- Sex hormones, including androgens, estrogens, and progestogens, are important biomarkers for various diseases. Quantification of sex hormones is typically conducted by LC-MS/MS. At present, most methods require liquid-liquid extraction or solid phase extraction for sample preparation. However, these pretreatments are prone to compromise LC-MS/MS throughput. To improve on the current standard practices, we investigated cold-induced phase separation for sex hormone extraction. After protein precipitation with acetonitrile and adjusting the solution constitution with water, samples were stored at −30°C for 10 min to generate two distinct phases: an acetonitrile-rich layer on top of a water-rich layer.
- FFAs display pleiotropic functions in human diseases. Short-chain FAs (SCFAs), medium-chain FAs, and long-chain FAs are derived from different origins, and precise quantification of these FFAs is critical for revealing their roles in biological processes. However, accessing stable isotope-labeled internal standards is difficult, and different chain lengths of FFAs challenge the chromatographic coverage. Here, we developed a metabolomics strategy to analyze FFAs based on isotope-free LC-MS-multiple reaction monitoring integrated with dual derivatization.
- Analyzing global steroid metabolism in humans can shed light on the etiologies of steroid-related diseases. However, existing methods require large amounts of serum and lack the evaluation of accuracy. Here, we developed an LC/MS/MS method for the simultaneous quantification of 12 steroid hormones: testosterone, pregnenolone, progesterone, androstenedione, corticosterone, 11-deoxycortisol, cortisol, 17-hydroxypregnenolone, 17-hydroxyprogesterone, dehydroepiandrosterone, estriol, and estradiol. Steroids and spiked internal standards in 100 μl serum were extracted by protein precipitation and liquid-liquid extraction.
- Quantification of malondialdehyde (MDA) as a marker of lipid peroxidation is relevant for many research fields. We describe a new sensitive and selective method to measure free and total plasmatic MDA using derivatization with 2,4-dinitrophenylhydrazine (DNPH) and ultra-HPLC-high-resolution MS. Free and total MDA were extracted from minute sample amounts (10 μl) using acidic precipitation and alkaline hydrolysis followed by acidic precipitation, respectively. Derivatization was completed within 10 min at room temperature, and the excess DNPH discarded by liquid-liquid extraction.