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 Experimental procedure
 Results and discussion
 Summary
 Literature cited






Group Title: Animal husbandry and nutrition mimeograph series - University of Florida Animal Husbandry and Nutrition Dept. ; no. 61-5
Title: The Effect of ante-mortem injection of enzymes on the tenderness of beef cattle
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 Material Information
Title: The Effect of ante-mortem injection of enzymes on the tenderness of beef cattle
Series Title: Animal husbandry and nutrition mimeograph series
Physical Description: 10 leaves : ; 28 cm.
Language: English
Creator: Huffman, D. L.
Palmer, A. Z.
Carpenter, J. W.
Shirley, R. L.
Publisher: Florida Agricultural Experiment Station
Place of Publication: Gainesville, Fla.
Publication Date: 1961
 Subjects
Subject: Beef -- Quality -- Florida   ( lcsh )
Enzymes -- Research -- Florida   ( lcsh )
Genre: bibliography   ( marcgt )
government publication (state, provincial, terriorial, dependent)   ( marcgt )
non-fiction   ( marcgt )
 Notes
Statement of Responsibility: D.L. Huffman ... et al..
Bibliography: Includes bibliographical references (leaf 10).
General Note: Caption title.
General Note: "January, 1961."
Funding: Animal husbandry and nutrition mimeograph series - University of Florida Animal Husbandry and Nutrition Dept. ; no. 61-5
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Bibliographic ID: UF00072904
Volume ID: VID00001
Source Institution: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
Resource Identifier: oclc - 77078510

Table of Contents
    Experimental procedure
        Page 1
        Page 2
    Results and discussion
        Page 3
    Summary
        Page 4
        Page 5
        Page 6
        Page 7
        Page 8
        Page 9
    Literature cited
        Page 10
Full Text



Animal Husbandry and Nutrition Florida Agricultural
Mimeograph Series No, 61-5 Experiment Station
January, 1961 Gainesville, Florida

THE EFFECT OF ANTE-MORTEM INJECTION OF ENZYMES
ON THE TENDERNESS OF BEEF CATTLE I/

D. L. Huffman, A. Z. Palmer, J. W. Carpenter and R. L. Shirley /


The ability of proteolytic enzymes to tenderize beef is well established.
Gottschall and Kles (1942), Wang and Maynard (1955), Tappel etj al (1956) and
Welr et al. (1958) have indicated that one of the problems inherent in the use
of papain in tenderizing beef cuts is that of uniform penetration of the enzyme
through the tissue. Palmer et al. (1957) investigated the effect of crude papain
solutions injected intraperitoneally, 12 and 24 hours ante-mortem on the tender-
ness of chickens and noted a tenderizing effect when the aqueous solution of
crude papain was injected at the rate of 100 ppm of live animal weight. Buek
et al. (1959) working with cattle, sheep, goats and chickens reported a signifi-
cant tenderizing effect when papain solutions were injected intravenously from
one minute to two hours prior to slaughter. Huffman et al. (1961) reported a
significant tenderizing effect from the injection of 100 ppm of crude papain
into chickens six hours ante-mortem; further, ash of crude papain showed a
tenderizing effect and when the ash of crude papain was combined with one ppm
crystalline papain an apparent synergistic effect was noted.

The purpose of this study was to determine the effect of ante-mortem
injections of crude and crystalline papain on the tenderness of beef cattle.
A further objective was to determine whether or not an enzyme activator was
present in the crude papain and if present, whether or not the activator would
be effective as a tenderizing agent alone or only when injected with enzyme.

ExDerimenta! Procedure

Crude and crystalline papain solutions were assayed for activity, in vitro,
by the method described by Weiner (1956). Injection levels were based on the
enzyme activity determined by this assay. The crystalline papain used in this
study was prepared by the method of Balls and Lineweaver (1939). The crystalline
preparation was in a 1:100 dilution of a 0.1 M cysteine solution.

