Sensitive UHPLC-MS/MS technique for monitoring Levothyroxine (T4) in human serum against endogenous thyroxin level

Background: Levothyroxine is a synthetic thyroid hormone that is chemically identical to Thyroxine (T4), which is secreted by the follicular cells of the thyroid gland. Levothyroxine is used to treat deficiency of thyroid hormone and to prevent the recurrence of thyroid cancer. Levothyroxine is present endogenously in human body. Method: It requires treated matrix for the preparation of calibration curve standard and quality control samples. The method was developed using LC-MS/MS and validated in human charcoal stripped serum. Charcoal stripped matrix was used for the preparation of Calibration curve standards and Quality control samples. Method involves Solid-Phase Extraction technique. Levothyroxine D3 used as an internal standard (ISTD). Result: Chromatographic separation was achieved using reversed phase analytical column Gemini NX-C18 110Å, 3µm (50x3.6) mm. Mobile phase consisted of acetonitrile and water in a ratio of 70:30 with 150µL of formic acid in 1000 mL of mobile phase. Mobile phase achieved shorter run-time of 0.9 minute due to use of Ultra-high performance liquid chromatography (UHPLC). Positive electro-spray ionization technique detected MRM ion pair transitions 777.60→731.65 for Levothyroxine and 780.70→734.6 for Levothyroxine- D3 (ISTD) were used. AB SCIEX Triple Quad™ API-4000 LC-MS/MS system and the bioanalytical method with 10ng/mL as limit of quantification has been applied successfully to pharmacokinetics studies. Conclusion: Chromatographic separation was achieved using reversed phase analytical column Gemini NX-C18 110Å, 3µm (50x3.6) mm. Mobile phase consisted of acetonitrile and water in a ratio of 70:30 with 150µL of formic acid in 1000 mL of mobile phase. Mobile phase achieved shorter run-time of 0.9 minute due to use of Ultra-high performance liquid chromatography (UHPLC). Positive electro-spray ionization technique detected MRM ion pair transitions 777.60→731.65 for Levothyroxine and 780.70→734.6 for Levothyroxine- D3 (ISTD) were used. AB SCIEX Triple Quad™ API-4000 LC-MS/MS system and the bioanalytical method with 10ng/mL as limit of quantification has been applied successfully to pharmacokinetics studies.

Fabricating an Electrospray Ionization Chip Based on Induced Polarization and Liquid Splitting

The coupling of the microfluidic chip to mass spectrometry (MS) has attracted considerable attention in the area of chemical and biological analysis. The most commonly used ionization technique in the chip–MS system is electrospray ionization (ESI). Traditional chip-based ESI devices mainly employ direct electrical contact between the electrode and the spray solvent. In this study, a microchip ESI source based on a novel polarization-splitting approach was developed. Specifically, the droplet in the microchannel is first polarized by the electric field and then split into two sub-droplets. In this process, the charge generated by polarization is retained in the liquid, resulting in the generation of two charged droplets with opposite polarities. Finally, when these charged droplets reach the emitter, the electrospray process is initiated and both positive and negative ions are formed from the same solution. Preliminary experimental results indicate that the coupling of this polarization-splitting ESI (PS-ESI) chip with a mass spectrometer enables conventional ESI-MS analysis of various analytes.

Ambient Electric Arc Ionization for Versatile Sample Analysis Using Mass Spectrometry

A novel, convenient ambient electric arc ionization (AEAI) device was developed as mass spectrometry ion source for versatile sample analysis. AEAI could be considered as a soft ionization technique in...

UPLC-MS/MS method for determination of retinol and α-tocopherol in serum using a simple sample pretreatment and UniSpray as ionization technique to reduce matrix effects

BackgroundOur goal was to develop a simple, rapid and precise ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the determination of retinol and α-tocopherol in serum. Currently published LC-MS/MS methods either require complex extraction procedures (liquid-liquid or solid-phase) or do not meet desirable specifications for imprecision in serum (coefficient of variation [CV] <6.8% and 6.9%, respectively).MethodsSample preparation consisted of a simple protein precipitation with ethanol and acetonitrile. Stable isotope-labeled internal standards (IS) and a homemade calibration curve were used for quantification. The analysis was performed using an Acquity I-class Xevo TQ XS LC-MS/MS. Chromatographic runtime was 6.0 min using a reversed phase gradient elution. UniSpray (US) as an ionization technique was compared to electrospray ionization (ESI). Analytical validation included matrix effect, recovery and trueness compared to National Institute of Standards and Technology (NIST) standards and United Kingdom National External Quality Assessment Service (UK NEQAS) samples.ResultsIntra- and inter-run CVs were <4.9% for retinol and <1.7% for α-tocopherol, both complying with desirable specifications for imprecision. Bias compared to NIST standards was <3.1% for both compounds. The method was linear over the entire tested range. The lower limit of quantification (LLOQ) with US was lower than with ESI for both retinol (0.022 vs. 0.043 mg/L) and α-tocopherol (0.22 vs. 0.87 mg/L). Matrix effects were not significant (<15%) for retinol. However, for α-tocopherol matrix effects of on average 54.0% were noted using ESI, but not with US.ConclusionsWe developed a fast, precise and accurate UPLC-MS/MS method for the determination of retinol and α-tocopherol in human serum using a single-step sample pretreatment. Ionization using US eliminated the matrix effects for α-tocopherol.

Simultaneous ultrafast determination of six alkaloids in mainstream cigarette smoke by DART-MS/MS

To determine nicotine, nornicotine, myosmine, anatabine, anabasine and nicotyrine in mainstream cigarette smoke simultaneously, a novel method based on Direct Analysis in Real Time (DART) Ionization technique combined with mass spectrometry was established by optimizing parameters such as injection rate, type of extraction solvent and extraction solvent dosage.