Development of chiral liquid chromatography-tandem mass spectrometry isotope dilution methods for the determination of unconjugated and total S-equol in human plasma and urine.
Liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods for the determination of unconjugated and total (conjugated plus unconjugated) S-equol in human plasma and urine were developed and validated. The separation of R and S enantiomers was achieved with a Chiracel OJ-H column operated in a normal phase mode using ethanol/hexane mobile phase components. Ionization of S-equol by negative ion electrospray generated the [M-H](-) ion whose response was augmented by post-column addition of ammonium hydroxide. A triple stage quadrupole mass spectrometer was used to measure the ion current generated from the dissociative transitions m/z 241→m/z 121 (S-equol) and m/z 245→m/z 123 (equol-d(4)). The determination of total S-equol included an additional deconjugation step involving incubation of the sample with sulfatase and glucuronidase. Average recovery for both unconjugated and total S-equol was 85% with no observable matrix effects. Linearity was established for unconjugated S-equol from 0.025ng/mL to 10ng/mL (plasma) and 0.20ng/mL to 200ng/mL (urine). The average coefficient of variation and accuracy per occasion was within ±15% of the theoretical concentration of S-equol. The method was used to measure the pharmacokinetics of S-equol in human plasma after an oral administration of a single 20mg dose of S-equol to three normal healthy volunteers.
Charles River Laboratories Preclinical Services, Montreal, Canada.
Jeffry B
JB
Plomley
Richard L
RL
Jackson
Richard J
RJ
Schwen
Jeffrey S
JS
Greiwe
D000293:Adolescent; D000328:Adult; D002482:Cellulose / Q000031:analogs & derivatives / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D004355:Drug Stability; D060754:Equol; D005260:Female; D005966:Glucuronidase / Q000378:metabolism; D006801:Humans; D007201:Indicator Dilution Techniques; D007529:Isoflavones / Q000097:blood* / Q000378:metabolism / Q000493:pharmacokinetics / Q000652:urine*; D057230:Limit of Detection; D008297:Male; D048789:Phytoestrogens / Q000097:blood* / Q000378:metabolism / Q000493:pharmacokinetics / Q000652:urine*; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization; D013237:Stereoisomerism; D013429:Sulfatases / Q000378:metabolism; D053719:Tandem Mass Spectrometry; D013678:Technology, Pharmaceutical; D055815:Young Adult
C007610:4',7-dihydroxy-3,4-dihydroisoflavone; D007529:Isoflavones; D048789:Phytoestrogens; C070231:Chiracel OJ; D060754:Equol; D002482:Cellulose; D013429:Sulfatases; D005966:Glucuronidase