A pilot trial was conducted to learn injection techniques and determine
if harmful effects resulted from ante-mortem injection. Eight cattle were
injected intravenously with varying levels of crude papain and crude papain
ash 30 minutes ante-mortem. Inspection made prior to, and following slaughter
showed no apparent effect of treatment. Livers of animals receiving crude
papain were over tenderized, but edible.

SThe cooperation of Central Packing Co., Center Hill, Florida, in supplying
the cattle used in this study is acknowledged with appreciation. The
assistance of Dr. L. E. Swanson, Department of Veterinary Science, Univer-
sity of Florida and Dr. R. H. Bennett, Chief of Meat Inspection Section of
the Florida Livestock Board in this study is acknowledged.
2/ Huffman, Research Assistant; Palmer, Associate Animal Husbandman; Carpenter,
Assistant Animal Husbandman; R. L. Shirley, Animal Husbandman; Department
of Animal Husbandry and Nutrition, University of Florida, Gainesville,
Florida. The assistance of Mrs. Normal Bowles and Mr. Howard Povey is
acknowledged.








Experimental Animals


Fifty-six cattle of Brahman and Brahman X Angus breeding were randomly
divided into eight iots with seven cattle in each lot. The cattle were out of
a single commercial feed lot, averaged 640 pounds in weight and graded from
low Standard to top Good on the rail.

Treatments

The cattle were randomly assigned to the following treatments: Lot I -
non-injected control; Lot 2 0.1 ml. physiological saline/lb. live wt.; Lot 3 -
10 mgs, crude papain/Ib. live wt.; Lot 4 10 mgs. crude papain ashed/Ib. live
wt.; Lot 5 20 mgs. crude papain ashed/Ib. live wt.; Lot 6 10 mgs. denatured
crude papain/lb. live wt.; Lot 7 0.2 mg. crystalline papain/Ib. live wt.;
Lot 8 0.2 mg. crystalline papain plus 10 mgs. crude papain ashed/Ib. live wt.

Preparation of Solutions

Solutions for treatments 3, 4, 5 and 6 were 10% solutions of crude papain
in 0.9% saline; solution injection was at the rate of IC mis. per hundred
pounds live weight. The solution for treatment 7 was prepared by mixing 150.94
mgs. crystalline papain with 100 ml. of 0.9% saline; solution injection was
at the rate of 13.25 mis. per hundred pounds liveweight. The solution for
treatment eight was prepared by mixing 150.94 mgs. crystalline papain with
each 100 mis. of solution previously prepared for Lot 4; solution injection was
at the rate of 13.25 mis. per hundred pounds live weight.

The ashed crude papain for treatments 4, 5 and 8 was prepared by weighing
out the desired amount of crude papain, ashing in a 6000 C. furnace for 12
hours, then the solution was prepared from the resultant ash. The crude papain
denatured solution for treatment six was prepared from a 10% solution of crude
papain in 0.9% saline by placing the solution in a 970 C. water bath for 30
minutes. The pH of all solutions was adjusted to 7.3 with 0.1 N HCI and Na2CO3.

Injection Technique

The cattle were restrained in a squeeze chute and the head held to the side
exposing the jugular vein. The injection was made with an 18 guage, one-inch
needle and a glass syringe. Each animal was held 30 minutes in a holding pen
from time of injection to time of slaughter.

Slaughter and inspection

Veterinarian inspectors provided thorough ante-mortem and post-mortem
inspection.

Tenderness Determinations

Warner-Bratzler shear values on steaks and roasts were obtained on one-half
inch cores. Taste panel tenderness values were on a one to nine hedonic scale,


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with I being inedible, 5 average tenderness, 7 very tender, 8 extremely tender -
grainy and 9, mushy over tenderized.

Liver slices, taken from the livers after a 24 hour chill, were pan fried
and tested for tenderness by a four member panel. Kidney slices were cooked
in covered petri dishes in a 3600 F. oven for one hour. The cooked samples were
quartered and tested for tenderness by a four member panel.

