Ute Gerhard | University of Hertfordshire (original) (raw)
Papers by Ute Gerhard
Journal of Organic Chemistry, Jul 19, 2000
Journal of Mass Spectrometry, Aug 1, 2004
Techniques such as mass spectrometry and NMR spectroscopy form an essential part of the medicinal... more Techniques such as mass spectrometry and NMR spectroscopy form an essential part of the medicinal chemist's toolbox for characterizing and assessing the purity of new molecules. Empowering medicinal chemists to gain early insight into their reaction products has a direct impact on productivity. Devolution of cutting-edge techniques from the specialist to the bench chemist also frees the specialist to concentrate on solving the more demanding of analytical problems. For open-access techniques to be taken up, they must be robust and be able to handle differing sample concentrations and varying sample complexities. This paper details the implementation of high-resolution liquid chromatography/mass spectrometry in open access to aid the medicinal chemist in characterizing desired products and identifying unexpected rearrangements, by-products and complete unknowns.
Proceedings of the National Academy of Sciences, 1993
An approach toward the estimation of binding constants for organic molecules in aqueous solution ... more An approach toward the estimation of binding constants for organic molecules in aqueous solution is presented, based upon a partitioning of the free energy of binding. Consideration is given to polar and hydrophobic contributions and to the entropic cost of rotor restrictions and bimolecular associations. Several parameters (derived from an analysis of entropy changes upon the melting of crystals and from the binding of cell wall peptide analogues to the antibiotic ristocetin A) which may be useful guides to a crude understanding of binding phenomena are presented: (i) amide-amide hydrogen bond strengths of -(1 to 7) +/- 2 kJ.mol-1, (ii) a hydrophobic effect of -0.2 +/- 0.05 kJ.mol-1.A-2 of hydrocarbon removed from exposure to water in the binding process, and (iii) free energy costs for rotor restrictions of 3.5-5.0 kJ.mol-1. The validity of the parameters for hydrogen bond strengths is dependent on the validity of the other two parameters. The phenomenon of entropy/enthalpy compen...
Journal of the American Chemical Society, 1994
It is demonstrated that the presence of bacterial cell wall analogues may either enhance or, in t... more It is demonstrated that the presence of bacterial cell wall analogues may either enhance or, in the case of ristocetin A, oppose dimerization of glycopeptide antibiotics. These observations may imply that dimerization plays a role in the mode of action of these antibiotics, and a mechanism is proposed to take account of this possibility. The glycopeptide dimers are also found to be formed more exothermically in the presence of cell wall analogues, and the nature of biological signaling events is discussed in this context. It is pointed out that binding enthalpy (rather than simply binding free energy, AG) may be an important quantity in signaling events. If this is so, then oligomers may be abundant in signaling processes partly because the extended aggregates they form are able to cooperatively amplify the conformational changes which are incurred on ligand binding, which occur through relatively small changes in free energy but larger opposing changes in enthalpy and entropy.
Journal of the American Chemical Society, 1994
A procedure for the determination of association constants in aqueous solution using hydrogendeut... more A procedure for the determination of association constants in aqueous solution using hydrogendeuterium exchange has been developed and used to measure the dimerization constant, &im, for a number of strongly dimerizing glycopeptide antibiotics. Thesevalues provide further insight into the thermodynamic contributions of various structural epitopes to the dimerization of these antibiotics. Consideration of ligand binding affinities together with dimerization potentials provides evidence that dimerization is implicated in the physiological mode of action of these antibiotics.
