Quantitation of Tacrolimus in Dried Blood Spots Using Paper Spray Mass Spectrometry

Paper spray mass spectrometry (MS) allows for the direct analysis of pharmaceuticals, drugs of abuse, and other small molecules from blood, urine, and other biofluids (1,2). Described here is the application of a Velox 360 Paper spray autosampler and ion source for the direct analysis of tacrolimus, an immunosuppressant drug, from dried blood spots and its comparison to HPLC–MS-MS.

Paper spray is performed by depositing the sample directly onto a porous cellulosic substrate contained within a disposable cartridge (Velox Sample Cartridge). The sample is allowed to dry, and the cartridge is inserted into the automated ion source for analysis. The Velox 360 performs all steps necessary to perform the analysis, including loading the cartridge, depositing extraction solvent onto the cartridge, positioning the cartridge in front of the mass spectrometer inlet, and applying ionization voltage. Ions are generated from the sharp tip of the porous substrate contained within the cartridge. The Velox 360 provides an automated, cost effective, and efficient method for therapeutic drug monitoring (TDM) using mass spectrometry with minimal sample preparation.

Experimental

A 200 µL blood sample was mixed with 10 µL of internal standard solution containing 150 ng/mL of [¹³C, ²H₂]-tacrolimus from Toronto Research Chemicals. 10 µL was spotted onto the cartridge and allowed to dry either at room temperature for 90 min or at 40 °C for 20 min. Sample analysis was performed using a Velox 360 prototype interfaced with a Thermo Scientific TSQ Vantage.

Results

Selective detection of the analytes was confirmed by requiring that the ratio between the quantifier and the qualifier SRM transitions be within 20% of an authentic sample. The accuracy of the Velox 360 MS-MS assay was confirmed by validating it against an HPLC–MS-MS assay at a reference lab. The results for 60 incurred samples, shown in Figure 1, show excellent agreement between the two methods.

Conclusion

This application note demonstrates the suitability of paper spray MS for the quantitative analysis of tacrolimus in dried blood spots. Paper spray MS-MS maintains the excellent sensitivity and specificity of mass spectrometry while being better suited for laboratories with limited expertise in the maintenance and troubleshooting of complex HPLC–MS-MS systems. The following advantages are realized using paper spray MS:

1. No sample preparation: Analysis is done directly from a DBS.

2. Rapid analysis: Less than 2 min per sample for the entire analysis.

3. No carryover, because the analysis is done from a single use cartridge

4. No chromatography: That means there are no columns to replace, no carryover on the column, and no chromatography methods to develop.

References

(1) H. Wang, J. Liu, R.G. Cooks, and Z. Ouyang, Ang. Chem. Int. Ed., 49, 877–880 (2010).

(2) N.E. Manicke, Q.A. Yang, H. Wang, S. Oradu, Z. Ouyang, and R.G. Cooks, Int. J. Mass Spectrom. 300, 123–129 (2011).

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