Differential Mobility Spectrometry of Derivatized Steroid Hormones: Examining the Relationships between Structures and Solvation Chang Liu, J. Larry Campbell, Yves Le Blanc, Subhakar Dey, Subhasish Purkayastha, Tim Hoffman ASMS 2014, Baltimore
Ion Solvation The interaction of ions with solvent molecules One of the most important physicochemical properties to explore in drug discovery and development. A measure of the ability of studied ions to distribute in biological systems. Ion-solvent interaction in the gas-phase studied with differential mobility spectrometry (DMS). 2 2014 AB SCIEX
Differential mobility spectrometry (DMS) Ion species in gas stream + High field Declustering High mobility High field Declustering High mobility The magnitude of the compensation voltage is a measure of the ion solvation strength. 3 2014 AB SCIEX Low field Clustering Low mobility Low field Clustering Low mobility Krylov et al., Int. J. Mass Spectrom. 2009, 285, 149.
Differential mobility spectrometry (DMS) Separation voltage (SV) stepped from 0 to 4000 V. Compensation voltage (CV) scanned from -20 V to +15 V. The optimal CV for the ion transmission was determined. e.g. SV=4000 no chemical modifier 1.5% acetonitrile added into the DMS cell Reletive intensity 1.0 0.8 FWHM=2.7 0.6 0.4 0.2 0.0-20 -10 0 10 Reletive intensity 1.0 0.8 FWHM=2.7 0.6 0.4 0.2 0.0-20 -10 0 10 4 2014 AB SCIEX
Dispersion plots TYPE C Bad clustering, hard-sphere model Lower mobility at high-field TYPE B Mixed mechanism TYPE A Good clustering Higher mobility at high-field 5 2014 AB SCIEX Purves and Guevremont, Anal. Chem. 1999, 71, 2346
6 2014 AB SCIEX Experimental conditions
Steroid hormones dihydrotestosterone testosterone boldenone mesterolone 17a-methyltestosterone androsterone dehydroandrosterone 7 2014 AB SCIEX
Aminooxy derivatization reagents * * Amplifex TM Keto reagent, commercially available from AB SCIEX 8 2014 AB SCIEX Star-Weinstock et al., Anal. Chem. 2012, 84, 9310
Experimental conditions e.g. Derivatization DMS-MS system + methanol, acetic acid room temp, 1h AB SCIEX SelexION TM Differential Mobility Spectrometer with QTRAP 5500 Mass Spectrometer system. 9 2014 AB SCIEX Star-Weinstock et al., Anal. Chem. 2012, 84, 9310
Results and Discussion The effect of chemical modifiers The structure of aminooxy derivatization reagents The structure of steroids 10 2014 AB SCIEX
Ion solvation strength The effect of chemical modifier e.g. 10 5 no chemical modifier methanol 0-5 isopropanol -10 acetonitrile acetone 11 2014 AB SCIEX -15 0 1000 2000 3000 4000 SV (V) The ion solvation strength: methanol < isopropanol < acetonitrile < acetone
10 Ion solvation strength Chemical modifier: selectivity 2D Graph 4 10 10 best selectivity 10 derivatized testosterone derivatized boldenone derivatized dihydrotestosterone derivatized 17a-methyltestosterone derivatized mesterolone derivatized androsterone derivatized dehydroandrosterone no chemical modifier methanol isopropanol acetonitrile acetone 10 5 5 5 5 5 0-5 0-5 0-5 0-5 0-5 -10-10 -10-10 -10-15 0 1000 2000 3000 4000 SV (V) -15 0 1000 2000 3000 4000 SV (V) -15 0 1000 2000 3000 4000 SV (V) -15 0 1000 2000 3000 4000 SV (V) -15 0 1000 2000 3000 4000 SV (V) 12 2014 AB SCIEX
Results and Discussion The effect of chemical modifiers The structure of aminooxy derivatization reagents The structure of steroids Example: derivatized testosterone 13 2014 AB SCIEX
