Effects of Chronological Age and Postmortem Aging on Thermal

Shrinkage Temperature of Bovine Intramuscular Collagen

M. D. Judge and E. D. Aberle
J ANIM SCI 1982, 54:68-71.

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EFFECTS OF CHRONOLOGICAL AGE AND POSTMORTEM AGING

ON THERMAL SHRINKAGE TEMPERATURE OF BOVINE
INTRAMUSCULAR COLLAGEN 1'2
M. D. Judge and E. D. Aberle
Purdue University, West Lafayette, IN 47907
sues are changed in some way during postmortem aging to diminish their recovery. Kruggel
and Field (1971) and Pfeiffer et al. (1972)
showed that two processes that tenderize meat,
i.e. prerigor stretching and postmortem aging,
change the molecular structure of collagen.
They observed high correlations between the
quantity of each collagen chain component and
muscle shear value.
Davey and Gilbert (1975) studied the
toughening that occurs during meat cooking
and observed that shear resistance increases in
two phases, the second of which is accompanied by muscle shortening. They referred to
this shortening as "cooking shortening" and
suggested that it occurs as a consequence of
thermal shrinkage of the intramuscular collagen. On the other hand, the earlier work of
Machlik and Draudt (1963) indicated that collagen shrinkage is a tenderizing action in meat.
These conflicting reports might be explained by
the development of "residual strength" in
collagen after shrinkage and the resultant
aggregation and hardening of myofibrillar proteins, as proposed by Bailey and Sims (1977).
In their view, varying amounts of tension may
be generated in mature collagen after thermal
shrinkage because of the presence of thermally
stable intermolecular crossqinks. These crosslinks may be responsible for the tendency for
relatively tough muscles to have intramuscular
collagen with relatively high shrinkage temperature (Field et al., 1970).
Judge et al. (1981) reported that beef carcass electrical stimulation reduces the thermal
shrinkage temperature (Ts) of muscle collagen
in some cattle and, on the basis of tenderizing
action observed, suggested that the stimulation
may result in cleavage of some heat-stable
collagen cross-links.
Information is needed on the extent to
which properties of intramuscular collagen are
influenced by antemortem or postmortem
events known to be associated with tender-

Summary

The thermal shrinkage temperature of intramuscular collagen from cattle of widely different ages was determined at different times
postmortem. Differential scanning calorimetry
performed on physically isolated connective
tissue samples revealed that intramuscular
collagen of relatively old animals shrinks at
slightly higher temperatures than that of relatively young animals. These age-related differences probably resulted from the existence of
varying numbers of total and(or) heat-stable
intermolecular cross-links. A postmortem decline in thermal shrinkage temperature was observed in samples obtained at 45 min, 24 h and
7 d postmortem. The decline may have resulted
from changes in number of total and(or) heatstable cross-links but apparently was not caused
directly by muscle pH decline. Collagenolytic
cathepsins may have attacked collagen structures or ion shifts in the postmortem period
may have disrupted collagen structures by
direct effects on or by dehydration of the
collagen fibril.
(Key Words: Collagen Shrinkage, Chronological
Age, Postmortem Aging.)
Introduction

Recent reports of collagen alterations by

numerous meat processing procedures have provided a basis for renewed interest in connective
tissue. McClain et al. (1970) showed that the
amount of intramuscular connective tissue that
can be isolated by physical means from bovine
and porcine muscles declines with postmortem
aging. They concluded that the connective tis-

1Journal Paper No. 8476 of the Purdue Agr. Exp.

Sta., Dept. of Anim. Sci.
2The technical assistance of Anna Marie Bracker is
gratefully acknowledged.
68

JOURNAL OF ANIMAL SCIENCE, Vol. 54, No. 1, 1982

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THERMAL SHRINKAGE OF BOVINE MUSCLE COLLAGEN

69

TABLE 1. THERMAL SHRINKAGE TEMPERATURE OF BOVINE

LONGISSIMUS INTRAMUSCULAR COLLAGEN

Group

Angus cows
Holstein cows
Hereford, Angus and
Hereford Angus
steers and heifers
Holstein steer calves

Animal
age, mob

Time postmortema
45 rain

6
9

77 (20)
53 (22)

70.2 t .4cxy
70.4 -+ .3 cx

13
4

16 (1)
4 (0)

69.3 +- .3cy
67.0 -+ .5 cz

24h

7d

Shrinkage temperature, C
66.4 t .4dx
62.3 -+.4ex
67.6 + .3dx
63.7 +-.3ey
64.9 -+.3dy
61.1 + .5 dz

61.1 -+.3ex
59.1 -+.5ez

aMean +-standard error.

bMean (standard deviation).
c d e .

