| Experiments were performed to develop methodology for utilizing 13C-labeled fatty acids to investigate the activity of the Delta 9-desaturase enzyme in vivo in lactating dairy cattle and women. Initially, it was necessary to determine the type of 13 C-labeled fatty acid metabolic tracer, uniformly or singly labeled, that would be most appropriate for quantitative measurements. When rumen microbial cultures were incubated with oleic, oleic-1-13C, and oleic-U- 13C, the biohydrogenation of oleic acid was affected by the increasing mass. Based on these results, use of singly 13C-labeled fatty acids is most appropriate in quantitative research. However, in examinations identifying substrate conversions, uniformly labeled compounds would be advantageous due to ease of analysis. Subsequent experiments evaluated in vivo desaturation of the four major fatty acid substrates of the Delta 9-desaturase enzyme, myristic-1-13C acid, palmitic-1- 13C acid, stearic-1-13C acid, and vaccenic-1- 13C acid, in lactating dairy cattle. Total yield of the corresponding Delta 9-desaturase enzyme substrate:product pairs in the milk fat is necessary for calculating the optimum dose of 13C-labeled fatty acid. In each instance, no 13C-enrichment was detected in the Delta 9-desaturase enzyme products in plasma but was detected in the milk fat. Thus indicating that in the short term infusions the majority of the Delta 9-desaturase enzyme activity is attributed to the mammary gland. Due to the potentially beneficial properties in humans of the Delta9 -desaturase enzyme substrate:product pair vaccenic acid and cis-9, trans-11 conjugated linoleic acid, their concentrations in human milk fat was determined. The existence of these fatty acids in human milk was confirmed and offered the prospect for examining the desaturation of vaccenic acid in lactating women using the techniques developed in lactating dairy cattle. Limited 13C-enrichment was detected in the Delta9-desaturase enzyme products in plasma but was detected in the milk fat. Thus again indicating that the majority of Delta 9-desaturase enzyme activity occurred in the mammary gland. In conclusion, this set of experiments in lactating dairy cattle and women confirm that 13C-labeled fatty acids may be used as a tool to measure the activity of the Delta9-desaturase enzyme in vivo. This methodology will provide a powerful tool in assessing dietary impacts on Delta9-desaturase enzyme activity. |
