Trace level quantification of deuterated 17β‐estradiol and estrone in ovariectomized mouse plasma and brain using liquid chromatography/tandem mass spectrometry following dansylation reaction

Abstract

A sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method coupled with dansylation was developed for the simultaneous quantification of exogenously administered deuterated 17beta-estradiol-d4 (E2-d4) and its metabolite, estrone-d4 (E1-d4), in mouse plasma and brain homogenates. The dansylation reaction was simple, fast, and sensitive, and a lower limit of quantification of 50 pg/mL was achieved by using 50 microL of mouse plasma. Interference from endogenous 17beta-estradiol and estrone in plasma and brain samples was minimized by the use of deuterated-E2 as well by utilizing ovariectomized (OVX) mice. The recovery of dansylated derivative exceeded 83% and the reaction was completed within approximately 3 min. The intra- and inter-day assay precision were better than 12.9% and assay accuracy ranged between 92-104% for E1-d4 and E2-d4 in plasma, respectively. The absorption of E2-d4 at both 1 and 3 mg/kg P.O. was rapid, reaching peak plasma concentrations (Cmax) at 5 min post-dose that was the earliest time point obtained, and were 1.1 and 13.8 ng/mL, respectively; the Cmax values for the estrone metabolite, E1-d4, were 1.1 and 43.2 ng/mL, respectively. The area-under-the-plasma-time curve (AUC(0-2 h)) values were determined to be 0.65 and 2.90 ng. h/mL for E2-d4 and 0.77 and 6.74 ng. h/mL for E1-d4, respectively, at 1 and 3 mg/kg. The mean brain-to-plasma ratio for E1-d4 and E2-d4 after P.O. administration of E2-d4 to the OVX mice at 1 and 3 mg/kg indicated that both E1-d4 and E2-d4 were present in the brain as well as in the circulation.

Oleoyl-estrone is a precursor of an estrone-derived ponderostat signal

“…However, the concentrations needed to show measurable peaks were much higher than those found in serum for most estrogens. Consequently, a modified dansyl-chloride (Sigma) derivatisation method was used to measure the serum levels of estrone and its oxidised derivatives [44].…”

Recent progress in the development of derivatization reagents having a benzofurazan structure

“…One of the most often used reagents for LC/ESI-MS/MS is dansyl chloride [5-(dimethylamino)-1-naphthalenesulfonyl chloride; DNS-Cl]. DNS-Cl is often used for the derivatization of aromatic hydroxyl group of steroids such as estradiols (Anari et al, 2002;Xu et al, 2005;Xia et al, 2004;Zhang et al, 2004). The generated DNS-estradiols efficiently produce a distinct fragment ion at m/z 171 by CID and thus these compounds have been sensitively analyzed by LC/ESI-MS/MS.…”

Synthesis of benzofurazan derivatization reagents for carboxylic acids in liquid chromatography/electrospray ionization–tandem mass spectrometry

“…One of the most often used reagents for LC/ESI-MS/MS is dansyl chloride [5-(dimethylamino)-1-naphthalenesulfonyl chloride; DNS-Cl]. DNS-Cl was used for the derivatization of aromatic hydroxyl groups of steroids such as estradiols (Anari et al, 2002;Xu et al, 2005;Xia et al, 2004;Zhang et al, 2004). The generated DNS-estradiols efficiently produced a distinct fragment ion at m/z 171 by CID and thus these compounds were sensitively analyzed by LC/ESI-MS/MS.…”