One inch steaks were removed from the short loin and top round of each
carcass, wrapped and frozen until tested. The club steaks were broiled to a
medium well done degree (1600F.). Round steaks were braised well done.

A four-inch rib roast from each carcass was wrapped and frozen until tested.
Roasts were cooked in a 3500 F. oven to an internal temperature of 1600 F.

Portions of the livers from the crude and crystalline papain treatments
were made into liver sausage to determine if the livers would make a product
acceptable to consumers. Livers from the non-injected treatment were used
to make an equal amount of liver sausage to serve as a control. Samples of each
formulation of liver sausage were distributed among 32 families to rate on a
one to nine scale as to color, flavor and texture.

Statistical Analysis

The experiment was based on a completely randomized design. Data were
statistically treated by analysis of variance as described by Snedecor (1956).
Duncan's Multiple Range Test according to Duncan (1955) was used to compare
treatment means when significant differences were noted by analysis of variance.

Results and Discussion

Ante-mortem and post-mortem inspection of the cattle, carcasses and parts
did not reveal abnormalities which could be attributed to treatment.

The results of taste panel evaluations of the livers are presented in
Table 1. Livers from cattle injected with crude papain (treatment 3) were
significantly more tender (P<0.01) than all other treatments. Livers from
cattle receiving crystalline papain plus crude papain ash were tenderized
compared to the saline injected controls (P<0.05).

The results of taste panel evaluation of the kidneys are reported in
Table 2. In general, kidneys from cattle treated with active enzyme were some-
what over tenderized and they lacked body and texture to be acceptable by the
taste panel.

The results of the taste panel evaluation of the club steaks are reported
in Table 3. Treatments 3, 4, 7 and 8 were significantly more tender than the
non-injected control (P<0.05). Warner-Bratzler shear values of club steaks
showed that ail injected treatments were significantly more tender (P4 than the non-injected control group.


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The results of taste 'panbl and shear '"dt~hMinations of the braised round
steak are presented in TWabl 4. D'i frerices were not significant among
the mean scores.

The treatment means for panel and shear determinations on rib roasts are
presented in Table 5. Differences noted were not significant. It is of
interest to note the similarity of treatment effect on tenderness between rib
roast, braised round and club steak tenderness data.

In the consumer preference study with liver sausage, 23 families preferred
the flavor of the treated liver sausage, seven preferred the flavor of the
non-treated and two had no preference. Seventeen families preferred the color
of the treated liver sausage, II preferred the non-treated and four had no
preference. Seventeen families preferred the texture of the treated liver
sausage, 12 preferred the non-treated and three had no preference. These
results indicate that the livers from cattle injected with enzymes could be
processed into a marketable liver sausage.


Summary

Fifty-six crossbred cattle were randomly divided into control and treat-
ment groups; treated groups were injected intravenously with varying levels
of papain and papain derivatives to compare the effects of crude and crystalline
papain and crude papain derivatives on the tenderness of beef cattle. Ante-
mortem and post-mortem inspection did not reveal abnormalities attributable
to treatment. The liver and kidneys of cattle injected with active enzymes
were somewhat over-tenderized. Liver sausage made from livers of papain
injected cattle was found acceptable in flavor, color and texture.

Club steak, round steak and rib roast taste panel and Warner-Bratzler
shear data showed similar effects of ante-mortem treatment on tenderness.
The crude and crystalline papain treated and crystalline papain with ashed
crude papain treated lots of cattle were generally more tender than lots
receiving other treatments. Ashed crude papain produced a degree of teoder-
ization when injected at the same level as the crude papain. An inhibition
of this tenderizing effect was noted with a higher level of crude papain ash.
When the crude papain ash was combined with the crystalline papain a com-
plimentary effect was noted indicating that an enzyme in the crystalline
papain solution may have been activated by a component of the ashed crude
papain. Heat denaturation of crude papain did not result in as marked degree
of tenderization as that noted from the ashed crude papain. This fact lends
additional support to the theory that an enzyme activator is present in the
crude papain ash.


