and 3, 6, 10, 14 were cooked by broil-

ing and deep fat frying, respectively.


Cooking

Two of the four L. dorsi steaks, removed at 1, 24, 48 and 192 hr post-

mortem, were used for cooking. The steaks were trimmed of all exterior fat

and connective tissue and weighed to the nearest gram. The steaks were cooked

by two methods, broiling and deep fat frying.

In broiling, a meat thermometer (Weston model 2261, 5 in. stem, cali-

brated from 0F to 2200F) was inserted into the most central position (depth,

height and width being considered) of the steak for use in recording initial and

final internal temperature. The steaks were then broiled in a pre-heated electric

oven. As the internal temperature of 76.6C was reached, the steaks were re-

moved from the oven, placed on plates, immediately weighed and data recorded.

The difference between raw and cooked weights was used to calculate per cent

cooking loss. Individual steak broiling times were recorded.

A General Electric (Model CK20, 230 volt, A. C.) deep fat fryer was used

for frying steaks to an internal temperature of 76. 60C. The commercial type

hydrogenated vegetable oil was pre-heated to 1490C and that temperature auto-

matically maintained within a plus or minus range of 1C.


Organoleptic panel and Warner-Bratzler shearing

The cooked, weighed steaks were cooled to room temperature for taste








panel and shear determinations. Two cores 1.27 cm in diameter were removed

from the dorsal, medial and lateral areas of the steaks. Using the W-B shear,

each core was then sheared twice, giving a total of twelve values per steak.

The twelve values were averaged to give the shear score for the steak.

After cores for shear determinations were taken, the remaining portion

of the steak was divided into four sections with numbered portions going to num-

bered members of the taste panel. Each panel member received a portion from

the same position of each different steak. The taste panel members determined

tenderness, flavor and juiciness using the palatability scale shown in Appendix

Table 45.


Water-holding capacity determinations

In the present study, water-holding capacity (WHC) is expressed in terms

of the amount of free or expressable water and bound water relative to the total

moisture content of the muscle. The pressing method of Grau and Hamm (1953)

with the modifications of Ritchey and Hostetler (1964) was used for the WHC deter-

minations of this study. The following procedures were used.

One of the four steaks cut from the L. dorsi muscle on the left side of each

carcass at the post-mortem intervals of 1, 24, 48 and 192 hr was used for this

determination. The removed L. dorsi muscle was trimmed of external fat and

connective tissue. Using a Hamilton Beach meat grinder, the sample was ground

three times and mixed thoroughly after each grinding. The food grinder was

washed and dried thoroughly between each sample grinding.

Total water was determined by the AOAC (1960) method using triplicate

10-g samples.








In determining bound water, triplicate sub-samples of 5 + 0. 1 g were

weighed on aluminum foil 7 cm in diameter; a second piece of foil was placed

on top of the sample. The sample enclosed in the two pieces of foil was placed

between two Whatman No. 41, 11.0-cm filter papers. Using a Carver laboratory

press (Model B), the sample was immediately pressed for one min at 5,800 kg

force. After pressing, the two pieces of filter paper were removed from the

foil enclosed sample. The fragments of meat extending beyond the foil were

trimmed away, and the foil coverings were pulled apart. The pressed sample

was scraped into a previously dried and weighed drying dish. The dish and sam-

ple were weighed to determine the sample weight. Residual moisture or "bound

water" was determined by the AOAC (1960) method.

Free water was determined by subtracting bound water from total water.


Muscle proteins studies

The frozen, stored L. dorsi steaks of 20 steers (5 steers randomly

selected from each treatment) numbered 1, 8, 12 and 16 (taken at 1, 24, 48 and

192 hr post-mortem, respectively) were used for this study.

Preparation of samples

The L. dorsi steak was removed from the freezer and placed in a 3. 3C

refrigerator for approximately 10 hr before extraction. Fat and connective tissue

were separated as completely as possible from the muscle while still partially

frozen. The trimmed, diced muscle was then ground three times in a Hamilton

Beach meat grinder and mixed thoroughly after each grinding. From the paste

thus obtained, the following sub-samples were taken: two g for total tissue nitro-

gen analysis, five g for the extraction and quantitative determination of nitrogen in








buffer soluble and buffer insoluble protein extracts, five g for the extraction

and quantitative determination of nitrogen in water soluble protein extracts,

five g for the extraction and qualitative study of buffer soluble protein extracts

by starch gel electrophoresis and five g for the extraction and qualitative study

of water soluble protein extracts by starch gel electrophoresis.

Extraction and fractionation of buffer soluble, buffer insoluble and water

soluble proteins

The classification and solubility properties of muscle proteins as out-

lined by Szent-Gyorgyi (1960) were the bases for the methods of extraction used

in this study. The extractability of these proteins as determined by Khan (1962)

and Helander (1957) was also considered. By combining and modifying the tech-

niques used by other workers (Weinberg and Rose, 1960; Sayre et al. 1963;

Hegarty et al. 1963; Fujimaki and Deatherage, 1964; Goll et al. 1964 and Khan

and Van den Berg, 1964) a method for the extraction and fractionation of buffer

soluble, buffer insoluble and water soluble proteins was adapted and used as

shown in Figs. 2 and 3 respectively.

To avoid protein denaturation, an ice bath and a cold room at 7 + 20C

were used for the extraction and centrifugation procedures.

Nitrogen analysis

All nitrogen determinations were made using a macro-kjeldahl method

(AOAC, 1960). Protein values were estimated by multiplying the nitrogen value

by a factor of 6.25. Nitrogen determinations were made in duplicate.

Preparation of potassium chloride-potassium phosphate buffer

This high ionic strength buffer was used for the extraction of buffer soluble















Sub- ample of 5 9 -ioo0 into 30 ml Virllo flask


30 0,1 0 5M 0C0phosphale buIfer

Pf00 8. 0-l 071 added


fvl lr o fo r 5 min In a VirtS "I45' maCro

h omoleoon ol0P r at a CstOolcOl $Pll ng Of 30


r anleffd to fnIr I Iu qe ttiboo s lIh 70 m- KCI

ph asphalp bull@ r used for rnlsing


Left in centrifuge tubrt% for I he


CILorllged for 15 mi n In Servall superspeed

0000 Sluo 0 ely SS- I P 20. 100 G using a

S poeoeaol Rool We irindoofmer

11 ype ;61 -tIh selling of 75


S ul r05a I0a 0 Residue


10 0 1h0e5s1a0

K Cj-phoiphato huller add

e I Io r e sdup


SlIOpo000 dod an ah 00

20 ser


pI o000, n a 1a 01l t fe

u, )uqu X layers


00rr r% FP 900 S Wo a
25 rI
290 010 ,ihar t:n




-a~hel -Ilt 20 01

t he a00 I l-itm00 000 built


0- 1 ............ 'l T I
Duplicate aoOO samplit s %,105000p~ ikb saIl

ol 50 ml or 5 ri o0 2 m,

for nlItfogen for lor u Il r -

analyisl 0 1000 S~ 401el

defef,,inalion eatorboame

measuremenf


tell In c0nlrilugo lubes

for I 0,


Centrifuged for 10 rmn at

20, 300 -0G


Supernatant e Vs .due


Res due IrSacslerre

10 p0r P00019h, p I0, I sI

at h 40 ml d0,s7 led

aster used for rinsing









Residue drie, 24 h, in

Oven at 105 C


1`,11 d0,0 ana cOnfetls

Sodfra lo, 0 rlre to d alor

to coo.


