55
CORROSION
OF BONE BY SOLUTIONS
RAPTOR GASTRIC JUICE*
SIMULATING
by
J. H. Cummings,G. E. Duke, and A. A. Jegers
Department of Veterinary Biology
Collegeof Veterinary Medicine
University of Minnesota
St. Paul, MN
55108
*This studywaspartiallysupportedby NationalScienceFoundationGrant No. NSF-GB37254.
ABSTRACT. To determinewhetherfalconiformsdigestthe bonesof their prey morethoroughlythan strigiformsbecauseof greatergastricacidity in falconiforms,mouseboneswere
incubatedin solutionssimulatinggastricjuice from the two orders.Solutionssimulatingthe
gastricjuice of falconiformswith a pH of 1.66 corrodedbones more extensivelythan
solutionssimulatingstrigiformgastricjuice with a pH of 2.35. Pepsin,at concentrations
rangingfrom 0 to 4 mg/ml, alsowereslightlyinvolvedin bonecorrosionat both pH's.
Introduction
Previousstudies have shown that hawks digest bone more exensivelythan do owls
(Errington 1932, Gladinget al. 1943, Clark 1972, Duke et al. 1975). Duke et al. (1975)
found that the gastricjuices of hawks and owls had approximatelyequal proteolyticactivities,but that hawk gastricjuice had a much lower pH than that of owls.They hypothesized that the difference in acidity may account for the greater corrosionof bones that
occursin the stomachof hawks.The purposeof this studywasto testthishypothesis.
Methods
To simulategastricjuices of falconiformsand strigiforms,two liters of Avian Ringer's
solutionwere treated in the followingmanner.Usingconcentratedhydrochloricacid, one
liter wasadjustedto pH 1.66 and the secondliter to pH 2.35, the meanpH's of samples
of
gastricjuice collectedpreprandiallyfrom severalspeciesof hawksand owls, respectively
(Duke et al. 1975). Solutionscontaining4 mg/ml of bovinepepsinwere preparedfrom
aliquotsof the Ringer'ssolutions,
and the aliquotswerereadjusted
to theirrespective
pH's.
This concentrationof pepsinapproximates
the proteolyticactivity of raptor gastricjuice
(Duke et al. 1975). Aliquotsof theselatter solutionswerethen dilutedto a concentrationof
2 mg/mlof pepsinusingthe Ringer'ssolutionswith pH's of 1.66 and 2.35. Thisprocedure
provided
threesolutions
at eachpH containing
(1) 4 mg/mlof pepsin,(2) 2 mg/mlof pepsin,
and(3) no pepsin.Five experiments
wereperformedusingthe latter two solutions,
andsix
experiments
wererun usingthe solutioncontaining4 mg/mlof pepsin(table 1).
In each experiment,bonesfrom 3 adult laboratorymice (Mus musculus),cleanedof
tissue by a dermestidbeetle colony, were used. The cleanedbones were washedin cold
water, oven-dried,
anddividedinto 2 groups:groupA contained1 femur, 1 scapula,2 ribs,2
each of caudal and lumbar vertebrae,and 1 tibia fibula; group B contained1 humerus,
Raptor Research10(2):55-57, 1976
56
RAPTOR RESEARCH
Vol. 10, No. 2
one-halfmandible,1 oscoxae,2 ribs,and2 eachof caudalandlumbarvertebrae.Eachgroup
of boneswasweighedon a numbered,preweighedfilter paper, and bone weight wascalculated. In order to minimize differencesin degreeof digestiondue to variation in bone size
and shape,bones from eachgroupwere addedto duplicate,numbered100 ml aliquotsof
eachof the six solutions(table 1), and the solutionswereincubatedfor 4 hoursin a heated
waterbath
at 39øC(theapproximate
bodytemperature
of hawksandowlsasdetermined
in
our laboratory).After incubation,the remaining
bonesin eachgroupwerecollectedon their
corresponding
numberedfilter papers(seeabove)by filtration. Filter paperandboneswere
driedat 60øFovernight
andweighed.