Journal of the American Chemical Society, 1991
An expression is presented for the estimation of approximate binding constants for bimolecular as... more An expression is presented for the estimation of approximate binding constants for bimolecular associations in solution. The consequences of the approach have been examined for the bimolecular association of two peptide components in aqueous solution: specifically for the binding of two vancomycin group antibiotics, vancomycin itself and ristocetin A, to the peptide cell wall analogue N-Ac-D-Ala-D-Ala and related ligands. Uncertainties in the treatment are relatively large, but the physical insights gained into the binding process (in part with the aid of calorimetric data obtained by others) are enlightening. We conclude that for amide-amide hydrogen bond formation in aqueous solution at room temperature, the intrinsic binding energy is ca. 24 kJ mol-' (an intrinsic binding constant of ca. lo4); this process is almost completely driven by a favorable entropy change associated with the release of water molecules from the amide NH and CO groups involved in hydrogen bond formation. The bimolecular association of N-Ac-D-Ala-D-Ala with ristocetin A has a remarkably small entropy change at 298 K (TAS = 3 f 1.5 kJ mol-'). We conclude that the release of water from polar and hydrocarbon groups involved in the binding almost exactly compensates for (i) the unfavorable entropy change due to the freezing out of four rotors of N-Ac-DAla-D-Ala upon binding and (ii) the unfavorable entropy change of a bimolecular association. A crude quantitation of these effects is presented. We also present an estimate of the increase in translational plus rotational free energy, as a function of the ligand mass, occurring when a ligand binds to a larger receptor. This quantity, fundamental to all binding processes, is relatively insensitive to the shape of the ligand. Extension of the approach will allow, in those cases where there is good complementarity between ligand and receptor, the prediction of approximate peptide-peptide binding constants in aqueous solution. 'Cambridge Centre for Molecular Recognition. $Smith Kline Beecham Pharmaceuticals.
The Journal of Antibiotics, 1995
The clinically important vancomycin group glycopeptide antibiotics, which act by blocking cell wa... more The clinically important vancomycin group glycopeptide antibiotics, which act by blocking cell wall synthesis, are crucial in the treatment of methicillin resistant Staphylococcus aureus. All of the group membersstudied so far, with the apparent exception of teicoplanin, enhance their antibiotic action by the formation of an asymmetric homodimer. Teicoplanin exists in two main conformers which differ by a rotation of approximately 1 80°ofa sugar residue. From NMRstudies and molecular modelling, we present structures for the two conformers and conclude that they have different binding affinities for cell wall analogues. The two conformers of teicoplanin are closely analogous to those adopted by each half of the asymmetric dimers of the other vancomycin group antibiotics. The vancomycin family of glycopeptide antibiotics1~4) are clinically important compounds, being active against Gram-positive bacteria. The emerging methicillin resistant Staphylococcus aureus (MRSA), which are re
Drug Metabolism and Disposition, 2002
[3R,5R,6S]-3-(2-cyclopropyloxy-5-trifluoromethoxyphenyl)-6-phenyl-1-oxa-7-azaspiro[4.5]decane is ... more [3R,5R,6S]-3-(2-cyclopropyloxy-5-trifluoromethoxyphenyl)-6-phenyl-1-oxa-7-azaspiro[4.5]decane is a substance P (Neurokinin 1 receptor) antagonist. Substance P antagonists are proven in concept to have excellent potential for the treatment of major depression, and they allow superior and sustained protection from acute and delayed chemotherapy-induced emesis. The metabolism of this compound was investigated in rat hepatocytes, and circulating rat plasma metabolites were identified following oral and intravenous dosing. The turnover in rat hepatocytes within 4 h was about 30%, and the major metabolites were identified as two nitrones and a lactam associated with the piperidine ring. Although these metabolites were also observed in rat plasma, the major circulating metabolite was a keto acid following oxidative de-amination of the piperidine ring. Liquid chromatography/tandem mass spectrometry and nuclear magnetic resonance were used to confirm the structure of the latter metabolite. A mechanism leading to the formation of the keto acid metabolite has been suggested, and most intermediates were observed in rat plasma.