' ' means mtne same row bearing different superscripts differ (P<.05).

x'Y'ZMeans in the same column bearing different superscripts differ (P<.05).

ness. This research was designed to determine

the Ts of muscle collagen with presumably different numbers or types of intermolecular
cross-links (from animals of different ages) and
to determine the effects of postmortem aging
on the Ts of muscle collagen.
Materials and Methods

Six Angus cows (54 to 113-too-old), nine

Holstein cows (27 to 92-too-old), 13 Angus,
Hereford and Angus Hereford crossbred
slaughter steers and heifers (13 to 17-too-old)
and four Holstein steer calves (4 mo old) were
transported directly from production facilities
to the Purdue University Meat Science Laboratory for slaughter. The carcasses were chilled
at approximately 0 C for 24 h and stored at
approximately 3 C thereafter.
At 45 rain, 24 h and 7 d postmortem,
longissimus muscle samples were obtained from
the second to third lumbar region of the carcasses. The samples were frozen in liquid
N after removal of the epimysial connective
tissue and stored at - 2 5 C for as long as 4
wk.
The frozen samples were subjected to the
connective tissue isolation procedure of McClain (1969). Samples were warmed to approxi-

Dupont Model 990, E. I. DuPont de Nemours and

Co., Inc., Concord Plaza McKean Bldg., Wilmington,
DE 19898.

mately - 5 C, mixed with ground Dry Ice and

powdered in a Waring blender. The powdered
samples were passed through 10-, 14-, 1 6 - a n d
25-mesh sieves. Connective tissue was recovered
from each sieve and stored at - 2 5 C for as long
as 4 wk.
The Ts of the collagen was determined as
described by Judge et al. (1981) with a thermal
analyzer 3 equipped with a differential scanning
calorimeter cell.
Prerigor connective tissue samples from six
randomly selected samples taken from the 13
to 17-too-old steers and heifers were used in
additional experiments conducted to determine
the effects of pH and NaC1 on collagen Ts.
Samples were placed in .05 M KH2PO4-NaOH
buffer adjusted to pH 5.5, 5.9, 6.3, 6.7 and 7.1.
Likewise, samples were placed in pH 7.1 buffer
to which NaCI had been added at concentrations of 1, 2, 3, 4 and 5% (w/v). The samples
were exposed to the respective buffer solutions
for 5 min at approximately 5 C after which the
Ts was determined. Preliminary experiments
demonstrated that immersion in the buffer
solutions for as long as 24 h did not alter
T s beyond that observed for 5 min exposure
time.
The data were subjected to the Harvey
(1960) least-squares analysis over the different
animal breed-age groups and over the three
times postmortem.
Significant differences
among means were determined by the NewmanKeuls sequential range test as described by
Snedecor and Cochran (1967).

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70

JUDGE AND ABERLE

TABLE 2. EFFECT OF pH ON THERMAL
SHRINKAGE TEMPERATURE OF
BOVINE LONGISSIMUSPRERIGOR
I N T R A M U S C U L A R COLLAGENa
Shrinkage
temperature, C

Treatment

TABLE 3. EFFECT OF NaCI CONCENTRATION

ON THERMAL SHRINKAGE TEMPERATURE
OF BOVINE LONGISSIMU8PRERIGOR
INTRAMUSCULARCOLLAGENa

Treatment

Shrinkage
temperature, C

Control b

69.4 .3e

Control b

67.3 .3cd

Buffer pH
5.5
5.9
6.3
6.7
7.1

66.6
66.8
66.8
67.3
67.3

NaCI, %
1
2
3
4
5

66.5
66.6
66.9
67.5

+ .3d
.3d
.3d
.3d
+ .3d

aMean standard error of least squares mean; n =

6.

68.2

+ .3 c
.3c
.3c
+ .3cd
.3 d

aMean -+ standard error of least squares mean; n =

6.

bsamples in distilled water.

C'dMeans bearing different superscripts differ
(P<.01).

