Influence of thyroidectomy on thyroxine metabolism and turnover rate in rats

Little is known about the kinetics and metabolism of thyroid hormones in the hypothyroid state. To investigate these factors, we developed a reliable method for measurement of serum thyroxine (T4), triiodothyronine (T3), reverse-T3 (rT3) and stable isotope-labeled T4 ([13C9]T4), using online solid-phase extraction liquid chromatography–mass spectrometry/mass spectrometry (online SPE LC–MS/MS). We measured supply and turnover rates of T4 in thyroidectomized (Tx) rats using [13C9]T4 as a tracer. In rats, serum T4, T3 and rT3 were decreased but not completely ablated after surgical Tx. Endogenous T4 and T3 levels in Tx rats were maintained at a constant low level throughout the experimental period. [13C9]T4 levels declined with a half-life of ∼1.2 days after it was administered to Tx rats intravenously. These findings strongly suggest that serum T4 levels in Tx rats are maintained by T4 supplied by extra-thyroidal tissues (e.g. secretion of extra-thyroidal storage, enhancement of enterohepatic recirculation, and production in extra-thyroidal tissues). Moreover, the turnover rate of T4 in Tx rats was approximately twofold lower than in controls. This finding suggests that degradation of serum T4 is repressed by Tx. In conclusion, serum T4 is maintained at a constant low level by T4 supply from extra-thyroidal tissues and repression of T4 degradation in Tx rats. The powerful online SPE LC–MS/MS tool can be used to investigate thyroid hormones kinetics and metabolism, and thus has the potential to be used as a diagnostic tool and to investigate the pathogenesis of thyroid disease.

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Development of a High-Throughput Online Solid-Phase Extraction/Tandem Mass Spectrometry Method for Cytochrome P450 Inhibition Screening

CrossRef Listing of Deleted DOIs ◽

10.1177/1087057110362581 ◽

2010 ◽

Vol 15 (4) ◽

pp. 447-452 ◽

Cited By ~ 31

Author(s):

Kheng B. Lim ◽

Can C. Özbal ◽

Daniel B. Kassel

Keyword(s):

Mass Spectrometry ◽

Cytochrome P450 ◽

Tandem Mass Spectrometry ◽

Solid Phase Extraction ◽

High Throughput ◽

Solid Phase ◽

Tandem Mass ◽

Phase Extraction ◽

Online Spe ◽

Online Solid Phase Extraction

A high-throughput online solid-phase extraction/tandem mass spectrometry (online SPE/MS/MS) system has been developed to support rapid evaluation of drug discovery compounds for possible drug-drug interaction (DDI). Each compound is evaluated for its DDI potential by incubating over a range of 8 test concentrations and against a panel of 6 cytochrome P450 (CYP) enzymes, 1A2, 2C8, 2C9, 2C19, 2D6, and 3A4. Previously, a postassay pooling and a 2-min gradient LC/MS/MS method had been reported to increase sample throughput, allowing for a 96-well plate of samples to be analyzed in under 4 h. The development of a new online SPE/MS/MS system has reduced the analysis time to less than 15 min per 96-well plate, translating to a 15-fold time savings compared to the 2-min LC/MS/MS method. Sampling precision without internal standard correction ranged from 3.1% to 5.6% relative standard deviation, and the carryover was determined to be between 1.0% and 4.1%. One hundred twenty in-house compounds were assayed and pooled for analyses using both the online SPE/MS/MS and LC/MS/MS, and the correlation coefficients ranged from 0.89 to 1.13, when comparing the IC50 results obtained from the 2 approaches for each of the CYP enzymes.

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A Single LC-MS/MS Analysis to Quantify CoA Biosynthetic Intermediates and Short-Chain Acyl CoAs

Metabolites ◽

10.3390/metabo11080468 ◽

2021 ◽

Vol 11 (8) ◽

pp. 468

Author(s):

Anthony E. Jones ◽

Nataly J. Arias ◽

Aracely Acevedo ◽

Srinivasa T. Reddy ◽

Ajit S. Divakaruni ◽

...

Keyword(s):

Mass Spectrometry ◽

Sample Preparation ◽

Biosynthetic Pathway ◽

Solid Phase ◽

High Sensitivity ◽

Short Chain ◽

Extraction Column ◽

Chain Acyl ◽

Precision And Accuracy ◽

Mass Spectrometry Mass Spectrometry

Coenzyme A (CoA) is an essential cofactor for dozens of reactions in intermediary metabolism. Dysregulation of CoA synthesis or acyl CoA metabolism can result in metabolic or neurodegenerative disease. Although several methods use liquid chromatography coupled with mass spectrometry/mass spectrometry (LC-MS/MS) to quantify acyl CoA levels in biological samples, few allow for simultaneous measurement of intermediates in the CoA biosynthetic pathway. Here we describe a simple sample preparation and LC-MS/MS method that can measure both short-chain acyl CoAs and biosynthetic precursors of CoA. The method does not require use of a solid phase extraction column during sample preparation and exhibits high sensitivity, precision, and accuracy. It reproduces expected changes from known effectors of cellular CoA homeostasis and helps clarify the mechanism by which excess concentrations of etomoxir reduce intracellular CoA levels.

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