Journal of Pharmaceutical and Biomedical Analysis, Jul 1, 2003
Examples of the use of extraction-NMR, an efficient and rapid method to obtain structural informa... more Examples of the use of extraction-NMR, an efficient and rapid method to obtain structural information on metabolites without prior separation, are described. Crude ethyl acetate extracts of in vitro microsomal incubations were analysed by NMR spectroscopy. The region downfield of 5.5 ppm in the proton spectra of these crude extracts was found to be essentially clear of endogenous resonances. As a consequence, sites of aromatic hydroxylation can often be determined without prior separation of the crude extracts. Sites of metabolism close to the aromatic region may also be accessible via two-dimensional (2D) homonuclear experiments (e.g. COSY, NOESY, TOCSY). One-dimensional (1D) and 2D fluorine experiments also can provide additional information on the structure of metabolites. Depending on the complexity of the aromatic region of the parent compound, signal overlap and the relative abundance of the individual components, extraction-NMR has the potential to provide information for unambiguous structure elucidation of two or three major metabolites. Should extraction-NMR produce inconclusive results, i.e. too many metabolites are present or metabolism occurred exclusively on aliphatic regions, it is possible to re-use the extraction-NMR sample and proceed with traditional methods of analysis.
Journal of the Chemical Society, Perkin Transactions 2
ABSTRACT 13C NMR chemical shifts of vancomycin have been used to obtain the association constants... more ABSTRACT 13C NMR chemical shifts of vancomycin have been used to obtain the association constants for a number of ligands which bind weakly to the antibiotic. The change in these 13C chemical shifts upon stepwise addition of ligands provides evidence that the ligands bind in a manner analogous to natural cell-wall precursor analogues. The binding constants obtained are in good agreement with those determined earlier by other methods. Where the ligands are amino acids (glycine or alanine), a potential alkylammonium to amide carbonyl interaction does not promote binding.
![Research paper thumbnail of Synthesis and Spectral Characterization of Benzo-[6,7][1,5]diazocino[2,1-a]isoindol-12-(14H)-one Derivatives](https://attachments.academia-assets.com/75924376/thumbnails/1.jpg)
](Molecules (Basel, Switzerland), Jan 23, 2016
A simple synthetic route affording 27%-85% yields of benzo[6,7][1,5]diazocino[2,1-a]isoindol-12(1... more A simple synthetic route affording 27%-85% yields of benzo[6,7][1,5]diazocino[2,1-a]isoindol-12(14H)-one ring systems from readily available 3-(2-oxo-2-phenylethyl) isobenzofuran-1(3H)-ones and 2-(aminomethyl)aniline starting materials in toluene and catalysed by p-toluene-sulfonic acid is developed. The ¹H- and (13)C-NMR spectra of the final products were assigned using a variety of one and two-dimensional NMR experiments. The distinction between the two potential isomers of the final products was made on the basis of heteronuclear multiple bond connectivity (HMBC) NMR spectra.
Bioorg Medicinal Chem Letter, 2005
Chemical communications (Cambridge, England), Jan 14, 2016
An anthracene molecular probe has been synthesised and shown to target mephedrone, a stimulant dr... more An anthracene molecular probe has been synthesised and shown to target mephedrone, a stimulant drug from the cathinone class of new psychoactive substances (NPS). A protocol has been developed to detect mephedrone via the probe using NMR spectroscopy in a simulated street sample containing two of the most common cutting agents, benzocaine and caffeine.
J Am Chem Soc, 1993
ABSTRACT Evidence is presented for the formation of dimers in aqueous solutions of the glycopepti... more ABSTRACT Evidence is presented for the formation of dimers in aqueous solutions of the glycopeptide antibiotics eremomycin, A82846B, vancomycin, and eremomycin-PSI. The dimerization constant K(dim) is determined by H-1 NMR spectroscopy for the last two compounds and also for the related compound ristocetin-psi, for which dimerization has previously been reported in mixed solvents. Values of K(dim) are obtained for these compounds over a range of temperatures, and thus DELTAH(dim) and DELTAS(dim) are calculated. In addition, a lower limit for K(dim) in the case of eremomycin is calculated (10(5) M-1). This is a remarkably large value, and it may be that dimerization is implicated in antibiotic action. The possibility that natural selection has led to adaptations which promote dimerization (such as the nature and sites of attachment of the sugars and a ring 2 chlorine atom) is discussed.
