solution (40 ml.) is used; use of so much solu- 
tion usually insures that the final acid concen- 
tration does not exceed that found in the ani- 
mal (Tilley and Terry 1963). 
Inoculum 
The in vitro system utilizes undiluted or only 
slightly diluted rumen liquor, whole liquor di- 
luted with mineral solutions, or various frac- 
tions of rumen liquor such as centrifuged cells 
or washed cell suspensions. Several investiga- 
tors have discussed these various inoculum 
preparations (Johnson 1963; Shelton and Reid 
1960; Van Dyne 1962). Aeration of the rumen 
liquor inhibits micro-organism activity by 40 
percent in 15 minutes and by 100 percent in 30 
minutes (Johnson et al. 1958). 
Questions have been raised concerning the 
validity of using the inoculum from an animal 
fed on a different kind of feed than the feed 
being evaluated. Several workers who have 
studied this subject recommend that the inocu- 
lum source animal either be fed on the same 
forage to be evaluated, or be fed a standard 
forage that is analyzed simultaneously with 
other forages and used as a correction factor.* 
In our laboratory, we have found differences in 
forage digestibilities due to inoculum collection 
delays where inoculum source animals were 
grazing the same range species being analyzed 
but during a different vegetative growth period 
(Pearson 1967a). 
We investigated the effect of analyzing na- 
tive range species with inoculum from animals 
grazing introduced species, and vice versa. In 
vitro digestibilities of six forage species and a 
diet mixture from ponderosa pine range were 
determined with inoculum from animals graz- 
ing native and introduced species. Analysis of 
variance revealed no overall difference in the 
forage digestibilities due to inoculum source 
(table 1), but examination of the individual di- 
gestibility samples revealed differences of up to 
10 percent. Apparently the species-inoculum 
source interaction variance was _ sufficiently 
high to mask any significant individual differ- 
ences. 
Careful examination of the individual inter- 
actions reveals that Festuca arizonica and 
Muhlenbergia montana had higher in vitro di- 
gestibilities when inoculum source animals 
grazed native range, while Agropyron interme- 
dium and the diet mixture, 65 percent of which 
consisted of introduced species, were lower. 
Other species responded differently between 
dates. Since individual values are important, 
these results substantiate use of inoculum from 
animals grazing the forage to be analyzed. 
Other inoculum sources may be used only after 
*Van Dyne 1962 Bezeau 1965; Johnson 1966; Brugge- 
mann et al. 1968. 
86 
information has been obtained concerning di- 
gestibility variances. 
TABLE 1.—Percent digestibility of forages col- 
lected from pine ranges and digested 
with inoculum from animals grazing 
native and introduced species 
Inoculum source 
Introduced Native 
Forage species species 
June 1967: 
Festuca arizonica ________ 56.8 59.7 
Muhlenbergia montana ____ 43.6 53.1 
Sitanion hystrix _________ 64.4 62.9 
Carex geophila .2..2-...- 54.1 57.1 
Agropyron intermedium ___ 67.5 60.0 
A cristatum (22 220%. ok * *60:77 62.7 
Dietemixture tee See. Se 66.4 59.6 
August 1967: 
FO OVIZONIee, 22 Seek JSS eeeee 51.3 52.4 
M. Montana 2. x22. 85a! 57.8 58.2 
Sie RU SUR a hea: 2 ee 65.2 65.7 
Gz Geopnid) = 22-2 22-3 =< = 61.3 55.4 
A. intermedium ________- 62.0 61.6 
A. cristatum __._.-.--__-- 60.0 59.2 
Dietimixture. 2. oP Ate 57.0 52.4 
Rumen fluid containing micro-organisms 
was frozen to preserve inoculum for delayed 
forage digestibility trials. Trials using frozen 
and fresh inoculum indicated that the micro- 
organisms die or at least become inactive since 
digestibilities using frozen inoculum are lower 
(table 2). 
TABLE 2.—Percent digestibility of forces col- 
lected during January 1966 on chap- 
arral range. Forages digested with 
fresh and frozen inoculum, with and 
without carbon dioxide flushing, 
prior to tube closure 
Treatment 
Forage Fresh Frozen Fresh 
inocu- inocu- inocu- 
lum, lum, lum, 
CO: CO: not 
flush flush flushed 
Ceanothus greggii 
(leaves) "223 haa 47.9 41.2 31.9 
Quercus turbinella 
(leaves) ______ ew est 44.6 36.3 37.8 
Cercocarpus breviflorus _. 34.6 30.2 35.7 
CROTCGOW x tee 31.6 25.2 17.4 
Rhus trilobata (stems) 41.7 35.2 21.7 
Poa longiligula ________- 52.6 37.9 47.8 
Sitanion hystrix _______- 48.9 32.6 47.1 
Eragrostis lehmanniana _ 39.5 18.8 21.9 
Bouteloua hirsuta _____- 38.8 19.4 29.4 
Be OVUCUIS te Meee ee 31.9 NED, 27.5 
Diet mixture 
(steerii§)* eee es 32.1 20.8 82.6 
Diet mixture 
(steer 4) ________ ie See eS 26.1 16.1 
Mean? 2.5. 2 Lata 39.3a 28.2b 30.6b 
* Diet mixture refers to forages removed from the 
rumen of cattle in rumen evacuation trials. 
*Means with same letter in common are not signifi- 
cantly different at the 0.1 significance level. 
