An average of 11 (range, 2 to 47) μmoles of formate per g per hr was produced and used in whole bovine rumen contents incubated in vitro, as calculated from the product of the specific turnover rate constant, k , times the concentration of intercellular formate. The latter varied between 5 and 26 (average, 12) nmoles/g. The concentration of formate in the total rumen contents was as much as 1,000 times greater, presumably owing to formate within the microbial cells. The concentration of formate in rumen contents minus most of the plant solids was varied, and from the rates of methanogenesis the Michaelis constant, K m , for formate conversion to CH 4 was estimated at 30 nmoles/g. Also, the dissolved H 2 was measured in relation to methane production, and a K m of 1 nmole/g was obtained. A pure culture of Methanobacterium ruminantium showed a K m of 1 nmole of H 2 /g, but the K m for formate was much higher than the 30 nmoles for the rumen contents. It is concluded that nonmethanogenic microbes metabolize intercellular formate in the rumen. CO 2 and H 2 are the principal substrates for rumen methanogenesis. Eighteen per cent of the rumen methane is derived from formate, as calculated from the intercellular concentration of hydrogen and formate in the rumen, the Michaelis constants for conversion of these substrates by rumen liquid, and the relative capacities of whole rumen contents to ferment these substrates.

The factors influencing the digestion of pebble-milled cellulose by enzymes were studied by using several strains of Ruminococcus albus including a mutant characterized by a more eccentric location of its colony in the clearing produced by digestion of the cellulose in the thin layer lining the wall of a culture tube. Most of the cellulase is extracellular. As much as 65% of the cellulose could be digested by the cell-free enzymes provided the quantity of cellulose was small. Fresh enzyme was repeatedly administered or the digestion experiment was arranged in a dialysis bag through which digestion products could diffuse. Cellobiose and, to a lesser extent, glucose inhibited digestion. Pebble-milled filter paper, moist crystalline cellulose from cotton, and dry crystalline cellulose (Sigmacel) were digested, but in decreasing rapidity, respectively. Carboxymethylcellulose was digested more rapidly than pebble-milled cellulose but to approximately the same final extent as judged by Cu reduction values. Cell walls from alfalfa were digested. The enzyme preparation was active over the pH range 6.0 to 6.8 and showed most rapid cellulose digestion at 45 C. Part of the cellulolytic activity was irreversibly destroyed by exposure to oxygen. Much of the enzyme was absorbed on cellulose. The absorption and desorption characteristics, as well as the partial inhibition by oxygen, indicate that multiple enzymes are involved. 729

No abstract

It is generally accepted that the climacteric phase of ripening fruits is well suited for the study of cellular senescence (1). Since a surge in respiratory activity is the dominant, readily measurable phenomenon, mitochondria are implicated as the sites of causal events. This view was embodied in the early work of Millerd et al. (19) with the resulting postulate that uncoupling of oxidative phosphorylation permitted an increased respiratory rate. Along similar lines, Hulme (7) and Pearson and Robertson (21) attributed the rise in respiration to the availability of phosphate acceptor (ADP) resulting from increased protein synthesis and accelerated demands for energy. However, with specific reference to mitochondrial protein, data on the yield of particulate protein (27) and on the incorporation of amino acids into mitochondrial protein (28) indicate a decline in synthetic activities as fruit reach the climacteric peak.Richmond and Biale (23), Sacher (37), and especially Young and Biale (44) have questioned the prevalence of "uncoupling" or "ADP control" in ripening fruit cells. Moreover, the work of Lance et al. (10), Hulme et al. (8,9) and the experiments reported below, reveal that mitochondria remain functionally sound and "cotipled" throughout the climacteric phase.