Dynamic binning peak detection and assessment of various lipidomics liquid chromatography-mass spectrometry pre-processing platforms

Liquid chromatography-mass spectrometry (LC-MS)-based lipidomics generates large datasets that need to be interpreted using high-performance data pre-processing tools such as XCMS, mzMine, and Pro genesis. These pre-processing tools rely heavily on accurate peak detection, which depends on proper setting of the peak detection mass tolerance (PDMT). The PDMT is usually set with a fixed value in either ppm or Da units. However, this fixed value may result in duplicates or missed peak detection and inaccurate peak quantification. To improve the accuracy of peak detection, we developed the dynamic binning method, which considers peak broadening described by the physics of ion separation and sets (m)(2) the PDMT dynamically in function of m/z. In our method, the PDMT is proportional to z for Fourier (m)(15) transform ion cyclotron resonance (FTICR), to z for Orbitrap and to m/z for Quadrupole time-of flight (Q-TOF), and is a constant for Quadrupole mass analyzer. The dynamic binning method was implemented in XCMS [1,2], and the adopted source code is available in GitHub at https://github.com/xiaodfeng/DynamicXCMS. We have compared the performance of the XCMS implemented dynamic binning with different popular lipidomics pre-processing tools to find differential compounds. We generated set samples with 43 lipid internal standards that were differentially spiked to aliquots of one human plasma lipid sample using Orbitrap LC-MS/MS. The performance of various pipelines using matched parameter sets was quantified by a quality score system that reflects the ability of a preprocessing pipeline to detect differential peaks spiked at various concentrations. The quality score indicated that our dynamic binning method improves the quantification performance of XCMS (maximum p-value 9.8$10-3 of two-sample Wilcoxon test) over its original implementation. We also showed that the XCMS with dynamic binning found differential spiked-in lipids better or with similar performance as mzMine and Progenesis do. (C) 2021 The Author(s). Published by Elsevier B.V

An antibody-free LC-MS/MS method for the quantification of intact insulin-like growth factors 1 and 2 in human plasma

Insulin-like growth factors 1 and 2 (IGF-1 and IGF-2) are important biomarkers in research and diagnosis of growth disorders. Quantitative analysis is performed using various ligand-binding assays or enzymatic digestion LC-MS/MS methods, whose widespread adoption is hampered by time-consuming sample preparation procedures. We present a simple and fast antibody-free LC-MS/MS method for the quantification of intact IGF-1 and IGF-2 in human plasma. The method requires 50 mu L of plasma and uses fully N-15-labelled IGF-1 as internal standard. It features trifluoroethanol (TFE)-based IGF/IGF-binding protein complex dissociation and a two-step selective protein precipitation workflow, using 5% acetic acid in 80/20 acetone/acetonitrile (precipitation 1) and ice-cold ethanol (precipitation 2). Detection of intact IGF-1 and IGF-2 is performed by means of a Waters XEVO TQ-S triple quadrupole mass spectrometer in positive electrospray ionisation (ESI+) mode. Lower limits of quantification were 5.9 ng/mL for IGF-1 and 8.4 ng/mL for IGF-2. Intra-assay imprecision was below 4.5% and inter-assay imprecision was below 5.8% for both analytes. An excellent correlation was found between nominal and measured concentrations of the WHO reference standard for IGF-1. Comparison with the IDS-iSYS IGF-1 immunoassay showed good correlation (R-2 > 0.97), although a significant bias was observed with the immunoassay giving substantially higher concentrations. The LC-MS/MS method described here allows for reliable and simultaneous quantification of IGF-1 and IGF-2 in plasma, without the need for enzymatic digestion. The method can be readily implemented in clinical mass spectrometry laboratories and has the potential to be adapted for the analysis of different similarly sized peptide hormones

Correction to:An antibody-free LC-MS/MS method for the quantification of intact insulin-like growth factors 1 and 2 in human plasma