0(lqhl 0of01h residue




Oried r-s-lue qro,, ith

,ortar ind petlal

to o po 01,

ft-ulle, Insolutle proteinsi


Sun Jar,,ple of

0 5 10 1 0 9 for

nitrogen anaifso


1F 2 S it9rm o1r 1 tI, 1I ,r j 1U i d h0 0 I I Ia llo,
0510iub0 p0rot 0a rId ofahr quantitaloie and qulolal voe 0j 01001 0on 0









Sub-sample of 5 g weighed into 30 ml Virtis flask
1
30 ml distilled water added
I
Extracted for 5 min in a Virtis"45" macro
homogenizer at a speed control setting of 30

Transferred to centrifuge tubes
with 70 ml distilled water used for rinsing
1
Left in centrifuge tubes for I hr

Centrlfuged for 15 min In Servall
superspeed centrifuge IType SS-I) at
20, 300 x G using a powerstat variable
transformer (Type li61 with setting of 75
1 1


Supernatant
1
Supernatant filtered
through 8 layers
of Klmwipes disposable
wipers (Type 900-S) into a
250 ml graduate cylinder

Disposable wipers
washed with 20 ml dis-
tilled water
1
The volume of filtrate recorded
(water soluble proteins)


Residue
I
ml distilled water
dded to residue


Stoppered and shaken
20 sec
I
Left In centrifuge tubes
for I hr

Centrlfuged for 15 min at
20, 300 x G
1 I


SS1


Duplicate suo-samples suD-sample suo-sample
of 50 mi of 5 mi of 2 mi
for nitrogen for for ultra-
analysis viscosity violet
determination absorb-
ance
measurement


upernatant
J


Residue
discarded


Fig. 3. Scheme for the extraction of water soluble proteins
and other quantitative and qualitative analysis conducted.







proteins. This buffer was made by dissolving the following reagents (ACS) in a

liter of distilled water;

a. 17.69183 g (0.13M) monobasic potassium phosphate (KH PO )

crystals.

b. 22.64379 g (0.13M) dibasic potassium phosphate (K2HPO4) powder.

c. 37.27850 g (0.5M) potassium chloride (KC1) crystals.

The calculated ionic strength (u) of this buffer was 1.02, and the measured pH

of this buffer solution was 6.8.

In order to minimize experimental error in making up the buffer, and

due to the large quantities of buffer required throughout the study, it was desir-

able to make the above buffer in stock solution. The stock solution was kept in

25-L bottles and diluted accurately to the desired concentration when used.

Viscosity determinations

The viscosities (n ) of the buffer and water soluble protein extracts were

determined by the use of a Cannon-Fenske viscometer (size 100). Viscosity

measurements were conducted on all the buffer and water soluble protein extracts

obtained from the 1, 24, 48 and 192-hr post-mortem L. dorsi samples of the 20

animals used in this study.

The temperature of the extracted solutions was adjusted to 170C before

each measurement. Using a volumetric pipette, 5 ml of the solution was then

transferred to the viscometer. The time in seconds required for the liquid to

flow through the capillary tube was recorded. The final reading for each sample

was obtained by averaging the two readings that agreed within one second. The

time required for 5 ml of distilled water at 170C was also determined.








For the calculation of the viscosity of the buffer soluble protein extracts,

a density measurement of the phosphate buffer at 170C was required; a Westphal

balance (No. 683) was utilized for this determination.

The viscosity in centipoise (cps) for the extracted solutions was calcu-

lated using the ratio of viscosity coefficients given below:

1. Viscosity calculation for the buffer soluble protein extracts

n P t
1 11


Where: 12 2 t 2

n = viscosity in cps for buffer soluble protein extracts at 170C.

n 2 = viscosity in cps for H20 at 170C = 1.0828 (Hodgman, 1961-1962).

P1 = density in g/ml for buffer soluble protein extracts at 170C =

1.0536 (as measured).

p = density in g/ml for H20 at 170C = 0.99880 (Hodgman, 1961-1962).
2
t1 = time in sec required for buffer soluble protein extracts to flow through

the capillary tube of Cannon-Fenske viscometer.

t2 = time in sec required for II20 to flow through the capillary tube of

Cannon-Fenske viscometer.

2. Viscosity calculation for the water soluble protein extracts

The above ratio of viscosity coefficients was also used for the calculation

of the viscosity of the water soluble proteins extracted. Because water was used

in the extracting of these proteins and because of the low nitrogen concentration

in the extracted solutions, it was assumed that p =02 Therefore, the final

coefficient ratio used was
ni t1

n2 t2








Where:

n = viscosity in cps for water soluble protein extracts at 170C.

n2 = viscosity in cps for H20 at 170 C = 1.0828 (Hodgman, 1961-1962).

t = time in sec required for water soluble protein extracts to flow

through the capillary tube of Cannon-Fenske viscometer.

t = time in sec required for H20 to flow through the capillary tube of

Cannon-Fenske viscometer.

Ultraviolet absorbance measurements

The ultraviolet absorbance of the buffer and water soluble protein extracts

was determined by the use of a Beckman spectrophotometer (Model DU) with silica

cuvettes. Ultraviolet absorbance measurements were conducted on all the buffer

and water soluble protein extracts obtained from the 1, 24, 48 and 192 hr post-

mortem L. dorsi samples of the 20 animals used in this study.

For determination of ultraviolet absorbance, a 0.2 ml aliquot was diluted

with 2.5 ml distilled water (1:12.5 dilution) and examined at 260 and 280 millimi-

crons. Distilled water was used as a blank for both buffer and water soluble

protein extracts.

Starch gel electrophoresis studies

The electrophoretic method utilized in this study was that of Smithies (1955)

in which he introduced starch gel as the supporting material. The improved

(vertical) method (Smithies, 1959), and the discontinuous system of buffers with

platinum electrodes of Poulik (1959) was used in this qualitative study. Some of

the modifications and procedures used by Pert et al. (1959); Pierce and Free (1961);

Tsuyuki et al. (1962) and Neelin and Rose (1964) were also employed.








Preparation of muscle protein extracts for starch gel electrophoresis

analysis

Two sub-samples of 5 g of the paste obtained from the frozen stored L.

dorsi steaks numbered 1, 8, 12 and 16, taken respectively at 1, 24, 48 and 192 hr

post-mortem, were used for the extraction of buffer and water soluble proteins

for starch gel electrophoresis studies. The scheme used for the extraction of

buffer and water soluble proteins in this qualitative study is shown in Figs. 4

and 5 respectively.

Precautions against protein denaturation were the same as previously stated

for the extraction and fractionation of buffer and water soluble proteins.

