Although translational research is referred to clinical chemistry measures, correct weighting factors for linear and quadratic calibration curves with least-squares regression algorithm have not been carefully considered in bioanalytical assays yet. The objective of this study was to identify steroidogenic roles in preeclampsia and verify accuracy of quantitative results by comparing two different linear regression models with weighting factor of 1 and 1/x 2 . A liquid chromatography-mass spectrom-etry (LC-MS)-based adrenal steroid assay was conducted to reveal metabolic signatures of preeclampsia in both serum and pla-centa samples obtained 15 preeclamptic patients and 17 age-matched control pregnant women (33.9 ± 4.2 vs. 32.8 ± 5.6 yr, respectively) at 34~36 gestational weeks. Percent biases in the unweighted model (w i = 1) were inversely proportional to concen- trations (-739.4 ~ 852.9%) while those of weighted regression (w i = 1/x 2 ) were < 18% for all variables. The optimized LC-MS combined with the weighted linear regression resulted in significantly increased maternal serum levels of pregnenolone, 21-deoxycortisol, and tetrahydrocortisone (P < 0.05 for all) in preeclampsia. Serum metabolic ratio of (tetrahydrocortisol + allo-tet-rahydrocortisol) / tetrahydrocortisone indicating 11β-hydroxysteroid dehydrogenase type 2 was decreased (P < 0.005) in patients. In placenta, local concentrations of androstenedione were changed while its metabolic ratio to 17α-hydroxyprogester-one responsible for 17,20-lyase activity was significantly decreased in patients (P = 0.002). The current bioanalytical LC-MS assay with corrected weighting factor of 1/x 2 may provide reliable and accurate quantitative outcomes, suggesting altered ste- roidogenesis in preeclampsia patients at late gestational weeks in the third trimester.