Liquid outflow rate and rumen volume were measured
at each LS period. One hundred grams of polyethylene
glycol 4000 were used as marker and added to the rumen, via
the fistula, one hour before feeding and 50 mL liquid rumen
samples were removed immediately before the PEG addition
and at one hour and 24 hours after feeding. The PEG
concentration in the samples was measured by the
turbidimetric method according to the procedure by
Hyden (1955). The liquid outflow rate was calculated by
regression from the natural logarithm of PEG concentration
samples and rumen volume was estimated by the total
amount added to the rumen and antilogarithm of intersection
regression.