Simultaneous determination of 11 phytoestrogens in human serum using a 2 min liquid chromatography/tandem mass spectrometry method
ABSTRACT A rapid 2 min liquid chromatography-tandem mass spectrometry (LC-MS/MS) method operating in multiple reaction ion monitoring mode was developed and validated that allows for the characterization and simultaneous quantification of 11 phytoestrogen metabolites with mass transitions m/z 241/119 (equol), 253/132 (daidzein), 255/149 (dihydrodaidzein), 257/108 (O-desmethylangolesin), 269/133 (genistein), 283/184 (glycitein), 267/191 (formononetin), 289/109 (biochanin A), 267/91 (coumestrol), enterodiol (301/253), and enterolactone (297/253). The method was demonstrated to be specific and sensitive, and a linear response for each phytoestrogen was observed over a range of 1-5000 ng/mL in human serum with the exception of dihydrodaidzein, whose lower limit of quantification was 2 ng/mL. The separation was carried out on a Synergi Polar-RP 2.5 micron (50 mm x 2.0 mm i.d.) column at 50 degrees C with water and acetonitrile (both containing 10 mM ammonium acetate) as the mobile phase under gradient conditions at a flow rate of 0.75 mL/min. This LC-MS/MS method is very useful for high-throughput analysis of phytoestrogens and proved to be simple, sensitive, reproducible, and reliable.
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ABSTRACT: Soy foods are the richest sources of isoflavones, mainly daidzein and genistein. Soy isoflavones are structurally similar to the steroid hormone 17β-estradiol and may protect against breast cancer. S-(-)equol, a metabolite of the soy isoflavone daidzein, has a higher bioavailability and greater affinity for estrogen receptor β than daidzein. Approximately one-third of the Western population is able to produce S-(-)equol, and the ability is linked to certain gut microbes. We hypothesized that the prevalence of breast cancer, ductal hyperplasia, and overall breast pathology will be lower among S-(-)equol producing, as compared with nonproducing, postmenopausal women undergoing a breast biopsy. We tested our hypothesis using a cross-sectional study design. Usual diets of the participants were supplemented with 1 soy bar per day for 3 consecutive days. Liquid chromatography-multiple reaction ion monitoring mass spectrometry analysis of urine from 143 subjects revealed 25 (17.5%) as S-(-)equol producers. We found no statistically significant associations between S-(-)equol producing status and overall breast pathology (odds ratio [OR], 0.68; 95% confidence interval [CI], 0.23-1.89), ductal hyperplasia (OR, 0.84; 95% CI, 0.20-3.41), or breast cancer (OR, 0.56; 95% CI, 0.16-1.87). However, the mean dietary isoflavone intake was much lower (0.3 mg/d) than in previous reports. Given that the amount of S-(-)equol produced in the gut depends on the amount of daidzein exposure, the low soy intake coupled with lower prevalence of S-(-)equol producing status in the study population favors toward null associations. Findings from our study could be used for further investigations on S-(-)equol producing status and disease risk.Nutrition research 02/2014; 34(2):116-25. DOI:10.1016/j.nutres.2013.12.002 · 2.59 Impact Factor
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ABSTRACT: This study was conducted to assess the value of a high resolution, high mass accuracy time-of-flight analyzer in combination with nanoliquid chromatography for the analysis of polyphenols and their metabolites. The goal was to create a method that utilizes small volumes of biological fluids and provides a significant improvement in sensitivity compared with existing methods. Accordingly, nanoLC-MS and nanoLC-pseudo-multiple reaction monitoring (MRM) methods were developed that had a lower limit of quantification of 0.5 nM for several polyphenols and were linear over 2-3 orders of magnitude (R2>0.999). Using urine samples, the ability to observe and quantify polyphenols in such a complex biological fluid depended on much narrower mass windows (0.050 amu or less) on a TOF analyzer than those used on a quadrupole analyzer (0.7 amu). Although a greater selectivity was possible with the low mass resolution of a triple quadrupole instrument using the MRM approach, for the daidzein metabolite O-DMA, a chromatographically resolvable second peak could only be substantially reduced by using a 0.01 amu mass window. The advantage of a TOF analyzer for product ion data is that the whole MSMS spectrum is collected at high mass accuracy and MRM experiments are conducted in silico after the analysis.Archives of Biochemistry and Biophysics 10/2014; DOI:10.1016/j.abb.2014.06.014 · 3.04 Impact Factor
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ABSTRACT: Human clinical trials targeted at preventing gains in body weight using soy protein and isoflavones are limited to adults and yield conflicting results. We hypothesized that daily intake of soy protein/isoflavones would attenuate gains in body weight to a greater extent than a casein-based control in 18 to 19 year-old females. To test this hypothesis, we conducted a randomized, double blind, placebo-controlled trial over 16 weeks to examine the effects of a soy protein/isoflavone-based meal replacement (experimental group) versus a casein-based meal replacement (control group) on body weight and body composition variables in female college freshmen (N = 120). Fat mass (FM), fat-free soft tissue mass (FFST), and percent body fat (%BF) were measured using dual energy X-ray absorptiometry (DXA; Delphi A). Repeated measures mixed models were used to determine the effects of treatment on anthropometric and body composition variables (body weight, waist circumference, FM, FFST, and %BF). No significant group×time interactions were observed, even when body mass index was controlled for in the analysis. Over 16 weeks, body weight, FM, FFST, and %BF significantly increased in both groups (P < .05). Our findings show that female college freshmen gained a significant amount of weight over the course of the 16-week study. Gains in body weight and FM were similar among participants assigned to the soy protein/isoflavone- and the casein-based meal replacements. Future research is warranted to determine the effects of soy protein/isoflavone- and casein-based meal replacements versus a non-intervention (i.e., non-protein based) control.Nutrition research 01/2014; 34(1):66-73. DOI:10.1016/j.nutres.2013.09.005 · 2.59 Impact Factor