Overcoming matrix effects in electrospray: Quantitation of β-agonists in complex matrices by isotope dilution liquid chromatography–mass spectrometry using singly 13C-labeled analogues

In this work, the implementation of isotope dilution mass spectrometry (IDMS) using minimal labeling and isotope pattern deconvolution (IPD) is evaluated as a strategy for the minimization of matrix effects during trace determination of β2-agonists in complex matrices by liquid chromatography electrospray ionization mass spectrometry (LC–ESI-MS). First, the parameters affecting the measurement of isotopic composition of organic compounds by liquid chromatography electrospray ionization high resolution mass spectrometry with a time-of-flight analyzer were evaluated using as a case of study three different β2-agonists: clenbuterol, clenproperol and brombuterol. Then, a calibration graph-free IDMS methodology was evaluated in order to overcome matrix effects in LC–ESI-MS in complex samples. In this procedure singly 13C-labeled analogues of clenbuterol, clenproperol and brombuterol were employed in combination with IPD. Using this approach accurate and precise results were obtained in the simultaneous quantification of β2-agonists in human urine and bovine liver, even at the sub ng g−1 and particularly in spite of the previously reported matrix effects. Recovery rates in the range of 97–114% in fortified human urine and from 95% to 111% in fortified bovine liver were obtained with RSD (%) of independent recovery experiments always lower than 6%. These results demonstrate that the proposed methodology based on the use of 13C1-labeled standards and IPD is a reliable approach for accurate LC–MS quantitation of small molecules and compatible with full-scan high-resolution mass spectrometry.

► A novel strategy based on isotope dilution mass spectrometry for LC–MS is proposed.
► Significant minimization of matrix effects occurring in electrospray LC–MS.
► Study of parameters affecting the measurement of isotope abundances by LC–TOFMS.
► Increased accuracy and precision for ultratrace level detection in complex matrices.
► Applied successfully to ultratrace detection of β-agonists in urine and liver.