Glucagon like peptide-1 analogues analysis in whole blood samples by the use of LC-ESI-HRMS method

IF 1.8 Q4 TOXICOLOGY
Nadia Arbouche , Alice Ameline , Laurie Gheddar , Jean-Sébastien Raul , Pascal Kintz
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引用次数: 0

Abstract

Objective

The use of GLP-1 analogues has been increasing worldwide in recent years due to their benefits in treating type II diabetes. Thanks to their effects on appetite regulation, in many countries they are also used, especially semaglutide, to treat obesity. However, due to their promotion by social media and celebrities as a weight-loss treatment, GLP-1 analogues are misused by a non-diabetic and non-obese population and also by a young public, which is the main target of these media. Following the alert by the ANSM (Agence nationale de sécurité du médicament) in France and the FDA (Food and Drug Administration) in the United States, which imposed the addition of fatal effects to the list of side effects for semaglutide, the misuse seems to become a public health problem. For this reason, it seems important that a toxicology laboratory is able to evidence the presence of these drugs in blood. In this study, the authors have developed and validated a method for the identification and quantification of semaglutide and liraglutide in whole blood using a LC-HRMS method.

Methods

LC separation was achieved using a Waters Acquity HSS C18 column (150 × 2.1 × 1.8 μm) with a controlled temperature maintained at 50 °C. A 5-μL injection with a 0.25 mL/min flow of waters with 0.1% of formic acid (solvent A) and acetonitrile with 0.1% of formic acid (solvent B) was used. A Xevo G2-XS Q-TOF high-resolution mass spectrometer (Waters corporation, Milford, MA, USA) was used, operating in positive ion mode and in sensitivity mode. In MS scanning, data were acquired from 500 to 2000 m/z. For semaglutide, the 4-fold charged [M + 4H]4+ molecule was observed at m/z 1029.29752 (which deconvolutes to 4117.70139), while for liraglutide the 4-fold charged ion [M + 4H]4+ was observed at 938.76490 (which deconvolutes to 3754.53707).
The extraction was performed by blood protein precipitation using a mix of ACN/MeOH (70:30), after the addition of 50 ng/mL of internal standard (bovin insulin). The method was applied to authentic whole blood samples following a hospital request for GLP-1 analogs determination and to postmortem blood samples. The blood samples were stored at + 4 °C until analysis.

Results and discussion

Usual therapeutic blood concentrations of both drugs are in the range 50-150 ng/mL. The validation procedure demonstrated an acceptable linearity between 2 and 500 ng/mL. LOD and LOQ were 1 and 2 ng/mL, respectively. Intra and inter-day precision were below 20% at three concentrations. The method was successfully applied to the blood samples of 3 diabetic patients under treatment of semaglutide (concentrations ranged from 31 to 70 ng/mL) and to one postmortem blood sample which tested positive for liraglutide at 12 ng/mL. These concentrations fall within the limits of therapeutic blood concentrations described in the literature.

Conclusion

The use of semaglutide and liraglutide is increasing for the treatment of type II diabetes but along with its therapeutic use, its misuse for weight loss is also increasing. Because of their side effects, which can be lethal, it seems essential to be able to document an intake of these drugs. The method developed aims to highlight a therapeutic administration of semaglutide and liraglutide, to be able to also evidence the presence of traces in case of improper administration and also to ensure a forensic application on whole blood, which is the most common matrix in this field.
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来源期刊
CiteScore
0.90
自引率
33.30%
发文量
393
审稿时长
47 days
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