Serum Metabolome and Proteome Effects of Phytoestrogenic Dietary Supplements in Postmenopausal Women

A Phase II clinical trial assessing the efficacy of two dietary botanical supplements, black cohosh (Actaea racemosa) and red clover (Trifolium pretense) at reducing vasomotor symptoms in postmenopausal women was completed. The study found no statistically significant difference from placebo in women treated with black cohosh and red clover. In order to further understand the effects that these botanicals have on the postmenopausal system, a study to investigate the metabolomic and proteomic changes was undertaken. Metabolomic profiles were generated by preparing serum samples by liquid-liquid extraction before analysis using a Shimadzu IT-TOF LC-MS/MS system. The XCMS online database was used to identify many significant features in each group as being upregulated and downregulated. Native isoflavone compounds from the red clover were compared to compare to the treatment group and several matched the retention time and HRMS spectra. The proteomic profiles were generated by labelling serum samples from each group using ThermoFisher TMT-labeling reagents, fractionation using an Agilent OFFGEL fractionator, and analysis by a Thermo Orbitrap LC-MS/MS system. These results were searched using Mascot and the relative abundances in each group showed many proteins upregulated and downregulated. These results were then analyzed using MetaboAnalyst and NCBI Biosystems databases to identify the pathways affected by the treatments. The experiments identified several features and proteins of interest for further analysis in order to determine the role played in vasomotor symptoms.