Quantitation of Urinary 6β-Hydroxycortisol and Free Cortisol by Ultra- Performance Liquid Chromatography-Quadrupole Time-of-Flight Mass Spectrometry
Bibliographic record
Abstract
A new UPLC-MS method that combined 0.01% formic acid/isopropanol as the mobile phase and an ESI-QTOF instrument has been developed for reliable quantitation of urinary 6β-hydroxycortisol (6βHC) and free cortisol (FC).The use of 0.01% formic acid/isopropanol for binary gradient elution on a reversed-phase C18 column resulted in 3.1 and 5.0 times the peak areas for 6βHC and FC, respectively, as compared to 0.1% formic acid/acetonitrile.Liquidliquid extraction with ethyl acetate at pH 5 was superior to solid-phase extraction for sample preparation.A mass window of 50 ppm was used for quantitative monitoring of 6βHC (m/z 379.212) and FC (m/z 363.217) using full-mass detection, with the on-column limits of detection as 4.0 and 1.4 fmol, and the lower limits of quantitation as 13.6 and 6.9 fmol for 6βHC and FC.The accuracy of quantitation ranged from 93.3% to 102.3% recoveries at three spiking levels, with the maximum intraday and interday %CV being 3.7 and 5.3 for both analytes.This LC-MS method was then applied to the quantitation of 6βHC and FC in the urine pairs collected during the follicular and premenstrual phases of menstrual cycle from each of sixty-one premenopausal women -forty-one with ovulatory cycles and twenty with subclinical anovulatory cycles.Paired two-tailed T-tests showed no significant difference (p>0.05) between the metabolic ratios of urinary 6βHC/FC during the follicular and premenstrual phases in both ovulatory and anovulatory subjects, indicating no correlation of the urinary 6βHC/FC ratios between the two physiological phases of menstrual cycle in these subjects.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.001 | 0.001 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 0.001 |
| Bibliometrics | 0.002 | 0.001 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.001 | 0.001 |
| Open science | 0.001 | 0.001 |
| Research integrity | 0.001 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".