Preparation of electrolyte buffers

Two different electrolyte buffers were required for the electrophoretic

fractionation of proteins investigated.

The tris-citrate buffer electrolyte solution, used in making the starch gels,

was made by dissolving 9.2 g tris hydroxymethyll) amino methane (Trizma base,

Sigma Chemical Company, St. Louis, Missouri) and 1.05 g anhydrous citric acid

(H3C6H507) in 1 L of distilled water. The pH of this buffer solution was 8.8.

The borate buffer electrolyte solution, used in the electrode vessels, was

made by dissolving 18.5 g boric acid (H3BO3) crystals (ACS) and 2. 0 g sodium

hydroxide (NaOH) pellets (ACS) in 1 L of distilled water. The pH of the solution

obtained was 7.0.

For the same reasons as previously stated, it was desirable to make the

above electrolytes in stock solutions. The stock solutions were kept in 10 to 20 L

bottles and solutions were diluted accurately to the desired concentration when used.







Sub-sample of 5 g weighed into 30 ml Virtis flask


25 ml of 0.5M KCl-phosphate buffer (pH6.8, = 1.02) added


Sample extracted for 5 min in a Virtis "45" macro homogenizer

at a speed control setting of 30


Transferred to certrifuge tubes


Left in centrifuge tubes for 1 hr


Centrifuged for 15 min in Servall superspeed

centrifuge (Type SS-1) at 20,300 x G using a powerstat vari-

able transformer (Type 116) with setting of 75

1-- ------------
Residue Supernatant

discarded
Supernatant filtered through

Whatman No. 41 filter paper


Filtrate (buffer soluble pro-

teins) stored in freezer until

analysis


Fig. 4. Scheme for the extraction of buffer soluble proteins
for starch gel electrophoresis analysis.








Sub-sample of 5 g weighed into 30 ml Virtis flask


15 ml of deionized water added


Samples extracted for 5 min in a Virtis "45" macro homo-

genizer at a speed control setting of 30


Transferred to centrifuge tubes


Left in centrifuge tubes for 1 hr


Centrifuged for 15 min in Servall superspeed

centrifuge (Type SS-1) at 20,300 x G using a powerstat

variable transformer (Type 116) with setting of 75


Residue Supernatant

discarded
Supernatant filtered through

Whatman No. 41 filter paper


Filtrate (water soluble pro-

teins) stored in freezer until

analysis


Fig. 5. Scheme for the extraction of water soluble proteins
for starch gel electrophoresis analysis.








Preparation of the starch gel

Gels were prepared from hydrolysed starch especially prepared for starch

gel electrophoresis (Connaught Medical Research Laboratory, Toronto, Canada).

The standard procedure used in making up the gels was as follows:

Exactly 500 ml of the tris-citrate buffer was added to 70 g hydrolysed

starch in 1000-ml Erlenmeyer flask. The starch and buffer suspension was gently

heated over a burner with continuous vigorous agitation until the granules were

ruptured and a clear solution obtained. Degassing was then conducted by suction

through a one-hole rubber stopper inserted into the neck of the flask and connected

through a suitable trap to a water aspirator. The resulting viscous, translucent

fluid was then poured into a rectangular plastic tray (32 cm long, 12.2 cm wide

and 0.7 cm deep). The surface of an 8-sample gel saw cuts cover (Otto Hiller,

Madison, Wisconsin) was layered with a few drops of mineral oil and then heated

in an oven at 700C. The preheated gel cover was used to cover the hot gel.

Weights were placed on the four corners and the middle of the plastic cover to en-

sure a uniform gel thickness and to prevent the formation of bubbles. The gel was

then allowed to cool and solidify overnight at room temperature.

Application of samples and electrophoresis conditions

After removing the cover and trimming the excess of gel from all sides

of the tray, the thawed protein extracts were introduced carefully into the sample

slots by means of disposable capillary pipettes. Each sample slot held approxi-

mately 0. 06 ml of the protein extract. A liquid was made of approximately 227 g

paraffin and 100 g vaseline, heated in an oven at 700C; a portion of this liquid was

carefully poured to form a thin film on the entire surface of the gel. The end plates









of the gel tray were removed, and the gel was placed vertically between two

buffer chambers. Electrical contact between the gel and the sodium hydroxide-

borate buffer in the chambers was made through wicks composed of 3 layers of

Whatman No. 2 filter paper. Platinum electrodes were used in the buffer cham-

bers.

Electrophoresis was carried out at 7 + 2 C with a current of approximately

45 22 ma and voltage of 18 32 V/cm across the gel for periods of 6 to 8 hr. A

regulated power supply unit and a vacuum tube voltmeter (Model IP32, and IM11,

respectively, Heath Company, Benton Harbor, Michigan) were utilized for supply-

ing, regulating and checking the voltage and current.

Staining and washing procedures

After completion of electrophoresis, the paraffin-vaseline film was re-

moved, and the gel carefully transferred to a slicing tray. The gel was sliced

lengthwise with a fine fishing thread. The horizontally sliced strips, with the cut

surface up, were stained for approximately 1 min with a saturated solution of

Amido black 10B (Naphthol blue black, E. Merckag, Darmstadt, Germany) in

methanol-water-acetic acid solvent (5:5:1 by volume). To remove excess stain

from gel strips, the strips were washed with solvent for a period of 20 to 30 hr by

placing them in a tank of an automatic gel washing machine (Otto Hiller, Madison,

Wisconsin). By means of a centrifugal pump provided with this washing apparatus,

the stain was continuously removed from the solvent by recirculation through a

drum of activated charcoal. A gentle stream of colorless solvent over the gel

strips was provided by means of a multiple hole tubing in the washing tank.

The stained, washed gel was then diagrammatically sketched, wrapped in








Saran Wrap and identified with the following data: gel run, slaughter number,

animal number and type of extract. The identified gels were stored in the

refrigerator at 3.30C.

System of sample analysis and data recording for the starch gel

electrophoresis

Water and buffer soluble protein extracts were analysed by the starch gel

electrophoresis technique on alternate days. Starting at the right end of the gel

(slot no. 1) the following sample order was used in filling the eight sample slots:

1, 24, 48, 192, 1, 24, 48 and 192-hr post-mortem samples. Data were recorded

as shown in Fig. 6.

Diagrammatic sketch for the gel

The faint appearance of several patterns on the gel strips and the notice-

able fading of those patterns shortly after removing the gel from the washing solvent

were recognized in preliminary experimentation with the technique. The above two

factors, combined with the difficulty of handling the fragile gel strips, made it

necessary to make a diagrammatic representation for the different patterns ob-

tained on the gel.

For each sample four locations on the two gel strips were obtained by

running the sample in duplicate per gel; the clearest separation from these four

locations was sketched in the following manner:

A base line representing the original sample slots in the gel was drawn on

the data sheet. A portion of this horizontal line was divided into four 0.9-cm parts

with each part representing 1, 24, 48 and 192-hr post-mortem intervals respective-

ly. With the use of a ruler (0.1-cm divisions), the distance of each pattern on the

gel from the origin at the four intervals examined was measured, and a line








Starch gel No.