Tetrahedron, 1993
... Wall Peptides to Vancomycin-Group Antibiotics: Factorising Free Energy Contributions to Bindi... more ... Wall Peptides to Vancomycin-Group Antibiotics: Factorising Free Energy Contributions to Binding Stephen E. Holroyd, Patrick Groves, Mark S. Searle, Ute Gerhard and Dudley H. Williants ... Rodriguez-Tebar, A.; Vazquez, D.; Perez Velazquez, JL; Laynez, J.; Wadso, I., J. Antibiot. ...
Analytical Chemistry, 2015
A novel approach is described for the quantitative bioanalysis of drugs in blood samples by ioniz... more A novel approach is described for the quantitative bioanalysis of drugs in blood samples by ionization of the analytes collected on solid-phase microextraction (SPME) fibers by mass spectrometry (MS). The technique combines the attractive features of SPME microsampling using minimal sample volumes with the speed, selectivity, and sensitivity capabilities of MS detection. The method reported in this study involved generating gas-phase ions directly from SPME fibers without the need for any additional sample preparation or chromatographic separation; the entire process was completed within 5 min. Traditionally, analytes extracted by SPME fibers are desorbed by washing with suitable solvents followed by a transfer into a sample vial and subsequent liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis to quantify the amount of analyte extracted and thereby determining the analyte concentration in the matrix. These sample preparation steps are completely eliminated by inserting the SPME fiber directly into the MS. Physiologically relevant concentrations of metoprolol and propranolol in blood samples were measured over several orders of magnitude down to concentration levels of 10 ng/mL. This preliminary assessment of direct SPME-MS showed high sensitivity (ng/mL), acceptable reproducibility (<30%), and lack of carryover. This novel approach simplifies current bioanalytical procedures providing time and cost savings. It demonstrates considerable potential for qualitative and quantitative pharmaceutical bioanalysis as well as other areas of challenging environmental and food analysis.
Journal of Organic Chemistry, Jul 19, 2000
Journal of Mass Spectrometry, Aug 1, 2004
Techniques such as mass spectrometry and NMR spectroscopy form an essential part of the medicinal... more Techniques such as mass spectrometry and NMR spectroscopy form an essential part of the medicinal chemist's toolbox for characterizing and assessing the purity of new molecules. Empowering medicinal chemists to gain early insight into their reaction products has a direct impact on productivity. Devolution of cutting-edge techniques from the specialist to the bench chemist also frees the specialist to concentrate on solving the more demanding of analytical problems. For open-access techniques to be taken up, they must be robust and be able to handle differing sample concentrations and varying sample complexities. This paper details the implementation of high-resolution liquid chromatography/mass spectrometry in open access to aid the medicinal chemist in characterizing desired products and identifying unexpected rearrangements, by-products and complete unknowns.
Proceedings of the National Academy of Sciences, 1993
An approach toward the estimation of binding constants for organic molecules in aqueous solution ... more An approach toward the estimation of binding constants for organic molecules in aqueous solution is presented, based upon a partitioning of the free energy of binding. Consideration is given to polar and hydrophobic contributions and to the entropic cost of rotor restrictions and bimolecular associations. Several parameters (derived from an analysis of entropy changes upon the melting of crystals and from the binding of cell wall peptide analogues to the antibiotic ristocetin A) which may be useful guides to a crude understanding of binding phenomena are presented: (i) amide-amide hydrogen bond strengths of -(1 to 7) +/- 2 kJ.mol-1, (ii) a hydrophobic effect of -0.2 +/- 0.05 kJ.mol-1.A-2 of hydrocarbon removed from exposure to water in the binding process, and (iii) free energy costs for rotor restrictions of 3.5-5.0 kJ.mol-1. The validity of the parameters for hydrogen bond strengths is dependent on the validity of the other two parameters. The phenomenon of entropy/enthalpy compen...