Results and Discussion

Small, b u t statistically significant differences

in mean collagen Ts among the different breed
and age groups were observed, and, in agreement with the results of Goll et al. (1964), the
Ts was higher in tissues from older animals
(table 1). Field et al. (1970) found no agerelated differences in Ts of epimysial collagen
but noted that epimysial collagen shrinks at
lower temperatures than intramuscular collagen.
The differences in Ts value among the
various breed-age groups were modest compared to the differences observed with time
postmortem (table 1). A reduction in collagen
Ts occured both before and after 24 h postmortem. The changes occurring before 2 4 h
postmortem might have resulted from cold
shortening-induced disruption of collagen crosslinking (O'Shea et al., 1974) if cold shortening
occurred in the samples. Otherwise, the alteration in collagen T s may not have been directly
associated with early postmortem events, since
glycolytic activity and rigor mortis onset are
largely complete within 24 h postmortem in
bovine muscle. In fact, the exposure of prerigor collagen to phosphate buffers of various
pH lowered the Ts by amounts that were independent of pH (table 2). A simple pH effect
on collagen cross-linking and Ts value, therefore, does not seem likely in intact muscle tissue, but it is possible that certain collagenolytic

bsamples in pH 7.1 phosphate buffer.

C'dMeans bearing different superscripts differ
(P<.01).

cathepsins that attack cross-links in the nonhelical region of the molecule were activated
by acidic conditions within the muscle (Burleigh et al., 1974).
Ions other than hydrogen i o n s in the
variable-pH buffer solutions may have effectively reduced the collagen Ts. In contrast,
increasing concentrations of NaC1 ions at constant pH increased the Ts of the collagen (table
3). Further research is needed to evaluate the
effects of specific ions on collagen Ts.
McClain et al. (1970) reported a slight, but
nonsignificant reduction in Ts with time postmortem up to 72 h in bovine longissimus collagen. However, the yield of intramuscular connective tissue declined significantly with time
postmortem when extracted by the method
used in the present study. Although not quantitared, the yield of connective tissue in our
study likewise appeared to be greater in samples
secured at 45 rain postmortem than in those
secured at later times. Aging may have rendered
the collagen network more easily fragmented
and consequently, more difficult to recover.
McClain and Wiley (1972) reported that the
transition temperature of intact bovine muscle
collagen is several degrees higher than that of
tropocollagen extracts of the same tissue.
We suggest that the collagen Ts differences
among the breed-age groups were due to the
presence of different numbers of total and(or)
heat-stable intermolecular cross-links. These

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THERMAL SHRINKAGE OF BOVINE MUSCLE COLLAGEN

n o n r e d u c i b l e c o v a l e n t b o n d s d e v e l o p slowly
f r o m heat-labile r e d u c i b l e b o n d s , w h i c h increase in n u m b e r f r o m b i r t h to 12 to 18 m o o f
age a n d decrease t h e r e a f t e r in b o v i n e muscle
collagen ( S h i m o k o m a k i et al., 1972).
T h e findings p r o v i d e c i r c u m s t a n t i a l e v i d e n c e
t h a t collagen c r o s s 4 i n k i n g is d i m i n i s h e d d u r i n g
p o s t m o r t e m aging o f b o v i n e muscle, as sugg e s t e d b y Kruggel a n d Field ( 1 9 7 1 ) a n d Pfeiffer
et al. ( 1 9 7 2 ) . O n t h e o t h e r h a n d , we c a n n o t
i g n o r e t h e possibility t h a t t h e p o s t m o r t e m muscle e n v i r o n m e n t i n c l u d e d factors t h a t l o w e r e d
Ts b y degrading t h e m o l e c u l a r s t r u c t u r e o f
collagen. Wu ( 1 9 7 8 ) o b s e r v e d t h a t muscle tissues i n c u b a t e d at pH 5 yielded h i g h e r q u a n t i ties o f n e u t r a l salt soluble collagen t h a n t h o s e
i n c u b a t e d at pH 7 a n d t h e o r i z e d t h a t t h e differe n c e m i g h t have b e e n d u e to a release o f m o r e
collagen m o n o m e r s o r t o an increased susceptib i l i t y of d e p o l y m e r i z e d collagen to p r o t e a s e
d e g r a d a t i o n . Privalov a n d T i k o p u l o ( 1 9 7 0 )
r e p o r t e d t h a t t h e a d d i t i o n of NaC1 t o t r o p o collagen s o l u t i o n s r e s u l t e d in t w o discrete
stages o f h e a t u p t a k e u p o n h e a t i n g at a rate o f
.2 C / m i n . T h e y suggested t h a t t h e s t a b i l i z a t i o n
o f collagen p r o b a b l y is d e p e n d e n t o n a regular
w a t e r s t r u c t u r e a d j a c e n t t o t h e m o l e c u l e . In t h e
p r e s e n t s t u d y , t h e loss o f s a r c o l e m m a i n t e g r i t y
a n d s u b s e q u e n t r e d i s t r i b u t i o n o f ions postm o r t e m m a y have e x p o s e d t h e collagen to a
c h a n g i n g ionic e n v i r o n m e n t or to d e h y d r a t i o n .
A d d i t i o n a l research is n e e d e d to e l u c i d a t e t h e
cause(s) o f t h e p o s t m o r t e m changes o c c u r r i n g
in Ts o f b o v i n e muscle collagen.
Literature Cited

