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The fatty-acid composition of C. roseus and N. tabacum cell suspension cultures was unaffected by subculture on Wood and Braun, Murashige and Skoog, or Gamborg B5C media. However, placing the cultures - which were normally grown at 25° C - at 15° C reduced growth but resulted in enhanced formation of oleic and linolenic acids in C. roseus cultures and increased levels of linoleic and linolenic acids in cultures of G. max and N. tabacum, respectively. The incorporation of [(14)C]acetate into [(14)C]linoleic acid was more rapid in N. tabacum cells than in G. max cells, but was very poor in C. roseus where the [(14)C] label was distributed mainly between palmitic and oleic acids.

A cell-free extract containing the enzymes for de-novo synthesis, elongation and desaturation of fatty acids was prepared from cultured cells of Catharanthus roseus G. Don. (14)C-Fatty acids synthesized by the extract from [2-(14)C]malonyl CoA substrate were palmitic (16:0), stearic (18:0) and oleic (18:1). Dialyzed extract was active and stable at room temperature and at 4° C, but was inactivated on boiling. There was an absolute requirement for NADPH for incorporation of [2-(14)C]malonyl CoA into total fatty acids. Escherichia coli acyl carrier protein stimulated total fatty-acid synthesis without affecting the relative ratio of individual fatty acids. Total fatty-acid synthesis at a rate of 45 nmol·mg(-1) protein·h(-1) occurred at a substrate level of 73 μM malonyl CoA, cofactor levels of 500 μM NADPH, 30 μg·ml(-1) E. coli ACP, and 1.0 mg·ml(-1) extract protein. Total fatty acid synthesis was also sensitive to cerulenin and CoA levels. Variations in the relative abundance of individual (14)C-fatty acids were regulated by concentrations of [(14)C]malonyl CoA. NADPH and ferredoxin, as well as by pH, temperature and length of incubation. Fatty-acid synthetase enzymes responsible for [(14)C]palmitic acid were rapidly saturated at a low substrate level (0.3 μM malonyl CoA). Increasing the level of [2-(14)C]malonyl CoA permitted further synthesis of [(14)C]stearate and [(14)C]oleate. Desaturation of [(14)C]stearate to [(14)C]oleate was stimulated by increasing the levels of NADPH and ferredoxin. The desaturase and elongase enzymes were sensitive to acidic pH. The desaturase was also unstable at 41° C, although fatty acid synthetase and elongase were unaffected by this temperature.

Cell suspension cultures of Catharanthus roseus G. Don, Glycine max (L.) Merr. and Nicotiana tabacum L. were incubated with [(14)C]acetate, [(14)C]oleic acid and [(14)C]linoleic acid at five different temperatures ranging from 15 to 35° C. When the incubation temperature was increased, [(14)C]acetate was incorporated preferentially into [(14)C]palmitate, with a concomitant drop in [(14)C]oleate formation. Between 15 and 20° C, [(14)C]oleic acid accumulated in C. roseus cells. In all cultures, optimum desaturation of [(14)C]oleic acid to [(14)C]linoleic acid occurred between 20 and 25° C, and in G. max this was also the optimal range for desaturation of [(14)C]linoleic acid to [(14)C]linolenic acid. Elongation of [(14)C]palmitic acid was inhibited when cultures grown at 15° C for 25 h were subsequently incubated with [(14)C]acetate at 25° C. [(14)C]oleic acid accumulated in G. max and C. roseus cultures grown at 35° C for 25 h and subsequently incubated at 25° C. Desaturation of [(14)C]oleic acid increased up to 25° C, but then decreased or leveled off depending on the cell line and on the temperature prior to incubation.