Theweightof thepaperplusbonewassubtracted
from their initial weight after dryingto estimatethe amount digested.
To determineif incubationor the presenceof bonesin the solutionmay havealteredthe
proteolyticactivityof the solutionswith 4 mg/ml of pepsin,proteolyticactivitywasdeterminedin the solutionsat eachpH (1)beforeincubationor the additionof bones,(2)after 4
hoursof incubationwith no bonesaddedto the solution,and (3) after additionof bonesand
4 hoursof incubation
at 39øC.Therelease
of tyrosinefromhemoglobin
uponadditionof
hemoglobinto the simulatedgastricjuice solutionswas usedas a measureof proteolytic
activity of the pepsin in the solutions.Tyrosine releasewas determinedby a modified
colorimetricmethod(Duke et al. 1975).
Results
and Discussion
Solutionsof pH 1.66 corrodedbonessignificantly
morethansolutions
of pH 2.35 (table
1) at all pepsinconcentrations.
Pepsinappearedto havea slighteffecton bonecorrosion
at
eitherpH.
Proteolyticactivitywas slightlygreaterin solutionsat pH 1.66 than in solutionsat pH
2.35 (table2). Fourhoursof incubation
andthepresence
of bonesin solutioneachappeared
to decreasethe proteolytic activity of the solutions.
The resultsof thesestudiessupportthe hypothesis
that the lowerpH of gastric
juiceis
principally
responsible
for greater
bonecorrosion
in falconiform
stomachs
thanin strigiform
stomachs
(Dukeet al. 1975).Theproteolyticactivityof thesimulated
gastric
juicesolution
apparentlywasalsoslightlyinvolvedin bone corrosion.
Literature
Cited
Clark, R. J. 1972. Pellets of the Short-earedOwl and MarshHawk compared.J. Wildl.
Manage. 36:962-964.
Duke, G. E., A. A. Jegers,G. Loft, and O. A. Evanson.1975. Gastricdigestionin some
raptors.Comp.Biochem.Physiol.50A:649-656.
Errington,P. L. 1932. Techniqueof raptor food habitsstudy.Condor34:75-86.
Glading,B. D., D. F. Tillotson,andD. M. Selleck.1943. Raptorpelletsasindicatorsof food
habits. Calif. Fish and Game 29:92-121.
Summer
1976
Cummings et. al. - Corrosion of Bone
TABLE
1
Effect of pH or pepsinon digestionof bone.
Contents/100 ml
Bone
Group
Pepsin
(rag)
A
0
A
"
B
•
B
•
A
2OO
A
g
•
•
g
•
A
4OO
A
•
B
•
B
•
pH
Boneloss(%)*
1.66
2.35
1.66
2.35
70.7 + 10.7 (5)
25.2 + 14.7 (5)
64.8 + 14.8 (5)
24.5 + 8.7 (5)
1.66
2.35
1.66
2.35
81.9 + 10.7 (5)
32.6 + 18.3 (5)
75.6 + 10.1 (5)
34.0 + 11.0 (5)
1.66
2.35
1.66
2.35
81.8 + 10.5 (6)
37.0 + 19.8 (6)
80.8 + 11.4 (6)
32.9 + 14.1 (6)
* Mean+ S. D. with numberof experimentsin parentheses.
TABLE
2
Effectsof incubationand/or digestionon
Proteolyticactivity of simulatedraptor gastricjuice
solutions
containing
4 mg/mlof pepsin.
Treatment
mg/ml of tyrosinereleased*
pH 1.66
pH 2.35
not incubated; no
4.13 (2)**
4.43 (2)
3.21 (2)
2.67 (2)
2.73 (2)
1.96 (2)
bones added
4 hr. of incubation;
no bones added
4 hr. of incubation;
bones added
* see text.
** meanwith numberof samples
testedin parentheses.
57