Culture of eukaryotic cells with macro-reticulate buffers: fermentation of cellulolytic fungi.

Fermentation of fungi for large-scale production of extracellular cellulolytic enzymes requires a strict control of pH. At the lab scale, where bioreactors are not available, a culture in the exponential growth phase requires frequent manual pH adjustments. When fungi are grown in the presence of macroreticulate buffers, the culture is stable and does not require any pH control for as long as two weeks. These insoluble buffers are polyacrylamide beads (e.g., 10%T, 8%C) containing acrylamido weak acids and bases in such ratios as to unequivocally define a single pH value along the pH sale. At such pH, the macroreticulate buffers possess a strong buffering power (up to 100 milliequivalent liter-1 pH-1). In the present example, a Trichoderma sp. strain is grown in the presence of 12% beads (v/v) with an isoelectric point of 5.6, containing 100 mM of a pK 6.2 weak acrylamido base and 89 mM of a pK 4.6 weak acrylamido acid. Enzyme production (exoglucanase, endoglucanase, xylanase, beta-glucosidase) is as good as (and often better than) the control in which the pH is adjusted manually 2-3 times/day.