18
In the final step of the DBS sampling process, the sample extract obtained from Figure 5b is aspirated into the pipette tip and transferred to the inlet of the microfabricated ESI chip in the same manner as described above. This extract is then sprayed through a nanospray nozzle in the ESI chip while collecting the mass spectrometric response of the analyte of interest using either full-scan mass spectral acquisition or selected reaction monitoring (SRM) procedures for quantitative analysis applications. In the present version each of the four sample spots on the card may be robotically sampled and analysed followed by manual removal of the card from the card holder and placement of the next card into the holder for its similar analysis. This strategy will be useful when separate time points are collected on adjacent card spots. It should be noted that this DBS approach involves no punching of individual spots which is customarily described in the literature [9, 15], nor does it require additional card treatment to alter the surface hydrophobicity.
Results
To demonstrate proof-of-principle for the DBS card holder/clamp described above human Li heparin whole blood samples were fortified with amlodipine across a standard curve concentration range of 10 ng/mL to 10,000 ng/mL. Duplicate 15 microliter samples at two different concentrations were manually dispensed via a 25 microlitre syringe onto Whatman blue ‘DMPK’ paper cards. As an example, standard one was dispensed in the centre of the first two circled regions on the DBS card while standard two was dispensed in duplicate within the second two circled regions of the same DBS card. Thus a set of four DBS cards contained the first set of eight standards for the calibration curve. A second set of four identical cards were prepared with these eight standards to allow for the acquisition of a duplicate set of standard curve entries.
Figure 6 shows the calibration curve for the quantitative determination of amlodipine in fortified human whole blood using tetradeuteriated amlodipone as the internal standard. The dynamic range was linear from 10 ng/mL to 10,000 ng/mL amlodipine with a correlation coefficient of 0.9999. Since this work was exploratory, for proof-of-principle at this early stage, the additional studies common to regulated bioanalysis such as recovery, stability, and reproducibility, were not undertaken in this study.
Conclusions
The card holder described in this report provides a confined sample extraction region located within the sample spot for a dried blood or similar biological matrix. The described approach does not depend upon the porosity of the paper or other substrate upon which the sample matrix is applied. The LESA sampling pipette allows micro liquid extraction of a 3mm area of the sample from a porous substrate followed by direct infusion nanoelectrospray mass spectrometric analysis. Inter sample extraction and mass spectrometric analysis of the four spots contained on a given sample card is fully automated.
Acknowledgements
We thank Gary van Berkel and Vilmos Kertesz of Oak Ridge National Laboratories (ORNL) for helpful discussion and AB SCIEX for a generous loan of the 4000 QTRAP mass spectrometer employed in this work.
References
1.Yamashita, M. and J.B. Fenn, Electrospray ion source. Another variation on the free-jet theme. J. Phys. Chem, 1984. 92: p. 4451-4459.
2. Fenn, J.B, et al, Electrospray ionization for mass spectrometry of large molecules. Science, 1989. 246(October): p. 64-71.
3. Whitehouse, C.M, et al, Electrospray interface for liquid chromatographs and mass spectrometers. Anal. Chem, 1985. 57: p. 675-679.
4. Covey, T.R, E.D. Lee, and J.D. Henion, High-speed liquid chromatography/tandem mass spectrometry for the determination of drugs in biological samples. Anal Chem, 1986. 58(12): p. 2453-60.
5. Henion, J.D, The origins of ion spray liquid chromatography-tandem mass spectrometry. Clin Chem, 2009. 55(6): p. 1234-5.
6. Henion, J.D, The reality of lab-on-a-chip technology for the mass spectrometry laboratory. LCGC North America, 2009. 27(10): p. 1-10.
7. King, R, et al, Mechanistic investigation of ionization suppression in electrospray ionization. J. Am. Soc. Mass Spectrom, 2000. 11: p. 942-950.
8. Dethy, J.M, et al, Demonstration of direct bioanalysis of drugs in plasma using nanoelectrospray infusion from a silicon chip coupled with tandem mass spectrometry. Anal Chem, 2003. 75(4): p. 805-11.
9. Spooner, N, R. Lad, and M. Barfield, Dried Blood Spots as a Sample Collection Technique for the Determination of Pharmacokinetics in Clinical Studies: Considerations for the Validation of a Quantitative Bioanalytical Method. Analytical Chemistry, 2009. 81(4): p. 1557-1563.
10. Chace, D.H, Mass spectrometry in newborn and metabolic screening; historical perspective and future directions. J. Mass Spectrom, 2008. 44: p. 163-170.