Slaughter No.

Animal No.

Type of extract




Voltage reading (power supply unit)

ma reading (power supply unit)

Voltage across starch (vacuum tube
voltmeter)


Time


Slot No.


Fig. 6. Starch gel electrophoresis data sheet.


Date


Start


Finish


Sample No.








representing each band was drawn at a distance from the base line equal to that

measured on the gel strip. The intensity and width of the different bands were

also maintained in this diagrammatic sketching. In many cases it was necessary

to run the extracts more than one time to get an adequately clear separation.


Statistical analysis

All data were punched on cards for electronic computing on the IBM 709

computer. The statistical methods (Snedecor, 1959) used in analyzing the data

were:

1. Analyses of variance and covariance were used to determine if

significant differences existed among the different traits studied.

2. Simple correlation coefficients were computed to determine if relation-

ships between certain variables existed.

3. The stepwise regression analysis was also employed in this study.

The stepwise regression analysis which was used picks out the independent vari-

able having the highest simple correlation with the dependent variable. It then

proceeds in a stepwise manner to add one independent variable at a time which,

when combined, gives the best estimate of the dependent variable. The next

variable selected is that variable having the highest partial correlation with the

dependent variable and independent of those factors in the equation. This process

continues until it is ascertained that addition of any of the remaining independent

variables will not significantly improve the fit of the regression equation.

Partial regression coefficients show the effect of an independent variable

on a dependent variable, with other variables held constant.













RESULTS AND DISCUSSION


The effect of feeding treatment on rate of gain, in-transit shrink and slaughter
characteristics

Rate of gain, per cent in-transit shrink and slaughter characteristics of

the four lots are summarized in Table 3. The significance of differences in

response to the four treatments are shown also in the same table.

Only slight differences in average daily gain, carcass weight, dressing

per cent, liver per cent, and carcass shrink were found between the four lots.

The differences lacked statistical significance, however. Small differences in

per cent in-transit shrink were observed between the four groups; analysis of

variance showed that the differences were not significant.


The effect of feeding treatment on carcass characteristics

Table 4 gives the mean values and the significance of differences between

the four lots in carcass characteristics.

Only insignificant differences in carcass characteristics were found

among the four treatments.

Feeding treatment had no effect on degree of marbling, conformation,

carcass maturity or color, texture and firmness of the lean. Carcass grades

were similar for all lots and the color of the external finish was not influenced

by feeding treatment. Differences between lots in rib eye area, fat thickness

over the rib eye and estimated kidney knob were not significant. Differences














02Idi
zz


r-1~ c
CT> t- .4
C D r -
CO *^ *-


to 00 r-4
m 1f4 -4


ic121020


0)t- ~ccl-
e -4- ,-


C4 r-4
oCm
r-l C~


C c)


qw*^ 0 cc
(D D r-4
t r-4
o d o
CDD
c~ -~


'-I 0
LOM
1 (


IN Cr)
- .-4
eq 0





tfi m






o Lfn



cor
000












I-< rQ
fln











P^ a

!
;!
l -)


N WH
m 't r4 0C4


-y-





'a
*o c
' C g '
- ^i c3 *













Cd




C) 0
2


CO





4-'
t.










o
$a










,

F o
.0











o
"-4
I CO












o -











Cd


Z~Zzzzzz










C4 CD CD C) C) CQ
4 atO l)-4












o C') '' ^ Lc)C C')









CN C CO CO C- M C '
C' NtC - C~D CM R













o C C )tf C) C
oI- ,o-
Q)ZZ Z





r4d4r


o a( o CD
0 0





ZooZZ
S0) 0 CO(D







CM C; CM
"t


















LO


Q Q






N.o


o 0

c- s



0 0 0
a o Po.O
Cd >1 4 -4 m I


U C < rMz>rx







between lots in estimated per cent of boneless, fat-trimmed meat from the

round, rump, loin, rib, and chuck lacked significance.


Post-mortem changes in the pH of the Longissimus dorsi

Fig. 7 shows the pH curve obtained by plotting the average pH values of

the 36 animals against the post-mortem time.

The graph illustrates that the average pH values of the L. dorsi decreased

with time post-mortem, rapidly for the first 15 hr and slowly thereafter; min-

imum values were obtained at the 192 hr post-mortem observation. These

results are in agreement with the reports of Paul et al. (1952) and Paul and

Bratzler (1955).

The correlations between average pH values of the L. dorsi at 1, 24,

48 and 192 hr post-mortem are presented in Table 5. The correlations between

the average pH value at 1 hr and the pH values at 24, 48 and 192 hr were low

and insignificant. pH values obtained at either 24 or 48 hr post-mortem were

not closely related to pH values obtained at 192 hr post-mortem. These results

support the report of Briskey (1959) and the finding of Marsh (1954) who found

little relationship between initial and ultimate pH values. Huffman (1962) found

a highly significant negative correlation of -. 92 between initial and ultimate pH

values in sheep, but the lack of agreement between his work and that reported

herein may be attributed to either species differences or to Huffman's (1962)

attempt to alter rate of pH drop or rate of rigor development by pre-rigor

treatments.

A significant correlation (P<.01) of .64 was found between average pH






























































i I
0


I I
0
re)


SI

O


- 0









LO



LLI
-0 -

0






0 c

o


0
I


-0





0


c=
7D


E -









ul
0c






L
)- CD


0 O







o I




LA-
a


C3
aM
U cn

** -





a0


Hd


0
I)


i


--~i


i








values at 24 and 48 hr post-mortem. This highly significant relationship

might be explained by the very small change in pH values from 24 hr to 48 hr

post-mortem as shown in Appendix Table 49. This result agrees with the

work of Lewis et al. (1963), who found a correlation coefficient of .70 between

pH values obtained at 24 and 48 hr post-mortem in pork L. dorsi muscle.


Table 5. Simple correlation coefficients between pH values
of the L. dorsi at four post-mortem intervals.


pH at:

24 hr 48 hr 192 hr


pHat 1 hr .21 .24 .22

pH at 24 hr .64** .23

pH at 48 hr .31


*P<.05 = .33
**P<. 01 = .42


The relationship between pH and tenderness

Simple correlation coefficients between average pH and tenderness values

obtained 1, 24, 48 and 192 hr post-mortem are presented in Table 6.

Correlations between pH of the L. dorst taken 1 hr post-mortem and

shear tenderness values of broiled and deep fat fried steaks, cooked at 1 hr

post-mortem, were -.50 and -.65 respectively; these correlations were highly

significant (P, 01). Lower correlation values were obtained between the 1 hr

pH value and panel tenderness scores of the steak cooked at 1 hr post-mortem.