Insulin-like growth factors 1 and 2 (IGF-1 and IGF-2) are important biomarkers in research and diagnosis of growth disorders. Quantitative analysis is performed using various ligand-binding assays or enzymatic digestion LC-MS/MS methods, whose widespread adoption is hampered by time-consuming sample preparation procedures. We present a simple and fast antibody-free LC-MS/MS method for the quantification of intact IGF-1 and IGF-2 in human plasma. The method requires 50 mu L of plasma and uses fully N-15-labelled IGF-1 as internal standard. It features trifluoroethanol (TFE)-based IGF/IGF-binding protein complex dissociation and a two-step selective protein precipitation workflow, using 5% acetic acid in 80/20 acetone/acetonitrile (precipitation 1) and ice-cold ethanol (precipitation 2). Detection of intact IGF-1 and IGF-2 is performed by means of a Waters XEVO TQ-S triple quadrupole mass spectrometer in positive electrospray ionisation (ESI+) mode. Lower limits of quantification were 5.9 ng/mL for IGF-1 and 8.4 ng/mL for IGF-2. Intra-assay imprecision was below 4.5% and inter-assay imprecision was below 5.8% for both analytes. An excellent correlation was found between nominal and measured concentrations of the WHO reference standard for IGF-1. Comparison with the IDS-iSYS IGF-1 immunoassay showed good correlation (R-2 > 0.97), although a significant bias was observed with the immunoassay giving substantially higher concentrations. The LC-MS/MS method described here allows for reliable and simultaneous quantification of IGF-1 and IGF-2 in plasma, without the need for enzymatic digestion. The method can be readily implemented in clinical mass spectrometry laboratories and has the potential to be adapted for the analysis of different similarly sized peptide hormones

Plasticity in metabolism of maternal androgens in avian embryos

Mothers can influence offspring phenotypes by transferring non-genetic information to the young, which provides them with a flexible tool to adjust the developmental trajectory of the young in fluctuating environments. Mothers can differentially deposit their resources in the same reproductive attempt in relation to the offspring position in the sibling hierarchy. However, whether embryos from different positions can be plastic in their response to the maternal signals, potentially leading to a mother–offspring conflict, is yet unclear. We used Rock pigeons (Columba livia), that lay two egg clutches where maternal androgen levels in second laid eggs at oviposition are higher than in first laid eggs, and investigated the plasticity of embryonic metabolism of maternal androgens. We experimentally elevated androstenedione and testosterone levels in first eggs to that present in second eggs and measured the change in androgen levels and its main metabolites (etiocholanolone and conjugated testosterone) after 3.5 days of incubation. We found that eggs with increased androgens show a different degree of androgen metabolism depending either on the egg laying sequence or initial androgen levels or both. Our findings indicate that embryos have certain plasticity in response to maternal androgen levels depending on maternal signals.</p

Residual endogenous corticosteroid production in patients with adrenal insufficiency

Objective This study aimed at comparing precursors of endogenous corticosteroid production in patients with primary adrenal insufficiency and in secondary adrenal insufficiency. Design Twenty patients with primary adrenal insufficiency and matched controls and 19 patients with secondary adrenal insufficiency participated in this ancillary analysis of two different studies. Patients and measurements Patients with primary adrenal insufficiency were on stable hydrocortisone and fludrocortisone therapy. Patients with secondary adrenal insufficiency received two different doses of hydrocortisone in a randomized crossover study. Main outcome measures were concentrations of precursors of cortisol and aldosterone measured by LC-MS/MS Results Compared to controls, progressively lower concentrations of the glucocorticoid precursors 11-deoxycortisol, 11-deoxycorticosterone and corticosterone concentrations were found in patients with secondary adrenal insufficiency on lower hydrocortisone dose, secondary adrenal insufficiency on higher hydrocortisone dose and primary adrenal insufficiency, respectively. Half of the primary adrenal insufficient patients showed evidence of residual endogenous cortisol or aldosterone synthesis, as determined by quantifiable 11-deoxycortisol, 11-deoxycorticosterone and corticosterone conce ntrations. In secondary adrenal insufficient patients with higher endogenous cortisol production, as indicated by 11-deoxycortisol concentrations above the median, no increased cortisol exposure was observed both by plasma pharmacokinetic parameters and 24-hour free cortisol excretion in urine. Conclusions Adrenal corticosteroid production is likely to continue during treatment in a considerable percentage of patients with both primary and secondary adrenal insufficiency. In patients with secondary adrenal insufficiency, this synthesis appears to be sensitive to the dose of hydrocortisone. However, the residual corticosteroid concentrations were quantitatively low and its clinical significance remains therefore to be determined