11. Kertesz, V. and G.J. Van Berkel, Fully automated liquid extraction-based surface sampling and ionization using a chip-based robotic nanoelectrospray platform. Journal of Mass Spectrometry. J. Mass Spectrom, 2010. 45: p. 252-260
12. Stankovich, J.J, et al, Liquid Microjunction Surface Sampling Analysis of Dried Blood Spots Using an automated chip-based nano-ESI Infusion Device (LMJ-SS/Nano-ESI/MS) Rapid Communication in Mass Spectrometry (in preparation), 2011.
13. Edwards, R, et al, Hemoglobin Variant Analysis via Direct Surface Sampling of Dried Blood Spots Coupled with High-Resolution Mass Spectrometry. Anal. Chem., 2011. 83(6): p. 2265-2270.
14. Alpha, C, et al, Liquid Extraction Surface Analysis (LESA) of Dried Blood Spot Cards via Chip-Based Nanoelectrospray for Drug and Drug Metabolite Monitoring Studies, in 58th Conference on Mass Spectrometry and Allied Tipics 2010: Salt Lake City, UT.
15. Abu-Rabie, P. and N. Spooner, Direct Quantitative Bioanalysis of Drugs in Dried Blood Spot Samples Using a Thin-Layer Chromatography Mass Spectrometer Interface. Analytical Chemistry, 2009. 81(24): p. 10275- 10284.
Reproduced from Mass Matters 66th edition 2011, courtesy of the British Mass Spectrometry Society. Dr Frank Pullen’s article was page 30 and the Advion article on dried blood spot analysis.
Landmark Imaging System Sale
Paraytec Ltd have sold their 50th ActiPix UV Imaging System. This milestone sale was achieved in a new applications area for Paraytec in the pharmaceutical industry. Paraytec’s latest system, the SDI300 allows pharmaceutical formulators to directly image drugs dissolving from a tablet surface, to enable more complete understanding of the drug behaviour in the body. This 50th system was sold in the United States by Paraytec’s distribution partner, Distek Inc, to a large pharmaceutical company in California. The addition of this system to their laboratory will enable the scientists to avoid pitfalls in the development process, by providing essential understanding of an API (Active Pharmaceutical Ingredient) at an earlier stage than possible with traditional dissolution techniques.
Jeff Seely, Global Manager, Sales, Marketing & Technical Service for Distek, Inc said of the SDI300 system: “The new dimension the SDI300 brings to pharmaceutical API testing further enhances our offering to our customers.” Mark Vaux of Paraytec Ltd, commented: “The in-depth knowledge Distek has of the US pharmaceutical industry enables the rapid uptake of Paraytec’s technology, as we move towards having multiple systems in the major pharmaceutical companies.”
The SDI300 is designed to provide key drug dissolution information on the new generation of sparingly soluble drug types, which require a new approach in formulation science. Circle no. 48
Anytime, Anywhere...
Online INTERNATIONAL LABMATE - JANUARY/FEBRUARY 2012
Page 1 |
Page 2 |
Page 3 |
Page 4 |
Page 5 |
Page 6 |
Page 7 |
Page 8 |
Page 9 |
Page 10 |
Page 11 |
Page 12 |
Page 13 |
Page 14 |
Page 15 |
Page 16 |
Page 17 |
Page 18 |
Page 19 |
Page 20 |
Page 21 |
Page 22 |
Page 23 |
Page 24 |
Page 25 |
Page 26 |
Page 27 |
Page 28 |
Page 29 |
Page 30 |
Page 31 |
Page 32 |
Page 33 |
Page 34 |
Page 35 |
Page 36 |
Page 37 |
Page 38 |
Page 39 |
Page 40 |
Page 41 |
Page 42 |
Page 43 |
Page 44 |
Page 45 |
Page 46 |
Page 47 |
Page 48 |
Page 49 |
Page 50 |
Page 51 |
Page 52 |
Page 53 |
Page 54 |
Page 55 |
Page 56 |
Page 57 |
Page 58 |
Page 59 |
Page 60 |
Page 61 |
Page 62 |
Page 63 |
Page 64 |
Page 65 |
Page 66 |
Page 67 |
Page 68 |
Page 69 |
Page 70 |
Page 71 |
Page 72 |
Page 73 |
Page 74 |
Page 75 |
Page 76 |
Page 77 |
Page 78 |
Page 79 |
Page 80 |
Page 81 |
Page 82 |
Page 83 |
Page 84 |
Page 85 |
Page 86 |
Page 87 |
Page 88 |
Page 89 |
Page 90 |
Page 91 |
Page 92
