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Electron micrographs of thin sections of nuclear, microsomal, and mitochondrial fractions obtained from a carrageenin-induced granuloma showed considerable contamination of the heavier by the lighter fractions. Striated collagen fibrils could be identified in the nuclei + debris fraction. Only a few striated fibrils occurred in the mitochondrial fraction; very fine filaments (diameter 50 A) could be seen in this fraction, but could not be distinguished with certainty from fibrillar material derived from broken nuclei. 35 per cent of the mitochondrial and 80 per cent of the microsomal collagen was extractable by 0.2 M NaCl and could be purified by the standard methods of solution and reprecipitation. The amino acid composition of these collagen fractions determined by ion exchange chromatography was within the range normally found for collagen and gelatin from other mammalian species, allowing for 10 to 20 per cent of some non-collagenous contaminant of the microsomal collagen. Hydroxyproline and proline were isolated by chromatography on paper from hydrolysates of the nuclear, mitochondrial, and microsomal collagen fractions, after incubation of tissue slices with L-(14)C-proline. The specific activities of the hydroxyproline from these collagens were in the approximate ratio 1:2:6, while that of bound hydroxyproline derived from the supernatant was only 1, indicating primary synthesis of collagen in the microsomes. Attempts to demonstrate incorporation of L-(14)C-proline into collagen or into free hydroxyproline in cell free systems were unsuccessful, nor was it possible to demonstrate non-specific incorporation of L-(14)C-valine into TCA-insoluble material by various combinations of subcellular fractions.

Chick embryos were sacrificed at intervals after simultaneous injection of BAPN and proline-C(14), the collagen separated into neutral salt-extractable and residue fractions, and total hydroxyproline and hydroxyproline specific radioactivity determined in each fraction. Extractable collagen, measured as hydroxyproline, increased markedly and had a higher specific activity in BAPN-treated embryos than in corresponding controls. Hydroxyproline of the residue collagen in the treated animals, however, had a lower specific activity. When proline-C(14) was injected 24 hours prior to BAPN, the specific radioactivity of the soluble collagen of treated embryos was similar to that of controls, in spite of the fact that the specific activity of the residue fraction was higher than that of the soluble fraction at the time of BAPN administration. These results suggest that the increased amount of soluble collagen in lathyrism induced by administration of BAPN does not arise from the collagen insoluble prior to administration of the drug, but rather that BAPN acts by blocking the formation of mature collagen fibers, perhaps by preventing the formation of cross-linkages between alpha-collagen chains.