Bailey, A. J. and T. J. Sims. 1977. Meat tenderness:

Distribution of molecular species of collagen in
bovine muscle. J. Sci. Food Agr. 28:565.
Burleigh, M. S., A. J. Barrett and G. S. Lazarus. 1974.
Cathepsin 81 : A lysosomal enzyme that degrades
native collagen. Biochem. J. 137:387.
Davey, C. L. and K. V. Gilbert. 1975. Cooking shortening and the toughness of beef. J. Food Technol. 10:333.

71

Field, R. A., A. M. Pearson and B. S. Schweigert.

1970. Hydrothermal shrinkage of bovine collagen. J. Anita. Sci. 30:712.
Goll, D. E., W. G. Hoekstra and R. W. Bray. 1964.
Age-associated changes in bovine muscle connective tissue. II. Exposure to increasing temperature. J. Food Sci. 29:615.
Harvey, W. R. 1960. Least-squares analysis of data
with unequal subclass numbers. USDA, ARS 20:
8.
Judge, M. D., E. S. Reeves and E. D. Aberle. 1981.
Effect of electrical stimulation on thermal shrinkage temperature of bovine muscle. J. Anim. Sci.
52:530,
Kruggel, W. G. and R. A. Field. 1971. Soluble intramuscular collagen characteristics from stretched
and aged muscle. J. Food Sci. 36:1114.
Machlik, S. M. and H. N. Draudt. 1963. The effect of
heating time and temperature on the shear of
beef semitendinosus muscle. J. Food Sci. 28:
711.
McClain, P. E. 1969. Isolation of intramuscular connective tissue. Nature 221 : 181.
McClain, P. E., G. J. Creed, E. R. Wiley and I. Hornstein. 1970. Effect of postmortem aging on isolation of intramuscular connective tissue. J. Food
Sci. 35:258.
McClain, P. E. and E. R. Wiley. 1972. Differential
scanning calorimeter studies of the thermal transitions of collagen. J. Biol. Chem. 247:692.
O'Shea, J. M., P. V. Harris, W. R. Shorthose and P. E.
Bouton. 1974. Changes in the thermal stability of
intramuscular connective tissue and mechanical
properties of bovine muscle concomitant with
changes in myofibrillar contraction state. J. Food
Sci. 39:1221.
Pfeiffer, N. E., R. A. Field, T. R. Varnell, W. G. Kruggel and I. I. Kaiser. 1972. Effects of postmortem
aging and stretching on the macromolecular properties of collagen. J. Food Sci. 37:897.
Privalov, P. L. and E. I. Tikopulo. 1970. Thermal conformational transformations of tropocollagen. I.
Calorimetric study. Biopolymers 9:127.
Shimokomaki, M., D. F. Elsden and A. J. Bailey.
1972. Meat tenderness: Age related changes in
bovine intramuscular collegen. J. Food Sci. 37:
892.
Snedecor, G. W. and W. G. Cochran. 1967. Statistical
Methods, (6th Ed.). The Iowa State Univ. Press,
Ames.
Wu, J. ]. 1978. Characteristics of bovine intramuscular
collagen under various postmortem conditions.
Ph.D. Dissertation, Texas A&M Univ., College
Station.

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