Journal of the American Chemical Society, 1994
It is demonstrated that the presence of bacterial cell wall analogues may either enhance or, in t... more It is demonstrated that the presence of bacterial cell wall analogues may either enhance or, in the case of ristocetin A, oppose dimerization of glycopeptide antibiotics. These observations may imply that dimerization plays a role in the mode of action of these antibiotics, and a mechanism is proposed to take account of this possibility. The glycopeptide dimers are also found to be formed more exothermically in the presence of cell wall analogues, and the nature of biological signaling events is discussed in this context. It is pointed out that binding enthalpy (rather than simply binding free energy, AG) may be an important quantity in signaling events. If this is so, then oligomers may be abundant in signaling processes partly because the extended aggregates they form are able to cooperatively amplify the conformational changes which are incurred on ligand binding, which occur through relatively small changes in free energy but larger opposing changes in enthalpy and entropy.
Journal of the American Chemical Society, 1994
A procedure for the determination of association constants in aqueous solution using hydrogendeut... more A procedure for the determination of association constants in aqueous solution using hydrogendeuterium exchange has been developed and used to measure the dimerization constant, &im, for a number of strongly dimerizing glycopeptide antibiotics. Thesevalues provide further insight into the thermodynamic contributions of various structural epitopes to the dimerization of these antibiotics. Consideration of ligand binding affinities together with dimerization potentials provides evidence that dimerization is implicated in the physiological mode of action of these antibiotics.
Journal of the American Chemical Society, 1991
An expression is presented for the estimation of approximate binding constants for bimolecular as... more An expression is presented for the estimation of approximate binding constants for bimolecular associations in solution. The consequences of the approach have been examined for the bimolecular association of two peptide components in aqueous solution: specifically for the binding of two vancomycin group antibiotics, vancomycin itself and ristocetin A, to the peptide cell wall analogue N-Ac-D-Ala-D-Ala and related ligands. Uncertainties in the treatment are relatively large, but the physical insights gained into the binding process (in part with the aid of calorimetric data obtained by others) are enlightening. We conclude that for amide-amide hydrogen bond formation in aqueous solution at room temperature, the intrinsic binding energy is ca. 24 kJ mol-' (an intrinsic binding constant of ca. lo4); this process is almost completely driven by a favorable entropy change associated with the release of water molecules from the amide NH and CO groups involved in hydrogen bond formation. The bimolecular association of N-Ac-D-Ala-D-Ala with ristocetin A has a remarkably small entropy change at 298 K (TAS = 3 f 1.5 kJ mol-'). We conclude that the release of water from polar and hydrocarbon groups involved in the binding almost exactly compensates for (i) the unfavorable entropy change due to the freezing out of four rotors of N-Ac-DAla-D-Ala upon binding and (ii) the unfavorable entropy change of a bimolecular association. A crude quantitation of these effects is presented. We also present an estimate of the increase in translational plus rotational free energy, as a function of the ligand mass, occurring when a ligand binds to a larger receptor. This quantity, fundamental to all binding processes, is relatively insensitive to the shape of the ligand. Extension of the approach will allow, in those cases where there is good complementarity between ligand and receptor, the prediction of approximate peptide-peptide binding constants in aqueous solution. 'Cambridge Centre for Molecular Recognition. $Smith Kline Beecham Pharmaceuticals.