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TABLE I

THE EFFECT OF ANTE-MORTEM INJECTION OF PAPAIN AND PAPAIN DERIVATIVES ON


Ml. of 0.9%
Treatment Saline/100 lbs.
Live Wt.


Non-injected control

Saline-injected control 10

Crude papain, 10 mgs./lb. live wt. 10

Crude papain, ashed, 10 mgs./lb.
live wt. 10

Crude papain, ashed, 20 mgs./lb.
live wt. 10

Crude papain, denatured, 10 mgs./lb.
live wt. 10

Crystalline papain, 0.2 mg./lb. live
wt. 13.25

Crystalline papain, 0.2 mg./lb. plus
crude papain ashed, 10 mgs./Ib. live wt. 13.25


More tender than all other treatments (P<0.01)

More tender than 2 (P<0,05)


Number
of
Animals


7

7

7


7


7


7


7


7


TENDERNESS OF LIVER


Average
Taste Panel
Scores


6.61

6.42

8.62 1


6.93


6.64


6.89


7.00


7.11 2


- -~~- -- --- ----cl----~-







TABLE 2

THE EFFECT OF ANTE-MORTEM INJECTION OF PAPAIN AND PAPAIN DERIVATIVES ON TENDERNESS OF KIDNEY


MI. of 0.9% Number Average
Treatment Saline/100 lbs. of Taste Panel
Live Wt. Animals Scores


1. Non-injected control -- 7 6.30

2. Saline-injected control 10 7 6.70

3. Crude papain, 10 mgs./lb. live wt. 10 7 8.20 I/

4. Crude papain, ashed, 10 mgs./ib.
live wt. 10 7 6.34

5. Crude papain, ashed, 20 mgs./Ib.
live wt. 10 7 6.01

6. Crude papain, denatured, 10 mgs./lb.
live wt. 10 7 6.01

7. Crystalline papain, 0.2 mg./lb. live
wt. 13.25 7 7.41 2.

8. Crystalline papain, 0.2 mg./lb. plus
crude papain ashed, 10 mgs./lb. live wt. 13.25 7 7.14./


More tender than 1, 2, 4, 5, 6 & 8 (P-0.01) and 7 (P<0.05)

More tender than 1, 4, 5, & 6 (P<0.01) and 8 (P<0.05)

More tender than 5 & 6 (P,0.0I) and I & 4 (P<0.05)







TABLE 3

THE EFFECT OF ANTE-MORTEM INJECTION OF PAPAIN AND PAPAIN DERIVATIVES


ON TENDERNESS OF CLUB STEAK


Treatment


MI. of 0.9%
Saline/100 Ibs.
Live Wt.


1. Non-injected control

2. Saline-injected control

3. Crude papain, 10 mgs./lb. live wt.

4. Crude papain, ashed, 10 mgs./lb
live wt.

5. Crude papain, ashed, 20 mgs./Ib.
live wt.

6. Crude papain, denatured, 10 mgs./Ib.
Ii ve wlt.

7. Crystalline papain. 0.2 mg./lb. live
wt.

8. Crustalline papain. 0.2 mg./lb. plus
crude papain ashed, 10 mgs./Ib. live wt.


I/ More tender than I (P<0.05)


10

10


10


10


10
I0


13.25


13.25


Number
of
Animals


7

7

7


7


7


7


7


7


Average
Taste Panel
Scores


3.86

4.91

5.54 -/


5.21 -/


4.79


4.89


5.11 L/


5.89 I/


Average
Shear
Scores


17.42

12.83 L/

II.64 I


12.15 J1


13.57 1/


12.80 1/


13.04 1/


10.87 I/







TABLE 4

THE EFFECT OF ANTE-MORTEM INJECTION OF PAPAIN AND PAPAIN DERIVATIVES ON TENDERNESS OF BRAISED


ROUND STEAK


Ml. of 0.9% Number Average Average
Treatment Saline/100 Ibs. of Taste Panel Shear
Live Wt. Animals Scores L/ Scores -1


