Wolfe, Henry R.2019-03-262019-03-261984http://hdl.handle.net/10919/88595A rapid, chemical method for the detection and quantitation of Iipoic acid has been developed. Lipoic acid produces a yellow color when reacted with PdCl₄⁻² in 1 N HCI. The colored complex formed is extractable into methylene chloride, which can be readily concentrated, increasing the color intensity. The limit of sensitivity of this assay for lipoic acid detection in supernatants from Escherichia coli K-12 cell cultures is 2.5 x 10⁻⁵ M, giving an absorbance of 0.10 at 408 nm. Using this assay, it would be possible to screen for a mutant of Escherichia coil K-12 which excretes 100-fold more lipoic acid than the parent strain. Cellular lipoic acid in an anaerobic Escherichia coli K-12 culture remained constant (5 to 6 µg/g dry weight) during growth in a. minimal salts medium containing 1% glucose. However, cellular lipoic acid in an aerobic Escherichia coli K-12 culture increased from 15 µg/g dry weight to 20 to 25 µg/g dry weight during cell growth in the same medium. Aeration (2 L air/L medium/ minute) of a mid-log phase anaerobic Escherichia coli K-12 culture resulted in a doubling of cellular lipoic acid levels within the first thirty minutes and a four fold increase over the next five hours of aerated cell growth. Chinese hamster ovary cells were found not to incorporate the two known bacterial precursors, [²H₃]-acetate and [U-²H₁₅]-octanoate, into lipoic acid. Further studies with both Chinese hamster ovary and mouse fibroblast cells which were designed to demonstrate that these two transformed eel 1 1 ines require lipoic acid for maximal growth were inconclusive.ix, 66 leavesapplication/pdfen-USIn CopyrightLD5655.V855 1984.W643BiosynthesisEscherichia coliThesis