03090naa a2200313 a 450000100080000000500110000800800410001902400300006010000210009024501570011126000090026852021400027765000160241765000110243365000110244465000150245565000180247065000130248865000150250165000230251665000140253965300210255365300220257465300320259665300320262865300270266065300210268777300680270815202332023-05-03       1978    bl uuuu     u00u1 u    #d7 a10.1093/jn/108.2.1812DOI1 aBRODERICK, G. A.  aIn vitro procedures for estimating rates of ruminal protein degradation and proportions of protein escaping the rumen undegraded.h[electronic resource]  c1978  aAbstract: An in vitro procedure, in which rates of ruminai protein degradation and proportions of protein escaping ruminai degradation were estimated from release of amino acid plus ammonia in the presence of hydrazine sulfate ( HS ), was developed and applied to casein. An equation was derived to estimate proportion of protein escaping ruminai degrada tion: estimated % escape = (kr/(kr + kd)) X 100, where kr and k~a are fractional rate constants for ruminai protein turnover (washout) and degradation, respectively. HS, when added at 1.0 mM to an incubation medium consisting of strained ruminai liquor (SRL) and McDougall's buffer, was found to effectively inhibit removal of added amino acids and ammonia. Thus, protein degradation may be estimated from accumulation of these end-products. Casein was incubated in the medium in small quantities (2 mg/ml SRL) and its degradation estimated by this procedure. Casein degradation was found to be a first-order process, with plots of log fraction undegraded versus time being linear up to 3 hours, and with mean kd = 0.242/hour. Assuming kr = 0.04/hour, a value of 14.2^0 ( (0.04/(0.04 + 0.242)) X 100) ruminai escape was calculated. Alkali-labile phosphorus (ALP) naturally bound to casein was used in an alternative approach to quantitate casein disappearance. In vitro studies indicated 1.0 HIM HS did not inhibit casein degradation (measured by ALP disappearance) up to 3 hours, and both the amino acid plus ammonia release and ALP pro cedures gave comparable results. Casein kd value determined simultane ously in vivo and in vitro were, respectively, 0.462/hour (by ALP) and 0.300/hour (by amino acid plus ammonia release). Use of cetyltrimethyl- ammonium bromide when stopping incubations increased apparent amino acid plus ammonia release rates, presumably by rupturing microbial cell walls. Alternative kd measurement was made from the ratio of Michaelis- Menten kinetic constants, k'd = Vâ??,ax/Km. Results suggest that this approach may give more accurate kd estimates. Implications of the described theory and techniques are also discussed. J. Nutr. 108: 181-190, 1978.  aAmino acids  aCasein  aCattle  aHydrazines  aHydroxylamine  aKinetics  aPhysiology  aRumen fermentation  aRuminante  aDietary proteins  aNutricao proteica  aRuminal in vitro incubation  aRuminal protein degradation  aRuminal protein escape  aRuminal turnover  tThe Journal of Nutritiongv. 108, n. 2, p. 181-191, Feb., 1978.