I. Non- injected control -- 7 4.32 11.87

2. Saline-injected control 10 7 5.07 12.01

3. Crude papain, 10 mgs./lb. live wt. 10 7 5.58 9.96

4. Crude papain, ashed, 10 mgs./Ib.
live wt. 10 7 4.75 12.43

5. Crude pepain, ashed, 20 mgs./Ib.
live wt. 10 7 4.39 13.72

6. Crude papain, denatured, 10 mgs./lb.
live wt. 10 7 4.93 12.14

7. Crystalline papain, 0.2 mg./Ib. live
wt. 13.25 7 5.07 11.70

8. Crycta!line papain, 0.2 mg./lb. plus
crude papain ashed, 10 mgs./lb. live wt. 13.25 7 5.25 11.18


1/ Differences were not significant.







TABLE 5

THE EFFECT OF ANTE-MORTEM INJECTION OF PAPAIN AND PAPAIN DERIVATIVES


MI. of 0.9% Number
Treatment Saline/100 Ibs. of
Live Wt. Animals


I. Non-injected control -- 7

2. Saline-injected control 10 7

3. Crude papain, 10 mgs./Ib. live wt. 10 7

4. Crude papain, ashed, 10 mgs./lb.
live wt. 10 7

5. Crude papain, ashed, 20 mgs./lb.
live wt. 10 7

6. Crude papain, denatured, 10 mgs./Ib.
live wt. 10 7

7. Crystalline papain, 0.2 mg./lb. live
wt. 13.25 7

8. Crystalline papain. 0.2 mg./Ib. plus
crude papain ashed, 10 mgs./lb. live wt. 13.25 7


I/ Differences were not significant.


ON TENDERNESS OF RIB ROAST


Average
Taste Panel
Scores -


4.75

5.71

6.13


5.82


5.71


5.96


5.89


6.14


Average
Shear
Scores -1


11.70

9.47

8.20


8.95


10.75


8.74


8.48


9.85


I _









Literature Cited


Balls, A. K. and H. Lineweaver. 1939. Isolation and properties of crystalline
papain. Journ. Bioi. Chem., 130:669.

Beuk, Jack F., Alfred L. Savich, Paul A.Goeser and John M. Hogan. 1959.
Method of tendering meat. U. S. Patent 2,903,362. Issued September 8,
1959.

Duncan, D. B. 1955. Multiple range and multiple F tests. Biometrics J :1.

Gottschall, G. Y. and M. W. Kies. 1942. Digestion of beef by papain. Food
Res. 7:373.

Huffman, D. L., A. Z. Palmer, J. W. Carpenter and R. L. Shirley. 1961. The
effect of ante-mortem injection of papain on tenderness of chickens. Fla.
Agr. Exp. Sta.,An. Husb. and Nutr. Mimeo Series No. 61-4.

Palmer, A. Z., R. H. Alsmeyer and J. W. Carpenter. 1957. Effect of ante-
mortem injection of papain on the tenderness of mature chickens and beef.
Ann. Report, Fla. Agr. Exp. Sta.

Snedecor, G. W. 1956. Statistical Methods. (5th Edition). The Iowa State
College Press, Ames, Iowa.

Tappel, A. L., D. S. Miyada, Clarence Sterling and V. P. Maier. 1956.
Meat Tenderization. II. Factors affecting the tenderization of beef
by papain. Food Res. 21:375.

Wang, H. and Nehema Maynard. 1955. Studies on enzymatic tenderization of
meat. I. Basic technique and histological observations of enzymatic
action. Food Res. 20:587.

Weiner, S. 1956. Papain, a review of the literature. Paul Lewis Laboratory
publication.

Weir, C. Edity, H. Wang, Marion L. Birkner, Jean Parsons and Betty Ginger.
1958. Studies on the enzymatic tenderization of meat. II. Panel and
histological analyses of meat treated with liquid tenderizers containing
papain. Food Res. 23:441.

I0


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