The Journal of Antibiotics, 1995
The clinically important vancomycin group glycopeptide antibiotics, which act by blocking cell wa... more The clinically important vancomycin group glycopeptide antibiotics, which act by blocking cell wall synthesis, are crucial in the treatment of methicillin resistant Staphylococcus aureus. All of the group membersstudied so far, with the apparent exception of teicoplanin, enhance their antibiotic action by the formation of an asymmetric homodimer. Teicoplanin exists in two main conformers which differ by a rotation of approximately 1 80°ofa sugar residue. From NMRstudies and molecular modelling, we present structures for the two conformers and conclude that they have different binding affinities for cell wall analogues. The two conformers of teicoplanin are closely analogous to those adopted by each half of the asymmetric dimers of the other vancomycin group antibiotics. The vancomycin family of glycopeptide antibiotics1~4) are clinically important compounds, being active against Gram-positive bacteria. The emerging methicillin resistant Staphylococcus aureus (MRSA), which are re
Drug Metabolism and Disposition, 2002
[3R,5R,6S]-3-(2-cyclopropyloxy-5-trifluoromethoxyphenyl)-6-phenyl-1-oxa-7-azaspiro[4.5]decane is ... more [3R,5R,6S]-3-(2-cyclopropyloxy-5-trifluoromethoxyphenyl)-6-phenyl-1-oxa-7-azaspiro[4.5]decane is a substance P (Neurokinin 1 receptor) antagonist. Substance P antagonists are proven in concept to have excellent potential for the treatment of major depression, and they allow superior and sustained protection from acute and delayed chemotherapy-induced emesis. The metabolism of this compound was investigated in rat hepatocytes, and circulating rat plasma metabolites were identified following oral and intravenous dosing. The turnover in rat hepatocytes within 4 h was about 30%, and the major metabolites were identified as two nitrones and a lactam associated with the piperidine ring. Although these metabolites were also observed in rat plasma, the major circulating metabolite was a keto acid following oxidative de-amination of the piperidine ring. Liquid chromatography/tandem mass spectrometry and nuclear magnetic resonance were used to confirm the structure of the latter metabolite. A mechanism leading to the formation of the keto acid metabolite has been suggested, and most intermediates were observed in rat plasma.
Journal of Pharmaceutical and Biomedical Analysis, Jul 1, 2003
Examples of the use of extraction-NMR, an efficient and rapid method to obtain structural informa... more Examples of the use of extraction-NMR, an efficient and rapid method to obtain structural information on metabolites without prior separation, are described. Crude ethyl acetate extracts of in vitro microsomal incubations were analysed by NMR spectroscopy. The region downfield of 5.5 ppm in the proton spectra of these crude extracts was found to be essentially clear of endogenous resonances. As a consequence, sites of aromatic hydroxylation can often be determined without prior separation of the crude extracts. Sites of metabolism close to the aromatic region may also be accessible via two-dimensional (2D) homonuclear experiments (e.g. COSY, NOESY, TOCSY). One-dimensional (1D) and 2D fluorine experiments also can provide additional information on the structure of metabolites. Depending on the complexity of the aromatic region of the parent compound, signal overlap and the relative abundance of the individual components, extraction-NMR has the potential to provide information for unambiguous structure elucidation of two or three major metabolites. Should extraction-NMR produce inconclusive results, i.e. too many metabolites are present or metabolism occurred exclusively on aliphatic regions, it is possible to re-use the extraction-NMR sample and proceed with traditional methods of analysis.
Journal of the Chemical Society, Perkin Transactions 2
ABSTRACT 13C NMR chemical shifts of vancomycin have been used to obtain the association constants... more ABSTRACT 13C NMR chemical shifts of vancomycin have been used to obtain the association constants for a number of ligands which bind weakly to the antibiotic. The change in these 13C chemical shifts upon stepwise addition of ligands provides evidence that the ligands bind in a manner analogous to natural cell-wall precursor analogues. The binding constants obtained are in good agreement with those determined earlier by other methods. Where the ligands are amino acids (glycine or alanine), a potential alkylammonium to amide carbonyl interaction does not promote binding.
