SummaryGlucose-limited chemostat cultures of Candida utilis were cultivated at various pH levels (3.0-7.5), temperatures (15-37.5"C), dilution rates (0.06-0.42 hr-l), and with different nitrogen sources (NH4+ and NO,-). The ratio of total nucleic acid to protein increased with increase in dilution rate at constant temperature and decreased with increase in temperature at constant dilution rate. The pattern of these variations is consistent with the hypothesis that the nucleic acid to protein ratio is a function of the ratio of the actual dilution rate to the critical dilution rate corresponding to each one of the cultivation temperatures. This ratio is called "reduced dilution rate." A basis is proposed on which various microorganisms may be compared with respect to the ratios of cell protein to nucleic acid, RNA, ribosomal RNA, and polysomes.The rate of balanced growth of microorganisms in a batch culture can be varied either by adjusting the medium composition or by changing the incubation temperature. With the first approach, the amount of RNA per unit biomass increases with growth rate whereas protein remains relatively constant. 1-6 Based on such observations, Schaechter et al.7 and Ecker and Schaechter' have calculated that in growing organisms the rate of protein synthesis per ribosome particle is constant and independent of growth rate. With more refined methods, Rosset et al.4 and Sykes and Young* have shown that the efficiency of ribosomes in protein synthesis increases with an increase in growth rate.The effect of cultivation temperature on RNA and protein contents of microorganisms has been studied by several authors. Schaechter et al.? demonstrated that although the growth rate of Salmonella 239

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Given the considerable time and expense invested in a single bioprocess (fermentation) batch, variability and losses must be identified quickly. We propose that "sniffing" the odor of cultivation media and broth using instruments could provide a rapid and early indication of bioprocess performance. The human sensation of odor is related to the molecular composition of the vapor phase. The traditional approach to characterize volatile compounds has been sample extraction followed by GC-MS analysis. This approach is very tedious and requires some knowledge of the molecules involved. A new, alternate approach based on an "electronic nose" is now available which, like the human nose, can directly characterize the odor without reference to chemical composition. Here, an array of "conductive polymer" sensors with different chemical sensitivities produces a set of different responses to the same odor. The responses are analyzed mathematically, using pattern recognition techniques, to differentiate between different odors with a high level of sensitivity. In this report, we demonstrate the feasibility of using a commercially available electronic nose for the following applications: monitoring lot-to-lot variation in bioprocess medium ingredients, detecting microbial contamination early, and evaluating bioprocess performance during cultivation of microorganisms at inoculum and production stages.

Human IL-4 (hIL-4) has been cloned from a human T cell line based on its homology to the murine IL-4 cDNA sequence. We have compared cytoplasmic and extra-cytoplasmic expression of this basic protein in Escherichia coli using various combinations of promoters, replicons and host strains. Strains producing a cytoplasmic product were most successful at heterologous protein expression, producing up to 500 mg/l of an inactive aggregated form of the protein. The biological activity of the protein could be restored by refolding the protein with guanidine hydrochloride and glutathione giving a specific activity identical to that of IL-4 derived from CHO cell lines stably transformed with an hIL-4 expression plasmid. Strains designed to secrete human IL-4 into the periplasmic space produced far less protein (approximately 5 mg/l). However, a significant fraction of this protein was detected in the culture medium. This fraction appeared to be soluble after ultracentrifugation, and demonstrated high specific activity without refolding. Leakage of heterologous protein into the culture medium may be a viable way to recover biologically active products without relying on the denaturation and refolding in vitro that can, at times, yield incorrectly folded gene product.

Glucose-limited chemostat cultures of Candida utilis were cultivated at various pH levels (3.0-7.5), temperatures (15-37.5 degrees C), dilution rates (0.006-0.42 hr-1), and with one of two nitrogen sources (NH+4 or NO-3). Enterobacter aerogenes was also cultivated in the chemostat under nitrogen and phosphorus limitations. The amino acid profile of total cell protein is expressed as the content of each amino acid relative to the sum of all amino acids recovered ater acid hydrolysis. Cell residues obtained after hot trichloracetic acid extraction display small variations in amino acid profile. Some of these variations correlate with the growth rate at satisfactory levels of statistical significance. In C. utilis, the correlations cover increased levels of lysine, arginine, and leucine and decreased levels of serine and glutamic acid with increased "reduced dilution rate" (D/Dc). In E. aerogenes, increased levels of lysine and arginine and a decreased level of glutamic acid correlate with increased dilution rate. The directions of most of these correlations and the extents of those pertaining to lysine and arginine are consistent with the change predicted to occur simultaneously in the relative level of the ribosomal protein group.