Farm Animal Metabolism and Nutrition

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more water-soluble gas such as CO 2 and
a less water-soluble gas such as CH 4
(Schofield and Pell, 1995b).


Open automated method

● Pro. Will accomodate larger samples.
The sample size may be limited, how-
ever, by the buffering capacity of the
medium. Measurements are made at an
approximately constant pressure close
to atmospheric and thus pressure effects
on digestion are not an issue.
● Con. If the method is to be sensitive, the
excess pressure needed to trigger valve
opening must be small. The pressure
sensors used by Cone et al. (1996)
measure the pressure difference between
the atmosphere and the digestion vessel
and must record this difference over a
50 h period. Changes in atmospheric
pressure may significantly change the
volume of gas needed to trigger valve
opening. Sudden changes will affect
only a few readings, but a slow, steady
change can affect many. One simple
solution to this problem would be to use
a ‘differential’ rather than a ‘gauge’ type
of sensor and refer all pressure measure-
ments to a closed standard (see
Appendix for a description of these
different types of sensors). This app-
roach insulates the digestion bottle from
any external pressure changes. It could
also be used for the closed system.


Problems with mixtures of CO 2 and
CH 4 are no different in the open system
than in the closed automated method (see
above). A small excess pressure must
accumulate before venting. The volume of
gas needed to generate this pressure
increase will be greater for CO 2 than for
CH 4 because of the difference in water
solubility of these gases. The calibration
factor (volume of gas needed to trigger
valve opening) will thus change if the com-
position of the headspace gas changes. The
error for a single reading is small but must
be evaluated as a fraction of a total reading
that also is small.


Appropriate Use of Blanks

The usual procedure in all gas systems is to
subtract, at each time point, the gas
produced in a ‘blank’ container from that
produced in the sample bottle or syringe.
The blank contains buffered RF in the same
proportions as the sample. Some gas does
appear in the blank because the RF
inoculum (unless centrifuged, washed and
re-suspended) inevitably contains feed
particles. The size of this blank contribu-
tion to the total gas volume depends on the
proportion of RF used. Some groups use a
1:2 ratio of RF to buffer (Menke and
Steingass, 1988; Cone et al., 1996), some a
1:4 ratio (Pell and Schofield, 1993) and
others a 1:9 ratio (Theodorou et al., 1994).
For the 1:2 ratio, the blank contributed
about 30% of the total reading after 10 h
incubation of grass samples (Cone et al.,
1997). These authors have demonstrated
that microbial turnover in the blank begins
after about 1 h and that about 30% of the
maximum blank reading can be attributed
to this turnover. In the presence of sub-
strate, turnover is delayed so that the blank
does not reflect accurately what happens in
the sample. For this reason, Cone et al.
(1997) suggest omitting the blank correc-
tion altogether.
An alternative, less draconian, app-
roach is to use a more diluted inoculum that
will reduce the contribution of the blank. A
second reason to retain the blank correction
appears in the ‘closed’ system where pres-
sure accumulates. The blank plays a dual
role in this system. Besides correcting for
fermentation products from the inoculum,
the blank also corrects for atmospheric pres-
sure changes occurring during the run (Pell
and Schofield, 1993).

Equipment Needed for

Computerized Gas Measurement

Digestion vessels

Figure 10.2 shows an example of serum
bottles as digestion vessels and an incubator
used for in vitrogas measurements. Any

Gas Production Methods 215
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