Methodological Effects on Neutral Detergent Fiber Disappearance and Degradation Kinetics in In Situ and Bag Based In Vitro Systems

Abstract

Neutral detergent fiber (NDF) is a major component of dairy cattle diets, and its degradation characteristics influence rumen function, feed intake, and milk production. The nutritive value of NDF depends on the rate and extent of ruminal degradation, which are commonly estimated using in situ and in vitro techniques. However, estimates obtained from these methods can be influenced by methodological factors associated with sample preparation, incubation, and post-incubation handling. The objective of this thesis was to evaluate the effects of methodological factors on neutral detergent fiber disappearance (NDFD) and degradation kinetics in bag based in situ and in vitro systems. Chapter 2 evaluated the effects of grinding method (1-mm cutter mill vs. cyclone mill), bag preparation (acetone-rinsed vs. untreated), and washing procedures (no wash, hand wash, and machine wash) on in situ NDFD and degradation kinetics. In the absence of ruminal incubation, cyclone-milled samples showed greater NDFD than cutter-milled samples (10.0 vs. 2.8%; P < 0.01), with the greatest disappearance observed under machine washing (up to 16.4% NDFD. During early ruminal incubation (<12 h), NDF disappearance did not differ from zero for no-wash and hand-wash treatments but was greater than zero under machine washing, including at 0 h (P < 0.05). Methodological factors also affected degradation kinetics, with differences observed in lag time and uNDF, and interactions between grinding and washing influencing the estimated degradation profile. Chapter 3 evaluated in vitro NDFD between closed flask-based system and open bag-based Daisy incubator system. Forage samples were incubated in buffered rumen fluid for up to 240 h, and NDFD was used to estimate degradation kinetics. NDFD at time 0 was not different from zero in the flask system but was greater than zero in the open system, particularly under machine washing (14% NDFD; P < 0.01). The open system had greater fractional degradation rates (kd; ~0.08 vs. 0.04 h⁻¹) and lower uNDF (42% vs. 58%) compared with the flask system. Overall, the results from this thesis demonstrate that estimates of NDFD and degradation kinetics are influenced not only by forage characteristics but also by methodological factors. Early NDF disappearance observed in bag-based systems is not consistent with the expected biology of fiber degradation and is more likely associated with physical loss of particles. These findings highlight the importance of methodological standardization and careful interpretation of NDF degradation data to ensure that estimates more accurately reflect biological processes.