56




*
mx. C) c t~- 0 0


r- * *
C1~lc ~C)CV~~C11









00 CD I m ~ r Lf
[I-I Ic I
o
.0 OS <1M 0 0




Q Tc C 0 0 C0 L OC 00





o












0
0 1' LO














.4-
-o




0 0
a)r)
0











$4
0 =

LO,
0

0





mO O L m 4




00 0








C0 *
*i*

















o b o b
00 E
00














a 'a





aN 0










0 =, n 0 0'a,1:
T3J


























I4 N -t ri %.0 C.4 -4 44 -
J 44 40 0 0 0 0
0 <








S 1I l4 l. 4. a) ')
4 a a 0 W P %
a S SS














-o 1.4
.0 .0.0.0.












o~d C) C) 0
.2 E a .















0C'000 0 *


0 0 . 4. 4. 4
o 0 0. 0 c S
p<- &q u u +
F3 bfl cg d
1- *- *cd~~












,-< 0 c-4 mM C) ell ell


*


I I



I
*





c 0










**
0 C


0 0 ell


00 v m
C C)


o 0 01 0I
OD t- oC
00hleq


- -4 cl -


*
c-< .-^ C



c.' cr- CT
o *v oD
01 01 M






C)~
hlhq~


a *







0o


I I





o c~
o -e


4-4 4-4 4-4 '- 4-4





,-4 $ $.4 4 $4 $.4
0 0 m m 0
4.4 C13 1, v-4 44 -4 -4 4 -
d)0 0 0 o 4




Cd Cd Cd Cd -4 r4 -4-4
W c c c c
04 04 04 0. 0
a a a a a t rt rt
Qt~~ ~ Q.Q .K


S0
0 0


I






rZ4



Cd



pq


C

0
0
U


CT0)

II II

oo



* *
*x








However, the panel tenderness score of the broiled steaks was significantly

(P,.05) correlated with the initial pH value, and the correlation approached

the same level of significance for those steaks cooked by deep fat frying 1 hr

post-mortem. These correlations clearly indicate that the higher the initial

pH value the more tender the meat at 1 hr post-mortem.

Although it can be demonstrated that rigor may proceed without

glycolysis, the drop in pH caused by the conversion of muscle glycogen to

lactic acid may be used as an indication of rate of rigor development or as an

indication of the severity and extent of rigor. In other words it may be assumed

that a rapid rate of pH drop or a rapid rate of glycolysis from slaughter to 1 hr

post-mortem is associated with either a rapid rate of rigor development or

more severe rigor. Since initial pH was found by Bate-Smith and Bendall

(1949), Briskey (1959), and Beecher et al. (1965) to depend on the severity of

the death struggle, the results obtained in this study would lead to the theory

that the more relaxed the animal was before or during death, the less rapid and

less severe the rigor and the more tender the meat should be at 1 hr post-mortem.

These relationships are explained by Davies' (1963) theory on muscle contraction

and are in agreement with the report of Locker (1960 a) who found that the relaxed

muscles were more tender than partly contracted muscles of beef.

The most striking phenomenon observed in the relationships between

initial pH value and tenderness at the four post-mortem intervals was the differ-

ence in the direction of the signs of the correlations. Initial pH and tenderness

of steaks cooked 24, 48 and 192 hr post-mortem were inversely related, which

was contrary to the 1-hr post-mortem pH-tenderness relationship. Initial pH







was significantly (P<. 01) correlated with the 24-hr panel tenderness values;

correlations of -.43 and -. 49 for broiled and deep fat fried steaks, respec-

tively, were obtained. The correlation between initial pH and 24-hr shear

tenderness was .38 (P,. 05) for the deep fat fried steaks and approached that

level of significance for the broiled steaks. The correlations between initial

pH and 48-hr tenderness were significant only with panel tenderness. The

relationships between initial pH and 192-hr tenderness were low but approached

the 5 per cent level of significance.

The correlation coefficients showed no relationship between the 24-hr

shear and panel tenderness values and pH value at that time. While a low re-

lationship existed between the 24-hr pH value and the 48-hr tenderness value,

the relationship between the same pH value and 192-hr tenderness value was

highly (P<. 01) significant.

The correlation coefficients between the 48-hr pH value and shear

tenderness values at 48 hr were either approaching the 5 per cent level of

significance (for broiled steaks) or significant at that level (for deep fat fried

steaks). Higher relationships between the same pH and the 48-hr panel tender-

ness with correlations of -.40 and -.55 for the broiled and deep fat fried steaks,

respectively, were obtained. These correlations were significant at the 5 per

cent and one per cent levels of significance, respectively. The results agree

essentially with work performed by Judge et al. (1960) who found significant

negative correlation between panel tenderness and pH of pork loins. Correlation

coefficients between the 48-hr pH value and 192-hr tenderness values were

highly (P<. 01) significant; higher pH values were associated with less tender

steaks by both panel and shear tests.








Low correlation coefficients of .19 and .17 between final pH value and

shear tenderness values at 192 hr for the broiled and deep fat fried steaks,

respectively, were found. However, the correlations of -. 33 and .30 between

the same pH value and panel tenderness were significant (P<. 05) for the broiled

steaks and approached the same level of significance for the deep fat fried steaks.

These data tend to agree with those reported by Husaini (1950 a) who found no

correlation between pH and tenderness at 14 days post-mortem. Walter et al.

(1965) found negative correlations of -. 230 and -. 375 between W-B shear values

and pH for the broiled and deep fat fried L. dorsi steaks at 5 days post-mortem;

steaks with higher pH tended to be more tender. In this study, however, those

correlations were positive, indicating that higher pH steaks tended to be less

tender.

Generally, the correlation coefficients obtained between the rate of change

in pH from 1 hr to 3, 5, 7, 9 and 11 hr post-mortem and tenderness values at

24, 48 and 192 hr post-mortem were either low or irregular. However, the most

uniform and significant correlations were found between rate of pH change from

1 to 9 and from 1 to 11 hr post-mortem and tenderness values at 24 hr post-

mortem. The correlations between the rate of pH changes and final tenderness

were either low or approaching zero.


Temperature of the Longissimus dorsi during the chilling period

The curve in Fig. 8 shows the post-mortem time-temperature relation-

ship. The curve represents the average temperature values of the 36 carcasses

during the first 48 hr post-mortem. The graph illustrates that the internal

temperature fall of the L. dorsi was slow for the first hour and very rapid












48



40i



32



S24



16-



8-



0-

0 10 20 30 40 50
HOURS POST-MORTEM
Fig. 8. Post-mortem changes in temperature
of L. dorsi muscle as presented by average values
with standard deviations.








thereafter for a period of approximately 20 hr post-mortem. Temperature

change was slow after the 20th hr post-mortem, and the rate of chilling decreased

with time until the internal L. dorsi temperature reached the chill room temper-

ature of 0.0+.60C at approximately 28 hr post-mortem.


The relationship between pH and rate of chilling of the Longissimus dorsi

A simple correlation coefficient analysis was used to study the relation-

ships and/or the influence of temperature decline on pH change post-mortem;

correlations are presented in Table 7.