Molecules (Basel, Switzerland), Jan 23, 2016
A simple synthetic route affording 27%-85% yields of benzo[6,7][1,5]diazocino[2,1-a]isoindol-12(1... more A simple synthetic route affording 27%-85% yields of benzo[6,7][1,5]diazocino[2,1-a]isoindol-12(14H)-one ring systems from readily available 3-(2-oxo-2-phenylethyl) isobenzofuran-1(3H)-ones and 2-(aminomethyl)aniline starting materials in toluene and catalysed by p-toluene-sulfonic acid is developed. The ¹H- and (13)C-NMR spectra of the final products were assigned using a variety of one and two-dimensional NMR experiments. The distinction between the two potential isomers of the final products was made on the basis of heteronuclear multiple bond connectivity (HMBC) NMR spectra.
Bioorg Medicinal Chem Letter, 2005
Chemical communications (Cambridge, England), Jan 14, 2016
An anthracene molecular probe has been synthesised and shown to target mephedrone, a stimulant dr... more An anthracene molecular probe has been synthesised and shown to target mephedrone, a stimulant drug from the cathinone class of new psychoactive substances (NPS). A protocol has been developed to detect mephedrone via the probe using NMR spectroscopy in a simulated street sample containing two of the most common cutting agents, benzocaine and caffeine.
J Am Chem Soc, 1993
ABSTRACT Evidence is presented for the formation of dimers in aqueous solutions of the glycopepti... more ABSTRACT Evidence is presented for the formation of dimers in aqueous solutions of the glycopeptide antibiotics eremomycin, A82846B, vancomycin, and eremomycin-PSI. The dimerization constant K(dim) is determined by H-1 NMR spectroscopy for the last two compounds and also for the related compound ristocetin-psi, for which dimerization has previously been reported in mixed solvents. Values of K(dim) are obtained for these compounds over a range of temperatures, and thus DELTAH(dim) and DELTAS(dim) are calculated. In addition, a lower limit for K(dim) in the case of eremomycin is calculated (10(5) M-1). This is a remarkably large value, and it may be that dimerization is implicated in antibiotic action. The possibility that natural selection has led to adaptations which promote dimerization (such as the nature and sites of attachment of the sugars and a ring 2 chlorine atom) is discussed.
Tetrahedron, 1993
... Wall Peptides to Vancomycin-Group Antibiotics: Factorising Free Energy Contributions to Bindi... more ... Wall Peptides to Vancomycin-Group Antibiotics: Factorising Free Energy Contributions to Binding Stephen E. Holroyd, Patrick Groves, Mark S. Searle, Ute Gerhard and Dudley H. Williants ... Rodriguez-Tebar, A.; Vazquez, D.; Perez Velazquez, JL; Laynez, J.; Wadso, I., J. Antibiot. ...
Analytical Chemistry, 2015
A novel approach is described for the quantitative bioanalysis of drugs in blood samples by ioniz... more A novel approach is described for the quantitative bioanalysis of drugs in blood samples by ionization of the analytes collected on solid-phase microextraction (SPME) fibers by mass spectrometry (MS). The technique combines the attractive features of SPME microsampling using minimal sample volumes with the speed, selectivity, and sensitivity capabilities of MS detection. The method reported in this study involved generating gas-phase ions directly from SPME fibers without the need for any additional sample preparation or chromatographic separation; the entire process was completed within 5 min. Traditionally, analytes extracted by SPME fibers are desorbed by washing with suitable solvents followed by a transfer into a sample vial and subsequent liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis to quantify the amount of analyte extracted and thereby determining the analyte concentration in the matrix. These sample preparation steps are completely eliminated by inserting the SPME fiber directly into the MS. Physiologically relevant concentrations of metoprolol and propranolol in blood samples were measured over several orders of magnitude down to concentration levels of 10 ng/mL. This preliminary assessment of direct SPME-MS showed high sensitivity (ng/mL), acceptable reproducibility (<30%), and lack of carryover. This novel approach simplifies current bioanalytical procedures providing time and cost savings. It demonstrates considerable potential for qualitative and quantitative pharmaceutical bioanalysis as well as other areas of challenging environmental and food analysis.