The Metabolism of Starch, Maltose, Glucose and Some Other Sugars by the Rumen Ciliate Entodinium Caudatum

Abstract

SUMMARYWashed suspensions of Entodinium caudatum grown in vitro and incubated anaerobically incorporated 14C from [14C]glucose into the cell pool as [ 1 4 C ] g l~~~~e , maltose and hexose phosphate and into protozoal polysaccharide and intracellular bacteria as a glucose polymer. The uptake of sugars into the pool was by an active process (predominant at low sugar concentrations) probably into the protozoal endoplasm, and by a passive process (predominant at high concentrations) into another part of the cell. Uptake of glucose by the active process was not sufficiently rapid to increase the level of glucose in the pool by more than 17% per hour; this was probably insufficient to increase appreciably the rate of glucose utilization. These protozoa hydrolysed starch to maltose and glucose and the level of these sugars in the pool of protozoa metabolizing starch grains was controlled in part by the inhibitory effect of these sugars on the enzymes that formed them. Studies on the enzymes involved in the metabolism of starch showed that the glucose in starch was hydrolysed to free glucose before phosphorylation. In experiments on the effect of carbohydrates on the survival of protozoa, starch was the most effective in prolonging protozoal life, followed by maltose and glucose in that order. The greater effectiveness of maltose was correlated with the finding that the rate of uptake of maltose carbon was always greater than that for glucose when the two sugars were present in equimolar concentrations. However, the rate for both sugars was markedly decreased in the presence of penicillin and neomycin. Evidence is presented that the protozoal ectoplasm may be freely permeable to sugars in the medium and that there is a barrier between the ectoplasm and endoplasm. 14C from r4C]starch grains was incorporated by the protozoa but there was no synthesis of protozoal protein from carbohydrate.

The Metabolism of Cellulose, Glucose and Starch by the Rumen Ciliate Protozoon Eudiplodinium maggii

“…Protozoa were fractionated as described by Coleman (1969). The number of protozoa was estimated as described by Coleman et a/.…”

“…The work was carried out with four separate isolates (c02, 4~, c and w) all of which behaved similarly, although they varied in size from 102 x 62 pm (isolate c) to 150 x 75 ,am (isolate c02). Washed suspensions were prepared as described by Coleman (1969) except that mineral salt solution D [containing (g 1-l): K2HP04, 6.3; KH2P04, 5.0; NaCI, 0.65; CaCI, (dried), 0.045; MgS04. 7&0,0.09] was used throughout.…”

“…After incubation, intact protozoa were gently centrifuged (see above) and washed ( x 3) in salt solution D at room temperature. The washed protozoa were then either plated out for the determination of radioactivity (Coleman, 1969) or suspended in 2 ml H 2 0 and disrupted (see below). Any bacteria present and most of the cellulose or starch grains that had not been engulfed were sedimented from the fist supernatant fluid obtained after removal of the protozoa and washed once (2000 g, 20 min, 4 "C, on an angle-head centrifuge).…”

“…4) although the addition of CaCl, or MgCl, (each at 1 or 10 mM) to the latter buffer was not stimulatory. Glucose was the major (over 90 %) digestion product unless cellobiose was added to the digestion mixture when, after 4 h, 60 % of the 14C was present as cellobiose (for method of identification see Coleman, 1969).…”

Factors Affecting the Uptake and Metabolism of Soluble Carbohydrates by the Rumen Ciliate Dasytricha ruminantium Isolated from Ovine Rumen Contents by Filtration

“…To determine the distribution of 14C within the protozoa, a sample of the washed cell suspension removed from the incubation flask after 60 min was fractionated with trichloroacetic acid (TCA) as described by Coleman (1969). A similar sample was refluxed with CHCl,/CH,OH (2 : I, v/v) to extract the lipids.…”