At 1 hr post-mortem, a correlation coefficient of -.52 (Pe. 01) between

temperature and pH was found; higher temperatures were associated with lower

pH values and lower temperatures were associated with higher pH values. Since

Forrest et al. (1965) reported correlations of .33 (Pc. 05) and .35 (P,. 05) between

muscle temperature at death and respiration and heart rate, respectively,

immediately prior to death, and since heart and respiration rates are associated

with blood and oxygen supplies to the muscle, it may be assumed that variations

in initial muscle temperature are partially due to muscle activity preceding death

and/or during the death struggle. Bate-Smith and Bendall (1949) found that the

initial pH value of the muscle was the result of muscle activity immediately pre-

ceding death and/or of the death struggle. Beecher et al. (1965) found that the

pH of the Semitendinosus was higher in pigs insensibilized by sodium pento-

barbital injection (exhibited no death reaction) than in animals insensibilized

with a captive-bolt pistol.

Average pH values at 24, 48 and 192 hr post-mortem were not related

to the 1 hr temperature value. These data substantiate the observations of





63






t 0 0 CO CL O0
e I In in I" in




S * *
m 0 CO




c





0 0
* *









a .














0.
co co




Lf C>

I I






























4,-,4
4- .m o a t- 0 a
o g n 0
m I I I -c












o 0
S 4












s2 2 sf. .


( DCc2 CC
ot oSl *
0. 0
2 in
^ i

























a tf K o tf t








Bate-Smith and Bendall (1949) and of Briskey (1959) that ultimate or post-rigor

pH depends on the animal's state of fatigue and level of feeding prior to slaughter.

Correlations between L. dorsi temperature at 1 hr and the rate of pH

change from 1 hr to 3, 5, 7, 9 and 11 hr were -.56 (P<.01), -.46 (P,.01),

-.41 (P<. 05), -. 30 and -. 16, respectively.

This decrease in correlation could be explained by the fact that the vari-

ations between carcasses in rate of pH change decreased with increasing time

post-mortem.

Average pH values at 24 and 192 hr post-mortem were independent of

the rate of chilling. However, average pH values at 48 hr were significantly

(P< .05) related to the rate of chilling from 1 to 3 and from 1 to 9 hr post-mortem.

While the correlations between the rate of pH change from 1 to 3 and

from 1 to 5 hr and rate of temperature change from 1 to 3 hr were low, all

other correlations between pH and rates of temperature change were either sig-

nificant (P,.05) or highly significant (P<. 01). These negative correlations

indicate that rate of pH change or rate of rigor development is inversely depen-

dent on rate of chilling. These data are in agreement with the report of Beecher

et al. (1965) who found that lowering the post-mortem holding temperature from

37 to 40C slowed post-mortem glycolysis (retarded pH decline) in porcine

Semitendinosus.


The relationship between tenderness and rate of chilling of the Longissimus dorsi
post-mortem

A simple correlation coefficient analysis was used to investigate the

relationship between tenderness and post-mortem change in temperature, as








an index of rate and extent of rigor development; results are shown in Table 8.

The correlation coefficients between L. dorsi temperature at 1 hr and

tenderness values of the broiled steaks at 1 hr post-mortem were .54 and -. 55

for shear and panel scores, respectively. The relationships were highly signif-

icant (P<.01). Highly significant (P<. 01) correlations obtained between the 1-hr

temperature and tenderness values of deep fat fried steaks were .65 and -.49

for shear and panel, respectively.

The correlation coefficients between the L. dorsi 1-hr temperature and

tenderness values at 24, 48 and 192 hr post-mortem were low and insignificant.

The correlation coefficients between rate of temperature change from

1 to 3, 5, 7, 9 and 11 hr post-mortem and tenderness values at 24, 48 and 192

hr post-mortem were also low and of no significance.

According to these results, no direct relationship between tenderness

and rate of chilling was found. This result was expected, since all carcasses

were chilled under the same temperature. The significant correlations between

the L. dorsi temperature and tenderness values at 1 hr post-mortem were ex-

plained previously by the significant relationship found between temperature and

pH at 1 hr post-mortem.


Post-mortem changes in bound and free moisture of the Longissimus dorsi
muscle

Average values for per cent total, bound and free moisture for the 144

raw L. dorsi steaks utilized at the four post-mortem intervals (36 steaks per

interval) are graphically presented in Fig. 9. Also, shown in the same figure

are average pH values at 1, 24, 48 and 192 hr post-mortem.














00 i N -4i i C

S I IM IM I


0
I' I


o tO t- 0
4 0 0 0-


LO 0
0 0


0 0


II II

00

v V

*











00 m 00


C


LO -4 0 ) -

I I I 1


I I


0 0
C>I


LN






CO C1
C. C.


N C) c0
0 r 0


0


CM C" CO 00
C C


bfl bfl bfl
C C C




.d 1.4 1$C -C C

a) Cl aLO a)t-
0. 0

3 3 3 B
= 4 s a s a4





H 0


a)

0)


a)


a)




0
0
u


a)





aC
.d






0
as4
-1
d W


Cd


a)

2

C C

s
E


I2


co eI


II II



* *
*













80-





60-


40-


20-





O-


cii

bc3 tx


I 24 48 192
HOURS POST-MORTEM


L TOTAL


j BOUND FREE


-7.0





-6.5





6.0 0





-5.5





5.0


ED pH


Fig. 9. Post-mortem changes in total, bound
and free moisture of raw steaks from L. dorsi muscle.








Variations in per cent of total moisture among the four post-mortem

intervals were very small. The average values of total moisture at 1, 24, 48

and 192 hr were, respectively, 74.27, 74.87, 74. 84 and 75.18 per cent. The

difference between average values was less than one per cent. Since the amount

of free moisture was determined in this study as the difference between total

and bound moisture values, the observation that variations between post-mortem

intervals in the amount of total moisture were minute was important because

post-mortem changes in free moisture was a primary objective of this study.

In general, bound moisture decreased with time post-mortem. The

decrease in amount of bound moisture with time post-mortem was significant

at the .005 level of probability. The highest average value (67. 10 per cent)

was observed at 1 hr post-mortem while the lowest value (61.73 per cent) was

noticed at 192 hr post-mortem. An unexpected increase of .42 per cent in the

amount of bound moisture was observed between 24 and 48 hr post-mortem.

Average pH values at 24 and 48 hr were 5.71 and 5.69, respectively. This small

increase in the amount of bound moisture may have been due either to experi-

mental error or to some biochemical changes that caused an increase in net

charge of muscle proteins (Hamm, 1963).

The trend in post-mortem change in amount of free moisture was

opposite to that observed for the bound moisture. A marked increase in amount

of free moisture from 7.17 per cent at 1 hr to 11.97 per cent at 24 hr was found.

A rather slow change in amount of free moisture was observed after the 24th

hr; a maximum average value of 13.45 per cent was found at 192 hr post-mortem.

A small decrease in amount of free moisture (.45 per cent) was also observed








between the 24th and 48th hr. This decrease could be explained by the same

factors discussed for the bound moisture. Here, also, post-mortem changes

in amount of free moisture were significant at the .005 level of probability.

