ABSTRACT
During the last decade phenomenal advances have taken place in large-scale mammalian cell culture both for microcarriers and suspension methods. The cost of serum and product quality require that such systems be examined in terms of both their physical and chemical parameters. Data are presented as to the method used to more than double the final harvest cell density of 100-liter batch culture reactors by simple temperature measurements and 12-liter reactors by increasing the oxygen potential of the liquid. The results are interpreted in terms of physical transport phenomena of momentum, heat and mass via the theoretical relationships of Navier -Stokes, Fourier and Fick respectively. In addition, data are presented of 3-, 12-, and 100 liter reactors that the spent cell growth media were analyzed for ten different chemistries. The Monod cell growth curve was used to interpret the results. The cost savings and product quality improvements can be enormous by approaching both the technology and physiology of mammalian cell culture from the vantage points of chemistry, thermodynamics and transport phenomena.
Subject(s)
Carcinoma 256, Walker/physiopathology , Culture Techniques/methods , Animals , Cell Division , Culture Media , Kinetics , Models, Biological , Rats , Temperature , Time FactorsABSTRACT
Glycine-derivatized polystyrene beads were prepared and used as microcarriers to grow normal cells of human embryonic kidney, rhesus monkey kidney, an human foreskin fibroblasts in suspension cultures. Growth of the cells on polystyrene beads derivatized with other amino acids, peptides, and carboxylic acids also was investigated.
Subject(s)
Cell Adhesion , Cells, Cultured , Animals , Cell Division , Fibroblasts , Glycine , Humans , Kidney , Macaca mulatta , Methods , Microspheres , PolystyrenesABSTRACT
Thei nhibition of the thermolysin catalyzed hydrolysis of FA-Gly-Leu-NH2 and FA-Gly-Phe-NH2 has been reported. The results suggest a model for substrate and inhibitor binding involving the hydrophobic specificity pocket, Arg-203 and Glu-143.
Subject(s)
Leucine/pharmacology , Oligopeptides/pharmacology , Phenylalanine/pharmacology , Thermolysin/antagonists & inhibitors , Binding Sites , Hydrogen-Ion Concentration , Kinetics , Oligopeptides/metabolism , Thermolysin/metabolismSubject(s)
Dipeptides/pharmacology , Thermolysin/antagonists & inhibitors , Acrylates/metabolism , Amines/metabolism , Amino Acid Sequence , Amino Acids/pharmacology , Arginine/analysis , Bacillus/enzymology , Binding Sites , Binding, Competitive , Dipeptides/metabolism , Drug Stability , Furans/metabolism , Hydrogen-Ion Concentration , Hydrolysis , Protein Conformation , Structure-Activity Relationship , Temperature , Thermodynamics , Thermolysin/analysisABSTRACT
Cell-free extracts of Bacillus stearothermophilus have been shown to exhibit proteolytic activity toward casein as well as specific activity to catalyze the hydrolysis of furylacryloylglycyl-l-leucine amide, furylacryloylglycine, and carbobenzoxyl-glycine-p-nitrophenyl ester, indicating the presence of a neutral proteinase, a carboxypeptidase-like enzyme, and an alkaline proteinase. The neutral proteinase and carboxypeptidase-like activities were separated by gel filtration over Bio-Gel P-60, and both were reversibly inhibited by 1, 10-phenanthroline. The esterase activity was inhibited by diisopropylfluorophosphate, which did not affect other enzymatic activities and was insensitive to 1, 10-phenanthroline and ethylenediaminetetra-acetic acid.
