[Simultaneous rapid determination of 37 alkaloids in whole blood by high performance liquid chromatography-mass...

Objective: To establish a rapid and sensitive high performance liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) method for the simultaneous determination of 37 alkaloids in whole blood, in order to meet the needs of clinical alkaloid toxicity detection. Methods: From October 2024 to January 2025, whole blood samples were precipitated with acetonitrile and methanol (V∶V=1∶1) to remove proteins, followed by centrifugation, filtration and sample injection. A gradient elution was performed using a mobile phase containing 0.1% formic acid acetonitrile, 0.1% formic acid, and 5 mmol/L ammonium acetate aqueous solution. The injection volume was 1 μl at a flow rate of 0.4 ml/min. Separation was achieved on Waters XSelect(®)CSH(TM) C18 column. Mass spectrometry was operated in positive electrospray ionization with multiple reaction monitoring (MRM) mode, and quantification was performed using a matrix working curve external standard method. Results: The linear relationships between 37 alkaloids in whole blood were good within the corresponding concentration range, with correlation coefficients (r) all greater than 0.990. The detection limits (S/N) were 0.01-2.50 ng/ml, and the quantification limits were 0.03-7.50 ng/ml. The recovery rates were 81.1%-122.1%, and the intra and inter batch accuracies were 1.0%-13.9% and 2.3%-17.9%, respectively. Conclusion: This study establishes a rapid and sensitive LC-MS/MS for the simultaneous determination of 37 alkaloids in whole blood. The method is concise, rapid and sensitive, and can be used for the quantification of alkaloid poisoning toxins, monitoring of blood concentrations of medicinal alkaloids, and the detection of emergency response to sudden food poisoning incidents.