These data are in agreement with that reported by Hashimoto et al. (1959) and

Wierbicki and Deatherage (1958) who found that meat hydration dropped very

markedly within a few hours after slaughter and reached a minimum in 24 to

48 hr; these data are contrary to the above two reports in that the increased

hydration found with aging in those studies was not observed in this study. In

fact, meat hydration was found to decrease with aging for 8 days. This obser-

vation was supported by the noticeabletdecrease in pH with aging; minimum pH

values were obtained at 192 hr post-mortem.


The effect of pH and time post-mortem on free moisture of the Longissimus
dorsi muscle

The effect of pH and time post-mortem on per cent free moisture of the

L. dorsi muscle is shown by simple correlation and step-wise regression

analysis. Table 9 presents the simple correlation coefficients obtained between

these variables of pH and time using the 144 L. dorsi steaks removed at four

post-mortem intervals.

Free moisture showed a significant (P<.01) relationship to pH with a

high correlation coefficient of -.73. This result is in line with the report of

Hamm (1960) who found that the minimum hydration of meat was around pH 5.0.

Also, this result supports the findings of Swift and Berman (1959), Judge et al.

(1960), Kauffman et al. (1961) and Lewis et al. (1963) who found a high relation-

ship between pH and free moisture.








Table 9. Simple correlation coefficients between time
post-mortem, pH, free and bound moisture.!/


Independent
variables Time pH


Free moisture, % .53** -.73**

Bound moisture, % -.48**

pH -.55**

I/Using values recorded at 1, 24, 48 and 192 hr post-mortem.

**P. 01 = .22


Time post-mortem significantly (P<. 01) influenced the amount of free

moisture in muscle, with a correlation coefficient of .53. This relationship

was substantiated by the negative significant correlation of -.48 found between

time post-mortem and bound moisture.

A negative significant (P<.01) correlation of -.55 between pH and time

post-mortem was found. This relationship could be explained by the fact that

average pH values decreased with time post-mortem, with minimum average

pH values at 192 hr post-mortem.

Simple and multiple correlations and regression coefficients for pH and

time post-mortem influencing or related to muscle free moisture are presented

in Table 10.

In this study muscle pH was responsible for 53.29% of the variability

in per cent free moisture in the L. dorsi muscle and was the first variable

entering the step-wise regression analysis. If the predictive value of per cent

free moisture during 1 to 192 hr post-mortem was Y then the predictive equation








for Y from the knowledge of muscle pH at that particular time and during this

specific (1 to 192 hr) post-mortem period becomes: Y = 36.93694 4.34375

(muscle pH). From the predictive equation, a .23 pH unit increase would de-

crease free moisture one per cent.


Table 10. Coefficients for predicting free moisture from piH
and time post-mortem.1/


Partial regression
Independent r coefficients
variable XY R Constant bl b2


pH -.73**a .73**b 36.93694 -4.34375

Time .53**a .75**c 32.70938 -3.72526 .00813


/ Using values recorded at 1, 24, 48 and 192 hr post-mortem.

**ap<. 01 = .22

**bP<. 01= .22

**c .01 = .25



Time (1 to 192 hr) post-mortem had a significant simple correlation co-

efficient of .53 with per cent free moisture and accounted for 28.09% of the var-

iability in the amount of free moisture in the muscle. When time post-mortem

was combined with pH, the multiple correlation coefficient was increased from

.73 to .75. This accounted for an additional 2.96% of the variation in free-moisture

per cent, and the predictive equation was: Y = 32.70938 3.72526 (muscle pH)

+ .00813 (hr post-mortem). Holding pH constant, a one hour increase during

the post-mortem period of 1 to 192 hr would increase free moisture content







.008 per cent. The addition of time, therefore, did not result in a significant

increase in the estimate of per cent free moisture in the L. dorsi muscle.


The effect of pH, initial steak temperature, time post-mortem, free and bound
moisture, cooking time and cooking method on cooking loss

Simple correlation coefficients between the most important factors in-

fluencing or associated with cooking loss as of 288 L. dorsi steaks are presented

in Table 11. The relationship or the influence of pH, free moisture and cooking

time on cooking losses are demonstrated in Figs. 10, 11 and 12, respectively.


Table 11. Simple correlation coefficients between cooking loss
and pH, initial internal steak temperature, time post-mortem, free and
bound moisture, cooking time and cooking method.

Cooking loss, Cooking time
Independent variable % (Min/100 g wt.)


pH of L. dorsi at time
of cooking

Initial internal steak
temperature

Time post-mortem, hr

Free moisture, %

Bound moisture, %

Cooking time (Min/100g wt.)

Cooking method


-.63**


-.63**

.52**

.46**

-.43**

.40**

.26**


-.54**


-.52**

.46**

.51**


**P.01 = .15



The pH of the L. dorsi steaks at time of cooking was closely related to

per cent cooking loss, with a highly significant (P<. 01) correlation coefficient













Hd

(D


9 "7 16 "'























8' 98"'9


>-
LL

0-

U

c\


F-
c:
oO 0


'-
(I)
0
0-


I
-.

rl


z
IJ
0


0 ( cOD c OO
I rO rO c\ CM

% 'SS07 9NIYNO0














% '3tynls/oIm 33Yj4


I I


0I


18 Z6 Z ::::
/9 0 : 9 tlf .. ...........
18. E;26ZT ~i~: ~ ...................j::


16'1 if//g L _




02T4f ....... .\\....

;9ZZ'1 6 '


I I
(0
rr>


I I

rf)


I
No


% 'SS07 ONINOOO


>-




LL
0.

rL
LJ




LJ


_o2
0

w

rr
LU


LU


2

)-

0
n-




0
Qa


V)




cL


0
0





CD-




E


E
L>I
0










0 0

0.J
c0








-







L-0


r (j
z

cr_
m


El














/M 001/ u/ '31Ml/ 9ANIN00O




-
(\J



IC -T IC -f ------------------------- ,,j ,,
26 f


0.
NZ 09
Z^^T m ^: ^ji iiijis1i iliii o





90O" | .06: : ) ._
9I-- c


CO
00 w









% SOz E.
189/ 1 66 Z ..,





O~z T i 80 -r 0....
cC



f02 f 0 0 9 7 F. O






r) r N

% 'SS07 9NIO003







of -.63. This result substantiates the observations of Hamm and Deatherage

(1960) who found that the pH of fresh meat markedly influenced water-holding

capacity of cooked meat. Also, this result supports the report of Kauffman

et al. (1961) who found that curing and cooking shrinkage was lower in pork

muscle with high pH values.

The pH of the L. dorsi steaks at time of cooking and cooking time

(Min/100 g wt.) were negatively and significantly (P<.01) related with a corre-

lation coefficient of -.54.

Correlation coefficients between initial internal steak temperature and

cooking loss and cooking time were -.63 and -.52, respectively; both corre-

lations were highly significant (P<. 01).

Time post-mortem showed significant (P<.01) relationships with cooking

loss and cooking time, with correlation coefficients of .52 and .46, respectively.

Per cent cooking loss was significantly (P<. 01) related to free and bound

moisture, with correlations of .46 and .43, respectively. A significant (P<.01)

correlation of .51 between free moisture and cooking time was found.

Cooking loss was also influenced by the cooking method; this was demon-

strated by the significant (P<.01) correlation of .26 found between cooking

method and per cent cooking loss; though significant, the correlation was low.

