A collaborative study comparing an in situ protocol with single time-point enzyme assays for estimating ruminal protein degradability of different forages

C. P. Mathis, R. C. Cochran, E. S. Vanzant, I. E.O. Abdelgadir, J. S. Heldt, K. C. Olson, D. E. Johnson, J. Caton, D. Faulkner, G. Horn, S. Paisley, R. Mass, K. Moore, J. Halgerson

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Seven institutions conducted a collaborative trial to compare three methods for estimating effective degradability (ED) of forage protein from alfalfa (Medicago sativa; 2.9% N), bermuda (Cynodon dactylon; 1.4% N), brome (Bromus inermis; 0.9% N), forage sorghum (Sorghum bicolor; 0.8% N), and prairie (mixture of native tallgrass prairie species; 0.9% N) hays. To facilitate the collaborative nature of this study, an adaptation of an abbreviated in situ procedure proposed by Broderick [Quantifying forage protein quality. In: Fahey, G.C., Collins, M., Mertens, D.R., Moser, L.E. (Eds.), Forage Quality, Evaluation, and Utilization. ASA, CSSA, and SSSA, Madison, WI, USA, pp. 200-228] and subsequently validated by Vanzant [J. Anim. Sci. 74 (1996) 2773] was adopted for comparison with other procedures. The abbreviated in situ procedure (conducted at four institutions) assumed first-order degradation kinetics and entailed not incubating (0 h) as well as ruminally incubating (2, 8, and 96 h) duplicate polyester bags (10 cm × 20 cm) of each forage (5 g) in the rumens of two steers per location during each of two periods. Rinsing, drying, and microbial correction procedures were standardized. The in situ estimates of ED were compared with two single time-point enzyme assays (using Streptomyces griseus protease) derived from the work of Roe [Techniques for measuring protein fractions in feedstuffs. In: Proceedings of the Cornell Nutrition Conference, Department of Animal Science, Cornell University, Ithaca, NY, pp. 81-88] and Krishnamoorthy et al. [Br. J. Nutr. 50 (1983) 555] with adaptations based on the work of Abdelgadir et al. [J. Anim. Sci 75 (1997) 2215] and Coblentz et al. [J. Diary Sci. 82 (1999) 343]. For the protease assays, duplicate forage samples (15 mg N) were incubated (at seven institutions) in 40 ml of a buffer solution for 1 h. Subsequently, 10 ml of a S. griseus protease solution (0.33 U of activity/ml) were added to the buffer plus forage and incubated for 48 h. In addition, a shorter procedure was evaluated (at six institutions) wherein an increased concentration (33.0 U of activity/ml) of S. griseus concentration was used to compensate for shorter incubation time (4 h). Average ED values obtained from both the 48 and 4 h S. griseus assays explained a large portion of the variation observed for the average ED values from the in situ technique (r2 = 0.87 and r2 = 0.88, respectively). However, there did appear to be some evidence that the S. griseus assays tested overestimated in situ ED in forages with low degradability. The 4 h/high-concentration and 48 h/low-concentration S. griseus assays were closely related (r2 = 0.99) with little bias evident in the relationship. These assays were also less variable than the abbreviated in situ approach. In conclusion, single time-point S. griseus assays with appropriately balanced incubation lengths and enzyme concentrations appear promising for estimating ED under routine laboratory conditions.

Original languageEnglish (US)
Pages (from-to)31-42
Number of pages12
JournalAnimal Feed Science and Technology
Issue number1-2
StatePublished - 2001
Externally publishedYes


  • Degradability
  • Forage
  • In situ
  • Protease
  • Protein

ASJC Scopus subject areas

  • Animal Science and Zoology


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