The use of stable isotope labelling for the analytical chemistry of drugs.

This perspective reviews the potential for stable isotope labelling to examine the metabolic transformations of drugs. The increased sensitivity and widespread availability of modern nuclear magnetic resonance (NMR) and high-resolution mass spectrometers will increase the application of stable isotopes to study drug metabolism. Creating mass doublets by mixing a natural isotopic abundance compound with a labelled isotopomer and applying stable isotope filtering to high resolution mass spectrometry allows one to rapidly identify drug metabolites in very complex samples, such as blood or urine. Applying this approach to drug metabolism will require a significant synthesis effort. The relatively small number of (13) C, (15) N, or (17,18) O-labelled precursors exacerbates this problem, making the synthesis of the labelled drug often more difficult than that of the parent compound. We have developed new strategies for stable isotope labelling of complex molecules based on the rich chemistry of [(13) C]methyl phenyl sulfide, where the phenylthio group acts as a stable, non-volatile carrier for the valuable (13) C-label. For example we have used [(13) C]methyl phenyl sulfide to prepare the three possible (13) C-isotopomers ([1-(13) C]-, [2-(13) C]-, [1,2-(13) C(2) ]) of the two carbon precursors, ethyl 2-(phenylthio) acetate and ethyl N,N-dimethyl oxamate. In each case, these two-carbon labelling precursors are asymmetric and the differential reactivity of the carbons allows for either/or (13) C-labelling in the products. We demonstrate the utility of these two carbon precursors in the synthesis of aromatic ring-labelled N-(4-hydroxyphenyl)acetamide (acetaminophen or paracetamol).

[1]  N. Botting,et al.  First synthesis of [1,3,5-(13)C3]gallic acid. , 2009, Organic & biomolecular chemistry.

[2]  A. Nier,et al.  HEAVY CARBON AS A TRACER IN HETEROTROPHIC CARBON DIOXIDE ASSIMILATION , 1941 .

[3]  Christian Ludwig,et al.  Ligand based NMR methods for drug discovery. , 2009, Frontiers in bioscience.

[4]  T. Baillie,et al.  Recent Advances in the Use of Stable Isotopes in Drug Metabolism Research , 1986, Journal of clinical pharmacology.

[5]  S. Grzesiek,et al.  Measurement of homo- and heteronuclear J couplings from quantitative J correlation. , 1994, Methods in enzymology.

[6]  Hao Chen,et al.  Application of stable isotope labeled glutathione and rapid scanning mass spectrometers in detecting and characterizing reactive metabolites. , 2005, Rapid communications in mass spectrometry : RCM.

[7]  P. Hajduk,et al.  NMR-based screening in drug discovery , 1999, Quarterly Reviews of Biophysics.

[8]  Jie Chen,et al.  A generic method to detect electrophilic intermediates using isotopic pattern triggered data-dependent high-resolution accurate mass spectrometry. , 2008, Rapid communications in mass spectrometry : RCM.

[9]  L. Kay,et al.  Heteronuclear 3D NMR and isotopic labeling of calmodulin. Towards the complete assignment of the 1H NMR spectrum. , 1990, Biochemical pharmacology.

[10]  Ad Bax,et al.  Methodological advances in protein NMR , 1993 .

[11]  O. Pelkonen,et al.  Rapid detection and characterization of reactive drug metabolites in vitro using several isotope-labeled trapping agents and ultra-performance liquid chromatography/time-of-flight mass spectrometry. , 2009, Rapid communications in mass spectrometry : RCM.

[12]  D. Chace.  Mass spectrometry in the clinical laboratory. , 2001, Chemical reviews.

[13]  M. Mccomish,et al.  Characterization of diaminouracil metabolites of caffeine in human urine. , 1983, Drug metabolism and disposition: the biological fate of chemicals.

[14]  D. Cane,et al.  The biosynthesis of streptonigrin. 5. Biosynthesis of streptonigrin from [U-13C6]-D-glucose. Origin of the quinoline quinone , 1982 .

[15]  H. Seto,et al.  Utilization of carbon-13 carbon-13 coupling in structural and biosynthetic studies. Alternate double labeling method , 1973 .

[16]  C. Unkefer,et al.  On the biosynthetic origin of methoxymalonyl-acyl carrier protein, the substrate for incorporation of "glycolate" units into ansamitocin and soraphen A. , 2006, Journal of the American Chemical Society.

[17]  M. Vestal,et al.  Methods of ion generation. , 2001, Chemical reviews.

[18]  S. A. McLuckey,et al.  Mass analysis at the advent of the 21st century. , 2001, Chemical reviews.

[19]  Jie Chen,et al.  Metabolite identification by data-dependent accurate mass spectrometric analysis at resolving power of 60,000 in external calibration mode using an LTQ/Orbitrap. , 2007, Rapid communications in mass spectrometry : RCM.

[20]  C. Unkefer,et al.  Biosynthesis of pyrroloquinoline quinone. 2. Biosynthetic assembly from glutamate and tyrosine , 1991 .

[21]  Gary W Caldwell,et al.  Stable-isotope trapping and high-throughput screenings of reactive metabolites using the isotope MS signature. , 2004, Analytical chemistry.

[22]  M. Kainosho,et al.  Hydrogen exchange rate of tyrosine hydroxyl groups in proteins as studied by the deuterium isotope effect on C(zeta) chemical shifts. , 2009, Journal of the American Chemical Society.

[23]  Swapan Chowdhury,et al.  An accurate-mass-based spectral-averaging isotope-pattern-filtering algorithm for extraction of drug metabolites possessing a distinct isotope pattern from LC-MS data. , 2009, Analytical chemistry.

[24]  A. Bax,et al.  An efficient NMR approach for obtaining sequence‐specific resonance assignments of larger proteins based on multiple isotopic labeling , 1990, FEBS Letters.

[25]  R. Mortishire-Smith,et al.  Extracting metabolite ions out of a matrix background by combined mass defect, neutral loss and isotope filtration. , 2009, Rapid communications in mass spectrometry : RCM.

[26]  J. Castro-Pérez.  Current and future trends in the application of HPLC-MS to metabolite-identification studies. , 2007, Drug discovery today.