These data are in agreement with those reported by Paul et al. (1952).

Simple and multiple correlations and regression coefficients for factors

influencing and/or associated with cooking loss are presented in Table 12.

In this study pH of the L. dorsi at time of cooking was associated with

39.69% of the variability in per cent cooking loss and was the first variable en-

tering the step-wise regression analysis. The partial regression coefficient for





78





o
t-










0 0 -- 0 0
m u O
-a 0-








0
b w, Lo < O










-00 0
o 0 t 0
(n0 .0 .o .
U z o O)

$d C- 0 0 c'
CD t: C'1 C
SOn C o t
CU 4 .0 t 0 Id 0


0 ; c4 C: C' C


0o 00 0, CO to




-c CI 0 'UD m 11





0o Co Co O 0 C
CU




0! os in c cl


0 0















0 0*
u u1


FS . S o d.

o a c a iu




s 0 0 *tiE





0. U U E-C H







the relationship of pH and cooking loss indicated that pH would have to be de-

creased .5 unit to increase cooking loss by 3.43%. When cooking method was

added to the equation, an increase in accountable variability of 6.55% was noted.

The addition of cooking time in the analysis accounted for an additional 8.52%

of the variability in estimating per cent cooking loss. Finally, the addition of

time post-mortem and internal steak temperature to the equation would account

for an additional 3 per cent of the variability, which is quite low.


Post-mortem changes in juiciness of the Longissimus dorsi steaks

Mean values of panel juiciness scores for the broiled and deep fat fried

steaks are presented in Table 13. Statistical analysis (Table 14) of the taste

panel scores showed that the differences in juiciness between the four treatment

groups were not significant. Average post-mortem changes in juiciness scores

for the 36 animals used are shown graphically in Fig. 13.

Differences in average juiciness scores between the 1, 24, 48 and 192-hr

post-mortem time were significant at the .005 level of probability. Average

juiciness scores at the four post-mortem intervals were higher for the broiled

than for the deep fat fried steaks. This observation was statistically significant

at the .01 level of significance. However, the time post-mortem X cooking

method interaction was insignificant.


The relationship between juiciness and bound moisture, cooking loss, cooking
method and time post-mortem

The relationships between average panel juiciness scores, for the 288

broiled and deep fat fried L. dorsi steaks, and factors that had been reported





80







44 m 00 to 00 0
00 co 0 00 oo







-I-
SC- 0 00 0-0






0))

Cd LO Lo 1 4 O

0 CO


o C 0
2 LO t i O








2(O O LO LO (O co 1

0 t

0






0 (0 10 10 C ,10-
o




CO 10 C- C lO lO C-


a) )c 0m
3 C0
0C
Cu 02 t' 10 02 01 o 0
0 0 OOC .

c a
. r_ Eo eq m l .







41 cl 0






E) 0 a 2 U) V)
Cu Cu
C) 1444.1
Q~oo5 00




5o 0 )
0 ~ 3
H U Hm
















r-4


Lo n
cIJ ( 0
~z ~ Z


4 -
0
3 0

cI
Cd
. *^ < ,

c~r


oo o~G t


mC m-I mM ( mT 0) C o CM) CD
SC C











CdS
0)







0 d Cd o c
4 0)0
0





I 1K^




r-4 0 6)
S S S S *s




0 Cd 0
< uH H H m XH
S8 a ix' tt ^ '~ *



o 'B ed X r
'o ; a a 'o ^ ^ 'o
M ~~~~~~~ k* Q, Q,( <> *-a <> ?


In
o
o








o


*
0 00 M CM 0 C 0
0o m On t- co N > t- m
C .-4 iLO '-4


0o














-S


"C
















r-4
CO




0
CO
-


h

,.




o
ct

Oi
t-





0


o

0
0




0
02















0
o



O



ed

o






o
.4
rt




03
.0




C








U,
0
S
'.4










U,
V
0
T~3




















Q,
U,
'.
a,

















0
ho
























4-'
C
4-I
rt















38039S SS3N/ID/n














S...... ................ .......
V)

>- m




19'7 ;0/ o,



0 w -


co







*-
N ccr
u o,



: E.

10 o 0 _-



- m Q'-

Sa.





..,,,-,,


0 0 10 0
JO0 Sn L3NI F


3800S SS3N13InP








to influence and/or to be associated with juiciness, are presented in Table 15.


Table 15. Simple correlation coefficients between juiciness
and bound moisture, cooking loss, cooking method and time post-mortem.


Juiciness

Bound moisture, % -.26**

Cooking loss, % -.16**

Cooking method -.16**

Time -.11


*P<.01= .15
**P<.05 = .12


The simple correlation between average juiciness scores and bound

moisture of -.26 was highly significant (P<. 01) indicating a definite relationship

with juiciness. This result substantiates Hamm (1960) who reported that bound

water may be related to juiciness of meat. Also, this finding agrees with that of

Hardy and Noble (1945), who found a highly significant correlation between percent-

age of press fluid and juiciness scores of pork loin roasts. Also, Gaddis et al.

(1950) and Ritchey and Hostetler (1964) found low correlation coefficients between

subjective scores for juiciness and either free or bound water. However, this re-

sult is in contrast to the report of Ritchey (1965) who found no significant correlation

between subjective scores for eating quality in two beef muscles and either bound

or free water.

A low, but significant (P<.01) correlation of -.16 between average

juiciness scores and cooking loss was found.








The simple correlation between juiciness and cooking method of -.16

was low but highly significant (P<. 01) indicating that cooking method had some

effect on juiciness.

Even though the differences in average juiciness scores between the

four post-mortem times were significant, time post-mortem was found to have

no significant relationship with juiciness, with a low negative correlation of -.11.

Simple and multiple correlations and partial regression coefficients for

the most important factors influencing or associated with average juiciness scores

are presented in Table 16.

Bound moisture was the first variable entered and accounted for 6.76

per cent of the variability in juiciness. A one-unit increase in juiciness score

would require 6.24 per cent increase in bound moisture. When cooking loss was

combined with bound moisture in the step-wise regression analysis, the multiple

correlation coefficient was increased from .26 to 40. This accounts for an

additional 9. 24 per cent of the variation in juiciness scores. When time post-mortem

and cooking method were combined with bound moisture and cooking loss in the step-

wise regression analysis, the multiple correlation coefficient was increased from

. 40 to .45. The final coefficient of 45 would account for 20.25 per cent of the

variability in juiciness.


Post-mortem changes in flavor of the Longissimus dorsi steaks

Mean values and analysis of variance for panel flavor scores are presented

in Tables 17 and 18, respectively. Analysis of variance revealed that treatment had

no significant effect on the average flavor scores. Therefore, the flavor data ob-

tained from the 36 animals were considered as one group. Figure 14 graphically




University of Florida Home Page
© 2004 - 2010 University of Florida George A. Smathers Libraries.
All rights reserved.

Acceptable Use, Copyright, and Disclaimer Statement
Last updated October 10, 2010 - - mvs