Relationships between structures and solvation The structure of aminooxy derivatization reagents Group A R= Relative intensity Chemical modifier: isopropanol; SV=4000 1.0 0.8 0.6 0.4 A1-testosterone A2-testosterone 0.2 A3-testosterone 0.0-20 -10 0 10 The presence of the intramolecular hydrogen bond A1 A2 A3 The solvation strength of A2-contained derivatized steroids was weaker than corresponding ions containing A1/A3. ΔG = -13 kcal/mol 14 2014 AB SCIEX
Relationships between structures and solvation The structure of aminooxy derivatization reagents Group B R= B1 B2 B3 Chemical modifier: isopropanol; SV=4000 Relative intensity 1.0 0.8 0.6 0.4 0.2 B1-testosterone B2-testosterone B3-testosterone The solvation strength of B2-contained derivatized steroids was stronger than corresponding ions containing B1/B3. 0.0-20 -10 0 10 The charge site of B2 derivatized steroids is least steric hindered. 15 2014 AB SCIEX
Relationships between structures and solvation The structure of aminooxy derivatization reagents Group C R= Relative intensity Chemical modifier: isopropanol; SV=4000 1.0 0.8 0.6 0.4 0.2 C1-testosterone C2-testosterone C3-testosterone 0.0-20 -10 0 10 The protonation site C1 C2 C3 The solvation strength of C2-contained derivatized steroids was stronger than corresponding ions containing C1/C3. 16 2014 AB SCIEX
Results and Discussion The effect of chemical modifiers The structure of aminooxy derivatization reagents The structure of steroids Example: steroids derivatized by 17 2014 AB SCIEX
Relationships between structures and solvation The structure of steroids: the additional methyl group Chemical modifier: isopropanol; SV=4000 testosterone 1 17a-methyltestosterone Reletive intensity 1.0 0.8 0.6 0.4 0.2 0.0-10 -5 0 5 10 derivatized testosterone derivatized 17a-testosterone 1.0 dihydrotestosterone mesterolone Reletive intensity 0.8 0.6 0.4 0.2 0.0-10 -5 0 5 10 The additional methyl group reduces the solvation strength of derivatized steroids derivatized dihydrotestosterone derivatized mesterolone 18 2014 AB SCIEX The steric effect
Relationships between structures and solvation The structure of steroids: double bonds dihydrotestosterone testosterone boldenone Relative intensity 1.0 0.8 0.6 Chemical modifier: isopropanol; SV=4000 0.4 derivatized dihydrotestosterone 0.2 derivatized testosterone derivatized boldenone 0.0-10 -5 0 5 10 Double bonds reduced the ion solvation strength. The electron density on N is enhanced by the conjugated double bond 19 2014 AB SCIEX
Relationships between structures and solvation The structure of steroids: the ketone position 1 dihydrotestosterone 1 androsterone Relative intensity 1.0 0.8 0.6 0.4 Chemical modifier: isopropanol; SV=4000 0.2 derivatized dihydrotestosterone derivatized androsterone 0.0-10 -5 0 5 10 The solvation strength of derivatized dihydrotestosterone was stronger than corresponding derivatized androsterone. The charge site of derivatized androsteorne is more steric hindered. 20 2014 AB SCIEX
Conclusions Differential mobility spectrometry (DMS) is a useful tool to study the ion solvation strength. Structurally similar derivatized steroid hormones could be differentiated in DMS, with the addition of chemical modifier. The structure of intramolecular hydrogen bond was predicted (weak ion solvation strength). The steric effect is another important factor of influencing the ion solvation strength. Results of DMS analysis agrees well with basic theoretical modeling. More intricate ion-solvent cluster modeling is underway*. *Campbell et al., J. Am. Soc. Mass Spectrom. 2014, in press; Poster: ThP 505 21 2014 AB SCIEX
Acknowledgements Bradley Schneider Tom Covey Deolinda Fernandes Adam Latawiec Michal Star-Weinstock Brain Williamson Sasi Pillai 22 2014 AB SCIEX
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